Clinical trial NCT03202511
Conditions: Pre-Exposure Prophylaxis
Sponsor: Indiana University
Start date: 2017-06-23; Completion date: 2018-07-02
Enrollment: 15 participants; Allocation: RANDOMIZED; Intervention model: CROSSOVER

Brief summary:
The study will be a randomized, open-label, cross-over clinical pharmacokinetic trial to investigate a strategy for probenecid "boosting" in the setting of tenofovir disoproxil fumarate (TDF)/emtricitabine (FTC) for HIV pre-exposure prophylaxis (PrEP). The study will be conducted at the Indiana University Clinical Research Center. All samples will be processed and the amount of tenofovir/FTC in plasma, blood, and urine, and tenofovir diphosphate and emtricitabine in peripheral blood mononuclear cells will be determined using validated analytical methods developed by the investigators at the University of Colorado. Probenecid plasma and urine concentrations will also be measured using an in-house assay. Following completion of the study, the secondary aim will be accomplished via analysis of selected samples collected at baseline and following treatment. Those selected samples will be assessed for urinary markers of proximal tubulopathy (urine total protein, albumin, creatinine, phosphorus, retinol binding protein, and beta-2-microglobulin) and serum alkaline phosphatase, osteocalcin, procollagen type 1 N propeptide, cystatin C, and creatinine to determine if the probenecid boosting strategy does indeed lead to less potential renal and bone toxicity.

Detailed description:

Arms: arm 1: The control phase will consist of subjects taking 600/400 mg oral tenofovir disoproxil fumarate/emtricitabine (TDF/FTC) on day 1 (2 tablets) followed by 300/200 mg (1 tablet) doses on days 2 and 3. arm 2: The treatment phase will consist of subjects taking 600/400 mg oral tenofovir disoproxil fumarate/emtricitabine (TDF/FTC) (2 tablets) along with a 2 gram oral probenecid (PRO) dose (4 tablets, 500 mg each) on day 1.
Interventions: intervention 1: Probenecid Oral Tablet intervention 2: Tenofovir disoproxil fumarate/Emtricitabine
Intervention descriptions: intervention 1: Included in arm/group descriptions. intervention 2: Included in arm/group descriptions.

Protocol documents:
Prot_SAP_000.pdf: 
PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
 
 
 
 
 
 
 
Assessment of a new ‘boosting’ strategy for 
HIV pre-exposure prophylaxis in healthy 
volunteers. 
 
 
Principal Investigator: 
Brandon T. Gufford, PharmD, PhD 
Assistant Professor of Medicine 
Division of Clinical Pharmacology 
Indiana University School of Medicine 
 
950 West Walnut Street | Research Institute II (R2), Room 466 
Indianapolis, IN 46202 
317.274.2847 tel 
bgufford@iu.edu 
 
Co-Investigators: 
Zeruesenay Desta, PhD 
Research Professor of Medicine, Division of Clinical Pharmacology 
Adjunct Senior Research Professor, Department of Pharmacology and 
Toxicology 
Indiana University School of MedicineSamir K. Gupta, MD, MS 
 
Vice Chair for Research, Department of Medicine 
Associate Professor of Medicine, Division of Infectious Diseases 
Director of Indiana Center for AIDS Research 
Indiana University School of Medicine 
 
Peter Anderson, PharmD 
Professor, Department of Pharmaceutical Sciences 
Director of the Pharmaceutical Sciences Graduate Program 
University of Colorado Skaggs School of Pharmacy 
 
 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
Table of Contents: 
 
Study Schema 
1.0 
Background 
2.0 
Rationale and Specific Aims 
3.0 
Inclusion/Exclusion Criteria 
3.1 
Selection of study participants 
3.2 
Inclusion Criteria 
3.3 
Exclusion Criteria 
4.0 
Enrollment/Randomization 
4.1 
Enrollment 
4.2 
Randomization 
5.0 
Study Procedures 
5.1 
Study Design 
5.2 
Study Procedures 
5.3 
Study Calendar 
5.4 
Sample Collection Schematic 
5.5 
DNA Genotyping 
5.6 
Analyses of Study Drugs 
6.0 
Reporting of Adverse Events or Unanticipated Problems involving Risk 
to Participants or Others 
6.1 
Risk Summary 
6.2 
Minimizing Potential Risks 
6.3 
Adverse Event Reporting 
6.4 
Data Safety and Monitoring Plan 
7.0 
Study Withdrawal/Discontinuation 
8.0 
Statistical Considerations 
9.0 
Privacy/Confidentiality Issues 
10.0 Follow-up and Record Retention 
11.0 References 
 
 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
1.0 
Background 
 
Prevention of HIV transmission is a worldwide imperative. On-demand HIV pre-exposure 
prophylaxis (PrEP, giving antiretroviral drugs to persons NOT infected with HIV) has 
been recently reported in the IPERGAY trial results as an effective approach to reduce 
the risk of HIV transmission among men who have unprotected sex with men.1 This 
study utilized a combination of antiretroviral drugs, Tenofovir Disoproxil Fumurate (TDF) 
and emtricitabine (FTC), taken before and after sexual activity. Although effective, this 
treatment regimen relies on adherence to intermittent drug administration regimens 
both before and after sexual activity. Perhaps of more practical importance, sexually 
active individuals (>2 sexual encounters per week) would require nearly daily drug 
administration to maintain protection. Poor adherence, especially in the setting of 
participants not currently in a monogamous relationship, has been suggested as a 
possible reason why some daily treatments may have failed in preventing HIV infections. 
As such, methods to improve adherence to pre-exposure prophylaxis regimens with oral 
antiretrovirals in HIV-negative individuals would be of great value.2 
 
TDF, a potent antiretroviral used in combination with FTC for PrEP, rapidly releases 
tenofovir (TFV) following oral administration, which subsequently is eliminated from the 
body entirely by the kidney. Part of this elimination process involves active transport of 
TDF into the urine by organic anion transporter (OAT) 1 and 3 (OAT1 and OAT3). 
Therefore, drugs inhibiting these transporters would be expected to reduce the renal 
excretion of TFV and prolong the time that the drug remains active in the body (i.e. 
prolong the elimination half-life). 
 
