Clinical and Translational Science Institute Prospective Longitudinal Assessment of Coinfected Subjects With HIV/Hepatitis C for Endothelial Function Study
Conditions: HIV, Hepatitis C
Sponsor: University of California, Los Angeles

Clinical and Translational Science Institute Prospective Longitudinal Assessment of 
Coinfected Subjects with HIV/Hepatitis C for Endothelial Function Study (CTSI-PLACE 
Study) 
   
 
 
A Prospective, Longitudinal Study of Endothelial Function in HIV/HCV Coinfected 
Subjects 
 
 
 
 
Sponsored by: 
 
UCLA AIDS Institute 
UCLA Center for AIDS Research 
UCLA Clinical and Translational Science Institute 
Merck Investigator Studies Program 
 
NCT02597790 
 
 
 
 
 
 
 
Protocol Co-Chairs: 
Kara W. Chew, MD 
 
 
 
Debika Bhattacharya, MD, MS 
 
 
 
 
W. David Hardy, MD 
 
 
 
Eric S. Daar, MD 
 
 
 
Wilbert C. Jordan, MD, MPH 
 
 
 
 
 
Version 4.0 
March 22, 2015 
 
 

CTSI-PLACE 
 
 
 
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3/22/15 
 
 
 
CONTENTS 
 
 
Page 
 
SITES PARTICIPATING IN THE STUDY ..................................................................................... 4 
PROTOCOL TEAM ROSTER ....................................................................................................... 5 
STUDY MANAGEMENT ............................................................................................................... 7 
PROTOCOL-SPECIFIC GLOSSARY OF TERMS ........................................................................ 8 
SCHEMA ..................................................................................................................................... 10 
1.0 
HYPOTHESIS AND STUDY OBJECTIVES .................................................................... 12 
1.1 
Hypothesis ........................................................................................................... 12 
1.2 
Primary Objectives .............................................................................................. 12 
1.3 
Secondary Objectives .......................................................................................... 12 
1.4  
Exploratory Objectives ......................................................................................... 12 
2.0 
INTRODUCTION ............................................................................................................. 13 
2.1 
Background ......................................................................................................... 13 
2.2 
Rationale ............................................................................................................. 14 
3.0 
STUDY DESIGN ............................................................................................................. 15 
4.0 
SELECTION AND ENROLLMENT OF SUBJECTS ........................................................ 16 
4.1 
Inclusion Criteria .................................................................................................. 16 
4.2 
Exclusion Criteria ................................................................................................. 17 
4.3 
Study Enrollment Procedures .............................................................................. 19 
4.4 
Coenrollment Guidelines ..................................................................................... 19 
5.0 
STUDY MEDICATIONS .................................................................................................. 19 
6.0 
CLINICAL, LABORATORY, AND ENDOTHELIAL FUNCTION EVALUATIONS ............ 20 
6.1 
Schedule of Events .............................................................................................. 20 
6.2 
Timing of Evaluations .......................................................................................... 21 
6.3 
Instructions for Evaluations ................................................................................. 22 
7.0 
CLINICAL MANAGEMENT ISSUES ............................................................................... 28 
7.1 
Toxicity ................................................................................................................ 28 
7.2 
Pregnancy ........................................................................................................... 28 
8.0 
CRITERIA FOR DISCONTINUATION ............................................................................. 28 
8.1 
Premature/Permanent Study Discontinuation ..................................................... 28 
9.0 
STATISTICAL CONSIDERATIONS ................................................................................ 28 
9.1 
General Design Issues ........................................................................................ 28 
9.2 
Endpoints ............................................................................................................. 28 
9.3 
Sample Size and Accrual .................................................................................... 29 
9.4 
Data Analysis Plan .............................................................................................. 29 

CTSI-PLACE 
 
 
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CONTENTS (Cont’d) 
Page 
 
10.0 
DATA COLLECTION AND MONITORING AND ADVERSE EVENT REPORTING ........ 30 
10.1 
Records to Be Kept ............................................................................................. 30 
10.2 
Role of Data Management ................................................................................... 31 
10.3 
Clinical Site Monitoring and Record Availability .................................................. 31 
11.0 
HUMAN SUBJECTS ....................................................................................................... 31 
11.1 
Institutional Review Board (IRB) Review and Informed Consent ........................ 31 
11.2 
Subject Confidentiality ......................................................................................... 31 
11.3 
Study Discontinuation .......................................................................................... 32 
12.0 
PUBLICATION OF RESEARCH FINDINGS ................................................................... 32 
13.0 
BIOHAZARD CONTAINMENT ........................................................................................ 33 
14.0 
REFERENCES ................................................................................................................ 32 
 
 
 
 

 
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SITES PARTICIPATING IN THE STUDY 
 
Multisite study open to all UCLA Clinical and Translational Science Institute (CTSI) sites.   
 
 
 
 
 
 

 
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PROTOCOL TEAM ROSTER 
 
Co-Chairs 
Kara W. Chew, MD 
Division of Infectious Diseases 
David Geffen School of Medicine at UCLA 
10833 Le Conte Avenue 
37-121 CHS 
Los Angeles, CA 90095 
Phone:  (310) 825-7225 
Fax:  (310) 825-3632 
E-mail: kchew@mednet.ucla.edu 
 
Debika Bhattacharya, MD 
Division of Infectious Diseases 
David Geffen School of Medicine at UCLA 
10833 Le Conte Avenue 
37-121 CHS 
Los Angeles, CA 90095 
Phone:  (310) 825-7225 
Fax:  (310) 825-3632 
E-mail: debikab@mednet.ucla.edu 
 
W. David Hardy, MD 
Division of Infectious Diseases 
Cedars-Sinai Medical Center 
8700 Beverly Blvd, Becker 221 
 
Phone: (310) 423-3896 
Fax : (310) 423-4599  
Email : david.hardy@cshs.org  
 
Eric S. Daar, MD 
Division of HIV Medicine 
Los Angeles Biomedical Research Institute 
at Harbor-UCLA Medical Center 
1124 W. Carson St., CDCRC-205 
Torrance, CA 90502 
Phone: (310) 222-2467 
Fax: (310) 533-0447 
Email:  EDaar@LABioMed.org 
 
Wilbert C. Jordan, MD 
OASIS Clinic 
King-Drew Medical Center 
Charles R. Drew University of Medicine and 
Science 
1807 E. 120th St., Los Angeles, CA 90059 
Phone: (310) 668-4213 
Email: tojo44@aol.com 
 
 
Statistician 
Chi-hong Tseng, PhD 
Division of General Internal Medicine and 
Health Services Research 
UCLA David Geffen School of Medicine 
Address: 
Phone: (310)794-3116 
Fax: (310)794-8149 
E-mail: ctseng@mednet.ucla.edu 
 
 
Study Coordinators 
Sten Witzel, RN BSN BA 
UCLA Clinical and Translational Research 
Center 
10833 Le Conte Avenue, BE-131A CHS 
Los Angeles, CA 90095 
Phone: (310) 267-1039 
Fax: (310) 206-5604 
Email: switzel@mednet.ucla.edu 
 
Gregg Sannes, RN-MN  
Division of Infectious Diseases 
Cedars-Sinai Medical Center 
8700 Beverly Blvd, Becker 221 
(310) 423-3755 
(310) 423-4599 
gregg.sannes@cshs.org 
gilda.robles@cshs.org 
 
Mario Guerrero 
Division of HIV Medicine 
Los Angeles Biomedical Research Institute 
at Harbor-UCLA Medical Center 
1124 W. Carson St., CDCRC-206 
Phone: (310) 222-3848 
 
Karen Herrigan 
OASIS Clinic 
King-Drew Medical Center 
1807 E. 120th St., Los Angeles, CA 90059 
Phone: (310) 668-4213 
Email: froglvn@aol.com 

 
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Investigators 
Judith S. Currier, MD, MSc 
Division of Infectious Diseases 
David Geffen School of Medicine at UCLA 
9911 W. Pico Blvd, Suite 980 
Los Angeles, CA 90035 
Phone: (310) 557-1891 
Fax: (310) 557-1899  
Email: jscurrier@mednet.ucla.edu 
 
Jordan Lake, MD, MSc 
Division of Infectious Diseases 
David Geffen School of Medicine at UCLA 
9911 W. Pico Blvd, Suite 980 
Los Angeles, CA 90035 
Phone: (310) 557-9679 
Fax: (310) 557-1899 
Email: jlake@mednet.ucla.edu 
 
