Conditions: Acute Coronary Artery Syndrome
Keywords: plaque LDL statin endothelial apheresis acute coronary artery syndrome
Allocation: RANDOMIZED | Intervention model: PARALLEL
Enrollment: 270
Healthy volunteers: no

Brief summary:
The purpose of this randomized, multi-site, clinical trial is to determine whether intensive therapy consisting of cholesterol-lowering statin drugs plus apheresis to cleanse the blood of low-density lipoprotein (LDL) cholesterol is more effective than statin therapy alone in reducing plaque volume in heart arteries of patients who have already suffered an acute coronary syndrome (ACS). The study will also investigate whether this intensive approach can help increase the presence of endothelial progenitor cells (EPC), stem cells that have been shown to reduce cardiovascular (CV) events in ACS patients. This study has two phases and FDA approval for phase II has been received and all information has been updated to reflect PREMIER Phase II.

Detailed description:
Using statins to lower blood cholesterol, and specifically LDL, is well established as a long-term strategy to reduce CVs and even death. But the most intensive pharmacologic lipid-lowering therapy with statins, though proven superior to standard dose regimens, is still associated with an unacceptably high rate of recurrent CV events early after an ACS. This study hypothesizes that for ACS patients undergoing percutaneous coronary intervention (PCI), intensive lipid-lowering therapy consisting of statins and LDL-apheresis (ILLT) will significantly reduce the total coronary atheroma volume of vulnerable plaque and augment mobilization of peripherally circulating EPC colony forming units, compared to guideline statin monotherapy (SMT). ILLT will lead to fewer CV events for these patients.

Patients presenting at four VA sites with ACS will be screened and consented before undergoing uncomplicated PCI (balloons or stents) and intravascular ultrasound with virtual histology (IVUS-HS). They will then be randomized into the ILLT arm or SMT arm of the study. The ILLT group will receive one treatment of LDL-apheresis plus a daily oral 40- 80mg dose of Atorvastatin or equivalent statin; the SMT group will only get 40-80mg Atorvastatin or equivalent. Patients will again undergo IVUS-HS 12 weeks after enrollment to measure atheroma volume; EPC level will also be checked.

The three-year duration of the study includes 24 months of accrual, six months of follow-up, and 12 months of study closure and data analysis. A two-sample t-test of mean difference with 90% power and 0.65 Cohen's D effect size provides a total sample size estimate of 102. Counting 20% drop-out rate, the sample size increases to 128.

The recent FDA recommendations regarding the design of the study have been included in the revised study protocol:

1. The safety data will be submitted to the FDA.
2. Patients will be randomized to both LDL-apheresis and an oral daily dose of 40-80mg Atorvastatin or equivalent statin (Intensive LDL-lowering therapy/ILLT) or a daily dose of 40-80mg of Atorvastatin or equivalent statin without LDL-apheresis (standard statin monotherapy/SMT) following an uncomplicated PCI.

Arms:
arm 1: Patient of acute coronary syndrome (ACS) undergoing percutaneous coronary intervention (PCI) is randomized to LDL-apheresis and an oral daily dose of 40-80mg of Atorvastatin or equivalent. arm 2: Patient of acute coronary syndrome (ACS) undergoing percutaneous coronary intervention (PCI) is randomized to an oral daily dose of 40-80mg of Atorvastatin or equivalent without LDL-apheresis

Interventions: intervention 1: intensive LDL-lowering therapy intervention 2: standard statin monotherapy
Intervention descriptions: intervention 1: The intensive LDL-lowering therapy uses LDL-apheresis in addition to the standard statin therapy. The device used in this study is the LIPOSORBER LA-15 System, manufactured by Kaneka Pharma America LLC. A filter separates plasma from whole blood, the Liposorber -columns remove LDL from the plasma. The system recombines plasma and blood cells and returns them into the patient's body. This procedure typically takes about 3 hours. The procedure provides an immediate reduction in a patient's lipid levels. A single apheresis treatment can lower LDL by more than 80%, but levels return to baseline within 3 weeks. intervention 2: The standard statin therapy of 40-80mg oral daily dose of Atorvastatin or other equivalent types of statin to lower LDL in blood for both randomized groups.

Protocol document:
Prot_000.pdf: 
 
PREMIER Pivotal Study Protocol Version 2.4 – 06/22/16 
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Plaque Regression and Progenitor Cell Mobilization with  
Intensive Lipid Elimination Regimen  
(PREMIER) 
Version 2.4 (Pivotal Study) 
 
June 22, 2016 
 
 
 
 
 
 
 
 
 
 
 
Principal Investigator: 
Subhash Banerjee, MD 
Dallas VAMC 


 
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1. 
Rationale 
 
1(a) 
Background 
The most intensive pharmacologic lipid lowering therapy with statins, though 
proven superior to standard dose regimens, is still associated with an unacceptably high 
rate of recurrent cardiovascular (CV) events early after an acute coronary syndrome 
(ACS). Progression or rupture of lipid rich necrotic core (NC) elements of atherosclerotic 
vulnerable plaque (VP) leads to a majority of recurrent CV events. Vascular healing by 
endothelial progenitor cells (EPC) plays a crucial role in repair following ischemic injury 
primarily by endothelialization of VP and neovascularization of ischemic myocardium. In 
fact, EPC mobilization while on statin therapy has been shown to enhance coronary 
blood flow in patients with stable coronary artery disease (CAD), and reduce myocardial 
ischemia and CV events in patients with ACS within a few weeks of treatment. This has 
prompted a continuous drive towards lowering of total cholesterol and specifically low-
density lipoprotein (LDL). However, what still remains uncertain is whether the most 
intensive LDL-lowering therapy (ILLT) with LDL-apheresis could lead to a rapid and 
detectable reduction in VP atheroma volume, along with a more robust EPC 
mobilization compared to standard statin therapy in ACS patients. 
 
 
1(b) 
Hypothesis 
We hypothesize that in ACS patients undergoing percutaneous coronary 
intervention (PCI), ILLT with LDL-apheresis plus statin therapy will significantly reduce 
the total atheroma volume of VP and augment mobilization of peripherally circulating 
endothelial progenitor cell colony forming units (EPC-CFU/ml), compared to guideline 
based standard statin monotherapy alone (SMT). We also hypothesize that the total 
number of and the percentage of patients with major peri-PCI procedure adverse events 
will not be a safety concern for the study. 
 
 
1(c) 
Specific Objectives 
This is the second phase of a multi-center trial of ACS patients. The first phase 
was a safety study involving 31 patients and was completed in December 2012. US 
Food and Drug Administration (FDA) reviewed the safety and efficacy data from the first 
phase and approved proceeding to this phase. In this trial, 128 participants will be 
randomized in a 1:1 ratio to either initial LDL-apheresis and an oral daily dose of 40-
80mg of Atorvastatin or equivalent (ILLT group) vs. a daily dose of 40-80mg of 
Atorvastatin or equivalent without LDL-apheresis (SMT group) following an 
uncomplicated PCI. Intravascular ultrasound with virtual histology (IVUS-VH) derived 
coronary atheroma volume and composition will be obtained at baseline and 90 days 
after enrollment, while peripheral blood sampling will be performed at enrollment, and at 
24 hours, 30 days, and 90 days post-PCI, along with a four-month and a six-month 
clinical follow-up visits to determine whether the ILLT group will affect: 
 
1. The total number of and percentage of patients with major peri-PCI procedure 
adverse events (primary safety endpoint) 


 
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2. The total number of and percentage of patients with statin-related abnormal liver 
function test events and statin-related muscle injury events (secondary safety 
endpoint) 
3. The total atheroma volume within a ≥ 20 mm long segment of the target coronary 
artery at 90 days IVUS-VH follow-up (primary effectiveness endpoint) 
4. The %NC component within a ≥ 20 mm long segment of the target coronary 
artery at 90 days IVUS-VH follow-up (secondary effectiveness endpoint) 
5. The EPC-CFU/ml of peripheral blood, compared to SMT group from baseline to 
30 days and 90 days post-PCI (secondary effectiveness endpoint) 
6. The major adverse CV events (MACE) at 90 days and at six months follow-up 
(secondary effectiveness endpoint) 
 
 
 


 
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2. 
Background and Significance 
 
2(a) 
Background 
2(a)(i). Lowering blood cholesterol with statins is well established as a long-term 
strategy to reduce death and ischemic CV events in patients with stable CAD1-3. Major 
mechanisms by which lipid lowering is thought to improve outcome include preventing 
the development of new atherosclerotic lesions and stabilizing existing atherosclerotic 
plaques4. In addition, statins can reduce vascular inflammation5, decrease platelet 
aggregability and thrombus deposition6, and increase endothelium-derived nitric oxide 
production7. Most recently, statins have been reported to promote the 
neovascularization of ischemic tissue in normocholesterolemic animals by up regulating 
EPC mobilization and differentiation8. 
 
2(a)(ii). Aggressive lipid lowering pharmacotherapy with statins, though proven 
superior to standard dose regimens, still is associated with an unacceptably high rate of 
recurrent CV events (28.8%)1,9. Moreover, intensive statin regimens are significantly 
underutilized (5%) in ACS patients undergoing PCI, who are at highest risk for recurrent 
ischemic events10. ACS patients, who survive through the presentation, have a high rate 
of recurrent ischemic events, early after an index event3. This has prompted a 
continuous drive towards lowering of LDL goals. 
 
2(a)(iii). Current analysis of statin studies also cannot definitively determine 
whether the benefit seen with high-dose statins is because of the high statin dose used 
or because low LDL levels are achieved. Our study will provide crucial information in 
this area by comparing ILLT which achieves a rapid drop in LDL levels using initial LDL-
apheresis, followed with 40-80mg daily Atorvastatin or equivalent to SMT with 40-80mg 
daily Atorvastatin or equivalent. This has particular relevance in the context of our 
recent publication that showed ACS patients lack the ability to mobilize EPC in 
response to an endovascular injury11. Thus, up-regulation of EPC mobilization with ILLT 
in ACS patients may define a novel mechanism (Figure 1) and therapeutic strategy to 
improve neovascularization of ischemic myocardium, and endothelialization of VP and 
endovascular stent prosthesis, especially drug-eluting stents. Circulating EPC levels 
also correlate to CV risks and predict CV events12. 
 
2(a)(iv). Clinical studies like the Myocardial Ischemia Reduction with Aggressive 
Cholesterol Lowering (MIRACL) trial have demonstrated that intensive cholesterol 
lowering with Atorvastatin administered immediately after hospitalization for ACS rapidly 
reduced the incidence of recurrent ischemic events over the first 16 weeks13. The 
Pravastatin or Atorvastatin Evaluation and Infection Therapy–Thrombolysis in 
Myocardial Infarction 22 (PROVE IT–TIMI 22) trial demonstrated that among patients 
who have recently had an ACS, an intensive lipid-lowering statin regimen provides 
greater protection against death or major CV events than does a standard regimen9. It 
showed a 16% reduction in the hazard ratio favoring intensive therapy. These early 
clinical benefits have been accompanied by only modest changes in intra-coronary 
plaque burden and are thought to be due to the atheroprotective property of mobilized 
EPC14,15. This early effect of statin treatment on clinical endpoints can be partly 


 
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explained by a 1.5-fold increase in the number of circulating EPC within one week after 
initiation of statin treatment, followed by sustained increased levels to three-fold 
throughout the four-week study period in patients with stable CAD16. There are currently 
no studies of EPC mobilization in ACS patients with intensive lipid lowering therapy. 
 