Probenecid is known to inhibit human OATs and thereby increases plasma 
concentrations and prolongs the half-life of many OAT substrates including ampicillin, 
cefamandole, cefazolin, cefoxitin, cephalothin, nalidixic acid, indomethacin, naproxen, 
and methotrexate at doses ranging from 500mg to 2g.3 Probenecid also improves the 
safety profile of drugs whose nephrotoxicity is mediated via uptake by organic anion 
transporters (e.g. the antiviral agent cidofovir). Over the past 50 years, probenecid has 
been combined with antibiotics and antivirals used in the treatment of gonorrhea, 
influenza, typhoid fever, mycobacterial diseases, and HIV.3 Peak action (as determined 
by the effect on penicillin plasma concentrations) after an oral dose of probenecid 
occurs at about 2 hours and the action lasts for approximately 8 hours with a dose 
dependent half-life (4 to 12 hours).4 
 
It follows that coadministration of TDF with PRO would be expected to reduce the 
elimination of TFV through the kidney and increase the length of time that TFV remains 
active in the body. Thus a combination of PRO, a well-tolerated oral drug, with TDF may 
allow simplified less frequent PrEP dosing, enhance TDF safety, substantially reduce 
costs, and thus make TDF-based PrEP more available and attractive for the intended 
population. A boosted single dose strategy of PRO-TDF given before anticipated sexual 
activity instead of the 4 doses of TDF given before and after currently being 
implemented would be more practical and may possibly improve adherence. Thus, this 
strategy should improve PrEP efficacy and compliance. Overall, PRO taken with TDF may 
drastically improve the likelihood of widespread uptake of pre-exposure prophylaxis in 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
both resource-replete and resource-limited settings and may greatly reduce the risk of 
HIV transmission around the world. 
 
In this pivotal proof of concept pharmacokinetic study, we propose to investigate a 
single dosing strategy for probenecid “boosting” of TDF for on-demand HIV pre-
exposure HIV prophylaxis. We estimate that approximately 14 healthy volunteers will 
need to complete this clinical study to determine if the single boosted PRO-TDF dosing 
strategy is viable. Data gathered from this study may be used to support future 
pragmatic trials of PRO-boosted TDF in individuals at high risk for HIV acquisition. 
Ultimately, the goal of this research is to create a simple and effective on-demand HIV 
PrEP regimen to reduce the incidence of HIV transmission worldwide. 
 
2.0 
Rationale and Specific Aims 
 
Research Aims 
Hypothesis: Probenecid “boosting” can be used to increase TFV concentrations, 
decrease TDF dosing requirements, and simplify TDF dosing regimens in the setting of 
on-demand HIV PrEP 
 
Primary Aim: Test whether TFV blood and blood cell concentrations of tenofovir 
diphosphate (TFV-DP) of a single probenecid-boosted TDF regimen are comparable to 
those achieved with the current ‘on-demand’ unboosted TDF dosing regimen of two 
tablets before and two tablets after sexual activity. 
 
Secondary Aim: Compare the short-term safety and tolerability of the single PRO-TDF 
regimen compared to the unboosted TDF regimen.  
 
Rationale 
 
Evidence of probenecid boosting of anti-infective agents: 
Probenecid (1 gram orally) increases amoxicillin systemic exposure, measured by the 
area under the plasma concentration-time curve (AUC) by 53%5 and in one small study, 
reduced dosing requirements by two-thirds.6 Probenecid also increases acyclovir plasma 
AUC by 48% following simultaneous administration of 1g probenecid with 1g 
valacyclovir.7 Probenecid increased the plasma AUC of cidofovir 24% for male monkeys 
and 86% for female monkeys administered 30mg/kg probenecid with 2.5mg/kg of 
cidofovir.8 One small study compared cidofovir pharmacokinetics in serum using 
standard probenecid dosing (2g before the cidofovir infusion, then 1g at 2 and 8 hours 
after the cidofovir infusion) vs. a lower probenecid dose, 2g 1-hour before cidofovir 
infusion with no subsequent doses, and found the cidofovir pharmacokinetics to be the 
same.9 This suggests lower doses of probenecid, such as the 2 gram dose utilized in this 
study, may offer the same pharmacokinetic “boosting” benefits of higher doses.  
 
3.0 
Inclusion/Exclusion Criteria 
 
3.1 Selection of study participants: A total of 16 healthy male volunteers (18 to 55 
years old) who meet all inclusion and exclusion criterion listed below will be enrolled to 
participate in this study. Based on our previous experience, a 15% drop out rate (n=2) 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
is expected and will be considered to ensure that at least 14 volunteers complete the 
entire study. Approval to conduct the study will be obtained in advance from the 
Institutional Review Board (IRB) of the Indiana University. All subjects will sign an IRB 
approved written informed consent prior to participation in the study Each subject will 
be provided a copy of the informed consent and allowed sufficient time to read, 
understand, and formulate questions about any aspect of the study. A member of the 
study team will carefully review the consent with each subject and answer any 
questions. Then, potential subjects will undergo a pre-enrollment screening examination 
(to be accomplished within a maximum of six weeks prior to enrollment into the study) 
for any medical abnormalities which will include medical histories, vital signs.  
demographic variables, standard laboratory blood and urine tests, and screening for HIV 
and HBV infection. The screening will be performed at the Indiana Clinical Research 
Center (ICRC). During the screening, a blood sample (15 ml) will be collected from 
each subject for laboratory tests and urine for urine analyses. An additional ∼10 ml will 
be obtained from each subject to extract genomic DNA for genotyping purposes. 
 
