Clinical trial NCT02956278
Conditions: Chronic Gout Hyperuricemia
Keywords: allopurinol
Sponsor: University of California, San Francisco
Start date: 2016-11-01; Completion date: 2018-03-01
Enrollment: 27 participants; Allocation: NON_RANDOMIZED; Intervention model: PARALLEL

Brief summary:
Subjects will undergo a placebo and allopurinol phase to better understand the effects of the reduced function BCRP Q141K variant on allopurinol pharmacokinetics and pharmacodynamics.

Detailed description:

Arms: arm 1: Participants that are homozygous reference for BCRP Q141K receive at least one 300 mg dose of allopurinol followed by blood/urine collection for up to 72 hours post-dose. arm 2: Participants that are heterozygous for the BCRP Q141K allele receive at least one 300 mg dose of allopurinol, with blood/urine collections up to 72 hours post-dose. arm 3: Participants homozygous for the BCRP Q141K allele receive at least one 300 mg dose of allopurinol with blood/urine collection for up to 72 hours post-dose.
Interventions: intervention 1: Allopurinol intervention 2: Placebo
Intervention descriptions: intervention 1: None intervention 2: lactose placebo pill

Protocol documents:
Prot_SAP_001.pdf: 
 
The Effects of BCRP Q141K on Allopurinol Pharmacokinetics and 
Dynamics 
 
NCT02956278 
Sept 2017 


 
 
 
PURPOSE: To investigate the potential association of polymorphic genetic variants 
ABCG2 with altered response to allopurinol, we will employ a genotype to phenotype 
strategy. Specifically, we will evaluate this hypothesis in Asians and Europeans, 
populations with rapidly growing incidence of hyperuricemia and gout that carry these 
genetic variants. To assess the effects of these variants on allopurinol response, we will 
measure serum uric acid and renal uric acid clearance as well as serum and urinary 
concentrations of allopurinol and its active metabolite, oxypurinol, to determine 
pharmacokinetic parameters. Other analyses may be done in the calculation of these 
parameters, such as serum creatinine and creatinine clearance.  
 
BACKGROUND: 
1. Gout is a major medical condition with a high prevalence in the U.S. and an 
increasing prevalence in global populations. Gout is an inflammatory arthritic disease 
caused by elevated serum uric acid (UA) levels. In humans, UA is the end-product of 
purine metabolism, and high UA levels are associated with gout and increased risk for 
cardiovascular events including stroke and myocardial infarction. Historically, because of 
protein-rich diets, gout was termed a “rich man’s disease” and confined to Western 
populations. However, with the global epidemic of metabolic syndrome, the prevalence 
of gout in other populations has increased dramatically1,2, e.g., certain populations in 
Asia, Polynesia and Africa have prevalence rates exceeding 10%3,4. 
2. Allopurinol (ALLO) is first line therapy for gout prevention with emerging uses 
in other diseases associated with metabolic syndrome. Discovered in the 1940s by 
researchers at Burroughs Wellcome ,including Nobel laureates Elion and Hitchings, 
ALLO has been first-line therapy for the treatment of hyperuricemia and gout for many 
years. ALLO is considered a pro-drug with weak pharmacologic effects; it is converted to 
an active metabolite, oxypurinol (OXY) via aldehyde oxidase, AOX1. OXY is a potent 
inhibitor of xanthine oxidase, the primary enzyme involved in the synthesis of UA. 
Though its major use is in the treatment of gout, ALLO is used for other diseases such as 
tumor lysis syndrome and kidney stones5-7, and is being repurposed for the treatment of 
diseases associated with metabolic syndrome, e.g., heart failure, stroke and cardiac 
shock8-11. Other xanthine oxidase inhibitors are also on the market or in clinical drug 
development. 
3. Recent data from our laboratory challenge the widely accepted mechanism of 
action of ALLO and OXY as its sole mechanism and suggest a major role for BCRP. 
In the first GWAS of ALLO response, we observed a single locus, in ABCG2, strongly 
associated with response to ALLO in 1300 patients of European ancestry. Previous 
genomewide association studies have shown ABCG2 and other UA transporters (e.g. 
GLUT9, URAT1) to be associated with high UA levels12-15. However, in our population 
of gout patients on ALLO, only ABCG2 (and not the other UA transporters) was 
significantly associated with drug response, i.e., ALLO-induced changes in UA levels. 
These results suggest that the gene may act on OXY directly. In fact, in exciting follow-
up studies in our laboratory, we found that both ALLO and OXY are excellent substrates 
of ABCG2 in HEK293 cells over-expressing the transporter suggesting that ABCG2 may 


