Clinical trial NCT01756833
Conditions: Aneurysm
Keywords: abdominal aortic aneurysm
Sponsor: University of Maryland, Baltimore
Start date: 2013-05-01; Completion date: 2019-07-23
Enrollment: 261 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
The primary aim of this study is to determine if doxycycline (100 mg bid) will inhibit (by at least 40%) the increase in greatest transverse diameter of small abdominal aortic aneurysms (3.5-5.0 cm in men, 3.5-4.5 cm in women) over a 24-month period of observation in comparison to a placebo-treated control group.

Detailed description:
N-TA\^3CT is a randomized, double-blind, placebo-controlled test of the hypothesis that doxycycline 100 mg bid, will reduce the rate of increase of maximum transverse diameter of small (3.5-5.0 cm among men and 3.5 to 4.5 cm among women) abdominal aortic aneurysms. The primary outcome is abdominal aortic aneurysm (AAA) maximum transverse diameter determined by CT scans at two-year follow-up with allowance for baseline (pre-randomization) diameter. Based on an anticipated growth rate of 2.5 mm per year in the placebo group and the current threshold at which surgical intervention will be offered to trial participants, (5.5 cm in men, 5.0 cm in women), the upper limit of AAA size for inclusion has been set at 5.0 cm for men and 4.5 cm for women. Among these subjects, the threshold for repair would be exceeded only by those exhibiting persistent growth. Secondary outcomes will determine if doxycycline affects other measures, e.g., MMP-9 levels in plasma and whether these effects are related to aneurysm growth. Nineteen clinical sites have identified pools of over 1600 patients with small aneurysm who meet the proposed inclusion/exclusion criteria. Two hundred fifty-eight patients will be randomized to placebo or doxycycline and their aneurysms followed for change in diameter at six-month intervals using CT imaging. The alternative hypothesis is that doxycycline will inhibit the expansion rate by 40% during the two years of observation. Patients enrolling in N-TA\^3CT must be able to give consent for their participation themselves and meet study eligibility criteria.

Arms: arm 1: 100 mg capsules, twice a day, for a period of two years. arm 2: 100 mg capsules, twice a day, for a period of two years.
Interventions: intervention 1: Doxycycline intervention 2: Placebo
Intervention descriptions: intervention 1: 100 mg po bid intervention 2: capsule identical to the doxycycline capsule

Protocol documents:
Prot_003.pdf: 
 
 
 
 
 
 
 
 
Non-Invasive Treatment of Abdominal Aortic Aneurysm Clinical Trial 
(N-TA3CT) 
 
 
 
 
 
 
 
 
 
 
 
PROTOCOL 
(Version 1.7.1) 
 
 
 
 
 
 
 
 
 


 
 
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Table of Contents 
 
CHAPTER 1 – BACKGROUND AND SIGNIFICANCE ......................................................... 6 
1.1 
BACKGROUND AND SIGNIFICANCE ....................................................................... 6 
1.2 
PRELIMINARY STUDIES ............................................................................................. 19 
CHAPTER 2 – STUDY OBJECTIVES .................................................................................... 27 
2.1 
PRIMARY AIM .............................................................................................................. 27 
2.2 
SECONDARY AIMS ...................................................................................................... 27 
2.3 
EXPLORATORY AIMS ................................................................................................ 27 
2.4      ACCOMPLISHING OBJECTIVES .............................................................................. 27 
CHAPTER 3 – OVERVIEW OF STUDY DESIGN AND METHODS ................................. 28 
3.1 
STUDY POPULATION .................................................................................................. 29 
3.1.1 
Inclusion Criteria .................................................................................................. 29 
3.1.2 
Exclusion Criteria ................................................................................................. 29 
3.2 
STUDY DESIGN ............................................................................................................. 29 
3.3 
STUDY TREATMENTS ................................................................................................ 29 
3.4 
END POINTS ................................................................................................................... 29 
3.4.1 
Primary Outcome .................................................................................................. 29 
3.4.2 
Secondary Outcomes ............................................................................................. 29 
3.4.3 Sample Size and Data Analysis ......................................................................................... 30 
3.4.4 Time Line ............................................................................................................................ 31 
CHAPTER 4 – PATIENT ELIGIBILITY, ORIENTATION AND RETENTION .............. 33 
4.1 
INCLUSION CRITERIA ............................................................................................... 33 
4.2 
EXCLUSION CRITERIA .............................................................................................. 33 
4.3 
INFORMED CONSENT ................................................................................................ 33 
4.4  
IDENTIFICATION OF POTENTIAL PATIENTS ..................................................... 35 
4.5 
PATIENT COMPETENCE TO GIVE INFORMED CONSENT .............................. 35 
4.6 
PROCEEDING FROM INFORMED CONSENT TO RANDOMIZATION ........... 36 
CHAPTER 5 – METHOD OF RANDOMIZATION ............................................................... 37 
5.1 
ELIGIBILITY ASSESSMENT ...................................................................................... 37 
5.2 
RANDOM TREATMENT ALLOCATION PROCEDURES ..................................... 37 
5.3  
CLINICAL SITE RANDOMIZATION START-UP ................................................... 37 
CHAPTER 6 – STUDY TREATMENTS .................................................................................. 37 
6.1 
INTRODUCTION ........................................................................................................... 37 
6.2 
TREATMENT PREPARATION ................................................................................... 38 
6.3 
BLINDING ....................................................................................................................... 38 
6.4 
EMERGENCY UNBLINDING ..................................................................................... 38 
6.5 
TREATMENT INTERRUPTIONS ............................................................................... 39 
6.6 
ASSESSMENT OF COMPLIANCE ............................................................................. 39 
6.7 
MISSED VISITS AND DROP OUTS ............................................................................ 39 
6.8 
DURATION OF STUDY TREATMENT ..................................................................... 39 
CHAPTER 7 – CONCOMITANT CARE ................................................................................. 39 
7.1 
INTRODUCTION ........................................................................................................... 39 
7.2 
HOSPITAL ADMISSIONS ............................................................................................ 39 
7.3 
BLOOD PRESSURE AND LIPID CONTROL ............................................................ 40 
7.3.1 
Anti-Hypertensives ................................................................................................ 40 
7.3.2 
HMG-CoA Reductase Inhibitors (“Statins”) and Other Lipid Lowering Agents
 
……………………………………………………………………………………..40 


 
 
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7.4 
TREATMENTS WHICH MAY INTERACT WITH DOXYCYCLINE ................... 40 
7.4.1 
Anti-Infectives ........................................................................................................ 40 
7.4.2 
Wafarin .................................................................................................................. 40 
7.5 
SMOKING AND OTHER TOBACCO USE CESSATION ........................................ 41 
7.6  
SURGERY ........................................................................................................................ 41 
7.6.1 
Introduction ........................................................................................................... 41 
7.6.2 
Retroperitoneal Surgery ....................................................................................... 41 
CHAPTER 8 – FOLLOW-UP PROCEDURES ....................................................................... 41 
8.1 
INTRODUCTION ........................................................................................................... 41 
8.2 
FOLLOW-UP VISITS .................................................................................................... 41 
8.2.1 
CT Scan Acquisition at Enrollment and Follow-up ........................................... 42 
8.2.2 
Biomarker Blood Specimen Collection at Enrollment and Follow-up ............. 43 
8.2.3 
Ascertainment of Adverse Events ........................................................................ 43 
8.3 
PATIENT MANAGEMENT DURING MEDICAL CONTACTS ............................. 43 
8.4 
 SECONDARY ASSESSMENTS ................................................................................... 44 
8.4.1 
Psychosocial Evaluation ........................................................................................ 44 
8.5 
 RETENTION .................................................................................................................. 44 
8.5.1  
Retention Promotion Efforts ................................................................................ 44 
8.5.2  
Monitoring Recruitment and Retention .............................................................. 45 
8.6  
FINAL FOLLOW-UP ASSESSMENT AND CLOSE-OUT ....................................... 45 
CHAPTER 9 – STUDY END POINTS ..................................................................................... 46 
9.1 
INTRODUCTION ............................................................................................................ 46 
9.2 
PRIMARY END POINT ................................................................................................. 46 
9.3 
STATISTICAL ANALYSIS OF THE PRIMARY END POINT ................................ 46 
9.4 
SECONDARY AND INTERMEDIATE OUTCOMES ................................................ 47 
9.4.1 
Clinical Secondary and Intermediate Outcomes ................................................ 47 
9.4.2 
Biomarker Secondary and Intermediate Outcomes .......................................... 48 
9.4.3 
CT-Scan Secondary and Intermediate Outcomes .............................................. 48 
CHAPTER 10 – DATA MANAGEMENT ................................................................................ 48 
10.1 INTRODUCTION ............................................................................................................ 48 
10.2 FORM DATA ................................................................................................................... 48 
10.3 CT SCANS ........................................................................................................................ 49 
10.4 BIOMARKER SPECIMENS .......................................................................................... 49 
10.5 QUALITY CONTROL .................................................................................................... 49 
10.5.1 
Training of Study Personnel ................................................................................. 50 
10.6 QUALITY MONITORING ............................................................................................. 50 
10.6.1 
Data Collection ...................................................................................................... 50 
10.6.2 
Recruitment and Randomization Rates .............................................................. 51 
10.6.3 
Response to Protocol Violations ........................................................................... 51 
10.6.4 
Response to Lapses in Clinical Site Performance .............................................. 51 
CHAPTER 11 – DATA ANALYSIS .......................................................................................... 52 
11.1 INTRODUCTION ............................................................................................................ 52 
11.2 INTERIM MONITORING ............................................................................................. 52 
11.3 PRIMARY ANALYSIS ................................................................................................... 53 
11.4 SUBGROUP ANALYSES OTHER SECONDARY OUTCOMES AND ANALYSES .      
…………………………………………………………………………………………………... 54 
11.5 STATISTICAL POWER ................................................................................................. 55 
CHAPTER 12 – STUDY ORGANIZATION ............................................................................ 59 


 
 
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12.1 PARTICIPATING UNITS .............................................................................................. 59 
12.1.1 
Clinical Site ............................................................................................................ 59 
12.1.2 
Clinical Coordinating Center ............................................................................... 59 
12.1.3 
Data Coordinating Center .................................................................................... 60 
12.1.4 
Imaging Core Laboratory .................................................................................... 60 
12.1.5 
Biomarkers Core Laboratory .............................................................................. 60 
12.1.6 
Study Chairman .................................................................................................... 60 
12.2 
ADMINISTRATIVE STRUCTURE ............................................................................. 60 
12.2.1 
Steering Committee ............................................................................................... 60 
12.2.2 
Management Committee ....................................................................................... 61 
12.2.3 
Publications and Ancillary Studies Committee .................................................. 61 
12.2.4 
Data and Safety Monitoring Board ..................................................................... 61 
12.2.5 
Role of DCC Staff .................................................................................................. 61 
12.2.6 
Data Coordinating Center Interface with Clinical Coordinating Center ........ 61 
12.2.7 
Data Coordinating Center Role in Training of Study Personnel ...................... 62 
12.3 
STUDY TIME LINE ....................................................................................................... 62 
CHAPTER 13 – POLICY MATTERS ...................................................................................... 63 
13.1 
PUBLICATION POLICY .............................................................................................. 63 
13.1.1 
General Statement of Editorial Policy ................................................................. 63 
13.1.2 
Types of Research .................................................................................................. 64 
13.1.3 
Authorship ............................................................................................................. 65 
13.1.4 
Purpose of Procedural Guidelines ....................................................................... 65 
13.2 
DESIGN AND METHODS REPORTS, MAINLINE FINDINGS, DATA BANK, 
ANCILLARY, AND INDEPENDENT STUDIES ........................................................ 65 
13.2.1 
Design Papers and Reports on Methodology ...................................................... 65 
13.2.2 
Reports of Mainline Findings ............................................................................... 65 
13.2.3 
Data Bank Studies ................................................................................................. 65 
13.2.4 
Ancillary Studies ................................................................................................... 65 
13.2.5 
Independent Studies .............................................................................................. 66 
13.3 
GUIDELINES FOR PREPARATION OF PROPOSALS FOR DATA BANK AND 
ANCILLARY STUDIES ................................................................................................. 66 
13.3.1 
Data Bank and Ancillary Studies ......................................................................... 66 
13.4 PROCEDURES FOR INITIATION AND APPROVAL OF STUDIES ..................... 66 
13.4.1 
Reports on Mainline Findings .............................................................................. 66 
13.4.2 
Data Bank Studies ................................................................................................. 67 
13.4.2.1 Submission of Proposals ............................................................................................... 67 
13.4.2.2 Conduct of Data Bank Studies ..................................................................................... 67 
13.4.2.3 Priorities for Work ........................................................................................................ 67 
13.4.2.4 Authorship ..................................................................................................................... 68 
13.4.2.5 Review and Approval of Abstracts and Manuscripts Prior to Presentation or 
Publication……………………………………………………………………………………...  68 
13.4.3 
Ancillary Studies ................................................................................................... 68 
13.4.3.1 Submission of Proposals ............................................................................................... 68 
13.4.4 
Independent Studies .............................................................................................. 68 
13.5 RELEASE OF N-TA3CT DATA OR SPECIMENS TO NON- N-TA3CT  
INVESTIGATORS ...................................................................................................................... 68 
13.6 CONFLICT OF INTEREST POLICY .......................................................................... 69 
13.6.1 
General Principles ................................................................................................. 69 


 
 
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13.6.2 
Individuals to be Governed by These Guidelines ............................................... 69 
13.6.3 
Time Period of the Policy ...................................................................................... 69 
13.6.4 
Financial Guidelines .............................................................................................. 69 
13.6.5 
Reporting of Financial Disclosures and Other Activities .................................. 70 
13.6.6 
Review of Policy Statement .................................................................................. 70 
13.6.7 
Relationship to Institutional Policies on Conflict of Interest ............................ 70 
13.7 HUMAN SUBJECTS TRAINING .................................................................................. 70 
Appendix – N-TA3CT Procedures and Treatments ................................................................. 80 
 


 
 
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CHAPTER 1 – BACKGROUND AND SIGNIFICANCE 
1.1 
BACKGROUND AND SIGNIFICANCE 
Scope of the Clinical Problem. Abdominal aortic aneurysms (AAAs) are a common and 
life-threatening condition. The natural history of AAA is one of progressive expansion. The rate 
of aneurysm growth and the risk of rupture are both exponentially related to aneurysm size. 
Aortic diameter is therefore the single best predictor of rupture risk.1,2 Ninety-five percent of 
aortic aneurysms occur in the aortic segment below the renal arteries; the normal aortic diameter 
at this level is 2.0-2.5 cm and risk of aneurysm rupture is low until aortic diameter increases 
beyond 5.5 cm.3-5 AAAs are typically asymptomatic and are often undetected until late in their 
natural history, when the weakened aortic wall is prone to rupture. Spontaneous rupture of an 
aortic aneurysm is usually fatal and is currently responsible for at least 15,000 deaths each year 
in the U.S.6 This is a conservative estimate of AAA-associated deaths. 
Following AAA rupture, most patients (90%) do not survive long enough to reach 
medical attention, and the mortality rate remains high for those who undergo emergency surgical 
repair.7 Despite numerous advances in surgery, anesthesia and critical care, it is unclear that 
operative mortality for ruptured aneurysms has been significantly impacted. The vast majority of 
ruptured AAAs are in patients previously undiagnosed with aneurysm disease and thus are 
preventable through early detection and elective surgical repair.8-10 Elective AAA repair is 
indicated for patients with AAAs at significant risk of rupture, a clinical decision currently based 
on aneurysm diameter greater than 5.5 cm for men.4,5  
Efforts to limit the mortality from AAAs are dependent on early detection and elective 
surgical repair of the diseased aorta. Although we know of approximately 55,000 patients with 
AAAs that come to clinical attention each year (15,000 with ruptured AAAs and 40,000 
undergoing elective AAA repair), the overall incidence of AAAs appears to be much higher than 
these figures would suggest.6,11  Prospective screening studies using abdominal ultrasound 
indicate that AAAs occur in 4-9% of the population over the age of 65, as defined by an 
infrarenal aortic diameter greater than 3.0 cm.12,13 There is also evidence that the general 
prevalence of AAAs is steadily increasing, both in the U.S. and other industrial nations. Even by 
conservative estimates, extrapolation of these figures to U.S. census data indicates that aneurysm 
disease currently affects at least 1.7 million individuals.14 Moreover, the incidence of AAAs can 
be expected to exceed 3 million by the year 2025 because of the aging US population.  
Several factors help explain the discrepancy between the known incidence of AAAs and 
the incomplete number of patients that are currently treated for this disease. For example, AAAs 
are usually asymptomatic until rupture and routine physical examination is insensitive in 
detecting nonruptured AAA. Thus, many aortic aneurysms are either unsuspected or simply 
missed, even in populations at risk. Deaths from ruptured AAAs are significantly 
underrepresented in statewide mortality data, as sudden death is typically attributed to other 
cardiopulmonary disease in the absence of autopsies, with autopsy rates below 5%.15 Secondly, 
abdominal imaging studies are extremely sensitive in detecting AAAs of all sizes, such that most 
AAAs requiring treatment are simply found by serendipity during studies performed for 
unrelated reasons. Smaller AAAs identified in this fashion may not be reported since specific 
treatment is not carried out. Finally, up to 90% of the AAAs detected in ultrasound screening 
programs, which are below a threshold that warrants immediate repair at the time of initial 
diagnosis (i.e., less than 5.5 cm diameter).14,16,17  
Detection of a small aneurysm raises a considerable management dilemma, in that the 
natural history of small asymptomatic AAAs is one of gradual expansion (at rate of 2.6-5.4 mm 


 
 
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per year) and eventual rupture (see Table 1 below). Because there are no proven medical 
interventions capable of suppressing aneurysm growth, the detection of small AAAs only leads 
to the need for serial measurements until repair is indicated. This "watchful waiting" approach is 
particularly unsettling given that aneurysms grow in an uneven and unpredictable fashion and 
that the biological factors influencing aneurysm expansion are incompletely understood. The 
absence of medical interventions by which to treat the large number of patients with small AAAs 
has diminished enthusiasm for ultrasound screening programs on the basis of cost. It is clear, 
however, that better approaches will be needed over the next decade for the evaluation and 
management of patients with small asymptomatic AAAs. 
Defining aortic aneurysms for a clinical trial. Aortic diameter is normally distributed, 
changes with age and is related to gender.3 Understanding the changes and variation in normal 
aortic diameter is important for making the distinction between aortic dilatation and the initiation 
of the aneurysmal process. The most pronounced increase in normal aortic diameter occurs 
during childhood and adolescence but there is also a 25 percent increase in aortic diameter 
between ages 25 and 75. This increase is greater for males than females, but since there is no 
significance difference between the sexes when data are normalized to body surface area, body 
mass rather than gender may be the important factor. At age 75, the normal infrarenal aortic 
diameter can range from 12.4 mm in a small woman to 27.6 mm in a large man. Aortic 
aneurysms are usually described by their greatest diameter. Several definitions of AAA relative 
to other segments of the aorta have been proposed, but because an infrarenal aortic diameter in 
excess of 29 mm is above the upper limit of normal regardless of age, sex and body surface area, 
a diameter in excess of 3.0 cm is the simplest and most practical definition of an aortic 
aneurysm. From the earliest studies of AAA size relative to rupture risk, the greatest transverse 
diameter has been used. The use of multichannel CT imaging now allows for volume 
determination within an aneurysm and there has been significant interest and controversy 
regarding the sensitivity of changes in volume compared to diameter. Since each aneurysm has a 
unique shape, there are no guidelines that define AAA by volume. All clinical decisions 
regarding intervention and rupture risk are based on diameter. To summarize, we have chosen 
the lower threshold of 3.5 cm for several reasons including: 1) aneurysms between 3.0 and 3.5 
cm demonstrate significantly lower expansion rates with greater variability; 2) a 3.5 cm 
infrarenal aortic diameter distinguishes aneurysmal disease from any form of diffuse 
arteriomegaly which may have a different clinical course; 2) the expansion rates for aneurysms 
between 3.5 and 5.0 cm are well-defined and relatively homogeneous. The rationale for choosing 
an upper threshold of 5.0 cm is relatively straightforward. Our primary endpoint is growth rate 
over a 2 year time frame. The study protocol allows for intervention when the threshold of 5.5 
cm is reached. Based on the average anticipated growth rate of 2.5 mm/year in a man enrolled 
with a 5.0 cm AAA, the threshold of 5.5 cm will be reached and detected at the time of the two 
year follow-up CT scan.   
The natural history of aortic aneurysms-defining growth rates. The natural history of an 
untreated AAA is one of progressive expansion and eventual rupture. Retrospective studies using 
ultrasound or CT scan have defined mean expansion rates. Results from a number of the larger 
published trials are shown in the following table. 
 
 
 
 


 
 
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Study 
AAA size 
Patient number 
Primary 
i
i
Growth mm/yr. 
PAT18 
3.0-5.0 
272* 
US 
2.6 
UK trial5 
4.0-5.5 
321* 
US 
3.3 
ADAM4 
4.0-5.4 
567* 
CT 
3.2 
Schlosser19 
3.5-5.0 
  92* 
US 
3.1 
 
*Number of patients in control group (placebo treated or nonsurgical); PAT = Propanolol Aneurysm Trial 
This includes the three largest prospective trials and recent prospective trials and 
demonstrate a consistent expansion rates at 2.6-3.3 mm/yr. The lower growth rate in the trial by 
Laupacis reflects to inclusion of small aneurysms in the 3.0-3.5 range which exhibit a growth 
rate of less than 2 mm/year with significant variability. This is partly related to the fact that a 
significant number of aortas in this size range will not progress. The lower threshold for our 
proposed trial is 3.5 cm. Conversely, the UK trial and ADAM trial have the highest growth rates 
since they included aneurysms up to 5.5 cm. We have set the upper limit of our study at 5.0 cm 
for men and 4.5 cm for women so that the patients can be followed for some time before 
reaching the surgical threshold (5.5 cm for men and 5.0 cm for women). A recent study by 
Schlosser et al included patients in a screening and surveillance program with aneurysms 
between 3.0 and 5.5. Dr. Schlosser was kind enough to provide the data set of patients between 
3.5 and 5.0 and this is shown in the table. We included these data to show that growth rates have 
not changed over time since publication of the larger prospective trials. For the purposes of this 
trial, we anticipate a growth rate of 2.5 mm/yr. We believe that this is an accurate (if not 
conservative) estimate considering the size of aneurysms to be included and followed in this 
trial.  
 
Screening for AAA. Using an approximate growth rate of 2.5 mm/yr. for an aneurysm 
that is 3.5 cm at the time of detection, the typical threshold for intervention (5.5) would be 
reached within 7 years of diagnosis (Figure 1). 
 
The current management if a small AAA is detected consists of follow-up imaging to 
detect expansion, coupled with elective surgical repair when the aorta reaches 5.5 cm diameter 
(i.e., "watchful waiting"). Because there are no proven medical interventions capable of 
suppressing aneurysm growth, the detection of small AAAs leads to dilemma for the patient and 
physician. They are faced with a problem that requires serial observation but for which there is 
no therapy until repair is indicated. The rationale for waiting is bases on two important facts: 1) 
the risk of repair exceeds the risk of rupture for AAA less than 5.5 cm in man and less than 5.0 
cm in women; 2) and because of age and comorbid disease many patients die of other causes 
before their AAA reaches the threshold for repair.20 Most aneurysms detected through population 
screening studies are small at the time of discovery. While this fact would seem to be of benefit 
to the patient, the fact that only 10% of aneurysms detected by screening are above the threshold 
for immediate repair is the precise reason screening is not now considered cost-effective except 
in select groups. Because of this low incidence of aneurysms requiring immediate intervention, 
the cost of screening per life saved has been estimated to be as high as $78,000.21,22 If, however, 
an inexpensive medical treatment (such as doxycycline) were available for the 90% of patients 
with aneurysms below the operative threshold, ultrasound screening would be considered highly 
cost effective. One time screening is now recommended only for, men over age 64 who have 
ever smoked. CMS now covers the cost of screening for AAA in this group at the time of their 
Medicare introductory physical examination. 


 
 
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Factors influencing aneurysm progression. AAA have been typically detected by 
serendipity when abdominal or pelvic imaging studies are obtained for an unrelated problem. 
The mean growth rate for small AAAs (≤ 5.5 cm) is 2.5 mm/y, increasing with aneurysm 
diameter.(see table above) Studies of AAA expansion, and the factors associated with expansion, 
have been limited by sample size or a limited number of serial observations. In the UK small 
aneurysm trial, AAA expansion in 1,743 patients followed for up to 7 years was most strongly 
associated with diameter at baseline.23 No association with growth rate was noted between age or 
gender. Self-reported cigarette smoking status was associated with an incrementally increased 
growth rate of 0.4 mm/year which persisted after adjustment for potential confounding variables. 
Of other potential risk factors considered in the UK SAT, including hypertension, peripheral 
arterial occlusive disease (PAD), total or HDL plasma cholesterol concentration and diabetes, 
only the presence of PAD or diabetes influenced aneurysm growth; PAD decreasing it by 0.2 
mm per year for each 0.2 change in ankle brachial index (95% CI -0.03 to 0.25), and diabetes 
reducing the growth rate by 0.79 mm/year (95% CI 0.27 to 1.33). Based on these data, 
investigators calculated that screening intervals of 36, 24, 12, and 3 months for patients with 
AAA diameter of 35, 40, 45, and 50 mm, respectively, yield less than a 1% chance of patients 
unexpectedly exceeding 55 mm in diameter between examinations.24 In clinical practice, 
examination intervals vary but rarely exceed more than 12 months with increasing frequency 
associated with progressive enlargement. Part of the reason for the more frequent studies is 
reassurance for both the patient and physician. Quality of life surveys indicate that diagnosis 
without treatment of AAA is usually associated with significant anxiety.  
Although not considered in the analyses of most AAA trials, lifelong patterns of lower 
extremity exercise may provide some protection from AAA. Computational flow modeling 
studies of hemodynamic conditions in the distal aorta suggest that the decreased flow from 
prolonged sedentary existence may promote aneurysmal disease.25 Indirect clinical evidence in 
Aneurysm Growth 
Diameter (cm)
Years
Normal Growth
40%  Reduction
0 1 2 3 4 5
6 7
8
9 10
Intervention
12
11 
3 
4 
5
6 
7 
8 
9 
1
Figure 1 


 
 
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support of this concept include the fact that above knee traumatic amputation and chronic spinal 
cord injury are associated with increased AAA risk independent of other risk factors including 
cigarette smoking.25,26 
Tobacco. Tobacco smoking as a specific risk factor for AAA disease prevalence, 
incidence and progression deserves special mention. The relative risk of AAA in individuals who 
have ever smoked is 2.5 times greater than the relative risk for coronary heart disease.27 AAA is 
more closely associated with cigarette smoking than any other tobacco-related disease except 
lung cancer. 27 Nearly all (>90%) AAA patients relate a history of smoking, however, only about 
half of those continue to smoke at the time of diagnosis.28 Several small studies have associated 
continued cigarette smoking with more rapid aneurysm expansion, Lindholt evaluated and 
prospectively followed 117 AAA patients. He found a positive correlation between continued 
smoking and the rate of expansion.29 In the UK SAT itself, smoking and initial aneurysm size 
were the only two factors positively associated with aneurysm growth although they did not find 
a dose response between self-reported smoking habits or serum cotinine levels and aneurysm 
growth rate.24 Animal studies have confirmed accelerated aneurysm growth with cigarette smoke 
exposure.30 When the studies are considered together, the best evidence suggests that continued 
smoking is associated with a relatively small (15%) increase in growth rate that, when 
compounded over several years, has important implications. At the present time, 
recommendations for and assistance with smoking cessation is the standard of care although 
initial success rates are low.   
-blockers. Several animal studies have indicated that propranolol might have beneficial 
effects on aneurysmal disease based on both its hemodynamic properties and its biochemical 
effects on matrix proteins. Two clinical studies used retrospective analysis to assess the impact of 
-blockers in aneurysm growth rates.31,32 Both identified a significant inhibitory effect of -
blockers. These studies were the basis for two multicenter randomized trials testing propranolol 
in aneurysm patients. Propranolol did not inhibit aneurysm expansion in a trial reported by 
Lindholt et al.33 These results were compromised by low compliance in the propranolol arm; 
only 22% of patients continued the medication for 2 years. A Canadian trial that recruited 552 
patients suffered similarly from compliance problems.18 The growth rate in the placebo group 
and in the propranolol group did not differ although there was a slight trend in favor of 
propranolol. Quality of life, assessed by the SF-36 questionnaire, showed that propranolol had a 
significant negative effect as one would anticipate from the low compliance rate. A randomized 
trial of selective beta-blockers has not been performed. 
Statins. Statin therapy reduces the progression of atherosclerosis and improves clinical 
outcomes in cardiovascular diseases. Although effective in reducing atherogenic lipoproteins, 
statins also demonstrate additional biologic effects (i.e., pleiotropic effects), including reduction 
of high sensitivity C-reactive protein (CRP) levels, that may be relevant to the pathogenesis of 
AAA disease.34 Several studies have found an association between the presence of AAA and 
total cholesterol.35,36 There is, however, no clear relationship between total cholesterol and AAA 
expansion rate. Despite the absence of a relationship between cholesterol and growth rate, there 
is evidence from a number of studies suggesting that statins may influence aneurysm growth 
rate, presumably via these pleiotropic effects. Simvastatin therapy at 2 mg/kg/day reduces both 
aortic diameter and the percentage of mice with aneurysms after elastase infusion.37 No changes 
in effect size were noted by repeating these experiments in hypercholesterolemic apoE-deficient 
mice.  