 
Figure 1: Proposed Study Mechanism 
 
                  
 
 
2(a)(v). Coronary atherosclerosis imaging: Although trials using clinical endpoints 
are the gold standard for evaluating novel atherosclerosis treatments, they are lengthy 
and resource-intensive. Surrogate endpoints, such as coronary angiography and IVUS-
VH, allow a more cost-effective evaluation of novel treatments. IVUS-VH is a novel 
technology which allows for the identification of discrete plaque components using 
radiofrequency backscatter data17. IVUS can visualize the coronary artery wall and 
measure atherosclerosis volume. The reliability and reproducibility of total atheroma 
volume by traditional grey scale IVUS have been well validated, and we have published 
the first report on reproducibility of repeated IVUS-VH measurements in a clinical 
practice setting32. We reported excellent agreement between the two pullback 
measurements for lumen area vessel area and plaque burden. The Spearman rank-
order correlation coefficients were (0.96, 0.96, and 0.95) for lumen area, vessel area, 
and plaque burden, respectively. (The data are presented in section 3(d) under "Work 
Accomplished”). 
IVUS has already been used as an endpoint in long-term trials (REVERSAL, or 
Reversal of Atherosclerosis with Aggressive Lipid Lowering18 and ASTEROID, or Study 
to Evaluate the Effect of Rosuvastatin on Intravascular Ultrasound-Derived Coronary 


 
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Atheroma Burden19) and in short-term IVUS follow-up trials with Apo-A1 Milano20 and 
delipidated high density lipoprotein (HDL) treatments21. Currently there are two clinical 
studies that have evaluated early (≤12 week) IVUS derived coronary atheroma 
regression and have demonstrated significant regression from baseline with pre-beta 
HDL21 and ApoA-1 Milano20 infusions. The change in atheroma volume in the control 
arm was 2.8mm3 (increase) and that in the treatment arms were -12.18mm3 and -
14.1mm3 with pre-beta HDL and ApoA-1 Milano therapies respectively. We also expect 
a robust response to ILLT with LDL-apheresis at 90 days. Our estimates of effect have 
been described in detail in Section 4(g) under "Biostatistical Considerations." 
 
2(a)(vi). Endothelial progenitor cells play a crucial role in repair following 
ischemic injury22. This is achieved primarily by neovascularization and endothelialization 
of vulnerable atherosclerotic plaque. Thus, augmentation of circulating EPC in the 
setting of an ACS may significantly contribute to endothelialization of coronary VP, and 
limit myocardial ischemic injury22. Increasing the number of circulating EPC by 
transplantation of hematopoietic stem cells or by injection of in vitro differentiated EPC 
has been shown to improve neovascularization of ischemic hind limbs23, accelerate 
blood flow in diabetic mice24, and improve cardiac function25. Therefore, augmentation 
of circulating EPC in the setting of an ACS may significantly contribute to the stimulation 
of neovascularization after tissue ischemia. Recently, EPC mobilization with statin 
therapy has not only been shown to rapidly enhance coronary blood flow in patients with 
stable coronary artery disease26, but also to reduce myocardial ischemia in patients with 
ACS within a few weeks of treatment27, along with a significant increase in circulating 
EPC. The current studies however, cannot definitively determine whether the EPC 
mobilization seen with statins is because of the statin dose used or the degree of LDL 
reduction achieved. 
 
2(a)(vii). LDL-apheresis is an extracorporeal blood processing system to acutely 
remove LDL-cholesterol from the plasma28. The approved LIPOSORBER® system will 
be used for the study. It contains dextran sulfate cellulose beads that selectively bind 
Apo-B containing lipoproteins (Lp(a), LDL and VLDL). During this procedure, plasma is 
separated from whole blood, LDL cholesterol removed from the plasma, and plasma 
and blood cells are recombined and returned to the patient28. LDL-apheresis treatment 
can lower LDL cholesterol levels by 85% after a single treatment. In familial 
hyperlipidemia (FH) patients, this significant lowering of the LDL cholesterol level is not 
maintained due to a metabolic defect that causes the overproduction of LDL cholesterol. 
The LDL cholesterol level begins to increase (or rebound) after treatment, eventually 
returning to baseline in about one to three weeks, requiring recurrent treatments28. We 
do not expect non-FH ACS patients to rebound rapidly, especially on continued statin 
therapy. We therefore propose using only one initial treatment with LDL-apheresis to 
achieve a rapid reduction in LDL-cholesterol, with follow-up guideline based lipid 
lowering pharmacotherapy with statins. 
LIPOSORBER® treatment is very selective in removing LDL cholesterol, and 
does not affect HDL and triglyceride (TG) levels. A mean reduction in plasma proteins 
(15%), fibrinogen (29%), and platelets (5%) may be observed and rarely pose a risk to 
the patient28,29. Approximately 300,000 treatments with the LIPOSORBER® system 


 
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have been performed worldwide on over 2,500 patients. Possible adverse reactions 
include hypotension, nausea/vomiting, flushing, chest pain, fainting, lightheadedness, 
anemia, abdominal discomfort, numbness/tingling, tachycardia, headache, shortness of 
breath, hemolysis, bradycardia, itching/hives, bleeding (due to the use of unfractionated 
heparin) and chills28. Hypotension is the most common, and in US trials, occurred in 
<1% of all treatments and was treated effectively with intravenous fluids. If an ACE 
(angiotensin converting enzyme)-inhibitor is used to manage the patient's blood 
pressure; it should be held 36 hours prior to LDL-apheresis. 
A single treatment with LDL-apheresis has been demonstrated to have significant 
effect on arterial blood flow. Tamai et al. demonstrated that a single LDL-apheresis 
treatment increased nitrous oxide production and forearm blood flow29. A single 
treatment with LDL-apheresis has also been shown to significantly reduce modified LDL 
particle, remnant-like particle-cholesterol, and C-reactive protein30. In the LACMART 
study, one LDL-apheresis treatment over one year lowered the LDL of FH patients 
aggressively and demonstrated a significant reduction in the plaque area and increase 
in the minimum luminal diameter (MLD) with an annual angiographic and IVUS follow-
up31. It also demonstrated that the benefits of LDL-apheresis are substantial when used 
in the setting of a statin pharmacotherapy. It is important to point out that in the FH 
population, the LDL cholesterol level in the medication-only arm did not change 
significantly from baseline (174±39mg/dl to 181±53mg/dl), and with LDL-apheresis it 
decreased from 213±25mg/dl to 140±27mg/dl, which is much higher than the expected 
post-LDL-apheresis LDL levels expected in non-FH ACS patients. The details of 
atheroma regression are described in section 4(g), “Biostatistical Considerations”. 
Thus, we believe that the proposed ILLT strategy to jumpstart aggressive LDL 
reduction post-ACS is expected to have significant pathophysiologic and biochemical 
effects, in addition to significant atheroma regression expected with nearly 85% 
reduction in serum LDL levels. Though the vasoactive effects of LDL-apheresis are well 
known, its effect on EPC mobilization has never been studied. Our proposal will for the 
first time test this hypothesis and lay the foundation for a dedicated mechanistic study in 
the near future. 
 
 
2(b) 
Relevance to Veterans Health 
We believe that our proposed strategy would provide the basis for a large-scale 
study to evaluate clinical endpoints as a primary objective and ultimately lead to the 
development of a new strategy to attain more aggressive therapeutic lipid goals, and 
reduce CV events post-ACS, reduce the socioeconomic burden of ischemic heart 
disease, and improve the quality of life of our veterans. 
 


 
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3. 
Work Accomplished 
This proposal is a logical extension of our ongoing work. We have: 
i. Reported for the first time, failure of ACS patients to mobilize EPC in 
response to PCI in contrast to those with stable coronary lesions who 
demonstrate a robust EPC response to endovascular injury 
ii. Successfully isolated and identified EPC-CFU from peripheral blood 
samples of patients undergoing PCI 
iii. Performed LDL-apheresis in patients with CAD 
iv. Performed and reported IVUS-VH validation study to detect coronary 
atheroma volume and its components 
v. Demonstrated clinical trial experience by completing and publishing the 
SOS (Stenting of Saphenous vein graft) multi-center randomized trial with 
12-month angiographic and IVUS follow-up 
 
 
3(a) 
EPC Study in ACS 
We have reported that EPC levels rise in response to PCI in patients with stable 
coronary lesions. In contrast, patients with unstable coronary lesions or ACS failed to 
mobilize EPC in response to PCI (Figure 2). 
These data, the first in humans undergoing PCI, are consistent with a protective 
role for EPC mobilization. In this study, we investigated the effect of a discrete 
endovascular insult on human EPC recruitment into the peripheral circulation11. By 
focusing on a defined clinical procedure, we were able to compare EPC levels before, 
and early after the manipulation at reproducible time points, avoiding possible 
nonspecific perturbations. 
 
Figure 2: Baseline EPC levels and mobilization after PCI 
                                
 
  


 
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3(b) 
EPC Isolation 
We have successfully isolated and identified EPC from peripheral blood samples 
of patients undergoing PCI using: 
i. Cell culture technique (Figure 3) 
ii. Flow-cytometry (Figure 4) 
 
3(b)(i). EPC assay, using the cell culture method: EPC form typical colony 
forming units morphologically, and transform into cobblestone monolayer after 2-3 
weeks (Figure 3a-b). Surface markers characteristic for EPC are von Willebrand Factor 
(vWF+), CD31+, CD34+, CD45- (Figure 3c-f)11. 
 
 
Figure 3 (a-f): Confirmation of EPC-CFU phenotype by cell culture 
 
 
3(b)(ii). Using flow cytometry, we have also measured and confirmed isolated 
EPC in peripheral blood samples. The results of such an analysis in two patients 
recently performed in our laboratory are shown below (Figure 4), and demonstrate 
CD34+, CD133+, and CD45 weakly+ circulating EPC, similar to those identified by cell 
culture technique (purple) (unpublished data). 
 
 
 
 
 
 
 
 
 


 
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Figure 4: Circulating EPC Levels by Flow Cytometry 
 
 
 
 
3(c) 
LDL-apheresis in Patients with CAD 
Our group, under the directorship of Dr. Ravindra Sarode (Director LDL-
apheresis center, University of Texas Southwestern Medical Center, Dallas, TX), has 
performed 12 uneventful LDL-apheresis sessions in two patients with CAD. One patient 
underwent LDL-apheresis every week, and the second every two weeks. Average 
reduction of LDL was approximately 84% after each session. Both patients have also 
been maintained on low dose statin therapy. 
From our experience in the first phase of the PREMIER trial, there was one 
transient hypotension episode recorded in 20 patients randomized to LDL-apheresis 
after PCI for ACS. The event resolved spontaneously and was attributed to micturition. 
 
3(d) 
Identification of Coronary Atheroma with IVUS-VH in ACS Patients 
undergoing PCI 
The principal investigators (PI's) perform over 2,300 cardiac catheterizations and 
500 coronary interventions annually, of which >80% are IVUS-VH guided. Coronary 
angiographic image acquisition, quantitative coronary angiographic analysis (QCA) 
using the Phillips proprietary software is performed by the PI’s. The quantitative IVUS-
VH image acquisition, live measurements of vessel diameter, cross-sectional area, 
plaque volume, and plaque composition assessments (QIVUS) are made by the PI’s32. 
Figure 5 demonstrates typical QIVUS assessments and measurements made during a 
typical study. These procedures can be performed reproducibly and safely by the PI’s. 
The overall annual complication rate of our cardiac catheterization laboratory is 1.6%, of 
which 85% consist primarily of minor access site complications. 
 


 
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Figure 5: Representative IVUS-VH frames from the present study demonstrating 
use of side branches to match regions of interest between successive pullback 
trials 
 
 
 
 
The reliability and reproducibility of total atheroma volume by traditional grey 
scale IVUS has been well validated, and the PI's have published the first report on 
reproducibility of repeated IVUS-VH measurements in a clinical practice setting32. We 
reported excellent Bland-Altman plots demonstrating reproducibility data for mean 
vessel area (a), lumen area (b), fibrous tissue (c), fibrofatty tissue (d), necrotic core (e), 
and dense calcification (f), respectively, agreement between the two pullback 
measurements for lumen area vessel area, and plaque burden (data not shown). The 
Spearman rank-order correlation coefficients were (0.96, 0.96, and 0.95) for lumen 
area, vessel area, and plaque burden respectively. 
VH analysis Spearman rank-order correlation coefficients between pullback trials 
for the fibrous, fibrofatty, necrotic core, and dense calcium measurements were (0.97, 
0.90, 0.90, 0.90) respectively and indicated a high level of reliability (data not shown). 
Coefficients ranged from 0.90 to 0.97 (p<0.0001). The Bland-Altman plots (Figures 6a-
f, above) indicated proportional error for the differences of the four measurements 
between the pullback trials. Accordingly, the CR values were less than z=1.96, ranging 
from 0.33 to 1.66. 
 