function directly in the pharmacokinetics of ALLO/OXY. These results are consistent 
with other pharmacogenomic studies that show that genetic variants in membrane 
transporters that affect pharmacokinetics are associated with altered drug response (e.g., 
SLCO1B1 and methotrexate and statin toxicities, and ABCG2 and rosuvastatin 
response)16-19. Because of its location on the apical membranes of intestinal and renal 
epithelia, the data suggest that ABCG2 functions in reducing the intestinal absorption and 
enhancing the renal secretion of the drugs as it does for several other drugs20-22. 
4. A common reduced function variant of ABCG2, Q141K, paradoxically leads to 
reduced response to ALLO. The ABCG2 locus identified in our GWAS included the 
nonsynonymous variant, Q141K (rs2231142), which is known to have reduced transport 
function due to reduced protein expression23,24. This variant had the lowest p-value of all 
ABCG2 nonsynonymous variants that were either genotyped or imputed. Interestingly, 
Q141K has also been associated, at genomewide level significance, with response to 
rosuvastatin19. Because our preliminary studies indicate that ABCG2 is an efflux 
transporter for OXY and ALLO, we expected that the variant allele would be associated 
with higher drug levels through increased absorption and reduced renal secretion 
mechanisms resulting in higher drug levels and improved response. The fact that we 
found that the variant was associated with a worse response suggests another mechanism. 
We propose that Q141K results in lower drug levels in the renal tubule, consistent with 
reduced renal secretion of the drugs. The lower levels of OXY in the renal tubule result in 
reduced inhibition of UA reabsorptive transporters through competitive inhibition 
mechanisms. In support of our hypothesis, we recently found that OXY and ALLO 
inhibit UA uptake via URAT1. Our data are consistent with previous studies showing that 
OXY is a substrate of URAT125. Also, reviewing data in the older literature we noted 
that ALLO treatment was associated with an increased renal clearance of UA in many 
patients after initiation of ALLO treatment26, supporting our hypothesis that ALLO 
and/or OXY inhibit UA reabsorptive transporters. Therefore we propose that ABCG2-
mediated secretion of OXY results in sufficiently high levels of the drug in the renal 
tubule to competitively inhibit uric acid reabsorptive transporters, e.g., URAT1. 
Competitive inhibition of uric acid reabsorption leads to increased UA excretion and 
therefore reduced serum levels of UA. Notably, ABCG2 can concentrate its substrates 
>20-fold20,27 and maximum plasma unbound concentrations of OXY and ALLO are as 
high as 100 µM. Therefore, estimated proximal tubule concentrations of OXY and ALLO 
could be in the mM range28. In contrast, lower tubule levels of OXY, which are achieved 
in individuals with the Q141K variant, result in less inhibition of UA reabsorption and 
therefore, poorer response. If our hypotheses are proven, our studies will lead to a new 
understanding of the mechanisms of action of ALLO and its active metabolite OXY. 
5. There is a major gap in our understanding of the genetic factors that determine 
response to OXY in all ethnic groups including under-represented populations. In 
contrast to the multiple GWAS studies identifying genetic risk factors for hyperuricemia 
and gout, there has been no published GWAS related to ALLO response. In preliminary 
studies using samples from individuals of European ancestry, we conducted the first 
GWAS of response to ALLO. We attempt to fill this gap by evaluating the effects of 
ABCG2 gene variants on the pharmacodynamics and pharmacokinetics of ALLO. This 
effort is a mechanistic genotype to phenotype pharmacogenetic study designed to 
supplement ongoing investigations of the contribution of genetic variability in drug 


transporters to inter-individual variation in drug response. Comparing the variability in 
uric acid levels and renal clearance of ALLO in individuals with different ABCG2 
genotypes will provide information about the extent to which genetic factors contribute to 
inter-individual differences in the renal elimination and response to ALLO. Investigating 
these polymorphic variants, which occur at relatively high frequencies within Asians and 
Europeans, will provide vital information for the purpose of guiding antihyperuricemic 
drug regimens in these populations. 
 
NUMBER OF SUBJECTS: 15 subjects will participate in the study. 5 from each 
genotype: homozygous recessive, homozygous dominate, heterozygous. This number has 
been selected so that our study will be 80% powered to detect a 40% change in 
allopurinol clearance.  
 
GENDER OF SUBJECTS: Both males and females will be enrolled for this study. 
Because allopurinol has been shown to have detrimental effects on unborn fetuses, 
pregnant women will not be allowed to participate in the study. 
 
AGE OF SUBJECTS: Healthy volunteers over 18 years of age will be enrolled 
 
RACIAL AND ETHNIC ORIGIN: Subjects of Asian and European descent will be 
enrolled in this study. These ethnicities are known to carry the genetic variant of interest 
for this study.  
 