 
 
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In human AAA specimens explanted for organ culture, addition of cervistatin reduces 
tissue levels of both total and active MMP-9 in a concentration-dependent manner. Evans et al 
treated AAA patients with a three week pre-operative course of simvastatin and showed that 
MMP-9 levels in excised aneurysm tissue were decreased.38 It is unclear, however, that these 
observations translate into an effect on the expansion rate of an existing aneurysm. The VA 
ADAM study, by far the largest prospective study with serial longitudinal observations, did not 
find an effect on expansion rate from lipid lowering medication.27 Several cohort studies have 
found decreased aneurysm expansion in patients on statins.19,39 Historically, these types of cohort 
studies overestimate this effect, a lesson learned from cohort studies of beta-blockers. Felix 
Schlosser provided data from their recent cohort study showing that patients with 3.5-5.0 cm 
AAA and taking statins grew at 2.8 mm/year which is not significantly different from growth 
rates in these cohorts collected before widespread use of statin.(personal communication) We 
hypothesize that growth in our control group will be 2.5 mm/year. These data from Schlosser 
demonstrate that even if statins exhibit some small inhibitory effect, growth rates in those 
patients taking statins are still expected to be greater than the 2.5 mm/year we have chosen as a 
conservative estimate of expected growth rate. If growth rates in our control groups are greater 
than 2.5 mm/year, the power of our study to detect 40% inhibition will increase significantly. 
Finally, we provide evidence in the section on preliminary data showing that doxycycline 
exhibits inhibitory effects on a wide array MMPs and inflammatory signaling pathways that are 
not or are only minimally affected by statins. 
Because AAA, CAD and PVD share common risk factors including dyslipidemia, there 
will be clear indications for statin use in most AAA patients. Efforts to meet the National Heart, 
Lung, and Blood Institutes (NHLBI) increasingly stringent Adult Treatment Protocol (ATP) 
guidelines for LDL management has led to wider use of statins. The Women’s Health Initiative 
has categorized AAA as a PVD equivalent, a relative indication for statins.40 A high prevalence 
of statin use among AAA patients will make it impossible to perform a randomized trial of 
statins that could definitively determine whether statins impact aneurysm growth rate. At present, 
the best assessment of the statin effects on AAA expansion will be a prospective cohort study 
with longitudinal follow-up. The study design we have proposed will address this important 
question by information by correlating statin use with the precise measurements of aneurysm 
expansion over a two year period.  
ACE inhibitors and angiotensin receptor blockers. ACE inhibitors have shown to both 
stimulate and inhibit MMPs depending on cell type or animal model. Losartan does not appear to 
have a direct effect on MMPs. A number of animal experiments using different models of 
aneurysmal disease have suggested an important role for the angiotensin/renin axis in aneurysm 
development. Captopril, but not losartan, an angiotensin receptor blocker, prevents aneurysm 
formation in the rat elastase model of AAA.41 Another commonly studied aneurysm model is 
based on chronic infusion of angiotensin II into apo E deficient mice resulting initially in 
midaortic dilatation and eventual rupture.42 Losartan prevents aneurysm formation in this model. 
This effect of losartan is consistent with observations in genetically engineered mice with 
Marfan Syndrome (MFS). Work done in these mice has suggested that the inability of mutated 
fibrillin to sequester TGF- plays a role in the progression of tissue changes associated with 
MFS.43 In a series of studies, TGF- antagonism by losartan was effective in preventing 
progressive matrix degradation.44 The reason for the discrepant effects of losartan -- ineffective 
in the elastase aneurysm model and effective in the angiotensin and Marfan models -- may relate 
to differences among the models. In the angiotension infusion model, initial dissection of the 


 
 
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12 
 
upper abdominal aorta is followed by dilatation.45 This process may have more similarities to the 
Marfan syndrome models where the thoracic aorta is affected. Clinical trials of losartan in MFS 
have recently begun enrollment.  
Hackam et al recently published results of an analysis of a linked administrative data base 
from Ontario Canada analyzing ruptured (n=3379) and nonruptured aortic aneurysms (n=11,947) 
from 1992-2002.46 ACE inhibitor use within the prior 3-12 months was less frequent among 
those admitted for aneurysm rupture (OR 0.82; CI 0.74-0.90). Beta-blockers, lipid lowering 
agents, angiotensin receptor blockers showed no relationship to rupture. In a published response 
to the paper, Lederle and Taylor note that among those patients who discontinued ACE inhibitors 
within the past 3-12 months, there is a harmful effect in favor of aneurysm rupture.47 The case 
control study by Schouten et al and post hoc analysis of the UK aneurysm trial data did not find a 
relationship between ACE inhibitors and aneurysm expansion rates.24,48 Most patients presenting 
with aneurysm rupture have large, undetected aneurysms while patients with known aneurysms 
typically undergo repair long before their rupture risk becomes significant. Thus, this 
information regarding ACE inhibitors and rupture risk might find its most practical application 
among the small number of patients deemed unfit for repair. The proposed trial may provide 
more insight into the effects of ACE inhibitors. Information regarding the use of ACE inhibitors 
can be correlated with aneurysm expansion rate in post hoc analysis. 
Macrolides. A number of antibiotics have been proposed as a treatment for AAA with 
varying rationales. One line of reasoning is that AAA progression is enhanced by secondary 
infection within the aortic wall. Chlamydia pneumonia has been found in atherosclerotic plaque 
and the wall of abdominal aortic aneurysms. There was once great enthusiasm for the hypothesis 
that treatment of the secondary Chlamydial infection could slow progression of atherosclerosis. 
This has been diminished by subsequent prospective randomized trials showing no 
cardiovascular benefit to a year of a treatment with azithromycin in patients with stable CAD.49 
A small study by Lindholt et al. suggested that serologic evidence of a Chlamydia pneumonia 
infection was associated with an increased rate of aneurysm expansion.50 This led to a 
randomized clinical trial in which 43 patients received a one month course of roxithromycin 
while 49 patients received placebo.51,52 Patients in the treatment arm had an expansion rate at the 
end of the study of 1.56 mm per year compared to a rate of 2.75 mm per year in the placebo-
treated group. The inhibition was greater in the first year than the second year. The study did not 
clarify the mechanism of effect since there was no correlation between Chlamydia titers and the 
ability of roxithromycin to inhibit aneurysm expansion. 
Tetracyclines. The tetracycline antibiotics have been studied because of their known 
inhibition of matrix metalloproteinases (MMP). Petrinic et al were the first to demonstrate that 
doxycycline could suppress aortic wall MMP activity, elastin degradation, and aneurysm 
development in the elastase-induced rat model.53 They achieved similar results using non-
antimicrobial (chemically-modified) tetracyclines and nonselective hydroxamic acid derivatives 
as MMP inhibitors, indicating that the aneurysm-suppressing effects of doxycycline are most 
likely related to its activity as an MMP inhibitor.54 Longo et al characterized a second murine 
aneurysm model using calcium chloride applied to the ablumenal surface to induce the 
aneurysm.55 In this model, doxycycline demonstrates the same dose-dependent inhibition of 
aneurysm expansion.56 The plasma doxycycline levels achieved in these animal studies were in 
the same range as those seen in AAA patients receiving doxycycline (100 mg bid).57 These 
murine studies suggest that inhibition can still be achieved at plasma levels in the 1-2 ug/ml 
range.56 


 
 
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A number of studies in patients have suggested that doxycycline can inhibit MMPs in 
aneurysm tissue. Curci et al treated a series of patients with a three week course of doxycycline 
prior to open aneurysm repair.58 Tissue levels of MMP-9 were significantly reduced by 
doxycycline in comparison to untreated patients. Baxter et al. in a small series of 36 patients on a 
six-month course of doxycycline, showed plasma MMP-9 levels decreased significantly in 
comparison to baseline levels.57 This work has been followed by a small prospective randomized 
trial of doxycycline in which 32 patients were randomized with 17 receiving doxycycline (150 
mg/day) for 3 months.59 Chlamydia pneumonia titers were assessed but found not to be affected 
by doxycycline treatment. The calculated growth rate at the end of the 18 month period of 
observation was 1.5 mm per year in the doxycycline treated group versus 3.0 mm per year and 
the placebo treated group. This difference did not achieve statistical significance but the 6 and 12 
month time periods did show a significant difference in favor of doxycycline treatment. Level B 
evidence (small randomized trials) suggests that roxithromycin or doxycycline will decrease the 
rate of aneurysm expansion. 
Potential Medical Therapies for Treating Small AAA 
Intervention 
Effect on AAA growth Level of evidence 
Class of 
recommendation 
Propranolol18,33 
no inhibition 
A 
III 
Macrolides52 
inhibition 
B 
IIa 
Tetracycline60*,90 
Inhibition 
No inhibition 
B 
A 
IIa 
III 
Statins19,39 
inhibition 
B 
IIb 
ACE inhibitors24 
No inhibition 
B and C 
IIb 
AR blockers42 
Animal data 
C 
IIb 
 
*Inhibition at 6 and 12 months following 3 months of treatment 
Level A = Large randomized clinical trials. Level B = Small randomized clinical trials. Level C 
= Observational studies and expert opinion. Class III = Ineffective or harmful, recommended 
against. Class IIa = Possibly efficacious. Class IIb = Unknown efficacy. 
Potential impact of pharmacotherapy on aneurysm expansion. While the natural history 
of AAAs is one of progressive expansion, the rate of aneurysm growth and the risk of rupture are 
both exponentially related to aneurysm size. Aortic diameter is therefore the single best predictor 
of rupture risk.1,2 Ninety-five percent of aortic aneurysms occur in the segment below the renal 
arteries; the normal aortic diameter at this level is 2.0-2.5 cm and risk of aneurysm rupture is low 
until aortic diameter increases beyond 5.5 cm.4,5 A small 3.5 cm AAA growing at an average rate 
(7%/yr.) will reach 5.5 cm in 7 years. Ideally, a drug might be found that would completely 
arrest growth so that repair would never be required. More realistically, drug therapy may prove 
to be less than 100% effective. The proposed trial is designed to test the hypothesis that 
doxycycline will reduce the rate of AAA expansion by 40%. This threshold was chosen for 
several reasons. Physicians (internists, family practitioners and surgeons) were surveyed and 
asked to indicate a threshold for AAA growth inhibition that they would consider clinically 
useful. They reported that they would prescribe such a medication only if it exhibited at least 30-
50% efficacy in reducing growth. They recognized that inhibiting AAA expansion to some lesser 
degree would not obviate the need for intervention but simply postpone it to a time when the 
individual will be older and, perhaps, less fit. A 40% reduction results in a clinically significant 


 
 
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14 
 
delay (≥ 4 yr.) in the median time it would take for a 3.5 cm AAA to reach 5.5 cm. Using the 
proposed study design, we have a 90% chance to detect a 40% decrease in AAA growth.  
An aneurysm detected at 3.5 cm and growing at an average rate cited in earlier studies 
(0.3 cm/year) will reach the 5.5 cm (a recommended threshold for intervention) in 7 years 
(Fig.1). We have conducted a Monte Carlo simulation study to project the effect on use of 
surgical procedures to treat small abdominal aortic aneurysms among patients whose presenting 
aneurysm diameters and ages have the same distributions as those of patients screened in our 
clinical sites. This simulation study is based on a conservative anticipated growth rate of 2.5 
mm/year and age-stratified mortality of the U.S. male population from 2006. Time to surgery is 
presented in Figure 2, and counts of patient outcomes out of 60,000 screened for an untreated 
group and a group treated with an agent that reduces growth by 40% (to 1.5 mm/year) are 
presented in the accompanying table. 
The additional time it would take a small 3.5 cm AAA to reach 5.5 cm is 4 years 
(median), for 40% inhibition of normal growth. A reduction in growth rate of 40% would have 
impact because, excluding patients who are found on screening to have aneurysms larger than 
5.5 cm, the 40% reduction in growth would be associated with an increase in the proportion of 
small abdominal aortic aneurysm patients who never undergo surgical intervention from 22% 
(10,311 ÷ (35,929 + 10,311)) to 35% (16,176 ÷ (30,064 + 16,176)), a 59% relative increase. 
Establishing a 40% reduction in growth rate would clearly demonstrate the biological effect of 
inhibition and have a monetary value of about $0.5 billion/year in reduced surgical interventions 
in the U.S. where approximately 1.5 million men and 1.5 million women will reach age 65 
annually over the next decade, and about 8% of men and 2% of women will have small 
aneurysms (19,500 surgical interventions prevented per year x $30,000/intervention = $5.85 
hundred million). Growth inhibition of 50% or more as observed by Mosorin would have a larger 
impact because as the efficacy of inhibition increases, the percent of patients who will die of 
other causes before reaching 5.5 cm is further increased.60 Also, because of comorbid 
cardiovascular disease and other conditions, the mortality rate from all causes in patients with 
AAA is likely to be larger than the rate used in our conservative projections. A 30% growth rate 
reduction would demonstrate a biological effect but have much less clinical impact. Again, the 
proposed trial is designed to give us a 90% chance to detect a 40% decrease in AAA growth rate 
at the 95% confidence level.  
New insights into the pathobiology of aortic aneurysm. It is of interest to note that all 
current treatment approaches to aortic aneurysms are based upon a "mechanical" concept of the 
disease (i.e. segmental graft repair of the diseased aorta), yet investigations emerging over the 
past several years have emphasized the complex cellular and molecular nature of this disorder. 
These studies have indicated that with greater knowledge and a conceptual shift in focus, aortic 
aneurysms might also be amenable to alternative, "mechanism-based" treatment strategies. . In 
seeking molecular targets by which to control the progression of aneurysm growth, an 
accumulation of studies has led to the belief that pharmacologic strategies to inhibit matrix 
metalloproteinases (MMPs) are now a feasible means to suppress the progression of aneurysmal 
degeneration. Although the pathophysiologic events underlying the initial development of AAA 
are still incompletely understood, it is clear that the progression of aneurysmal degeneration 
involves destructive remodeling of aortic wall connective tissue. Recent studies have implicated 
three processes in this pathologic pattern of remodeling: chronic mononuclear inflammation, 
progressive destruction of structural matrix proteins associated with excessive local production 
of matrix-degrading proteinases, and impaired connective tissue repair. 
 


 
 
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15 
 
Figure 2. Product-limit estimates from two growth rates: X axis: years 
 
Growth rate=0.25 cm per year 
 
Growth rate=0.15 cm per year 
 
Excludes patients whose AAA diameter≥5.5cm at screening. 
 
 
Number of Surgeries that Could Be Prevented in a Cohort of 60,000 Patients 
Follow-Up Interval=6 months 
Stratum 
Total No. of 
simulated 
patients 
No. of screened 
patients whose 
diameter were  
larger than 5.5 
cm at baseline 
(time of the 
referral) 
No. of patients 
whose diameter 
grows ≥5.5 cm 
during follow-up 
No. of patients, 
never operated, 
dying of causes 
other than 
aneurysm 
rupture* 
Growth rate=1.5 
cm per year 
60,000 
13,760 
30,064 
16,176 
Growth rate=2.5 
cm per year 
60,000 
13,760 
35,929 
10,311 
Change 
 
 
-16.3% 
+56.9% 
* Mortality determined according to U.S. general population age and gender rates. 
 
Matrix-degrading Proteinases. Through numerous studies conducted over the past 
decade, a pathophysiologic concept has emerged that proteolytic degradation of medial elastin is 
responsible for weakening and dilatation of the aortic wall, and that collagen degradation is 
responsible for aneurysm rupture.55,61-63 Because elastin is one of the most durable structural 
proteins of the extracellular matrix, the dissolution of elastic fibers requires the presence of 
0 . 0 0
0 . 2 5
0 . 5 0
0 . 7 5
1 . 0 0
t i me
0
5
1 0
1 5
2 0
2 5
3 0
ST RAT A:
Gr o wt h = 0 . 1 5
Ce n s o r e d  Gr o wt h = 0 . 1 5
Gr o wt h = 0 . 2 5
Ce n s o r e d  Gr o wt h = 0 . 2 5


 
 
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16 
 
specific proteinases. Abundant evidence now suggests that the elastin and collagen degradation 
in aneurysm tissue is mediated in large part by members of the MMP family, including 
collagenase-1 (MMP-1), stromelysin-1 (MMP-3), the 72-kDa and 92-kDa gelatinases (MMP-2 
and MMP-9, respectively), macrophage elastase (MMP-12), and collagenase-3 (MMP-13).62-67 
MMPs constitute a large family of structurally-related metalloendopeptidases that are 
collectively capable of degrading all components of the extracellular matrix. They play important 
roles in normal tissue development and remodeling, particularly during embryonic growth, 
uterine involution and wound healing. Abnormal expression of MMPs also contributes to a 
variety of pathological processes, including osteoarthritis and rheumatoid arthritis, tumor 
invasion and metastasis, pulmonary emphysema and atherosclerosis.68-72 Local regulation of 
MMP activities is critical to prevent widespread tissue destruction during normal remodeling and 
in disease. MMPs are controlled at several different levels, including transcriptional induction 
and suppression of MMP genes, extracellular processes required for proenzyme activation, and 
interactions with naturally-occurring MMP inhibitors. The activation of the proMMPs is thought 
to be mediated by other MMPs, serine proteases, or oxidative processes. Physiological inhibitors 
of MMP activity include plasma-derived alpha2-macroglobulin and at least four specific tissue 
inhibitors of metalloproteinases (TIMPs). 
Of the MMPs implicated in human aneurysm disease, MMP-2, MMP-9, and MMP-12 
have attracted particular interest because they have activity against insoluble elastin fibers.62,63,73 
MMP-9 is the most abundant elastolytic proteinase produced by human AAA tissues in vitro, it 
is expressed in situ by aneurysm-infiltrating macrophages located at the site of tissue damage, 
and it is synthesized in AAA tissue in a manner correlating with increased aneurysm 
diameter.55,73 Whereas MMP-2 is found in close association with the extracellular matrix in 
aneurysm tissues, MMP-9 is elevated in the circulating plasma of patients with AAAs.63,74 
MMP-12 is also thought to play a critical role in aneurysm development, because it is selectively 
expressed by macrophages within the elastic media of AAA tissue and it is specifically localized 
to elastin fiber fragments by immunohistochemistry.62,66 These observations have fostered the 
notion that MMP-2, MMP-9 and MMP-12 provide useful biological markers of aortic aneurysm 
disease, and perhaps more importantly, potential targets for pharmacologic therapy. 
Use of tetracyclines as MMP inhibitors. During studies on the mechanisms of 
periodontitis in diabetic rats, Golub and colleagues made the discovery that tetracyclines have 
substantial metalloproteinase-inhibiting effects.75 In subsequent studies they suggested that this 
inhibition took place through a mechanism similar to that found with the endogenous tissue 
inhibitors of metalloproteinases.76 Non-antibiotic chemically-modified tetracyclines (CMTs) 
have a similar efficacy as MMP inhibitors, demonstrating that the MMP-inhibiting property of 
tetracyclines is unrelated to their antimicrobial activity.54 Tetracyclines prevent matrix 
degradation in many animal models of disease, and because of their safety profile, they have 
been successfully tested in several conditions associated with elevated MMP activity and 
connective tissue destruction.  Furthermore, it has now been clearly demonstrated that 
doxycycline can effectively penetrate and suppress MMPs in the complex tissue environment of 
degenerative human aortic aneurysms.58,77 
 
 
 
 
 


 
 
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17 
 
Drug 
Medical Condition 
Comments 
Doxycycline Atherosclerotic Carotid 
Plaques 
Doxycycline decreases MMP expression in carotid 
plaque removed during endarterectomy.78 
Minocycline 
Rheumatoid Arthritis (RA) 
Trial 1 
Minocycline improved laboratory and clinical 
parameters in RA.79 
 
RA Trial 2 
Improved laboratory and clinical parameters in RA.80 
 
RA Trial 3 
Minocycline superior to placebo in clinical 
parameters.81 
 
RA Trial 4 
On 2 year follow-up minocycline treatment resulted 
in improved outcomes compared to standard 
therapy.82 
Minocycline 
Acute Stroke 
Minocycline treatment was associated with a better 
stroke score days 7 and 30.83 
Tetracycline 
RA 
Tetracycline and clindamycin improved outcomes in 
refractory RA.84 
 
Why doxycycline is the best choice for a clinical trial. Following modification of 
chlortetracycline, which was discovered in 1944, a number of useful antibiotics including 
tetracycline were created. Because of the relatively short half-life of these compounds, two new 
compounds with increased liposolubility were created (doxycycline and minocycline). These 
compounds exhibited rapid intestinal absorption, longer half-life, superior tissue penetration and 
reduced toxicity. Doxycycline is almost completely absorbed from the upper GI tract (95%) 
which prevented many of the lower GI tract side effects associated with the earlier 
tetracyclines.85 Since doxycycline is 80-95% protein bound and is lipid soluble, blood levels are 
sustained allowing it to be administered on a 12-24 hour dosing regimen. Only 30-40% of 
doxycycline is eliminated by renal mechanisms. The majority is eliminated by hepatic and 
intestinal clearance and this path of excretion is enhanced with renal failure. Thus, no 
accumulation occurs even when patients are anuric. All of these features of doxycycline make 
appealing for a trial involving elderly patients with AAA. Minocycline shares many of the 
favorable pharmacologic features of doxycycline including a high percentage of absorption in the 
proximal GI tract, a long half-life and combined GI and renal excretion.77 In fact, the half-life of 
minocycline is longer than doxycycline so that it could be administered once daily. A major 
drawback to the chronic administration of minocycline has been identified in trial of rheumatoid 
arthritis where it has been found to cause greenish discoloration of the skin, a problem which is 
usually but not always reversible.86 
Of the other potential candidate drugs that may impact AAA, propranolol has been 
eliminated by negative prospective studies. As discussed above, a majority of AAA patients 
(67% of eligible patients surveyed at the clinical sites for our trial) will be on statins for other 
reasons. Because of the proven benefits of statins in preventing cardiac events, a randomized trial 
of statins in AAA would be unethical since it would require withdrawal of statin therapy.  
Although macrolide antibiotics show promise, enthusiasm was diminished by the negative results 
of trials in coronary disease.52 Furthermore, these drugs are first and second line therapy as 
antibiotics so that their long term use in such a trial raises concerns about promoting the 
development and growth of antibiotic resistance. Because of past widespread use of tetracyclines 


 
 
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18 
 
in humans, they are relegated to the treatment of a very limited number of infections. Testing 
other antihypertensives such as ACE inhibitors and AARB inhibitors is extremely complex in a 
patient population where hypertension is common and specific indications exist for the use of 
each class of antihypertensive medications. And, while there are a number of promising 
proteinase inhibitors in development, there will be some period of time before these drugs are 
proven safe, especially with regard to potential for inducing fibrosis. If, however, doxycycline 
provides the initial proof of concept that medical intervention can slow aneurysm progression, 
the search for more effective agents can proceed in ernest. 
Clinical evidence that doxcycline will inhibit AAA. The first randomized, placebo 
controlled trial using doxycycline to inhibit AAA expansion was published in October 2001.60 A 
total of 32 patients were randomized with 17 receiving doxycycline (200 mg/day) for 3 months. 
While there was no decrease in the overall expansion during the trial, the interval analysis at 6-
12 months and 12-18 months indicated significantly less expansion in the doxycycline treated 
patients. While an accompanying editorial pointed out the weaknesses related to the small 
sample size especially as this relates to the use of multiple interim analyses, this trial provides 
the first clinical evidence suggesting that doxycycline could inhibit aneurysm expansion. This 
information is particularly important in the context of the proposed clinical trial given that some 
of the preliminary data supporting the trial has been generated in animal models that cannot 
precisely recapitulate chronic human diseases. It also highlights the importance of moving 
forward with a large properly controlled trial. Unfortunately, many clinicians will not identify 
the inadequacy of such studies. In the absence of other medical options for AAA and the 
relatively low incidence of side-effects, the data from small trials such as this will lead to 
increased use of doxycycline in AAA patients without adequate proof of efficacy. With 
dissemination of information from these small trials, it may become increasingly difficult to get 
patients to consent to a trial where they have a 50% chance of receiving placebo. 
Two recent articles emphasize the diverse and potentially beneficial effects of doxycycline on 
aneurysm tissue.77,87 Both papers are based on data from a study of 60 patients scheduled for 
open abdominal aortic aneurysm repair who were randomly assigned to treatment with one of 3 
daily doses of doxycycline (50, 100 or 150 mg daily) or placebo. Treatment was begun 2 weeks 
prior to the planned surgical procedure, with aortic tissue harvested at the time of procedure. As 
in prior studies, a reduction in the total MMP-9 protein was seen in the aortas taken from 
individuals treated with doxycycline.58 In addition, the expression of MMP-3 and MMP-25 were 
significantly reduced, and there were significant increases in protease inhibitors, Cystatin C and 
TIMP-1, particularly at higher doses of doxycycline. In addition, there appeared to be a dose 
dependent decrease in the activation of MMP-8 and MMP-9 extracted from the tissue. 
The neutrophil content of the aortic wall was found to be significantly reduced as well as the 
numbers of CD8+ T-lymphocytes, but not other inflammatory cell types. They also found 
significant reductions in tissue levels of Interleukin (IL)-6 and IL-8, as well as consistent effects 
on the upstream regulators of these interleukins including AP-1 activation and C/EBP 
expression. Although some effects of doxycycline appeared to be dose independent, others, 
particularly the effects on aortic MMP activity, appeared to be greater at the higher doses of the 
drug.   
These new findings in human tissue further suggest that doxycycline treatment may have 
multiple beneficial effects on the progression of aneurysm disease. The reduction of aortic 
neutrophils is a particularly encouraging effect. While there has been some indirect evidence that 
neutrophils may play an important role in aneurysm development, several recent publications 


 
 
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19 
 
have now clearly demonstrated that neutrophil activity has an important role in an animal model 
of aneurysm disease.88,89  
These new findings support the rationale for N-TA3CT based on biologic effects of 
doxycycline. These new dose ranging studies, combined with our prior Phase II safety studies, 
also provide some additional assurance that the dosing choice of 100 mg twice daily is the most 
appropriate to develop a maximal biologic effect with minimal side effects. 
1.2 
PRELIMINARY STUDIES 
Preliminary studies supporting this application  demonstrate that doxycycline can inhibit 
aneurysm development and expansion in two distinct experimental models of AAA through the 
inhibition of MMPs. While we understand that information obtained from such models cannot 
precisely recapitulate the chronic disease we see in patients, we present additional lines of 
evidence suggesting that this experimental work is relevant: 1) we show that serum 
concentrations of doxycycline that inhibit experimental aneurysm formation can be achieved in 
AAA patients with relatively few side-effects; 2) we demonstrate that doxycycline, at the dose 
proposed for this trial (200 mg/day), inhibits MMP production in human AAA tissue. In 2011, 
however, the Pharmaceutical Aneurysm Stabilization Trial (PHAST) was completed in the 
Netherlands. This randomized, placebo-controlled, clinical trial of doxycycline (100 mg daily) 
for the treatment of abdominal aortic aneurysms larger than 3.5 cm on ultrasound (including 
aneurysms larger than 5.5 cm) was registered on the Netherlands Trial Register 
(http://www.trialregister.nl/trialreg/admin/rctview.asp?TC=1345). PHAST reported a small 
increase in the rate of growth of aneurysms associated with doxycycline but did not observe an 
increase in adverse aneurysm outcomes associated with doxycycline. 90 This information 
combined with the data cited above from the Mosorin clinical trial showing decreased AAA 
expansion with doxycycline, demonstrate the need for a large, well-controlled trial to definitively 
answer this important question. 
Pilot Study: Doxycycline treatment in patients with small asymptomatic AAA. Despite 
the promising preliminary data suggesting that doxycycline would inhibit the growth of aortic 
aneurysms and the long record of safety with the clinical use of doxycycline, several critical 
questions had to be answered before a larger and more definitive trial could be designed. The 
pilot trial addressed three important questions; 1) How would high dose doxycycline be tolerated 
and what would the level of compliance be in the elderly AAA patient population? 2) What 
would the plasma concentrations of doxycycline be in patients taking high dose doxycycline and 
how would these levels compare to the therapeutic levels required in animal models? 3) What 
effect would doxycycline have on plasma MMP-9 levels? Our attempts to recapitulate human 
disease in acute animal models cannot be entirely successful since we cannot mimic the chronic 
atherosclerotic features of AAA. We have previously shown that AAA is a dynamic remodeling 
process. Based on one known effect of doxycycline, its ability to decrease MMP levels, we 
expect that it will inhibit aneurysm growth. Doxycycline has other properties such as its ability to 
inhibit proliferation and induce apoptosis in some cell lines. These actions may not be beneficial 
in a complex remodeling process such as AAA. Thus, a final goal of the pilot trial was to be sure 
that doxycycline was safe and did not cause an unexpected increase in aneurysm growth. 
Because our primary endpoint was not aneurysm growth rate, we were more lenient with 
regard to initial aneurysm size and imaging modality. Among the 36 patients enrolled and 
followed for six months, the initial range of aneurysm diameter was 3.3 to 5.5 cm.57 Patients 
were recruited from the practices of the trial investigators. All patients were placed on high-dose 
(100 mg p.o. b.i.d.) doxycycline for six months. Since there was no funding for the imaging 


 
 
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20 
 
studies, we allowed that patients could be studied using CT scan or ultrasound as long as the 
initial and final studies were done using the same technique. IRB approval was obtained and 
patients were enrolled from six of the centers that will be participating in the larger trial.  
Patient enrollment began with review of the imaging study and recording of greatest 
transverse aneurysm diameter. The compatibility of the patient’s current medications with 
doxycycline was analyzed (Drug-reax Micromedex Health Care Series, Englewood, CO) and 
patients were questioned about allergies to tetracyclines. The Euroqual health survey, physical 
examination and history were obtained followed by a venipuncture for plasma (EDTA) MMP-9 
and doxycycline levels. The plasma samples were frozen for batched analysis. The patients were 
given a three-month supply of medication and were told to take 100 mg b.i.d with food. They 
were asked to keep and return the medication that they did not take at their three-month follow 
up. At the first follow-up, patients were questioned about side effects, onset of new medical 
problems or exacerbation of existing medical problems. Blood was drawn for plasma MMP-9 
and doxycycline levels. They were then given the final three months supply of medication. At six 
months, the Euroqual health survey, a physical examination and history were obtained followed 
by a venipuncture for plasma MMP-9 and doxycycline levels. The medication was counted as a 
measure of compliance. An imaging study (CT scan or ultrasound) was obtained. The trial was 
complete on June 1, 2000.  
Of the 36 patients initially enrolled, three chose to discontinue the study before the end of 
the 6-month period of follow-up, for a retention rate of 92%. The patients who chose to withdraw 
cited tetracycline-induced photosensitivity (n = 1) and discoloration of teeth (n = 1) as the 
principal reasons for dropping out, while the third patient cited new concerns about being 
involved in clinical research based on an article in a weekly newsmagazine. These problems 
arose between months 2-3 in two of the patients and between months 3-4 in the other.  
With regard to other adverse events related to doxycycline treatment, two additional 
patients reported one or more episodes of tetracycline-induced photosensitivity. In each case, 
these episodes were controlled by limiting sun exposure and did not require withdrawal from the 
study. While a second patient also reported mild discoloration of the teeth, she chose to continue 
in the trial. One patient suffered an acute myocardial infarction 2 months after starting the study, 
but chose to continue on doxycycline treatment through the duration of the trial. By patient self-
report and assessment of medication counts at each follow-up visit, a high rate (94% of 
medication taken) of compliance with doxycycline treatment was observed throughout the 6-
month period of the pilot study. This was confirmed by plasma doxycycline levels. The mean 
value of 4.62 +/-0.67 g/ml (range 1.31-14 g/ml) was obtained. 
Patient characteristics 
Number 
Age 
Men/Women 
Caucasian 
Black 
Hispanic 
36 
69+/-1.9 yrs 
30/6 
35 
1 
0 
   
We originally planned to enter 30 patients in the pilot study, but were inundated with 
requests by patients to participate. We allowed six additional patients to enroll. Investigators at 
several of the sites also chose to put patients on doxycycline outside the trial because of the large 
number of requests to participate after patients heard about the trial. This enthusiasm for 
enrolling in a trial reflects the anxiety of patients with AAA the fact that there are no other 
medical therapies available.  


