Conditions: NAFLD NASH - Nonalcoholic Steatohepatitis HIV Infections
Keywords: Fatty liver NAFLD NASH HIV Vitamin E
Allocation: RANDOMIZED | Intervention model: PARALLEL
Enrollment: 3
Healthy volunteers: no

Brief summary:
The purpose of this study is to see how taking Vitamin E daily affects fatty liver in persons living with HIV. Subjects will have both HIV and a fatty liver and the purpose of the study is to learn if underlying liver condition (fatty liver) gets better, worse, or stays the same from taking Vitamin E.

Detailed description:
The investigators will conduct a proof-of-concept clinical trial to evaluate the efficacy of vitamin E for treatment of non-alcoholic steatohepatitis (NASH) in persons living with HIV. Hypothesis: Vitamin E will improve radiographically measured hepatic fat content and circulating markers of liver inflammation and injury in persons living with HIV who have NASH.

A. Perform a pilot randomized placebo controlled trial of vitamin E 800 IU/daily for 6 months in 56 persons living with HIV with biopsy-proven NASH B. Measure change in liver fat content by magnetic resonance proton-density fat fraction (Primary outcome) C. Determine the impact of vitamin E treatment on noninvasive markers of hepatic and systemic inflammation, hepatic fibrosis, and systemic oxidative stress (Secondary outcomes) D. Define baseline hepatic gene expression signatures predictive of response to therapy.

Upon completion, the proposed clinical trial may establish vitamin E as an excellent and inexpensive candidate for further development as a treatment for NASH in persons living with HIV.

Arms:
arm 1: Vitamin E 800 IU/daily for 24 weeks arm 2: Matching placebo for 24 weeks

Interventions: intervention 1: Vitamin E intervention 2: Placebo
Intervention descriptions: intervention 1: Vitamin E 800 IU/daily intervention 2: Matching placebo daily

Protocol document:
Prot_002.pdf: 
Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       1 
 
 
Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Principal Investigator 
  
Samer Gawrieh, MD 
Indiana University School of Medicine 
Department of Medicine 
Division of Gastroenterology/Hepatology 
702 Rotary Circle, Suite 225 
Indianapolis, IN 46202 
 
 
 
 
NCT number:  03669133 
 
Date of Document:  September 20, 2020 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       2 
 
 
 
 
1.0 
Background 
 
The prevalence of nonalcoholic fatty liver disease (NAFLD) is not well defined in 
U.S. adults with HIV infection in the current era. Chronic liver disease is a major 
cause of morbidity and mortality among individuals with HIV infection (1, 2). 
With the availability of effective therapies for hepatitis B and C, NAFLD is 
predicted to become the leading cause of liver disease in the aging HIV 
population (3).  In the non-HIV population, NAFLD is associated with a) 
increased overall and liver-related mortality (4); b) increased risk for 
cardiovascular disease and type 2 diabetes (5-9); and c) increased risk for liver 
cancer even in absence of cirrhosis or significant fibrosis (10-14).  In HIV 
infected patients, the prevalence of NAFLD has not been well established. 
Reported prevalence has varied by study inclusion criteria and modality of NAFLD 
diagnosis, ranging from 15-54% when assessed by imaging modalities (15-17) 
and up to 73% in studies including liver biopsy (18).  These studies suffered 
several limitations, including the inclusion of only a small number of HIV infected 
individuals (18, 19), inclusion of patients with concurrent hepatitis C infection 
(20-23) or limiting the HIV population assessed to only military personnel and 
their dependents or to men alone (15, 16).  
 
The risk factors for NAFLD and NASH in HIV infected individuals have not been 
examined in depth. First, traditional risk factors for NAFLD in the general 
population may not apply to NAFLD in those with HIV infection. While most 
studies suggest an influence of insulin resistance, obesity and dyslipidemia on 
the risk of NAFLD in HIV infected individuals, this may not apply to male HIV 
patients who tend to be more physically fit, have similar or lower BMI and lower 
incidence of fatty liver compared to controls (16, 35, 40). Thus, it is essential to 
comprehensively evaluate NAFLD and NASH in both women and men with HIV 
infection and stratify by the metabolic syndrome and BMI to accurately 
determine prevalence in different risk groups.   
 