Steven-Huy Han, MD 
Hepatology Clinical Research Center 
Pfleger Liver Institute 
David Geffen School of Medicine at UCLA 
200 UCLA Medical Plaza, Suite 214 
Los Angeles, CA 90095-7302 
Phone: (310) 794-5970 
Fax: (310) 206-4197 
E-mail: sbhan@mednet.ucla.edu 
 
Zhaoping Li, MD, PhD 
Division of Clinical Nutrition 
UCLA Center for Human Nutrition 
David Geffen School of Medicine at UCLA 
12-217 Warren Hall 
900 Veteran Ave 
Los Angeles, CA 90095 
Phone: (310) 206-1987 
Fax: (310) 267-2036 
Email: ZLi@mednet.ucla.edu 
 
C. Noel Bairey Merz, MD, FACC, FAHA 
Heart Institute 
Cedars-Sinai Medical Center 
444 S. San Vicente Blvd, Suite 600 
Los Angeles, CA 90048 
Phone: (310) 423-9680 
Fax (310) 423-9681 
Email: merz@cshs.org 
 
Regulatory 
Irma Franco-Gonzalez 
UCLA CARE Center 
9911 W. Pico Blvd, Suite 980 
Los Angeles, CA 90035 
Phone: (310) 557-1895 
Fax: (310) 557-1899 
Email: ifranco@mednet.ucla.edu 
 
 
Laboratory Technologist 
Ofelia A. Berina 
UCLA Clinical and Translational Research 
Laboratory 
Phone: (310) 825-0825 
Email: OBerina@mednet.ucla.edu 
 
 
 
 
 

 
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STUDY MANAGEMENT 
 
All questions concerning this protocol should be sent to the study Co-Chairs (Kara Chew, 
Debika Bhattacharya, David Hardy, Eric Daar, and Wilbert Jordan) via e-mail.  The team 
members should generally respond within 48 hours. 
 
Clinical Management  
For questions concerning entry criteria, toxicity management, concomitant medications, and 
coenrollment, contact the protocol co-chairs via email.  Include the protocol name, patient 
identification number (PID), and a brief relevant history. 
 
Laboratory/Peripheral arterial tonometry 
For questions specifically related to the protocol-specific assays or procedures, investigators 
should contact the co-chairs via email. 
 
Data Management 
For nonclinical questions about inclusion/exclusion criteria, case report forms (CRF), the CRF 
schedule of events, delinquencies, REDCap data entry, and other data management issues, 
contact Kara Chew (kchew@mednet.ucla.edu).  
• 
Include the PID and a detailed question. 
 
Subject Registration  
For subject registration questions or problems:  
• 
Send an e-mail message to Kara Chew (kchew@mednet.ucla.edu) and Debika 
Bhattacharya (debikab@mednet.ucla.edu).  
 
REDCap/Database/Data Entry Problems  
Contact Kara Chew (kchew@mednet.ucla.edu) or Martin Lai, Informatics Core Manager, UCLA 
Clinical & Translational Research Center (email mylai@mednet.ucla.edu, phone 310-794-9396). 
 
Protocol Document Questions 
For questions concerning the protocol document, contact the study Co-Chairs.  
 
Phone Calls 
Sites are responsible for documenting any phone calls made to protocol team members. 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
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PROTOCOL-SPECIFIC GLOSSARY OF TERMS 
 
ACE-I  
angiotensin-converting enzyme inhibitor 
ANC 
 
absolute neutrophil count 
AI 
 
AIDS Institute 
AE 
 
adverse events 
ALT 
 
alanine aminotransferase 
ARB 
 
angiotensin-receptor blocker 
ART 
 
antiretroviral therapy 
AST   
aspartate aminotransferase 
CAD 
 
coronary artery disease 
CAP 
 
College of American Pathologists 
CCB 
 
calcium-channel blocker 
CDU 
 
Charles Drew University 
CFAR  
Center for AIDS Research 
CHD 
 
coronary heart disease 
CI 
 
confidence interval 
CLIA  
Clinical Laboratory Improvement Amendments 
CRF   
Case Report Form 
CTSI  
Clinical and Translational Science Institute 
CVD 
 
cardiovascular disease 
DAA 
 
direct acting agent (ie antiviral drug) 
EAE   
expedited adverse event 
EC 
 
ethics committee 
EOT 
 
end of treatment 
FLP 
 
fasting lipid panel 
GLP 
 
good laboratory practice 
HCV   
hepatitis C virus 
HIV 
 
human immunodeficiency virus 
HDL 
 
high-density lipoprotein cholesterol 
HOMA-IR 
Homeostasis Model of Assessment - Insulin Resistance 
hsCRP high-sensitivity C-reactive protein 
IATA  
International Air Transport Association 
ICF 
 
informed consent form 
ICH  
 
International Conference on Harmonisation 
IFN 
 
interferon 
IL-6 
 
interleukin-6 
INR 
 
international normalized ratio 
IRB 
 
institutional review board 
LAB 
 
LA Biomed/Harbor-UCLA 
LDL 
 
low-density lipoprotein cholesterol 
LPC 
 
Laboratory Processing Chart 
Lp-PLA2 
lipoprotein-associated phospholipase A2 
NIH 
 
National Institutes of Health 
OHRP  
Office for Human Research Protections 
OI 
 
opportunistic infection 
PAT 
 
peripheral arterial tonometry 
PEG-IFN 
pegylated interferon 

 
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RBV   
ribavirin 
RHI 
 
reactive hyperemia index 
RNA 
 
ribonucleic acid 
SAE   
serious adverse event 
sICAM-1 
soluble ICAM-1 
sP-selectin 
soluble P-selectin 
SVR 
 
sustained virologic response 
TC 
 
total cholesterol 
TG 
 
triglycerides 
WBC  
white blood cell 
UCLA-WW 
UCLA-Westwood 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
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SCHEMA 
 
Clinical and Translational Science Institute Prospective Longitudinal Assessment of 
Coinfected Subjects with HIV/Hepatitis C for Endothelial Function Study (CTSI-PLACE 
Study) 
 
DESIGN 
 
Prospective, observational, longitudinal cohort of HIV/HCV coinfected 
subjects matched to HIV monoinfected controls with primary cross-sectional 
analysis of baseline endothelial function and circulating cardiovascular 
biomarkers.   
 
DURATION 
Subjects will be on study for a minimum of 52 weeks.  Those who initiate 
HCV treatment will be on study for at least 24 weeks after end of HCV 
treatment. 
 
SAMPLE SIZE         A total of 120 subjects will be enrolled in the study (80 HIV/HCV coinfected  
                                 and 40 HIV monoinfected subjects).  
 
POPULATION  
Group A: 80 HIV/HCV coinfected subjects 
 
Group B: 40 HIV monoinfected subjects 
 
 
 
All subjects ≥ 18 years of age without known CVD at baseline with CD4 cell 
count ≥200 cells/mm3 and HIV-1 RNA <50 copies/mL within 120 days of 
study entry and on stable antiretroviral therapy (ART) since the time of last 
CD4 and HIV-1 RNA measurement.  Those with a HCV infection must have 
HCV RNA ≥ 10,000 IU/mL.  The two groups will be frequency matched 
based upon age, race, sex, diabetes, smoking status, hormonal therapy 
and statin use, and CD4 cell count at baseline. 
 
PROCEDURES       Group A (HIV/HCV coinfected) 
 
Baseline: All subjects will undergo baseline endothelial function testing and 
laboratory assays on study entry. 
 
Follow-up: Subjects who do not initiate HCV treatment will have follow-up 
endothelial function and laboratory testing at 1 year.  Subjects who initiate 
HCV treatment will have follow-up endothelial function and laboratory 
testing at 4 weeks after completing HCV treatment.   
 
 
Group B (HIV monoinfected) 
 
Baseline:  All subjects will undergo baseline endothelial function testing and 
laboratory assays on study entry. 
 
Follow-up: All subjects will have follow-up endothelial function and 
laboratory testing at 1 year.