 
 
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Plasma MMP-9 levels were measured before treatment and at the 3- and 6-month interval 
evaluations. The mean plasma MMP-9 level before treatment was 140.91 ± 45.42 g/ml, a level 
somewhat higher than that measured in our previous analysis of patients with AAAs undergoing 
surgical treatment. This may reflect an even greater degree of connective tissue remodeling in 
small AAAs as compared to those already large enough to require surgical repair. Importantly, 
MMP-9 levels decreased during a short course of treatment with doxycycline, with a mean 
decrement of 64.1% ± 8.3% from the pre-treatment baseline. This difference did not achieve 
significance at three months but was significant at 6 months compared to baseline. There was no 
apparent correlation between plasma MMP-9 concentrations and doxycycline levels. The 
observations support the concept that serial measurements of plasma MMP-9 may be of use in 
evaluating the response of individual patients to doxycycline treatment. The effects of treatment 
with doxycycline on plasma MMP-9 levels are comparable to successful surgical repair and 
substantially greater than those seen with incomplete endovascular aneurysm exclusion. The 
larger trial proposed will provide much greater information regarding the relationship between 
doxycycline levels, MMP-9 levels and inhibition of aneurysm expansion. 
Effects of doxycycline on experimental AAA. There has been longstanding interest in the 
feasibility of MMP inhibition as a strategy for the suppression of aneurysmal degeneration. To 
examine this possibility, Petrinec et al. characterized the development of AAA following 
elastase-induced injury to the rat aortic wall.53 Under the proper experimental conditions, there is 
only minimal structural damage to the medial elastic lamellae immediately after elastase 
perfusion and the aorta only dilates about 30-50 percent over normal. Aortic wall structure and 
diameter also remain stable for several days after elastase perfusion, yet the damaged aorta 
begins to progressively expand thereafter, enlarging to aneurysmal proportions (> 100 percent of 
normal diameter) within 7 to 14 days. Importantly, the delayed onset of aortic dilatation is 
temporally and spatially associated with aortic wall infiltration by mononuclear phagocytes, 
increased local expression of elastolytic metalloproteinases (including MMP-2 and MMP-9), and 
pronounced destruction of the medial elastic lamellae. The aortic wall response to elastase 
perfusion therefore recapitulates many of the morphologic and biochemical events evident by 
histology and biochemical assays of human AAA, encouraging the use of this model for 
pathophysiologic and pharmacologic investigations. 
Petrinec et al. used doxycycline to suppress aortic wall MMP activity, elastin 
degradation, and aneurysm development in the elastase-induced rat model.53 Whereas treatment 
with doxycycline (60 mg/kg/day) reduced the extent of aortic dilatation by about 60%, this effect 
was subsequently shown to be dose-dependent with half-maximal suppression at clinically-
achievable, but relatively high doses (6 mg/kg/day).54 They achieved similar results using non-
antimicrobial (chemically-modified) tetracyclines and nonselective hydroxamic acid derivatives 
as MMP inhibitors, indicating that the aneurysm-suppressing effects of doxycycline are most 
likely related to its activity as an MMP inhibitor.54 These and other studies suggested that 
tetracyclines might have significant advantages as a clinically-applicable strategy for achieving 
MMP inhibition in patients with AAAs. 
More recent studies have focused on whether treatment with doxycycline might also 
influence the growth of established aortic aneurysms. To address this question, a series of rats 
underwent elastase perfusion. In a subset of animals sacrificed at day 7, the mean increase in AD 
was 111% ± 8%, indicating that aneurysms had largely developed by this interval. The remaining 
animals were then provided either normal drinking water or water supplemented with 


 
 
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22 
 
doxycycline (30 mg/kg/day) and sacrificed on day 14 or 21. Animals treated with doxycycline 
exhibited a marked attenuation of the late progression of aneurysmal dilatation.53  
AAA in the mouse. To further examine if MMPs might play a functionally significant 
role in the development of experimental AAAs, Petrinec et al. developed and characterized an 
elastase-induced model of AAAs in the mouse.53 In parallel, Longo et al. has characterized a 
second murine aneurysm model using calcium chloride applied to the ablumenal surface to 
induce the aneurysm.55 This injury produces an inflammatory response leading to increased 
MMP expression, matrix degradation, and aortic dilatation. As in previous studies in the rat, 
doxycycline was used as a non-selective MMP inhibitor. In mice treated with doxycycline 
beginning the day after elastase perfusion, there was a significant reduction in mean diameter on 
day 14 compared to untreated controls. The incidence of AAAs in individual animals was also 
significantly reduced by doxycycline treatment, from 91% to 50%, associated with preservation 
of the elastic media.54 Using the CaCl model, doxycycline demonstrates the same dose-
dependent inhibition of aneurysm expansion.56 The plasma doxycycline levels achieved in these 
animal studies (see figure below) were in the same range as those seen in AAA patients receiving 
high dose (100 mg bid) doxycycline (patient levels mean 4.62 ug/ml, SE 0.67, range 1.31-14). 
These murine studies suggest that inhibition can still be achieved at plasma levels in the 1-2 
ug/ml range.  
Suppression of aortic aneurysms by targeted gene disruption of MMP-9. Based on 
previous studies suggesting that MMP-9 might play a particularly important role in the 
pathophysiology of aortic aneurysms, one of the important goals of these studies was to discern 
if this enzyme is required for the development of experimental AAAs. The response to elastase 
perfusion was therefore examined in mice with targeted disruption of the MMP-9 gene.62 While 
functional abnormalities in MMP-9 (-/-) mice have been described, these animals do not exhibit 
spontaneous cardiovascular abnormalities. Experiments with these animals revealed that 
elastase-induced aneurysmal dilatation is substantially reduced in MMP-9 (-/-) mice, with an 
increase in aortic diameter of only 87 ± 10% on day 14 compared to 132 ± 8% in the 129/SvEv 
background controls (P < 0.05).17 AAAs developed in only 40% of the MMP-9 (-/-) animals 
compared to 94% in wild type controls (P < 0.05). The suppression of aneurysmal dilatation 
achieved by genetic deficiency in MMP-9 was similar to that achieved in wild-type mice treated 
with doxycycline. These studies demonstrate that expression of MMP-9 is required in the process 
of elastase-induced aneurysmal degeneration and that treatment with doxycycline can reduce 
aneurysm development to an equivalent extent in this model of AAAs. Using the CaCl murine 
model of AAAs, Dr. Baxter's laboratory has independently corroborated the effects of MMP-9 
deficiency on aneurysm development. Their studies also demonstrated that there is some degree 
of aneurysm suppression in MMP-12 (-/-) mice.66  


 
 
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23 
 
 
 
Broad protease inhibitory effects of doxycycline. While it is clear that doxycycline 
inhibits MMP-9, there is evidence of much broader MMP inhibition. We have demonstrated that 
doxycycline inhibits MMP-2, another key MMP in AAA. Using gene array studies with cultured 
macrophages, we have recently identified spectrum of MMPs and MMP promoting pathways 
impacted by doxycycline.(unpublished observations) 
Treatment with doxycycline reduces MMP expression and activation in human AAA 
tissue. To begin translating these experimental observations to patients with AAAs and to further 
examine the mechanisms by which doxycycline might act in vivo, 15 patients scheduled to 
undergo elective AAA repair were examined.58 Eight of these patients were treated with oral 
doxycycline (100 mg bid) for one week prior to operation, while the others served as untreated 
but contemporaneous controls. Aneurysm tissues excised at operation were used for analysis of 
MMP protein (gelatin zymography and immunoblot analysis) and mRNA expression 
(competitive RT-PCR). The two groups were indistinguishable with respect to age, gender and 
AAA size, but there was a 2.5-fold reduction in extractable MMP-9 protein in the doxycycline-
treated patients vs. controls (P < 0.05). See Figure below. It was also found that aortic wall 
expression of MMP-9 mRNA was reduced by 5.5-fold in the doxycycline-treated group (P < 
0.05) (not shown). Although there was no significant difference in the amount of extractable 
MMP-2 protein or mRNA, there was a reduction in the activated fraction of MMP-2 in the 
doxycycline-treated patients (not shown). These studies indicate that even brief treatment with 
doxycycline can effectively reduce MMP-9 production in human aortic tissues from patients with 
AAAs. They also indicate that the effects of doxycycline occur through a combination of 


 
 
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24 
 
favorable mechanisms (i.e., a reduction in MMP expression and activation, as well as direct 
pharmacological inhibition of enzyme activities), suggesting that doxycycline might be a 
particularly useful agent with which to inhibit MMPs in patients with AAAs. 
Plasma MMP-9 levels in patients with AAA. To assess biological events more directly 
involved in the enzymatic degradation of matrix proteins, McMillan et al. examined the 
production of MMP-9 in AAA tissue and its presence in the circulation.74 Their initial studies 
involved 22 patients with AAA, 9 patients with aortoiliac occlusive disease (AOD), and 8 normal 
controls. Using a sensitive enzyme-linked immunosorbent assay (ELISA) to measure MMP-9 in 
peripheral venous blood plasma samples, they found that plasma MMP-9 was significantly 
higher in patients with AAA (85.66 ng/ml ± 11.64; mean ± SEM) than in AOD (25.75 ng/ml ± 
4.16; p < 0.001) or normal controls (13.16 ng/ml ± 1.94; p < 0.001); the difference between 
AOD and normal controls was not significant. Interestingly, patients with multiple aneurysms 
had significantly higher plasma MMP-9 levels than patients with an isolated AAA (134.68 ng/ml 
± 17.5 versus 71.03 ng/ml ± 10.7; p < 0.04). The production of MMP-9 was also significantly 
increased in organ cultures of AAA and AOD compared to normal aorta, suggesting that the 
diseased aortic tissue was the source of the elevated circulating MMP-9. These intriguing results 
provided the first evidence that circulating MMP-9 levels might provide a sensitive biomarker of 
aneurysm disease, a notion confirmed and extended by the preliminary studies described below. 
Thus, one of the aims of the proposed study will be to examine prospectively if circulating levels 
of MMP-9 might correlate with aneurysm growth rates or therapeutic effects in suppressing 
aneurysm growth. The relevant biologic samples will be obtained during the course of the study. 
These data will be valuable in the future as potential measures by which to monitor therapeutic 
efficacy during pharmacological treatment, especially if changes in MMP-9 levels precede 
changes in the rate of aneurysm growth. 
Following the demonstration that plasma levels of MMP-9 are elevated in patients with 
AAAs, Hovsepian et al. sought to extend these observations.91 Peripheral venous blood was 
sampled in 25 patients with AAAs (mean age 66.6 ± 3.9 yrs), 15 patients with aortoiliac 
occlusive disease (AOD; mean age 65.4 ± 3.9 yrs), and 5 normal healthy controls (mean age 31.2 
± 2.2 yrs). Additionally, the patients with AAA had arterial blood samples taken above and 
below the aneurysm. Plasma MMP-9 levels were again determined using an ELISA kit. In this 
study, the mean (±SE) plasma MMP-9 concentration was 36.10 ± 7.71 ng/ml in 5 normal 
controls, 54.71 ± 10.45 ng/ml in 15 patients with AOD (P=NS vs. normal), and 99.38 ± 17.38 
ng/ml in 25 patients with AAAs (P<.05 vs. normal and AOD). With an upper limit of normal 
defined as 87.8 ng/ml (mean + 3 SDs), elevated levels of plasma MMP-9 were found in only 
1/15 (6.7%) of patients with AOD, but in 12/25 (48%) with AAAs (P<.05). The positive 
predictive value of an elevated plasma MMP-9 level was 92.3%. Non-elevated plasma MMP-9 
levels were an unreliable indicator of the absence of AAA, with a negative predictive value of 
59.4%. Neither age, gender, nor aneurysm diameter correlated with plasma MMP-9 levels, 
although the highest values were observed in patients with large AAAs. The results of this study 
parallel those reported by McMillan et al.74 suggest that plasma levels of MMP-9 might be a 
useful marker for assessing the success of medical or surgical intervention for AAA. 
Plasma MMP-9 levels are decreased by successful surgical repair. Based on the 
hypothesis that plasma MMP-9 levels reflect the biological events occurring within the 
aneurysmal aorta, it would be predicted that elevated levels of plasma MMP-9 should fall 
considerably after aneurysm repair. A newer and less invasive technique of aneurysm repair 
involves placing an expandable stent graft (endografting) within an aortic aneurysm using a 


 
 
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25 
 
femoral artery approach. Occasionally, this technique fails to exclude blood flow into the 
aneurysm sac (endoleak). We also considered whether the presence or absence of such a 
decrease in plasma MMP-9 levels might be useful as a functional biomarker if an endovascular 
repair were not successful. To test this possibility, plasma MMP-9 levels were measured in a 
series of patients scheduled to undergo elective treatment for infrarenal AAAs.92 Eleven patients 
were identified who had elevated plasma MMP-9 levels prior to aneurysm repair (PreOp). 
Postoperative (PostOp) plasma MMP-9 levels were re-measured 3 to 10 months after open repair 
(OR; n = 6), endovascular repair without endoleak (ER/NoL; n = 3), or endovascular repair with 
endoleak (ER/wL; n = 2). Data were expressed as the mean ±SEM for each group and compared 
by the Student's t-test (PreOp vs. PostOp). The percent decrease in plasma MMP-9 following 
treatment was also determined for each patient and correlated with the success or failure of repair 
at 6 and 12-month follow-up intervals. 
  
 
The upper panel is a Western blot showing the MMP-9 levels in aortic tissue from patients 
receiving doxycycline and a control group that did not receive oral doxycycline prior to operation 
and several normal aortas from transplant donors. The lower panel shows the densitometric 
quantification of these studies.13 
 


 
 
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26 
 
Prior to aneurysm repair, the mean plasma MMP-9 concentration for all patients was 
290.19 ± 45.46 ng/ml (compared to 87.8 ng/ml as the upper limit of normal in our laboratory) 
with no significant difference between groups (open repair, 328.05 ± 73.85 ng/ml; all 
endovascular repair, 244.77 ± 47.31 ng/ml; endovascular repair with no leak, 253.54 ± 76.70 
ng/ml; endovascular repair with leak, 231.66 ± 66.73 ng/ml). Plasma MMP-9 levels decreased in 
all 11 patients following AAA repair, reaching an overall mean of 48.41 ± 19.35 ng/ml (P < 
0.001, preoperative vs. postoperative). The mean postoperative plasma MMP-9 level was 36.99 
± 23.45 ng/ml in the 6 patients who had undergone open repair (P < 0.01 vs. preoperative) and 
62.11 ± 33.90 ng/ml in the 5 patients who had undergone endovascular repair (P < 0.05 vs. 
preoperative). In the endovascular repair group, postoperative plasma MMP-9 levels were 7.08 ± 
3.70 ng/ml for 3 patients without endoleak (P < 0.05 vs. preoperative) and 144.66 ± 9.69 ng/ml 
for 2 patients with endoleaks (P = NS vs. preoperative). These preliminary observations support 
the hypothesis that the aneurysm is the source of plasma MMP-9. In addition, they suggest that 
serial measurements of plasma MMP-9 concentrations can be used to follow the effects of 
medical treatment of AAA. We demonstrate below that we did observe a decrease in plasma 
MMP-9 levels in AAA patients taking doxycycline in 6 month pilot trial performed by the 
investigators involved in this proposed trial. We expect this to be an important biomarker in the 
trial. 
Summary of background and preliminary data. It is of interest to note that all current 
treatment approaches to aortic aneurysms are based upon a "mechanical” concept of the disease 
(i.e., segmental graft repair of the diseased aorta), yet investigations emerging over the past 
decade have emphasized the cellular and molecular nature of this disorder. These studies have 
indicated that with greater knowledge and a conceptual shift in focus, aortic aneurysms might 
also be amenable to alternative "mechanism-based" treatment strategies. In seeking molecular 
targets by which to control the progression of aneurysm growth, accumulating information leads 
to the conclusion that pharmacologic strategies to inhibit matrix metalloproteinases (MMPs) are 
now a feasible means to suppress the progression of aneurysmal degeneration. We have 
demonstrated that doxycycline: 1) inhibits experimental aneurysm formation in a dose dependent 
fashion in three different animal models of AAA; 2) decreases MMP levels in human aneurysm 
tissue when given preoperatively; 3) decreases circulating MMP-9 levels in patients with 
untreated aneurysms; 4) is well-tolerated in AAA patients in doses that achieve significant 
inhibition of aneurysms in animal models.  
The PHAST results are a warning that doxycycline treatment may be harmful instead of 
beneficial. We have selected the dose of 100 mg doxycycline twice a day because our pre-
clinical, pharmacokinetic and pharmacodynamics data (presented above) suggest that the 100 mg 
daily dose used in PHAST could not achieve circulating levels of doxycycline that would be 
efficacious but a 100 mg twice daily dose (total 200 mg per day) could. PHAST results provide 
evidence that a 100 mg daily dose of doxycycline will not reduce the growth of small abdominal 
aortic aneurysms. In consideration of the possibility of accelerated growth of small abdominal 
aortic aneurysms due to doxycycline, we will be monitoring N-TA3CT findings closely. We will 
perform repeated analyses of six-month CT scan findings anticipating a halt to study treatment if 
doxycycline accelerates aneurysm growth. We have planned formal futility analyses at the time 
of the second interim analysis for efficacy.  
The use of CT scans for precise measurement of maximal transverse diameters will allow 
us to monitor patient safety and provide information for clinical decisions with greater certainty 
than has been possible in previous clinical trials (including PHAST) that were based on 


 
 
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27 
 
ultrasound measurements. The combination of N-TA3CT dose selection and measurement 
methods and monitoring of doxycycline and MMP-9 levels will allow us to address our clinical 
trial question definitively. The definitive answer to this question is in the best interest of our 
patients and of the public which would not be well served by a clinical trial now with a lesser 
dose of doxycycline, less precise measurements of abdominal aortic aneurysm maximal 
transverse diameters, or less information on circulating levels of doxycycline or MMP-9. 
If the primary hypothesis of this study proves to be correct, and doxycycline inhibits 
expansion of small aneurysm, the entire approach to this common disease will be altered. Proof 
of the concept that AAA can be controlled medically will lead to trials designed to find more 
effective medications. Screening for AAA would become highly cost effective and the standard 
of care. The vast majority of deaths related to rupture of undiagnosed AAA would be prevented.  
CHAPTER 2 – STUDY OBJECTIVES 
The overall goal of the proposed study is to determine whether treatment with 
doxycycline 100 mg p.o., b.i.d., in patients with small abdominal aortic aneurysms (3.5 to 5.0 cm 
in diameter among men, 3.5 to 4.5 cm among women) is associated with reduced growth of the 
aneurysm in diameter compared to growth with treatment with placebo over the course of two 
years. 
2.1 
PRIMARY AIM 
1. Determine if doxycycline (100 mg bid) will inhibit the increase in greatest transverse 
diameter of small abdominal aortic aneurysms (3.5-5.0 cm in men, 3.5-4.5 cm in women) over a 
24-month period of observation in comparison to a placebo-treated control group. 
2.2 
SECONDARY AIMS 
1. Determine if doxycycline will inhibit circulating MMP-9 levels compared to a control 
(placebo) group. 
2. Determine if doxycycline (100 mg bid) will prevent the increase in aneurysm volume 
of small abdominal aortic aneurysms (3.5-5.0 cm in men, 3.5-4.5 cm in women) over a 24-month 
period of observation in comparison to a placebo-treated control group. 
3. Determine if doxycycline (100 mg bid) will inhibit the increase in greatest transverse 
diameter or volume of small aortic aneurysms (3.5-5.0 cm in men, 3.5-4.5 cm in women) using 
CT data from all 6 month time points. 
4. Determine the relationship between MMP-9 levels and interval aneurysm expansion 
rate. 
5. Determine if doxycycline treatment will adversely affect perceived quality of life. 
2.3 
EXPLORATORY AIMS 
1. Determine if doxycycline will reduce circulating interferon gamma levels compared 
to a placebo-treated group.  
2. Determine the relationship between interferon gamma and aneurysm expansion rate. 
2.4      ACCOMPLISHING OBJECTIVES  
      Pools of over nine hundred patients likely to be eligible have been identified at the 15 clinical 
sites. CT images obtained serially at 6 month intervals at the clinical sites will be transmitted for 
central reading to a specialized Imaging Core Laboratory (ICL). The blood samples collected at 
6 month intervals will be shipped to the Biomarkers Core Laboratory (BCL) for MMP-9 and 
Interferon-γ levels. The Data Coordinating Center (DCC) will receive and process data from the 
clinical sites, the ICL and the BCL for analysis. A brief description of how the aims will be 
accomplished follows: 
 
Primary Aim- This aim will be accomplished by quantitative assessment and comparison 


 
 
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28 
 
of the greatest transverse diameter on CT scan obtained at baseline and at 24-month follow-up. 
The data will be compared by methods that account for all patients ranking them based on 
aneurysm diameter and informative outcomes such as aneurysm repair, aneurysm rupture and 
death. 
Secondary Aim 1- This aim will be accomplished by quantitative assessment (g/ml) of 
plasma MMP-9 levels obtained at baseline and during follow-up at six month intervals in the 
doxycycline and placebo treated patients. Data analysis will be accomplished with longitudinal 
methods for comparison of the multiple values at different time points for time by treatment 
interactions. 
Secondary Aim 2- This aim will be accomplished by quantitative assessment and 
comparison of aneurysm volume on CT scan obtained at baseline and at 24-month follow-up. 
The data will be compared by methods that account for all patients ranking them based on 
aneurysm volume and other informative outcomes such as aneurysm repair, aneurysm rupture 
and death. 
Secondary Aim 3- This aim will use CT images obtained at each six month time point 
rather than using only baseline and 24 month values as described for the primary aim. Analysis 
will be done with longitudinal methods for comparison of the multiple follow-up points for time 
by treatment interactions.   
Specific Aim 4- This aim will use the plasma MMP-9 levels obtained at baseline and 6 
month intervals and analyze these levels in relationship to change in transverse diameter during 
the corresponding periods. Data will be analyzed with longitudinal methods for comparison of 
the multiple follow-up points. 
Specific Aim 5- This aim will be accomplished by comparing the results of the Medical 
Outcomes Study (MOS) Short Form 36 (SF-36) questionnaire assessed at baseline and during 
treatment comparing the doxycycline and placebo-treated groups. Data analysis will be 
accomplished with longitudinal methods for comparison of the multiple follow-up points for 
time by treatment interactions. 
Exploratory Aims 1 and 2 - This aim will be accomplished by quantitative assessment 
(g/ml) of plasma interferon-gamma levels obtained at baseline and during follow-up at six-
month intervals in the doxycycline and placebo groups. Data analysis will be the accomplished 
with longitudinal methods for comparison of the multiple values at different time points for time 
by treatment interactions. 
CHAPTER 3 – OVERVIEW OF STUDY DESIGN AND METHODS 
The Non-invasive Treatment of Abdominal Aortic Aneurysm Clinical Trial (N-TA3CT) 
is a randomized, double-blind, placebo-controlled clinical trial designed to test the hypothesis 
that doxycycline 100 mg p.o., b.i.d., will reduce the rate of increase of maximum transverse 
diameter of small (3.5-5.0 cm among men and 3.5 to 4.5 cm among women in the largest 
diameter) abdominal aortic aneurysms. Maximum transverse diameter will be determined by 
Computer Tomography (CT) scans. Men who are found on CT scan to have 3.5-5.0 cm 
abdominal aortic aneurysms and women who are found on CT scan to have 3.5 to 4.5 cm aortic 
aneurysms that do not involve the renal arteries are eligible. Patients will be randomly assigned 
to receive doxycycline capsules, 100 mg, p.o., b.i.d., or indistinguishable placebo capsules p.o., 
b.i.d. The primary outcome is abdominal aortic aneurysm maximum transverse diameter at two-
year follow-up with allowance for baseline (pre-randomization) diameter. Secondary outcomes 
will determine if doxycycline decreases other measures of aneurysm status, MMP-9 levels in 
plasma and whether this decrease corresponds to inhibition of aneurysm growth. 


 
 
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29 
 
3.1 
STUDY POPULATION 
3.1.1 Inclusion Criteria 
The study population will include patients 55 years of age or older who are found to have 
small (3.5-5.0 cm among men and 3.5 to 4.5 cm among women in the largest transverse 
diameter) abdominal aortic aneurysms on quantitative computer tomography (CT) scans. 
3.1.2 Exclusion Criteria 
Patients will be excluded from the study if they are unable to give their own informed 
consent to participate; have symptoms related to abdominal aortic aneurysm; have other intra-
abdominal vascular pathology that may require repair within 24 months (e.g., renal artery 
stenosis, large iliac artery aneurysms, iliac occlusive disease, aneurysmal involvement of the 
renal artery); have had previous abdominal aortic aneurysm repair by open surgical or 
endovascular technique; have an active malignancy with life expectancy less than two years; 
have an allergy to tetracycline; are currently or have been recently treated (previous six months) 
with tetracycline derivatives; they are currently taking anti-seizure medicines metabolized by 
pathways influenced by doxycyclines (e.g., carbamazepine, phenytoin, and barbiturates); stage II 
hypertension (patients whose blood pressure is persistently in the range of systolic > 160 mm Hg 
or diastolic > 100 mm Hg despite primary physician’s best effort to achieve adequate therapy); 
have dialysis dependent renal failure or impending dialysis treatment for renal insufficiency; 
have a chronic infection requiring long-term (> 2 weeks) antibiotics, have known genetic 
syndromes responsible for the abdominal aortic aneurysm (e.g., Marfan’s Syndrome), are under 
treatment with systemic immunosuppressive agents, could become pregnant, are not good 
candidates for clinical trial participation or are enrolled in another clinical trial. 
3.2 
STUDY DESIGN 
We will conduct a randomized, double-blind, parallel, two-group multicenter trial 
(Exhibit 3-1). A schedule of study procedures and treatments is found in the Appendix. 
3.3 
STUDY TREATMENTS 
Patients will be assigned to doxycycline 100 mg p.o., b.i.d., or matching placebo. Patients 
randomly assigned to doxycycline will receive bottles containing a sufficient supply of 100 mg 
doxycycline capsules to take one capsule twice a day until the next appointment (about 100 
days). Patients randomly assigned to placebo will receive a similar appearing supply of placebo 
capsules. 
3.4 
END POINTS 
3.4.1 Primary Outcome 
The primary outcome is change in abdominal aortic aneurysm maximum transverse 
diameter on CT scan from baseline to the follow-up assessment two years after randomization as 
measured in the Imaging Core Laboratory (ICL). 
3.4.2 Secondary Outcomes 
Secondary outcomes will derive from central, ICL analyses of the CT scans performed 
every six months on patients and from the clinical follow-up of randomized patients, from 
clinical observation, local laboratory findings, study visit quality of life assessments, and from 
biomarker analyses to be performed in the Biomarkers Core Laboratory (e.g., changes from 
initial MMP-9 levels, and MMP-9 levels at 24 months of follow-up). 
3.4.2.1 Clinical Outcomes of Interest 
Clinical outcomes of interest related to cardiovascular disease include death, acute 
myocardial infarction, unstable angina, aortic aneurysm rupture, and aortic aneurysm repair. 
Clinical outcomes related to chronic tetracycline therapy will include gastrointestinal side effects 


 
 
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30 
 
(e.g., nausea, vomiting and diarrhea), dermatologic side effects (e.g., rashes and photosensitivity) 
and occurrence of infectious diseases (e.g., C. difficile colitis). 
3.4.2.2 CT Scan Outcomes 
We will compare treatment groups on central, ICL assessments of infrarenal aneurysm 
neck length and diameter, aneurysm volume, characteristics of the neck (e.g., suitable or not for 
endovascular repair, neck angulation), maximum diameter at times other than two years after 
randomization, serial measurements of aneurysm diameter growth patterns, and wall stress. 
3.4.2.3 Clinical Laboratory Studies 
Liver function tests (alkaline phosphatase, alanine aminotransferase and aspartate 
aminotransferase), blood urea nitrogen (BUN), creatinine and complete blood count will be 
assessed annually at Clinical Site laboratories, reported on study forms and compared between 
the two treatment groups. 
3.4.2.4 Quality of Life 
Quality of life will be assessed with the SF-36 administered every six months. 
3.4.3 Sample Size and Data Analysis 
The study will include 258 randomized patients. Approach to no more than 1600 eligible 
patients is anticipated to be necessary to obtain consent from 258 (16%). 
The primary analysis will be performed to test the null hypothesis of no difference in 
growth of abdominal aortic aneurysms between the two treatment groups (doxycycline and 
placebo) after 24 months of follow-up as measured by CT scans analyzed in the Imaging Core 
Laboratory. This analysis will be performed according to the principle of intention-to-treat. In 
order to include patients whose 24-month abdominal CT scans are missing because of death, 
endovascular repair or for other reasons in the primary analysis, we will base our analysis on 
normal scores for percentile of rank status. We will assign worst ranks to deaths, in order of time 
from study entry to death, next worst ranks to rupture or endovascular repair with evidence of 
imminent rupture, next worst ranks to endovascular repair for reasons of aneurysm growth or 
other indication free of evidence of rupture, all in order of time from study entry within category, 
and impute from previous CT scan measurements and other patient characteristics (using 
standard SAS procedures) to 24 months for those few patients whose 24-month CT scans were 
not performed for technical reasons or are not useful for study purposes. Other patients will be 
assigned the remaining ranks according to the amount of change in largest, transverse diameter 
of the aneurysm. 
 
A Data and Safety Monitoring Board (DSMB) will review the accumulating data for 
early, convincing evidence of benefit or harm. We anticipate reviews at approximately six-month 
intervals with the first review of primary outcome data 30 months after the first patients are 
randomly assigned study treatment. There will be two interim analyses of the primary outcome 
data and our final report for the DSMB. Interim analyses will be conducted at a one-sided alpha 
level of 0.0005, leaving alpha of 0.0247 for the final efficacy analysis. This adjustment of alpha 
level is made after the fashion of Haybittle-Peto. 
 
Study power calculations are based on a 0.025 (one-tailed) alpha level and 90% power 
against the alternative hypothesis of differences in size that reflect growth less than 40% between 
the two treatment groups. The projected number of patients to be recruited allows for a sum of 
10% of patients crossing over (percentage of patients assigned to doxycycline who do not take 
their study treatment plus percentage of patients assigned to placebo who start taking 
doxycycline) and 15% missing at random data for the 24-month CT scan assessment of 
maximum cross sectional aneurysm diameter. 


 
 
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31 
 
 
We propose as a stopping guideline a Pocock-type boundary of Z=1.645, corresponding 
to a nominal one-sided 5.0% type I error rate in the direction of more aneurysm growth in 
subjects receiving doxycycline, at each of the reviews of six-month scan data. No adjustment of 
alpha or the p-value would be necessary in the primary efficacy analysis, because the study 
would be stopped on the basis of the analysis of six-month scans only if doxycycline were doing 
worse than placebo. The statistical analysis method for six-month CT scans would be the same as 
for 24-month CT scans, including the way any deaths or surgeries would be handled. 
To take into account the multiplicity of hypotheses being tested in secondary and 
exploratory analyses, a p-value < 0.01 will be required to consider there to be evidence of 
differences present, and p<0.001 for strong evidence. 
3.4.4 Time Line 
The study time line includes six months for planning, training and administrative 
approvals (e.g., DSMB approval of the final protocol, IRB approvals of the final protocol and 
final contract negotiations), eighteen months for recruitment, twenty-four months minimum for 
follow-up, and twelve months for close-out analysis and publications (Exhibit 3-2). 
 
N-TA3CT Design 
Exhibit 3-1 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Exclusion Criteria 
Randomization 
Consent 
Target Population  
Men and Women with Small Abdominal Aortic Aneurysms 
N>2000 Available to Scan  
Eligible Population 
N=950 
Study Population 
N = 258 
Patients Who are Not Screened and 
Screened Patients Who Decline to 
Give Consent
 
Follow-Up Visits Every Three Months and CT Scan Evaluation Every Six Months for Two Years 
Excluded Patients
Doxycycline 
N1 = 129 
Placebo 
N2 = 129 
 
30 Month Final Outcome Assessment, Analysis, Close-Out Visit 


 
 
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Exhibit 3-2 
N-TA3CT Study Timeline 
 
Planning, 
Training, 
Administrative 
Approvals 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Recruitment 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Follow-Up 
 
 
 
 
 
 
 
 
 
 
 
 
 
Close-Out 
 
 
 
 
 
 
 
 
 
 
 
 
 
Analysis and 
Publications 
 
 
 
 
 
 
 
 
 
 
 
 
 
            Award    
               Date 
December 31, 2012 
   March 31, 2016 
  March 31, 2018 
December 31, 2018 
      


 
 
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CHAPTER 4 – PATIENT ELIGIBILITY, ORIENTATION AND RETENTION 
4.1 
INCLUSION CRITERIA 
1. 
Patients 55 years of age or older, women post-surgical menopause or at least two 
years since last menses if natural menopause. 
2. 
CT scan documented infrarenal abdominal aortic aneurysm with maximum 
transverse diameter larger than 35 mm and no greater than 50 mm, in men, and 
larger than 35 mm and no greater than 45 mm in women. 
4.2 
EXCLUSION CRITERIA 
1. 
Prior repair of the abdominal aortic aneurysm. 
2. 
Renal artery involvement or suprarenal extension of the aneurysm. 
3. 
Documented failure of the aneurysm to increase in size over the two years prior to 
enrollment if the aneurysm is < 4.0 cm in maximal transverse diameter. 
4. 
Iliac artery aneurysm > 2.9 cm in diameter. 
5. 
Iliac artery occlusive disease planned for repair. 
6. 
Renal artery stenosis with planned open repair. 
7. 
Known thoracic aortic aneurysm > 3.5 cm, for aneurysms that are saccular or 
observed to be expanding at a rate greater than ordinary and > 4.0 cm, for 
aneurysms that are not saccular and are expanding at accustomed rates. 
8. 
Known connective tissue disease (e.g., collagen vascular disorder), heritable or 
genetic syndrome (e.g., Marfan Syndrome, Ehlers-Danlos Syndrome) underlying 
the abdominal aortic aneurysm. 
9. 
Stage II hypertension (patients whose blood pressure is persistently in the range of 
systolic > 160 mm Hg or diastolic > 100 mm Hg despite personal physician’s best 
effort to achieve adequate therapy). 
10. 
Creatinine > 2.0 g/dL or creatinine clearance < 30 ml/min. 
11. 
Allergy or intolerance of tetracyclines. 
12. 
Use of tetracyclines within past six months. 
13. 
Taking anti-epileptic pharmaceutical agents (e.g., carbamazepine, 
diphenylhydantoin). 
14. 
Current or planned treatment with chemotherapy or radiation therapy for cancer 
other than squamous cell cancer of the skin. 
15. 
Current or planned treatment with systemic immunosuppressive agents (e.g., 
prednisone, azathioprine, methotrexate, cyclosporine for autoimmune disease or 
following transplantation of bone marrow, heart, liver, lung or other solid organ). 
16. 
Chronic infection managed with long-term antibiotics, frequent courses of 
antibiotic therapy or self-administration of antibiotic therapy. 
17. 
Personal physician/surgeon is not willing to follow the protocol. 
18. 
Prognosis less than two-year survival or other reason the Clinical Site director 
believes the patient is not a suitable candidate for N-TA3CT (e.g., history of 
repeatedly missing follow-up appointments or regular residence outside of the 
U.S.) 
19. 
Enrollment in another, concurrent clinical trial study. 
20. 
Refusal or inability of patient to provide written informed consent. 
4.3 
INFORMED CONSENT 
Informed consent will be obtained by properly trained (in human subjects’ research and 
N-TA3CT Protocol) clinical site investigators (e.g., vascular surgeons), residents, fellows or 


 
 
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Clinical Site Coordinators. The patient must be competent and freely willing to give informed 
consent to participate in N-TA3CT. The patient will be told about the purpose of the study, the 
treatments – doxycycline or placebo – follow-up every three months including CT scan follow-
up every six months for two years. In patients who sign informed consent, a progress note will be 
written that identifies the patient as willing to participate in the trial. 
No Clinical Site will begin enrolling patients before its consent form is acceptable to and 
is on file at the Clinical Coordinating Center and the Data Coordinating Center. The exact 
language used on a Clinical Site’s consent form may vary from institution to institution, but the 
text must be comprehensible to persons with an 8th grade reading level, and no form will be 
considered as having been given final approval until it has been reviewed at the Clinical 
Coordinating Center. Items relevant to the presentation of each consent form are listed below: 
1. 
The purpose of the study is to determine whether doxycycline administration can 
decrease the rate of growth of small abdominal aortic aneurysms. The effect of 
doxycycline may be to decrease the activity of an enzyme which weakens the strength of 
the wall of the aorta. There is a 50-50 chance of getting drug or placebo (“look-alikes”). 
2. 
The extent of patient involvement is one out-patient visit every three months for up to 
two years and if feasible one more visit at two and a half years. Selected (9- , 15- , 21-
month) visits may be conducted by telephone if the patient prefers and the clinical site 
chooses to exercise this option. The patient should not join unless he/she is prepared to 
continue for two years. No more than three tablespoons of blood (45 ml) will be taken 
twice a year; over the course of the entire study, no more than 13 tablespoons (195 ml 
maximum) of blood will be collected. Each clinic visit will take about half an hour, and 
two times a year a member of the clinical site staff will administer a questionnaire 
concerning the quality of the patient’s life and assure that the patient receives a CT scan 
for clinical and research follow-up of the aneurysm size. 
3. 
The potential benefit is that aneurysm growth may be reduced. It may take some months 
for a beneficial effect to be noticed, if a beneficial effect exists. The inherent variability 
in rate of aneurysm growth may increase or decrease independent of any treatment effect. 
        There is a small risk of aneurysm rupture which is not thought to be altered by treatment. 
4. 
The potential risks of treatment are primarily photo sensitivity, darkening of skin 
(especially scars) or eyes, headache or G.I. disturbance (nausea, vomiting, diarrhea). 
Rarely the treatment may interact with medicine the patient is given for other conditions 
or result in changes to the bacteria naturally in the patient’s body that can result in serious 
infection (e.g., C. difficile colitis), dermatological abnormalities (including skin rash), 
and liver or kidney dysfunction. There may be other risks, but the drug is not new, and 
they should be very unlikely to occur. 
5. 
The CT scans required involve radiation exposure which might increase the risk of 
developing a fatal cancer. Patients could receive a maximum of six CT scans while in the 
study. The risk increment is small (< 1/1000; for example from a lifetime risk of 23.1% 
to 23.2% of fatal cancer), and is not entirely due to research because imaging studies are 
recommended at six-month intervals. Bi-annual imaging is routine. Some doctors use CT 
scans; others use ultrasound or some combination of the two modalities. The patient 
should be aware of the small risk associated with the CT scans. 
6. 
There is also a risk of teratogenesis. Women who could become pregnant should not join 
the study. Natural or surgical menopause is a requirement for eligibility. 