The impact of novel antiretroviral agents on risk for NAFLD and NASH has not 
been comprehensively evaluated. Some antiretroviral agents, particularly 
nucleoside or nucleotide reverse transcriptase inhibitors (NRTI) and protease 
inhibitors (PI), impair autophagy and mitochondrial function leading to 
lipodystrophy, insulin resistance, steatosis and steatohepatitis. (38, 41-46). There 
are, however, conflicting data from clinical studies on the contribution of ART to 
this risk and the impact of new antiretroviral agents with better metabolic 
profiles, such as integrase inhibitors, on the risk of NAFLD (15, 18, 20, 35, 47, 
48).  
 
Safe and effective therapies for NASH in HIV infected individuals are absent. 
Over the last decade, there have been several large randomized clinical trials 
(with more than a hundred ongoing Phase 2 and Phase 3 currently registered on 
Clinicaltrials.gov) to treat NASH in the general population, but the majority of 
them systematically excluded HIV infected individuals (74). Weight loss, exercise 
and therapy with vitamin E or pioglitazone have shown efficacy in improving 
NASH and are recommended for HIV uninfected patients with NASH (75). 
However, whether these measures will be effective in HIV infected individuals 
with NASH is unknown. A pilot study showed that in patients with HIV and 


Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       3 
 
abdominal adiposity on stable ART, Tesamorelin, a growth hormone–releasing 
hormone analog, resulted in reduction of visceral and liver fat (76). Whether this 
expensive agent will be useful to treat NASH in HIV infected patients has not 
been established. Vitamin E is an attractive and inexpensive therapy for NASH in 
HIV infected individuals. Oxidative stress contributes to the progression of NASH 
(77-82). Based on its known function as an antioxidant (83), we and others 
have studied vitamin E used alone or with other compounds in multiple trials to 
treat NASH or NAFLD in the non-HIV population (84-93). Variable dosages 
ranging from 100-1200 IU/daily have been used with reported beneficial effects 
on liver enzymes, steatosis, inflammation, ballooning, and hepatic fibrosis. 
Duration of vitamin E monotherapy in these trials ranged from 4 to 96 weeks 
(92, 94). Vitamin E can decrease ALT levels in as early as 4 weeks (92) and 
improve histology in 6 months (87, 88). In the largest randomized trial, PIVENS 
(90), vitamin E was given at a dose of 800 IU/daily for 96 weeks and compared 
to pioglitazone or placebo. Vitamin E and pioglitazone improved ALT, steatosis, 
lobular inflammation, and ballooning and resulted in resolution of NASH in many 
patients. Vitamin E, however, was not associated with weight gain as was 
pioglitazone. Of note, low vitamin E levels were found in 7% of HIV infected men 
in one study (95), and certain ART regimens have been associated with lower 
vitamin E levels (96). Earlier small studies suggested potential reduction in 
oxidative stress and HIV viral loads and increased viability of circulating 
lymphocytes with vitamin E supplementation (97, 98). Thus, there is a strong 
rationale to investigate vitamin E as a therapy for NASH in the HIV infected 
individuals. 
 
Vitamin E for NASH in the non-HIV infected population: The efficacy of vitamin E 
for NASH was best established with the PIVENS study, one of the largest 
randomized trials in NASH to date (90). Indiana University was a major 
participating center in this study. Vitamin E was given at dose of 800 IU/daily for 
96 weeks and compared to pioglitazone or placebo. Vitamin E and pioglitazone 
improved ALT, steatosis, lobular inflammation, and ballooning and resulted in 
resolution of NASH in a significant number of patients (Figure.4). Vitamin E has 
not been tested previously as a therapy for NASH in HIV infected individuals.  
 
 
 
 
 
 
2.0 
Rationale and Specific Aims 
 
The aim of this study is to conduct a proof-of-concept clinical trial to evaluate the 
efficacy of vitamin E for treatment of NASH in HIV infected individuals.   
 
