Analysis of Cellular Kinases and Aging in PBMCs and Colorectal Tissue
Conditions: HIV Infections, Aging
Sponsor: Johns Hopkins University

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Analysis of Cellular Kinases and Aging in PBMCs and Colorectal Tissue 
 
 
 
 
 
Principal Investigators: 
 Heng Zhu, PhD 
 
 
 
 
 
Version 3.0 
21 January 2025 
 
 
 
Clinical Trials.gov ID: NCT6132386

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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Table of Contents 
 
List of Abbreviations .............................................................................................................3 
I. PROJECT OVERVIEW.........................................................................................................4 
II. INTRODUCTION ..................................................................................................................4 
III. RATIONALE ........................................................................................................................5 
Study Objectives:.....................................................................................................................5 
IV. STUDY DESIGN .................................................................................................................6 
Overview .................................................................................................................................6 
Study Population .....................................................................................................................6 
Inclusion Criteria ...............................................................................................................6 
Exclusion Criteria ..............................................................................................................7 
Study Procedures ....................................................................................................................8 
Visit 1: Screening Visit......................................................................................................8 
Visit 2: Study Visit (Screening Visit ≤ -42 days)...........................................................9 
V. RECRUITMENT AND OUTREACH .................................................................................10 
VI. COMPENSATION TO SUBJECTS .................................................................................11 
VII. COSTS TO SUBJECTS ..................................................................................................11 
VIII. PROTECTION OF HUMAN RESEARCH PARTICIPANTS .......................................11 
Risks ................................................................................................................................11 
Benefits ............................................................................................................................12 
IX. STATISTICAL ANALYSIS ...............................................................................................12 
X. CONFIDENTIALITY...........................................................................................................12 
XI. REFERENCES .................................................................................................................13 
APPENDIX I: SCHEDULE OF EVENTS ...........................................................................................15 
 
 
 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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List of Abbreviations 
 
ALT 
 
Alanine transaminase 
AK2 
 
Adenylate kinase 2 
AST  
 
Aspartate aminotransferase 
BUN 
 
Blood urea nitrogen 
CBC 
 
Complete Blood count 
CKM 
 
Creatine kinase 
EMR 
 
Electronic medical record 
FTA-ABS 
Fluorescent treponemal antibody test absorption 
FTC 
 
Emtricitabine 
GC/CT  
Gonorrhea/Chlamydia 
HIV 
 
Human immunodeficiency virus 
INR 
 
International normalized ratio 
NRTIs  
Nucleotide reverse transcriptase inhibitors 
PBMC  
Peripheral blood mononuclear cells 
PLWH  
People living with HIV 
PrEP 
 
Pre-exposure prophylaxis 
PTID 
 
Participant ID 
RPR 
 
Rapid plasma reagin 
STI 
 
Sexually transmitted infection 
TDF 
 
Tenofovir disoproxil fumerate 
TFV 
 
Tenofovir 
TFV-DP 
Tenofovir diphosphate 
TFV-MP 
Tenofovir monophosphate 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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I. PROJECT OVERVIEW 
 
This project involves obtaining peripheral blood mononuclear cells (PBMCs) and colorectal 
tissue samples from study participants to measure adenylate kinase 2 (AK2) and muscle-type 
creatine kinase (CKM) enzyme levels in an “older adult” group (65-80) when compared to a 
“younger adult” group (18-50). Both AK2 and CKM has been demonstrated to be vital enzymes 
in converting the prodrug tenofovir (TFV) into its active form, tenofovir diphosphate (TFV-DP).1-4 
However, no study has yet investigated the effect of aging on AK2 or CKM in colorectal tissues 
relevant to HIV infection and prevention. This study will investigate the AK2 and CKM variability 
in individuals from the younger and older age groups in preparation for a follow-up study 
investigating how the pharmacokinetics (PK) of tenofovir (TFV) vary with altering levels of 
cellular enzymes in different age populations. A finding of clinically significant differences in the 
activity of  AK2 or CKM, may indicate the need for dose adjustment of TFV for HIV treatment or 
prevention depending upon age to either reduce toxicity or enhance efficacy. 
 