INCLUSION CRITERIA: 
1. Subject self-identify racial background; identify themselves, their parents, and their 4 
grandparents as European or Asian; 
2. Subjects are generally healthy with approved (normal) laboratory values for: 
• CBC Panel: WBC, RBC, Hemoglobin, Hematocrit, MCV, MCH, MCHC, RDW, 
Platelet Count, MPV and Differential (Absolute and Percent - Neutrophils, 
Lymphocytes, Monocytes, Eosinophils, and Basophils)  
• HFP: Total Protein, Albumin, Globulin (calculated), Albumin/Globulin Ratio 
(calculated), Total Bilirubin, Direct Bilirubin, Indirect Bilirubin (calculated), 
Alkaline Phosphatase, AST, ALT 
• RFP: Albumin, BUN/Creatinine Ratio (calculated), Calcium, Carbon Dioxide, 
Chloride, Creatinine, Estimated Glomerular Filtration Rate (calculated), Glucose, 
Phosphate (as Phosphorus), Potassium, Sodium, Urea Nitrogen 
• Uric Acid 
3. Subjects with the ABCG2 genotype, homozygous, heterozygous or homozygous for 
the major allele of rs2231142 will be recruited; 
4. Subjects >18 years of age and <40 years of age. 
 
EXCLUSION CRITERIA: 
1. Patients with vascular disease; 
2. Patients with renal impairment (GFR < 60 ml/min); 
3. Patients taking medications/supplements that affect uric acid levels; 
4. Pregnant or lactating women; 


5. Prior history of any allergic reaction to allopurinol/testing positive for HLA-B*5801 
allele; 
6. Risk of urinary or gastric retention or narrow-angle glaucoma (by medical history 
examination); 
7. Impaired hepatic function (> 1.5 times the upper limit of normal); 
8. Evidence of anemia (hemoglobin < 10 g); 
9. Evidence or diagnosis of congestive heart failure; 
10. Smokers; 
11. Subjects with a mutation other than rs2231142 in the ABCG2 genotype; 
12. Subjects taking hormonal contraceptives or other hormonal medications; 
13. Evidence of recreational drug use as determined by questionnaire; 
 
VULNERABLE SUBJECTS: Students may be enrolled in this study. This study poses 
little to no risks for healthy students. Only students over 18 will be enrolled and informed 
consent will be obtained. 
 
METHODS AND PROCEDURES: 
 
Questionnaire: 
 
Effect of the Q141K BCRP variant on the pharmacokinetics and pharmacodynamics of 
Allopurinol – This questionnaire will be used to assess the subject’s inclusion or 
exclusion from the study. The questionnaire will ask ancestry, health history, 
medications, habits, and family history. This questionnaire will help determine whether 
the subject is eligible to participate in the cheek swab portion of the screening.  
 
Procedures, Timeline, and Biological Measures: 
 
Screening Procedures: Part 1: Once eligible subjects have expressed their interest to 
participate and have had their questions answered by clinical personnel, the potential 
subject will be asked to email or call the research staff at Open Medicine Institute, Inc. If 
the eligible subject is already in the presence of the research staff they may proceed with 
the visit at the moment. During this visit the research staff will perform a screening 
evaluation for genotyping to determine eligibility. If email correspondence is preferred a 
secure network will be used for all correspondence between potential subjects and 
research staff.  
 
Subjects will be asked to come to Open Medicine Institute, Inc in Mountain View, 
California, or other site determined by Kathleen M. Giacomini, Ph. D (P.I.), to complete 
a screening visit. The initial screening will include a review of the volunteer's medical 
history and history of drug use to identify any status that would determine eligibility. If 
the information on the health, drug use and ethnicity questionnaires are accurate, this will 
suffice for the inclusion criteria. Changes in medical or drug use history will be noted 
during the study. The purpose of these questionnaires is to characterize health status and 
ethnicity as accurately as possible (see Inclusion/Exclusion Criteria). If the volunteer is 
determined to be eligible to participate after the questionnaire, a cheek swab to determine 


the subject's eligibility based on their ABCG2 genotype will be performed. 
 
Part 2: If the genotype group of the subject is not yet full of enrolled subjects, the subject 
will then be asked to return to Open Medicine Institute, Inc. and an additional 
venipuncture of 10 ml will be drawn to screen for liver and renal insufficiency within 2 
weeks of scheduled study visit. Subject's DNA will also be extracted (either from blood 
or additional cheek swab) to test for HLA*B 5801 allele, which is associated with 
allopurinol sensitivity. Female subjects' blood samples will be used for pregnancy testing 
(see Inclusion/Exclusion Criteria). Remaining saliva or blood sample may be stored for 
future genome-wide genotyping/sequencing.  
 
The subject may not exercise, drink alcohol, or drink coffee. Strenuous exercise can 
affect uric acid excretion and alcohol and coffee, both caffeinated and decaffeinated, have 
been shown to affect uric acid levels. 
 