This project is a proof-of-concept, randomized controlled trial to test the 
hypothesis that vitamin E improves hepatic steatosis and markers of hepatic 
necroinflammation in HIV infected individuals with biopsy-proven NASH. In this 
placebo-controlled, double-blind, randomized pilot clinical trial we will randomize 
56 eligible participants at two different sites to receive 800 IU/daily of vitamin E 
or matching placebo for 6 months. The primary endpoint is the change in hepatic 
fat content from baseline to end of treatment in the two groups.  Subjects will 
undergo MR imaging to measure hepatic fat content using MR-PDFF at baseline 
and at the end of treatment. In the randomization visit, all participants will 
receive standard recommendations about lifestyle modifications including healthy 


Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       4 
 
dieting, exercise, and weight loss, and use of alcohol and dietary and herbal 
supplements. 
 
 
Vitamin E is a well-established therapy for NASH in the non-HIV population (see 
section B.5 in Significance). While variable dosages ranging from 100-1200 
IU/daily have been used with reported beneficial effects on liver enzymes and 
histology, including steatosis, the dose chosen in this trial is the same used in 
PIVENS, the largest randomized clinical trial establishing the efficacy of vitamin E 
(90) and is the dose recommended by the multi-society guidelines for treatment 
of NASH in the non-HIV population (75). As vitamin E effects on ALT could be seen 
in as early as 4 weeks (92) and on histology in 6 months (87, 88), we believe the 
6 month duration of the proposed proof-of-principle study is rational. Further, 
there is emerging data to suggest low vitamin E level may be seen is small subset 
(7%) of HIV infected men (95) and in association with use of certain ART (96). 
Benefits reported with vitamin E use in the HIV infected patient in earlier small 
studies include reduction in oxidative stress and HIV viral load and increased 
viability of circulating lymphocytes with (97, 98). Finally, vitamin E is relatively 
inexpensive and has no interactions with any ART drugs. Thus, there is a strong 
rationale to investigate vitamin E as a therapy for NASH in the HIV infected 
individuals. 
 
A total of 56 subjects at two sites will be enrolled in this trial, with approximately 
28 individuals included at IU and 28 at Massachusetts General Hospital.  Subjects 
will be randomly assigned to either Group A or Group B. 
 
Group A:  Vitamin E (RRR alpha-tocopherol) 800 IU/daily for 24 weeks.  
Group B: Matching placebo for 24 weeks. 
 
 
3.0 
Inclusion/Exclusion Criteria 
 
             Inclusion criteria:  
1. males and females ≥21 years with biopsy-proven NASH within 6 months prior 
to enrollment 
2. histological diagnosis of NASH will be confirmed by an experienced liver 
pathologist before study entry 
3. HIV infection  
4. stable dose of anti-diabetic agents and ART in the 3 months preceding 
enrollment and expected by the physician treating diabetes and HIV to remain 
on stable medications during the study 
5. stable dose of tesamorelin in the 6 months preceding enrollment 
6. willingness to participate in the study 
7. ability to understand and give informed consent for participation 
8. willing to stop use of Vitamin E and/or multivitamins containing Vitamin E at 
time of enrollment and for the duration of participation in the study 
 
 
Exclusion Criteria: 
1. Presence of other chronic liver diseases (hepatitis B or C, autoimmune 
hepatitis, cholestatic liver disease, Wilson disease, hemochromatosis, etc.) 
2. average alcohol consumption >3 drinks/day for men or >2 drinks/day for 
women in the 6 months prior to enrollment.  


Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       5 
 
3. Alcohol Use Disorder Identification Test (AUDIT) score 8 or more  
4. ongoing use of medications known to cause hepatic steatosis (e.g., 
corticosteroids, amiodarone, methotrexate, tetracycline, tamoxifen, estrogens 
at doses greater than those used for birth control, anabolic steroids, or 
valproic acid) 
5. prior bariatric surgery 
6. severe co-morbidities (e.g., advanced cardiac, renal, pulmonary, or 
psychiatric illness, as defined as at least one suicide attempt in the past 12 
months) 
7. allergy to vitamin E 
8. use of drugs with potential effect on NASH such as ursodeoxycholic acid, S-
adenosylmethionine (SAM-e), betaine, pentoxifylline, or milk thistle in the 
three months prior to enrollment.  
9. changing doses of statins (simvastatin, pravastatin, atorvastatin, fluvastatin, 
lovastatin, rosuvastatin) or fibrates (clofibrate, fenofibrate) in the three 
months prior enrollment.  
10. illicit substance abuse within the past twelve months, except for marijuana 
11. breast feeding, pregnancy, inability or unwillingness to practice contraception 
for the duration of the study 
12. contraindications for the MRI procedure (e.g., prostheses, severe 
claustrophobia). 
13. poorly controlled diabetes with A1C >9 within in the last six months 
14. use of total parenteral nutrition in the 6 months preceding liver biopsy or 
enrollment 
15. has decompensated cirrhosis, as defined by the presence of ascites, variceal 
bleeding, and/or encephalopathy 
 