 
 
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7. 
Compliance is very important if the study is to be a success; blood samples will be 
collected at the routine appointments and tested to see if the study drug in the capsules is 
getting into the blood system at the expected levels. 
8. 
Patients will be reimbursed at least $40/visit and additional reimbursement for allowable, 
documented travel costs (e.g., mileage in excess of that accommodated by $40 for the 
visit). 
9. 
If patients become ill, because of the abdominal aortic aneurysm or some other illness, 
the study will not pay for medical care. Patients and their insurance will be responsible 
for this care. 
10. By signing the consent, the patient gives the N-TA3CT investigators permission to get 
records from any medical facility attended during the study. The study records will be 
kept confidential; patients will not be identified by name; but, data may be shared with 
the National Institute on Aging (NIA)/National Institutes of Health (NIH). 
11. The alternative to participating in the study is for the patient’s medical care to continue as 
before. Doxycycline may be added to usual care, but because of uncertainty about 
whether it will work, it is not used routinely to treat aneurysms. Patients may withdraw 
from the study at any time, without prejudice. 
12. By signing the consent, the patient acknowledges that he understands what he/she has 
been told, and that questions regarding the study have been answered. 
13. In accord with local institutional requirements, means for seeking more information about 
patient protection and redress from injury due to the study, must be spelled out. 
14. This protocol has been approved by the local Institutional Review Board (IRB). 
4.4  
IDENTIFICATION OF POTENTIAL PATIENTS 
At the beginning of the study, each lead investigator at a Clinical Site will be personally 
contacted by the Clinical Site Coordinator. Permission will be sought from each clinical site 
investigator to recruit patients who have a small, abdominal aortic aneurysm. Each site will 
maintain a list of clinical colleagues not wishing their patients to participate in the trial. Patients 
of these surgeons will not be approached for recruitment. 
 
Clinical Site Coordinators and other clinical site staff will identify patients eligible for N-
TA3CT by review of CT scan findings and medical records of vascular surgery practices 
affiliated with their clinical site. Once a potentially eligible patient is identified, the Clinical Site 
director, Clinical Site coordinator or other clinical site staff will contact the patient, preferably at 
the time of a return visit for routine abdominal aortic aneurysm follow-up, after having assured 
that the patient’s physician/surgeon is in agreement with approaching the patient. 
4.5 
PATIENT COMPETENCE TO GIVE INFORMED CONSENT 
The individual obtaining informed consent for the patient for N-TA3CT will explain the 
study to the patient and after providing the explanation ask a brief series of questions about the 
study (e.g., purpose, treatments, follow-up plans, method to communicate concerns about the 
conduct of the study or request withdrawal). If the patient answers these questions correctly, the 
patient will be accepted as competent to give informed consent. The individual obtaining 
informed consent will keep notes of the patient’s answers to the questions on an Evaluation of 
Competence to Give Informed Consent worksheet kept with the informed consent in the patient’s 
study file. A draft Evaluation of Competence to Give Informed Consent is included in Exhibit 4-
1. 
 
 


 
 
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4.6 
PROCEEDING FROM INFORMED CONSENT TO RANDOMIZATION 
Patients who give written informed consent to enroll in N-TA3CT may proceed to 
random treatment assignment after satisfactory completion of baseline evaluations (including 
review of pre-randomization CT scan in the Imaging Core Laboratory) to assure eligibility and 
collection of pre-randomization specimens (blood, plasma and serum) for later shipment to the 
Biomarkers Core Laboratory. 
 
Exhibit 4-1 
NON-INVASIVE TREATMENT OF ABDOMINAL AORTIC ANEURYSM 
(Consent Evaluation Form) 
Patient Identification Number ___ ___ ___ - ___ ___ ___      Letter Code ___ ___ ___ 
Name ________________________________________ 
To evaluate a patient’s competency to give informed consent, start by making a subjective 
judgment regarding item 1 below. If a patient is alert and able to communicate answer question 
2. If the patient is responsible for consent of medical care, ask the patient questions 3-6. Record 
each response. The evaluator may select the language to use in asking the questions in order to 
help the patient understand them. 
Items: 
 
1. Is the patient alert and able to communicate with the examiner? 
______ yes     ______ no 
If NO, patient will not be allowed to give consent. 
 
2. Is the patient responsible for consent for his or her own medical care? 
______ yes     ______ no 
If NO, the patient will not be allowed to give consent. 
3. Ask the patient to briefly describe the study treatments. 
4. Ask the patient to describe the follow-up imaging studies (“pictures of the artery bulge”) 
5. Ask the patient to explain what he/she would do if he/she decides that he/she no longer 
wishes to participate in the study. 
6. Ask the patient to name a potential risk of participation in the study. 
Signatures: 
I certify that the above patient is alert, able to communicate and able to give acceptable answers 
to items 3, 4, 5, and 6 above. 
_____________________________________    _____________________________________ 
Evaluator                                            Date             Witness                                               Date 
 
ACCEPTED RESPONSE TYPES 
1. Yes 
2. Yes 
3. Two treatments, one with a drug (doxycycline) and one with an inactive, look alike 
(placebo). 
4. X-ray pictures (CT scan) every six months. 
5. Tell the study nurse, study doctor, personal doctor, or a medical center/hospital official that 
he/she does not wish to participate. 
6. Complications of placebo or doxycycline (skin rash, dizziness, change in eye sight, nausea) 
or loss of privacy. 


 
 
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CHAPTER 5 – METHOD OF RANDOMIZATION 
5.1 
ELIGIBILITY ASSESSMENT 
At the time an eligible patient provides informed consent, Clinical Site staff will complete 
all sections of the pre-randomization forms, collect pre-randomization blood, plasma and serum 
specimens, and submit the qualifying CT scan to the Imaging Core Laboratory by overnight 
courier. Within two working days of receipt of the CT scan by the Imaging Core Laboratory, a 
confirmation or denial of patient’s eligibility according to CT scan criteria will be sent to the 
Clinical Site and DCC. If confirmed, the Clinical Site Coordinator must contact the patient’s 
vascular specialist prior to randomization. If the vascular specialist confirms his/her willingness 
to follow the protocol and the patient meets all eligibility criteria, the Clinical Site Coordinator 
will complete randomization to obtain the patient’s assigned study treatment kit number. 
5.2 
RANDOM TREATMENT ALLOCATION PROCEDURES 
The DCC staff will prepare randomization schedules for each Clinical Site participating 
in N-TA3CT. The program for generating randomization schedules will have the following 
characteristics: 1. There will be two randomization strata – one for men and one for women – at 
each clinical site; 2. Treatments are assigned in random order within blocks sizes two, four, six 
or eight with equal numbers of patients assigned to the doxycycline or placebo within each 
block. 3. Block sizes (two to eight patients per block) are randomly selected with the probability 
of each block size specified by DCC staff. 
The DCC staff will maintain a web-based randomization system for Clinical Site staff to 
request treatment allocations as eligible patients are identified. The randomization system is 
accessible only to study personnel who enter the password for the Clinical Site and his/her 
assigned personal identification number (PIN). An individual can request a treatment allocation 
after he/she has passed training in the use of the system. Questions include specification of 
gender, confirmation that the patient meets all inclusion criteria and has no exclusion criteria and 
that the patient has given informed consent for enrollment. Depending on the answers to these 
items, the next available treatment allocation is issued. The date and time of the completion of 
the treatment assignment is the time of study entry for each patient. 
5.3  
CLINICAL SITE RANDOMIZATION START-UP 
 
Each clinical site will be randomizing patients one by one during the course of the 
clinical trial. To assure that no clinical site randomizes too few patients to be included in data 
analyses, randomization of the patients into the study at any given site can begin once clinical 
site staff  have supplied the Data Coordinating Center with a list (without direct identifiers) of 16 
eligible, pre-screened patients based on clinical practice records. 
CHAPTER 6 – STUDY TREATMENTS 
6.1 
INTRODUCTION 
 
For patients enrolled in N-TA3CT dosage will be fixed at one capsule p.o., b.i.d. (either 
100 mg doxycycline or placebo). Capsules should be taken in the morning (e.g., after brushing 
teeth, with morning coffee) and in the evening (e.g., after dinner). These capsules will be 
provided to the study patients in individually marked study treatment kits containing supplies of 
doxycycline or placebo according to the patient’s random assignment. 
 
The Treatments Distribution Center (Catalent) coordinator will ship study treatment kits 
to the Clinical Sites as directed by the Data Coordinating Center. Clinical Sites will acknowledge 
receipt of medication kits by means of a form sent to the Data Coordinating Center. 
 
 


 
 
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6.2 
TREATMENT PREPARATION 
 
Doxycycline 100 mg capsules will be over-encapsulated at Catalent. Placebo (Starch 
1500) will be packed in identically appearing, over-encapsulated capsules. The capsules will all 
be the same size, and will contain a white powder. The Treatments Distribution Center will count 
out a three-month supply of each type of capsule in numbered kits, bottle them in child-proof 
containers, and label them with a unique, study assigned number and instructions including the 
“Investigational Drug” warning, “Non-invasive Treatment of Abdominal Aortic Aneurysm 
Clinical Trial”, instructions on how to take the capsules, and an “emergency call” telephone 
number. Study treatments will be shipped in three-month batches, in sufficient time to assure that 
the Clinical Sites will receive the bottles before the patient’s next scheduled visit. Inventory 
records for drug and placebo will be kept by Treatments Distribution Center staff. 
6.3 
BLINDING 
 
In the Clinical Sites, Clinical Coordinating Center, Imaging Core Laboratory and 
Biomarkers Core Laboratory, the patients, directors (i.e., Clinical Site lead investigator), 
coordinators, and other study staff will be blinded to treatment assignments. Staff of the 
Treatments Distribution Center and Data Coordinating Center will have access to individual 
patient treatment assignment on a “need-to-know” basis. The Treatments Distribution Center and 
Data Coordinating Center will maintain records of each patient’s treatment assignment. 
 
Plans have been made to prevent unblinding of patient treatments. Despite these 
precautions, if the Clinical Site director thinks he/she inadvertently has become unblinded, 
patient contact must be carefully managed to avoid any comment to the patient or coordinator 
regarding unblinding. 
 
The Clinical Site directors will assert at the outset the intention to avoid seeking 
information that may unblind them with regard to individual patient’s treatment assignments. 
Clinical Site coordinators will conduct patient follow-up visits and process and maintain files of 
study documents blind to treatment assignment. Discussions among Clinical Site staff or with 
patients regarding possible patient treatment assignment are inappropriate. As long as official 
unblinding has not been done and the patient notified, the Clinical Site coordinator must avoid 
seeking any information that may unblind him/her. 
6.4 
EMERGENCY UNBLINDING 
 
Every patient will be given an identification card describing his participation in the study, 
listing emergency study telephone numbers (e.g., the Clinical Coordinating Center telephone 
number, Treatments Distribution Center telephone number, the Clinical Site director’s telephone 
number, and the Data Coordinating Center telephone number). 
 
In an emergency, arrangements will be made so that the patient’s medication can be 
disclosed to the Clinical Site director (or a party designated to share this responsibility) after 
consultation between the Clinical Site director and a Clinical Coordinating Center or other N-
TA3CT leadership physician (one of whom will always be available). Reasons for unblinding are 
limited, are based on clinical grounds, and unblinding must be initiated by the Clinical Site 
director. Reasons for unblinding include accidental ingestion of study medications by another 
person and clinical conditions in which management might be changed if the nature of the study 
drug were known. 
 
If a patient’s therapy is unblinded, the Clinical Coordinating Center must send a report on 
the reasons for unblinding to the Data Coordinating Center. The nature of the patient’s 
medication will be withheld from the Clinical Site coordinator. 
 


 
 
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6.5 
TREATMENT INTERRUPTIONS 
 
There may be instances of treatment interruption either related to medical conditions 
(e.g., acute, intercurrent illnesses such as an infection when it may be advisable to interrupt study 
therapy without unblinding) or for other reasons (e.g., study treatments lost in a robbery). Non-
emergent interruptions for medical conditions should be allowed only with the advice of the 
Clinical Site director and not at the discretion of local medical doctors. The Clinical Site director 
is responsible for notifying the Data Coordinating Center of treatment interruptions. 
6.6 
ASSESSMENT OF COMPLIANCE 
 
At approximate six-month intervals each patient’s plasma samples will be assayed for 
doxycycline at the Biomarkers Core Laboratory. 
 
Capsule counts will be done at each regular follow-up visit. If capsule counts are not 
consistent with regular compliance, Clinical Site staff will discuss the nature of any difficulties 
with the patient. The importance of compliance will be emphasized for all patients. 
 
Even if patients are repeatedly considered to be non-compliant, they will continue to be 
followed and will continue to receive study reimbursements (e.g., for travel and parking). 
6.7 
MISSED VISITS AND DROP OUTS 
 
Each regularly scheduled clinic visit missed by a patient will be reported to the Data 
Coordinating Center. Patients who do not wish to continue attending clinic visits in N-TA3CT 
will continue to be followed as much as possible for CT scan evaluations, identifiable events and 
vital status. 
6.8 
DURATION OF STUDY TREATMENT 
 
The goal of the study will be to maintain all patients on protocol for two years.  
CHAPTER 7 – CONCOMITANT CARE 
7.1 
INTRODUCTION 
Except for study treatments and CT scan monitoring, care of study patients will be 
determined by their personal (referring) physicians. Personal physicians will be blind to study 
treatment and will be advised to select concomitant care to be appropriate whether the patient is 
receiving doxycycline or placebo. Cooperation of treating physicians and house officers or 
hospitalists will be solicited as much as possible to enhance adherence to study protocol at the 
same time as assuring best practices and standard of care treatment for N-TA3CT patients. 
Patients will be issued study identification cards requesting treating physicians and house 
officers/hospitalists to contact the patient’s N-TA3CT Clinical Site director with questions 
concerning N-TA3CT patient treatment. 
7.2 
HOSPITAL ADMISSIONS 
During hospital admissions for any cause, the patient’s care must be directed to address 
the patient’s main, acute medical condition. If a patient is admitted to a hospital, study drug is to 
be continued if possible. If a patient forgets to bring medication with him when admitted, a 
family member or friend should bring it to the hospital as soon as possible. Exact arrangements 
used to dispense a study drug to an inpatient may differ between Clinical Sites; such 
arrangements must be explored before recruitment begins, and the Clinical Coordinating Center 
is available for assistance in such administrative matters. For Clinical Sites which may have 
patients admitted to one of several hospitals, such arrangements should be made, in advance, for 
all likely possibilities. 
 
 
 


 
 
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7.3 
BLOOD PRESSURE AND LIPID CONTROL 
7.3.1 Anti-Hypertensives 
At the time or randomization, N-TA3CT patients must have normal blood pressure or be 
undertaking reasonable treatment for Stage I hypertension (e.g., on one or more appropriate anti-
hypertensive agents with BP ≤ 160/100 mm Hg). Because of the prevalence of high blood 
pressure among patients who have abdominal aortic aneurysms, it is important that blood 
pressure eligibility criteria not exclude patients who would receive doxycycline if it proved 
efficacious. All patients must receive appropriate standard of care therapy. Stage II hypertension 
is not considered to be adequately managed for participation in this clinical trial. Patients with 
Stage II hypertension can be evaluated for enrollment if their blood pressures can be brought 
under control. Personal physicians will be encouraged to maintain adequate control of their 
patients’ blood pressures, adding or changing medication to restore control when blood pressures 
become elevated, or undesirable side effects appear, in accordance with the reports of the Joint 
National Committee on Prevention, Detection, Evaluation and Treatment of High Blood 
Pressure. Letters reporting blood pressure findings will be sent to personal physicians for every 
patient who is found to have high blood pressure (BP > 120/80) on study follow-up visits for use 
in patient management. Control of anti-hypertensive therapy, as other clinically indicated 
therapies, will remain with the patient’s personal physician. 
7.3.2 HMG-CoA Reductase Inhibitors (“Statins”) and Other Lipid Lowering Agents 
The American Heart Association and American College of Cardiology recommend that 
abdominal aortic aneurysm be considered equivalent to atherosclerotic cardiovascular disease 
(ASCVD) based upon historical studies showing that cardiac mortality is not an uncommon 
cause of death in abdominal aortic aneurysm patients. In order to achieve lipid level treatment 
goals, the preferred treatment for LDL-cholesterol reduction being HMG-CoA reductase 
inhibitors (“statins”). Personal physicians will be encouraged to establish adequate control of 
their patients’ lipid levels making use of prescription for diet, exercise and lipid-lowering agents 
(statins if well tolerated or other agents, such as niacin, if statins are not well tolerated) as 
appropriate for their patients, prior to randomization. Clinical Site directors will encourage study 
patients’ personal physicians to maintain adequate control of their patients’ lipid levels, adding 
or changing lipid-lowering medication(s) only to restore control when lipid levels become 
elevated or undesirable side effects appear, in accordance with National Cholesterol Education 
Program guidelines. 
7.4 
TREATMENTS WHICH MAY INTERACT WITH DOXYCYCLINE 
7.4.1 Anti-Infectives 
Personal physicians will be encouraged to select anti-infectives, should they become 
necessary for patient management (e.g., to treat an intervening pneumonia or urinary tract 
infection) as if the patient had been taking doxycycline – i.e., to select bacteriocidal non-
tetracycline antibiotics as much as possible. 
7.4.2 Wafarin 
Personal physicians will be advised that if they plan to initiate warfarin therapy for study 
patients there is a possibility that the study treatment (which is being maintained at a fixed dose) 
may influence the dose of warfarin used to achieve desired international normalized ratios 
(INRs). Thus, INR levels should be monitored closely until the personal physician has 
established that a stable therapeutic dose of warfarin has been achieved. 
 
 


 
 
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7.5 
SMOKING AND OTHER TOBACCO USE CESSATION 
Clinical Site directors will advise study patients’ physicians that avoidance of tobacco 
use is an important part of management of abdominal aortic aneurysms and encourage the 
personal physicians’ efforts to achieve smoking and other tobacco use cessation in their patients. 
7.6  
SURGERY 
7.6.1 Introduction 
With the exception of retroperitoneal procedures in the central body area, there are no 
study-specific recommendations concerning the performance of surgery. 
7.6.2 Retroperitoneal Surgery 
 
Because of the potential risk of aneurysm rupture consequent to retroperitoneal surgical 
procedures, the attending vascular interventionalist may elect to repair the aneurysm before 
retroperitoneal surgery using an endograft or at the time of the retroperitoneal procedure. 
CHAPTER 8 – FOLLOW-UP PROCEDURES 
8.1 
INTRODUCTION 
Patients will be scheduled for follow-up visits every three months. At the time of each 
scheduled visit, a medical review is conducted including ascertainment of clinical outcomes, 
study treatment compliance and adverse effects. 
 
At semi-annual follow-up visits, outcome CT scans will be collected, specimens for 
Biomarkers Core Laboratory assay and storage will be collected, and study forms evaluating 
medical review and quality of life will be administered. At annual visits blood specimens will be 
collected for local evaluation of CBC, liver function and renal function. The results will be 
included on study forms. 
8.2 
FOLLOW-UP VISITS 
At the three-month follow-up visits, a medical review will be conducted including 
ascertainment of possible adverse effects, current therapies, and collection of the previous study 
medication and capsule count. The prescribed study medication for the following three months is 
dispensed. 
 
In addition to all three-month follow-up visit procedures, at the six-month visits and at 
the final follow-up assessment the patient will complete an SF-36 for an assessment of health-
related quality of life, plasma and serum will be collected and prepared for shipment to the 
Biomarkers Core Laboratory, and a CT scan of the abdomen will be performed, anonymized and 
sent to the Imaging Core Laboratory. 
 
When a patient requests that a follow-up be conducted by telephone the following 
applies. 
Telephone Follow-up Visits 
At the discretion of clinical site investigators and coordinators, patients who cannot without 
undue burden travel to the N-TA3CT clinical sites may conduct visits which do not coincide with 
an N-TA3CT required CT scan or specimen collection with a study team member via telephone. 
The only exception to this approach would be the three-month visit for which a clinic visit is 
mandatory for thorough assessment of subject’s tolerance and compliance with study treatment. 
Telephone contact will include: 
1. Identify yourself and state that you are recording form data portions of the conversation 
for quality control purposes.  
2. Verification of subject identity.  
3. Review of all required data fields on Subject Follow up form. 
4. Review of all current medications. 


 
 
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5. Assessment of subject’s compliance with taking medication by self-report.  
6. Review of proper dosing, side effects, documentation of missed doses on study diary. 
7. Instruct subject new study drug will be sent via mail or overnight courier. Upon receipt of 
new study drug subject is to stop taking medication from existing supply and begin taking 
from new bottle. Encourage subject to tape old bottle closed and to set aside. Remind 
subject not to destroy any study bottles or the box in which they came. All will need to be 
returned at time of next clinic visit. It is recommended similar instructions be included 
with the shipment of new drug. 
8. Recording of the telephone contact with a device approved by the study leadership (e.g., 
Olympus VN-3100PC). 
Completion of Case Report Forms: 
1. Follow Up form: Complete with information obtained from telephone call. 
2. Vital Signs: Mark as not done. 
3. Concomitant Medications: Complete with information obtained from telephone call. 
4. Drug Adherence: Complete Item #4 with information provided by subject. The remainder 
of the form is to be completed at next clinic visit when study bottles are returned. 
5. Drug Dispensing: Complete at time new study treatment kit is assigned and dispensed. 
Do not record date sent to subject or date subject receives. Keep tracking record for pick-
up and delivery of study treatment kit with source documents. 
8.2.1 CT Scan Acquisition at Enrollment and Follow-up 
The standardized acquisition protocol for computed tomography (CT) examination of the 
abdominal aorta and iliac arteries in N-TA3CT is as follows for multidetector, helical CT: 
1) Narrow Field-of-View centered on aorta, < 36 cm encouraged. 
2) Tube current and energy to be set to best clinical standards for image quality. 
3) Obtain CT scout without contrast. 
4) Scan from top of diaphragm to symphysis pubis. 
5) Section thickness less than or equal to 2.5 mm. 
6) Reconstruction overlap minimum 50%, i.e., 1.25 mm intervals, 2.5 mm thickness. 
If requested by ordering physician (suggested but not required by study protocol), infuse 
nonionic contrast, 300 mg iodine per/ml, total volume of 150 ml through arm vein at a rate of 3 
ml per second. Delay individualized with test dose program. Each participating clinical site may 
have different CT contrast dosage protocols and CT scanner automatic exposure control systems. 
These individual measures can be taken to reduce radiation exposure and contrast dosage 
providing there is preservation of diagnostic-quality images. Coach breath holding instructions. 
Scan 2.5 mm section thickness from top of diaphragm to symphysis pubis (about 300 mm). 
Specific performance standards for different equipment (e.g., Siemens 64 slice CT 
Scanner) will be provided in the Imaging Core Laboratory’s Procedures Manual. 
CT Scans should be transferred from clinical sites to the Imaging Core Laboratory in 
DICOM format on CD. Each CD should contain one study, on one patient only and be sent to the 
Imaging Core Laboratory by traceable Federal Express (FedEx) or United Parcel Service (UPS) 
courier. 
The scan recorded in the CD should be de-identified (e.g., no patient name, address, 
medical record number or birth date appears on the headers or images) so that only the study 
assigned patient ID number (PID) and Alphabetic Code (Letcode) appears with the images. The 
PID/Letcode and specific visit (baseline, 6-month, 12-month, 18-month, etc.) should be in the 
electronic header and recorded manually (e.g., with a label provided by the Imaging Core 


 
 
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Laboratory) on the non-recording side of the CD. The study to be recorded on the scan should be 
the full dataset of axial series, without contrast required and with contrast encouraged for best 
practices but not required because contrast may be contraindicated in particular patients for 
clinical reasons. Only physicians and surgeons at the clinical site are able to judge whether a 
contrast study is acceptable for any particular patient. 
8.2.2 Biomarker Blood Specimen Collection at Enrollment and Follow-up 
 
Clinical sites must develop a brief but specific plan for the procurement, initial 
processing and temporary storage of peripheral blood samples from all enrolled patients during 
the study. Each site will ship specimens to the Biomarkers Core Laboratory. Blood will be 
collected in plasma collection tubes which contain lithium heparin and serum collection tubes. 
Samples will be processed into sterile 0.25 ml aliquots of serum, plasma with timely transfer of 
specimen on ice between collection and processing areas, if necessary. Short-term storage of the 
sample aliquots in -70 C freezers (or colder) is required at the clinical sites. Shippers of the 
materials must have taken the appropriate training for the shipment of DOT/IATA Biological 
Substances Category B and Dry Ice. The Biomarkers Core Laboratory will provide shipping 
labels and labels for each aliquot to the clinical sites. Batch shipment of the specimens packed in 
dry ice will be made at three-month intervals. The specimens will be maintained frozen at the 
Biomarkers Core Laboratory, and some aliquots analyzed in batches per this protocol and the 
remainder will be stored for future studies.  
 
Subjects enrolled in the study may, optionally, participate in the banking of genetic 
information at the Biomarkers Core Laboratory of the study. Each enrolled subject will be 
offered this opportunity. After a detailed discussion of all of the risks, including the unique 
privacy risks of genetic material, subjects may provide consent for the collection and storage of 
their genetic information. At one of the study visits, the subject will have whole blood drawn 
(approximately 10 ml) which will be frozen as described above for transport to the Biomarkers 
Core Laboratory. The samples will be processed for DNA at the Biomarkers Core Laboratory, 
and this material will be placed into aliquots. There are no specific studies planned to evaluate 
the genetic information in this protocol, however this material will be maintained with the 
plasma and serum samples at the Biomarkers Core Laboratory and will be made available for 
future studies. 
8.2.3 Ascertainment of Adverse Events 
All medical contacts indicating possible serious or unexpected adverse events (SAEs), 
whether treated on an out-patient or in-patient basis, will be reviewed by the Management 
Committee. These SAEs will be classified by an independent committee of vascular surgeons 
who are in institutions that are not in the N-TA3CT clinical consortium, blind to study treatment, 
the N-TA3CT Event Classification Committee. Their classifications of aneurysm rupture, repair 
of aneurysm indicated by impending rupture and repair of aneurysm for reasons of size as well as 
other less severe indication will be used in the primary outcome analysis. SAEs will be reported 
to the Data and Safety Monitoring Board. 
8.3 
PATIENT MANAGEMENT DURING MEDICAL CONTACTS 
Each patient will be given an identification card to be carried at all times and to be 
presented at any medical contact, stating that he/she is enrolled in the N-TA3CT, that his/her 
medication must be continued if at all possible, that endograft devices etc. should be deployed 
only in accordance with N-TA3CT protocol, that N-TA3CT will require records of medical 
contacts, and that the N-TA3CT Clinical Site director and Clinical Coordinating Center are to be 
contacted in case of need to know the study treatment assignment. 


 
 
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Medical staff at N-TA3CT Clinical Sites should be aware of study requirements for the 
management of patients presenting with abdominal aortic aneurysm or its complications, and the 
requirements of documenting all medical contacts. The patient should be maintained on study 
medication if hospitalized. N-TA3CT Clinical Site directors will avoid seeking information 
which may unblind them with respect to the N-TA3CT treatment assignment. 
8.4 
 SECONDARY ASSESSMENTS 
8.4.1 Psychosocial Evaluation 
Data from the Medical Outcomes Study Form SF-36 will be used in secondary analyses. 
Questionnaires will be administered prior to enrollment and every six months during the study 
and at the close-out visit. 
8.5 
 RETENTION 
 
Maintaining participation post-randomization is especially relevant in studies with older 
adults who may be lost to follow-up due to changes in living situation, acute illness, cognitive 
changes, or death. At the time of enrollment, the participant will be asked to provide contact 
information (names, addresses, phone numbers) for two people that know them well and who 
would likely know the whereabouts of the participant if s/he could not be contacted for a follow-
up assessment.  
8.5.1  Retention Promotion Efforts 
 
At the time of randomization, participants will receive clear, easy-to-follow, written 
instructions about the schedule of follow-up assessments. Reviewing these instructions with the 
participant prior to departure will be a priority. If demonstrated compliance problems exist, study 
staff will review the importance of the visits and the remainder of the study schedule. Involving 
the participant’s spouse or other family members in these reviews can be useful. Attempts wil be 
made to maintain continuity of follow-up care, so that, whenever possible, the same staff 
member contacts/sees the participant throughout the study. Every attempt will be made to make 
all contact with the participant pleasant. Minimizing waiting time, reimbursement for 
transportation costs and comfortable waiting room facilities makes the visit more pleasant, 
thereby enhancing participant retention at follow-up assessments. 
 
Clinical sites will be advised to keep detailed records of rescheduled or broken study 
assessment appointments for each participant. Participant retention will be monitored, and efforts 
will be made to identify those participants who need support and encouragement. Summary 
reports of such difficulties help to identify problems. Critical review of such problems may offer 
potential solutions. 
 
The following procedures will be implemented to prevent missed follow-up: 
 Sending out pre-visit reminders (e.g., postcards and phone calls) for upcoming 
assessment visits.  
 Participants will be called the night before a follow-up assessment to remind them of 
the visit and the time for transportation pick-up, if appropriate. 
 Clinical sites will track attendance so that staff will be immediately alerted to a 
missed intervention or assessment visit. 
 Immediately contacting participants (or significant others) when they miss a visit. 
 Rescheduling the visit within the same window, if possible. Follow-up assessments 
that fall outside of the target window remain important and will be used in all 
analyses. Whenever an intervention visit or follow-up assessment is missed, it will be 
recorded as such on the appropriate form with a corresponding reason. This will also 
be documented in the Tracking Database.


 
 
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Some randomized participants may not actively participate. Regardless of the reason(s), 
these participants will be followed until the end of the study, and study staff attempts to make 
contact every four weeks after randomization. These contacts are intended to remind the 
participant that they are welcome to rejoin the study at any time. Considerable effort will be 
expended to collect main outcome data at each of the follow-up assessments. 
 