 
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Baseline assessments2 
Study Week 
1 Group A, HIV/HCV coinfected; Group B, HIV monoinfected 
2 Baseline assessments as described in section 6.1, including vital signs, medical history, medications, medical record abstraction, serum/plasma collection and 
processing for laboratories and storage, and PAT testing 
3 HCV treatment may be initiated at any point following study entry 
4 The chart abstraction visit will consist of data abstraction by study staff from the medical record for the preceding 6 months (from time of study entry). 
5 Follow-up assessments as described in section 6.1.  PAT and biomarker testing at 1 year for all subjects except those that initiate HCV treatment.  For those 
that initiate HCV treatment, PAT and biomarker testing should occur at 4 weeks after end of HCV treatment.  HCV-treated subjects should then continue to be 
followed until 24 weeks after end of HCV treatment and HCV viral load at 12 and 24 weeks after end of HCV treatment recorded (through medical record 
abstraction).   
HCV treatment course3 – Follow-up
Follow-up assessments5 
Group A1 
Group B1 
Follow-up
 0                                                  12                                                    24                                            36                                              52           
Study Week 
Baseline assessments2 
No HCV treatment – Follow-up   
Follow-up 
assessments5 
Follow-up 
assessments5 
  0                                                  12 
                                   24 
                                          36                                               52           
Baseline assessments2 
Chart 
abstraction4 
Chart 
abstraction4 

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1.0 
HYPOTHESIS AND STUDY OBJECTIVES 
 
1.1 
Hypotheses:  
 
1.1.1  HCV is additive with HIV in promoting CVD risk through increased immune 
activation and inflammation. 
 
1.1.2 Markers of endothelial function, inflammation, and thrombosis that are not 
produced by the liver will provide more accurate and novel biomarkers for 
assessment of CVD risk in HIV/HCV coinfected persons. 
 
1.1.3 Treatment of HCV in HIV/HCV coinfected persons will improve endothelial 
function and reduce chronic immune activation and inflammation. 
 
1.2 
Primary Objectives 
 
 
1.2.1 To establish a UCLA CTSI-wide cohort of demographically diverse HIV/HCV 
coinfected and matched HIV monoinfected men and women with HIV suppressed 
on stable antiretroviral therapy (ART). 
 
1.2.2.   To determine the effect of HCV coinfection on endothelial function in the HIV-1- 
infected cohort at baseline. 
 
1.2.3 To assess whether non-hepatic biomarkers of endothelial dysfunction,  
inflammation, and thrombosis (sICAM-1, sE-selectin, Lp-PLA2, IL-6, and D-
dimer) are better correlates of endothelial function than traditional hepatic-
derived biomarkers (hsCRP and lipids) in HIV/HCV coinfected in cross-sectional 
assessment at baseline. 
 
1.3 
Secondary Objectives  
 
1.3.1 To determine the effect of HCV coinfection on endothelial function in the HIV-1- 
infected cohort over one year of follow-up. 
 
1.3.2 To assess whether non-hepatic biomarkers of endothelial dysfunction,  
inflammation, and thrombosis (sICAM-1, sE-selectin, Lp-PLA2, IL-6, and D-
dimer) are better correlates of longitudinal endothelial function than traditional 
hepatic-derived biomarkers (hsCRP and lipids) in HIV/HCV coinfected persons  
 
1.3.3 To determine the effect of HCV coinfection on metabolic parameters (insulin 
resistance, cholesterol) in the HIV-1 infected cohort at baseline. 
 
1.3.4 To determine the effect of HCV coinfection on metabolic parameters (insulin 
resistance, cholesterol) in the HIV-1 infected cohort over one year of follow-up. 
 
1.4  
Exploratory Objectives  
 

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1.4.1  To explore the effect of HCV viremia on endothelial function in HIV/HCV  
coinfected persons. 
 
1.4.2 To explore the effect of HCV viremia on circulating CVD biomarkers in HIV/HCV 
coinfected persons.  
 
1.4.3 To explore the effect of HCV viremia on metabolic parameters (insulin resistance, 
cholesterol) in HIV/HCV coinfected persons.   
 
1.4.4 To explore the effect of HCV coinfection on circulating plasma levels of 
endothelial, platelet, and T-cell microparticles in HIV-infected persons.  
 
 
2.0 
INTRODUCTION 
 
2.1 
Background 
 
Hepatitis C virus (HCV) infection is a leading cause of morbidity and mortality among 
those with HIV-1.  Non-liver related morbidity and mortality, including cardiovascular 
outcomes, are poorly characterized in HIV/HCV coinfection. All-cause mortality is 
increased in HIV patients with HCV coinfection, independent of HIV disease 
progression.1 Not only liver-related, but non-liver related deaths may be reduced with 
HCV treatment.2,3 However, non-liver related outcomes in HIV/HCV coinfection remain 
poorly characterized.  Large retrospective observational studies have suggested an 
increased risk of CVD events in HCV-infected individuals, both with and without HIV 
infection.4-6 The contribution of HCV to elevated CVD risk in HIV patients is unknown.  
Multiple potential mechanisms for increased CVD risk with HCV infection exist, including 
hepatic steatosis, insulin resistance, and diabetes mellitus.7,8 Chronic immune activation, 
implicated in HIV-associated accelerated atherosclerosis, is further increased with HCV 
coinfection.9,10  
 
Endothelial function, a predictor of CVD risk, has not been adequately characterized in 
HIV/HCV coinfection.  Endothelial dysfunction mediates development and progression of 
atherosclerosis.11  HIV-infected patients have impaired endothelial function compared to 
non-HIV-infected patients.12 In a single small cross-sectional analysis by Masia et al, no 
difference was seen in endothelial function measured by brachial flow-mediated dilation 
(FMD) in HIV/HCV coinfected versus HIV monoinfected persons, but interpretation of 
their findings is limited by confounding related to inconsistent HIV suppression, which 
has a clear impact on these measures.13  Peripheral arterial tonometry (PAT) is an 
established, non-invasive method for assessment of endothelial function that has been 
shown to both correlate with known CVD risk factors and predict CVD risk in multiple 
non-HIV infected populations.14-16 Its characteristics of reproducibility and reduced inter-
operator variability in comparison to FMD make it an attractive tool for the assessment of 
CVD risk in HIV-infected persons.17   
 
Routine CVD risk markers are altered with HCV infection and non-hepatically produced 
markers of endothelial dysfunction and inflammation may be better predictors of CVD 
risk in this patient population.  Lipid and hsCRP levels are widely used for CVD risk 

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assessment.18,19 However, hsCRP and lipids are reduced in the setting of HCV-
associated chronic liver disease,20,21 related to impaired hepatic function and an 
incompletely understood interaction between HCV and host lipid metabolism.22 Thus, 
routine markers may underestimate CVD risk in HCV coinfected persons.  Non-hepatic 
CVD biomarkers which may have improved predictive utility in HIV/HCV coinfection 
include: soluble ICAM-1 (sICAM-1), soluble E-selectin (sE-selectin), lipoprotein-
associated phospholipase A2 (Lp-PLA2), D-dimer, and interleukin-6 (IL-6).  In large 
prospective studies, levels of these biomarkers predicted future CVD events and were 
consistently associated with classical risk factors.23-34 Lp-PLA2 has never been 
described in HCV and minimal data exist for HIV-infected individuals.35 A few studies 
have compared sICAM-1 and sE-selectin levels in HIV/HCV versus HIV- or HCV-
monoinfected controls and found higher baseline levels with HIV/HCV coinfection,29,30,36 
but none have appropriately controlled for key confounders such as HIV viremia or 
traditional CVD risk factors.  One study attempted to correlate levels of vascular 
adhesion molecules with endothelial dysfunction by FMD, but interpretation of their 
findings is similarly limited by the heterogeneity of their cohort and significant 
confounding.13 Thus, the predictive utility of these markers remains unknown. 
 
Microparticles (MPs) are a novel marker of endothelial dysfunction.  MPs are cell 
membrane blebs produced by activated and injured cells that retain the cellular 
hallmarks of their parent cells.37  While a stimulus such as infection or tissue injury is 
believed to trigger MP formation, MPs stimulate a “sterile,” self-propagating, 
inflammatory cascade. MPs are understudied in HIV infection, but T cell MP numbers 
correlate with severity of hepatic disease in HCV mono-infection,38 and endothelial cell 
MP numbers correlate strongly with degree of endothelial dysfunction in the general 
population.39,40 Similar to peripheral blood mononuclear cells (PBMCs), MP phenotyping 
and quantification via flow cytometry allows for assessment of immune activation and 
cellular dysfunction, but MPs can be measured on frozen plasma and are not subject to 
the same viability concerns as PBMCs.  
 
Successful HCV therapy may improve CVD outcomes. CVD risk factors such as hepatic 
steatosis and insulin resistance improve with HCV clearance.41-44 A growing, though still 
small, body of literature describes reduction of levels of endothelial markers and 
inflammation in subjects who responded to HCV treatment as compared to those who 
did not.29,30,36 These studies did not include an accompanying, more direct surrogate 
vascular measure of CVD risk such as endothelial dysfunction by PAT or FMD.  The 
clinical significance of the observed change in circulating biomarkers is unknown.  Our 
study will be the first to address both gaps.   
 