 
4.0 
Enrollment/Randomization 
 
Participants found to have NASH will be reviewed for their eligibility to participate in this 
protocol.  Eligible subjects will be contacted about this study and interested individuals 
will enter the screening phase of the protocol.   
 
Screening period: Eligible subjects will be screened within six weeks prior to 
randomization.   The following procedures will take place at the screening visit: 
 
1. Complete explanation of the study, including all study visits and procedures and 
administration of the informed consent statement and HIPAA Authorization form. 
2. An AUDIT questionnaire to determine alcohol consumption will be administered.  
Eligible subjects will be required to obtain a score of less than 8 on the 
questionnaire.   
3. Current social and medical history will be thoroughly reviewed.  
4. Inclusion and exclusion criteria will be verified during this visit.  
5. Female subjects of child-bearing potential will have a urine pregnancy test and 
the pregnancy test must be negative to participate in the study 
6. Information to be collected includes: 
a. age 
b. sex 
c. race 
d. ethnicity 
e. past medical and medicinal history 
f. BMI 


Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       6 
 
g. waist and hip circumference measurements 
h. vital signs, including blood pressure, respiratory rate, heart rate, 
temperature 
i. 
historical laboratory values from medical records, including: 
i. serum electrolytes 
ii. liver profile 
iii. renal parameters 
iv. insulin 
v. glucose 
vi. A1C level  
vii. lipid profile 
viii. HIV viral load 
ix. CD4 count 
  
Randomization-baseline evaluation visit 
 
1. Subjects will return no more than six weeks after the completion of the 
screening visit for a baseline evaluation and randomization into the study.  All 
eligible subjects who have provided informed consent and who have met all 
the inclusion criteria and none of the exclusion criteria will have multiple 
evaluations performed prior to randomization.  This baseline evaluation will 
be conducted in a single study visit.  Subjects will be asked to fast overnight 
and will undergo the following activities: review of medical history 
2. detailed review of medications history 
3. administration of the AUDIT questionnaire to capture alcohol use with a score 
less than 8 in order to participate in the study 
4. collection of vital signs 
a. height 
b. weight   
c. blood pressure 
d. heart rate 
e. respiratory rate 
f. temperature 
5. focused physical examination 
6. measurement of waist and hip circumference 
7. approximately 50mL of blood during a fasting state will be collected for 
       the following tests: 
a. liver profile 
b. complete blood count 
c. lipid 
d. insulin 
e. glucose 
f. INR 
g. MDA 
h. K-18 
i. TNF-alpha 
j. IL-10 
k. alpha and gamma-tocopherols  
8. female subjects of child-bearing potential will have pregnancy test and the 
pregnancy test must be negative to participate in the study 
9. a positive urine pregnancy test will be confirmed with a serum pregnancy test 


Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       7 
 
10. transient elastography (FibroScan®) will be performed by a trained 
technician to measure steatosis by Controlled Attenuation Parameter (CAP) 
and fibrosis by liver stiffness (LSM) 
11. MRI will be performed to measure hepatic fat content by PDFF according to 
the established protocol 
 
Randomization  
At the same visit, after  the baseline evaluation is completed, subjects will be randomly 
assigned to either Group A:  vitamin E in the form of all natural RRR-alpha tocopherol at 
800 IU/d for 6 months (n = 28) or Group B:  matching placebo for 6 months (n = 28). 
We will use a stratified randomization plan based on one factor, diabetes, similar to the 
recent FLINT trial (142). This will ensure two treatment groups having similar 
distribution of diabetes. A stratum specific computer-generated randomization list will be 
prepared by the study statistician to randomly assign study subjects in one of the two 
treatment groups. A block size of four will be used to ensure balance in number of 
subjects recruited in the two groups.   Participants will be given sufficient drug for daily 
dosing and will be given detailed instructions about how to take the medicine. 
 