In this study, individuals who are interested in participating will be screened for eligibility. If 
eligible, participants will return for a single  study visit. At the study visit, blood will be drawn and 
colorectal tissue will be obtained via flexible sigmoidoscopy. Participants can participate in an 
optional PK sub-study if they are already taking a tenofovir (TFV)-based regimen for prevention 
or are willing to take a daily TFV-based regimen for 7 days prior to the flexible sigmoidoscopy. 
Subsequently, AK2 and CKM will be analyzed in PBMCs and colorectal tissue using mass 
spectrometry.  
 
II. INTRODUCTION 
 
The prevalence of HIV in older adults (≥50 years of age) living with HIV has increased, in part to 
the success of antiretroviral therapy extending life expectancy, with nearly half of the people 
living with HIV (PLWH) in the United States over the age of 50, and 16% ≥65 years of age (data 
from the Centers for Disease Control and Prevention). However, each year, there are nearly 
40,000 new HIV diagnosis infections in the United States and 1.7 million infections globally.5,6  
Individuals over the age of 50 account for 17% of new HIV diagnoses annually. In 2020, men 
accounted for 80% of new HIV diagnosis in the United States, with nearly 70% in gay, bisexual, 
or other men who reported male-to-male sexual contact. With this in mind, there is a need to 
develop and optimize efficacious strategies for prevention of HIV infection in this population5.  
 
The recent development of pre-exposure prophylaxis (PrEP) for the prevention of HIV can 
significantly decrease the risk of HIV infection with proper adherence.7 Tenofovir is a prodrug 
that is the main antiviral component of current PrEP regimens.8-10 TFV is from the drug class 
known as nucleotide reverse transcriptase inhibitors (NRTIs) in which chain termination occurs 
during DNA synthesis by HIV reverse transcriptase, therefore, suppressing HIV replication. 
However, only the diphosphate form of tenofovir (TFV-DP), generated by phosphorylation of 
TFV by intracellular kinases, is capable of inhibiting HIV replication.  
 
In key tissues related to HIV acquisition, it has been found that adenylate kinases and creatine 
kinases play a significant role in TFV activation.1 Specifically, in colorectal tissue, it has been 
found that adenylate kinase 2 (AK2) is the primary kinase in the initial phosphorylation of TFV to 
tenofovir monophosphate (TFV-MP). Moreover, muscle-type creatine kinase (CKM) has been 
demonstrated to be the enzyme that is vital for the second phosphorylation, transforming TFV-
MP to TFV-DP.  
 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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Previous studies have found that age has an effect on CKM expression, post-translational 
modifications, and activity in multiple tissues.11-16 Thus, it is hypothesized that expression levels 
and post translational modifications of AK2 and CKM may differ depending upon age, therefore, 
altering the amount of active TFV-DP and increasing variability of protection. Therefore, it is 
imperative to investigate expression and post translational modifications of AK2 and CKM in 
colorectal tissue and PBMCs from various age populations to see if theses cellular enzymes 
change with increasing age.   
 
III. RATIONALE 
 
While PrEP has been shown to be highly effective in reducing the risk of HIV infection, despite 
adherence to PrEP, some individuals become infected with HIV. To gain an understanding of 
CKM phosphorylation activity and examine whether CKM might also contribute to variability of 
TFV-DP activation in colonic CD4+ cells in the presence of HIV infection, we will also measure 
levels of CKM and TFV-DP in colonic CD4+ T cells of younger and older individuals infected 
with HIV taking tenofovir-based HIV treatment, as well as those taking tenofovir-based PrEP 
who are not infected with HIV. 
 
This study is designed to investigate the intracellular AK2 and CKM enzymes in both colorectal 
tissue and PBMCs in a younger and older adult population. It is hypothesized that age alters 
expression and post translational modifications and expression of AK2 and CKM in colorectal 
tissue and PBMCs causing variability in TFV-DP levels in individuals taking tenofovir-based 
medications.  
 