The following strategies will be used to promote adherence to the protocol, in terms of 
intervention adherence and follow-up assessment attendance. 
 Participant-staff relationship A key element contributing to participants’ continued 
commitment to the trial will involve fostering positive, respectful relationships 
between study participants and individual members of the staff. 
 Continuity The number of study staff making contact with a participant will be 
controlled. In general, the same study staff contacting participants to schedule the 
baseline and follow-up assessments will also conduct the study visits. This will 
ensure consistency of study staff contact across participants. 
 Clinic Environment A clinic environment that is warm and pleasant and oriented to 
the comfort of the participant will be maintained. 
 Participant-staff communications Good and consistent communication will be 
essential. Instructions will be clear and interactions will be friendly and 
individualized. The participant will be reminded of the benefits of study participation. 
 Convenience and accessibility An easily accessible location at the clinical sites, 
availability of transportation, and convenient hours all serve to facilitate study 
adherence. Clinical sites will take steps to ensure that follow-up assessment 
attendance is not compromised by lack of transportation, unsuitable hours of 
operation, or any similar circumstance. If necessary, transportation will be provided 
for participants or reimbursed for parking. 
 Time in clinic will be kept to a minimum, consistent with maintaining quality. If 
waiting is necessary, the situation will be explained to the participant. However, 
participants will not be rushed or made to feel unwelcome. Clinical site staff will be 
trained to take time to visit with participants. Clinical sites may provide juice and 
snacks during the study assessment visits.  
 Reminder phone calls will be placed the night before a visit to enhance attendance for 
assessment. 
8.5.2  Monitoring Recruitment and Retention 
 
Retention will be monitored through completion rates of follow-up assessment visits. 
Regular reports will be available to clinical sites and the Clinical Coordinating Center. Members 
of the Clinical Coordinating Center maintain regular phone contact with clinic staff to review 
retention. 
8.6  
FINAL FOLLOW-UP ASSESSMENT AND CLOSE-OUT 
Patients who complete a two-year follow-up visit after March 29, 2016, and before May 
1, 2018, will discontinue study medication at that time. For patients who have completed at least 
24 months of therapy and had a two-year follow-up visit before the time of approval of Protocol 
Version 1.6.0, the patient will cease medication on the date of the next scheduled visit. There 
will be a final follow-up assessment in the 30-month window for those who completed two years 
of follow-up before May 1, 2018; those who complete drug treatment after 27 months will 
complete a final follow-up assessment between 3 and 6 months after discontinuation of therapy.  
No study visits are necessary between therapy discontinuation and final follow-up assessment. 


 
 
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Patients randomized after May 1, 2016, but before October 29, 2016, will have their 24-
month visits completed on or before July 31, 2018, and, in lieu of their 30-month follow-up, final 
assessment visits approximately 30 days after their 24-month visits to check for any adverse 
events after discontinuation of study drug. This final assessment may be done by telephone. 
Patients randomized after October 29, 2016, will have their final on-study visit at their 18-month 
follow-up, to be scheduled no later than July 31, 2018. They will discontinue study drug at this 
visit. Their final assessment contact will be conducted approximately 30 days after their 18-
month visits to check for any adverse events after discontinuation of study drug. Their primary 
outcome will be imputed as described in Section 11.3. To summarize, study treatment, collection 
of aneurysm measurements and biomarkers will be completed within the follow-up windows for 
July 31, 2018; final assessments for safety will be completed no later than September 1, 2018. 
After the conclusion of study follow-up for all patients, a close-out visit will be offered to 
each patient. When notified of study close-out, each site should arrange close-out visits for 
enrolled patients within a relatively short period of time. Every effort will be made to schedule 
enrolled patients for the close-out visit. Vital status will be ascertained on patients lost to follow-
up. At the close-out visit, patients will be informed of the primary results of the study and the 
recommendations of the N-TA3CT investigators. They will be informed of their individual study 
treatment assignments. 
CHAPTER 9 – STUDY END POINTS 
9.1 
INTRODUCTION 
 
The primary objective of the Non-Invasive Treatment of Abdominal Aortic Aneurysm 
Clinical Trial (N-TA3CT) is to determine the safety and efficacy of doxycycline (100 mg p.o. 
b.i.d.) in the prevention of growth of small abdominal aortic aneurysms. The primary analysis in 
this study will compare the change from baseline in size of abdominal aortic aneurysms at two 
years from randomization, using a normal-score transformation, between patients randomly 
assigned to doxycycline and those assigned to placebo with adjustment (co-variance) for size of 
aneurysm at study entry.  
Follow-up CT scans will be obtained every six months regardless of compliance with 
assignment study treatment. The analysis of the primary study outcome will be conducted on an 
“intention to treat” basis with two-sided statistical tests comparing the outcome between groups 
of patients defined at study entry by random assignment to the doxycycline or placebo groups. 
9.2 
PRIMARY END POINT 
 
For the purpose of defining the primary study outcome, the size of the abdominal aortic 
aneurysm will be defined as the maximum transverse diameter (adventitia to adventitia) 
measured on a quantitative CT-scan in the orthogonal plane. The measurements to be used in the 
primary end point assessment will be taken from CT scans obtained at baseline (study entry) and 
two years after random treatment assignment. A normal score will be assigned to each baseline 
and two-year follow-up measurement for analysis. If a two-year follow-up measurement is not 
available, the primary outcome will be defined as described in the following section. 
9.3 
STATISTICAL ANALYSIS OF THE PRIMARY END POINT 
 
An attempt will be made to measure for each patient the maximum transverse diameter of 
the patient’s abdominal aortic aneurysm two years after random treatment assignment. It is 
anticipated that during two years of follow-up a small percentage of the patients may die or be 
operated on for repair (endovascular or open) due to growth of the aneurysm beyond clinically 
tolerable limits (5.5 cm for men and 5.0 cm for women) or be operated on for symptoms or signs 


 
 
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including pain or signs of rupture or in less urgent circumstances. These patients’ two-year 
aneurysm diameters will be missing for cause and will be accommodated in the primary outcome 
analysis by classification (by the N-TA3CT Event Classification Committee) and rank order 
methods.  
 
A committee of vascular interventionalists not active in study clinical sites and blind to 
treatment assignments will review relevant medical records for all deaths and abdominal aortic 
aneurysm repairs to classify outcomes as related or not related to rupture of aneurysm. 
Patients’ outcomes will be ranked at two years following randomization as follows: 
patients who die will be ranked worst, in order of number of days from randomization to death; 
patients who experience rupture of the abdominal aortic aneurysm will receive the next worst 
ranks, in order of number of days from randomization; patients who undergo open or 
endovascular repair without evidence of rupture will receive the next worst ranks, in order of 
days from randomization to repair; patients with follow-up CT scans will be given the remaining 
ranks from worst (largest diameter) to best (smallest diameter).  
The primary analysis will follow the intention-to-treat principle; that is, all randomized 
patients will be included, assigned to treatment according to the treatment received. In the event 
a patient does not complete two years of CT scan follow-up for cause(s) other than death, open 
repair or endovascular repair (e.g., withdrawal of consent or loss to follow-up, expected to be 
rare events), the patient’s previous CT scans and other characteristics will be used for the 
imputation of a measurement of the abdominal aortic aneurysm diameter to be included in the 
primary analysis.93 The ranks for patients assigned to doxycycline will be compared to the ranks 
of patients assigned to placebo using normal scores.94 If N is the total number of randomized 
patients, a rank of k will be assigned the value corresponding to k/(N+1) in a standard normal 
distribution; for example, a rank of 100 out of 248 patients will receive the normal score 
corresponding to 0.4016, or -0.2492. The change in normal score from baseline -- i.e., the two-
year normal score for patients randomized to doxycycline and placebo minus the baseline normal 
score -- will be compared using a linear regression model (analysis of covariance -- ANCOVA), 
with variables for the randomized treatment assignment (0 for placebo, 1 for doxycycline), the 
normal score for baseline diameter, and gender in the model. 
9.4 
SECONDARY AND INTERMEDIATE OUTCOMES 
Secondary and intermediate outcomes will include a variety of clinical findings and 
quality of life assessments, Biomarker Core Laboratory findings and Imaging Core Laboratory 
findings. 
9.4.1 Clinical Secondary and Intermediate Outcomes 
The clinical components of the primary outcome, death, open repair and closed repairs 
will be ascertained and compared between the treatment groups. Other clinical outcomes that 
will be compared between the treatment groups will include myocardial infarction, 
cerebrovascular accident (stroke), peripheral arterial thromboembolic events, venous 
thromboembolic events, and infections (urinary tract infections, pneumonia, gastrointestinal 
infection, etc.). 
The frequency of adverse effects of doxycycline (e.g., clinical presentations such as 
photosensitivity rash, gastrointestinal symptoms and clinical biochemistry laboratory findings 
such as abnormalities of liver function tests or renal function tests) will be ascertained at each 
clinic visit and compared between treatment groups. 


 
 
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Health-related Quality of Life will be assessed every six months with the Medical 
Outcomes Study (MOS) Short Form-36 (SF-36) and compared between treatment groups and 
according to patient characteristics. 
9.4.2 Biomarker Secondary and Intermediate Outcomes 
Blood specimens collected for analysis in the Biomarker Core Laboratory will be used to 
assess circulating levels of MMP-9, IFN- and high sensitivity (hs) C-reactive protein (CRP) 
with comparisons between the treatment groups and according to baseline characteristics as well 
as cotinine and doxycycline levels for assessment of their relationship to aneurysm growth. 
Doxycycline levels will also be used to evaluate protocol adherence. Additional specimen 
analyses may be planned in ancillary studies with independent support. 
9.4.3 CT-Scan Secondary and Intermediate Outcomes 
CT Scans collected every six months will be analyzed in the Imaging Core Laboratory to 
make comparisons between the treatment groups of characteristics of abdominal aortic aneurysm 
dimensions assessed after randomization, other than the primary outcome, with adjustment for 
baseline characteristics. Secondary outcomes measured on two-year CT Scans and on other study 
CT Scans will include aneurysm volume, characteristics of the aneurysm neck, and wall stress. 
CHAPTER 10 – DATA MANAGEMENT 
10.1 
INTRODUCTION 
N-TA3CT data management occurs in two phases – pre-randomization and post-
randomization. The purpose of data management procedures is to establish in as timely a manner 
as feasible (as close to “real time” as practical) the set of information necessary to assess the 
status of every patient with regard to treatment assignment (randomization); treatment supply; 
follow-up visit completion; blood specimen collection, processing, shipment, receipt, analysis 
and storage; CT scan collection, processing, shipment, receipt, analysis and storage; and, clinical 
observations and inquiries. This timely and accurate set of information is necessary for 
monitoring study performance and progress and well as to perform data analyses for the 
assessment of study findings. 
10.2 
FORM DATA 
The form data will include information collected pre-randomization to establish patient 
eligibility, assign a study treatment and assign a numeric identification (PID) and alphabetic 
(Letcode) that will be used to track the patient’s study treatments, follow-up visits, clinical data, 
CT Scans and blood specimens throughout the study. Form data will include a record of all 
clinical information, whether or not blood specimens were collected (and for collected specimens 
their label numbers), and whether or not CT Scans were performed for each follow-up visit. 
Clinical Site staff will store all source documents in locked file cabinets. 
Clinical data will be entered by the site staff using a secure, Web-based 21CRF11-
compliant electronic data capture system (eDC). Data Coordinating Center (DCC) staff will train 
and test proposed clinical site staff in the basic conduct of N-TA3CT and in the use of the Web 
systems. Each clinical site staff member who successfully completes training and passes the 
DCC certification test will be accorded a unique Staff ID for access to the website. The staff 
members will be responsible for selecting their passwords adhering to requirements for password 
strength. Clinical site staff may use any computer (lap-top or desktop) allowed by their medical 
centers for entry of study data. They have permission for entry of their patients’ data based on 
their Staff ID. Data Coordinating Center staff will train clinical site staff to keep their passwords 
confidential. Passwords must be changed on a regular schedule to enhance confidentiality. The 


 
 
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important point for scientific integrity is that clinical site staff understand that they have 
responsibility for data entered under their Staff ID passwords. 
Clinical site staff must maintain a hard copy of each study form entered as a source 
document, on site. They may complete forms manually and then key in the data, or may enter 
data on the website and then print out the form entered, check the print out for accuracy and then 
sign the printed form.  
The eDC system will be programmed with extensive data edits including range and value 
checks, checks to make sure that all required fields are completed and logical consistency both 
within form page and across pages. Data edits will be responded to, tracked and closed within the 
eDC system. Further data edits based on monitoring or hand review of the data will be entered 
into the eDC system by DCC and CCC staff and similarly responded to, tracked and closed. 
10.3 
CT SCANS 
As described in Chapter 8, clinical site staff will transmit CT scans in DICOM format on 
CD’s to the Imaging Core Laboratory. 
Imaging Core Laboratory staff will assess the abdominal aortic aneurysm measurements 
in duplicate (one reader, two readings) and create a database of each patient’s CT scan 
measurements. This database will include dates of CT scan receipt, initial review (if any) and 
definitive evaluation. 
Upon receipt of a pre-randomization CT scan from a clinical site, the Imaging Core 
Laboratory will notify the clinical site (with copy to the Data Coordinating Center) whether the 
abdominal aortic aneurysm meets eligibility criteria for N-TA3CT (e.g., no renal artery or iliac 
artery involvement, maximum transverse diameter greater than 3.5 cm and less than 4.5 cm for 
women or 3.5 cm and 5.0 cm for men) within two working days of receipt. 
The Imaging Core Laboratory will enter pre-randomization screening data, quality 
assessment data and CT scan measurements from both the qualifying CT scan and follow-up 
scans directly into a secure Web application linked with the randomization application. 
Measurement and quality assessment data will be merged with forms data as needed for 
reporting. 
10.4 
BIOMARKER SPECIMENS 
As described in Chapter 8, clinical site staff will ship frozen blood, plasma and serum 
specimens to the Biomarkers Core Laboratory. 
Biomarkers Core Laboratory staff will inventory specimens according to unique label 
numbers on each vial received. The vials will be numbered uniquely with freezer proof labels 
designed by the Biomarkers Core Laboratory where there will be no other patient-specific 
information in the course of the study. Specimen label number and PID/Letcode will be linkable 
through the Data Coordinating Center or Clinical Site of origin, only. A system of replicate 
labeling and processing will be used for blinded replicate quality control. 
Biomarker Core Laboratory staff will assay specimens for doxycycline, cotinine, MMP-
9, IFN- and hs-CRP on the direction of the Data Coordinating Center and create a data file of 
doxycycline, MMP-9, IFN-, hs-CRP levels for each label number analyzed. At three-month 
intervals, on days negotiated by the Data Coordinating Center and the Biomarkers Core 
Laboratory, Biomarker Core Laboratory staff will transfer doxycycline, MMP-9, IFN-, hs-CRP 
and cotinine data according to label number to the Data Coordinating Center for integration into 
the main study database, monitoring of specimen collection, processing, shipment and evaluation 
and reporting of interim results to the Data and Safety Monitoring Board. 
10.5 
QUALITY CONTROL 


 
 
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The quality of the proposed trial is dependent upon 1) achieving the projected recruitment 
and randomization rates, 2) the complete and accurate collection of data, and 3) adherence to the 
protocol. The investigators recognize that effective quality control procedures are of particular 
importance in a multi-center study. Therefore, we have developed quality control procedures 
with significant central oversight and designed a governance structure specifically to support 
community-based sites. 
10.5.1 Training of Study Personnel 
The Clinical Site directors and Coordinators are certified in procedures of key importance 
to the study prior to initiation of recruitment. Initial training for certification is at the first of the 
annual collaborator’s meetings and on site initiation visits. The initial one-day coordinator’s 
meeting will include a thorough review of recruitment procedures; obtaining informed consent; 
randomization; protocol adherence; and data collection and reporting procedures including use of 
the data entry system. Training methods include a presentation on the Procedures Manual; 
question and answer sessions; and hands-on experience using the data collection forms, the 
randomization system and the data entry system. Participants will be provided with sample charts 
and asked to complete data entry forms. Recommended procedures include reviewing completed 
forms for missing information and checking for consistency of PID and Letcode between forms. 
In addition, we will describe the pilot and related study experiences with specific reference to 
identified problems and solutions, effective mechanisms for maximizing patient recruitment, and 
means of insuring protocol adherence. 
Certification requires passing an examination on the protocol and demonstrated 
proficiency in required tasks (e.g., completing study forms, using the randomization and data 
entry systems.) The DCC staff will work with CCC staff to prepare specific certification 
procedures. Recertification occurs in association with site visits and is based on the level of 
proficiency demonstrated in the on-going performance of study activities (e.g., a form failing edit 
rate less than 10%), and demonstrated expertise. Associate Clinical Site coordinators will be 
trained by the Clinical Site coordinator and Clinical Site director using materials developed by 
the CCC and DCC. Small secondary training sessions will be held at the CCC or DCC for 
Clinical Site staff in need of assistance to improve performance, who are from Clinical Sites 
joining the consortium after the training meeting or who are replacing previously trained staff 
under circumstances that do not permit adequate training at the Clinical Site alone (i.e., abrupt 
departure of certified staff).  
Each Clinical Site will be visited annually for the purpose of assessing quality and 
providing additional training. The site visit team will include a representative of the DCC, ICL, 
and CCC. During these site visits, we will provide additional review of the study protocol and 
procedures. We will also review any problems identified by the DCC quality monitoring 
procedures and/or during the site visit and recommend possible remedial actions. In addition, the 
Clinical Site Coordinators will participate in conference calls with the CCC Lead Coordinator 
every month. 
Collaborators' meetings will be held annually for all Clinical Site directors and 
coordinators. These meetings will include a review of study procedures, discussion of any 
problems and solutions, and an update on the status of the trial. It will also provide a forum for 
open discussion among the collaborators and generate continued enthusiasm for participation in 
the trial. 
10.6 
QUALITY MONITORING  
10.6.1 Data Collection 


 
 
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The DCC has primary responsibility for monitoring data collection, providing protocol 
adherence review, and initial editing of the reported data. All forms entered via the data entry 
system will be evaluated by a central edit program designed to detect out-of-range items, 
inconsistent sets of items and combinations of items that are incomplete. Results of the edit will 
be included in the DSMB Reports. 
In addition to these standard editing procedures, source documents of the first two 
randomized patients after the start of the study (or at the time the Clinical Site Coordinator 
changes) from each site and a random sample of 10% of patient source documents will be 
carefully reviewed by the CCC Lead Coordinator to insure the accuracy of the data entry 
process. The DCC will generate a list of the study identification numbers of the patients whose 
documents will be reviewed. This will be sent to the Clinical Site coordinator at the Clinical Site 
who will then copy the documents and forward to the CCC. The DCC will also forward the list 
of identification numbers along with a computer printout of the data items to the CCC. CCC staff 
will compare the data entered with the data in the source documents and record whether or not 
each data point was verified. This review will be designed to identify problems in recording data, 
data entry, data processing, as well as protocol adherence. 
10.6.2 Recruitment and Randomization Rates 
The DCC will carefully monitor recruitment, minority representation, and randomization 
rates, and reports will be prepared and distributed weekly for N-TA3CT leadership (Management 
Committee) conference calls and will be sent in monthly reports to each site. 
If recruitment is low for a site, we will review with the Clinical Site coordinator and 
Clinical Site director how recruitment is organized and implemented. If the CCC or DCC staff 
are unable to solve the problem, then study leadership will seek assistance from Clinical Site 
coordinators with excellent recruitment rates. If these steps do not resolve the problem, study 
leadership will visit the site. Besides meeting with all the key personnel, study leadership will 
walk through recruitment procedures. 
It may be helpful for the Study Chairman to present rounds for surgical and/or medical 
departments. This would provide the opportunity to answer questions and to generate interest in 
the trial. If the problem is that one of the opinion leaders at the hospital is not supportive of the 
study, the Study Chairman will communicate directly with Clinical Site opinion leaders as an 
educational tool to help them comprehend the nature and importance of the clinical trial. 
10.6.3 Response to Protocol Violations 
Major protocol violations are those that undermine the fundamental premise of the study 
(such as failure to administer the assigned treatment) or jeopardize patient safety (such as 
inappropriate administration of contrast in CT scan performance). The initial response to a major 
protocol violation will be remedial efforts such as conference calls, site visits and development 
of procedures to prevent the violation from reoccurring. However, if remedial efforts fail to 
address the problem (e.g., failure to maintain a treatment crossover rate < 10%), randomization 
function for that site will be suspended pending review by the study leadership, DSMB, and 
NIA. This review could result in either more aggressive remedial efforts or termination of the 
Clinical Site.  
10.6.4 Response to Lapses in Clinical Site Performance 
Lapses in Clinical Site performance include the failure to obtain the required CT scans or 
blood specimens as outlined in the protocol, failure to complete and submit the required forms, 
and failure to adhere to the assigned treatment. Both the DCC data editing process and the more 
thorough review of a subset of charts will be used to identify lapses in performance. These will 


 
 
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be discussed with the Clinical Site coordinators during conference calls with the CCC every 
month and reported in the DSMB. 
Minor lapses in Clinical Site performance are those that impede progress of the study 
such as not submitting data in a timely fashion (form delinquencies). Clinical sites will receive 
reduced payments if there are cases whose forms are intolerably delinquent or until those form 
delinquencies are corrected. 
CHAPTER 11 – DATA ANALYSIS 
11.1 
INTRODUCTION 
The primary analysis will be a linear regression analysis (analysis of covariance -- 
ANCOVA) to test the hypothesis of no difference in the mean change from baseline in 
abdominal aortic aneurysm maximum transverse diameter at two years after randomization, 
allowing for maximum transverse diameter at the time of randomization (i.e., comparing growth 
in maximum transverse diameter), between patients randomly assigned to doxycycline 100 mg 
bid and patients assigned to placebo (see Sections 9.3 and 11.3). The analysis will be done on 
normal scores derived from the ranks of the diameters at baseline and at two years after 
randomization. Quantiles (2.5, 25, 50, 75, 97.5 percentiles) will be reported for baseline and two-
year measurements. The primary analysis will be performed in accordance with the principle of 
intention-to-treat. 
11.2 
INTERIM MONITORING 
A Data and Safety Monitoring Board (DSMB) will review the accumulating data for 
early, convincing evidence of benefit or harm. We anticipate DSMB reports at approximately 
six-month intervals over the period of patient recruitment and follow-up, at least five interim 
reports followed by a final analysis after the conclusion of data collection. Since the first follow-
up CT scan will not be performed any earlier than six months after the start of recruitment, we 
anticipate at least five interim reports on CT scan findings prepared for the DSMB and one final 
analysis. To maintain the overall Type I error rate () at 0.025 (one-sided), while performing 
interim monitoring for the primary outcome, we propose to use a simple, Haybittle-Peto type 95,96 
(adding a small adjustment to the final alpha level) repeated analysis adjustment for efficacy. We 
will perform the first interim analysis when 1/3rd of the primary outcome data are available and 
the second when 2/3rds are available. With this approach, we will require stringent evidence of 
treatment efficacy. The final efficacy analysis will be performed at close to the overall -level 
planned for the study (one-sided 0.025). The critical p-value proposed for each of the two 
efficacy interim analyses is one-sided 0.0005 (corresponding to t=3.291), which allows a final 
analysis at a critical p value of 0.0247 (corresponding to t=1.965) in an approach similar to that 
proposed by Haybittle-Peto. 95,96 
The above approach will be used in monitoring for possible early termination of the trial 
because of a beneficial effect of doxycycline. The DSMB will assess the interim data for possible 
early stopping for harm. Also, we propose that the trial could be stopped early if there is 
evidence of futility at the second interim analysis such that the conditional probability of 
rejecting the null hypothesis given data observed to the time of the analysis and the assumption 
that the design alternative hypothesis is correct is less than 20%.97   
 
We propose as a stopping guideline a Pocock-type boundary of Z=1.645, corresponding 
to a nominal one-sided 5.0% type I error rate in the direction of more aneurysm growth in 
subjects receiving doxycycline, at each of the reviews of six-month scan data. No adjustment of 
alpha or the p-value would be necessary in the primary efficacy analysis, because the study 
would be stopped on the basis of the analysis of six-month scans only if doxycycline were doing 


 
 
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worse than placebo. The statistical analysis method for six-month CT scans would be the same as 
for 24-month CT scans, including the way any deaths or surgeries would be handled. 
Before making a recommendation to terminate the trial, either for a beneficial effect, 
futility, or to discontinue study treatment, the DSMB will consider the interim analysis results 
along with other relevant factors.  These factors will include enrollment and study conduct, the 
numbers and distributions of deaths and surgeries for aneurysm repair, the frequencies and 
distributions of other secondary outcomes described in Section 9.4, and findings from other 
relevant studies.  
 
11.3 
PRIMARY ANALYSIS 
A variety of analytic methods will be used for primary, secondary and other analyses 
(Table 1). The primary study outcome will be the change from baseline of abdominal aortic 
aneurysm maximum transverse diameter measured two years from randomization. This outcome 
will be compared according to assigned treatment (consistent with the principle of intention-to-
treat), using linear regression analysis (ANCOVA)98, with baseline abdominal aortic aneurysm 
maximal transverse diameter and gender, as well as an indicator of treatment assignment, in the 
model. The test for differences between treatments in the primary outcome will be conducted at 
an overall one-sided -level of 0.025.  (The significance level could also be given as two-sided 
0.05; stating it as one-sided 0.025 emphasizes the hypothesis that the investigators wish to 
“prove” statistically, and it also corresponds to the boundaries for interim monitoring for a 
beneficial effect of doxycycline.)  
Prior to performing the primary outcome analysis, an interaction term for the 
randomization stratum (gender) with treatment assignment will be assessed in the ANCOVA 
model. If there is significant (at two-sided =0.05) evidence of differences in the effect of 
treatment according to gender, other than a small quantitative difference between effects that are 
in the same direction for each gender, the analysis will be performed separately for men and for 
women. This gender-treatment interaction is of concern because the pathophysiology of 
abdominal aortic aneurysm in women could differ from that in men by more than just a simple 
shift in normal aortic diameter, and it would be important to know if a large treatment effect 
among men results in an overall treatment effect in the study but does not extend to women. 
If an interaction between treatment and gender exists, the assumptions of the ANCOVA 
test of overall, homogeneous, treatment effect will not be met and an overall estimate of the size 
differential will not make sense. In the event of a treatment by gender interaction being observed, 
we will review study results by gender to determine the nature of the differences in treatment 
effects. If there is no interaction, the ANCOVA will be used to estimate the overall treatment 
effects across all clinical sites and both strata (genders). The primary analysis will be performed 
on the normal scores corresponding to percentile rank of each patient at baseline and follow-
up.94,99-101 At baseline all scores will be based on rank of abdominal aortic aneurysm maximum 
transverse diameter; information that must be available for the patient to be enrolled and will be 
centrally checked for eligibility. At two-year follow-up, rank will be assigned for patients who 
did not undergo two-year CT scan according to survival status, classification of patient condition 
at the time of surgical intervention (evidence of rupture, or symptoms or signs of imminent 
rupture versus undergoing repair for reason of maximal transverse diameter exceeding 5.5 cm for 
men or 5.0 cm for women or other less urgent indication). Multiple imputation methods available 
in SAS will be used for those (presumed to be few) patients who do not, for unanticipated 
reasons, complete a CT scan at two years (e.g., because of a missed appointment). Two-year CT 


 
 
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scans missing for pre-specified causes such as death, rupture, surgical intervention will be 
assigned ranks as described above. 
The model for the primary outcome ANCOVA after removal of unimportant, interaction 
term(s) would be: 
Y =  + 1X1 + 2X2 + 3X3 + , where 
Y = the change in normal score from baseline, assessed after two-year follow-up, 
 = the study average change in normal score 
1 = the treatment effect coefficient where  
     X1 = 1 = doxycycline or 0 = placebo 
 
2 = the baseline normal score coefficient where 
X2 = the baseline normal score 
 
3 = the randomization stratum (gender) coefficient where 
X3 = 1 = female or 0 = male 
 = error. 
The interaction term to be tested (and dropped from the model if unimportant) would be 
4 X1X3 where 4 is the treatment by gender interaction coefficient. 
11.4 
SUBGROUP ANALYSES OTHER SECONDARY OUTCOMES AND ANALYSES 
Linear regression models will be used to explore age, statin use status (user versus non-
user), race/minority status and other factors for effects on growth of abdominal aortic aneurysms. 
Secondary analyses will be conducted to assess the effect of doxycycline in subgroups of 
interest, for example subgroups defined by clinical factors. 
To take account of the multiplicity of hypotheses being considered in secondary analyses, 
a p-value < 0.01 will be required to consider that some evidence of differences is present, and 
p<0.001 for strong evidence. Secondary outcomes are analyzed with more stringent alpha-levels 
to allow for multiple comparisons; a stringent alpha level reduces the likelihood of Type I error 
to some extent. This adjustment does not involve the primary outcome comparison or interim 
analysis plan. 
Secondary outcomes to be considered include the occurrence of clinical events such as 
death, the occurrence of myocardial infarction or death and thromboembolism, measures of 
health related quality of life, high sensitivity C-reactive protein, MMP-9 levels, Interferon  
(IFN-) levels, other measurements made on CT scans (at two-years or other six-month time 
points) and subgroup analyses of the primary outcome. 
 
With the data we plan to collect on 129 patients assigned to doxycycline and 129 patients 
assigned to placebo, we will have an opportunity to evaluate prognostic indicators that have not 
been adequately considered previously for abdominal aortic aneurysm outcomes. Among the 
characteristics that we evaluate are: gender, age, race/ethnicity, co-morbid states, baseline 
imaging characteristics of the abdominal aortic aneurysm, concomitant medications, and baseline 
MMP-9, high sensitivity C-reactive protein levels, etc. 
 
Binary outcomes will be considered as present/absent variables, and the effect of 
treatment on these outcomes will be analyzed initially with a comparison of two proportions and 
a chi-square test.101 Additional analyses, taking account of other patient characteristics, will be 
performed using logistic regression.93,103,104 Outcomes in multiple categories (e.g., death, non-
fatal myocardial infarction, acute coronary syndrome, no coronary artery disease) will be 
analyzed in 2 x k contingency tables using chi-square tests with more than one degree of 
freedom.102 Time to death will be analyzed with log rank statistics.105 


 
 
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55 
 
 
Losses-to-follow-up are expected to be few for vital status and surgical intervention 
status, based on our pilot experience and the required follow-up of abdominal aortic aneurysm 
patients for their clinical management. Missing two-year follow-up CT scans are expected to be 
less than 10%. Death and surgical intervention are accommodated in the proposed primary 
intention-to-treat analysis by rank order and normal scores methods, but if these outcomes are 
unrelated to treatment, they represent noise, lost opportunities to measure treatment effect on 
aneurysm size. CT scan data will be imputed using standard SAS procedures for participants 
who are missing two-year CT scans for reasons other than death or surgical intervention.93 
Sensitivity analyses will be performed to assess the impact of loss-to-follow-up. These will 
include, for example, assuming that all patients lost to follow-up have good outcomes, (e.g., no 
aneurysm growth) and assuming that all patients lost to follow-up have bad outcomes (e.g., 
maximal aneurysm growth). Allowance for losses of opportunity to measure treatment effects 
has been made in planning the number of patients to be enrolled in the study (See Section 11.5). 
 