5.0 
Study Procedures 
 
Subjects will take the study medicine daily.  They will return for a study visit at weeks 4, 
12, 24 and at end of study at week 26.  The study visits for weeks 4 and 12 should last 
about four hours and study visit at week 24 will last about eight hours, and the end of the 
study visit at week 26 will last about two hours.  Total amount of time in the study is 
about 32 weeks, with six weeks allowed between the screening and randomization visits. 
 
The following study procedures will take place at Week 4: 
 
1. study staff will review medical records with the subject 
2. subjects will be asked about any problems they are having 
3. subjects will bring back unused medicine to monitor medication compliance 
4. subjects will be asked about any medications taken, including supplements 
5. subjects will receive sufficient amount of medicine to last until the next study visit 
approximately eight weeks later 
6. AUDIT questionnaire will be administered to detail alcohol use and subjects must 
score less than 8 to continue participating in the study 
7. female subjects of childbearing potential will undergo a urine pregnancy test and 
the test must be negative to continue participating in the study 
8. vital signs will be taken, including height and weight 
9. waist and hip circumference will be measured 
10. subjects will undergo a focused physical examination 
11. transient elastography (FibroScan®) will be performed by a trained technician to 
measure steatosis by Controlled Attenuation Parameter (CAP) and fibrosis by 
liver stiffness (LSM) 
12. historical laboratory values will be captured from the medical records, including 
a. HIV viral load 
b. CD4 count 
 
 
The following study procedures will take place at Week 12:   
 
1. study staff will review medical records with the subject 
2. subjects will be asked about any problems they are having 
3. subjects will bring back unused medicine to monitor medication compliance 


Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       8 
 
4. subjects will be asked about any medications taken, including supplements. 
5. subjects will receive sufficient amount of medicine to last until the next study visit 
approximately twelve weeks later. 
6. AUDIT questionnaire will be administered and subjects must score less than 8 to 
continue participating in the study 
7. female subjects of childbearing potential will undergo a urine pregnancy test and 
the test must be negative to continue participating in the study 
8. vital signs will be taken, including height and weight 
9. waist and hip circumference will be measured 
10. subjects will undergo a focused physical examination 
11. transient elastography (FibroScan®) will be performed by a trained technician to 
measure steatosis by Controlled Attenuation Parameter (CAP) and fibrosis by 
liver stiffness (LSM) 
12. historical laboratory values will be captured from the medical records, including 
a. HIV viral load 
b. CD4 count 
13. approximately 50 mL of blood will be drawn to measure serum alpha- and 
gamma-tocopherols with the remaining amount used for future research. 
 
The following study procedures will take place at week 24 (end of study visit): 
 
1. study staff will review medical records with the subject 
2. subjects will be asked about any problems they are having 
3. subjects will bring back unused medicine to monitor medication compliance 
4. subjects will be asked about any medications taken, including supplements 
5. AUDIT questionnaire will be administered to detail alcohol use  
6. vital signs will be taken, including height and weight 
7. waist and hip circumference will be measured 
8. subjects will undergo a focused physical examination 
9. transient elastography (FibroScan®) will be performed by a trained technician to 
measure steatosis by Controlled Attenuation Parameter (CAP) and fibrosis by 
liver stiffness (LS).  
10. approximately 50mL of blood during a fasting state will be collected for the 
following tests: 
a. liver profile  
b. complete blood count 
c. lipid 
d. insulin 
e. glucose 
f. INR 
g. MDA 
h. K-18 
i. TNF-alpha 
j. IL-10 
k. alpha and gamma-tocopherols  
 
11. An MRI will be performed to measure hepatic fat content by PDFF according to 
the established protocol.   
 
 
Approximately two weeks after the Week 24 end of study visit, subjects will return 
for a follow-up visit.  The following procedures will take place at this visit: 
 
1. The study staff will review medical records with the subject. 


Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       9 
 
2. Subjects will be asked about any problems they are having. 
3. Subjects will be asked about any medications taken, including supplements. 
4. Subjects will undergo a focused physical examination. 
 