 
 
 


 
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Figures 6(a-f): Bland-Altman plots indicated proportional error for the differences 
of the four measurements between the two pullback trials 
 
 


 
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4. 
Work Proposed 
The study was proposed to be carried out in two phases. The first phase was a 
safety study, in which 31 ACS patients were randomized to either ILLT (device) group or 
SMT (control) group with a randomization scheme of 2:1 (20 ILLT patients and 10 SMT 
patients). After all 30 out of 31 randomized ACS patients in the first stage completed the 
6-month follow-up visit, the study data (for both safety and efficacy) was submitted to 
FDA for review. FDA approved continuation to the second phase for the pivotal study, 
which will include 128 ACS patients randomized to either ILLT (device) group or SMT 
(control) group with a randomization scheme of 1:1 (64 in ILLT device group and 64 in 
SMT control group). 
 
4(a) 
Study Objectives 
The safety objective of this study is to collect additional safety data for ACS 
patients who undergo the LDL-apheresis post-PCI procedure and the use of maximum 
dose of statin drugs.   
The primary effectiveness objective of this study is to compare the total atheroma 
volume within a ≥ 20 mm long segment of the target coronary artery in ACS patients 
with an uncomplicated PCI randomly assigned to LDL-apheresis with 40-80mg daily of 
Atorvastatin or equivalent (ILLT group) or 40-80mg daily of Atorvastatin or equivalent 
only (SMT group). The specific hypothesis being tested is: ILLT is superior to SMT in 
reducing the total atheroma volume at 90 days IVUS-VH follow up. 
The secondary effectiveness objectives of this study will be: (1) to compare %NC 
component of atheroma within a ≥ 20 mm long segment of the target coronary artery; 
(2) to compare the EPC-CFU/ml of peripheral blood in the two treatment groups; and (3) 
to compare MACE, associated with the two treatment groups at 90 days and at 6 month 
follow-up. 
Rationale for study effectiveness objectives: The rationale for the study 
effectiveness objectives includes the observation of increased recurrent CV events in 
ACS patients early (three months) post-index event, despite intensive statin 
pharmacotherapy, due to progression and rupture of non-critical coronary lesions. 
Aggressive LDL lowering using LDL-apheresis (ILLT) as proposed in this study will test 
a novel strategy of non-critical coronary atheroma regression at 90 days, and hence 
reduce early (12-week) MACE events in ACS patients, sustained at end of study (at 
least six months follow-up). We also propose to test whether EPC mobilization with 
aggressive LDL lowering correlates with coronary atheroma regression, change in 
atheroma composition (%NC), and MACE events. We acknowledge that the scope of 
this study does not test the mechanism of EPC targeting to site of coronary VP and its 
endothelialization, however, it would provide for the first time important initial evidence 
which could be tested in future dedicated studies. 
We expect to demonstrate a significant reduction in atheroma volume at 90 days 
by IVUS-VH (primary endpoint) because this study will have the most aggressive LDL 
lowering strategy in ACS patients ever tested in non-FH ACS patients. The 12-week 
time period is also the time point when early post-ACS CV events begin to increase, as 
seen in the PROVE IT–TIMI 22 trial9. Currently there are two clinical studies that have 
evaluated early (≤12 week) IVUS derived coronary atheroma regression and have 
demonstrated significant regression from baseline with pre-beta HDL21 and ApoA-1 


 
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Milano20 infusions. The change in atheroma volume in the control arm was 2.8mm3 
(increase) and that in the treatment arms were -12.18mm3 and -14.1mm3 with pre-beta 
HDL and ApoA-1 Milano therapies respectively. We also expect a robust response to 
ILLT with LDL-apheresis at 90 days. Our estimates of effect are much more 
conservative and have been described in detail in Section 4(g) under "Biostatistical 
Considerations". 
 
 
4(b) 
Study Outcome Measures 
The primary safety outcome measures will be the total number of and percentage 
of patients with major peri-PCI procedure adverse events, such as hypotension, angina, 
myocardial ischemia, myocardial infarction (if the patient is determined to have had 
unstable angina rather than non-ST-elevation MI at admission), cerebrovascular event (CVA), 
vermicular tachycardia, bleeding (at the PCI access and apheresis cannulation sites in the 
apheresis patients and at the PCI access site in the non-apheresis control patients), and 
death. The peri-PCI procedure is defined as encompassing the time of the PCI procedure 
and the time of the subsequent LDL-apheresis procedure for ILLT group vs. the time of the 
PCI procedure for SMT group. All LDL-apheresis-related adverse events in the ILLT group, 
including any minor expected events, will be recorded. 
The secondary safety outcome measures will be the total number of and percentage 
of patients with statin-related abnormal liver function test events and statin-related muscle injury 
events, which could occur due to the maximum dose of statin drugs being given to both the 
ILLT group and SMT group patients. The statin-related muscle injury is defined as a muscle 
injury which cannot be attributed to a non-statin cause and which is evidenced by symptoms 
(muscle soreness, pain, or tenderness) and/or lab tests such as serum total creatine 
phosphokinase (CPK), CPK-MM, or myoglobin. 
The primary effectiveness outcome measure will be the change in the total 
atheroma volume within a ≥ 20 mm long segment of the target coronary artery from 
baseline to 90 days post-PCI. The measurement will be done via IVUS-VH at both 
centers of investigation.  
The secondary effectiveness outcome measures will include the %NC 
component of atheroma, EPC-CFU/ml of peripheral blood, and major adverse 
cardiovascular endpoints (MACE) including death, myocardial infarction, coronary 
revascularization, and stroke during the follow-up periods. The cell culture assay and 
quantification of circulating EPC-CFU will be done for patients recruited at the Dallas VA 
center. The major adverse CV events will be collected via both clinical visit and 
searches of the Austin database for VA hospital utilization by Hines staff. Approvals for 
using the patients’ Social Security numbers for data searching will be obtained. 
 
 
4(c) 
Study Design and Procedures 
The proposed study is a multi-center, randomized controlled study comparing 
atheroma volume, %NC component, and peripherally circulating EPC-CFU/ml of blood 
in ILLT vs. SMT groups of ACS patients undergoing PCI (Figure 7). All patients will sign 
an informed consent during the index hospital admission for ACS but prior to any study-


 
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related procedure. The protocol will be approved by each site's institutional review 
board. 
 
Figure 7: Study design overview for primary endpoint: 
 
           
                
 
 
4(c)(i). Subject Screening Criteria (Figure 7, green arrow): Patients presenting 
with an ACS, LDL >70mg/dl on Atorvastatin ≤80mg or equivalent dose of another statin, 
normal liver function test (LFT) and referred for non-emergent cardiac catheterization 
(the procedure is not required to be performed within 3 hours after patient presentation) 
will be screened and consented during the index hospital admission for ACS. Patients 
on ACE inhibitors will either have it discontinued or switched to an angiotensin receptor 
blocking agent (ARB). All screen eligible patients will be subsequently asked to sign an 
IRB-approved informed consent form prior to clinically indicated PCI or any subsequent 
study related tests. 
ACS is defined as chest discomfort lasting at least 10 min within the last 24h, 
new ≥lmm ST-wave or dynamic T-wave changes in at least two contiguous EKG leads 
but without significant ST elevation, and/or troponin I or T levels above the 99th 
percentile. Abnormal LFT is defined as any liver transaminases (ALT or AST) ≥ 3 times 
the upper limit of the normal laboratory reference. Atorvastatin 40-80mg will be 
considered equivalent to 20-40mg of Rosuvastatin. Patients on Simvastatin at the time 
of enrollment will be switched to Atorvastatin 40-80 mg or Rosuvastatin 20-40 mg. 
After PCI of the target or culprit coronary artery (as determined by the operator) 
and IVUS-VH analysis of a ≥20mm segment of the target coronary artery are performed 
during the index hospitalization for the qualifying ACS event, the patients will be 
randomized to ILLT or SMT if they meet all inclusion and exclusion criteria (Figure 7, 
blue arrow). PCI will be performed using standard technique at the discretion of the 
operator. 40-80mg daily Atorvastatin or equivalent treatments (ILLT or SMT) will be 
initiated within 24 hours after randomization. Patients randomized to the ILLT arm will 
undergo LDL-apheresis within 36 hours after presentation unless the 36-hour window is 
mandatory for those who were on ACE inhibitors to allow for the complete washout of 
ACE inhibitors. Adjunctive anti-platelet and anti-thrombin therapies will be at the 
discretion of the primary operator and clinical team. Refer to Figure 8 for all study 
timelines. 
 
4(c)(ii). Inclusion Criteria: 
(1). Willing and able to provide informed consent (including HIPAA) 
(2). Age >30 years 
ACS: Acute coronary syndrome; PCI: percutaneous coronary intervention; ILLT: intensive lipid lowering therapy; 
SMT: standard lipid lowering therapy; IVUS-VH: intravascular ultrasound with virtual histology 
Patient Presenting 
With ACS 
Index cardiac 
Catheterization+ PCI + 
IVUS-VH 
ILLT 
SMT 
Atheroma 
volume by IVUS-
VH 
90 d follow-up 


 
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(3). Presenting with acute coronary syndrome (ACS), manifested as unstable 
angina or non-ST-elevation myocardial infarction 
(4). Referred for clinically indicated, non-emergent (the procedure is not 
required to be performed within 3 hours after patient presentation) 
coronary angiography and PCI with IVUS-VH of target coronary artery for 
ACS 
(5). Successful placement of two large-bore IV cannulas in bilateral upper 
extremities 
(6). Fasting (≥ 12 hrs) LDL ≥70mg/dl while on ≤80mg Atorvastatin or 
equivalent dose of other statin, performed at time of admission  
 
4(c)(iii). Exclusion Criteria: 
(1). Known allergy to aspirin, statins, or iodinated contrast 
(2). Positive pregnancy test, planning to become pregnant,  or breast-feeding 
(3). Coexisting conditions that limit life expectancy to less than six months or 
affect patient compliance 
(4). Uncontrolled fasting (≥ 12 hrs) triglyceride levels (≥500mg/dl) 
(5). Already participating in an investigational device or drug study 
(6). History of heparin induced thrombocytopenia (HIT) 
(7). Persons with estimated GFR less than 45 ml/min  
(8). ST-elevation myocardial infarction at admission 
(9). Abnormal liver function test (LFT) at time of admission, with abnormal 
LFT defined as any liver transaminases (ALT or AST) ≥ 3 times the 
upper limit of the normal laboratory reference 
(10). Pre-PCI or post-PCI left ventricular ejection fraction <25% by echo or 
cardiac catheterization done after admission 
(11). Pre-PCI, intra-PCI, or post-PCI hemodynamic instability with 
hypotension 
(12). Pre-PCI, intra-PCI, or post-PCI cardiac arrest 
(13). Pre-PCI or post-PCI acute heart failure with or without pulmonary edema 
(14). Intra-PCI or post-PCI sustained ventricular tachycardia 
(15). Complicated PCI, defined as PCI with any of the vascular access 
complications (large hematoma with lump > 5 cm or requiring medical 
treatment; AV fistula; pseudo aneurysm requiring treatment; 
retroperitoneal bleeding), or PCI with any of the procedural 
complications (abrupt vessel closure; no-reflow phenomenon; new 
angiographic thrombus; new major dissection with reduced flow; 
catheter-related thrombus), or PCI requiring further medical treatments 
(urgent CABG; endotracheal intubation; unplanned in-aortic balloon 
pump; LVAD; covered stent; unplanned temporary pacemaker wire; 
administration of inotropes; CPR) , or PCI resulting in clinical events 
(death; stroke; myocardial infarction; stent thrombosis) during or within 
24 hours after the index PCI  
(16). Post-PCI ongoing chest pain 
(17). Post-PCI severe groin pain and hematoma > 5cm in diameter 


 
PREMIER Pivotal Study Protocol Version 2.4 – 06/22/16 
Page 17 
(18). Persons whose hemoglobin is less than 9 grams following the index 
PCI/IVUS procedure, or who experience a drop in hemoglobin of greater 
than or equal to 2 grams following the procedure 
(19). Not able to comply with study protocol as determined by the 
investigators 
 
4(c)(iv). Baseline Data Collection: A complete history and physical, 12-lead 
electrocardiogram, lipid panel, basic metabolic panel (BMP) with LFT, complete blood 
count, and complete cardiac catheterization, PCI, IVUS-VH and adjunctive pharmaco-
therapy data will be collected on all eligible patients apart from demographic and 
contact information. 
 