2.1 
Rationale 
 
In this era of aging coinfected persons on effective ART, CVD and metabolic outcomes 
must be better characterized. There are no existing prospective, longitudinal cohorts 
designed to investigate endothelial function in this population.  It is not widely recognized 
by clinicians that the hepatically-produced CVD risk markers used in routine clinical 
practice may be unreliable in coinfected persons, and data characterizing non-hepatic 
markers in this population are lacking.  Through a unique, multi-disciplinary collaboration 
between the four UCLA CTSI institutions, we will establish a HIV/HCV coinfected cohort 

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diverse with respect to race/ethnicity, gender, socioeconomic class, and mode of HCV 
transmission, in a large catchment area, and for whom outcomes have not been 
described.  The HIV/HCV cohort will be matched with HIV monoinfected controls, all with 
HIV suppression on stable ART, for cross-sectional and longitudinal evaluation of 
endothelial function, as well as improved characterization of both novel and traditional 
CVD risk biomarkers.  We will conduct novel exploratory analysis of the effect of HCV 
treatment and virologic suppression on endothelial function.  The project is intended to 
generate preliminary data for application for R01 or program project funding to continue 
the cohort and improve our understanding of CVD and metabolic outcomes with chronic 
HIV/HCV coinfection. 
 
2.2      Study Population 
There will be two study groups: Group A will be HIV/HCV coinfected subjects and Group 
B will be HIV monoinfected subjects.  All HCV coinfected subjects will have evidence of 
ongoing HCV replication on study entry.  The HIV/HCV group may include both HCV 
treatment-naïve subjects and treatment-experienced subjects who have failed prior HCV 
therapy, as ongoing HCV replication, irrespective of prior HCV treatment, is believed to 
be the primary driver of HCV-associated chronic inflammation and immune activation 
and end-organ disease.  To control for the effect of HIV infection on the outcomes of 
interest, all subjects are to have undetectable HIV RNA (<50 copies/mL) and CD4 cell 
count ≥ 200 cells/mm3.  All HCV genotypes are permitted in the study. Subjects who 
initiate HCV treatment following study entry are included to allow exploratory analysis of 
the potential benefit of HCV treatment and virologic clearance on CVD.  
 
2.3 
Selection of PAT as measure of endothelial function and correlate for CVD risk.  
 
Endothelial dysfunction represents an early stage of coronary artery disease (CAD).45 
The presence of endothelial dysfunction in coronary or peripheral vessels constitutes an 
independent predictor of cardiovascular events.46  Noninvasive measures of peripheral 
vascular endothelial function have been developed, but most are limited by operator 
dependency or complexity.47-49 
PAT is a noninvasive technique to assess peripheral microvascular endothelial function 
by measuring changes in digital pulse volume during reactive hyperemia.  PAT has 
favorable characteristics when compared to other noninvasive measures of peripheral 
vascular endothelial function with respect to reduced complexity, operator 
independence, and reproducibility.17,49  
Endothelial dysfunction is reversible and early detection of endothelial dysfunction may 
have therapeutic and prognostic implications.   Multiple studies have demonstrated 
changes in endothelial function measured by PAT within 1 year or less, including with 
interventions.49-51 
 
3.0 
STUDY DESIGN 
 
This is a prospective, observational, longitudinal study of two groups, Group A, HIV/HCV 
coinfected subjects and Group B, HIV monoinfected subjects.   The HIV/HCV subjects 

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will be frequency-matched to the HIV monoinfected subjects by age (<40, 40-49, 50-54, 
and ≥55 years), race (white, black, other), sex at birth, diabetes (yes/no), smoking status 
(current or not), HMG Co-A reductase inhibitor (statin) and hormonal therapy use, and 
CD4 cell count (200-349 and ≥350 cells/mm3) at baseline.  All subjects will be followed 
for a minimum of 1 year.  A subset of the HIV/HCV coinfected group are expected to 
initiate HCV treatment during this time and will be followed through up to 24 weeks 
following end of HCV treatment.  All subjects will be included in baseline cross-sectional 
analysis of endothelial function, measured by peripheral arterial tonometry (PAT) and 
laboratory testing for circulating CVD biomarkers. All subjects will have follow-up PAT 
and laboratory testing and clinical follow-up.  Those HIV/HCV coinfected that do not 
begin HCV treatment and all HIV moninfected will have follow-up PAT and laboratory 
testing at 1 year.  Those HIV/HCV coinfected who begin HCV treatment will have follow-
up PAT and laboratory testing at 4 weeks after end of HCV treatment, but will be 
followed clinically for up to 24 weeks after end of treatment to capture complete HCV 
treatment response data. 
 
4.0 
SELECTION AND ENROLLMENT OF SUBJECTS 
 
4.1 
Inclusion Criteria  
 
4.1.1 Men and women ≥ 18 years of age.     
 
4.1.2 Hepatitis C status:  
4.1.2.1   For HIV/HCV coinfected participants:  
4.1.2.1.1    Presence of chronic HCV infection, defined by presence of  
serum HCV RNA in a subject with HCV antibody for ≥ 180 
days, or two documented HCV RNA positive results >180 days 
apart, or positive HCV RNA with biopsy demonstrating chronic 
hepatitis.  
 
NOTE: Two separate antibody tests > 180 days apart are not 
required; only that the positive HCV antibody test should be ≥ 
180 days ago, with current serum HCV viremia as outlined in 
section 4.1.2.1.2. 
 
4.1.2.1.2 Serum or plasma HCV RNA level ≥ 10,000 IU/mL at any point 
prior to study entry and in the absence of intervening HCV 
treatment, by any laboratory that has a Clinical Laboratory 
Improvement Amendments (CLIA) certification or its 
equivalent. 
 
NOTE: This must be a quantitative HCV RNA test, not a 
qualitative HCV RNA test. 
 
4.1.2.2 For HIV monoinfected participants: 
Absence of chronic HCV infection, defined as a negative HCV 
screening antibody (+/- undetectable HCV RNA) within 1 year prior to 

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4.1.3 HIV infection, documented by any licensed rapid HIV test or HIV enzyme or 
chemiluminescence immunoassay (E/CIA) test kit at any time prior to study entry 
and confirmed by a licensed Western blot or a second antibody test by a method 
other than the initial rapid HIV and/or E/CIA, or by HIV-1 p24 antigen, or plasma 
HIV-1 RNA viral load.  
 
WHO (World Health Organization) and CDC (Centers for Disease Control and 
Prevention) guidelines mandate that confirmation of the initial test result must 
use a test that is different from the one used for the initial assessment. A reactive 
initial rapid test should be confirmed by either another type of rapid assay or an 
E/CIA that is based on a different antigen preparation and/or different test 
principle (eg, indirect versus competitive), or a Western blot or a plasma HIV-1 
RNA. 
 
4.1.4 CD4+ T-cell count > 200 cells/mm3, and none less than 200 cells/mm3, during the 
12 weeks prior to study entry, at any laboratory that has CLIA certification or its 
equivalent. 
 
4.1.5   Plasma HIV-1 RNA < 50 copies/mL at screening, and none > 200 copies/mL, 
during the 12 weeks prior to study entry, at any laboratory that has CLIA 
certification or its equivalent.   
 
4.1.6 On continuous and stable ART for at least 12 weeks with CD4+ T cell count and 
plasma HIV-1 RNA measurements as in 4.1.4 and 4.1.5.  Changes in regimen 
prior to study entry are allowed as long as they were not for treatment failure, in 
which case subjects should be on the same ART regimen for at least 8 weeks 
prior to study entry with documented HIV RNA <50 copies/mL on this regimen.  
Subjects should have no plans to change ART in the next 52 weeks.  
 
4.1.7 Ability and willingness of subject to provide written informed consent. 
 
 
4.2  Exclusion Criteria   
 
4.2.1 Known cardiovascular disease, as defined by a self-reported history or medical 
record documentation of any of the following: myocardial infarction, angina 
pectoris, prior revascularization (coronary artery bypass graft, percutaneous 
coronary intervention), peripheral vascular disease, arrhythmias, congestive 
heart failure, congenital heart disease, cerebrovascular disease (stroke, transient 
ischemic attack). 
 