 
Upon completion of this visit, subjects will have completed their participation in the 
study. 
 
 
 
 
 
Study visits and data collection schedule 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
6.0 
Reporting of Adverse Events or Unanticipated Problems involving Risks 
to Participants or Others 
 
Data will be captured systematically with regards to new medications, over-the-counter 
medications and supplements, alcohol intake (if any), dietary intake, and changes in 
weight or physical activity. Patients will be instructed not to take any new medication 
unless reviewed with and approved by the study PI (wherever possible). During the trial, 
subjects will be instructed to avoid medications or supplements that may affect the 
primary outcome (as detailed in the inclusion and exclusion criteria). 
 
 
 
Treatment Phase (weeks) 
 
 
Screening 
Randomize 
4 
12 
24 
Follow-
up 
Consent & 
reaffirmation 
X 
X 
 
 
 
 
Baseline medical 
history 
X 
 
 
 
 
 
Interval medical 
history and 
anthropometric 
measurements 
 
X 
X 
X 
X 
 
AUDIT questionnaire 
X 
X 
X 
X 
X 
 
Assessment of 
compliance  
 
 
X 
X 
X 
 
Review of 
concomitant 
medicines 
X 
X 
X 
X 
X 
 
Review of adverse 
events 
 
 
X 
X 
X 
X 
Physical examination 
X 
X 
X 
X 
X 
X 
Dispensing study 
medicines 
 
X 
X 
X 
 
 
Blood sample 
collection 
 
X 
 
X 
X 
 
Urine pregnancy test 
X 
X 
X 
X 
X 
 
Liver stiffness and 
CAP by TE 
 
X 
X 
X 
X 
 
MR-PDFF 
 
X 
 
 
X 
 
Biobanking 
 
X 
X 
X 
X 
 
 
 


Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       10 
 
Risks of blood draw include pain and bruising at the site.  Blood will be drawn by 
experienced personnel. 
 
Risks of completing the questionnaires include embarrassment or being uncomfortable 
with the questions asked.  Subjects can refuse to answer any questions to which they do 
not wish to respond. 
 
Risks of transient elastography include being uncomfortable lying supine pain, soreness 
at the site and a small chance of allergic reaction to the ultrasound gel. These study 
procedures will be performed by experienced personnel. 
 
Risks of MRI include being uncomfortable lying still for a period of time and experiencing 
claustrophobia (feeling closed in or trapped). 
 
Risks of taking Vitamin E include allergic skin reactions (inflammation or itching), blurred 
vision, changes in cholesterol levels, changes in insulin resistance, diarrhea, dizziness, 
fatigue, flu-like symptoms, headache, heart conditions, increased risk of death, 
increased risk of fainting or falls, increased risk of heart failure, increased risk of high 
blood pressure in pregnancy, increased risk of stroke, increased risk of tuberculosis, 
kidney dysfunction, nausea, severe response to infection (in preterm babies), sexual 
dysfunction, stomach pain, vision loss, and weakness. 
 
There is a risk of loss of confidentiality. 
 
A Data Safety and Monitoring Board (DSMB) will be organized and instituted to review 
study and safety data as well as reporting adverse events/unanticipated problems and 
will meet as necessary for a thorough review of this study. The DSMB will consist of a 
hepatologist, an infectious disease specialist, and a biostatistician.   
 
 
7.0 
Study Withdrawal/Discontinuation 
 
It is anticipated that this study will be completed within the time frame proposed and 
that it will establish that vitamin E is beneficial in HIV infected patients with NASH. If 
vitamin E is found to exert beneficial effects on hepatic fat content and markers of 
NASH, it will justify undertaking larger multicenter randomized trials with histological 
end points to evaluate the efficacy and safety of vitamin E in this population.  
 