4(c)(v). Stratification and Randomization: Prior to randomizing a patient, all 
screening and baseline assessments must be completed and the patient determined to 
be eligible for inclusion in the study. Eligible patients will be randomized to one of two 
treatment groups (ILLT or SMT) following an uncomplicated index PCI. 40-80mg daily 
Atorvastatin or equivalent statin medication treatment (ILLT or SMT) will be initiated 
within 24h after randomization and LDL-apheresis for ILLT group will be initiated within 
36 hours of hospital admission unless the 36-hour window is mandatory for those who 
were on ACE inhibitors to allow for the complete washout of ACE inhibitors. Refer to 
Figure 8 for all study timelines. Screening and randomization will be performed by a 
study coordinator, and LDL-apheresis will be conducted by a contracted specialized 
apheresis nurse. 
 
4(c)(v)(1). Randomization Methods: The randomization scheme and associated 
codes will be developed by the Cooperative Studies Program Coordinating Center 
(CSPCC). Permuted block randomization with random block sizes will be employed to 
assign patients to either ILLT or SMT in a 1:1 ratio.  The randomization will be stratified 
by investigation sites only.  
 
4(c)(v)(2). Randomization and Blinding Procedures: Following an uncomplicated 
PCI procedure, patients who are eligible and willing to participate in the study will be 
randomly assigned to either the ILLT or SMT arm using an automated telephone-based 
randomization system. A touch-tone phone is necessary to use this Voice Information 
System (VIS) to randomize eligible subjects. CSPCC will assign each site a unique 6-
digit PIN that the unblinded staff can use to access the VIS system and randomize a 
subject (or add/update an SSN). The VIS will query the unblinded staff for subject 
information and confirm these data prior to determining the subject's treatment group 
assignment. A copy of the appropriately executed consent and HIPAA authorization 
documents must be on file at the Hines CSPCC within 24 hours of randomization. If the 
VIS is not working during normal business hours, the research coordinator can 
telephone the Hines CSPCC staff to randomize a patient. The subjects or Site 
Investigators will not be blinded to the study treatment assignment; however all follow-
up visits and endpoint adjudications will be performed by a dedicated study nurse 
blinded to treatment allocation. Patients will be cautioned not to reveal his/her treatment 
allocation to the blinded study nurse. This blinding precaution will be emphasized by the 


 
PREMIER Pivotal Study Protocol Version 2.4 – 06/22/16 
Page 18 
procedures written into the study Operations Manual and the communications to the 
Site Investigator, the Site Coordinator and other study personnel during the kick-off 
meeting and the teleconference calls throughout the study. Patients will be screened 
and consented before PCI, but will have the opportunity, after PCI, to decline 
participation in the study and will not be randomized if they choose not to participate. 
 
 
4(d) 
Intervention and Treatment 
The randomized treatment regimens of this study include: 
(i) ILLT 
(ii) SMT 
 
4(d)(i): ILLT: Patients randomized to ILLT will undergo LDL-apheresis within 36h 
after presentation with an ACS unless the 36-hour window is mandatory for those who 
were on ACE inhibitors. All ILLT patients will also be started on Atorvastatin 40-80mg 
daily or equivalent within 24h after randomization. Patients will be continued on aspirin 
81-325mg and clopidogrel 75mg daily. LDL-apheresis will be initiated in the intensive 
care unit or inpatient hemodialysis or plasmapheresis center, supervised by a dedicated 
contracted LDL-apheresis nurse and Dr. Banerjee and an apheresis expert. A 
hematologist or an apheresis specialist will supervise LDL-apheresis at the other study 
centers. Bilateral large bore brachial venous intravenous access will be obtained and 
the patient connected to the LIPOSORBER® machine. Pre-existing intravenous access 
sites can be used for this purpose to minimize patient discomfort associated with 
additional IV placements. Each machine has two apheresis columns. The system is 
primed with heparinized Lactated Ringer’s injection, and 30-60 units of heparin per 
kilogram body weight is administered intravenously to the patient followed by an 
infusion throughout the treatment. When the first apheresis column has completed 
adsorbing LDL, the computer-regulated machine automatically switches the plasma flow 
to the second column to continue LDL adsorption. Simultaneously, the plasma 
remaining in the first column is returned to the patient. The first column is regenerated 
using a 5% Sodium Chloride Injection, eluting the LDL, VLDL and Lp(a). When elution is 
completed and flushed through the waste lines to a waste bag, the column is reprimed 
completely and ready for the next cycle of adsorption, allowing continuous LDL-
apheresis. No additional fluids are given to the patient during column switchover, and 
only the treated plasma is returned. A typical procedure takes about three hours. 
Typical LIPOSORBER® settings are: 
Extracorporeal volume = approx. 400ml (170ml blood, 230ml plasma) 
Plasma volume treated = approx. 0.7 x kg body weight x hematocrit 
Treatment time = 2-4 hours 
LDL-C acute reduction = 73-83% 
Unfractionated heparin dose = 45 units/kg (of which 75% given as a single IV 
bolus dose at the initiation of LDL-apheresis, and the rest to be delivered via 
an IV infusion at the rate of 1 ml/hr [1000 units/ml formulation]). Heparin 
infusion to be stopped 30 minutes prior to the estimated end of treatment. 
 


 
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Machine settings ranges include: 
Treatment volume = 0-9950ml plasma 
Whole blood flow rate = 0-150ml/min 
Plasma flow rate = 0-40% of whole blood flow rate 
Heparin infusion rate = 0-5ml/hr 
Blood warmer temp = 35-40°C 
 
All patients randomized to ILLT will receive Atorvastatin 40-80mg daily or equivalent 
within 24 hours after randomization. 
 
4(d)(ii). SMT: Standard medical therapy will include Atorvastatin 40-80mg daily or 
equivalent initiated within 24h after randomization. Patients will be initiated on 
Atorvastatin 40-80mg if their outpatient medication included Atorvastatin ≤80mg or 
equivalent dose of statin. Atorvastatin 40-80mg will be considered equivalent to 
Rosuvastatin 20-40mg. In the event of a restrictive formulary change of preferred statin 
drug within the VA system, the drug equivalency statement will serve as the reference 
for statin drug changes to meet the inclusion/exclusion and study follow-up LDL-goals 
as per the National Cholesterol Education Program (NCEP) and the latest ACC/AHA 
Lipid Lowering Therapy guidelines33,37. 
The study will not enroll patients with known history of allergy or contraindications 
to statin therapy. If a patient is shown to be intolerant to the treatment of Atorvastatin 
40-80mg daily or equivalent after randomized into the study, the Site Investigator may 
use an alternative statin provided above (Simvastatin or Rosuvastatin). If the intolerance 
remains unresolved, the Site Investigator may adjust the doses or even discontinue the 
statin therapy per clinical needs. These patients will still be followed up and included in 
the final analysis based on the intent-to-treat principle. 
4(d)(iii). Follow-up and study terminations: All participants will return for their 
scheduled study follow-up visits for six months. Participants will have the right to 
withdraw from the study at any time. However, Site Investigators may not withdraw a 
participant from the study without just cause and without the approval of the Study 
Chair. 
 
 
 


 
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Page 20 
4(e) 
Follow-up Assessments 
All study patients will undergo IVUS-VH analysis at index PCI and at follow-up. A 
minimum distance of 10mm away from the PCI site is the preferred site to visualize the 
atheroma volume with IVUS-VH. Patients will return at 90 days from index PCI for 
repeat cardiac catheterization and ≥20mm matched target coronary artery segment 
IVUS-VH at the exact same site as that of baseline using similar projections and side 
branch and/or calcification reference (Figure 5) using the technique published by our 
group29. A core laboratory consisting of two experienced cardiologists blinded to the 
study treatment allocation will review baseline and follow-up coronary arteriograms and 
IVUS-VH of each patient in the study. These two cardiologists will be independent and 
have no direct relationship to the study team. If any disagreement occurs between these 
two cardiologists regarding the angiogram or IVUS-VH interpretation, a third cardiologist 
will be referred to review the data and break the tie. After visual evaluation, quantitative 
analysis (QCA) will be performed using a standard automated edge-detection method. 
Angiographic restenosis is defined as >50% reduction in minimum luminal diameter 
(MLD) of the intervened segment during index PCI. 
 
 
Figure 8: PREMIER study timelines 
 
             
 


 
PREMIER Pivotal Study Protocol Version 2.4 – 06/22/16 
Page 21 
Atheroma volumes within a ≥ 20 mm long segment of a target coronary artery 
segment will be compared to assess change in total atheroma volume and %NC 
volume. Vessel and lumen area will be calculated for each frame, using previously 
published methods18. Atheroma burden will be calculated as [(EEM area – Lumen 
area)/EEM area] or expressed as a percent change [(EEM area – Lumen area)/EEM 
area) x 100]18.VH analysis will be performed for each segment, and the area of each 
plaque constituent (fibrous, fibrofatty, calcific, and necrotic core) will be determined in 
an automated fashion using Volcano S5 software version 2.2.3.2236 (Volcano Corp., 
Rancho Cordova, CA)32 (Figure 9). Follow-up cardiac catheterization and IVUS-VH 
analysis as described above will be performed at 90 days from index PCI. This strategy 
will allow capture of any early change in coronary atheroma volume as a result of the 
most intensive LDL reduction ever achieved in ACS patients. All follow-up IVUS-VH 
analyses will be compared to baseline IVUS-VH obtained during the index cardiac 
catheterization. A more long-term (1 or 2 year) IVUS-VH follow-up is not expected to 
demonstrate the treatment effect instituted immediately after ACS and has been well 
described with Nissen et al. with aggressive long-term LDL lowering18. Acute changes in 
coronary atheroma have traditionally been studied by short-term IVUS 
assessments20,21. 
 
If a patient has a clinically indicated catheterization within two months of the 
scheduled study procedure, this would be considered as the follow-up procedure and 
the operator will be encouraged to procure all study-related data.  
 
Peripheral blood sampling for EPC quantification will be performed using cell 
culture only at VA North Texas center in Dallas. This will ensure an optimal number (at 
least 50) uniformly standardized and cost-effective EPC cell culture assays in this multi-
center study. A limited number (n=25) of FACS EPC analysis will be performed to 
correlate cell culture and FACS levels of circulating EPC. Such a correlation would be 
important to demonstrate if future large-scale use of this treatment strategy of EPC 
mobilization is to be implemented with the rapid and less cumbersome FACS instead of 
the established standard of EPC cell culture technique. Approximately 10ml of whole 
blood is needed for either assay. Cell culture assay based upon our published 
methodology11 will be performed ≤48 hours of collection. EPC flow cytometry will be 
performed using standard methodology34. All EPC sampling schedules are indicated in 
Figure 8 and in Table 1, along with all other scheduled clinical and laboratory tests. All 
EPC analyses will be compared to baseline pre-PCI levels. Table 1 summarizes all 
planned follow-up visits and tests. 
 