To test this hypothesis, colorectal biopsies and PBMCs will be collected to assess for changes 
in AK2 and CKM levels using mass spectrometry and changes in enzymatic activity to 
phosphorylate TFV from the following populations: 
Cohort A: 10 younger (age 18-50) and 10 older (age 65-80) adults who are not infected 
with HIV and not taking tenofovir-based PrEP (“Control Cohort”). 
 
Cohort B: 10 younger (age 18-50) and 10 older (age 65-80) adults who are not infected 
with HIV, but are taking a TFV-based regimen (either on daily TFV- based PrEP or agree 
to take a daily TFV-based regimen 7 days prior to procedure). (“PrEP Cohort”) 
 
Cohort C: 10 younger (age 18-50) and 10 older (age 65-80) adults who are infected with 
HIV, taking a TFV-based regimen. (“ARV Cohort”) 
 
Study Objectives: 
Objective 
Endpoint 
1. Determine if levels and activity of AK2 
and CKM in colorectal tissue differ 
between  
(a) younger adults (age range18-50 
years), and older adults (age range 65-
80) or  
• 
Proteomics will be used to measure 
intracellular protein levels and modification 
status of kinases that phosphorylate TFV.  
 
• 
In individuals taking a TFV-containing 
regimen, colorectal tissue TFV-DP will be 
measured at steady-state. 
 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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(b) between HIV negative and HIV 
positive persons. 
• 
In individuals not taking a TFV-based 
regimen, in vitro incubation of TFV with 
colorectal tissue biopsies will be performed 
to measure TFV activation as a function of 
age.  
2. Determine if levels and activity of AK2 
and CKM in PBMCs differ between: 
(a) younger adults, defined as 18-50 
years of age, and older adults, defined 
as 65-80 years of age or  
(b) HIV negative and HIV positive 
persons.. 
• 
Proteomics will be used to measure 
intracellular protein levels and modification 
status of kinases that phosphorylate TFV. 
 
• 
In individuals not taking a TFV-based 
regimen, in vitro incubations of TFV with 
PBMCs will be performed to measure TFV 
activation as a function of age. 
 
• 
In individuals taking a TFV-containing 
regimen, colorectal tissue TFV-DP will be 
measured at steady-state. 
 
IV. STUDY DESIGN 
Overview 
The design of the study will be a two-visit study, including a screening visit (Visit 1) and a 
sampling visit (Visit 2). During the screening visit, the individual will be assessed for eligibility. 
For participants scheduled for sigmoidoscopy or colonoscopy for a medical indication, the 
screening visit may be conducted by phone prior to the procedure. If eligible, the participant will 
return for the study visit < 42 days following the screening visit. During the study visit, 
approximately 100 mL of blood will be drawn for PBMC isolation. Following the blood draw, 
participants will undergo endoscopic evaluation in which 10 colorectal biopsy samples will be 
obtained between 10-20 cm from the anal verge. Thirty adults from each age population 
(younger and older) will be enrolled for this study. Three cohorts (each including young and old 
groups) will be defined by TFV administration and HIV status. 
 
Study Population 
Three cohorts of participants will be recruited, with 10 younger and 10 older in each cohort.  
Cohort A: Healthy Volunteers, not taking TFV based PrEP (Control Cohort) 
Cohort B: Healthy volunteers, steady state TFV-based regimen (either on daily TFV- based 
PrEP or taking daily TFV-based regimen 7 days prior to procedure) (PrEP Cohort) 
Cohort C: Persons infected with HIV taking TFV/FTC-based HIV treatment (ARV Cohort) 
 
Inclusion Criteria 
Participants who meet the following criteria are eligible for inclusion in this research: 
 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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1. English speaking male or female volunteers between the ages of 18-50 for younger adults 
or 65-80 for older adults 
2. Willing to provide written informed consent 
3. Willing to abstain from insertion of anything in rectum for 72 hours before and 72 hours after 
the endoscopic procedure for colorectal tissue collection. 
4. Not currently participating in other research studies involving drugs and/or medical devices. 
 