Inability to meet target enrollment in a timely fashion is always a concern for any clinical 
trial.  However, the sample size is realistic and enrollment is spread across several 
funding years and at two leading liver centers. People with cirrhosis have been excluded 
from this trial, as hepatic fat usually diminishes or disappears with progression to 
cirrhosis and will likely be difficult to measure at baseline in these patients. To minimize 
observer variability, MR-PDFF images will be centrally read by the study radiologist. 
Although concerns have emerged about increased risk for prostate cancer with vitamin E 
use in healthy men (144), this should be less of concern in HIV infected men who are at 
lower risk for prostate cancer than non-HIV infected men, possibly due to hypogonadism 
(145). Assuming 10% drop out rate with the clinical trial, about ten percent more 
subjects than needed to get the statistical power to evaluate the primary outcome will 
be recruited. 
 


Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       11 
 
 
8.0 
Statistical Considerations 
 
Because host genetic factors contribute to the response to therapy, pre-therapy baseline 
liver expression profiles will be studied to identify expression signature that predict 
response to vitamin E therapy. This approach has been used in other liver disease such 
as hepatitis C. For example, in one study, pre-treatment peripheral blood monocytes 
gene expression signature predicted response to hepatitis C therapy with 94% accuracy 
(143).  
 
Quantitative global gene expression analysis on fresh frozen liver tissue from subjects 
participating in the clinical trial will be performed.  Based on prior experience, it is 
expected to have a portion of the liver biopsy snap frozen and available for expression 
analysis on a sizable proportion of the subjects (>50%) undergoing liver biopsies as part 
of Specific Aim 2 who are the primary source for the proposed clinical trial. As such, 
HumanHT-12 v4 Expression BeadChip will be used for this aim. This chip provides 
genome-wide transcriptional coverage of well-characterized genes and their splice 
variants, and targets more than 47,000 probes derived from the NCBI RefSeq Release 
38 (November 7, 2009) and other sources.  
 
After detectable transcripts are identified, expression data will be standardized. The t- 
test statistic will be utilized to measure the expression difference between the sample 
means (responders vs. non-responders) in units of the standard deviation for which the 
difference can be tested using certain p value. For this, a significance cut-point of 
p<0.001 will be used. Then hierarchical clustering and gene network and pathway 
analysis will be performed to identify those predicting response to vitamin E therapy. A 
logistic multivariate-based analysis will be used to build up a receiver operating 
characteristic (ROC) curve, and the area under the ROC curve (AUC) will be calculated 
to assess the average sensitivity of the biomarker in discriminating the responders and 
non-responders.  
 
All endpoints are measured as the change from baseline to 24 weeks. The primary 
outcome measure is the change in hepatic fat content as measured by central reading of 
MR-PDFF performed at baseline and end of study. Secondary endpoints will be changes 
in inflammatory markers (liver enzymes, TNF-alpha, IL-10 levels), serum K-18, Liver 
stiffness and CAP measurement by TE, and MDA level as a marker of oxidative stress. 
 
The primary outcome is the improvement in liver fat as assessed by MR-PDFF at end of 
treatment compared to baseline. To our knowledge, there are no data on studying the 
effect of vitamin E on MR-PDFF in NASH patients with HIV. Using an alpha of 5% and a 
two-sample independent t-test and equal variance assumption, a sample size of 25 per 
group will have 91% power to detect an effect size of 0.97 and will have 80% power to 
detect an effect size as small as 0.81. To ensure this number is achieved, 28 subjects per 
group will be recruited, allowing for 10% drop-out during the trial. The primary purpose 
of this pilot study is to generate preliminary data to estimate the mean and variability of 
improvement in liver fat with vitamin E in HIV infected patients with NASH.  
 
Primary outcomes analysis: The primary analysis will be based on an analysis of covariance 
(ANCOVA) to compare the improvement in liver fat due to vitamin E relative to placebo. 
The model will include one dummy variable for treatment group indicator as well as 
baseline demographic characteristics and baseline liver fat.  The 95% confidence intervals 
for the difference between the two treatment groups will be reported.   
 


Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Protocol date:  September 2, 2020 
IRB study number 1807274844                                                       12 
 
Secondary outcomes analysis: Secondary efficacy end points are continuous and as such 
the ANCOVA model will be used as described for analyzing the primary outcome except 
that the model will include the baseline value of the response variable. 
 
 
9.0 
Privacy/Confidentiality Issues 
 
Data will be entered by authorized study personnel only into RedCap, which is a 
password protected and access restricted research data entry system.   
 