4.2.2 Diabetes requiring insulin therapy or hemoglobin A1c > 8% within 6 months prior 
to study entry. 
 

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4.2.3 Inability to conform to the following drug interruptions for PAT testing, whether 
due to safety (determined by the investigator) or willingness: No caffeine for 24 
hours prior; no nicotine for 4 hours prior; no recreational or prescription stimulant 
use for 24 hours prior; stopping of beta blockers, short-acting calcium channel 
blockers (CCBs), nitrates, angiotensin-converting enzyme inhibitors (ACE-Is), 
angiotensin-receptor blockers (ARBs), and renin-inhibitors for 24 hours prior; and 
stopping of long acting CCBs 48 hours prior to testing. 
 
4.2.4 Evidence of decompensated liver disease manifested by presence of or history of 
ascites, variceal bleeding, or hepatic encephalopathy.  
 
4.2.5 Other known causes of significant liver disease including chronic or acute 
hepatitis B, acute hepatitis A, hemochromatosis, or homozygote alpha-1 
antitrypsin deficiency. 
 
4.2.6 Serious illness including acute liver-related disease and malignancy requiring 
systemic treatment or hospitalization within 12 weeks prior to study entry.   
 
4.2.7 Presence of active or acute AIDS-defining opportunistic infections (OIs) within 12  
  
weeks prior to study entry, which may affect CVD biomarkers. A list of AIDS-
defining opportunistic infections as defined by the CDC, can be found in 
Appendix B of the following document: 
http://www.cdc.gov/mmwr/preview/mmwrhtml/00018871.htm 
 
4.2.8 History of major organ transplantation with an existing functional graft and on 
immunosuppressive therapy. 
 
4.2.9 History of known vascular disorder or autoimmune processes including Crohn’s 
disease, ulcerative colitis, severe psoriasis, rheumatoid arthritis, and 
cryoglobulinemia that may affect vascular studies.  
 
4.2.10 Pregnancy. 
 
4.2.11 HCV treatment (any approved or investigational agents) within 24 weeks prior to 
study entry. 
 
4.2.12 Use of immune-based therapies or systemic corticosteroids which may affect 
vascular studies or inflammatory/endothelial biomarkers within 12 weeks prior to 
study entry.  
 
NOTE:  Routine standard of care, including hepatitis A and/or B, human 
papilloma virus, influenza, pneumococcal, tetanus, and meningococcal 
vaccines are permitted if administered at least 14 days before biomarker 
blood collections and peripheral arterial tonometry examinations.   
 
4.2.13 Advanced renal insufficiency as defined by glomerular filtration rate (GFR) < 30 
mL/min/1.73 m2 or treatment by dialysis. 
 

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4.3 
Study Enrollment Procedures 
 
4.3.1 Prior to implementation of this protocol, and any subsequent full version  
amendments, each site must have the protocol and the protocol consent form 
approved, as appropriate, by their local institutional review board (IRB)/ethics 
committee (EC) and any other applicable regulatory entity (RE). Upon receiving 
final approval, sites will submit proof of IRB approval and site-specific informed 
consent forms (ICFs) to Kara Chew (kchew@mednet.ucla.edu) and UCLA-
Westwood (UCLA-WW) regulatory staff (Irma Franco-Gonzalez, ifranco-
gonzalez@mednet.ucla.edu).  They will ensure that all of the required documents 
have been received. 
 
All site-specific and primary UCLA-WW IRB documents should be retained in the 
site’s regulatory files.   
 
Upon receiving final IRB/EC and any other applicable RE approvals for an 
amendment, sites should implement the amendment immediately. Sites should 
submit proof of approved amendments for their site via email to Kara Chew and 
UCLA-WW regulatory staff (Irma Franco-Gonzalez).  A copy of the amendment 
should be retained in the site's regulatory files. 
 
Once a candidate for study entry has been identified, details will be carefully 
discussed with the subject. The subject (or, when necessary, legal guardian if the 
subject is under guardianship) will be asked to read and sign the approved 
protocol consent form.  
 
For subjects from whom a signed informed consent has been obtained, a CTSI-
PLACE Study Screening Checklist must be entered through REDCap, the study 
database. 
 
4.3.2 Subject Registration 
 
For subjects from whom informed consent has been obtained, but who are 
deemed ineligible or who do not enroll into the initial protocol step, a CTSI-
PLACE Study Screening Failure Result form must be completed and keyed into 
the database.  
 
4.4 
Coenrollment Guidelines.  Coenrollment into any study requires study team approval. 
 
5.0 
Study Medications.  No medications are being given as part of this study, but 
medications are relevant to the study questions and should be documented at each 
study visit. 
 
5.0.1   Concomitant Medications 
 
Whenever a concomitant medication is initiated or a dose changed, investigators 
must document the change in source documents and CRFs.  

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5.0.2 Required Medications 
 
All antiretroviral regimens are acceptable. 
 
5.0.3 Prohibited Medications 
 
 
There are no prohibited medications specific to this study. 
  
 
 
6.0 
CLINICAL, LABORATORY, AND ENDOTHELIAL FUNCTION EVALUATIONS 
 
6.1 
Schedule of Events 
 
Evaluation 
Screening 
 
Entry 
Week 
0 
Chart 
abstraction 
Study 
Visit 
Premature Study 
Discontinuation1 
Week  
Week 24 
Week 
522 
 
Window 
±28 days 
±28 days 
 
Documentation of HCV Status* 
X 
 
 
X 
X 
Documentation of HIV-1 Infection* 
X 
 
 
 
 
Medical History/Medication History 
X 
X 
X 
X 
X 
Clinical Assessment3 
X 
X 
  
X  
X 
Concomitant Medications 
 
X 
X 
X 
X 
Hematology/Chemistry/Liver 
Function Tests* 
 
X 
  
X 
X 
INR* 
 
X 
 
X 
X 
CD4+/CD8+ T-Cell Counts* 
X 
  
X 
X 
X 
Plasma HIV-1 RNA* 
X 
  
X 
X 
X 
Serum HCV RNA (screening)* 
X 
 
 
 
 
Serum HCV RNA (on-study)* 
 
  
X 
X 
X 
HCV Genotype Test* 
 
X  
 
 
 
Hemoglobin A1c* 
X 
 
 
X 
X 
Pregnancy test 
X 
 
 
X 
X 
Liver biopsy results* 
 
X 
 
X 
X 
Fasting Stored Serum/Plasma for 
Batched CVD Biomarkers4 
 
X 
 
X 
X 
Stored Serum/Plasma for Future 
Studies 
 
X 
 
X 
X 
Peripheral Arterial Tonometry  
 
X 
 
X 
X 
 
*Longitudinal values of these laboratory tests, if available, will be obtained via chart abstraction from the participants’ 
medical records. Specific tests will be performed in real-time only as indicated in the instructions for evaluations. INR 
should be collected for HCV coinfected subjects only.  Hemoglobin A1c should be collected for diabetic subjects only.   
Availability of liver biopsy results will be assessed and, if available, results recorded from the medical record. Liver 
biopsy will not be performed for this study. For HIV monoinfected subjects, screening HCV antibody should be 
performed at the week 52 visit.  

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1All subjects enrolled in the study for a minimum of 3 months but less than 1 year should have a premature study 
discontinuation visit, including repeat PAT and biomarker testing.  If this testing occurs while on HCV treatment, 
concurrent treatment status should be documented in source documents and on the CRF. For subjects enrolled in the 
study for fewer than 3 months, telephone interview and chart abstraction should be conducted for clinical data from 
the time of study entry to study discontinuation.   
2For coinfected subjects who initiate HCV treatment, week 52 assessments should occur at 4 weeks after end of 
treatment (EOT), but no earlier than 3 weeks and no later than 8 weeks after EOT.  If not available in the medical 
record, a serum HCV RNA should be performed at 4 weeks after EOT.  The medical record should be reviewed and 
HCV RNA at 12 and 24 weeks after EOT should be recorded.  
3Required elements for clinical assessments are described in section 6.3.   
4Batched biomarkers will include fasting lipid panel, fasting insulin, fasting glucose, hsCRP, IL-6, D-dimer, sICAM-1, 
sE-selectin, and Lp-PLA2. Fasting insulin and fasting glucose should only be performed on non-diabetic subjects. 
Plasma will also be tested for microparticles. 
 
 
 
6.2 
Timing of Evaluations 
 
6.2.1 Screening Evaluations  
 
Screening evaluations must occur within 4 weeks prior to the subject starting any 
study procedures. 
 
6.2.2 Entry Evaluations 
 
Entry evaluations must occur within 4 weeks of the screening visit.  
 