For participants not infected with HIV (Control Cohort A) 
1. No known risk for HIV exposure or a documented negative HIV-1/HIV-2 Ag/Ab test in the 
past 3 months (HIV risk, but not on PrEP) 
2. Documented negative HBsAg in those taking study TFV (i.e. cohort B) 
 
For participants not infected with HIV, but taking or willing to take TFV (PrEP Cohort B) 
1. Currently taking TFV-based oral PrEP daily or willing to take oral TFV-containing PrEP 
for one week 
 
2. Documented negative HIV-1/HIV-2 Ag/Ab test in the past 3 months 
3. Documented negative HBsAg 
 
For participants infected with HIV (ARV Cohort C) 
1. Virologically suppressed HIV for at least 6 months prior to screening. 
2. Taking tenofovir disoproxil fumarate or tenofovir alafenamide containing regimen to treat 
HIV 
 
Exclusion Criteria 
Participants who meet any of the following criteria at screening will be excluded from the study:   
 
1. History of inflammatory bowel disease or active inflammatory condition of the GI tract 
2. History of significant gastrointestinal bleeding 
3. Current medically-indicated use of warfarin or heparin or other anticoagulant medications 
associated with increased risk for bleeding following mucosal biopsy (e.g., daily high dose 
aspirin [>81 mg], non-steroidal anti-inflammatory drugs [NSAIDs], or Pradaxa®)  
4. Use of systemic immunomodulatory medications within 4 weeks of enrollment 
5. Use of rectally administered medications within 4 weeks of enrollment 
6. Use of product containing nonoxynol-9 within 4 weeks of enrollment 
7. Use of any investigational products within 4 weeks of enrollment 
8. Any other clinical condition or prior therapy that, in the opinion of the investigator, would 
make the participant unsuitable for the study or unable to comply with the study 
requirements. Such conditions may include, but are not limited to, current or recent history of 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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severe, progressive, or uncontrolled renal, hepatic, hematological, gastrointestinal, 
endocrine, pulmonary, neurological, or cerebral disease. 
9. Active rectal infection (GC, Chlamydia, HSV). Participants screening positive for GC/CT at 
the time of endoscopy will be excluded from analysis (and replaced). 
For participants not undergoing a concurrent endoscopic procedure for indication 
unrelated to this study: 
10. Hct <36% 
11. Platelet count <150/mm3 
12. INR > 1.2 
 
Study Procedures 
 
Note: This protocol requires colorectal biopsies to support the primary objectives. These are 
collected using an endoscope (e.g., flexible sigmoidoscope or colonoscope). We plan to use a 
flexible sigmoidoscope as the least invasive method to achieve our objectives. If, however, the 
research participant already has a colonoscopy scheduled for a medically indicated purpose, 
e.g., a preventive colon cancer screening, then we will coordinate with the clinical endoscopy 
team to obtain the biopsies by this method, in order to reduce the number of procedures related 
to our study. The protocol indicates some minor differences in procedures to accommodate this 
option.   
 
This section includes information on visit specific study procedures. This study includes two 
visits, a screening visit and a sampling visit. The breakdown of the procedures on each day are 
as follows: 
 
Visit 1: Screening Visit 
Prescreening via email or a phone call will take place with potential participants. Those 
interested in the study can request that the informed consent documents be emailed or mailed 
to them prior to the Screening Visit. 
 
Written informed consent will be obtained from study participants prior to any study procedures 
taking place. To obtain informed consent, an interactive question and answer process will occur 
in which the potential subject will be asked specific questions to determine their understanding 
of the purpose, procedures, risks and benefits of participating in the study. If, in the judgment of 
the investigator, the subject cannot demonstrate sufficient understanding, then that subject will 
be excluded from participation in the protocol. If participants do not meet the eligibility criteria, 
screening will be discontinued once ineligibility is determined.  
 