Phase I procedures: Subjects will be asked to come to the clinic after 8 hours of fasting to 
provide a 10 ml blood sample to determine baseline serum uric acid and creatinine 
concentrations. Female subjects will be asked to provide a urine sample (20 mL) to 
ensure no change in pregnancy status. Subjects will be asked to empty their bladder and 
will then be given a placebo (microcrystalline cellulose). Blood samples will be collected 
at the following time points via an IV catheter: 0.5, 1, 1.5, 2, 3, 4, 5, 6, 8, 10, 24 hours. 
Subjects will be asked to collect all urine samples during their stay. The volume of the 
urine will be recorded and 20 ml of the sample stored at -20°C for analysis of uric acid 
and creatinine content. The remaining urine will then be stored with all other urine 
samples from the subject until the last time point. Subjects will be asked to return in the 
morning for their 24 hour blood draw and final urine sample. Subjects will be asked to 
turn in any urine from overnight. 20 ml of the 24-hour collective urine sample will be 
stored at -20°C for analysis of uric acid and creatinine. 
 
Phase II Procedures: Day 1: Subjects will be asked to come to the clinic after 8 hours of 
fasting to provide a 10 ml blood sample to determine baseline serum uric acid and 
creatinine concentrations. Female subjects will be asked to provide a urine sample (20 
mL) to ensure no change in pregnancy status. Subjects will be asked to empty their 
bladders and will then be given 300 mg of allopurinol. Blood samples collected at the 
following time points via an IV catheter: 0.5, 1, 1.5, 2, 3, 4, 5, 6, 8, 10, 24 hours. Subjects 
will be asked to collect all urine samples during their stay. The volume of the urine will 
be recorded and 20 ml of the sample stored at -20°C for analysis of uric acid, drug, and 
creatinine content. The remaining urine will then be stored with all other urine samples 
from the subject until the last time point. Subjects will be asked to return in the morning 
for their 24 hour blood draw and final urine sample. Subjects will be asked to turn in any 
urine from overnight. 20 ml of the 24-hour collective urine sample will be stored at -20°C 
for analysis of uric acid, drug, and creatinine.  
 
Days 2-5: The subjects will receive 4 more doses of allopurinol to be taken once a day 
until their next visit. Their final inpatient visit must be on Day 6. Adherence will be 
measured via phone calls and patient diaries. Patients will be asked to keep a log of their 


food and drinks.  
 
Day 6: Subjects will be asked to return to the clinic after 8 hours of fasting for their final 
dose of allopurinol. Subjects will be asked to provide a 10 ml blood sample to determine 
steady-state levels of drug, creatinine, and uric acid. Subjects will be asked to empty their 
bladder and they will then be given 300 mg allopurinol. Following their final dose, blood 
and urine samples will be collected following Phase II: Day 1 procedures. Following the 
24-hour time point, subjects will return at least two more times for blood and urine 
collection up to 72 hours post-allopurinol. In total, subjects will provide approximately 
400 ml of blood. This is less than a typical blood donation of 500 ml and is below 
minimal risk of 550 ml for healthy subjects. 
 
Visit 
Timing 
Collection 
Screening Part 1 
Anytime 
Cheek Swab: ABCG2 genotype 
Questionnaire 
Screening Part 2 
Within two weeks of Placebo visit 
Blood: CBC, HFP, RFP, UA 
If female - Pregnancy 
Cheek Swab: HLA*B 5801 
Phase 1: Placebo 
 
Before – Fast 8 hours 
clinic visit 
Blood: Baseline- Creatinine, UA 
During at 0.5, 1,1.5, 2, 3, 4, 5, 6, 8, 
10 hours (Creatinine, UA, 
ALLO/OXY) 
Urine: Before – females 20 ml for 
pregnancy test. 
During- all urine during stay 
collected 
Phase 1: Placebo 
24 hours after placebo dose 
clinic visit 
 
Blood: Creatinine, UA, 
ALLO/OXY 
Urine: Turn in what was collected 
overnight 
Phase 2: Allopurinol 
– Oral 300 mg dose 
 
 
Before – Fast 8 hours 
Day 1—clinic visit 
 
*Must be started within 2 weeks of 
Day 0 (Placebo Phase) 
Blood: Baseline- Creatinine, UA 
During at 0.5, 1,1.5, 2, 3, 4, 5, 6, 8, 
10 hours (Creatinine, UA, 
ALLO/OXY_) 
Urine: Before – females 20 ml for 
pregnancy test. 
During- all urine during stay 
collected 
Phase 2: Allopurinol 
– Oral 300 mg tablet 
 