3.2 Inclusion criteria:  
 
1. 18 to 55 years old healthy (as decided from a pre-enrollment screening session 
described above) male participants within 32% of their ideal body weight. 
2. Individuals who agree to refrain from taking any prescriptions medications, over-the-
counter medications (including salicylates/aspirin), hormonal agents, and herbal, 
dietary, and alternative supplements that may interact with the metabolism of those 
study drugs at least 2 weeks prior to the start of the study and until study 
completion.   
3. Nonsmoker or individuals willing to refrain from smoking or use of tobacco or 
marijuana for at least one month prior to and until the completion of the study (the 
entire study lasts for approximately 49 days).  
 
3.3 Exclusion criteria:  
 
Subjects will not be allowed to participate in the study if they at time of study screening: 
1. Are underweight (weigh less than 52 kg or 114 lb) or overweight [body mass index 
(BMI) greater than 32].  
2. Females will be excluded to reduce study variability for this first proof of concept 
study. 
3. Have insufficient renal function (estimated Creatinine Clearance ≤ 90 mL/min). 
4. Have history of current alcohol or drug abuse (more than 4 alcoholic drinks per day 
on a regular basis). 
5. Have history of intolerance, allergic reactions (e.g. rash) or other forms of 
hypersensitivities to any of the study medications (tenofovir disoproxil fumurate/ 
emtricitabine, probenecid). 
6. Have taken TDF or FTC as part of PrEP within the past 6 weeks. 
7. Any current major illness or chronic illness such as (but not limited to) kidney 
disease, hepatic disease, diabetes mellitus, gout, hypertension (definition?), coronary 
artery disease, chronic obstructive pulmonary disease, cancer, chronic active HBV 
infection, or HIV. 
8. History of anemia or any other significant hematologic disorder. 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
9. Have history or current gastrointestinal disorders such as persistent diarrhea or 
malabsorption that would interfere with the absorption of orally administered drugs. 
10. Have a serious infection within the last week before study enrollment. 
11. Have donated blood within the past two months. 
12. Have blood results that do not fall in a healthy range (e.g., blood hemoglobin less 
than 12.0 mg/dl).  
13. Are taking on regular basis substances that may interfere with the metabolism 
(breakdown) of study medications by the body, including prescription medications, 
over-the-counter, herbal or dietary supplements, alternative medications, or 
hormonal agents (i.e. oral contraceptives, intra-uterine device with hormones). 
14. Have a life style that places subjects at a higher risk for contracting HIV during the 
study period (e.g. active illicit drug use, excessive alcohol drinking, sexually 
transmitted infection (including gonorrhea, chlamydia, syphilis, herpes, HPV) within 
the past one year, or having more than one sexual partner in the past 6 months). 
15. Positive HIV antibody test. 
16. Positive HBV surface antigen test. 
17. Have participation in a research study or use of an investigational drug in the last 
one month. 
18. Are employed or are student under supervision of any of the investigators of this 
study. 
19. Cannot state a good understanding of this study including risks and requirements; 
are unable to follow the rules of this study.  
20. Cannot commit the time requested for this study.  
 
4.0 
Enrollment/Randomization 
 
4.1 Enrollment: 
Subjects will be recruited from 1) IRB approved campus fliers affixed to community 
bulletin boards; 2) verbal script using the attached template; or 3) via email of the pre-
recruitment script. No subject will be excluded from the study on the basis of ethnicity 
or race. We will include all minorities. We expect that our recruitment of healthy 
volunteers will reflect the ethnic and racial demographics of the Indianapolis 
metropolitan based on current local clinic and hospital visits, state census and previous 
studies at the Indiana University School of Medicine. Having completed multiple similar 
trials in the past, we have developed experience in recruiting subjects and in managing 
the complex logistics of clinical trials. 
4.2 Randomization: 
Subjects will be randomly allocated to enter the study in either the control or treatment 
arm using a blocked design to ensure equal numbers of subjects in each sequence 
group (n=8, control to treatment; n=8, treatment to control). Following a washout 
period of at least 6 weeks, each individual will crossover to the next study phase. The 
randomization table will be constructed prior to subject enrollment by the Principal 
Investigator using the Statistical Analysis Software (SAS®) randomization procedure. In 
the event that an individual is withdrawn from the study, an alternate participant will 
replace that individual using an identical study sequence to maintain the block design. A 
total of 16 subjects will be initially enrolled in an effort to ensure that at least 14 
subjects complete the entire study. If needed, additional subjects will be screened and 
enrolled until the target enrollment of n=14 subjects have completed the entire study. 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
 
5.0
Study Procedures 
 
5.1 Study Design: 
The study will be a randomized, open-label, cross-over clinical pharmacokinetic trial to 
investigate a strategy for probenecid “boosting” in the setting of tenofovir 
DF/emtricitabine for HIV PrEP. The study will be conducted at the Indiana University 
Clinical Research Center. All samples will be processed and the amount of tenofovir/FTC 
in plasma, blood, and urine, and tenofovir diphosphate and emtricitabine in peripheral 
blood mononuclear cells will be determined using validated analytical methods 
developed by the investigators at the University of Colorado. Probenecid plasma and 
urine concentrations will also be measured using an in-house assay. Following 
completion of the study, the secondary aim will be accomplished via analysis of selected 
samples collected at baseline and following treatment. Those selected samples will be 
assessed for urinary markers of proximal tubulopathy (urine total protein, albumin, 
creatinine, phosphorus, retinol binding protein, and beta-2-microglobulin) and serum 
alkaline phosphatase, osteocalcin, procollagen type 1 N propeptide, cystatin C, and 
creatinine to determine if the probenecid boosting strategy does indeed lead to less 
potential renal and bone toxicity.  
 