Follow-up tests like fasting lipid (FL) will be performed immediately following 
LDL-apheresis in ILLT device group at about 12-24 hours post-PCI procedure. The 
same fasting lipid profile test for SMT control group will be done at 24 hours post-PCI 
procedure or prior to hospital discharge, whichever comes earlier. A post-PCI 30-day 
visit will include a complete blood count (CBC) and basic metabolic profile (BMP) with 
LFT. The 30-day post-PCI CBC, BMP and LFT are clinically indicated tests. A 30- and 
90-day FL in both treatment groups, along with EPC assay and an additional 60 day 
phone follow-up, will ensure strict lipid management and close follow-up for clinical 


 
PREMIER Pivotal Study Protocol Version 2.4 – 06/22/16 
Page 22 
events, and prevent any loss of recruited study subjects. After the 90-day follow-up 
cardiac catheterization and IVUS-VH analysis, all study subjects will return in 30 days 
for a 4-month study visit with CBC, BMP, LFT and FL. After an additional 60 days, or at 
6 months from index PCI, all patients will return for a study completion visit and a FL 
assay. Any MACE will also be collected via searching the Austin database of VA 
hospital utilization by Hines CSPCC staff. This dual MACE assessment will help avoid 
missing out on any participating site MACE.  
 
All adverse event, serious adverse event and MACE assessments during follow-
up will be performed during patient interviews or history and physical assessments and 
phone interviews, as indicated in Table 1. 
 
 
 
 
 


 
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Figure 9: Schematic (A) and on-line in vivo (B) IVUS-VH coronary analysis 
        
 
 
 
 
 
 
 
 
 


 
PREMIER Pivotal Study Protocol Version 2.4 – 06/22/16 
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Table 1: Follow-up study visits, clinical and laboratory testing schedule 
             
Visits (days)* 
H&P EKG 
Card. 
cath./ 
IVUS-
VH 
FL 
BMP 
+ 
LFT 
CBC EPC Phone 
Call 
Screening/Recruitment 
+ 
+ 
 
+ 
+ 
+ 
 
 
Pre-PCI 
 
 
 
 
 
 
+ 
 
Index PCI 
 
 
+ 
 
 
 
 
 
Post-PCI 
 
+ 
 
+ 
+ 
+ 
+ 
 
Hospital Discharge following index PCI for ACS 
30-day f/u visit 
+ 
+ 
 
+ 
+ 
+ 
+ 
 
60-day f/u visit 
 
 
 
 
 
 
 
+ 
90-day f/u IVUS-VH 
+ 
+ 
+ 
+ 
+ 
+ 
+ 
 
4-month f/u visit 
+ 
+ 
 
+ 
+ 
+ 
 
 
6-month f/u visit 
+ 
+ 
 
+ 
+ 
+ 
 
 
*These tests will be performed on all patients in study with “+” indicated when the 
tests are performed. All follow-up visits calculated from the day of study treatment 
randomization (dark gray line). Follow-up visits to be scheduled within  7 days of 
that time point in study. 90-day follow-up only to be scheduled  15 days of the 90-
day time point. 
H&P=history and physical; EKG=electrocardiogram; Card. cath./IVUS-VH=cardiac 
catheterization/PCI/intravascular ultrasound with virtual histology; FL=fasting lipids; 
BMP=basic metabolic profile; LFT=liver function test; 
CBC=complete blood count; EPC=endothelial progenitor cell; PCI=percutaneous 
coronary intervention; 
ACS=acute coronary syndrome; f/u=follow-up. 
 
 
4(f) 
Adverse Event (AE) and Serious Adverse Event (SAE) Assessments and 
Reporting 
 
4(f)(i). Role of the Local Site Investigator in Adverse Event Monitoring:  The local 
Site Investigator will be responsible for following adverse event reporting requirements: 
(1). Reviewing the accuracy and completeness of all adverse events reported 
(2). Complying with study policies as well as local IRB policies for reporting 
serious adverse events or problems involving risks to subjects or others 
in VA research. 


 
PREMIER Pivotal Study Protocol Version 2.4 – 06/22/16 
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(3). Reporting to the IRB any safety issues reported to the site by the 
Sponsor (Study Chair), and  
(4). Closely monitoring of study participants for any new Adverse Events 
(AE), Unanticipated Adverse Device Effects (UADE), or Serious Adverse 
Events (SAE). 
 
4(f)(ii). Definitions:  
 
An Adverse Event (AE) is defined as “any untoward medical occurrence in a 
patient or clinical investigation subject administered a pharmaceutical product and does 
not necessarily have a causal relationship with this treatment.” An AE can therefore be 
any unfavorable or unintended sign (including an abnormal laboratory finding), 
symptom, or disease temporally associated with the study intervention, whether or not 
related to the study intervention. 
 
A Serious Adverse Event (SAE) is one that results in any of the following 
outcomes: death, a life-threatening adverse event, inpatient hospitalization or 
prolongation of an existing hospitalization, a persistent or significant incapacity or 
substantial disruption of the ability to conduct normal life functions, or a congenital 
anomaly/birth defect.  Important medical events that may not result in death, be life-
threatening, or require hospitalization may be considered serious when, based upon 
appropriate medical judgment, they may jeopardize the patient or subject and may 
require a medical or surgical intervention to prevent one of the outcomes listed in this 
definition. 
 
An Unanticipated Adverse Device Effect (UADE) means any serious adverse 
effect on health or safety or any life-threatening problem or death caused by, or 
associated with, a device if that effect, problem, or death was not previously identified in 
nature, severity, or degree of incidence in the investigational plan or application, or any 
other unanticipated serious problem associated with a device that relates to the rights, 
safety, or welfare of subjects.  
 
All AE/SAE/UADE with a reasonable causal relationship to the investigative 
treatment should be considered at least “possibly related.” A definite relationship does 
not need to be established. For the purpose of this trial potential investigative-treatment-
related AE/SAE/UADE can be classified into three categories: 
 
(1). Related to LDL-apheresis: hypotension (most common, <1% of treatments in US), 
nausea/vomiting, flushing, chest pain, fainting, lightheadedness, anemia, abdominal 
discomfort, numbness/tingling, tachycardia, headache, shortness of breath, hemolysis, 
bradycardia, itching/hives, bleeding due to the use of unfractionated heparin or low 
platelet count (thrombocytopenia), low Vitamin E level, temporary decrease in blood 
protein level, vertigo (dizziness, unsteadiness) and excess sweating. 
(2). Related to statin use: muscle pain, generalized body pains, nausea/vomiting, 
abnormal liver function test (≥ 3 times above the upper limit of the normal laboratory 
reference values of the study institution), and rhabdomyolysis. Also, on February 28, 


 
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2012, the FDA notified clinicians of the following possible effects of statin use: cognitive 
(memory loss or confusion), which usually reverses after stopping the statin; and 
increased blood sugar levels (hyperglycemia) that could lead to a diagnosis of type 2 
diabetes mellitus. Specifically regarding Lovastatin (Mevacor©), the FDA said that statin 
in combination with some other medications—including HIV protease inhibitors (e.g. 
tipranavir plus ritonavir and lopinavir plus ritonavir) and hepatitis C protease inhibitors 
(e.g. telapravir), and drugs used to treat certain bacterial (e.g. erythromycin, 
clarithromycin) and fungal infections (e.g. fluconazole, itraconazole, posaconazole, and  
voriconazole)—can increase the risk of myopathy and rhabdomyolysis. The FDA said 
serious liver injury due to statin use is rare.  
(3). Related to follow-up cardiac catheterization and IVUS-VH: The risk of follow-up 
study-related cardiac catheterization and IVUS-VH include bleeding, groin pain, 
arrhythmia, hypotension, pericardial effusion, blood clots, infection, allergic reaction to 
the dye, perforated blood vessel, air embolism, myocardial infarction, stroke, renal 
failure, and death. Local groin discomfort and minor access site hematoma (<3 cm in 
diameter) are most common. Death is rare (<0.04%), and the risk of MI and stroke is 
<0.5%. 
(The examples in these categories are not exhaustive, and investigators should 
not feel limited by these examples in designating an event “related” or “possibly related” 
if other AE/SAE/UADE occurs that in their opinion could be potentially related to the 
investigative treatment.) 
 
4(f)(iv). AE and SAE Monitoring and Reporting:  AE/SAE/UADE will be monitored 
at the study sites throughout the whole period of the study at each clinic visit and 
telephone contact, beginning as soon as a study participant signs the Informed Consent 
and continuing through end-of-study for each participant. All AE/SAE/UADE including 
both those related to the study intervention and those not related to the intervention, will 
be collected and recorded on the appropriate case report forms (CRF). For the purpose 
of safety monitoring, the study intervention is defined as the use of 1) statins with or 
without the addition of LDL-apheresis, 2) LDL-apheresis, and 3) follow-up cardiac 
catheterization and IVUS-VH as described in this protocol. 
All SAEs require expedited reporting. The SAE report form should be completed 
and faxed to CSPCC/CSPCRPCC/Study Chair Offices within 48 clock-hours of initial 
awareness of the event. The CSPCRPCC AE Pharmacist will be responsible for 
evaluating all serious adverse events for patient safety concerns. In addition, a Clinical 
Events Committee (CEC) will be formed to adjudicate adverse events in the study. The 
committee involves a physician from CSPCC, a Q.A./safety monitor from CSPCC, and 
the AE Pharmacist from CSPCRPCC. The committee is independent from the Study 
Chair and the Site Investigators. 
Serious adverse events that are related to the investigative treatment and 
unexpected will be reported to the FDA, CSP Director, and Site Investigators after 
review by the Study Chair, the CSPCRPCC Director and the CSPCC Director. Site 
investigators should follow local policy to inform their IRB. The Hines CSPCC will 
generate tabulations of all AE/SAE/UADE for the Data Monitoring Committee annually 


 
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or on a more frequent schedule if requested by the Committee. All participants 
presenting to the VA hospital will be provided treatment for all SAEs and UADEs.  
 
While participants have the right to withdraw from the study at any time, they will 
continue to be monitored, via passive follow-up, for SAEs that occur within 30 days of 
leaving the study. All unresolved SAEs must be followed up at least every 30 days until 
resolved or when no further change is expected. All SAEs are followed until no changes 
are expected or follow-up ends on the last study participant, regardless of the 
participant’s status in the trial. 
 
 
4(g) 
Biostatistical Considerations 
 
4(g)(i). Expected Treatment Effect: There is only one study which examined the 
treatment of medication only (20mg Pravastatin or 10mg Simvastatin) vs. LDL-
apheresis with statin medication on coronary plaque regression in familial 
hypercholesterolemia and published in 2002 Journal of American College of 
Cardiology31. The trial reported the net change of plaque area from baseline to one year 
follow-up. The changes were -0.69±2.08mm2 in the LDL-apheresis group vs. 
0.88±1.75mm2 in the medication only group. Two studies, which have primarily 
mobilized coronary atheroma LDL using HDL or its apoprotein, have shown similar 
magnitude of atheroma regression at 6-10 weeks, without directly lowering LDL levels. 
Given the expected 85% reduction in LDL after LDL-apheresis, we expected to observe 
a similar treatment effect for the primary effectiveness outcome of this study at 90 days. 
 
4(g)(ii). Sample Size/Power/Level of Significance: To maintain 90% power to 
detect a Cohen’s D effect size of 0.65 (-0.38 ± 2.08 mm2 in the LDL-A group vs. 0.88 ± 
1.75 mm2 in the medication only group) with the randomization scheme of 1:1, α = 0.05, 
and a 20% drop-out rate, the pivotal study needs a sample size of 128. 
 
4(g)(iii). Study Duration/Number of Sites: Four sites are considered for the 
purpose of generalizability and improved enrollment. Based on the enrollment rate in 
stage 1 of this trial, sites would expect to randomize 18 patients per site per year with 
an average rate of 1.5 patients per site per month. The average annual PCI volume 
during FY 2012 – 2013 reported by VA Cardiovascular Assessment and the data from 
CSP#571 currently enrolled at these 4 sites indicate that they will be able to meet or 
exceed the target of 18 participants per year. The percentage of patients with PCI who 
need to be randomized ranges from 3.5% to 5.0% across the four sites based on these 
reported annual PCI volume. About 30% of eligible subjects who meet the 3 major 
inclusion/exclusion criteria (ACS, LDL, and eGFR) need to be randomized in order to 
meet the accrual target. Dallas and Oklahoma City sites have ongoing LDL-apheresis 
capability and equipment on-site at the VA campus, and the Nashville and Denver sites 
have approved LDL-apheresis sites supported by device manufacturer (Kaneka) that 
will expedite the induction of these sites into the study. 
 