24 hours after first dose 
Day 2—clinic visit 
Blood: Creatinine, UA, 
ALLO/OXY 
Urine: Turn in what was collected 
overnight – pregnancy test 
Patient given diaries and tablets to 
take home for Days 3-6 
Phase 2: Allopurinol 
– Oral 300 mg tablet 
48 hours after first dose 
Day 3 – at home 
Patient Diary and phone follow-ups 


 
Phase 2: Allopurinol 
– Oral 300 mg tablet 
72 hours after first dose 
Day 4 – at home 
Patient Diary and phone follow-ups 
Phase 2: Allopurinol 
– Oral 300 mg tablet 
96 hours after first dose 
Day 5 – at home 
Patient Diary and phone follow-ups 
Phase 2: Allopurinol 
Final Oral 300 mg 
tablet 
120 hours after first dose 
Before – Fast 8 hours 
Day 6 – clinic visit 
Blood: Baseline- Creatinine, UA, 
ALLO/OXY 
During at 0.5, 1,1.5, 2, 3, 4, 5, 6, 8, 
10 hours (Creatinine, UA, 
ALLO/OXY) 
Urine: Before – females 20 ml for 
pregnancy test. 
During- all urine during stay 
collected  
Phase 2: Allopurinol 
– Follow-up 
24 hours after final dose 
Day 7 – clinic visit 
Blood: Creatinine, UA, 
ALLO/OXY 
Urine: Turn in urine that was 
collected overnight 
 
Phase 2: Allopurinol 
– Follow-up 
Day 8 – clinic visit 
Blood: Creatinine, UA, 
ALLO/OXY 
 
Phase 2: Allopurinol 
– Follow-up 
As close as possible to 72 hours after 
injection 
Day 9 – clinic visit 
Blood:, Creatinine, UA, 
ALLO/OXY 
 
 
ANALYTICAL METHODS: Measurement of allopurinol and oxypurinol will be 
performed by high performance liquid chromatography and mass spectrometry (LC-
MS/MS). Measurement of uric acid and its precursors, xanthine and hypoxanthine, will 
be measured via Amplex Red and Abcam colorimetric assays. 
 
Genotyping: All recruited subjects will sign informed consents and HIPAA forms. At 
screening, a blood sample (10 mL) or saliva sample will be obtained for DNA extraction. 
Plasma will be stored for assays of endogenous compounds and drugs. The ABCG2 
variants will be genotyped at UCSF by a Taq Man Assay using primers and probes from 
ABI. The reaction will be run on an ABI 7900 HT. 
 
HLA genotyping and wellness panels will be analyzed by qualified personnel at OMI or 
other reputable company as approved by Kathy Giacomini or other study personnel. 
 
STATISTICAL ANALYSIS: 
Mechanistic pharmacokinetic modeling will be conducted to characterize the 
pharmacokinetics of allopurinol (ALLO) and oxypurinol (OXY). The mechanistic model 
will characterize details relevant for this study such as conversion mechanism from 
ALLO to OXY and estimation of fraction of metabolized drug; estimation of ALLO and 
OXY renal and secretory clearance; possible concentration-dependent saturation 


mechanism of secretory clearance and consequent longitudinal changes in OXY secretory 
CL and influence of pharmacogenomics (ABCG2) and demographic covariates 
(creatinine clearance, age, gender, race and body weight) on secretory and renal CL. By 
use of the physiological model, we will also attempt to estimate the drug concentration in 
the renal tubule; by knowing plasma and urinary concentration of OXY, and by 
estimating filtration and secretory clearance of OXY, we will be able to estimate the time 
profile of tubular concentrations of OXY. These tubular OXY concentrations represent 
concentration at the proposed site of action and they will further be linked with the 
pharmacodynamic response. A model describing concentration dependent inhibition of 
UA re-absorption from the renal tubule by OXY will be implemented. We will examine 
linear and non-linear (Emax type of model) relationships between tubular OXY and 
inhibition of UA reabsorption. In the pharmacokinetic modeling, we expect to see a 
significant influence of creatinine clearance on renal clearance and a significant influence 
of ABCG2 genotype on secretory CL and oral bioavailability, as described below. In the 
pharmacokinetic-pharmacodynamic analysis we expect that individuals with higher 
tubular concentrations of OXY will show the better response in terms of plasma UA 
lowering resulting in higher urine UA concentrations. Consequently, higher tubular 
concentrations will be mostly expected in individuals with high secretory clearance, e.g. 
individuals with ABCG2 141Q/Q, however the modeling may also account for other 
factors contributing to high tubular concentration, such as higher filtration rates and other 
factors, therefore providing more granularity and better mechanistic understanding in all 
processes leading to overall UA physiology. Although this specific PKPD analysis will 
be focused on the mechanism and role of ABCG2 in UA response to ALLO and OXY, 
the established mechanistic framework can be used to test the role of potential new 
pharmacogenetic variants. Longitudinal PKPD modeling using nonlinear mixed effects is 
a method carrying the most power to separate the signal from the noise; therefore 
modeling analysis will be associated with at least 80% power to detect differences in 
primary endpoints (net secretory clearance and change in UA) as described above. In 
addition to confirming the result of the primary analysis, this analysis will establish a 
mechanistic representation of the physiology underlying UA metabolism, therefore 
providing strong support and evidence to our mechanistic hypothesis. Pharmacokinetic 
parameters including renal clearance, and net reabsorptive or secretory clearance of UA, 
OXY and ALLO will be calculated from serum and plasma drug levels by standard 
pharmacokinetic methods. To assess net secretory or reabsorptive clearance of the three 
compounds, we will subtract fu*GFR (where fu is the unbound fraction of ALLO, OXY 
or UA and GFR is the glomerular filtration rate), as measured or estimated from serum 
and urine creatinine values, from renal clearance of ALLO, OXY and UA, directly 
measured from plasma and urine values. Previously we have used these methods to 
calculate net secretion (or reabsorption) of several drugs. For pharmacodynamics, 
differences in DSUA between the genotype groups will be evaluated first with an 
analysis of variance. If DSUA is not normally distributed, we will evaluate the 
differences between the genotype groups using a non-parametric Kruskal-Wallis test. We 
expect individuals with ABCG2 141K/K and 141Q/K will have a reduced response to 
ALLO (smaller DSUA) than individuals with ABCG2 141Q/Q. In addition, individuals 
with ABCG2 141K/K will have reduced renal clearance of ALLO and OXY in 
comparison to other genotypes since we propose that ABCG2 functions as a secretory 