5.2: Study Procedures: 
Overview: The study will be a randomized, open-label, cross-over clinical 
pharmacokinetic trial to investigate a strategy for probenecid “boosting” in the setting of 
tenofovir DF/emtricitabine for HIV PrEP in healthy volunteers. Potential subjects will 
undergo a screening appointment with those deemed eligible being enrolled and 
randomized to enter the study in either the control or treatment phase. Following 
completion of the first phase, subjects will crossover to the next phase after a washout 
period of at least 6 weeks. 
 
 
 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
Pre-Screening Recruitment: Subjects interested in study participation will be pre-
screened by a member of the study team. Each potential subject will be provided 
standardized information relating to the general study purpose, inclusion/exclusion 
criteria, time commitment, scheduling, and study expectations. This information will be 
provided via a standard recruitment script. Each subject will be asked to complete a pre-
screening questionaire. Based upon the pre-screening conversation and questionaire, 
those individuals believed to be eligible for participation will then be scheduled for 
screening. 
 
Screening: A copy of the informed consent document will be provided to each subject 
and each subject will have adequate time to review and formulate questions about the 
study. Each subject will be allowed to review the consent document in a private room or 
office. A member of the study team will then review the informed consent document, 
ensure subject understanding, and answer questions that the subject might have about 
participating in the study. Following this process, the subject will be asked if they agree 
to consent to participate in the study. Subjects will provide a written signature using 
indelible ink on a hard copy of the consent document. The orginal signed hard copy of 
the consent document will be retained by the investigator and a copy of the consent 
document will be given to each subject. 
 
After obtaining informed consent but during the initial screening, a blood sample (15 
ml) will be collected from each subject for laboratory tests and urine for urine analyses. 
An additional ∼10 ml will be obtained from each subject to extract genomic DNA for 
genotyping purposes. Only those subjects that complete the entire study will have their 
DNA analyzed. 
 
Control Phase: The control phase will consist of subjects taking a regimen mirroring 
that of the IPERGAY trial: 600 mg oral TDF/FTC on day 1 (2 tablets) followed by 300 mg 
(1 tablet) doses on days 2 and 3. 
 
Treatment Phase: The treatment phase will consist of subjects taking 600 mg oral 
TDF/FTC (2 tablets) along with a 2 gram oral probenecid (PRO) dose (4 tablets) on day 
1. 
 
5.3 Study Calendar:  
Day 1: 
 
Eligible subjects will be requested to arrive in the morning at the ICRC (about 7 
a.m.) on the first study day after fasting overnight. On the evening before 
admission to the ICRC inpatient study day (day 1), it is important that subjects 
have nothing to eat or drink after 11 pm except for water. 
 
Vital signs (e.g., sitting blood pressure, respiration rate, pulse rate, oral 
temperature) will be obtained prior to initiating study procedures.  
 
A sterile intravenous catheter will be inserted in one arm for blood collection.  
 
Prior to administration of study drugs, a baseline blood (6 ml) and urine sample 
will be collected. 
 
Subjects will receive 600 mg TDF/FTC (2 tablets) alone (control phase) or with 
probenecid (treatment phase) by mouth with ~250 mL water. A standard meal 
will be served approximately 4 hours after drug dosing. 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
 
Samples will be collected as outlined below. 
Day 2: 
 
After the 24 hour blood draw and urine collection, the venous catheter will be 
removed and subjects will be discharged to go home. One urine collection 
container will be provided to them to collect urine over the period of 24-48 
hours. They will be instructed to bring the collected urine with them on day 3 
when they visit the ICRC.  
 
During the control phase subjects will receive a second TDF/FTC tablet 
immediately following the 24 hour sample collection. 
Day 3: 
 
Subjects will be requested to return to the ICRC on the morning day 3 (48 hours 
post dose) for a brief outpatient visit and single blood draw. The outpatient 
blood sample (6 mL) will be collected via a peripheral venipuncture. An additional 
urine collection container will be provided to them to collect urine over the period 
of 48-72 hours. They will be instructed to bring the collected urine with them on 
day 4 when they visit the ICRC. 
 
During the control phase subjects will receive a third TDF/FTC tablet immediately 
following the 48 hour sample collection. 
Day 4: 
 
Subjects will be requested to return to the ICRC on the morning of day 4 (72 
hours post dose) for a brief outpatient visit and single blood draw. The 
outpatient blood sample (6 mL) will be collected via peripheral venipuncture.  
 
Subjects will be instructed to return to the ICRC for the next study phase 
following the 6 week washout period. 
 