A total of 22 months is needed for 4 sites to randomize the total sample size of 
128 in this second stage. The whole study duration for the second stage is expected to 


 
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be 46 months including 6 months of start-up, 22 months of enrollment, 6 months of 
follow-up,  and 12 months of database search, study closure, and final data analysis. 
Upon approval, it is expected that the Dallas and Oklahoma City sites will begin 
recruitment almost immediately, followed by Nashville and Denver sites. 
 
 
Estimated Enrollment for CCTA #0002 Phase II 
Site 
PCI Volume as 
Reported by VA 
Cardiovascular 
Assessment, Reporting, 
and Tracking System for 
Cath Labs (CART-CL) 
Annual 
ACS 
Indication, 
FY2014* 
LDL > 70 
mg/dL 
AND 
eGFR > 45 
ml/min, 
FY2014* 
Annual 
Eligible 
Patients 
Monthly 
Eligible 
Patients 
Monthly 
Accrual 
Target 
FY2013 
FY2014 
Dallas 
350 
419 
158 
94 
94 
7.8 
1.5 
Oklahoma 
City 
402 
482 
258 
138 
138 
11.5 
1.5 
Nashville 
250 
313 
173 
110 
110 
9.2 
1.5 
Denver 
132 
210 
115 
58 
58 
4.8 
1.5 
*Eligibility criteria. 
 
4(g)(iv). Statistical Analysis of Primary Effectiveness Endpoint:  
The primary effectiveness outcome will be analyzed with intent-to-treat (ITT) 
approach by including all randomized patients regardless of crossover or drop-out. The 
change in total atheroma volume within a ≥ 20 mm long segment of the target coronary 
artery from baseline to 90 days will be analyzed using the two-sample t-test. We will 
define their change scores to be 0 if they do not have a 90-day atheroma volume 
measurement. Analysis of Covariance (ANCOVA) will be performed to evaluate the 
treatment effect on primary effectiveness outcome adjusting for age, pre-existing CAD, 
pre-existing diabetes, and baseline LDL levels. 
 
4(g)(v). Statistical Analysis of Secondary Effectiveness Endpoints:  
The intent-to-treat principle will also be applied to all the secondary effectiveness 
outcomes analyses. 
(1). The change of %NC component of atheroma from baseline to 90 days 
will be analyzed using the two-sample t-test. Similarly, the change scores 
for those who miss the 90 days follow up assessment will be set to zero. 
Analysis of Covariance (ANCOVA) will be performed to evaluate the 
treatment effect on primary effectiveness outcome adjusting for age, pre-
existing CAD, pre-existing diabetes, and baseline LDL levels. 
(2). The EPC-CFU/ml of peripheral blood assessed at various time points 
(pre-PCI, post-PCI, 30 days of follow-up, and 90 days of follow-up) will be 
analyzed via Mixed Linear models with random intercepts. The treatment 
effect, time effect and their interaction will be considered with or without 


 
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adjusting for age, pre-existing CAD, pre-existing diabetes, baseline LDL 
levels, and other relevant covariates such as ACS, peripheral vascular 
disease, and chronic kidney disease. The statistical test of interest is the 
time by treatment interaction. 
(3). Contrasts will be used in the Mixed Linear models to compare the 
differences among any specifically interested time points.  
 
4(g)(vi). Statistical Analysis of Safety Endpoints and Complications: The primary 
safety endpoints, the secondary safety endpoints, and the incidence of major adverse 
cardiovascular endpoints (MACE) including death, myocardial infarction, coronary 
revascularization, and stroke will be analyzed in two ways. For each type of event, the 
percentage of people who experience the event at least once will be compared using 
the chi-square test. Logistic regression models will be performed to adjust for the 
covariates such as age, pre-existing CAD, and pre-existing diabetes. In addition, a 
similar analysis will be done for all AE and SAE that are possibly or probably attributable 
to the study intervention. 
An event-based analysis will also be done since adverse events can be 
recurrent. In this analysis, a non-parametric method called the mean cumulative 
function (MCF) 35,36 will be used as an alternative to the above crude incidence rate 
analyses. The overall safety profiles as well as the safety profiles in specific subgroups 
over the whole study period will be compared for the two randomization groups 
including the times of event recurrence and censoring mechanisms.     
Time to event (survival analysis) will also compare the time to first MACE 
between treatment groups. A second survival analysis will be performed to identify risk 
predictors and to evaluate whether the observed treatment effect is modified by 
adjusting for covariates such as pre-existing CAD and pre-existing diabetes.  
 
4(g)(vii). Interim Monitoring: Hines CSPCC will produce a progress report every 
six months for review by Data Monitoring Committee. The report will include figures for 
patient accrual, tables for baseline characteristics, site performance, data quality, 
treatment compliance, and safety issues. Group sequential methods will be used to 
specify the α-levels in order to maintain the overall significance level at α = 0.05 for the 
primary effectiveness hypothesis test. The sequential analyses will use O’Brien-Fleming 
boundaries with an overall α = 0.05 for significance and 90% for power. EAST software 
was used to obtain the interim monitoring rule and the adjusted sample size.  The look 
will be around 50% of the target sample size (about 64 patients) completing the 90-day 
follow-up visit as DMC recommended at its first meeting on September 25, 2009. The 
sample size remains at 128 to account for this extra interim look by assuming the 
groups mean difference of -1.26 and the pooled standard deviation of 1.922.    
The interim look will cost 0.003 for α. If the critical value is greater than 2.9626 or 
less than -2.9626, the study will be stopped to reject the null hypothesis. However, if the 
critical value is between -2.9626 and 2.9626, the study will continue until the target 
sample size is met for final analysis. 
 
4(g)(viii). Subgroup Analyses: Since acute coronary syndrome (ACS) is 
manifested as either unstable angina or non-ST-elevation myocardial infarction, both 


 
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safety and effectiveness outcomes will be analyzed for LDL-apheresis treatment effects 
in unstable angina subgroup and non-ST-elevation MI subgroup as well as the total 
ACS patients. In addition, separate analyses will be performed across 4 participating 
sites for each treatment group in order to evaluate the site effect and confirm the 
appropriateness of pooling the data for primary endpoint.       
 
4(g)(ix). Sensitivity Analysis for Missing Data: In addition to the above mentioned 
ITT principle for primary and secondary effectiveness outcomes in which the change 
scores will be set to zero if the follow-up assessment is missing, other approaches will 
be applied in order to assess the robustness of the study results to assumptions about 
the missing data. For outcomes about atheroma volume and % of NC component of 
atheroma which will be measured only at 2 time points, the sensitivity analysis 
approaches include: 1) performing analysis only on complete data, 2) applying the worst 
case scenario (set the lowest score at baseline and the highest score at 90-day), and 3) 
generating imputes for missing data with multiple imputation. For outcome about EPC-
CFU/ml of peripheral blood which will be repeatedly measured at four time points, the 
sensitivity analysis approaches include: 1) performing analysis only on complete data, 
and 2) using the mixed model for repeated measures. 
 
All tests will be two-sided. The critical level for all the primary and secondary 
outcomes will be 0.05. No adjustments will be made for multiple secondary endpoints. 
 
 
4(h) 
Quality Assurance, Oversight and Study Regulation 
 
4(h)(i) Training to assure accuracy, precision and validity of the data: Toward the 
end of the six-month start-up period, training of study personnel will take place during a 
two-day kick-off meeting. All participating investigators, site research 
coordinators/raters, and Hines CSPCC personnel will be in attendance. Prior to the 
meeting, case report forms will be finalized and a detailed manual of study operations 
will be written and circulated. This manual will serve as the training manual for the 
meeting and as a reference document following the meeting. The PI’s office and the 
data and pharmacy coordinating centers will provide the training. The general training 
will include the study treatment, patient screening and consent, baseline evaluation, 
follow-up procedures, and proper entry and maintenance of data. Cyber security 
awareness training and privacy training (VHA Privacy Policy, Protection of Human 
Research Subjects, and Good Clinical Practices) are required annually of all VA and 
VA-WOC employees. 
 
4(h)(ii). Data Management and Security 
 
4(h)(ii)(1). Data Collection: An operations manual and case report forms will be 
developed jointly with Hines CSPCC. The site study coordinators will collect all data. 
After a patient consents to participate in the study, the site coordinator will create a 
patient casebook, which will contain the consent forms, all relevant source documents, 
and any other information pertinent to the study. The completed case report forms will 


 
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be assembled and faxed to Hines CSPCC for further handling on a daily basis. These 
original completed paper forms will be kept in the Site Investigator’s study files as 
source document. 
 
4(h)(ii)(2). Data Quality Assurance: DataFax, a clinical trial data management 
system (by Clinical DataFax Systems, Inc.), will be used for data collection and 
management. The study coordinators from the sites and the Independent Assessors will 
complete case report forms (CRFs) in paper and fax them directly to the DataFax 
computer server, where data images of the CRFs are stored as files. The system uses 
an optical character recognition (OCR) paradigm to automatically process and store the 
information from the image as data into the study database. The original fax image is 
also stored. Data management staff at Hines CSPCC will review each CRF by 
comparing the faxed image with the OCR data and ensure that the two match. Data 
management staff at Hines CSPCC will also review CRFs for protocol adherence, data 
consistency, and add data queries to items that fail these checks. Extensive data 
checks, including missing values, out-of range entries, and consistency between 
variables, both within and across forms, will be performed manually and 
programmatically. On a regular basis, data management staff will produce site-specific 
Quality Control reports that list all unresolved data queries.  Data management staff will 
make the reports available to each site and work with the study coordinators and the 
Independent Assessors to help them resolve queries.  Queries will be resolved when 
the appropriate corrections to the CRF are made and data resent, or when an 
explanation is provided that allows for data management staff to resolve the query.  All 
corrections and changes to the data will be reviewed by data management staff.  In 
addition to the Quality Control report, Hines CSPCC may generate and distribute 
targeted data edit reports on an as-needed basis. The Hines CSPCC will monitor 
completeness and timeliness of the data discrepancy resolution made by study 
coordinators at participating sites. The Study Chair, the study coordinators and 
Independent Assessors will receive periodic reports regarding the quality and quantity of 
data submitted to the Hines CSPCC. Other quality control measures include periodic 
reports containing participant recruitment information and relevant medical data for 
review by the Study Chair.  The Hines CSPCC will also prepare summary reports for the 
Study Chair, the Data Monitoring Committee, and other monitoring groups of the data to 
track progress, and conduct final analyses of the study data. Study reports will be 
generated using DataFax, SAS, and other tools (e.g., Microsoft Excel and Access).  
SAS and other statistical software packages will be used to conduct data analysis for 
the study.  The Hines CSPCC is using SAS Version 9.3 in 2014 and will upgrade to 
newer versions once they are purchased and validated. 
 
4(h)(ii)(3) Data Confidentiality and Data Security: Handling and storage of study 
data will adhere to current VA policies. The analytical database will not contain 
information that can directly identify the study participant (such as name, address, etc.); 
however it will not be a completely de-identified database since age and study visit 
dates will be collected. The Hines CSPCC requires that a copy of the signed consent 
form and a participant contact sheet be on file at the Hines CSPCC. The consent form is 
required by CSP policy in order for the coordinating center to independently certify that 


 
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all study participants have been consented. Because the Hines CSPCC is the final data 
repository for the study, participant contact sheets are collected in the event study 
participants need to be contacted (such as for safety notices) after study sites have 
completed the study. Consent forms and participant contact sheets are stored 
separately from study data. All paper-based records, including source documents and 
paper case report forms, will be kept in locked file cabinets at the participating sites and 
Hines CSPCC. The servers housing the study databases will be located at a secure VA 
facility and housed behind the VA firewall on VA-owned and VA-maintained servers.  
The system will be monitored to ensure that all applicable VA regulations and directives 
are strictly followed. Access to the study data is restricted by the Hines CSPCC to 
properly credentialed research staff who have completed required VA security trainings.  
Only CSP-approved individuals (such as: staff at the study sites, CSPCC, and CSP 
Clinical Research Pharmacy Coordinating Center [CSPCRPCC]) will have access to the 
personal health information (PHI) of study participants. 
 