transporter for ALLO/OXY in the kidney. We will also determine whether the ABCG2 
141K carriers have a reduced renal clearance. 
 
DATA STORAGE AND CONFIDENTIALITY:  
All personal and medical data will be considered confidential. Subjects will be assigned a 
unique number that will code for all personal data collected. This code will be used for all 
stored samples. All electronic study data will be stored on a password-protected computer 
on a secure network in the PI's laboratory, and backup files of the data will be stored on a 
hard disk which will be kept in a locked file cabinet or office. 
 
Participation in research may involve a loss of privacy, but information about subjects 
will be handled as confidentially as possible and in compliance with HIPAA regulations. 
All information gathered in the study will be used collectively with information from 
other participants. All personal and medical data will be considered confidential. Upon 
the subjects enrollment in the study the researchers will assign them a number that will 
code for all of the personal data collected. Dr. Giacomini, her research associates, and 
team members will have access to information about each subject. Research staff, UCSF 
Committee on Human Research, and other University of California personnel may also 
review or receive information about the subject for quality assurance purposes (i.e. to 
review an adverse drug response). The subject's names or any identifying information 
will not be used in any published reports about this study, and data will be submitted and 
reported in consolidated, coded form. Furthermore, coded data obtained in the study will 
be deposited into the Pharmacogenetics Research Network Database (PharmGKB) on the 
World Wide Web. Researchers for further studies may access information on this 
database. However, the database will NOT include any information pertaining to the 
identity of the samples. Information identifying the subjects will not be available on the 
database. 
 
For subjects enrolled at Open Medicine Insitute (OMI), access to patients’ records will be 
limited to access by the study and clinic staff.  OMI and site staff will go over the 
questionnaire with the volunteer to determine initial inclusion/exclusion.  Each 
participating patient will be assigned a unique identification number.  All data collected 
on each subject will be labeled with the corresponding study ID number. Patients will be 
enrolled in the study by providing signed written informed consent or by completing the 
existing web-based consent process system 
 
Confidentiality of all records and materials will be maintained to the fullest extent 
possible.  Written informed consent forms will be kept in locked files at participating 
sites or, if completed online, password protected. 
 
All data generated by OMI will be stored on OMI's state of the art data center in Fremont, 
CA. This center runs a VMware distributed architecture with a large EMC2 VNX data 
store with their own proprietary OpenMedNet layer. This ensures data integrity and 
security. 
 
RISK CATEGORY: Minimal 


 
POTENTIAL RISKS:  
 
1. Allopurinol: Allopurinol is a xanthine oxidase inhibitor used to treat gout and 
hyperuricemia that can be dosed from 100 mg to 800 mg daily. The most frequent 
side effects are skin rash (1.5%), diarrhea (1.3%), nausea (1.2%), and acute gout 
attacks. Furthermore, these side effects occur in less than 1% of those patients 
dosed and at a dose of 300 mg/day, we do not expect to see any discomfort in the 
volunteers. 
a. Common but less serious: 
• Diarrhea 
• Drowsiness 
• Headache 
• Change in taste 
• Muscle pain 
b. Rare but serious 
• Renal Failure 
• Stevens-Johnson Syndrome* 
 
* Although some patients develop Stevens-Johnson syndrome on allopurinol, subject’s 
side effects will be monitored closely and dosing will be discontinued with any signs of 
skin rash. 
 