Control Phase 
Day 1: 
Inpatient ICRC 
Day 2: 
Discharge 
Day 3: 
Outpatient 
Day 4: 
Outpatient 
Administer 2 tablets of 
TDF/FTC 
Obtain 24 hour blood 
sample 
Obtain 48 hour blood 
sample 
Obtain 72 hour blood 
sample 
Obtain 
blood 
and 
urine 
samples 
as 
outlined below 
Administer 
1 
tablet 
TDF/FTC 
Administer 
1 
tablet 
TDF/FTC 
Collect urine collection 
containers 
from 
subject 
 
Discharge subject with 
urine 
collection 
container 
Discharge subject with 
urine 
collection 
container 
 
Treatment Phase 
Day 1: 
Inpatient ICRC 
Day 2: 
Discharget 
Day 3: 
Outpatient 
Day 4: 
Outpatient 
Administer 2 tablets of 
TDF/FTC 
and 
probenecid 
Obtain 24 hour blood 
sample 
Obtain 48 hour blood 
sample 
Obtain 72 hour blood 
sample 
Obtain 
blood 
and 
urine 
samples 
as 
outlined below 
Discharge subject with 
urine 
collection 
container 
Discharge subject with 
urine 
collection 
container  
Collect urine collection 
containers 
from 
subject 
 
5.4 Sample Collection Schematic: The sample collection schedule (outlined below) 
will be identical for the control and treatment study phases. The control and test phase 
will each consist of intensive pharmacokinetic sampling with serial blood draws occurring 
at baseline (0), 0.5, 1, 2, 4, 8, 12, 18, 24, 48, and 72 hours following administration of 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
the first dose TDF/FTC dose in each phase. Each phase will be separated by a minimum 
6 week washout period to ensure that any residual TFV-DP has been completely 
eliminated from PBMCs. The blood volume collected during the control and treatment 
phases will be approximately 66 mL/phase in addition to ~25 mL collected during 
screening to yield a total blood collection volume of ~157 mL or 2/3 cup per subject 
during the entire study. 
 
 
 
5.5: DNA Genotyping: 
Exploratory genotyping efforts will focus on known variants in transporter proteins that 
may alter the disposition of tenofovir and/or emtricitabine. Variants will include single 
nucleotide polymorphisms identified in the following genes: SLC22A6 (OAT1), SLC22A8 
(OAT3), ABCC2 (MRP2), and ABCC4 (MRP4). Genomic DNA will be extracted from 
human blood using standard operating procedures. Genotyping for these variants will be 
performed by use of the predeveloped TaqMan Assay-Reagents Allelic Discrimination 
Kits (Applied Biosystems, Foster City, CA) according to the supplier's instructions.  
 
5.6: Analyses of Study Drugs 
Plasma and Urine Analysis 
Tenofovir and Emtricitabine: Within 24 hours following collection, plasma (harvested 
from blood via centrifugation) and urine samples will be processed using standard 
operating procedures within the Indiana CTSI Specimen Storage Facility (SSF). 
Simultaneous determination of tenofovir and emtricitabine concentrations in plasma and 
urine samples will be accomplished using a validated liquid chromatography-tandem 
mass spectrometry (LC-MS/MS) assay established and routinely performed by co-
investigators at the University of Colorado.10  


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
Probenecid: We will develop an LC-MS/MS based assay for the determination of 
probenecid concentrations in plasma and urine samples. It is anticipated that probenecid 
can be determined in conjunction with tenofovir and emtricitabine by adapting the assay 
outlined above. 
 
Intracellular PBMC Analysis 
Tenofovir Diphosphate and Emtricitabine: Within 24 hours following collection, 
PBMCs will be recovered and counted from whole blood using standard operating 
procedures within the Indiana CTSI SSF. Due to extensive processing time and cost 
PBMCs will be isolated only from samples at selected time points. Isolated PBMC 
samples will be shipped (de-identified) to the University of Colorado for analysis using a 
validated and routinely performed LC-MS/MS assay.11 
 
Dry Blood Spot Analysis 
Dry blood spot (DBS) samples will be collected from whole blood prior to harvesting of 
plasma at each time point. DBS samples will be prepared according to standard 
operating procedures within the Indiana CTSI SSF and analyzed via LC-MS/MS using a 
validated method established by co-investigators at the University of Colorado.12 
 
Following processing, all samples will be stored at –80°C pending analysis. 
 
6.0 
Reporting of Adverse Events or Unanticipated Problems involving Risk 
to Participants or Others 
 
6.1 Risk Summary: 
 
Venipuncture: Blood samples will be collected through puncture of a vein by a needle. 
This may cause slight pain, bruising, and bleeding from the site of the needle puncture 
into the vein, and in rare cases fainting and infection. To minimize these discomforts 
and risks, only trained and experienced nurses or physicians will place the intravenous 
catheter in the vein and collect blood samples. Aseptic technique and hospital/nursing 
standards will be followed. 
 
Blood Loss: The maximum blood volume that will be collected from each subject over 
the course of the entire study (duration is ~50 days) is expected to be <200 mL. The 
total amount includes potential for repeat screening and exit laboratory tests which may 
be needed for safety reasons as deemed by the study physician. To minimize the impact 
of blood loss, subjects accepted into the study must have a hemoglobin count of 12.0 
g/dl or above. Given the total volume of blood sampling and the duration of the study, 
we would expect that the body will adequately replace this blood and should pose no 
risk to a healthy subject. 
 
Study Drugs: All drugs to be used in this study (TDF/FTC, probenecid) are FDA 
approved, commercially available, and are under wide clinical use. Study drugs will be 
purchased and handled through the Indiana University Hospital Pharmacy, 
Investigational Drug Service section. Potential risks associated with each study drug are 
outlined below. 
 
 
Tenfovir Disoproxil Fumurate / Emtricitabine 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
Tenofovir disoproxil fumurate (TDF) / Emtricitabine (FTC) is administered as a fixed 
dose combination tablet containing 300 mg TDF and 200 mg FTC. Truvada is the 
tradename for this FDA-approved fixed-dose combination. Truvada belongs to a class of 
antiretroviral drugs known as nucleoside/nucleotide reverse transcriptase inhibitors, 
commonly known as NRTIs. The most common risks and side effects from NRTIs include 
fatigue (tiredness), headache, mild stomach discomfort, nausea (upset stomach), 
insomnia (difficulty sleeping), rash, vomiting, and diarrhea.  
 