4(h)(ii)(4). Site Performance: Hines CSPCC evaluates recruitment and retention 
performance monthly. Other performance problems such as protocol deviations, poor 
data quality, missing or overdue data, and reasons for withdrawal from study will also be 
tracked. A monthly study conference call for study personnel including Hines CSPCC, 
CSPCRPCC, Chair’s office, and all sites will be held to review study recruitment, data 
quality, and protocol adherence. Study sites will be put on probation for poor 
performance, including under-recruitment. Typically the probationary period is three 
months, at which time the site may be taken off probation, have probation continued, or 
have funding stopped, depending on its performance during the probationary period. 
The Hines CSPCC Director is authorized to make these decisions. 
 
4(h)(iii). Study Monitoring: The groups charged with centrally monitoring the 
various aspects of the study will be the Executive Committee (EC) and Data Monitoring 
Committee (DMC). Both committees meet at study start-up and annually thereafter. The 
EC will also have quarterly conference calls. The DMC may elect to meet more 
frequently if it deems it necessary.  
The Executive Committee is the management and decision-making group for the 
operational aspects of the study. One of its major responsibilities is to monitor the 
performance of the participating medical centers. The EC considers the need for 
protocol modifications. The Executive Committee also reviews and approves all 
manuscripts and abstracts emanating from the study. Typically the EC is composed of 
the original study Planning Committee. It is chaired by the Study Chair. 
The Study Group, which consists of all participating investigators and site clinical 
research personnel, will meet annually to discuss the progress of the study and any 
problems encountered during the conduct of the trial. Study personnel will adhere to 
Office of Research & Development (ORD) policy on human subjects’ protections by fully 
completing approved Good Clinical Practices (GCP) training per VHA Handbook 
1200.05. Completed training certificates will be provided by site personnel to the Hines 
CSPCC to maintain on file throughout the study. SMART (Site Monitoring, Auditing, and 
Resource Team), a division of the CSP Clinical Research Pharmacy Coordinating 
Center (CSPCRPCC), may conduct a full audit of participating sites if requested by any 


 
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of the monitoring bodies. The study will be monitored utilizing centralized data and 
statistical monitoring methods and remote-based source verification; which will be 
based on risk assessment and management strategies. 
The Data Monitoring Committee will review the progress of the study, including 
patient intake, completeness of follow-up, data quality, protocol deviations, and safety. 
The DMC may also choose to implement a formal interim monitoring rule to monitor 
efficacy. The DMC will review any protocol modifications recommended by the EC. The 
DMC will establish criteria for study termination and make recommendations to the 
Director, CSR&D, through the Director, Hines CSPCC, as to whether the study should 
continue, be put on probation, or be terminated. Interim unblinded progress reports will 
be provided to the DMC by the study biostatistician. This study has been assigned to a 
central DMC organized and managed by VA Clinical Sciences Research and 
Development (CSR&D). The membership of DMC is approved by the VA CSR&D 
Director and has no direct relationship to any facility, study or investigator. The DMC 
has about 10 members, in which three members including one interventional 
cardiologist, one critical care physician, and one biostatistician were assigned to review 
this study. 
The Study Group, which consists of all participating investigators and study 
personnel at Hines CSPCC and Albuquerque CSPCRPCC, will meet annually to 
discuss the progress of the study and any problems encountered during the conduct of 
the trial. Finally, the study protocol and progress must be reviewed annually by the local 
R&D Committee and IRB at each site in order for that site to participate in the study. 
 
4(h)(iv). Study Performance Benchmarks: Study benchmark dates will be 
determined upon successful funding of the study. However, the following timelines will 
serve as a reference:  
(1). Enrollment of four VA sites: Dallas VA, and 3 other VA sites 
(2). Study site IRB submission and approval: four months 
(3). Study sample transportation and storage arrangements and contract: 
two months 
(4). Recruitment of ACS patients/month/site: one and a half 
(5). Study sites recruitment/year: 72 
(6). Subject follow-up and follow-up study blood sampling: 80% 
(7). Lost to follow-up: <20% 
 
 
4(i) 
Human Subjects: 
1. Human subject involvement and characteristics: As the study proposes to enroll 
ACS patients, strict enrollment criteria will be used to select eligible patients.  
 
2. Sources of material: The source of material will be information obtained during 
hospitalization, procedure logs, and laboratory values and tests. The laboratory 
values and tests will include research laboratory and patient relevant hospital 
laboratory data. LDL-apheresis data will be collected by the apheresis center 
nurse on paper forms. Patients’ names will not be divulged, and all data will be 
coded in the study records. The investigators will only use information gained 


 
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during the study for the evaluation of study results. The copies of signed 
informed consents will be kept on file by the site investigators, and no patient 
names or other identifying data will be used in future publications. All files will be 
secured in a locked filing cabinet in a locked office. 
 
3. Informed consent: The site investigators or the site study coordinators will 
interview and explain the details of the study to all potential subjects, who will be 
subsequently asked to sign an IRB-approved informed consent form prior to any 
study procedure.  
 
4. Provisions for managing adverse reactions: Only trained and accredited 
personnel will be performing PCI, IVUS, LDL-apheresis, peripheral blood 
sampling, and laboratory handling of specimens. Cardiac catheterization will be 
performed using standard clinical protocol, and all patients will be observed 
closely in hospital after the procedure and discharged when clinically indicated. 
Consented patients with a complicated PCI procedure will be excluded from 
participating in the study, but will be monitored for AEs/SAEs, and the safety data 
will be reported to the study safety and oversight committees. However, 
randomized patients with post-PCI complications will still be followed unless 
otherwise terminated from the study, performed as many assessments as 
possible, and included in the final analyses. Intensive patient follow-up protocol 
has already been described in Table 1. Patients will undergo LDL-apheresis 
using peripheral IV access exclusively. No more than two patients will be enrolled 
and will undergo LDL-apheresis on a given day at a center. The study will be 
terminated if recommended by the oversight committee at any point if potential 
harm to the patients is demonstrated. 
 
5. Potential benefit to subjects and others: All patients will be treated and followed 
following an ACS and will receive more intensive pharmacologic hyperlipidemia 
management which has traditionally resulted in improved patient outcomes. 
Patients will also be reimbursed $50.00 for participation in the study, $50.00 for 
the 90-day follow-up visit, and $40.00 for each of the other follow-up visits to 
improve compliance and attainment of National Cholesterol Education Program 
(NCEP) and the latest ACC/AHA Lipid Lowering Therapy guidelines. The 
information obtained from this study could certainly develop as a viable strategy 
for our veterans. Successful completion of this study with acquired necessary 
experience may lead to development of this treatment strategy at one of the 
largest VAMCs in the nation. 
 
6. Importance of the knowledge gained: The results of this study, apart from 
providing VP stabilization information from intensive LDL lowering, will principally 
advance the field of coronary atherosclerosis and VP healing. 
 
 
4(j) 
Publication Policies and Planned Manuscripts 


 
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The study findings will be presented at national meetings and published in peer-
reviewed medical journals. Before the publication phase, the study’s objectives and 
other related facts will be presented for informational (not recruitment) purposes on the 
Cooperative Studies Program website at www.research.va.gov/programs/csp/ccta0002. 
Only general information will be provided without any participant-specific references. 
The Executive Committee will approve all topics for presentation or publication. It will 
review and approve all manuscripts and abstracts prior to submission. The Director, 
Hines CSPCC, must also approve every manuscript and abstract prior to publication. 
 
 
4(k) 
Resources 
All work including LDL-apheresis will be conducted on the premises of four study 
sites except that the EPC cell culture analysis will be done only at the Dallas VAMC: 
 
1. Cardiac catheterization and PCI/IVUS-VH will be performed in the cardiac 
catheterization laboratory of the Dallas VAMC and other VA sites. 
 
2. Laboratory analysis of EPC by cell culture and flow cytometry will take place in 
the research laboratory of the PI at Dallas VAMC. The PI will be utilizing a 
dedicated laboratory space, which currently includes 850 sq ft of laboratory 
space, 200 sq ft of office space, and shared 400 sq ft dedicated to cell culture 
facility. The same building houses common-use instruments such as 
ultracentrifuges; inverted dual laser confocal microscope (Zeiss); inverted 
microscope (Nikon) with epifluorescence, Nomarski optics, and digital image 
acquisition (Metamorph); flow cytometer (BD FacsCaliber); cryotome and 
microtome, and molecular imager (BioRad). All equipment for performance of the 
project is available to the PI. 
 
3. Single LDL-apheresis in all study patients enrolled will take place at the intensive 
care and inpatient hemodialysis rooms. Dr. Banerjee has attended several 
apheresis sessions conducted at UT Southwestern Medical Center in Dallas and 
during the phase 1 of the study to obtain first hand experience and didactic 
education. A specialized LDL-apheresis nurse coordinator will be recruited and 
perform the apheresis procedure under the supervision of an apheresis expert 
identified as site co-investigator. Similar to Dallas, arrangement regarding the 
apheresis expert will be done at other participating sites. 


 
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Treatment of Blood Cholesterol to Reduce Atherosclerotic Cardiovascular Risk in 


 
PREMIER Pivotal Study Protocol Version 2.4 – 06/22/16 
Page 39 
Adults, Journal of the American College of Cardiology (2013), 
doi:10.1016/j.jacc.2013.11.002 
 

SAP_001.pdf: 
CCTA 0002 PREMIER – Statistical Analysis Plan V2.0 (09-30-2014)               Page 1 of 8 
 
 
Statistical Analysis Plan  
for 
VA Cooperative Clinical Trial Award Study #0002 
 
 
Plaque Regression and Progenitor Cell Mobilization  
with Intensive Lipid Elimination Regimen  
 
(PREMIER) 
 
 
Version 2.0 
 
 
September 30, 2014 
 
 


CCTA 0002 PREMIER – Statistical Analysis Plan V2.0 (09-30-2014)               Page 2 of 8 
CCTA 0002 PREMIER Study  
Statistical Analysis Plan Version 2.0 
 
Primary Safety Outcome Measure 
 
The primary safety outcome measures will be the total number of and percentage of 
patients with major peri-PCI procedure adverse events, such as hypotension, angina, 
myocardial ischemia, myocardial infarction (if the patient is determined to have had 
unstable angina rather than non-ST-elevation MI at admission), cerebrovascular event (CVA), 
vermicular tachycardia, bleeding (at the PCI access and apheresis cannulation sites in the 
apheresis patients and at the PCI access site in the non-apheresis control patients), and 
death. The peri-PCI procedure is defined as encompassing the time of the PCI procedure 
and the time of the subsequent LDL-apheresis procedure for ILLT group versus the time of 
the PCI procedure for SMT group. All LDL-apheresis-related adverse events in the ILLT group, 
including any minor expected events, will be recorded. 
 
 
Secondary Safety Outcome Measure 
 
The secondary safety outcome measures will be the total number of and percentage of 
patients with statin-related abnormal liver function test events and statin-related muscle injury 
events, which could occur due to the maximum dose of statin drugs being given to both the 
ILLT group and SMT group patients. The statin-related muscle injury is defined as a muscle 
injury which cannot be attributed to a non-statin cause and which is evidenced by symptoms 
(muscle soreness, pain, or tenderness) and/or lab tests such as serum total creatine 
phosphokinase (CPK), CPK-MM, or myoglobin. 
 
 
Primary Effectiveness Outcome Measure 
 
The primary effectiveness outcome measure will be the change in the total atheroma 
volume within a ≥ 20 mm long segment of the target coronary artery from baseline to 12 
weeks post-PCI. The measurement will be done via IVUS-VH at 2 time points (baseline 
during index PCI and 90-day follow-up). 
 