2. Blood loss and Venipuncture: Withdrawal of blood from a vein may cause pain, 
bruising, and in rare cases infection at the site of needle puncture. Some people 
may faint when blood is drawn. All subjects will provide approximately 400 mL 
of blood. Loss of 400 mL of blood is not harmful and will be replenished by the 
body within 2-3 weeks. 
 
3. Questionnaire: Some of the questions may make the participants feel 
uncomfortable. All participants will be informed that they have the right to 
decline to answer any questions. 
 
4. Genetic Testing: Participation in this study may involve a loss of privacy. All 
DNA testing will be performed using blood/saliva samples tagged with a unique 
identification number, to minimize the risk of loss of confidentiality. Only the 
research staff will have access to the samples and records and the key to the code. 
Information from this study used for scientific publication will not contain any 
identifying information. Subjects may withdraw consent for DNA storage at any 
time during or after the study by writing a letter to REDACTED 
5. Inconvenience of Study Visit: Subject may experience an inconvenience by 
having to attend multiple study visits while study procedures and questionnaire 
data are being collected. 
 
PROTECTION AGAINST RISKS: 
To minimize the risks/discomforts to subjects, subjects will be excluded from 


participation if: 
1. Under 18 years old 
2. Patients with vascular disease 
3. Patients with renal impairment (GFR < 60 ml/min) 
4. Patients taking medications that affect uric acid levels 
5. Pregnant or lactating women 
6. Prior history of any allergic reaction to allopurinol 
7. Risk of urinary or gastric retention or narrow-angle glaucoma (by medical history 
examination) 
8. Impaired hepatic function (> 1.5 times the upper limit of normal) 
9. Evidence of anemia (hemoglobin < 10 g) 
10. Evidence or diagnosis of congestive heart failure 
11. Evidence of recreational drug use 
 
To further minimize risks, OMI is equipped with emergency medical tools such as ACLS 
trained personnel, crash carts, and emergency drugs and supplies to stabilize any subject 
until emergency personnel arrive after 911 is called. El Camino Hospital is directly across 
the street in case of emergencies as well. 
 
BENEFITS TO SUBJECTS: 
The benefit of participating in this studies is that you are able to add to our knowledge 
about how the pharmacokinetic and pharmacodynamic mechanisms by which Q141K 
(rs2231142) associates with uric acid lowering in gout patients on allopurinol with 
relatively little inconvenience to you. While these types of studies are unlikely to offer 
direct benefit to your disease at the time of your involvement, the information gained 
may be used to develop new therapies to improve survival or quality of life for you or 
other patients like you in the future. 
 
ALTERNATIVES TO PARTICIPATION: 
The subject can choose not to participate in this study.  
 
SUBJECT IDENTIFICATION AND RECRUITMENT:  
 
Subjects will be recruited from Asian and European populations within reasonable 
driving distance of Open Medicine Institution (OMI), with the initial focus on Asian 
populations. OMI will also reach out to locations where Asian and European descent 
populations are prevalent to request permission to hand out or hang an IRB approved 
flyer. We will also post a booth with the flyer at other local sites. Interested individuals 
may contact OMI directly through the information on the flyer to begin the screening 
process. If research personnel are available in person the interested individual may also 
talk directly to the research personnel at that time in a private location. 
 
PROCESS OF CONSENT: 
 
At OMI, all research intervention will be conducted in a private room to maintain 
privacy. All subjects are required to sign an Informed Consent Form prior to participation 


in compliance with 21CFR50 Subpart B.  Open Medicine Institute (OMI) will receive 
approval of the Informed Consent Form from the Western Institutional Review Board 
(WIRB) as well as continuing review approval, as necessary, in compliance with 
21CFR812 Subpart D. 
All subjects must be provided a copy of the Informed Consent and the California 
Experimental Bill of Rights but the bill of rights and consent may be done either using 
the electronic informed consent on https://www.OpenMedNet.org or using written 
informed consent.  If the subject signs the electronic consent form this form is always 
available for view by the subject, or the subject may print it out, so a signed written copy 
of the informed consent is not necessary.  If the subject signs a written copy the subject 
must receive a copy of the signed and dated consent document.  The original copy of the 
consent form will be retained at OMI in a secure, locked location.  Each subject may be 
enrolled into the study only once. 
Subjects must be explained the purpose of the study in terms understandable to them with 
sufficient information for them to make an informed decision regarding their 
participation.  They must be informed and understand that participation is always 
voluntary and that participation in the study will not impact the clinical care that they 
currently receive.  The consent form will specify whom to contact if there are injuries as 
a result of the study and whom to contact with general questions about the study.  The 
consent form will notify subjects that blood, urine and DNA samples will be collected 
and will be used for research purposes and stored for the purpose of conducting future 
research by UCSF, including genetic DNA sequencing. 
 