The following list of activities will be completed during the screening visit: 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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Table 1: Visit 1 - Screening  
Component 
Procedure/Analysis 
Administrative 
 
• 
Obtain written informed consent 
• 
Assign participant ID (PTID) 
• 
Assess eligibility 
• 
Obtain demographic and locator information 
• 
Provide counseling: 
• 
HIV/STI risk reduction counseling 
• 
HIV pre- and post- test counseling, if indicated 
• 
Provide a copy of pre- and post- visit reminders 
• Participants not undergoing screening colonoscopy will be provided 
with a Enema and directions for use prior to Sampling Visit.  
Clinical 
• 
Obtain complete medical history, documenting pre-existing 
conditions  
• 
Review and record concomitant medications 
• 
Obtain vital signs – blood pressure (BP), heart rate (HR), height, and 
weight (height may be obtained from EMR, and weight captured in 
the EMR within 6 months of screening may be obtained from EMR 
 
Note: Medical history and physical examination data may be extracted 
from electronic medical record, or performed at the time the 
participant presents for an endoscopic procedure scheduled for 
clinical indication. 
Safety Labs and 
STI Screening (for 
participants not 
undergoing a 
concurrent 
endoscopy 
procedure only):  
• 
Perform general physical exam   
• 
Collect rectal swab for GC/CT 
• 
Collect blood specimens for: 
o Complete blood count (CBC) with differential1 
o AST, ALT, BUN, Creatinine1 
o HIV testing2 
o Syphilis testing3 
PrEP cohort only 
for those not 
already on TFV-
based PrEP prior 
to screening 
• 
HBsAg (result from prior test within 12 months acceptable) 
• 
Dispense F/TDF or F/TAF for daily dosing to begin 7 days prior to 
study procedure (PrEP Cohort B only, not already on PrEP) 
• 
Commence daily virtual DOT for 7 days prior to study procedure 
 
¹ Screening laboratory values are valid up to 42 days.   
2See Appendix II: HIV Testing Algorithm; HIV testing not to be done in participants known to be infected 
with HIV  
3Syphilis Chemiluminescence assay (with confirmatory RPR; FTA-ABS performed on RPR negative 
samples) 
 
 
Visit 2: Sampling Visit (≤ 42 days since Screening Visit) 
 
Note: Where applicable, Visit 2 may be combined with Visit 1 
 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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After eligibility is confirmed, participants will be contacted to schedule the study visit (Visit 2). 
Where possible, Study Visit 2 may be coordinated with a scheduled clinical procedure, such as 
a screening colonoscopy. 
 
The following activities will take place at the Study Visit:   
 
Table 2: Visit 2 – Sampling Visit 
 
Component 
Procedure/Analysis 
Administrative 
 
• 
Confirm participant understood and signed informed consent 
• 
Review/update locator information 
• 
Confirm eligibility 
• 
Provide counseling: 
• 
HIV/STI risk reduction counseling 
• 
Provide a copy of pre- and post- visit reminders 
Clinical 
• 
Review/update medical history 
• 
Review/update concomitant medications 
• 
Perform symptom-directed physical exam as needed 
• 
Obtain vital signs – blood pressure (BP), heart rate (HR),  
• 
Obtain 10 colorectal biopsies via flexible sigmoidoscopy or during 
clinically-indicated study procedure 
STI Screening 
(For participants 
undergoing a 
concurrent 
endoscopy for 
clinical 
indications):  
 
• 
Collect rectal swab for CT/GC testing 
Blood for PBMC 
Collection 
• 
Pre-endoscopic Procedure 
TFV PK (only in 
HIV uninfected 
participants 
taking TFV) 
• 
Blood plasma 
 
V. RECRUITMENT AND OUTREACH 
 
Sources of recruitment will be as follows: 
• 
Referral from treating clinicians at any regional clinic   
• 
Prior Hopkins/Affiliates study participants who have given informed consent to be 
reached for future studies 
• 
Individuals who learn about the study through advertisements or peer/network 
recruiting 
Approved recruitment flyers will be distributed throughout the community. In addition, approved 
messages for social media platforms, internet recruitment and department website will be used. 
Staff will also follow-up with all persons who express an interest in the study in a timely manner. 
 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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VI. COMPENSATION TO SUBJECTS 
 
Participants will be compensated for their time and inconvenience related to participation in this 
study. Compensation will be $425 for completion of the study and is based upon the schedule 
established for DDU studies. 
 