ANCOVA models will be fitted to assess associations between patient characteristics and 
abdominal aortic aneurysm growth (i.e., blood pressure, use of aspirin, use of beta-blockers).106  
GEE models will be used for assessments based on all CT scans collected.107,108 Cotinine levels 
will be treated as a time dependent covariate in the GEE models.107,108 
 
Protocol treatment adherence will be assessed according to capsule counts and 
doxycycline levels. Capsule counts will be analyzed for comparability of adherence between 
treatment groups and to assess the success of study performance against an absolute expectation 
of individualized patients achieving 80% or better adherence. Similarly, doxycycline levels will 
be assessed by comparing the distributions at the six-month visits against the blood levels known 
to be efficacious in animal models and anticipated from the proposed regimen. If there is failure 
to achieve anticipated levels, protocol revision to a new regimen (e.g., increased dose) could be 
considered. Cotinine levels will be used for objective confirmation of patient reports on smoking 
and use of nicotine patches. That correspondence within patient may bear on general adherence 
and accuracy of reporting. 
11.5 
STATISTICAL POWER 
In this section we consider sample size and power in two stages. First we determine the 
sample size required for 90% power to find a significant difference between doxycycline and 
placebo with respect to the change in abdominal aorta diameter from baseline to two years after 
randomization, at a one-sided 0.025 significance level, when the treatments are compared using a 
linear regression model (ANCOVA) for change from baseline in normal scores corresponding to 
abdominal aorta diameter, with adjustment for the normal score at baseline. Then we show that 
the power is still high (> 80%) after allowance for deaths and surgical interventions.   
The number of patients to be enrolled in N-TA3CT was planned based on information 
from three databases – a research database, kindly provided as a privileged communication by 
Frank Lederle, M.D., the pilot study conducted by B. Timothy Baxter, M.D., and colleagues, and 
a reconstruction of the database from which results were published by Mosorin and colleagues in 
their pilot clinical trial of six months of doxycycline therapy (compared to placebo) for effect on 
abdominal aortic aneurysm diameter.57,60 We have compared the aneurysms’ average annual 
growth rate in these databases to published data from other studies of small abdominal aortic 
aneurysm and found them to be consistent. The three databases we analyzed establish an annual 
aneurysm growth rate of 2.5 mm/year as a reasonable lower bound for placebo. 
Statistical power with our proposed method of analysis (ANCOVA) depends upon the 
correlation (r) of baseline measurements with follow-up measurements, the between-subject 


 
 
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variability (standard deviation, SD) of the measurement to be made, specified alpha-level (0.05, 
two-tailed) and the alternative hypothesis specified. We have specified the alternative hypothesis 
as a 40% reduction in aneurysm diameter growth rate (from 2.5 mm/year to 1.5 mm/year) 
because it is a difference in growth rate likely to be clinically important for our patients 
(prolonging growth to a size indicating intervention to repair by a median of four years, and 
close to but smaller than that observed by Mosorin (50% reduction).60  
We found correlations of r= 0.88 for 12 placebo patients at 18 months and r=0.94 for 
doxycycline patients (14) in Mosorin, r=0.94 for measurements one year apart in Lederle (664 
patients) and r=0.88 for all (31) treated patients in Baxter’s data. We expand this range of 
observations to a range of r=0.80 to r=0.96 for consideration in power calculations. We found 
standard deviations of abdominal aortic aneurysm diameter ranging from 4.6 mm in one of 
Lederle’s treatment groups at baseline and 6.2 mm in the other (average 5.4 mm), 5.0 mm and 
5.8 mm on follow-up (average 5.4 mm), to 5.8 mm at baseline and 7.5 mm at follow-up in 
Baxter’s data, and 7.5 mm among Mosorin’s placebo patients and 6.8 mm among the Mosorin 
doxycycline patients at baseline (average 7.2 mm), 9.6 mm and 7.6 mm (average 8.6 mm) on 
follow-up respectively. We used these data to set a 5.0 mm to 9.0 mm range of standard 
deviations of interest. In Lederle’s data there were 18 patients whose measurements were made 
at a two-year interval; r=0.82 and SD=4.3 mm at first measurement and 6.9 mm on the second. It 
seems reasonable (and conservative) to predicate initial (effective) sample size on a two-year 
correlation of 0.82 and a standard deviation of measurement of 7.0 mm and assess the sensitivity 
of the statistical power of our proposed clinical trial in the ranges of r=0.80 to 0.96 and SD=5.0 
mm to 9.0 mm. 
Data from published studies using doxycycline in patients who have small abdominal 
aortic aneurysms may be summarized as follows: 
 
 
 
 
Aneurysm Size 
 
 
 
 
Control 
Doxycycline 
Study 
N 
Doxycycline 
Daily Dose 
Imaging 
Modality 
Initial 
Final 
Initial 
Final 
Mosorin et al60 
32 
150 mg 
Ultrasound 
35 mm 
39 mm 
31 mm 
33 mm 
 
 
 
 
(31.0-40.0) 
(30.5-44.0) 
(27.5-38.5) 
(27.0-38.5) 
 
 
 
 
 
 
 
 
Baxter et al57 
36 
200 mg 
CT Scan and 
Ultrasound 
NA 
NA 
41.0 
42.7 
 
 
 
 
 
 
± 0.9 
± 1.3 
 
Using r=0.82 and SD=7.0 mm (corresponding to SD=4.2 mm for change in diameter), 
specifying desired statistical power to be 0.90, with a basic t-test formula and reduction of group 
size (n1=n2) for the efficiency of ANCOVA compared to t-test109, we arrive at treatment group 
sizes of 85 patients each. On consideration of the range of correlations and standard deviations in 
our planning data sets, we find that the statistical power of ANCOVA for 85 patients assigned to 
each treatment group and a two-group comparison will be as in the table below. 
 
Statistical Power of a Two Group Comparison at Two-Year Follow-Up 
                 Standard Deviation of Dependent Variable 
 
 
5.0 mm 
7.0 mm 
9.0 mm 
0.80 
0.99 
0.87 
0.67 


 
 
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Correlation of 
pre- and post-
treatment 
measurements 
0.88 
>0.99 
0.97 
0.86 
0.96 
>0.99 
>0.99 
>0.99 
 
We will have over 80% power in the event that our projected baseline to two-year follow-
up correlation in abdominal aortic aneurysm maximal transverse diameter is modestly optimistic 
(i.e., really should have been 0.80 instead of 0.82) or our projected variability (standard 
deviation) of maximal transverse diameter was too optimistic (i.e., really should have been 9.0 
mm instead of 7.0 mm), protecting study power against possible mistaken design assumptions. In 
the unlikely event that both assumptions are mistaken, we will still have close to 70% power.  
These power calculations are based on a t-test and ANCOVA on the CT scan aneurysm 
diameter measurements.  However, we plan to use ANCOVA on normal scores.  Since the power 
of the normal scores test is asymptotically equivalent to the power of the t-test if the aneurysm 
diameter measurements are normally distributed and is likely to be higher than the power of the 
t-test if the measurements are not normally distributed94, the power calculations should give a 
good approximation to the power of the normal scores test. 
The above calculations do not allow for the possibility of death, rupture of the abdominal 
aortic aneurysm, or open or endovascular repair without evidence of rupture in some patients 
within two years after randomization. If we assume one of these events will occur in 10% of 
randomized patients, then interpolation in Table 2 of McMahon and Harrell99 with survival = 
90% indicates that 90% power for a rank test on aneurysm diameter corresponds to power of > 
80% for a “worst-rank” test on a combined outcome of aneurysm diameter and the events noted 
above. (We actually expect these events to occur in less than 10% of patients, but in order to be 
conservative we have assumed 10% in the power calculation.)  
Since a normal scores test is as powerful as or more powerful than the corresponding 
rank test, and thus have the same power asymptotically94, power of > 80% also applies to our 
primary analysis, ANCOVA on normal scores. 
 We also want to allow for the possibility that 10% of  patients in each treatment group 
will not adhere to assigned study treatment (e.g., patients assigned to doxycycline do not take 
their medication because of gastrointestinal discomfort or skin rash, etc., and patients assigned to 
placebo end up taking doxycycline for an indication like periodontitis or rosacea) and for the 
possibility that two-year CT scans will be missing for 15% of patients for reasons unrelated to 
outcome and treatment effect (i.e., noise in a system with signal we are trying to detect).  
Taking into account all the above possibilities – death, aneurysm rupture, or open or 
endovascular repair without evidence of rupture; nonadherence; and missing scans for reasons 
unrelated to outcome or treatment effect – the number of patients to be randomized to each 
treatment group becomes 85 ÷ (1-0.1)2 ÷ (1-0.15) = 124 patients.110 We have been awarded a 
supplement to enroll 10 additional women to increase the statistical power and precision of our 
gender-based analyses. Thus, our recruitment goal is 258 patients (129 in each treatment group), 
a number that should give us high power to find a statistically significant effect if the true effect 
is a 40% reduction in the mean increase in aneurysm diameter from 2.5 mm/year to 1.5 mm/year, 
if our well documented design assumptions (correlation and standard deviation) are 
approximately correct for the population we enroll. If our assumptions are absolutely correct, 
adherence perfect and there are no patients who do not undergo CT scan of the abdomen at two-
year follow-up, we will have a very well powered study (~ 90% or more).  


 
 
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With 129 patients in each treatment group, setting alpha levels at the more stringent 0.01 
for some evidence in secondary outcome analyses we would find statistical significance for 
between-treatment group differences of approximately 0.5 standard deviation (about the same as 
observed in other studies10,91) with 90% power using ordinary Student’s t-tests. For clinical 
events that occur with a frequency of 15% we would have 80% power to find significance for 
reductions to 5%. Statistical power would not be appreciable for reductions in events occurring 
with lower frequency; lesser effect sizes would be detectable for more frequently occurring 
events. 
 
 
 
 
 
 
 
 
 
Table 5: Analysis Methods for Performing Primary, Secondary and Other Analyses 
Analyses 
                  Method  
1. Primary End Point Analyses 
 
a) Abdominal aortic aneurysm diameter 
ANCOVA of change from baseline in normal scores of 
ranked outcomes, allowing for baseline rank and 
randomization stratum (gender) 
b) Gender interaction with treatment 
ANCOVA 
2. Secondary Analyses (Treatment Group 
Comparisons) 
 
a) Baseline characteristics in each treatment group 
Descriptive statistics (means, standard deviations, 
percentages) 
b) Baseline characteristic and duration of treatment 
interactions with treatment effect                                      
ANCOVA of ranked outcomes, allowing for baseline 
rank, randomization stratum (gender) and including 
interaction term(s) for the baseline characteristic(s) of 
interest with treatment assignment 
c) Adherence 
Chi-square tests with one degree of freedom, T-tests 
d) Abdominal aortic aneurysm imaging (CT scan) 
findings on repeated studies 
Multiple logistic regression, Generalized Estimating 
Equations (GEE) and Mixed Models 
e) Clinical events such as death, aneurysm rupture, 
aneurysm repair, myocardial infarction, stroke, acute 
coronary syndrome, in univariate or combinations and 
the interaction of treatment effect with duration of 
treatment 
Log Rank Tests and Cox Proportional Hazards 
Regression. Comparison of two proportions using a 
chi-square test with one degree of freedom 
f) Biomarkers (MMP-9, IFN-) 
T-tests, ANCOVA, GEE, and Mixed Models 
g) SF-36 
T-tests, GEE 
3. Other Analyses 
 
a) Other characteristics’ (including Statins and 
Biomarkers) effect on aneurysm growth 
ANCOVA, GEE 


 
 
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b) Patient characteristics association with clinical 
events 
Log Rank Tests and Cox Proportional Hazards Tests. 
Chi-square tests with one degree of freedom, and with 
more than one degree of freedom 
 
CHAPTER 12 – STUDY ORGANIZATION 
12.1 
PARTICIPATING UNITS 
 
The participating units in N-TA3CT integrate with the N-TA3CT administrative structure 
to form the study organization (See Exhibits 12-1 and 12-2). 
12.1.1 Clinical Site 
At each collaborating hospital, a Clinical Site director and Clinical Site coordinator will 
be identified. These two people will work closely together to assure successful performance of 
the trial. Clinical Sites are expected to enroll an average of just over 16 patients each (16 Clinical 
Sites x 16.125 patients/Clinical Site = 258 patients). Eligible patients seen over a three month 
period in each Clinical Site participating in N-TA3CT planning are presented in Exhibit 12-1. 
Clinical Sites in the N-TA3CT organization are subject to change according to the feasibility of 
study conduct at each site. 
12.1.1.1 Clinical Site Director 
The responsibilities of the Clinical Site director include: 1) To insure that all vascular 
surgery staff involved with the care of abdominal aortic aneurysm patients are well informed 
about the trial; 2) To insure that all patients with abdominal aortic aneurysm are routinely 
considered for the trial; 3) To insure that the treatment assignment is followed; 4) To 
communicate with Clinical Coordinating Center staff and Data Coordinating Center staff any 
problems or concerns related to the study; 5) To assist the Clinical Site Coordinator as necessary; 
6) To ensure that all guidelines governing clinical research are adhered to. 
12.1.1.2 Clinical Site Coordinator 
The responsibilities of the Clinical Site Coordinator include: 1) To identify all abdominal 
aortic aneurysm patients for the trial; 2) To obtain informed consent from the patient if not 
obtained by the Clinical Site lead investigator or other investigators, fellow or resident; 3) To 
enroll the patient in the study by telephoning the randomization service; 4) To inform the 
surgeons and nurses caring for the patient of the patient’s randomization in N-TA3CT; 5) To 
complete data collection forms and process data edit queries; 6) To insure compliance with the 
study Protocol; 7) To e-mail or fax data forms to the Data Coordinating Center and arrange for 
copying and mailing of medical records to the Clinical Coordinating Center for quality 
assurance; 8) To insure the clinical site office and pharmacy are stocked with trial materials; 9) 
To participate in telephone calls with the Clinical Coordinating Center head nurse; 10) To train 
assistant site coordination and other staff at the Clinical Site as needed; 11) To maintain all 
Clinical Site study materials; and 12) To adhere to all guidelines governing clinical research. 
12.1.2 Clinical Coordinating Center 
 
The University of Nebraska Medical Center, Division of Vascular Surgery will be 
responsible for clinical coordination of the trial. The responsibilities of the Clinical Coordinating 
Center are to: 1) Provide administrative and fiscal support for the Clinical Sites; 2) Provide 
technical, patient assessment and Protocol adherence advice to Clinical Site staff; 3) Assist 
Clinical Sites to correct problems with recruitment, Protocol adherence and data collection; 4) 
Participate in site visits; 5) Provide advice about any aspect of the trial; 6) Direct performance of 
the Treatments Distribution Center; 7) Direct performance of the Biomarkers Core Laboratory; 
and 8) Lead presentation and publication of study results for the scientific and lay press. 


 
 
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12.1.3 Data Coordinating Center 
The Department of Epidemiology and Preventive Medicine, University of Maryland 
School of Maryland, Baltimore, Maryland, will be responsible for data coordination of the trial. 
The responsibilities of the Data Coordinating Center are to: 1) Provide all study materials to 
centers; 2) Provide a 24 hour randomization service; 3) Receive data collection instruments and 
to verify them for completeness, retrieving any missing data from the centers; 4) Enter data into 
a computer database and conduct routine data edits; 5) Monitor performance to detect problems 
with recruitment, Protocol adherence and data collection; 6) Participate in site visits; 7) Provide 
advice about any aspect of the trial; and 8) Perform interim and final analyses. 
12.1.4 Imaging Core Laboratory 
The Imaging Core Laboratory will be responsible for the standardized, quantitative and 
qualitative assessments of CT scans in N-TA3CT. Using labels provided by the DCC and 
Clinical Site programming to delete identifiers in the electronic file to obscure patient 
identification, Clinical Sites will send CT scan CDs to the Imaging Core Laboratory. Imaging 
Core Laboratory staff will send electronic files of CT scan assessments to the DCC. The DCC 
will direct a limited number of blinded CT scan resubmissions to the Imaging Core Laboratory 
for quality control. Imaging Core Laboratory CT scan measurements will be used for the primary 
outcome assessments in N-TA3CT. The Imaging Core Laboratory will provide initial, prompt 
review of each qualifying CT scan to the clinical site and to the Data Coordinating Center to 
confirm or deny each potential study patient’s eligibility on CT scan criteria, and will review 
(post hoc) all CT scans considered in the clinical sites to indicate vascular repair. 
12.1.5 Biomarkers Core Laboratory 
The Biomarkers Core Laboratory will be responsible for measurement of doxycycline 
levels, high sensitivity C-reactive protein levels and MMP-9 activity as well as preservation of 
specimens for future research. Specimens will be processed every 6 months and results will be 
forwarded to the Data Coordinating Center for incorporation into the trial database. The 
doxycycline results will be provided to the Data Coordinating Center for a check on protocol 
adherence. MMP-9 results will be provided to the Data Coordinating Center for incorporation 
into the study database. 
12.1.6 Study Chairman 
Study Chairman responsibilities will include: 1) To provide overall organization of the 
trial; 2) To serve as Chair, Steering Committee; 3) To administer the Treatments Distribution 
Center; 4) To work with the Clinical Sites and Data Coordinating Center to maximize 
collaboration and to arrange study meetings; 5) To provide collaborators with information about 
the progress of the trial; 6) To participate in visits to the Clinical Sites to assess quality and assist 
with problems; 7) To take a leadership role in defining the analysis of study data; and 8) To 
prepare the manuscripts describing the design and results of the study. 
12.2 
ADMINISTRATIVE STRUCTURE 
12.2.1 Steering Committee 
A Steering Committee will be responsible for overseeing the study. This committee will 
make major organizational and policy decisions. The committee will include the Study 
Chairman, Core Laboratory Directors, representatives of the Clinical Coordinating Center and 
the Data Coordinating Center, the Clinical Site directors, and two coordinators representing the 
Clinical Sites to be selected by the other Clinical Site Coordinators and the Clinical Site 
directors. The NIA project officer will be invited to participate. This group will have quarterly 


 
 
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conference calls and meet yearly prior to the annual Clinical Site directors and coordinators 
meeting. 
12.2.2 Management Committee 
A smaller committee involved with the day to day running of the study will have 
conference calls every two weeks to discuss the progress of the trial (including recruitment and 
protocol violations). This group will be composed of the Study Chairman, Director of the Data 
Coordinating Center and the Data Coordinating Center coordinator, Director of the Biomarkers 
Core Laboratory, Director of the Imaging Core Laboratory, and the Clinical Coordinating Center 
head nurse coordinator. 
12.2.3 Publications and Ancillary Studies Committee 
A Publications and Ancillary Studies Committee will be formed from the study 
leadership (Data Coordinating Center, Imaging Core Laboratory, Biomarkers Core Laboratory, 
and Clinical Coordinating Center) and leading Clinical Site directors. This committee will 
review all proposals for and final versions of research abstracts, presentations, and manuscripts 
to be submitted to journals and national meetings. The committee will also review proposals for 
ancillary studies. Since the success of the trial depends entirely upon the collaboration of the 
doctors and nurses in the participating centers, credit will be assigned to them (as the “N-TA3CT 
Research Group”) in the authorship of reports from the study. Each Clinical Site director and 
Clinical Site coordinator will be named personally in an appendix to the main report. 
 
12.2.4 Data and Safety Monitoring Board 
Members of the Data Safety and Monitoring Board (DSMB) will be appointed by and 
report to the National Institute on Aging (NIA) director. They will monitor accruing data in order 
to confirm that the patients in the trial are being cared for safely. The DSMB will be responsible 
for: 
1. Reviewing and analyzing the progress of the study. 
2. Approving amendments to the trial Protocol, if warranted. 
3. Monitoring the safety of the study treatments. 
4. Reviewing data quality. 
5. Reviewing interim analyses and recommending early stopping or continuation of the trial. 
The DCC and the Management Committee provide input to this committee as requested. 
The DSMB will review study data reports every six months and primary end point analysis every 
six months after the first patients complete two years of follow-up. The DSMB may convene in 
face-to-face meetings or on conference calls. 
12.2.5 Role of DCC Staff 
At least one DCC staff member will be assigned to each study committee and 
subcommittee to participate in all meetings and conference calls. The DCC director will serve as 
Executive Secretary of the Steering Committee; Management Committee; Publications and 
Ancillary Studies Committee; Event Classification Committee; and will work with the 
Chairperson of each committee to draft the agenda, summary notes, and lists of action items for 
each meeting or conference telephone call. 
The Executive Secretary of the Publications and Ancillary Studies Committee will 
supervise the development and implementation of a system to track preparation and review of 
study manuscripts and abstracts on main results, databank studies and ancillary studies to 
disseminate the design, methods and results of this trial. 
12.2.6 Data Coordinating Center Interface with Clinical Coordinating Center 


 
 
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Conference calls, facsimile transmission, e-mail and courier will be used to maintain 
frequent and regular communications between the Clinical Coordinating Center and the Data 
Coordinating Center. The investigators will discuss study activities in person or by telephone 
every two weeks during the Management Committee conference calls. These calls serve to 
review study progress and Protocol adherence at the participating Clinical Sites, review 
committee activities, and plan study meetings. In the first months of N-TA3CT, CCC and DCC 
staff will develop training sessions for Clinical Site staff, and in the last months they will lead 
manuscript preparation efforts. Documents and materials for training and draft manuscripts will 
be reviewed with the Management Committee as will plans for long-term archival of study data. 
12.2.7 Data Coordinating Center Role in Training of Study Personnel 
Prior to the start of recruitment and in conjunction with the training session planned by 
the Clinical Coordinating Center, DCC staff will organize training for Clinical Site staff to 
review enrollment of patients using the Interactive Web-based Randomization System (IWRS), 
data collection procedures, submitting forms by electronically to the DCC, and responding to 
edit queries. DCC staff will monitor each Clinical Site to assure that staff at each site have 
completed the required training, and that all other steps necessary to begin participant 
recruitment are completed. Clinical Site staff who have completed training procedures in Clinical 
Sites that are approved to participate in N-TA3CT will be given certification numbers that will be 
used to track the individual responsible for each data item and activity in N-TA3CT. 
 
12.3 
STUDY TIME LINE 
The study will last five years and be broken into the following phases. 1) Planning and 
Organization Phase 1 (months 1-4): Finalize the Manual of Operations and data collection 
instruments. Communicate with the Clinical Site directors at each study site. Obtain IRB 
approval at clinical sites with assistance from CCC staff. Develop computer software for 
randomization and data entry. This time period is feasible because we a) are implementing the 
protocol that was developed under the planning grant, b) the forms we will be using have been 
extensively tested, and c) staff from the CCC will assist Clinical Site staff in completing IRB 
applications. The development of computer software and manual of operations will be addressed 
quickly at the beginning of the study. 2) Planning and Organization Phase 2 (months 5-6): Obtain 
approval of Study Protocol from the DSMB. Communicate final study protocol and procedures. 
Plan and schedule training. Hire Clinical Site Coordinators. Hold annual Collaborators' Meeting 
for Clinical Site directors and coordinators. Distribute final forms. 3) Recruitment and Follow-up 
Phase (second half of Year 1, Years 2, 3, and 4): Initiate patient enrollment and continue for 1.5 
years or until patient recruitment is completed or the study is stopped early. Hold annual 
Collaborators' Meetings. Visit Clinical Sites once during the first year of patient recruitment and 
annually thereafter. CCC prepares and distributes quarterly newsletters. Bi-annual meetings 
(either in person or via conference call) of the Data and Safety Monitoring Board. 4) Close-out 
and Analysis Phase (Last Year): Complete follow-up and data cleaning. Perform data analysis. 
Discuss results at collaborators meeting. Prepare manuscripts and final reports. 
 
Exhibit 12-1 
Clinical Sites Participating in N-TA3CT Planning* 
Aneurysm Patients  
Without Repair 
Meeting Permanent 
Eligibility Criteria 
University of Nebraska Medical Center 
26 
17 
Northwestern University 
35 
18 


 
 
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Washington University School of Medicine 
48 
22 
Emory University School of Medicine 
24 
15 
Geisinger Medical Center 
143 
70 
New York Weill Cornell Medical Center 
28 
21 
University of Utah Health Sciences Center 
31 
15 
University of Arizona Health Sciences Center 
25 
11 
The University of Michigan Health System 
102 
35 
University of Cincinnati School of Medicine 
23 
16 
Oregon Health Sciences University 
19 
9 
Cleveland Clinic Lerner College of Medicine 
171 
89 
University of Maryland School of Medicine 
19 
16 
University of South Florida Health South 
12 
8 
Baptist Hospital of Miami 
24 
9 
Total 
730 
371 
*Other Clinical Sites Involved Include The University Of Virginia and The Beth Israel/Deaconess 
Medical Center (Boston, Massachusetts) 
Exhibit 12-2 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
CHAPTER 13 – POLICY MATTERS 
13.1 
PUBLICATION POLICY  
13.1.1 General Statement of Editorial Policy 
It is anticipated that the Doxycycline Treatment of Abdominal Aortic Aneurysm Clinical 
Trial (N-TA3CT) will generate considerable new data relative to patients with aortic aneurysms. 
The Steering Committee fosters and guides development of scientific reports originating from 
data obtained in the project. The scientific integrity of the project requires that all data from all 
Clinical Sites be analyzed study wide and reported as such. Thus, an individual center is 
N-TA3CT Clinical Trial Organization 
 
DSMB 
Biomarkers Core 
Laboratory 
 
NIA 
Clinical Coordinating  
Center 
 
Clinical Sites 
Pharmacy/Treatment 
Distribution Center 
Data Coordinating 
Center 
Imaging Core 
Laboratory 


 
 
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expected not to report and publish data collected from its center alone. Development of sub 
studies or data bank studies dealing with specific analyses are encouraged. All presentations and 
publications of any data collected by the N-TA3CT Research Group are expected to protect the 
integrity of the main objectives of the overall project. Major findings are not presented prior to 
release of "mainline" results of the study by agreement of all N-TA3CT Clinical Site directors. 
The Steering Committee determines the timing of presentation of mainline results (including 
papers on design and methods) and designation of the meetings at which they might be 
presented. 
Publications are grouped into five general categories (see Section 13.1.2). Topics for 
consideration to be developed into publications are generated from questions or hypotheses that 
are submitted to the Steering Committee by investigators, study coordinators and other study-
related staff. A writing group with a designated Chairperson is selected for each topic. 
The Publications and Ancillary Studies Committee has primary responsibility for 
reviewing and approving all abstracts and all manuscripts on mainline findings, special 
laboratory studies, data bank or ancillary studies submitted for presentation or publication. 
Abstracts and manuscripts are also reviewed by the NHLBI according to existing procedures. 
Investigators at all N-TA3CT Clinical Sites, the Clinical Coordinating Center (CCC), the 
Biomarkers Core Laboratory (BCL), the Imaging Core Laboratory (ICL), and the Data 
Coordinating Center (DCC) have equal status with regard to developing proposals, participating 
in such studies as approved by the Steering Committee, and collaborating in the development 
and publication of research papers based on study material. With the approval of the Principal 
Investigator, study coordinators and other staff at these centers are encouraged to participate in 
this process. The Management Committee has developed standards for regular evaluation of the 
submission and completion of these protocols. The Management Committee determines 
priorities for analyses among data bank study proposals which have been approved. 
N-TA3CT Investigators at Clinical Sites, the BCL, the ICL, the CCC or DCC proposing 
studies that require the collaboration of CCC, BCL, ICL, or DCC (e.g., specimen, image or data 
analysis) contact the appropriate individuals prior to submission of a given proposal. The 
appropriate staff in the CCC and DCC participate in drafting the proposal, indicate willingness to 
participate, and identify sources of funding to support the level of effort required for the project. 
The CCC, BCL, ICL, and DCC Investigators are consulted in the development and 
analysis of protocols that require review of accumulated data or data on file at the CCC, BCL, 
ICL, or DCC. The members of the CCC, BCL, ICL, and DCC collaborate in designing and 
carrying out all N-TA3CT research. 
13.1.2 Types of Research 
Research and the resulting presentations and publications are grouped into the following 
categories:  
1. 
Design paper(s) and reports on methods. 
2. 
Mainline findings. 
3. 
Data bank studies. 
4. 
Ancillary studies. 
5. 
Independent studies. 
Distinctions among these types of studies are given in Section 13.2. If possible, analysis 
of data may be conducted prior to the end of the N-TA3CT investigation and is strongly 
encouraged, so that the maximum information can be published from this study and so that the 
methods for evaluating and analyzing study data may be refined in preparation for later analyses. 


 
 
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13.1.3 Authorship  
The first publication(s) pertaining to the fundamental goals of N-TA3CT involving 
patients enrolled in the study will list the members of the writing team as the authors with “the 
N-TA3CT Research Group" as the last author. An appendix listing the Principal Investigator and 
Co-Investigators will be included at the end of the manuscript's text. It is intended that there will 
be more than one publication concerning the mainline goals; all publications will list the writing 
team as the authors on behalf of the N-TA3CT Investigators. 
13.1.4 Purpose of Procedural Guidelines 
The procedures adopted by the investigators for use of study data are intended to protect 
the interests of all participants in the study, to assure that study data conform to the requirements 
of study design and are accurately presented, that authorship is appropriately acknowledged, that 
the text of each publication is well-written, to ensure that all investigators are aware of ongoing 
analysis projects, to avoid duplication of analysis projects and to ensure that publication or 
presentation of study data does not occur without the knowledge and approval of the Publications 
and Ancillary Studies Committee and the Steering Committee. 
13.2 
DESIGN AND METHODS REPORTS, MAINLINE FINDINGS, DATA BANK, 
ANCILLARY, AND INDEPENDENT STUDIES 
13.2.1 Design Papers and Reports on Methodology 
Manuscripts concerning the overall design, protocol, procedures, or organizational 
structure of the study that do not involve mainline findings or data collected on study patients 
may be published prior to the end of the study. Such preliminary publications will be developed 
and reviewed according to the same guidelines used for other reports of mainline findings. 
Many public presentations about N-TA3CT that do not involve protocol data, special 
laboratory studies, data bank or ancillary study data (e.g., grand rounds talks concerning the 
study's general design and objectives) do not require formal preliminary review and approval by 
the Publications and Ancillary Studies Committee. However, if there is any doubt, investigators 
are asked to first consult with the Publications Committee indicating their intention to present the 
material, in order to avoid the premature release of study data or the inappropriate publication of 
confidential information. 
13.2.2 Reports of Mainline Findings 
A report on mainline findings is one addressing the fundamental goals of N-TA3CT or 
that involves protocol data -- such as changes in aneurysm size or MMP-9 levels over time in 
study patients -- which cannot be released prior to the end of the study. These studies will 
summarize the findings based on the entire study population and will be written at the conclusion 
of the project. These reports must be reviewed and approved by the Publications and Ancillary 
Studies Committee and ratified by the Steering Committee. 
13.2.3 Data Bank Studies 
A data bank study uses data or specimens (including banked specimens) which are 
routinely collected on patients who are recruited and/or enrolled in the N-TA3CT. Analysis of 
these data are used to answer specific scientific questions. Data used in this research are not 
directly related to the fundamental goals of the study. Data bank studies must be approved by the 
Management Committee and ratified by the Steering Committee. All presentations or 
publications of data bank studies are to be reviewed following the procedures outlined in Section 
14.4. 
13.2.4 Ancillary Studies 


 
 
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An ancillary study uses supplementary data that are collected on patients who are 
recruited and/or enrolled in N-TA3CT, over and above the data collection required by the 
protocol. Such studies are restricted to consideration of a specific test technique or involve only 
the supplemental data collected on study cases and controls. Ancillary studies are reviewed and 
approved by the Management Committee and ratified by the Steering Committee prior to 
initiation to ensure that they do not conflict with the main protocol. Review by the Publications 
and Ancillary Studies Committee is required for presentation or publication of an ancillary study. 
13.2.5 Independent Studies 
Independent studies of concern to N-TA3CT are studies conducted in potential patients 
who are not enrolled in the study, but data are collected at a Clinical Site. These data are not 
transmitted to the N-TA3CT Clinical Coordinating Center or Data Coordinating Center. 
It is understood that each Clinical Site has the right to conduct studies which are 
independent of N-TA3CT in patients with who do not meet criteria for enrollment into the study. 
Independent studies of patients who meet eligibility criteria but are not enrolled in N-TA3CT 
must be reviewed by the Management Committee. Study investigators agree not to conduct 
independent studies which would compete with or have a detrimental effect on the conduct of N-
TA3CT during the period of recruitment and follow-up. 
 
 
 
13.3 
GUIDELINES FOR PREPARATION OF PROPOSALS FOR DATA BANK AND 
ANCILLARY STUDIES 
13.3.1 Data Bank and Ancillary Studies 
Each proposal for an ancillary or data bank study should contain a brief description of the 
objectives, methods, analysis plans, significance of the study, and proposed collaborators. Full 
details should be given concerning any procedures to be carried out, such as cytokine 
evaluations, exercise tests or psychological testing, etc. Mention should be made of any 
substances to be injected or otherwise administered to the patients. Any observations to be made 
or procedures to be carried out on patients or on banked specimens outside of the Clinical Site 
should be described. Mention should be made of the extent to which the data bank or ancillary 
study requires extra clinic visits or prolongs the usual clinic visits. Information should be given 
concerning the extent to which the ancillary study requires specimens in addition to those 
presently required by the protocol. If blood specimens are to be obtained from the patients or 
banked specimens are required, mention should be made of the number of specimens as well as a 
description of all procedures to be carried out on these specimens. 
13.4 
PROCEDURES FOR INITIATION AND APPROVAL OF STUDIES 
13.4.1 Reports on Mainline Findings 
Reports on mainline findings from N-TA3CT generally involve the collaboration of many 
investigators. Proposals for these reports are introduced and developed by any N-TA3CT 
Investigator or staff member with the approval of the Clinical Site director. These reports are 
reviewed and approved by the Management Committee and ratified by the Steering Committee. 
13.4.1.1 Submission of Proposals 
Two copies of each proposal should be submitted to the Data Coordinating Center for inventory 
and transmission to the Management Committee. The Director of the DCC notifies the 
Investigator when the project is approved, disapproved or whether additional information is 
needed before a decision can be made. 