  
Secondary Effectiveness Outcome Measures 
 
The secondary effectiveness outcome measures will include: 
1. The %NC component of atheroma. The %NC component of atheroma will be 
obtained via IVUS-VH at 2 time points (baseline during index PCI and 90-day 
follow-up). 
2. EPC-CFU/ml of peripheral blood. The cell culture assay and quantification of 
circulating EPC-CFU will be performed for patients recruited at the Dallas VA 
center only. The assay will be done at 4 time points (pre-PCI, post-PCI, 30-day 
follow-up, and 90-day follow-up). 


CCTA 0002 PREMIER – Statistical Analysis Plan V2.0 (09-30-2014)               Page 3 of 8 
3. The major adverse cardiovascular endpoints (MACE) including death, myocardial 
infarction, coronary revascularization, and stroke during the follow-up periods. 
The major adverse CV events will be collected via both clinical visits for up to 6 
months post-PCI and searches of the VA Austin database for up to 1 year post-
PCI. Approvals for using the patients’ Social Security Numbers for data searching 
will be obtained via informed consent form. 
 
 
Sample Size Determination 
 
There is only one study which examined the treatment of medication only (20mg 
Pravastatin or 10mg Simvastatin) vs. LDL-apheresis with statin medication on coronary 
plaque regression in familial hypercholesterolemia and published in 2002 Journal of 
American College of Cardiology31. The trial reported the net change of plaque area from 
baseline to one year follow-up. The changes were -0.69±2.08mm2 in the LDL-apheresis 
group vs. 0.88±1.75mm2 in the medication only group. Two studies, which have 
primarily mobilized coronary atheroma LDL using HDL or its apoprotein, have shown 
similar magnitude of atheroma regression at 6-10 weeks, without directly lowering LDL 
levels. Given the expected 85% reduction in LDL after LDL-apheresis, we expect to 
observe a similar treatment effect for the primary effectiveness outcome of this study at 
12 weeks. 
 
The sample size of 30 patients for the first feasibility/pilot study leads to a power of 54% 
with α = 0.05 and the same effect size as reported in the published paper (Cohen’s D 
effect size = 0.82 for -0.69 ± 2.08 mm2 in the LDL-A group vs. 0.88 ± 1.75 mm2 in the 
medication only group). The power decreases to 38% with α = 0.05 and a more 
meaningful lower Cohen’s D effect size of 0.65 by decreasing the difference of two 
treatment means by 20% (-0.38 ± 2.08 mm2 in the LDL-A group vs. 0.88 ± 1.75 mm2 in 
the medication only group). The power decreases further to 31% by considering 20% 
drop-out rate with α = 0.05 and the Cohen’s D effect size = 0.65. 
 
The second stage of pivotal study is approved to obtain the 90% power to detect a 
Cohen’s D effect size of 0.65 (-0.38 ± 2.08 mm2 in the LDL-A group vs. 0.88 ± 1.75 mm2 
in the medication only group) with the randomization scheme of 1:1, α = 0.05, and a 
20% drop-out rate, the pivotal study needs a sample size of 128. 
 
 
 


CCTA 0002 PREMIER – Statistical Analysis Plan V2.0 (09-30-2014)               Page 4 of 8 
 
Baseline Characteristic Comparisons 
 
Patient characteristics and outcome measures at baseline will be compared between 
the two treatment groups (ILLT vs. SMT) to determine if the treatment arms differ on 
any important variables.  The chi-square test or Fisher’s exact test will be used for 
categorical variables and the t-test for two independent samples will be used for 
continuous variables as shown in the following sample table: 
 
 
  
 
ILLT 
 
SMT 
 Variable 
N 
Mean±SD (or %) 
N 
Mean±SD (or %) 
p-value 
 Age (yrs) 
 Male (%) 
 
 
 
The following data will be summarized using the table format above: 
• Demographics 
• Clinical Evaluations 
• Lab Tests 
• Medical History 
• Index PCI Results 
• Index PCI Complications 
• IVUS-VH Chart 
• EPC Counts 
• Medications Usage – Dosage 
 
 
Analysis of Safety Outcomes 
 
The primary safety endpoint and the secondary safety endpoints will be analyzed in two 
ways. For each type of event, the percentage of people who experience the event at 
least once will be compared using the chi-square test. Logistic regression models will be 
performed to adjust for the covariates such as age, pre-existing CAD and pre-existing 
diabetes. In addition, a similar analysis will be done for all AE and SAE that are possibly 
or probably attributable to the study intervention. An event-based analysis will also be 
done since adverse events can be recurrent. In this analysis, a non-parametric method 
called the mean cumulative function (MCF) will be used as an alternative to the above 
crude incidence rate analyses. The overall safety profiles as well as the safety profiles 
in specific subgroups will be compared for the two randomization groups including the 
times of event recurrence and censoring mechanisms.     
 
 


CCTA 0002 PREMIER – Statistical Analysis Plan V2.0 (09-30-2014)               Page 5 of 8 
Analysis of Primary Effectiveness Outcome 
 
The primary effectiveness outcome will be analyzed with intent-to-treat (ITT) approach 
by including all randomized patients regardless of crossover or drop-out. The change in 
total atheroma volume within a ≥ 20 mm long segment of the target coronary artery from 
baseline to 90-day follow-up will be analyzed using the two-sample t-test. We will define 
their change scores to be 0 if they do not have an atheroma volume measurement at 
90-day follow-up. Analysis of Covariance (ANCOVA) will be performed to evaluate the 
treatment effect on primary effectiveness outcome adjusting for age, pre-existing CAD, 
pre-existing diabetes, and baseline LDL levels. 
 
 
Analysis of Secondary Effectiveness Outcomes 
 
The intent-to-treat principle will also be applied to all the secondary effectiveness 
outcomes analyses. 
The change of %NC component of atheroma from baseline to 90-day follow-up will be 
analyzed using the two-sample t-test. The change scores for those who miss the 90-day 
follow-up assessment will be set to zero. Analysis of Covariance (ANCOVA) will be 
performed to evaluate the treatment effect on primary effectiveness outcome adjusting 
for age, pre-existing CAD, pre-existing diabetes, and baseline LDL levels. 
 
The EPC-CFU/ml of peripheral blood assessed at 4 time points (pre-PCI, post-PCI, 30-
day follow-up, and 90-day follow-up) will be analyzed via Mixed Linear models with 
random intercepts. The treatment effect, time effect and their interaction will be 
considered with or without adjusting for age, pre-existing CAD, pre-existing diabetes, 
baseline LDL levels, and other relevant covariates such as ACS, peripheral vascular 
disease, and chronic kidney disease. The statistical test of interest is the time by 
treatment interaction. Contrasts will be used in the Mixed Linear models to compare the 
differences among any specifically interested time points. 
 
The incidence of major adverse cardiovascular endpoints (MACE) including death, 
myocardial infarction, coronary revascularization, and stroke will be analyzed in two 
ways. The percentage of people who experience the event at least once will be 
compared using the chi-square test. Logistic regression models will be performed to 
adjust for the covariates such as age, pre-existing CAD and pre-existing diabetes. An 
event-based analysis will also be done since MACE can be recurrent. In this analysis, a 
non-parametric method called the mean cumulative function (MCF) will be used as an 
alternative to the above crude incidence rate analyses. The overall safety profiles as 
well as the safety profiles in specific subgroups over the whole study period will be 
compared for the two randomization groups including the times of event recurrence and 
censoring mechanisms. Time to event (survival analysis) will also compare the time to 
first MACE between treatment groups. A second survival analysis will be performed to 
identify risk predictors and to evaluate whether the observed treatment effect is modified 
by adjusting for covariates such as pre-existing CAD and pre-existing diabetes.  


CCTA 0002 PREMIER – Statistical Analysis Plan V2.0 (09-30-2014)               Page 6 of 8 
 
 
Handling Missing Data 
 
The Intent-to-treat (ITT) principle will be applied for primary and secondary 
effectiveness outcomes in which the change scores will be set to zero if the follow-up 
assessment is missing. 
 
Other approaches will also be applied in order to assess the robustness of the study 
results for assumptions about the missing data. For outcomes about atheroma volume 
and % of NC component of atheroma which will be measured only at 2 time points, the 
sensitivity analysis approaches include: 1) performing analysis only on complete data, 
2) applying the worst case scenario (set the lowest score at baseline and the highest 
score at 90-day follow-up), and 3) generating imputes for missing data with multiple 
imputation. For outcome about EPC-CFU/ml of peripheral blood which will be 
repeatedly measured at 4 time points, the sensitivity analysis approaches include: 1) 
performing analysis only on complete data, and 2) using the mixed model for repeated 
measures. 
 
 
Plan for Presenting Data to DMC 
 
Hines CSPCC will produce a progress report every six months for review by Data 
Monitoring Committee. The report will include figures for patient accrual, tables for 
baseline characteristics, site performance, data quality, treatment compliance, and 
safety issues. 
 
The frequency, reasons and timing for withdrawal from the study will be reported and 
compared by treatment group. The number of misrandomized patients, i.e. ineligible 
patients who were randomized will be reported by treatment group. The reasons for 
misrandomization will be listed. The number of patients who do not receive a study 
intervention, receive the wrong study intervention in error or intentionally receive the 
study intervention to which they were not randomly assigned will be reported by 
treatment group. Protocol deviations will also be compared by surgical site. 
 
 
  
 
ILLT 
 
SMT 
 Withdraw 
N 
% 
N 
% 
p-value 
 Reason ---  
 Too ill to participate 
  
  Timing --- 
 Post-PCI 
 30-day follow-up 
 


CCTA 0002 PREMIER – Statistical Analysis Plan V2.0 (09-30-2014)               Page 7 of 8 
 
  
# of Protocol Deviation 
# of Patients with Deviations 
 Protocol Deviation 
ILLT 
SMT 
ALL 
ILLT 
SMT 
ALL 
 
 Miss-randomization 
 F/U outside window 
 
 
 
 
The mean group change from baseline to follow-up visits of outcome measures will be 
compared between the two treatment groups using a two-sample t-test: 
 
 
  
 
ILLT 
 
SMT 
Change 
 Outcomes 
Time 
N 
Mean±SD 
N 
Mean±SD 
%Chg  p-value 
 Atheroma V 
Baseline 
  
90-day F/U 
 
 
 
 
AE and SAE will be analyzed by preferred term and system organ class. Additionally, 
incidence rates for AE and SAE will be analyzed by investigator attributed relatedness 
and severity of the events with the similar table format as the following: 
 
 
  
 
ILLT 
 
SMT 
 AE/SAE 
# of Event 
% of Patient 
# of Event 
% of Patient 
p-value 
 AE 
  
 
 SAE 
 
 
 
 
 
 
 


CCTA 0002 PREMIER – Statistical Analysis Plan V2.0 (09-30-2014)               Page 8 of 8 
Plan for Interim Analysis 
 
There will be no interim analysis for first stage of feasibility/pilot study due to the small 
sample size of 30 patients. Since DMC and FDA approve the study to continue to the 
second stage of pivotal study, there will be 2 interim looks, with the first look at the end 
of the first stage (30 patients completing the 6-month follow-up visit), and the second 
look will be 50% of the target sample size (about 64 patients) completing the 6-month 
follow-up visit as requested by FDA.  
 
Group sequential methods will be used to specify the α-levels in order to maintain the 
overall significance level at α = 0.05 for the primary effectiveness hypothesis test. The 
sequential analyses will use O’Brien-Fleming boundaries with an overall α = 0.05 for 
significance and 90% for power. EAST software will be used to obtain the interim 
monitoring rule and the adjusted sample size. The first interim look will cost 0.000 for α. 
If the critical value calculated for the interim data is greater than 4.637 or less than -
4.637, the study will be stopped with evidence that the null hypothesis H0: θ = 0 (no 
difference between two groups) is rejected and the ILLT group is either superior or 
inferior. However, if the critical value is between -4.637 and 4.637, the study will 
continue to the next interim look. The second interim look will cost 0.003 for α. If the 
critical value is greater than 2.963 or less than -2.963, the study will be stopped to reject 
the null hypothesis. However, if the critical value is between -2.963 and 2.963, the study 
will continue until the target sample size is met for final analysis.