All study visits will take place in a private room.  Informed consent will be obtained only 
after the study is thoroughly explained to the prospective subject and all his questions 
have been answered. 
 
10.0 Follow-up and Record Retention 
 
The study will last approximately three years.  Data will be stored in RedCap in 
accordance with NIH and Indiana University Guidelines or until such time as the records 
are no longer needed. 

SAP_003.pdf: 
 
 
 
 
 
 
Vitamin E for NASH Treatment in HIV Infected Individuals 
 
Principal Investigator 
  
Samer Gawrieh, MD 
Indiana University School of Medicine 
Department of Medicine 
Division of Gastroenterology/Hepatology 
702 Rotary Circle, Suite 225 
Indianapolis, IN 46202 
 
 
 
Statistical Plan 
 
NCT number:  03669133 
 
Date of Document:  September 20, 2020 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


Statistical Considerations 
 
Because host genetic factors contribute to the response to therapy, pre-therapy baseline liver 
expression profiles will be studied to identify expression signature that predict response to 
vitamin E therapy. This approach has been used in other liver disease such as hepatitis C. For 
example, in one study, pre-treatment peripheral blood monocytes gene expression signature 
predicted response to hepatitis C therapy with 94% accuracy (143).  
 
Quantitative global gene expression analysis on fresh frozen liver tissue from subjects 
participating in the clinical trial will be performed.  Based on prior experience, it is expected to 
have a portion of the liver biopsy snap frozen and available for expression analysis on a sizable 
proportion of the subjects (>50%) undergoing liver biopsies as part of Specific Aim 2 who are 
the primary source for the proposed clinical trial. As such, HumanHT-12 v4 Expression BeadChip 
will be used for this aim. This chip provides genome-wide transcriptional coverage of well-
characterized genes and their splice variants, and targets more than 47,000 probes derived 
from the NCBI RefSeq Release 38 (November 7, 2009) and other sources.  
 
After detectable transcripts are identified, expression data will be standardized. The t- test 
statistic will be utilized to measure the expression difference between the sample means 
(responders vs. non-responders) in units of the standard deviation for which the difference can 
be tested using certain p value. For this, a significance cut-point of p<0.001 will be used. Then 
hierarchical clustering and gene network and pathway analysis will be performed to identify 
those predicting response to vitamin E therapy. A logistic multivariate-based analysis will be 
used to build up a receiver operating characteristic (ROC) curve, and the area under the ROC 
curve (AUC) will be calculated to assess the average sensitivity of the biomarker in 
discriminating the responders and non-responders.  
 
All endpoints are measured as the change from baseline to 24 weeks. The primary outcome 
measure is the change in hepatic fat content as measured by central reading of MR-PDFF 
performed at baseline and end of study. Secondary endpoints will be changes in inflammatory 
markers (liver enzymes, TNF-alpha, IL-10 levels), serum K-18, Liver stiffness and CAP 
measurement by TE, and MDA level as a marker of oxidative stress. 
 
The primary outcome is the improvement in liver fat as assessed by MR-PDFF at end of treatment 
compared to baseline. To our knowledge, there are no data on studying the effect of vitamin E 
on MR-PDFF in NASH patients with HIV. Using an alpha of 5% and a two-sample independent t-
test and equal variance assumption, a sample size of 25 per group will have 91% power to detect 
an effect size of 0.97 and will have 80% power to detect an effect size as small as 0.81. To ensure 
this number is achieved, 28 subjects per group will be recruited, allowing for 10% drop-out during 
the trial. The primary purpose of this pilot study is to generate preliminary data to estimate the 
mean and variability of improvement in liver fat with vitamin E in HIV infected patients with NASH.  
 
Primary outcomes analysis: The primary analysis will be based on an analysis of covariance 
(ANCOVA) to compare the improvement in liver fat due to vitamin E relative to placebo. The 
model will include one dummy variable for treatment group indicator as well as baseline 
demographic characteristics and baseline liver fat.  The 95% confidence intervals for the 
difference between the two treatment groups will be reported.   
 
Secondary outcomes analysis: Secondary efficacy end points are continuous and as such the 
ANCOVA model will be used as described for analyzing the primary outcome except that the 
model will include the baseline value of the response variable.