 
 
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13.4.1.2 Preparation of Mainline Reports 
 After approval of a proposed topic for a mainline report, members are elected or invited 
to serve on an ad hoc Writing Subcommittee and a Chairperson is chosen. These investigators 
work with the CCC and DCC staff to conduct the data analysis needed to investigate the question 
at hand and prepare a manuscript based on these findings. Every effort is made by the 
Subcommittee to consider and incorporate in this manuscript the comments and suggestions from 
the Steering Committee. Often the Subcommittee members meet with staff from the CCC, DCC 
or other Clinical Sites for development of these papers. 
13.4.1.3 Review and Approval of Abstracts and Manuscripts Prior to Presentation and  
  Publication 
 
Every study manuscript considered suitable for publication is submitted by the 
Chairperson of the Writing Subcommittee to the DCC for distribution to the Publications 
Committee. The Chair of the Publications and Ancillary Studies Committee is responsible for 
arranging and implementing review according to the following procedures. 
1. The manuscript is forwarded promptly to at least two reviewers selected from the 
members of the Steering Committee or their associates, with the request to respond 
within two weeks with a detailed critical review of the manuscript. Outside reviewers are 
selected when appropriate. 
2. Reviews are forwarded to all members of the ad hoc Writing Subcommittee with a 
request for appropriate revision and response. 
3. The ad hoc Writing Subcommittee is expected to respond to the review in a reasonable 
period of time, forwarding to the CCC the revised manuscript and a letter commenting in 
detail on the points raised by the reviewers; DCC staff will distribute these materials to 
the Publications and Ancillary Studies Committee. 
4. After review by the Publications and Ancillary Studies Committee, the DCC staff return 
the manuscript to the ad hoc Writing Subcommittee with final comments or suggested 
changes. 
5. If acceptable to the study leadership (Management Committee), the completed manu-
script is submitted by the Chairperson of the Writing Subcommittee to the appropriate 
journal. A copy of the transmittal letter and copy of the manuscript are submitted to the 
DCC for distribution to the Steering Committee. 
13.4.2 Data Bank Studies 
13.4.2.1 Submission of Proposals 
Data bank studies must be approved by the Management Committee and ratified by the 
Steering Committee. Before beginning a data bank project, a proposal initiated by one or more of 
the Investigators and/or their associates is submitted to the DCC for inventory and distribution to 
the Management Committee for consideration. The Director of the DCC notifies the Investigator 
when the project is approved, disapproved or additional information is needed before a decision 
can be made. 
13.4.2.2 Conduct of Data Bank Studies 
After approval is given by the Steering Committee, the Investigators (on the data bank 
project) work with the CCC and DCC staff to conduct the data analysis. 
13.4.2.3 Priorities for Work 
Because of the routine work load at the CCC and DCC, it is necessary to establish 
priorities for data processing and analysis. Therefore, the DCC staff conduct analyses on data 


 
 
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bank studies in the order in which they have been approved or, as necessary, seek guidance from 
the Management Committee for determining priorities for analysis. 
13.4.2.4 Authorship 
After a data bank study proposal is approved by the Steering Committee, its research and 
development are the responsibility of the identified investigators on the project. Authorship 
decisions on data bank studies take into account the unique cooperative effort that has produced 
the results. For clinical papers in particular, individuals from Clinical Sites, CCC, DCC and NIA 
staff have the opportunity to join writing teams when their contributions are appropriate. On the 
other hand, there will be papers of more limited scope which probably do not warrant a large 
number of authors. The following mechanism is utilized to determine authorship: 
1.  The lead author proposes a list of co-authors, based on the above guidelines. 
2. The Management Committee reviews and approves, or makes recommendations 
regarding alterations in the proposed list of authors. 
The names of these investigators is followed by the designation "and N-TA3CT Research 
Group" on the byline. 
13.4.2.5 Review and Approval of Abstracts and Manuscripts Prior to Presentation or 
Publication 
The Publications and Ancillary Studies Committee reviews all data bank study abstracts 
and manuscripts prior to submission for presentation and publication. Recommendations are 
forwarded to the Management Committee for review and final decision. All abstracts must be 
received by the Publications and Ancillary Studies Committee members, all co-authors, DCC, 
and CCC at least two weeks prior to the submission deadline. Manuscripts prepared based on 
data bank studies must be submitted to the DCC at least one month (30 days) before the 
scheduled submission date. After review, the Publications Committee makes recommendations 
to the Management Committee in consultation with the DCC. The Director of the DCC notifies 
the authors and Steering Committee of the decision within one month of the receipt of a 
manuscript, within one week for abstracts. The approved manuscript or abstract is then 
submitted. 
13.4.3 Ancillary Studies 
13.4.3.1 Submission of Proposals 
Ancillary study proposals are reviewed by the Management Committee and are ratified 
by the Steering Committee to ensure that the proposed study does not conflict with the primary 
goals of N-TA3CT. 
Two copies of each proposal are submitted to the Data Coordinating Center for inventory 
and transmission to the Management Committee. The Director of the DCC notifies the 
Investigator when the project is approved, disapproved or additional information is needed 
before a decision can be made. 
Abstracts and manuscripts are to be submitted for review prior to submission. 
13.4.4 Independent Studies 
Results of independent studies which are approved as acceptable by the Management 
Committee may be published or presented at the discretion of investigators initiating the 
independent study. 
13.5 
RELEASE OF N-TA3CT DATA OR SPECIMENS TO NON- N-TA3CT 
 
INVESTIGATORS 
 
Requests for study results, study data, or banked specimens may be submitted by 
investigators who are not participating in N-TA3CT during the course of this investigation. These 


 
 
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requests will arise primarily from colleagues and researchers who are interested in abdominal 
aortic aneurysm. Each request should be submitted in writing and provide the same information 
as required for study data bank and ancillary studies submitted by N-TA3CT Investigators. The 
Management Committee reviews each request and the following principles are addressed in 
determining the disposition of each request. 
1. Overlap with the study major goals or approved data bank and ancillary studies. 
2. The scientific importance of the request. 
3. The efforts and costs of providing the information. 
4. The willingness of the individuals submitting the request to accept the limitations, as 
specified by the N-TA3CT Management Committee, on the uses that can be made of the 
data and data analysis. 
At least one month prior to the end of funding, the Data Coordinating Center staff will 
prepare data files and appropriate documentation for submission to the NIA Project Office. 
These files will not include personal identifiers of patients. The release of these data tapes will be 
based on the NIH Policy on Release of Data from Large Scale NIH Sponsored Studies. 
13.6 
CONFLICT OF INTEREST POLICY 
13.6.1 General Principles 
The N-TA3CT investigators have agreed to a policy on conflict of interest which has few 
specific restrictions, but a broad indication for disclosure of potential conflicts of interest. The N-
TA3CT investigators wish to endorse the spirit and content of the 21st Bethesda Conference: 
Ethics in Cardiovascular Medicine dealing with these issues, and seek to make this policy 
consistent with the record of that conference. 
To address actual or perceived conflict of interest in N-TA3CT, the participating 
investigators voluntarily agree to abide by the guidelines described in the policy statement 
developed for N-TA3CT. See Exhibit 14-1 for a copy of the Conflict of Interest Statement. 
13.6.2 Individuals to be Governed by These Guidelines 
Members of the N-TA3CT Study Group who will be governed by these guidelines 
include the Study Chairman, the director at each Clinical Site, key personnel in the Clinical 
Coordinating Center, Biomarkers Core Laboratory, Imaging Core Laboratory, and Data 
Coordinating Center. Co-Investigators and other staff who have major responsibility for 
enrollment, recruitment, follow-up or collection of data for N-TA3CT at Clinical Sites will also 
be governed by these guidelines. The Principal Investigator for each N-TA3CT Center or Core 
Laboratory will submit a list of individuals who will be governed by these guidelines at the 
beginning of the study. The leaders of each participating unit will review the guidelines with all 
appropriate staff prior to the start of patient recruitment and at least annually thereafter. 
13.6.3 Time Period of the Policy 
The guidelines set forth in this policy commence at the start of patient recruitment and 
will terminate at the time of initial public presentation or publication of the principal results. 
Investigators not privy to end point data who discontinue participation in the study during 
recruitment will be subject to these guidelines until their departure from the study. 
13.6.4 Financial Guidelines 
1. The investigators agree not to own, buy or sell stock or stock options during the 
aforementioned time period in any of the pharmaceutical companies or related medical 
equipment companies with products used in this study. In addition, the investigators 
agree not to have retainer-type consultant positions or positions of decision making 


 
 
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responsibilities (e.g., board of directors, policy advisory committee), hold patents or 
partnerships with these companies for the time period defined above. 
2. The Clinical Coordinating Center will maintain conflict of interest statements updated 
annually from each investigator. 
Activities not explicitly prohibited, but to be reported annually with information 
maintained by the Clinical Coordinating Center include: 
1. Participation of investigators in authorized educational activities (e.g., FDA approved 
workshops for training in use of devices) that are supported by the companies defined 
above. Note: general speaker bureaus or lecture/ continuing medical education activities 
sponsored by these companies are not allowed. 
2. Participation of investigators in other research projects supported by the companies 
defined above. 
Financial interests in the companies defined above, over which the investigators has no 
control, such as mutual funds or blind trusts are not restricted under these policies. 
13.6.5 Reporting of Financial Disclosures and Other Activities 
The investigators agree to update their financial disclosures and related activities as 
described above on an annual basis and submit these data to the Clinical Coordinating Center for 
storage. The Clinical Coordinating Center staff maintain the confidentiality of these records and 
prepare any reports indicating a potential conflict of interest for review by the Management 
Committee. In the case of actual or perceived conflict of interest, the Study Chairman brings it to 
the attention of the Management Committee, NIA Project Office, and the Data and Safety Moni-
toring Board. 
13.6.6 Review of Policy Statement 
The investigators agree to review these guidelines on an annual basis and take any 
additional steps to insure that the scientific integrity of the study remains intact. 
13.6.7 Relationship to Institutional Policies on Conflict of Interest 
Since existing policies on conflict of interest vary among participating institutions, in 
addition to the above policy, it is expected that investigators comply with the policies on conflict 
of interest which exist within their individual participating institutions (medical schools and 
hospitals). This is the responsibility of each individual investigator. 
13.7 
HUMAN SUBJECTS TRAINING 
 
All N-TA3CT investigators and staff who have any contact with N-TA3CT patients, with 
health care providers treating N-TA3CT patients, with individual patients’ N-TA3CT data or 
specimens must complete approved training in human subjects research and provide 
documentation of current training to the Clinical Coordinating Center annually. 
EXHIBIT 13-1 – Conflict of Interest Statement for N-TA3CT Investigators 
CONFIDENTIAL 
Except as noted below: 
 I am not a part-time, full-time, paid or unpaid employee of any organizations: 
(a) whose products or services will be used or tested in the study under review, or (b) 
whose products or services would be directly and predictably affected in a major way by 
the outcome of the study; 
 I am not an officer, member, owner, trustee, or director of such organizations; 
 I do not have any financial interests or assets in any organizations meeting the above 
criteria, nor do my spouse, dependent children, nor organizations with which I am 
connected. 
 


 
 
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PLEASE COMPLETE THE APPROPRIATE BOX BELOW 
 
NO RELEVANT INTERESTS OR ACTIVITIES. 
 
EXCEPTIONS ARE NOTED IN THE ATTACHED LETTER. 
I will notify the Clinical Coordinating Center Principal Investigator promptly if: 
 A change occurs in any of the above during the tenure of my responsibilities, or 
 I discover that an organization with which I have a relationship meets the criteria for a 
conflict of interest. 
I am aware of my responsibilities for maintaining the confidentiality of any non-public 
information that I receive or become aware of through this activity, and for avoiding using such 
information for my personal benefit, the benefit of my associates, or the benefit of organizations 
with which I am connected or with which I have a financial involvement. 
__________________________ 
________________________ 
________________ 
Investigator (type name) 
            Signature 
 
 
            Date 
 
 
 
 
 
 
 


 
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Appendix – N-TA3CT Procedures and Treatments 
Study Procedures/Treatments 
 
Study Visit 
Baseline 
3 
Months 
± 45 days 
6 
Months 
± 45 days 
9 
Months 
± 45 days 
1 
Year 
± 45 days 
15 
Months 
± 45 days 
18 
Months 
± 45 days 
21 
Months 
± 45 days 
2 
Years 
± 45 daysǂ 
30  
Months 
± 45 days 
Screen for Inclusion/Exclusion 
X 
 
 
 
 
 
 
 
 
 
Consent, Questions, Signature 
X 
 
 
 
 
 
 
 
 
 
Randomization 
X 
 
 
 
 
 
 
 
 
 
Vital Signs 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
Medical Review 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Death 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Compliance 
 
X 
X 
X 
X 
X 
X 
X 
X 
 
AE’s 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Current Therapies 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
SF-36 
X 
 
X 
 
X 
 
X 
 
X 
X 
CT Scan Performed 
X 
 
X 
 
X 
 
X 
 
X 
       X**** 
Send CT Scan To Imaging Laboratory 
X 
 
X 
 
X 
 
X 
 
X 
X 
Biomarkers Core Lab Assays* 
 
 
 
 
 
 
 
 
 
 
Doxycycline Levels 
X 
 
X 
 
X 
 
X 
 
X 
X 
hsCRP's 
X 
 
X 
 
X 
 
X 
 
X 
X 
MMP-9 
X 
 
X 
 
X 
 
X 
 
X 
X 
Cotinine Levels 
X 
 
X 
 
X 
 
X 
 
X 
X 
Interferon Gamma 
X 
 
X 
 
X 
 
X 
 
X 
X 
DNA** 
 
 
X 
 
 
 
 
 
 
 
Local Labs (standard of care)*** 
 
 
 
 
 
 
 
 
 
 
Complete Blood Count*** 
X 
 
 
 
X 
 
 
 
X 
 
Liver Function Tests*** (alanine 
aminotransferase; aspartate 
aminotransferase) 
X 
 
 
 
X 
 
 
 
X 
 
Blood Urea Nitrogen*** 
X 
 
 
 
X 
 
 
 
X 
 
Creatinine*** 
X 
 
 
 
X 
 
 
 
X 
 
Lipid Panel*** 
X 
 
 
 
X 
 
 
 
X 
 
Dispense Emergency Card 
X 
 
 
 
 
 
 
 
 
 
Dispense Study Drug 
X 
X 
X 
X 
X 
X 
X 
X 
 
 
Collect Study Drug 
 
X 
X 
X 
X 
X 
X 
X 
X 
 
Dispense Diary 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
 
* At Baseline and Year 2 visits draw 2 plasma (green) and 2 serum (red) kits. All other visits only require 1 plasma (green) and 1 serum (red) kit.                 
** DNA should be drawn at the 6-month visit. If it is missed it can be drawn at the 1 year or 18 month visit. 
*** These tests may be collected and data entered any time within one year prior to the indicated due date. Please see text on next page for clarification of events. 
**** CT scan if available. 
ǂ With the elimination of 27-month visits the window for the 2-year visit is extended forward to +90 days and for the 30-month visit backward to -90 days to 
maintain contiguity.


 
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Standard of care monitoring for patients who are treated for abdominal aortic aneurysm or coronary artery 
disease or peripheral arterial disease or who are treated with or may be treated with these medications may 
include an annual complete blood count and analyses of circulating blood urea nitrogen (BUN), creatinine, 
liver enzyme and lipid levels. 
Because personal physicians may be ordering the routine laboratory analyses independently of our clinical trial 
and to avoid the possibility of coverage refusals, recover the most recent complete blood count, BUN, 
creatinine, alanine aminotransferase (ALT) or aspartate aminotransferase (AST) and lipid levels that have been 
collected within one year. If lipid analyses have not been performed, the coordinators may let the lipid 
analyses go, and if a complete blood count has been performed but no white blood cell differential (neutrophil 
%, lymphocyte %, eosinophil % and basophil %) the differential cell count can be let go; they are of interest 
and may be data entered but are not essential for safety monitoring by investigators who are not taking 
responsibility for the patients' management outside of N-TA3CT.  
The complete blood count (CBC), BUN, creatinine and ALT or AST levels are necessary for our safety 
monitoring. If they have not been collected within a year prior to expectation, collect blood for these specific 
tests. Obtain a differential white blood cell count along with the CBC. These blood tests may be entered 
anytime within the one-year window. 
 

ICF_004.pdf: 
 
 
DRAFT MODEL 2/3/15 
1 
 
NON-INVASIVE TREATMENT OF ABDOMINAL AORTIC ANEURYSM CLINICAL TRIAL 
(Consent Form) 
1. Nature and Purpose of the Study 
The main artery (aorta) from your heart has a small bulge (aneurysm) in its wall as it 
passes through your abdomen. Because of its location this is called an abdominal aortic 
aneurysm or AAA. Small bulges can stay the same size for a time; most grow slowly; some 
grow quickly. No one knows exactly why there are these differences. When a small bulge is 
present, the usual course is to check its size every 6 months to see if it enlarges to a size that 
should be fixed. If the bulge grows to this larger size, there is a danger it will burst, which would 
be a threat to your life and well being. Your doctors would recommend fixing the bulge if it 
becomes large enough to pose a risk to you. 
The purpose of this study is to learn the effect of a medicine called “doxycycline” on the 
growth of this blood vessel bulge. If the drug slows the growth of the bulge in blood vessels, in 
future years it could be used to prevent the need for surgery or repair to the blood vessel.  
Doxycycline may block an enzyme that acts to take apart proteins in the bulge wall.  
The time it might take for the bulge to reach this large size could be short (6 months) or 
long (years). Because bulge growth varies, it can be hard to tell if a new medicine really works. 
To find out if doxycycline can make the bulge slow its growth, we have to test it along with an 
inactive “look-alike” capsule. Because these bulges typically grow at a slow rate, we will need to 
follow the growth for 2-3 years to see if there is a difference. 
Doxycycline is used to treat other sicknesses. The other sicknesses doxycycline is used 
to treat include rosacea (red face rash), gum loss, and infections. Doxycycline is an FDA 
approved drug for some of these uses, but not for treatment of bulges like the ones we will 
study.  
 


 
 
DRAFT MODEL 2/3/15 
2 
 
There is a lot of work in science labs that shows that the drug we will use in this study is 
good at slowing or stopping the growth of artery bulges in mice and rats. We do not know the 
effect of this drug on small bulges, like yours, in human patients. The tests that have been done 
in people, so far, have not been as detailed as this one. We know of two studies in people.   
• 
In one small study in Finland the drug seemed to slow growth of small bulges in human 
patients, but the doctors who led that study stated that the effect could have been due to 
chance.  
• 
A large, recent study in Holland tested doxycycline in people with both small and large 
bulges. The doctors who led the study report that the bulges grew slightly faster in the 
patients who got the drug instead of the placebo (inactive look alike) with no effect noted 
on rupture or other outcomes that are of concern for the care of patients. 
This study is a clinical trial funded by the National Institutes of Health. About 250 patients 
with small bulges of the main artery are expected to join this study. 
2.   What to Expect 
 
If you agree to join this study you will be asked to take two capsules every day, one in 
the morning and one at night. Half of the patients will get capsules of doxycycline and the other 
half will get a capsule with the inactive “look-alike”. This will be decided by chance, like a lottery. 
Neither you nor your doctor will know which of the two kinds of capsules you are taking. 
However, central study offices will keep a record of what you are taking should your doctors 
need to know this to take care of you. You will be asked to take your capsules twice a day for 2-
3 years or as long as you are in the study. You should not join the study unless you are ready to 
continue for 2-3 years. As long as you are in the study, you will be seen in our clinic every three 
months. If traveling to our offices is a hardship for you, you may ask us to conduct the visits at 9, 
15, 21, 27 and subsequent visits that do not call for a CT scan by telephone. We will record the 
study form parts of these calls for data quality control. Each visit will take about half an hour.  


 
 
DRAFT MODEL 2/3/15 
3 
 
Twice a year we will fill out a form about how much you can do and how well you feel 
and take a special X-ray study to measure the size of the bulge in your aorta, and collect a 
blood sample from a vein in your arm with a needle into a test tube to be sure that your 
treatment is safe or to check your blood for levels of the study drug or learn what the blood 
findings have to do with the growth of blood vessel bulges. Each sample will be no more than 
three tablespoons of blood. We will collect no more than three-quarters of a cup over the course 
of the study, seven blood collections at most. 
 
This blood will be frozen and may be tested later to find out how much study drug or 
what changes due to study drug are seen in any blood. These blood samples will be sent to and 
stored at Washington University in St. Louis. The frozen tubes of blood will not contain your 
name, birth date, or other data that could be used by people outside the study to recognize you.  
 
Dr. ____________ and the study staff will ask about your health in the past. You will be 
asked to sign forms so that we can get records from your private doctor or hospitals where you 
have been treated if we find out that you have been ill while in our study.  
3. Risks of Taking Doxycycline and This Study 
 
The most common undesirable effects seen in patients taking doxycycline are stomach 
and bowel problems such as nausea, vomiting, diarrhea, stomach discomfort or abdominal pain. 
Fewer than 10% of patients will have any one of these problems. Your skin will be more 
sensitive to the sun. So, you may be more likely to get sunburn. You can avoid or reverse most 
of these side effects by taking the medication as instructed and cutting down the amount of sun 
you get. Other less common side effects (1% or less) include tooth discoloration, skin rashes, 
liver injury or changes in blood count. As with any medication, there are very rare (<0.1%) 
problems that could lead to permanent, serious injury. You could have an allergic reaction or 
drug interaction which could be severe. We are going to ask you about the drugs you use before 
starting the study to keep people who take drugs that could cause bad reactions out of the 


 
 
DRAFT MODEL 2/3/15 
4 
 
study. We will review your other treatments with you in the course of this study to avoid drug 
reactions. 
 
As part of this study we will take an X-Ray picture (CT scan) of your aorta every six 
months (at least five and no more than seven times while you are in the study). Some doctors 
take CT scans at six-month intervals on their patients who have bulges in their aorta, others 
take CT scans less frequently or take other pictures that do not use X-rays (for example, 
ultrasound pictures). CT scans are recommended but are not the only choice for taking care of 
patients like you whether or not you are in a research project. The amount of radiation in seven 
CT scans could increase your lifetime risk of developing a fatal cancer by 1/1000. Cancer from 
radiation usually takes 20-30 years to develop.  Among the 250 patients who will join the trial, 
we do not expect to see any one develop a fatal cancer because of this study, but it could 
happen. If you do not want to have any radiation exposure related to research on your condition, 
you should not join this study. Your doctor may choose to use X-ray dye at the time of CT scans 
to see blood vessels better. This study does not ask for your doctors to use dye. If your doctor 
wants to use the dye for you (for example, because he or she thinks that the dye picture will be 
helpful in deciding how to treat you), this study will accept those images. 
Rarely (1% per year) a bulge in an artery will burst even though it is a small bulge. Some 
patients are expected to have this problem whether they join this study, take the study drug or 
look alike, or not. 
You may feel some pain or get a black and blue mark at the site of needle puncture for 
blood sample collection. Very rarely (less than 1% of the time) these puncture sites may 
become infected. 
 
As with studies of all new treatments, there may be other risks to using doxycycline that 
are not now known. If any new bad effects are observed, patients in the study will be told as 
soon as possible; we will take action to protect your safety as needed. 
4.   Benefits 


 
 
DRAFT MODEL 2/3/15 
5 
 
 
You may not receive direct benefit from participation in this study. The treatment used in 
this study may have no effect on growth of the bulge size, may decrease the rate of growth or 
increase it. You would not notice this yourself. Only on medical pictures (for example, X-ray 
pictures) could the change be seen. It may take up to two years to see any effect of being on 
the drug. We will send X-ray reports and any important medical information about you (for 
example, high blood pressure levels) that we found on a clinic visit to the doctors who take care 
of you. At its end, the doctors in this study will be able to use the data they collect from you to 
tell other patients who have small bulges in that artery whether or not taking doxycycline might 
help them. This advice will be useful to these patients, to their families and to their doctors. 
5.   Privacy 
 
In this study only your clinic will know your name. Clinic staff will make note of your age, 
ethnicity, sex, weight, and height.  That information will be stored in the study computer. You will 
not be identified personally in any report from this study. Your personal medical reports will be 
kept private. At the end of the study, a computer file of the study results will be made for future 
use. It will not include any information that could identify you directly. Information may be given 
to the National Institutes of Health, but your name will not be used in such files. 
 A description of this clinical trial will be available on http://www.clinicaltrials.gov, as required by 
U.S. Law. This web site will not include information that can identify you. At most, the web site 
will include a summary of the results. You can search this web site at any time. 
6.   Other Choices 
 
If you join this study, it is your own choice. You may refuse to take part in it or you may 
leave it at any time. The study doctors may decide to stop your study treatment any time they 
think it would be in your best interest or in the best interest of the study. If you do not join, or if 
you leave the study, doing so will not harm your present or future care at the hospital or clinic. 
Instead of taking part in this study you may go to your doctor for your usual treatment. Your 
doctor could add doxycycline to your treatment, but until studies such as this are done, your 


 
 
DRAFT MODEL 2/3/15 
6 
 
doctor will not know whether doxycycline will be helpful to you. There have been other studies 
that have examined the effect of doxycycline on small artery bulges. So far no study has been 
able to be sure whether the drug slows the growth of a small bulge or not. 
7.  Costs Paid for by the Study 
 
You will not be charged for any of the visits that are part of the study and not part of your 
routine care.  You will not be charged for the study drug.  Clinic visits and imaging studies that 
are part of your routine care will be billed to you and your insurance company as has been done 
in the past.  
8.   Costs Not Paid for by the Study 
 
If you need medicine other than doxycycline the study will not pay for it. Your routine CT 
scans are not paid for by the study. If you must visit your private doctor or emergency room, or 
must stay in the hospital, those costs will not be covered by this study. If the bulge in your aorta 
grows to a size for which stent-graft or surgery is the standard of care, your study doctor will 
notify you and your personal doctor. The cost of stent-grafts or surgery will not be covered by 
the study. 
9.  Payments to You 
 
You will be paid at least $40.00 every three months for travel costs. You will receive this 
money from the clinic after each of your study clinic visits. If you do not attend study clinic visits 
or become unable to take part in the study then you will no longer be paid that money. We will 
cover allowable, documented transportation costs that exceed the $10.00 that we anticipated in 
the $40.00 reimbursement. For example, travel of 100 miles by car at a cost approved for one 
recent year by the US government of $0.56/mile amounting to $56.00 exceeds $10.00. We will 
approve and reimburse those excess costs when you give us advance notice and receipts or 
other acceptable information such as a description of your driving route from current home to 
the clinic. In our example, $40.00 + $56.00 - $10.00 = $86.00 would be paid. 
10.  Use of Left Over Blood Samples  


 
 
DRAFT MODEL 2/3/15 
7 
 
   
If you agree, the left over blood sample may be stored and used in future research 
projects to help discover causes of or new treatments for bulges in arteries or other health 
concerns. 
Please initial each one of the following sentences that applies: 
(If you do not initial next to a statement we will assume that your answer is NO) 
_______   I agree to use my blood for future research. 
_______   I do not agree to use my blood for future research. 
I understand that if I do not want my samples used for future research this will not affect my 
ability to take part in this study or in future studies. 
11.  Studying Your Genetic Information  
To collect your DNA or perform genetic studies on your DNA, we need you to give us 
specific consent. There are different risks that go along with studying your genes that do not 
apply to the rest of your participation in the study. You do not have to participate in the genetic 
portion of the study to participate in the other portions of the study. 
Why are we collecting genetic information as part of the N-TA3CT? 
Genes are the basic “instructions” for the development and activities of our bodies. 
Genes are made out of DNA. The DNA of a person is about 99.9% the same as the DNA of any 
other person. But no two people have exactly the same DNA except identical twins. Differences 
in DNA are called genetic variations. They explain some of the differences among people, like 
eye colors and blood groups. They also partly explain why some people get diseases like 
cancer, diabetes, as well as aortic aneurysms. It may also partially explain why some 
aneurysms grow faster than others. 
How will the samples be used? 
The DNA samples will be stored with the other blood samples drawn over the course of 
the study.  We anticipate that researchers will study the samples to find places in the DNA 


 
 
DRAFT MODEL 2/3/15 
8 
 
where people are different. For each sample, this may include information on hundreds of 
thousands (eventually millions) of genetic variations.  
The sample will be linked to other data from the study. All of these data are put in one 
place called a database. All of this information together lets us see patterns that tell the story of 
how genes affect health. This database will include features of your health and your aneurysm 
disease such as the size and growth rate of the aneurysm over the course of the study. The 
final research database will not include any identifiable medical information about anyone who 
gave a sample.  
We may use the data on the differences between the genes of people in this study to 
map out the ways genes connect with each other to do their work. Researchers can then look 
for specific genes and study how they work. This will help them figure out better ways to 
prevent, diagnose, and treat aortic aneurysm disease. They can also learn how to make drugs 
that work better in more people.  
It is possible that studying your genes may reveal information about the risk of 
aneurysms or some other unrelated disease. However, it will be very hard for anyone to learn 
anything about you personally from any of this research because none of the samples or the 
database will include your name or any other information that could identify you or your family. 
What will happen if I decide to give a sample? 
At one of the routine blood draws as part of the N-TA3CT study, we will set aside one 
tablespoon of blood from your arm for DNA tests. It will not require any additional effort on your 
part.  We will send your sample to the N-TA3CT Biomarkers Core Laboratory (BCL) at the 
Washington University School of Medicine in St. Louis [U.S.A.], which is overseen by the 
National Institutes of Health (NIH), the main government agency in the U.S. that funds medical 
research.  
Who will be studying these samples? 


 
 
DRAFT MODEL 2/3/15 
9 
 
Any researchers who will study the samples, data obtained from the samples or data in 
the database will have to follow U.S. laws and guidelines that apply to research. All studies 
using the genetic information will have to be approved by an Institutional Review Board (IRB). 
An IRB is a committee similar to the one that approved the N-TA3CT study to make sure that 
your safety is protected.  Doctors who propose to study the samples must write up their plans 
for review by the leaders of the study. Their plans must be worthy of permission to use the 
samples.  
The results generated by these researchers may be shared, without any identifying 
information, with large national databases of genes and diseases such as the Database of 
Genotypes and Phenotypes (dbGaP) which is maintained by the NIH. 
How will you protect my privacy? 
While [insert name of institution] will keep your signed consent form, nobody else will 
see it. We will not keep your name with your sample or give your sample a code number that 
could identify you. So nobody at the BCL or who studies your sample will know that it came from 
you.  
What are the benefits of giving a sample? 
You probably will not directly benefit from giving a sample, because of the long time this 
research will take to produce results that would change treatment. But researchers will study 
these samples for many years to learn about health and disease. This research will eventually 
benefit the health of people around the world. 
What are the risks of giving a sample? 
The risks at the time of drawing the blood are as described in the main study consent 
form.  If your sample is used for genetic analysis, a large amount of genetic information from 
your sample may be generated, and many researchers may have access to that information. 
However, there are few ways anybody could trace the information back to you. One is if they 
thought your information might be in the database, got another sample from you, did many tests 


 
 
DRAFT MODEL 2/3/15 
10 
 
on that sample, and then compared the genetic information from those tests with the information 
in the database. The other way is if somebody compared genetic information from samples in 
the BCL with genetic information known to be from you that was in another database and 
figured out who you were. The risk of either of these things happening is very small, but it may 
grow in the future. 
We cannot always predict the results of research, so new risks to you may come up in 
the future that we are unable to predict now. Your sample will not be used to make a clone of 
you. 
Can I change my mind after I give a sample? 
Giving a sample is completely up to you. You will not lose any benefits if you choose not 
to give a sample, including the opportunity to participate in all other aspects of the N-TA3CT 
study. However, since nobody will know which sample came from you, after you give a sample 
you cannot take it back or take any information out of the database. 
How will I find out what happens with this project? 
We do not plan to give you individual results from any of the research performed on the 
genetic samples. There will be publication of the results of the studies in peer-reviewed medical 
journals and other publications about what they are learning about health and disease. You can 
obtain copies of these public reports by seeking them out on the Internet or asking the research 
staff here for help finding them. 
Consent and Signature 
Please read the paragraph below, think about your choice, and sign if you agree: 
I agree to give a blood sample to obtain my DNA, for researchers to use for future studies of the 
type described in the form. I have read or listened to the information, I have asked any 
questions I had, and all my questions were answered. I know that giving a sample is my choice. 
I understand that after I give a sample, I cannot withdraw my sample from the Repository. 
 


 
 
DRAFT MODEL 2/3/15 
11 
 
__________________________________________ 
   ___________________________ 
Signature   
 
 
 
 
 
 
    Date 
 I do not agree to give a blood sample to obtain my DNA, for researchers to use for future 
studies of the type described in the form. 
 
__________________________________________ 
   ___________________________ 
Signature   
 
 
 
 
 
 
    Date  
12.  Rights 
 
You will be given a copy of this consent form to keep. If at any time you have questions 
or concerns about the study you may call (give study staff name and telephone number) 
________________, the study staff, or (give IRB contact or  ombudsman name and telephone 
number) ________________, a person whose job is to watch over the well being of patients in 
medical research projects. Should you have any bad effect of treatment or medical emergency 
during the study, care will be provided to you. The cost of treating such bad effects is not 
covered in the study, and no money has been set aside to pay for these bad effects. 
13.  Questions 
 
This study has been explained to you by ____________________ and your questions 
were answered. If you have any other questions about this study you may call Dr. 
___________________ at ____________________. 
 