VII. COSTS TO SUBJECTS  
 
There will be no costs to the subjects outside of costs to travel to the study site. 
 
 
VIII. PROTECTION OF HUMAN RESEARCH PARTICIPANTS 
 
Risks  
 
Flexible Sigmoidoscopy: Endoscopy is uncomfortable for some persons and poses only a 
very small risk of perforation and bleeding when biopsies are taken. The most common adverse 
events associated with endoscopy are gas, bloating, and cramping which usually resolve within 
hours of the procedure. Taking the biopsies causes no discomfort due to the insensate epithelial 
lining of the colon. Pinch biopsies may cause minor bleeding which stops on its own. More 
significant and rare adverse events can include prolonged bleeding and/or perforation. 
Participants will be screened and excluded if taking any blood thinners or have conditions that 
may prolong bleeding. Risk of perforation, which is a medical emergency that may require 
surgery, will be minimized by enrolling participants who do not have significant bowel disease 
and having the procedure performed by a credentialed endoscopist. Where possible, the 
endoscopic procedure will be paired with a procedure that may have a clinical indication, such 
as screening colonoscopy for colon cancer. In this case, the only additional risk to the 
participant are the risks of collecting pinch biopsies, which are lower in risk than the endoscopy 
itself.. 
 
Phlebotomy is performed by drawing blood from a vein in the forearm or antecubital fossa. This 
may cause pain during the insertion of the needle or slight bruising afterward. 
 
TFV (administered as part of this research protocol): TFV disoproxil fumarate (TDF) and 
TFV alafenamide (TAF) are both TFV prodrugs licensed by FDA for the prevention and 
treatment of HIV. TFV-based PrEP regimens currently approved include emtricitabine / tenofovir 
alafenamide (F/TAF) or emtricitabine / tenofovir disoproxil fumarate (F/TDF). Both combination 
regimens are well tolerated. Risks of TFV include acute exacerbation of hepatitis B virus 
Immune Reconstitution Syndrome; New or worsening renal impairment; lactic acidosis; 
hepatomegaly with steatosis; lower bone mineral density; diarrhea; nausea; headache; fatigue; 
abdominal pain; rash. Risks will be minimized by confirming participants do not have hepatitis B 
or HIV and baseline safety labs will be monitored prior to receiving study medication. TFV-naïve 
participants who are not already prescribed TFV-containing regimens for PrEP or treatment will 
take only 7 doses, which will minimize the risks associated with chronic administration. 
 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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STI testing: Participants may become embarrassed, worried, or anxious when completing their 
HIV-related interviews and/or receiving STI counseling. Certain positive results (Chlamydia, 
gonorrhea) may need to be reported to appropriate Department of Public Health Agencies. The 
reporting only happens if the test result is positive and is always done confidentially.  
 
Incidental Findings:  Significant clinical findings may be discovered during study procedures, 
which may warrant referral for medical care and further testing. If significant clinical findings are 
discovered during the procedure, study subjects will be referred to the proper Johns Hopkins 
Division for additional evaluation and treatment as necessary. 
 
Social Risk: Although the study site will make every effort to protect participant’s privacy and 
confidentiality, it is possible that involvement in the study could become known to others, and 
that social harms may result (i.e., subjects could become known as HIV-infected or at "high risk" 
for HIV infection) and, consequently, be treated unfairly or discriminated against by families, 
friends and/or communities. 
 
Loss of confidentiality: Although every effort will be made to protect the privacy and 
confidentiality of all participants, it is possible that their involvement in the study could become 
known to others. All guidelines for the protection of Personal Health Identifiers will be followed.  
 
Benefits 
 
There are no direct benefits to subjects who participate in this study. However, there may be 
personal satisfaction in the knowledge that they are aiding in important medical research, and 
that the knowledge obtained from this study will advance the field of HIV prevention. 
Participants will be reimbursed for their time and inconvenience. 
 
The risks to individual participants are outweighed by potential benefits to the scientific 
community and to society – namely, contribution to our microbicide knowledge-bank.   
 