Rare but more serious risks and side effects include pancreatitis, severe liver problems, 
anemia (low red blood cells), lactic acidosis (buildup of lactate in the body), peripheral 
neuropathy (pain, tingling, and/or numbness in the hands and/or feet), lipodystrophy 
(alterations in fat distribution in your body), decreased kidney function, and metabolic 
disorders (changes in lipid and sugar levels in your body). 
 
 
Probenecid 
“Probenecid has been administered millions of times to hundreds of thousands of people 
with very few adverse effects and almost no serious adverse effects”.3 When used with 
cidofovir at high doses (2 g before the cidofovir infusion, then 1g at 2 and 8 hours after 
the cidofovir infusion) for the treatment of cytomegalovirus in persons with AIDS, 
nausea, vomiting, fever, headache, or rash occur in 44%-56% of patients and result in 
treatment discontinuations in 4%-7%.13,14 However, these side effects may be reflective 
of cidofovir use or underlying medical conditions rather than PRO itself. The safety 
profile of PRO in the setting of TDF-boosting in healthy individuals has not been 
determined. This may be important if the increased nausea and vomiting seen with TDF 
vs. placebo in other PrEP studies may be exacerbated with PRO.15 Furthermore, the 
effectiveness of PrEP using a PRO-boosted TDF strategy may be limited by tolerability. 
Thus, the proposed study will assess the short-term adverse event profile of PRO and 
inform future studies on the acceptance of this drug combination. Patients will be 
advised not to use salicylates (e.g. aspirin) during the course of the study due to a 
potential drug-drug interaction with probenecid. Subjects who require mild analgesia 
during the study will be advised to use acetaminophen (brand name: Tylenol). 
 
Other Possible Risks: The study drug treatment combination may involve risks that 
are currently unknown or unpredictable. Important new findings uncovered during the 
course of this study that may impact subject safety or willingness to participate will be 
provided to subjects. Study investigators and ICRC staff will routinely ask subjects about 
any potential side effects. 
 
Efforts will be made to keep all personal information confidential. However, a risk of loss 
of confidentiality still exists. 
 
6.2 Minimizing Potential Risks: Any changes in subject status that might be judged 
unsafe to the subject will immediately lead to discontinuation of therapy and early 
termination of the study for that individual. The research nursing staff, investigators, 
and subjects will not be blinded to treatment for reasons of safety. The patients will be 
asked to report any intolerance or bothersome side effects that occur during the study. 
All subjects participating in this study will be informed in detail about potential risks, 
followed closely, and encouraged to report any adverse reactions immediately. During 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
each inpatient clinical day advanced cardiac life support equipment and oxygen will be 
available to treat hypoventilation or apnea and intravenous fluids will be available to 
treat hypovolemia. Low wall suction equipment and resuscitation equipment including 
chemical code drugs will be available at the ICRC. In addition, medication orders for 
anti-emetic therapy will be provided in the event that any subject experiences nausea 
and/or vomiting as a result of the study medications. 
 
6.3 Adverse Event Reporting: Patients will be closely monitored for any evidence of 
adverse events. Any adverse effects associated with the study intervention (as deemed 
by the PI and/or study physician) will be documented and reported to appropriate 
bodies (IRB, FDA) using standard procedures. The subjects participating in this study 
will be informed in detail about the potential risk and encouraged to report any adverse 
reactions immediately.  
 
Adverse events will be assessed using the NCI Common Toxicity Criteria guidelines listed 
in (http://ctep.cancer.gov/forms/CTCAEv3.pdf) along with potential study drug specific 
toxicities as outlined above. 
Adverse events will be graded as follows: 
Grade 0 – No adverse effects 
Grade 1 - Mild adverse events 
Grade 2 – Moderate adverse events 
Grade 3 – Severe adverse events 
Grade 4 – Life threatening or disabling adverse events 
Grade 5 – Death related to adverse events 
 
Any serious and unexpected adverse events associated with the study intervention will 
be reported to Research and Sponsored Programs and the ICRC within 3 working days 
of notification of the event. Deaths will always be reported if they occur within 30 days 
of study intervention. The principal investigators will be responsible for reporting the 
adverse effects to the IRB and other relevant bodies. All adverse events that do not 
meet immediate reporting requirements will be submitted to the IRB at time of 
continuing review. 
 
6.4 Data and Safety Monitoring Plan: A Data and Safety monitoring board will 
created to provide additional study oversight and to ensure patient safety. The DSMB 
will be chaired by Michael Eadon, MD. Dr. Eadon is board-certified in internal medicine, 
nephrology, and clinical pharmacology. These credentials make Dr. Eadon an excellent 
individual to assess the safety of this study which focuses on modulation of renal drug 
transport. Additional members will include Samir Gupta, MD, MD, Brandon Gufford, 
PharmD, PhD, and Zeruesenay Desta, PhD. The DSMB will be charged with: (1) review 
of the research protocol and establishment of plans for data and safety monitoring, (2) 
study progress evaluation such as assessments of data quality, timeliness, subject 
recruitment, accrual, retention, and subject risk vs. benefit, (3) reviewing scientific or 
therapeutic developments that may have an impact on the safety of the participants or 
the ethics of the study, (4) reviewing major proposed modifications to the study prior to 
their implementation, including termination, dropping an arm of the study, or increasing 
the target sample size, (5) submitting reports of recommendations to the IRB or funding 
foundation that include the current status of the study, summary reports of adverse 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
events resulting from participation in the study, and determination of whether the study 
should continue as originally designed, should be changed, or should be terminated.  
 