              Signature 
 
 
 
    Print 
Patient: ______________________ Name: _____________________ Date: ______________ 
Investigator: __________________ Name: _____________________ Date: ______________ 
Witness: _____________________ Name: _____________________ Date: ______________ 

SAP_001.pdf: 
Non-Invasive Treatment of Abdominal Aortic Aneurysm Clinical Trial 
 
N-TA3CT 
STATISTICAL ANALYSIS PLAN 
 
 
 
 
Version:  1.0 
 
Page 1 of 18 
Date:  26-MAR-2017 
 
 
Statistical Analysis Plan for the Primary Endpoint 
Sponsor:  University of Maryland through a grant from the NIH/NIA 
Protocol Title:  Non-Invasive Treatment of Abdominal Aortic     
                                Aneurysm Clinical Trial 
Document Version / Date: 
1.0: 26-Mar-2017 
 
 
 
 


Non-Invasive Treatment of Abdominal Aortic Aneurysm Clinical Trial 
 
N-TA3CT 
STATISTICAL ANALYSIS PLAN 
 
 
 
 
Version:  1.0 
 
Page 2 of 18 
Date:  26-MAR-2017 
 
 
 
 TABLE OF CONTENTS 
1 SIGNATURE .......................................................................................... 4 
2 LIST OF ABBREVIATIONS AND DEFINITIONS OF TERMS .................. 5 
3 INTRODUCTION .................................................................................... 6 
3.1 
PRIMARY AIM:................................................................................................................................................ 6 
DETERMINE IF DOXYCYCLINE (100 MG BID) WILL INHIBIT THE INCREASE IN GREATEST TRANSVERSE DIAMETER OF 
SMALL ABDOMINAL AORTIC ANEURYSMS (3.5-5.0 CM IN MEN, 3.5-4.5 CM IN WOMEN) OVER A 24-MONTH PERIOD OF 
OBSERVATION IN COMPARISON TO A PLACEBO-TREATED CONTROL GROUP. ............................................................... 6 
3.2 
ENDPOINTS ..................................................................................................................................................... 6 
3.3 
STUDY DESIGN ............................................................................................................................................... 6 
3.3.1 
Study population .................................................................................................................................... 6 
3.3.2 
Treatment groups and dosing ................................................................................................................ 7 
3.3.3 
Study visits and assessments, study baseline ......................................................................................... 7 
3.3.4 
Randomization and Blinding or Masking .............................................................................................. 7 
3.3.5 
Sample Size and Power .......................................................................................................................... 8 
4 GENERAL CONSIDERATION FOR DATA ANALYSES ........................... 8 
4.1 
ANALYSIS POPULATIONS ................................................................................................................................ 8 
4.1.1 
Intention to Treat ................................................................................................................................... 8 
4.1.2 
Full Analysis Population ....................................................................................................................... 8 
4.1.3 
Per-Protocol Population ........................................................................................................................ 9 
4.2 
STATISTICAL ANALYSIS ISSUES ...................................................................................................................... 9 
4.2.1 
Strata and Covariates ............................................................................................................................ 9 
4.2.2 
Multiple Comparisons............................................................................................................................ 9 
4.2.3 
Multi-center Studies ............................................................................................................................... 9 
4.2.4 
Study Baseline and Visits ....................................................................................................................... 9 
5 INTERIM ANALYSIS AND DATA SAFETY AND MONITORING BOARD
 
10 
6 SUBJECT DISPOSITION ..................................................................... 10 
6.1 
DISPOSITION OF PARTICIPANTS .................................................................................................................... 10 
6.2 
EXTENT OF EXPOSURE .................................................................................................................................. 10 
6.3 
PROTOCOL VIOLATIONS ............................................................................................................................... 10 
7 BASELINE DATA ................................................................................. 11 
7.1 
DEMOGRAPHICS AND BASELINE CHARACTERISTICS ..................................................................................... 11 
8 EFFICACY ANALYSES ........................................................................ 11 
8.1 
PRIMARY EFFICACY ANALYSIS .................................................................................................................... 11 
8.1.1 
Imputation for the Primary Efficacy Analysis ...................................................................................... 12 
8.2 
ANALYSIS WITHOUT IMPUTATION ................................................................................................................ 15 
8.3 
ANALYSIS OF PER-PROTOCOL POPULATION ................................................................................................. 15 
8.4 
CHANGES FROM PROTOCOL-SPECIFIED EFFICACY ANALYSES ...................................................................... 15 


Non-Invasive Treatment of Abdominal Aortic Aneurysm Clinical Trial 
 
N-TA3CT 
STATISTICAL ANALYSIS PLAN 
 
 
 
 
Version:  1.0 
 
Page 3 of 18 
Date:  26-MAR-2017 
 
 
9 INTERPRETATION OF THE PRIMARY OUTCOME .............................. 15 
10 
SAP REVISION.................................................................................. 18 
 
 
 


Non-Invasive Treatment of Abdominal Aortic Aneurysm Clinical Trial 
 
N-TA3CT 
STATISTICAL ANALYSIS PLAN 
 
 
 
 
Version:  1.0 
 
Page 5 of 18 
Date:  26-MAR-2017 
 
 
 
 
2 List of Abbreviations and Definitions of Terms 
AAA: Abdominal Aortic Aneurysm 
ANCOVA: Analysis of covariance 
BMI: Body Mass Index 
CRP: C-reactive protein 
CT scan: Computerized tomography scan 
DCC: Data Coordinating Center 
DSMB: Data Safety and Monitoring Board 
GEE: Generalized Estimating Equations  
ICL: Imaging Core Laboratory 
IFN-γ: Interferon gamma 
MMP-9: Matrix metallopeptidase 9 
MTD: Maximum Transverse Diameter 
SAP: Statistical Analysis Plan 
SF-36: The Short Form (36) Health Survey   
 
 
 
 
 
 
 


Non-Invasive Treatment of Abdominal Aortic Aneurysm Clinical Trial 
 
N-TA3CT 
STATISTICAL ANALYSIS PLAN 
 
 
 
 
Version:  1.0 
 
Page 6 of 18 
Date:  26-MAR-2017 
 
 
 
 
 
 
3 Introduction 
This Statistical Analysis Plan (SAP) describes the analyses to be conducted for the primary 
endpoint of the Non-Invasive Treatment of Abdominal Aortic Aneurysm Clinical Trial (N-
TA3CT).   
3.1 
Primary Aim:  
Determine if doxycycline (100 mg bid) will inhibit the increase in greatest transverse 
diameter of small abdominal aortic aneurysms (3.5-5.0 cm in men, 3.5-4.5 cm in women) 
over a 24-month period of observation in comparison to a placebo-treated control group. 
3.2 
Endpoints 
The primary outcome is the change in abdominal aortic aneurysm maximum transverse 
diameter (MTD) on CT scan from baseline to the follow-up assessment CT scan two years after 
randomization, as measured in the Imaging Core Laboratory (ICL). 
Secondary outcomes will derive from central ICL analyses of the CT scans performed every six 
months on patients and from the clinical follow-up of randomized patients, from clinical 
observation, local laboratory findings, study visit quality of life assessments, and from 
biomarker analyses to be performed in the Biomarkers Core Laboratory (e.g., changes from 
initial MMP-9 levels, and MMP-9 levels at 24 months of follow-up). 
3.3 
Study Design 
3.3.1 
Study population 
The study population will include patients 55 years of age or older who are found to have 
small (3.5-5.0 cm among men and 3.5 - 4.5 cm among women in the largest transverse 
diameter) abdominal aortic aneurysms on quantitative CT scans. 
Patients will be excluded from the study if they are unable to give their own informed 
consent to participate; have symptoms related to abdominal aortic aneurysm; have other 
intra-abdominal vascular pathology that may require repair within 24 months (e.g., renal 
artery stenosis, large iliac artery aneurysms, iliac occlusive disease, aneurysmal 
involvement of the renal artery); have had previous abdominal aortic aneurysm repair by 


Non-Invasive Treatment of Abdominal Aortic Aneurysm Clinical Trial 
 
N-TA3CT 
STATISTICAL ANALYSIS PLAN 
 
 
 
 
Version:  1.0 
 
Page 7 of 18 
Date:  26-MAR-2017 
 
 
open surgical or endovascular technique; have an active malignancy with life expectancy 
less than two years; have an allergy to tetracycline; are currently or have been recently 
treated (previous six months) with tetracycline derivatives; they are currently taking anti-
seizure medicines metabolized by pathways influenced by doxycyclines (e.g., 
carbamazepine, phenytoin, and barbiturates); stage II hypertension (patients whose blood 
pressure is persistently in the range of systolic > 160 mm Hg or diastolic > 100 mm Hg 
despite primary physician’s best effort to achieve adequate therapy); have dialysis 
dependent renal failure or impending dialysis treatment for renal insufficiency; have a 
chronic infection requiring long-term (> 2 weeks) antibiotics, have known genetic 
syndromes responsible for the abdominal aortic aneurysm (e.g., Marfan’s Syndrome), are 
under treatment with systemic immunosuppressive agents, could become pregnant, are 
not good candidates for clinical trial participation or are enrolled in another clinical trial. 
3.3.2 
Treatment groups and dosing 
Patients will be assigned to doxycycline 100 mg p.o., b.i.d., or matching placebo. Patients 
randomly assigned to doxycycline will receive bottles containing a sufficient supply of 100 mg 
doxycycline capsules to take one capsule twice a day until the next appointment (about 100 
days). Patients randomly assigned to placebo will receive a similar appearing supply of placebo 
capsules. 
3.3.3 
Study visits and assessments, study baseline 
Under the original protocol, participants were to be followed in-clinic every 3 months until a 
common termination when the last participant randomized had been followed for 24 months.  
Under Protocol Revision 1.6.0, participants are expected to have 10 visits for this study: one 
baseline, eight at quarterly intervals while on treatment, and one post-treatment follow-up.  
Follow-ups will be conducted as in-clinic visits except that at 9, 15 and 21 months they may be 
conducted by telephone. The appendix of the protocol contains a summary of the timing of the 
visits and the assessments and data that are collected at each visit. 
3.3.4 
Randomization and Blinding or Masking 
The DCC staff prepared randomization schedules for each Clinical Site participating in N-
TA3CT. The program for generating randomization schedules has the following characteristics: 
1. There will be two randomization strata – one for men and one for women – at each clinical 
site; 2. Treatments are assigned in random order within blocks sizes two, four, six or eight with 
equal numbers of patients assigned to the doxycycline or placebo within each block. 3. Block 
sizes (two to eight patients per block) are randomly selected with the probability of each block 
size specified by DCC staff. 
The DCC staff maintained a web-based randomization system for Clinical Site staff to request 
treatment allocations as eligible patients are identified. Only study-certified clinic staff with 
individually assigned userid/password can access the randomization and drug dispensing 


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application.  At randomization, after selecting the participant ID to randomize, clinic site staff 
are asked to enter the participant’s “letter code”, gender and date of birth.  These are verified 
against the data entered into the electronic data capture application for this participant.  Only 
participants meeting the inclusion/exclusion criteria according to data entered in the eDC 
system, with a CT scan read by the Imaging Core Laboratory confirming eligibility, and with 
complete baseline data, including record of collection of the baseline biomarker sample are 
listed among the participant IDs eligible for randomization. The date and time of the completion 
of the randomization treatment assignment is the time of study entry for each patient. 
In the Clinical Sites, Clinical Coordinating Center, Imaging Core Laboratory and Biomarkers 
Core Laboratory, the patients, directors (i.e., Clinical Site lead investigator), clinical site 
coordinators, and other study staff involved with direct patient care will be blinded to treatment 
assignments. Staff of the Data Coordinating Center will have access to individual patient 
treatment assignment on a “need-to-know” basis. The DSMB, the Director of the Data 
Coordinating Center will remain blinded to individual participant treatment assignment and to 
by-treatment-arm interim results.  Only Data Coordinating Center staff involved in the 
production and distribution of the Closed Report to the DSMB or development and maintenance 
of the randomization system will have access to treatment code assignments and by-treatment-
arm results.   
The Data Coordinating Center will maintain records of each patient’s treatment assignment. 
3.3.5 
Sample Size and Power 
The study is designed to have greater than 90% power with a total sample size of 248 patients 
for a one-sided 0.025 test for an assumed reduction of 40% in the mean increase in aneurysm 
diameter from 2.5 mm/year to 1.5 mm/year.  The calculations allow for a 10% non-adherence 
rate in the doxycycline treatment group and for 15% of participants to have a missing two-year 
CT scans.  Additional details regarding the sample size and power calculations are contained in 
the protocol. 
4 General Consideration for Data Analyses 
4.1 
Analysis Populations 
4.1.1 
Intention to Treat 
The intention-to-treat population includes all patients randomized, analyzed by treatment 
assigned at randomization.   
4.1.2 
Full Analysis Population 
The full analysis population includes all patients randomized except those with no follow-up 
contact and no follow-up CT scan, analyzed by treatment assigned at randomization.  Because 


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treatment was never started and because the primary endpoint is not known and cannot be 
reasonably imputed in participants with no follow-up and no follow-up CT scan, analysis of the 
primary endpoint will be conducted on the full analysis population.  
4.1.3 
Per-Protocol Population 
A per-protocol population will be defined as all participants randomized who  
1. Complete 24 months of follow-up with a 2-year CT scan available for analysis; 
2. Or undergo repair within 24 months (+ 90 days) of randomization; 
3. Or die or suffer rupture of their AAA within 24 months (+90 days) of randomization; 
4. And reported to have taken at least 80% of their expected study drug doses over the 
course of their follow-up period. 
4.2 
Statistical Analysis Issues 
4.2.1 
Strata and Covariates 
Randomization is stratified by gender and site.  Gender will be included in the primary efficacy 
analysis model.   
4.2.2 
Multiple Comparisons 
For the primary analysis, we consider a one-sided p-value of 0.0247 to be significant.  It is less 
than 0.025 to account for the interim analyses.  To account for the multiplicity of hypotheses 
being tested in secondary and exploratory analyses, a p-value < 0.01 will be required to consider 
there to be evidence of differences present, i.e. to reject the null hypothesis.  We consider 
p<0.001 as strong evidence. 
4.2.3 
Multi-center Studies 
There are a total of 22 sites participating in the study.  Analyses will not adjust for site. 
4.2.4 
Study Baseline and Visits 
Study baseline corresponds to measurements collected at the baseline visit, prior to 
randomization and initiation of treatment.    Follow-up visits are scheduled based on the date of 
randomization and are expected to occur 45 days before or after the target time-point; 90 days 
are allowed for CT scans or primary outcome clinical events (i.e., death or surgical repair).   


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5 Interim Analysis and Data Safety and Monitoring Board 
Two formal interim analyses will be performed: one when approximately the first third of 
outcome data is available and the next when approximately the second third of outcome data is 
available.  Formal rules for stopping for efficacy, futility, or harm are described in the protocol. 
In addition, at each DSMB meeting an analysis of six-month CT scan results will be presented.  
The DSMB may recommend early termination if the aneurysm growth is higher in patients 
receiving doxycycline than in placebo recipients, with a one-sided p-value less than or equal to 
0.05 (Z = 1.645). 
6 Subject Disposition 
6.1 
Disposition of Participants 
A CONSORT diagram (https://www.consort-statement.org/consort-statement/flow-diagram)  
will depict the disposition of all participants screened and randomized. 
6.2 
Extent of Exposure 
Compliance with assigned treatment arm will be assessed by capsule counts at each follow-up 
visit.   For the full analysis set, for each treatment arm and visit, the percentages of patients who 
have taken greater than or equal to 80% of the expected capsules, based on the capsule count, 
will be reported. 
Plasma samples will be collected approximately every six months.  For patients assigned to 
doxycycline, these samples will be analyzed for measurement of doxycycline levels. At each of 
the four six-month time points, the percentages of patients on the doxycycline arm with detected 
doxycycline will be reported, along with mean, standard deviation, median and interquartile 
range of the doxycycline values. 
Time to permanent discontinuation of study drug will be depicted by Kaplan-Meier plot 
comparing the two treatment arms. 
6.3 
Protocol Violations 
Protocol violations will be reported for the full analysis population using counts and percentages 
of violations. The difference in count between the intention to treat population and the full 
analysis population will be presented.  


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7 
Baseline Data 
7.1 
Demographics and Baseline Characteristics 
Baseline characteristics by treatment group for the intention-to-treat population will be 
compared using counts, percentages and for continuous variables, descriptive statistics. The 
following characteristics will be reported: gender, age (mean, standard deviation, median and 
interquartile range), ethnicity, race, maximum diameter and volume of AAA separately by 
gender (most recent prior to baseline with mean, standard deviation, median and interquartile 
range), vital signs including systolic and diastolic blood pressure, height, weight and BMI by 
gender, medical histories, concomitant medications, baseline quality of life as measured by the 
SF-36 Quality of Life instrument, and baseline biomarker levels.  Characteristics will be 
reviewed for the full analysis population as well.  
8 Efficacy Analyses 
8.1 
Primary Efficacy Analysis 
The primary analysis will be performed to test the null hypothesis of no difference in growth of 
abdominal aortic aneurysms between the two treatment groups (doxycycline and placebo) after 
24 months of follow-up as measured by CT scans analyzed in the Imaging Core Laboratory. This 
analysis will use the full analysis population. In order to include patients whose 24-month 
abdominal CT scans are missing because of death, endovascular repair or for other reasons in the 
primary analysis, we will base our analysis on normal scores for percentile of rank status. We 
will assign worst ranks to deaths, in order of time from study entry to death, next worst ranks to 
rupture, next worst ranks to endovascular repair for clinical presentation, next worst ranks to 
endovascular repair for reasons of aneurysm growth beyond 5.5 cm in men or 5.0 cm in women 
or other presentation free of clinical indication, all in order of time from study entry within 
category.  The order of ranks are summarized in Table 1.  Clinical presentation will be assessed 
by the three vascular surgeon Principal Investigators, blinded to treatment assignment, 
independently; agreement by two of the three or all three will be taken as final. Outcomes for 
patients missing the 24-month CT scan will be imputed, as described in the next section. 
 
Table 1. Summary of ranks. 
Order of rank 
assignment 
(worst to best) 
Description 
1 
Death (from day of study entry to day of death) 
2 
Aneurysm rupture (from day of study entry to day of repair) 
3 
Endovascular repair for reasons related to clinical presentation (from day of 
study entry to day of repair) 


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4 
Endovascular repair for growth beyond 5.5 cm in men or 5.0 cm in women or 
other presentation free of clinical indication (from day of study entry to day 
of repair) 
5 
Change from baseline in largest, transverse diameter of the aneurysm, using 
normal scores as described below. 
 
The ranks for patients assigned to doxycycline will be compared to the ranks of patients assigned 
to placebo using normal scores. If N is the total number of randomized patients, a rank of k will 
be assigned the value corresponding to k/(N+1) in a standard normal distribution; for example, a 
rank of 100 out of 248 patients will receive the rank score corresponding to 0.4016, or a normal 
score z = 0.2492.  
 
The change in normal score from baseline -- i.e., the two-year normal score for patients 
randomized to doxycycline and placebo minus the baseline normal score -- will be analyzed 
using a linear regression model (analysis of covariance -- ANCOVA), with variables for the 
randomized treatment assignment (0 for placebo, 1 for doxycycline), the normal score for 
baseline diameter, and gender in the model. 
 
Prior to performing the primary outcome analysis, an interaction term for the randomization 
stratum (gender) with treatment assignment will be assessed in the ANCOVA model. If there is 
significant (at two-sided α=0.05) evidence of differences in the effect of treatment according to 
gender, other than a small quantitative difference (less than half a normal score difference in 
treatment effect) between effects that are in the same direction for each gender, the analysis will 
be performed separately for men and for women.  In the event of a treatment by gender 
interaction being observed, we will review study results by gender to determine the nature of the 
differences in treatment effects. If there is no interaction, the ANCOVA will be used to estimate 
the overall treatment effects across all clinical sites and both strata (genders). The primary 
analysis will be performed on the normal scores corresponding to percentile rank of each patient 
at baseline and follow-up. 
 
8.1.1 
Imputation for the Primary Efficacy Analysis 
In the event a patient does not complete two years of CT scan follow-up for cause(s) other than 
death, rupture, open repair or endovascular repair (e.g., withdrawal of consent or loss to follow-
up, expected to be infrequent events), the patient’s previous CT scans and other characteristics 
will be used for the imputation of a measurement of the abdominal aortic aneurysm diameter to 
be included in the primary analysis.  The model used for imputation will be developed as 
described below. 
 
For the multiple imputation of missing 24-month CT scans, we first assume that event status is 
known for all participants.  Therefore, we assume we are imputing the 24-month CT scan score 
only for patients who did not have an event.  First, we will assess the 24-month CT scans of 
patients who did not have an event for normality and transform them if necessary.  If the data 


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show egregious departures from normality, possible transformations are natural log 
transformations, square root transformations, or Box-Cox transformations.  Second, we will 
model the MTD from the 24-month scan using fully conditional specification, a type of multiple 
imputation using chained equations (MICE), implemented with SAS 9.4’s PROC MI under the 
FCS option.  At a minimum, we will include MTD from the baseline CT scan, gender, treatment, 
and gender by treatment interaction, and MTD from CT scans obtained at 6-month, 12-month 
and 18-month follow-ups.  The interaction of treatment by gender is included because the test of 
interaction is planned in the final analysis.   
 
Additional variables that will be considered for the multiple imputation model are baseline 
history of coronary artery disease, peripheral vascular disease, cancer, chronic obstructive 
pulmonary disease, diabetes mellitus, hypercholesterolemia, baseline use of beta blockers, ACE 
inhibitors or angiotensin receptor blockers, calcium channel blockers, diuretics, non-steroidal 
anti-inflammatory drugs, use of aspirin or anti-platelet, adherence score, and current smoking 
status.  Adherence scores over 100 will be censored to 100. A continuous variable will be added 
to the multiple imputation model if its Type III Wald Chi-Square test is significant at the 0.05 
level in linear regression of the MTD from the 24-month CT scan after adjusting for baseline 
MTD, gender, treatment, and gender by treatment for complete cases.  Alternatively, if a 
continuous variable’s partial correlation with 24-month CT scan is above 0.4 or above 0.75*R2 
after adjusting for baseline MTD, gender, treatment, and gender by treatment interaction, then it 
will be added to the multiple imputation model.  The R2 is the correlation of 24-month CT scan 
result with baseline MTD, gender, treatment and gender by treatment interaction.   
 
To be considered for the imputation model, a categorical variable will be required to have at least 
5 of each group in the data set.  Any of the above listed categorical variables that is missing in 
20% or more in the imputed cases will be excluded.  If the final list of variables includes more 
than 3 different types of medications, the list will be sent to the Director for the Data 
Coordinating Center for a check of collinearity between variables on a biological basis. This 
information and consultation will in no way unblind the Director of the Data Coordinating 
Center to the results of the analyses. If there are collinear variables, only the variable with the 
higher partial correlation will be included in the modeling process. 
 
Finally, using complete cases (i.e., cases with no missing values for the 24-month MTD or any of 
the candidate independent variables), we will start with a model for 24-month MTD that includes 
all the candidate independent variables and perform backward elimination to test whether or not 
there are variables that can be deleted. The initial model will include MTD from the baseline CT 
scan, 6-month scan, 12-month scan and 18-month scan, treatment, gender, treatment by gender 
variable, and any other variable that meets the above statistical requirements. Variables other 
than baseline MTD, gender, treatment and gender by treatment interaction will be included in the 
final imputation model if they have a p-value less than 0.05 after the backward elimination.   
  
The process of choosing candidate variables and developing a final model for imputing 24-
month MTDs will be implemented before each interim analysis of the primary endpoint, as well 


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as at the final analysis.  Thus, variables that are used for the multiple imputation at one analysis 
may or may not be used at a different stage, as data on more participants are available. 
 
Data from study participants who have a clinical endpoint event (death, rupture, or repair) before 
the 6-month follow-up CT scan will not be included in the fitting of the model for imputing the 
24-month MTD for other participants.  Data from participants who have an event after the 
earliest significant follow-up MTD will be included in the fitting of those models, but their rank 
for the primary analysis will be determined by the event they experienced – i.e., their data will be 
used in imputing the 24-month MTD for other participants, but their clinical event will be used 
in determining their rank in the primary analysis.  Data from participants excluded from the 
multiple imputation modeling will be added back into the imputed data set, so that they will be 
included in the primary analysis.   
 
In the imputation model, the MTDs from CT scans at 6, 12, and 18 months will be constrained to 
have a minimum value of 3.5 and a maximum value of 7, to reflect the plausible range of MTD’s 
in participants who do not experience a clinical event.  The maximum value for MTD’s was 
purposefully set high in order to not constrain the within and between imputation variance. All 
imputed MTD values will be rounded to the nearest hundredth (0.01) to reflect the same 
accuracy as the observed values. 
 
For the multiple imputation regression modeling, we will use the default SAS settings (linear 
regression for continuous variables, logistic regression for dichotomous variables, and 
discriminant function for nominal variables with more than 2 categories).  Since we are using the 
default, there is no need to specify a method on the FCS statement of PROC MI. The default 
number of burn-in iterations of 10 will be used (NBITER=10).  Variables other than final and 
intermediate follow-up MTDs will be listed on the VAR statement from most complete to least 
complete. MTDs from follow-up CT scans will be listed in chronological order, regardless of 
missingness, with MTD at 24 months listed last. A seed of 384830 will be used, and 100 imputed 
values will be created for each missing observation in the multiple imputation model.   
 
The multiple imputation data set will be used to test the significance of both the treatment 
variable and the treatment by gender interaction.  For both the test of the treatment by gender 
interaction and the subsequent test of treatment effect or treatment and treatment by gender 
interaction, the degrees of freedom will be adjusted.  The adjusted degrees of freedom is a 
function of the complete-data degrees of freedom and the between-imputation and within-
imputation variance estimates.  In the test of treatment by gender interaction, the complete-data 
degrees of freedom is the number of subjects in the analysis minus one.  If the test of interaction 
is not significant at the 0.05 level, then we test only the treatment effect and the complete data 
degrees of freedom is the same (number of subjects minus one).  If the test of interaction is 
significant at the 0.05 level, then we test the treatment and treatment by gender interaction, and 
the complete-data degrees of freedom is the number of participants minus two, as we are 
estimating two parameters in this case. 
 
 


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8.2 
Analysis Without Imputation 
Using the same normal scores for percentiles of rank methodology described above (section 
7.1), we will compare the treatment groups without imputation of any missing 24-month CT 
measurements of the MTD of the aneurysm. 
 
8.3 
Analysis of Per-Protocol Population 
An analysis will be performed using the methodology described above for the primary endpoint, 
based on the per-protocol population. 
8.4 
Changes from Protocol-Specified Efficacy Analyses 
The primary efficacy analysis population has been renamed the “full analysis population” rather 
than the “intention-to-treat population” to avoid dispute over whether exclusion of patients for 
whom there are no follow-up visits or follow-up CT scans represents an imperfection in an 
“intention-to-treat” analysis plan or a different study population.  
9 Interpretation of the Primary Outcome 
Interpretation of the primary outcome analysis must be cognizant of N-TA3CT as a Phase IIb 
clinical trial. The primary outcome is based primarily on anatomical measurements instead of 
clinical events. Benefit in the primary outcome could influence physician behavior for off label 
use, but is not going to result in a labeled indication. In particular it cannot be expected that we 
will know whether or not doxycycline effects on abdominal aortic aneurysm diameter are 
associated with the occurrence of rupture or the diameter at which risk of rupture outweighs the 
risk associated with repair. Effects on rupture or mortality could be observed fortuitously, and 
would have to be interpreted as secondary outcomes.   If the investigators reject the null 
hypothesis, they have a considerable responsibility to assure that they have not introduced bias  
and explain to the medical community their basis for believing that bias  is not the reason for 
rejecting the null hypothesis. If the investigators fail to reject the null hypothesis, they have a 
considerable responsibility to assure that they have performed a sensitive test, i.e., not introduced 
bias through defects in study design (especially power calculation assumptions) and performance 
(especially adherence or missing data), and explain to the medical community their basis for 
believing that bias is not the reason for failing to reject the null hypothesis. 
 
 
Interactions: 
With gender: If there is a statistically significant interaction (p<0.05) between the effect of 
doxycycline in men and the effect in women, there will be a search for explanations for the 
interaction. For example, adherence will be examined. The gender strata will be analyzed 
separately in the event of an important interaction. If there are important, non-significant 
differences between the gender strata, e.g., no effect in one stratum and a 40% effect size in the 


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other, there will be a search for explanations for the differences between the strata, and the 
separate strata will be analyzed according to the standards of secondary analyses.  
With Clinical Site: Influence of clinical site on treatment effects will be inspected. If there are 
outlier clinical sites, explanations for the inhomogeneity will be sought (e.g., differences in 
adherence) and any differences or inhomogeneities found will be presented with the primary 
analysis. 
 
The findings of the primary outcome analysis in the full analysis population may reject or fail to 
reject the null hypothesis. Rejection of the null hypothesis with a median growth >40% less in 
the doxycycline group than in the placebo group would be evidence of both a statistically 
significant and clinically significant effect.  
 
Results of analysis of the Full Analysis Population Conflict with Results from the Per-
Protocol Population 
If the primary outcome analyses based on the full analysis population and per protocol 
population are in agreement rejecting the null hypothesis or failing to reject the null hypothesis, 
we will reject or fail to reject the null hypothesis accordingly. If the analysis based on the full 
analysis population fails to reject the null hypothesis but the analysis based on the per protocol 
population rejects the null hypothesis, we will interpret the findings to indicate that doxycycline 
may be but has not been demonstrated to be efficacious for the reduction of growth of small 
abdominal  aortic aneurysms. N-TA3CT  would be inconclusive with regard to treatment effect 
but may be informative as a Phase IIb clinical trial with regard to defining the defects in 
performance, e.g., insufficient treatment adherence, that must be addressed in designing a Phase 
III clinical trial. If the full analysis population analysis leads us to reject the null hypothesis and 
the per protocol population analysis leads to an effect size of similar magnitude to the full 
analysis population analysis but does not reject the null hypothesis, we will conclude that the per 
protocol analysis population analysis was insensitive and reject the null hypothesis. If the full 
analysis population analysis and the per protocol population analysis are contradictory – i.e., no 
similarity of effect size and no agreement on rejection of or failure to reject the null hypothesis – 
we will examine the results closely for an explanation of these pathological findings. 
 
If we reject the null hypothesis with ≥ 40% less growth in doxycycline group: 
• An analysis without imputation will be performed. If the results of the analysis without 
imputation do not return an effect size similar to the primary outcome analysis, the 
rejection of the null hypothesis will be called into question as too highly dependent on 
imputation.  
• Also, a per protocol analysis with an effect size similar to or larger than 40% would be 
supportive of the interpretation that a clinically significant effect was observed in the full 
analysis population, but if the per protocol analysis effect size were less than 40%, the 
clinical significance of the finding should be viewed as questionable.  
 
If we reject the null hypothesis with a median growth less than 40% reduced in the 
doxycycline group as compared to the placebo group:  


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• Should the per protocol analysis be associated with an effect size greater than 40%, there 
could be reason for belief that a clinically significant benefit exists that would justify 
further investigation.  
• Also, rejection of the null hypothesis with an effect size less than 40% but a large 
difference (e.g., doubling) of the proportion of patients whose aneurysms grew less than 
1.0 mm or a large reduction (e.g., halving) of the proportion of patients whose aneurysms 
grew more than 6 mm would suggest the possibility of clinically significant benefit that 
would justify further investigation.  
 
An alternative interpretation to be kept in mind whenever the null hypothesis is rejected is the 
occurrence of a difference by chance.  
 
Failure to reject the null hypothesis: 
Failure to reject the null hypothesis in the primary outcome analysis would be interpreted to 
mean that doxycycline did not have a beneficial effect size >40% or that the design assumptions 
and performance were not of the quality necessary for a definitive test of the null hypothesis.  If 
the effect size is >40% but we do not reject the null hypothesis, we will examine the design 
assumptions and study performance carefully to report any deficiencies that may influence future 
investigations. 
 
 
 


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10 SAP Revision 
Identify major changes.  Only changes after sign-off need to be recorded.   
Revision Date  
Section 
Summary of Revision Reason for Revision 
 
 
 
 
 
 
 


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1 Signature: 
 
 
 
William C. Blackwelder, Ph.D. Senior Biostatistician 
Date 
 
 
 
Diana Lam, Ph.D. Biostatistician 
Date 
 
 
 
David Prince, Ph.D. Biostatistician 
Date 
 
 
 
Michael L.Terrin, MD, MPH, Director Data Coordinating Center 
Date 
 
 
 
B. Timothy Baxter, MD, Study Principal Investigator 
Date 
 
 
 
Jon Matsumura, MD, Director Imaging Core Laboratory 
Date 
 
 
 
John Curci, MD, Director Biomarkers Core Laboratory 
Date 
 
 
 
Ruth McBride, Director Data Management Unit 
Date