6.2.3 Post-Entry Evaluations 
 
On-Study Evaluations  
Evaluations should occur after registration. The week 24 visit consists only of 
review of the medical record and abstraction of data from the time since study 
entry.  The week 52 study visit should be scheduled within the visit window.  For 
HCV-infected subjects who initiate HCV treatment, the week 52 visit should occur 
at 4 weeks after end of HCV treatment, but not earlier than 3 weeks and no later 
than 8 weeks after EOT.   
 
Study Completion Evaluations 
Clinical assessment and laboratory evaluation, as outlined in section 6.1, will be 
performed at study completion (week 52 visit). 
 
6.2.4 Discontinuation Evaluations 
 
Evaluations for Registered Subjects Who Do Not Start Study Procedures 
All CRFs must be completed and keyed for the period up to and including week 
0. 
 
Subjects who discontinue the study prior to 1 year 
Subjects who permanently discontinue study prior to 1 year and have been 
enrolled for at least 3 months will have a study discontinuation visit including 

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repeat biomarker and PAT testing.  Subjects on study for < 3 months should be 
contacted by phone for a telephone interview and have medical record review 
and data abstraction for clinical events, including new diagnoses and change in 
medications, since the time of study entry.   
 
Pregnancy 
Female subjects who conceive after study entry will be discontinued from the 
study.   
 
 
6.3 
Instructions for Evaluations 
 
All clinical, laboratory, and tonometry information required by this protocol is to be 
present in the source documents.  All stated evaluations are to be recorded on the CRF 
and keyed into the database unless otherwise specified. This includes events that meet 
the following International Conference on Harmonisation (ICH) definitions for a serious 
adverse event (AE): 
 
• 
Results in death 
• 
Life-threatening 
• 
Requires inpatient hospitalization or prolongation of existing hospitalization 
• 
Results in persistent or significant disability/incapacity  
• 
Congenital anomaly/birth defect 
• 
Other important medical event (may not be immediately life-threatening or result in 
death or hospitalization, but may jeopardize the patient or may require intervention to 
prevent one of the events listed above).   
 
  
6.3.1 Documentation of HIV-1 Infection 
 
Please refer to section 4.0 regarding assay requirements for HIV-1 
documentation.   Year of HIV diagnosis and nadir CD4 cell count should be 
recorded on the CRF.   
 
6.3.2 Documentation of HCV Infection 
 
Please refer to section 4.0 regarding requirements for HCV documentation for 
both HIV/HCV coinfected and HIV monoinfected subjects.   Year of HCV 
diagnosis and mode of HCV transmission should be recorded on the CRF.   
 
6.3.3 Documentation of Liver Biopsy 
 
If a liver biopsy has been performed within 5 years prior to study entry or is 
performed during the course of the study, documentation of date of biopsy, liver 
fibrosis and activity scores, steatosis measurement, and scoring system (e.g. 
METAVIR, Knodell, etc.) should be recorded on the CRF.   
 
6.3.4 Medical History 

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The medical history must include all diagnoses, year of initial diagnosis, and 
status of diagnosis (ongoing condition or not). Any allergies to medications and 
their formulations must be documented.  Record tobacco, alcohol, and drug use 
history on the CRF.  Record menopause status on the CRF.  
 
6.3.5 Medication History 
 
A medication history must be present, including start and stop dates. All current 
medications should be recorded.  The table below lists the medications that must 
be included in the history.  
 
Medication Category 
Complete History or Timeframe 
Record on 
CRFs 
HIV treatment 
Within 120 days prior to entry 
Yes 
HCV treatment 
Complete History 
Yes 
Other prescription drugs 
Within 42 days prior to entry 
Yes 
Non-prescription drugs  
Within 42 days prior to entry and 
taken daily for at least 7 days 
Yes 
Complementary and 
alternative medicines 
Within 42 days prior to entry and 
taken daily for at least 7 days 
Yes 
 
6.3.6 Clinical Assessments 
 
Vital Signs  
Blood pressure and heart rate will be recorded at screening and entry.  
 
Height 
Height will be measured at entry. 
 
Weight 
Weight will be measured at entry and week 52 visit. 
 
Waist Circumference 
Waist circumference will be measured at entry and week 52/study 
discontinuation visit. Measure waist circumference at level of iliac crest in 
centimeters (cm).  
 
Signs and Symptoms 
At entry, all signs and symptoms that occurred within 42 days before entry must 
be recorded.  At week 52/study discontinuation visit, all signs and symptoms that 
occurred within 42 days prior must be recorded. 
 
6.3.7 Concomitant Medications 
 
Initiation and discontinuation of all concomitant medications should be recorded 
on the CRF with start and stop dates. All ART (including interruption of ART for at 
least 7 days), HCV treatment (approved or experimental), immune-based and 

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anti-inflammatory therapy, prescription drugs, vaccines, alternative therapies, 
dietary supplements, and any experimental therapies taken daily for at least 7 
days must be recorded. Changes only in dose or formulation do not need to be 
recorded. 
 
At the premature study discontinuation visit, record all concomitant medications 
and ART per above guidelines, including ongoing medications. 
 
6.3.8 Laboratory Evaluations (Note: the majority of these will be collected from the 
medical records and will not be drawn for the study) 
 
The following must be recorded on CRFs and keyed into the database unless 
otherwise indicated: 
 
Hematology 
Hemoglobin, hematocrit, white blood cell count (WBC), absolute neutrophil count 
(ANC), and platelets within 120 days prior to study entry and within 120 days 
prior to the week 52/study discontinuation visit. 
 
Chemistry 
Sodium, potassium, chloride, bicarbonate, blood urea nitrogen (BUN), creatinine, 
and glucose within 120 days prior to study entry and within 120 days prior to 
the week 52/study discontinuation visit. 
 
Liver Function Tests 
Total bilirubin, indirect bilirubin, direct bilirubin, aspartate aminotransferase 
(AST), alanine aminotransferase (ALT), and alkaline phosphatase, and albumin 
within 120 days prior to study entry and within 120 days prior to the week 
52/study discontinuation visit. 
 
INR 
INR obtained at a local laboratory within 120 days prior to study and within 120 
days prior to the week 52/study discontinuation visit.  
 
Pregnancy Test 
For women with reproductive potential: Serum or urine beta-human chorionic 
gonadotropin (β-HCG [urine test must have a sensitivity of at least 50 mIU/mL]) 
within 28 days prior to study entry and at week 52/study discontinuation visit.  
 
Hepatitis C Antibody Test 
For HIV-monoinfected subjects only, screening HCV antibody within 1 year 
prior to study entry and within 120 days prior to the week 52/study 
discontinuation visit.  
 
Hemoglobin A1c 
For diabetic subjects only, hemoglobin A1c obtained within 180 days prior 
to study entry and within 120 days prior to the week 52/study 
discontinuation visit. 

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6.3.9 Immunologic Studies 
 
CD4+/CD8+ T-cell counts 
Absolute CD4+/CD8+ T-cell count and percentages within 12 weeks prior to 
entry from a laboratory that possesses CLIA certification or an equivalent. CD4 
nadir should be recorded on the CRF.  At the week 24 and week 52 visits, the 
medical record should be reviewed and all post-entry values obtained since the 
last study visit should be recorded.   
 
 
 
 
6.3.10 Virologic Studies 
 
Plasma HIV-1 RNA 
Plasma HIV-1 RNA level within 12 weeks prior to study entry and within 28 days 
of week 52/study discontinuation visit, at any laboratory that has CLIA 
certification or its equivalent.   
 
Subjects who switch their ART regimen prior to study entry (see 4.1.6) should 
have an HIV-1 RNA level performed while on the new regimen, prior to study 
entry.  Record post-entry values obtained as part of routine care.   
 
The requirement of plasma HIV-1 RNA level within 28 days of the week 
52/study discontinuation visit is waived for subjects that meet ALL of the 
following criteria:  
 
1) HIV-1 RNA <50 copies/mL on all laboratory testing in the 
preceding 2 years  
2) HIV-1 RNA <50 copies/mL in the preceding 90 days 
3) No change in ART regimen since study entry 
4) No ART interruption of more than 7 consecutive days or 14 total 
days in the preceding 90 days 
5) No HCV treatment since study entry 
 
For subjects who meet ALL of the above criteria, the most recent HIV-1 
RNA level from the medical record, closest to the time of the week 52/study 
discontinuation visit, is acceptable.  
 
 
Serum HCV RNA (screening) 
At screening, the HCV RNA result must be obtained by any FDA-approved test 
for quantifying HCV RNA at any local laboratory that has a CLIA certification or 
its equivalent. Screening HCV RNA at any point prior to study entry is 
acceptable, in the absence of intervening treatment.  
 