The first DSMB meeting will be held once three subjects have completed the study in 
order to evaluate the data produced. Data quality, subject recruitment, accrual, adverse 
events, and results of related studies that impact subject safety will be evaluated. The 
voting members of the DSMB will determine the frequency of future DSMB meetings. 
The frequency may be based on accrual rates (i.e. after 8 volunteers are enrolled) or 
elapsed time (i.e. every two months).  
 
7.0 
Study Withdrawal/Discontinuation 
 
Taking part in this study is voluntary. Subjects can choose not to take part or to 
withdraw from this study at any time without penalty. Leaving the study will not result in 
any penalty or loss of benefits to which the subject is entitled. The decision of any 
subject to participate or not to participate will not affect: current or future relationship 
with the research team; the status as a volunteer, employee, student, etc; or medical 
care. The principal investigator reserves the right to withdraw a subject from the study 
without regard to subjects consent in the following circumstances: if risks in 
participating in this study outweigh their best interest in terms of safety; or if subject is 
unable to adhere to the study requirements. In addition, this study may be terminated 
by the IRB at their discretion. 
 
Subjects have the option of participating or not participating in the study. There are no 
other alternative procedures or courses of treatment that may be advantageous to 
them. Subjects will receive $225 for each 24 hour overnight stay (Day 1 of the Control 
and Treatment phase) and $50 for each outpatient visit (Days 3 and 4 of the Control 
and Treatment phase). Subjects who complete all phases of the study will receive a total 
of $650. Individuals that withdraw from the study (voluntarily or at the discretion of the 
PI) will be compensated for the portions of the study which were completed. 
Compensation will be issued at the completion/withdrawal of the study. 
 
8.0 
Statistical Considerations 
 
The area under the TFV plasma and TFV-DP PBMC concentration-time curve (AUC) will 
be calculated via routine non-compartmental approaches. The primary trial outcome will 
be the geometric mean ratio (GMR) calculated as the test phase/control phase TFV 
plasma and TFV-DP PBMC AUCs. A two one-sided statistical testing procedure, as 
recommended by the FDA, will be used for the primary endpoint analysis.16 If the 90% 
confidence interval for the geometric mean ratio is contained entirely within the bounds 
of 0.8-1.25 then the probenecid “boosting” strategy is determined to provide equivalent 
tenofovir diphosphate concentrations relative to the IPERGAY regimen. Using this study 
design allows each subject to serve as his/her own control to decrease variability and 
reduce the required sample size. Power and sample sizes were determined based on a 
study of daily tenofovir DF/emtricitabine in healthy volunteers by Peter Anderson.17 At 
day 30 (steady state), the mean (SD) TFV-DP concentration in the PBMCs of 18 
participants was 100.3 (±32.3) fmol/106 cells.  
 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
To demonstrate equivalence of TFV-DP concentrations in PBMCs with probenecid 
“boosting” strategies relative to the IPERGAY trial TDF/FTC regimen (i.e. control group) 
would require 14 total participants in a crossover design. A total of 14 evaluable subjects 
with complete data will provide 90% power to detect a mean difference of 25% in the 
primary outcome assuming a Type I error rate of 0.05. Given that dropouts are likely for 
any PK study of this duration, we estimate that 16 participants will need to be enrolled 
to achieve the final 14 participants who complete the entire crossover trial. In the event 
that an individual is withdrawn from the study, an alternate participant will replace that 
individual using an identical study sequence to maintain the randomized block design. 
Additional subjects will be screened and enrolled until the target of n=14 subjects have 
completed the entire study. 
 
9.0 
Privacy/Confidentiality Issues 
 
The study data and results are for research purposes only. To the extent allowed by law, 
information about study subject’s personal and medical data is considered confidential. 
Although we cannot guarantee absolute confidentiality, every effort will be made to 
keep all subject information strictly confidential. The samples collected from this study 
will be stored in locked freezers and only authorized personnel will have access to the 
samples, coded databases, and the results. The samples will be labeled with a code and 
not with subject names. All subject identified information will be stored in locked 
cabinets and on the PIs computer that is password protected. It will not be possible for 
anyone other than the investigators to determine subject specific test results without the 
explicit written consent of the subject or unless allowed by law. It is possible that data 
collected during the screening and exit lab tests, and inpatient care may become part of 
subject’s medical record. However, DNA genotyping results will not be recorded in 
medical records. All DNA samples will be marked with a unique code number and stored 
in an anonymous fashion in two different secured computer databases; one containing 
the DNA sample codes and the other with subject information (such as age, sex, ethnic 
group, health conditions, etc.) to maximize confidentiality.   
 
Personal information may be disclosed if required by law. People or organizations that 
may review or inspect personal information to verify the quality of the research and data 
analysis include the study principal investigator and his research team, the Indiana 
University Institutional Review Board or its designees, the study sponsor which is the 
Campbell Foundation or their designees, and the Indiana Clinical Research Center 
(ICRC). In addition, as allowed by law, state or federal agencies, specifically the Office 
for Human Research Protections (OHRP) and the Food and Drug Administration (FDA), 
may need to access medical and/or research records. 
 
Samples or information will not be released to anyone other than those listed above 
without obtaining a written consent from the subjects who participated in the study. 
When the results of study are published, no names or specific identifiers will be used. In 
addition, we will not use this sample to characterize subject’s genetic history or 
determine the likelihood that they will experience a disease or not. The genetic 
information gathered in this study will be used together with information from other 
participants. Therefore, while the possibility that participation in this study may cause a 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
loss of privacy cannot be excluded entirely, the steps outlined above will be 
implemented to significantly minimize the risk of loss confidentiality. 
 