IX. STATISTICAL ANALYSIS 
 
To compare protein levels between the younger adult and older adult groups, protein 
abundances will be averaged within age groups. Differences in protein levels will be determined 
by unpaired two-tailed t test using the Benjamini-Hochberg correction for multiple comparisons, 
with significant differences being defined as having an adjusted p-value of <0.05. Based on our 
previous studies of TFV phosphorylation we estimate ~80% power to detect a 1.5-fold 
difference between study groups using our measured outcomes and >95% power to 
detect a 1.75-fold difference between study groups at our sample size. 
 
X. CONFIDENTIALITY  
 
Given the sensitive nature of the proposed studies, the investigational team will keep the study 
information private to the extent possible by law. Where possible, clinical information will be 
identified by a unique number assigned to each individual research participant. The research 
team will make every effort to register research subjects in the hospital information systems for 
clinical testing using this identifier. Access to study records will be limited to the study team, the 
Office of Human Research Protections, and Johns Hopkins University officials, where applicable 
and required by law. Enrolled patients will be assigned study numbers for data collection and 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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evaluation. Publications in medical journals arising from this study will delete the names of the 
subjects involved.  
 
The key linking study to numbers will be kept as an electronic file. Electronic files will be 
maintained on the DDU’s computer server, which is password protected for access solely by 
study investigators at the level of the individual protocol. 
 
This study will not require a Certificate of Confidentiality. However, a Certificate of 
Confidentiality has been issued as a term and condition of this NIH-Funded research. All 
records are stored in a locked area and only research staff will have access to them. Paper files 
are maintained in a locked cabinet located in the Division of Clinical Pharmacology, again, with 
access limited to study investigators. 
 
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IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
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is a negative regulation on muscle creatine kinase. J Biol Chem 2007;282:12022–9. 
https://doi.org/10.1074/jbc.M610363200.  
16. Pierce A, DeWaal E, VanRemmen H, Richardson A, Chaudhuri A. A novel approach for 
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https://doi.org/10.1021/bi052031i. 
 
 

IRB00307426: Cellular Kinase and Aging                                                                                 PI: Zhu 
 
15 
APPENDIX I: SCHEDULE OF EVENTS 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
1After the screening visit, physical exam is symptom-directed; vital signs consist of heart rate and blood 
pressure only. 
2For participants not undergoing a concurrent endoscopy procedure.  
3 For PrEP cohort only in participants not already taking PrEP. 
4See Appendix II: HIV Testing Algorithm. Will not be completed for those with known HIV infection. 
5Syphilis Chemiluminescence assay (with confirmatory RPR; FTA-ABS performed on RPR negative 
samples)  
6For participants in PrEP or ARV Cohort taking TFV-based regimen for 7 days prior to procedure. 
7With the exception of rectal GC/CT screening, safety labs will not be obtained in participants undergoing 
screening colonoscopy. In those individuals, rectal GC/CT screening can be performed concurrently with 
Visit #2.  
 
 
Screening Visit 
Sampling Visit 
Visit Day 
< -42 
1 
Visit # 
1 
2 
ADMINISTRATIVE 
Informed consent 
x 
 
Assessment of Eligibility 
x 
 
HIV/STI Risk Reduction Counseling 
x 
 
Dispense and provide directions for enema2 
x 
 
Dispense TFV to TFV-naïve participants3 
 
 
CLINICAL 
History/Concomitant meds 
x 
 
Physical Exam, Vital signs, height, weight 1 
x 
 
Oral TFV-based regimen dosing and virtual DOT6 
Day -7 to -1 
 
SAFETY LABS6 
Blood CBC w/ diff, AST, ALT, BUN, Creatinine2 
x 
 
Blood HIV testing4 
x 
 
Blood syphilis screening5 
x 
 
Rectal swab GC/CT NAT 
x 
x6 
Blood HBsAg6 
x 
 
SPECIMEN COLLECTION 
Blood PBMC 
 
x 
Rectal biopsy 
 
x 
Blood for TFV6 
 
x