Serum HCV RNA (on-study evaluations) 
At the week 24 and week 52 visits, the medical record should be reviewed and all 
post-entry values obtained since the last study visit should be recorded.   

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Coinfected subjects who begin HCV treatment should have HCV RNA testing at 
4 weeks after EOT (“week 52” visit), but no earlier than 3 weeks and and no later 
than 8 weeks after EOT.  This may be abstracted from the medical record and if 
not done, should be performed using a CLIA-certified or equivalent laboratory.  
The medical record should be reviewed and HCV RNA at weeks 12 and 24 after 
EOT should be documented.   
 
HCV Genotype Test 
A documented result (serum or plasma) is a source document from a CAP/CLIA-
approved laboratory at any time prior to entry. For those subjects lacking a 
documented result, HCV genotyping should be performed by a local CAP or 
CLIA-approved (or its equivalent) laboratory. 
 
6.3.11 Stored Plasma and Serum for batched testing for biomarkers 
 
Stored plasma and serum will be collected at the indicated visits (section 6.1) for 
centralized, batched testing at the UCLA Clinical and Translational Research Laboratory 
(CTRL) for the following biomarkers: hsCRP, fasting glucose, fasting insulin, fasting lipid 
panel, hsCRP, sICAM-1, sE-selectin, Lp-PLA2, IL-6, and D-dimer.  Fasting glucose and 
fasting insulin will be performed for non-diabetic subjects only.  Subjects must be fasting 
at least 8 hours (nothing by mouth except medications and water). If subjects indicate a 
non-fasting state, they should be asked to return in a fasting state within the required 
visit window (please refer to section 6.1 for more information regarding visit window). If 
that is not possible, a non-fasting specimen should be obtained the same day, and the 
non-fasting state should be recorded on the CRF.  Stored plasma will be collected at the 
indicated visits (section 6.1) for centralized testing for endothelial, monocyte, and T-cell 
microparticles in the laboratory of Dr. Otto Yang (UCLA Division of Infectious Diseases).  
Samples should be collected, processed, and stored on-site frozen at -70°C until 
shipped to UCLA-WW in accordance with the CTSI-PLACE Study Lab Processing Chart 
(LPC) (Appendix II). Subjects should not have received vaccines within 14 days 
prior to plasma and serum collection.  
 
6.3.12 Stored Plasma and Serum for future studies 
 
Stored plasma and serum will be collected and processed at the indicated visits 
(section 6.1) for future studies.  Samples will be temporarily stored on site at -
70°C until transfer to UCLA-WW for central long-term storage.  Samples should 
be transferred to UCLA-WW every 1-2 months.  
 
6.3.13 Peripheral Arterial Tonometry testing 
 
Endothelial function will be measured by the FDA-approved method of Endo PAT 
2000 (Itamar® Medical Ltd, Caesarea, Israel) at baseline and 52 weeks in all 
subjects except those undergoing HCV treatment.  Those undergoing HCV 
treatment will have follow-up PAT at 4 weeks after end of HCV treatment.  PAT 
will be done at two sites: at CSMC by Dr. C. Noel Bairey Merz and at the UCLA 
Center for Human Nutrition by Dr. Zhaoping Li.  The device consists of 
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accompanying pulse waves.17,46 A blood pressure cuff will be placed on one 
upper arm (study arm), while the other arm will serve as a control (control arm).  
Peripheral arterial tonometry probes will be placed on one finger of each hand for 
continuous recording of the PAT signal (baseline arterial pulse wave amplitude, 
PWA).  After a 5-minute equilibration period, the blood pressure cuff will be 
inflated to suprasystolic pressures for 5 minutes (the ischemic stimulus).  The 
cuff will then be deflated while PAT recording is continued for 5 minutes (Figure 
1).  Reactive hyperemia index (RHI) is the primary measure of endothelial 
function.  The RHI is automatically derived using Endo PAT version 3.0.4 
software, as the ratio of the average PWA over a 1 minute interval starting 1 
minute following cuff release to the pre-occlusion PWA (average amplitude over 
3.5 minutes before cuff inflation), normalized to the control arm.  
 
                 
 
Figure 1 Representative reactive hyperemia peripheral arterial tonometry recordings of 
subjects with normal and abnormal reactive hyperemic response. Normal response is 
characterized by a distinct increase in the signal amplitude after cuff release compared 
with baseline.  
 
The choice to use the average 1-min PAT signal starting 1 min after cuff deflation 
to describe the magnitude of reactive hyperemia is based on the observation that 
this time interval provides the best information regarding detection of coronary 
endothelial dysfunction as determined by receiver operating characteristic (ROC) 
curve analysis.  
 
Subjects will be required to be fasting for 12 hours and to abstain from the following prior 
to PAT: caffeine for 24 hours; nicotine for 4 hours; beta blockers, calcium-channel 
blockers (CCBs), nitrates, angiotensin-converting enzyme inhibitors, angiotensin-
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prescribed or recreational stimulant use for 24 hours; vigorous exercise for 12 hours. 
Subjects should not have received vaccines within 14 days prior to PAT testing.  
 
7.0 
CLINICAL MANAGEMENT ISSUES 
 
No clinical management issues are expected as no treatment is being provided in this 
study.   
 
7.1 
Toxicity  
 
No toxicities are anticipated as no treatment or invasive procedures are being performed 
as part of this study.  
 
7.2 
Pregnancy 
 
Pregnancy is not a contraindication to PAT or laboratory testing, but may affect 
measurements of endothelial function.  Pregnancy status should be documented at each 
PAT visit for female subjects (week 0 and week 52 or 4 weeks after EOT for those with 
treated HCV).    
 
 
8.0 
CRITERIA FOR DISCONTINUATION 
 
8.1 
Premature/Permanent Study Discontinuation 
 
• 
Request by the subject to withdraw. 
• 
Request of the primary care provider if s/he thinks the study is no longer in the best 
interest of the subject. 
• 
Subject judged by the investigator to be at significant risk of failing to comply with the 
provisions of the protocol as to interfere with the validity of the study results. 
• 
At the discretion of the institutional review board (IRB), Office for Human Research 
Protections (OHRP), investigator, or sponsors.  
 
 
9.0 
STATISTICAL CONSIDERATIONS 
 
9.1 
General Design Issues 
 
The CTSI-PLACE Study consists of single-arm evaluation to assess endothelial function 
cross-sectionally and longitudinally in two study populations: HIV/HCV coinfected (Group 
A), a subset of which will undergo HCV treatment, and HIV monoinfected (Group B). The 
primary endpoint is endothelial function measured by peripheral arterial tonometry 
reactive hyperemia index (RHI) in both groups. 
 
9.2 
Endpoints 
 
9.2.1 Primary Endpoints 
 

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9.2.1.1    Reactive hyperemia index (RHI) by PAT at baseline. 
 
9.2.2 Secondary Endpoints 
 
9.2.2.1 Insulin resistance by HOMA-IR (fasting insulin (µU/mL) x fasting glucose 
(mg/dl)/405) at baseline and week 52. 
 
9.2.2.2 Framingham risk score for 10-year CHD and 10-year CVD risk at 
baseline and week 52.  
 
9.2.2.3 Cardiovascular biomarkers (fasting lipid panel, hsCRP, IL-6, D-dimer, 
sICAM-1, sE-selectin, Lp-PLA2) at baseline and week 52. 
 
9.2.2.4 RHI by PAT at week 52  
 
9.2.2.5  Change in RHI , biomarkers, HOMA-IR, and FRS from baseline to week 
52.  
 
 
 
9.3 
Sample Size and Accrual 
 
9.3.1 The primary comparison is of baseline RHI between the HIV/HCV coinfected and 
HIV monoinfected groups. The estimated difference in RHI between the 2 groups 
is assumed to be the difference between high-risk CVD (HIV/HCV) and 
moderate-risk CVD (HIV).  RHI is expected then to be 1.6 in HIV/HCV and 1.3 in 
HIV, with standard deviation (SD) of 0.37.52  With a sample size of 80 HIV/HCV 
and 40 HIV, we have an estimated power of 99% for detection of the expected 
difference in RHI between the HIV/HCV and HIV groups.  However, if we 
conservatively estimate the difference in baseline RHI to be 0.2 between the 2 
groups, we will have 80% power with this sample size. For select secondary 
analyses of differences in the CVD biomarkers of sICAM-1 (expected difference 
of 779 vs1284 ng/mL, SD=526) and sE-selectin (118 vs 223 ng/mL, SD=190),30 
we will have 99% and 81% power, respectively, with this sample size.  
 