Finally, a description of this clinical trial will be made available on ClinicalTrials.gov, as 
required by U.S. Law. This Web site does not include information that can identify 
individual subjects. At most, the site will include a summary of the protocol and results. 
Subjects have the opportunity to search this freely available online resource at any time. 
 
10.0 Follow-up and Record Retention 
 
This study is expected to be completed within one year. Records (documents) will be 
kept to the extent allowed by law per NIH recommendations after the study is closed 
(typically up to 7 years). All documents will be destroyed by shredding any document 
holding study information or through deletion of computer files using HIPPA compliant 
digital shredding utilities. Subjects retain (keep) the right to have any remaining 
biological sample material destroyed at any time by contacting the principal investigator. 
The principal investigator is responsible for the destruction of biological samples at the 
subject’s request. Any data from this study will be published without subject names. 
 
11.0 References 
 
1 
Molina JM, Capitant C, Spire B, Pialoux G, Cotte L, Charreau I et al. On-Demand 
Preexposure Prophylaxis in Men at High Risk for HIV-1 Infection. N Engl J Med 
2015; 373: 2237-2246. 
 
2 
Anderson PL, Garcia-Lerma JG, Heneine W. Nondaily preexposure prophylaxis for 
HIV prevention. Curr Opin HIV AIDS 2016; 11: 94-101. 
 
3 
Robbins N, Koch SE, Tranter M, Rubinstein J. The history and future of 
probenecid. Cardiovasc Toxicol 2012; 12: 1-9. 
 
4 
Cunningham RF, Israili ZH, Dayton PG. Clinical pharmacokinetics of probenecid. 
Clin Pharmacokinet 1981; 6: 135-151. 
 
5 
Barbhaiya R, Thin RN, Turner P, Wadsworth J. Clinical pharmacological studies of 
amoxycillin: effect of probenecid. Br J Vener Dis 1979; 55: 211-213. 
 
6 
Allen MB, Fitzpatrick RW, Barratt A, Cole RB. The use of probenecid to increase 
the serum amoxycillin levels in patients with bronchiectasis. Respir Med 1990; 
84: 143-146. 
 
7 
De Bony F, Tod M, Bidault R, On NT, Posner J, Rolan P. Multiple interactions of 
cimetidine and probenecid with valaciclovir and its metabolite acyclovir. 
Antimicrob Agents Chemother 2002; 46: 458-463. 
 
8 
Lacy SA, Hitchcock MJ, Lee WA, Tellier P, Cundy KC. Effect of oral probenecid 
coadministration on the chronic toxicity and pharmacokinetics of intravenous 
cidofovir in cynomolgus monkeys. Toxicol Sci 1998; 44: 97-106. 


PRINCIPAL INVESTIGATOR: Brandon Gufford, PharmD, PhD 
Revision Date: 07/05/2017 
 
IRB Protocol: 1705315090 
 
9 
Wolf DL, Rodriguez CA, Mucci M, Ingrosso A, Duncan BA, Nickens DJ. 
Pharmacokinetics and renal effects of cidofovir with a reduced dose of 
probenecid in HIV-infected patients with cytomegalovirus retinitis. J Clin 
Pharmacol 2003; 43: 43-51. 
 
10 
Delahunty T, Bushman L, Robbins B, Fletcher CV. The simultaneous assay of 
tenofovir and emtricitabine in plasma using LC/MS/MS and isotopically labeled 
internal standards. J Chromatogr B Analyt Technol Biomed Life Sci 2009; 877: 
1907-1914. 
 
11 
King T, Bushman L, Kiser J, Anderson PL, Ray M, Delahunty T et al. Liquid 
chromatography-tandem mass spectrometric determination of tenofovir-
diphosphate in human peripheral blood mononuclear cells. J Chromatogr B 
Analyt Technol Biomed Life Sci 2006; 843: 147-156. 
 
12 
Zheng JH, Rower C, McAllister K, Castillo-Mancilla J, Klein B, Meditz A et al. 
Application of an intracellular assay for determination of tenofovir-diphosphate 
and emtricitabine-triphosphate from erythrocytes using dried blood spots. J 
Pharm Biomed Anal 2016; 122: 16-20. 
 
13 
Lalezari JP, Holland GN, Kramer F, McKinley GF, Kemper CA, Ives DV et al. 
Randomized, controlled study of the safety and efficacy of intravenous cidofovir 
for the treatment of relapsing cytomegalovirus retinitis in patients with AIDS. J 
Acquir Immune Defic Syndr Hum Retrovirol 1998; 17: 339-344. 
 
14 
Lalezari JP, Stagg RJ, Kuppermann BD, Holland GN, Kramer F, Ives DV et al. 
Intravenous cidofovir for peripheral cytomegalovirus retinitis in patients with 
AIDS. A randomized, controlled trial. Ann Intern Med 1997; 126: 257-263. 
 
15 
Glidden DV, Amico KR, Liu AY, Hosek SG, Anderson PL, Buchbinder SP et al. 
Symptoms, Side Effects and Adherence in the iPrEx Open-Label Extension. Clin 
Infect Dis 2016; 62: 1172-1177. 
 
16 
Food and Drug Administration Center for Drug Evaluation and Research. Drug 
Interaction Studies - Study Design, Data Analysis, Implications for Dosing, and 
Labeling Recommendations (Draft Guidance) 2012; doi. 
 
17 
Baheti G, Kiser JJ, Havens PL, Fletcher CV. Plasma and intracellular population 
pharmacokinetic analysis of tenofovir in HIV-1-infected patients. Antimicrob 
Agents Chemother 2011; 55: 5294-5299.