Accrual of about 20 subjects per month is expected, to complete enrollment in 6 
months. 
 
9.3.2 With 80 HCV/HIV subjects, we will have 80% power to detect a correlation 
between any one marker and RHI of > 0.31.  With 40 HIV subjects, we will have 
80% power to detect a correlation >0.42, correlations that have been seen in 
other populations.53,54   
 
9.3.3 Among 80 HIV/HCV subjects, we expect about 40 (50%) will initiate HCV 
treatment. Out of 40 who initiate HCV treatment, we expect about 27 (67%) will 
be responders and 13 will be non-responders. To compare responders (n=27) 
with subjects without treatment (n=40), we will have 80% power to detect a 
difference of 0.7 in effect size. To compare HIV monoinfected controls (n=40) 

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and untreated HIV/HCV subjects, we will have 80% power to detect a difference 
of 0.63 in effect size.  
 
9.4       Data Analysis Plan  
 
9.4.1    Objective: To determine the effect of HCV coinfection on endothelial function in  
the HIV-1-infected cohort at baseline. 
 
Descriptive statistics, including mean, SD, median, minimum, maximum, and  
interquartile range, will be provided for the overall cohort, by HCV status, and by 
PAT performance site for baseline patient characteristics, RHI, CVD biomarkers, 
HOMA-IR, hemoglobin A1c, and FRS.  Histograms and scatter plots will be 
produced to visually inspect the distributions and pairwise association. Log-
transformation as necessary will be done for RHI, if its distribution is found to be 
skewed. ANOVA (analysis of variance) will be used to compare continuous 
variables by HCV status and site and Chi-square test will be used to compare 
categorical variables. All tests will be 2-sided and a p-value less than 0.05 will be 
considered statistically significant. 
 
Two sample t-test or Wilcoxon rank sum will be performed to compare RHI in the 
HIV/HCV vs HIV groups. Multiple linear regression analysis will also be carried 
out to compare RHI in HIV/HCV vs HIV, adjusting for PAT performance site and 
any differences in baseline characteristics including CD4 nadir and ART type.  
Subgroup analyses by race/ethnicity and gender will be conducted.  Similar 
analyses will be carried out to compare biomarkers between the HIV/HCV and 
HIV groups. Given multiple comparisons, in addition to unadjusted p-values, we 
will report significance according to Benjamini-Hochberg false discovery rate 
method.55    
 
 
9.4.2 Objective: To assess whether non-hepatic biomarkers of endothelial dysfunction,  
inflammation, and thrombosis (sICAM-1, sE-selectin, Lp-PLA2, IL-6, and D-
dimer) are better correlates of endothelial function than traditional hepatic-
derived biomarkers (hsCRP and lipids) in HIV/HCV-coinfected in cross-sectional 
assessment. 
 
 
Correlation analysis (Pearson or Spearman) will be conducted to evaluate the 
association of each circulating biomarker as the predictor and RHI as a 
continuous outcome variable.  Further correlation of biomarkers with each other 
and with RHI will be performed by use of correlation matrices.   Multiple linear 
regression models will be used while controlling for confounders as in Aim 1 
analysis, but with each biomarker as the primary independent variable to 
determine the contribution of each biomarker as a predictor of RHI.  A final model 
will be constructed by including all significant biomarkers to predict RHI. The 
adjusted R-squared value will be used to indicate the predictive ability of the 
model.  Similar analysis will be carried out in HIV-monoinfected subjects only to 
derive the RHI prediction model. The models for each cohort will be compared to 
each other for their predictive ability and differences in individual biomarker 

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significance in each cohort.  This analysis will allow us to define whether non-
hepatically-derived biomarkers have stronger correlation with RHI than 
hepatically-derived biomarkers.   
 
9.4.3 Objective: To determine the effect of HCV co-infection and HCV virologic 
suppression post treatment on endothelial function and CVD biomarkers in the 
HIV-1-infected cohort over one year of follow-up. 
 
Change in RHI and change in biomarkers from baseline to follow-up will be 
compared among 4 groups of subjects (HCV treatment responders and non-
responders, untreated HIV/HCV subjects, and HIV controls). Boxplots will be 
used to visually display the difference among 4 groups. Pair-wise comparison will 
be carried out based on 2-sample t-test or Wilcoxon test. ANCOVA will be carried 
out to compare among 4 groups adjusting for baseline values and demographic 
characteristics and other baseline confounders. Adjusted comparison between 
any 2 groups will be estimated based on appropriate contrast. Correlation 
analysis will be carried out between change in biomarkers and change in RHI.  
 
 
10.0 
DATA COLLECTION AND MONITORING AND ADVERSE EVENT REPORTING 
 
10.1 
Records to Be Kept 
 
Case report forms (CRF) will be provided for each subject. Subjects must not be 
identified by name on any CRFs. Subjects will be identified by an assigned patient 
identification number (PID) in the REDCap database.  Data must be keyed in to the 
study database, REDCap.   
 
No personally identifiable information (PII) may be entered into REDCap.  A code key 
must be maintained at each site for all subjects who sign the study informed consent 
form.   
 
Users of REDcap must have completed HIPAA training or access will not be assigned.  
All REDCap operations will be tracked through audit trail.  REDCap is a web-based data 
management system.  The REDCap web server uses SSL encryption and system 
updates are automatically applied when they are available.  For added security, the 
REDCap database server sits within a firewall.  Both the web and database servers are 
backed up daily. 
 
 
10.2 
Role of Data Management 
 
10.2.1 Instructions concerning the recording of study data on CRFs will be provided. 
Each CTSI site is responsible for keying the data in a timely fashion. 
 
10.2.2 It is the responsibility of the CTSI-PLACE Study Co-PIs to assure the quality of 
computerized data for the study. This role extends from protocol development to 
generation of the final study databases. 

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10.3 
Clinical Site Monitoring and Record Availability 
 
10.3.1 UCLA-WW investigators will serve as monitors for the other sites and at UCLA-
WW the CTU coordinator (Cara O’Connor) will serve as the site monitor.  
Monitors will review the individual subject records, including consent forms, 
CRFs, supporting data, laboratory specimen records, and medical records 
(physicians’ progress notes, nurses’ notes, individuals’ hospital charts), to ensure 
protection of study subjects, compliance with the protocol, and accuracy and 
completeness of records. The monitors also will inspect sites’ regulatory files to 
ensure that regulatory requirements are being followed and sites’ laboratories 
and freezers to review specimen storage and management.   
 
10.3.2 The site investigator will make study documents (eg, consent forms, code key, 
CRFs) and pertinent hospital or clinic records readily available for inspection by 
the local IRB, the site monitors, the OHRP, and the sponsors for confirmation of 
the study data.  
 
 
11.0 
HUMAN SUBJECTS 
 
11.1 
Institutional Review Board (IRB) Review and Informed Consent 
 
This protocol and the informed consent document will be reviewed and approved by the 
IRB responsible for oversight of the study. A signed consent form will be obtained from 
the subject (or legally guardian, or person with power of attorney for subjects who cannot 
consent for themselves). The consent form will describe the purpose of the study, the 
procedures to be followed, and the risks and benefits of participation. A copy of the 
consent form will be given to the subject’s legal guardian, and this fact will be 
documented in the subject’s record. 
 
11.2 
Subject Confidentiality 
 
All laboratory specimens, evaluation forms, reports, and other records that leave the site 
will be identified only by coded number to maintain subject confidentiality. All records will 
be kept locked. All computer entry and networking programs will be done with coded 
numbers only. Clinical information will not be released without written permission of the 
subject, except as necessary for monitoring by the IRB, OHRP, or sponsors.  
 
11.3 
Study Discontinuation 
 
The study may be discontinued at any time by the IRB, OHRP, or government agencies 
as part of their duties to ensure that research subjects are protected.  
 
 
12.0 
PUBLICATION OF RESEARCH FINDINGS 
 

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Publication of the results of this study will be determined by the study co-chairs. 
 
 
13.0 
BIOHAZARD CONTAINMENT 
 
As the transmission of HIV and other blood-borne pathogens can occur through contact 
with contaminated needles, blood, and blood products, appropriate blood and secretion 
precautions will be employed by all personnel in the drawing of blood and shipping and 
handling of all specimens for this study, as currently recommended by the Centers for 
Disease Control and Prevention and the National Institutes of Health. 
 
All dangerous goods materials, including diagnostic specimens and infectious 
substances, must be transported using packaging mandated by CFR 42 Part 72. Please 
refer to instructions detailed in the International Air Transport Association (IATA) 
Dangerous Goods Regulations.  
 
 
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