SUBJECT CAPACITY:  
Subjects will only be enrolled if they are able to give informed consent themselves.  
 
SUBJECT COMPREHENSION:  
Informed consent will be obtained using the form attached to this protocol. Subjects will 
be briefed on all procedures in the trial in plain speech and will have the opportunity to 
have all of their questions answered. Subjects will indicate that they understand the risks 
and benefits and agree to participate in the study by signing the informed consent form.  
 
DOCUMENTATION OF CONSENT:  
Signed consent forms will be retained on a secure server along with the subject 
information as previously described.  
 
COSTS TO THE SUBJECT:  
There will be no cost to the subject other than the time it takes to participate in this study 
(approximately 36.5 hours).  
 
PAYMENT FOR PARTICIPATION:  
If subjects agree to be screened, they will receive a $10 payment for the initial screening 
visit (cheek swab). If they qualify and decide to continue onto the secondary screening, 
they will receive an additional $15. If they are enrolled into the remaining phases of the 


study, they will be eligible to receive an additional $575 payment to offset their time and 
inconvenience. Only if they complete everything and comply with all aspects of the study 
will they be paid the entire amount of $575 (in addition to the $25 received for the 
screening for a total of $600). If they fail to comply any portion of the study visit or do 
not complete the entire visit then they will only receive a prorated payment based on the 
time of participation (approximately $10.0 /per hour). Payments will be paid in the form 
of a check or debit card at the completion of the study, which they should receive four to 
six weeks after the last visit. Screening visit may be paid in cash or check. Subjects must 
provide the researchers their Social Security number and mailing address so the check or 
debit card can be processed. Transportation and Parking fees may be provided if 
requested in advance. According to law, subjects will have to pay taxes on payment for 
research participation in excess of $600 per calendar year. 
 
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4. Portis, A.J. et al. High prevalence of gouty arthritis among the Hmong population in 
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Arthritis Care Res (Hoboken) 62, 1386-91 (2010). 
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cause, 
consequence, or just a not so innocent bystander? Endocrine 41, 350-2 (2012). 
11. Gotsman, I., Keren, A., Lotan, C. & Zwas, D.R. Changes in uric acid levels and 
allopurinol use in 
chronic heart failure: association with improved survival. J Card Fail 18, 694-701 (2012). 
12. Kottgen, A. et al. Genome-wide association analyses identify 18 new loci associated 
with serum 
urate concentrations. Nat Genet 45, 145-54 (2013). 
13. Karns, R. et al. Genome-wide association of serum uric acid concentration: 
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14. Yang, Q. et al. Multiple genetic loci influence serum urate levels and their 
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cardiovascular disease risk factors. Circ Cardiovasc Genet 3, 523-30 (2010). 
15. Dehghan, A. et al. Association of three genetic loci with uric acid concentration and 
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genome-wide association study. Lancet 372, 1953-61 (2008). 
16. Chu, A.Y. et al. Genome-wide association study evaluating lipoprotein-associated 
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17. Trevino, L.R. et al. Germline genetic variation in an organic anion transporter 
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18. Link, E. et al. SLCO1B1 variants and statin-induced myopathy--a genomewide study. 
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19. Chasman, D.I. et al. Genetic determinants of statin-induced low-density lipoprotein 
cholesterol 
reduction: the Justification for the Use of Statins in Prevention: an Intervention Trial 
Evaluating Rosuvastatin 
(JUPITER) trial. Circ Cardiovasc Genet 5, 257-64 (2012). 
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21. Mizuno, N. et al. Evaluation of the role of breast cancer resistance protein 
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multidrug resistance-associated protein 4 (MRP4/ABCC4) in the urinary excretion of 
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metabolites of edaravone (MCI-186; 3-methyl-1-phenyl-2-pyrazolin-5-one). Drug Metab 
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(2007). 
22. Mizuno, N. et al. Impaired renal excretion of 6-hydroxy-5,7-dimethyl-2-
methylamino-4-(3- 
pyridylmethyl) benzothiazole (E3040) sulfate in breast cancer resistance protein 


(BCRP1/ABCG2) knockout 
mice. Drug Metab Dispos 32, 898-901 (2004). 
23. Woodward, O.M. et al. Gout-causing Q141K mutation in ABCG2 leads to instability 
of the nucleotidebinding 
domain and can be corrected with small molecules. Proc Natl Acad Sci U S A 110, 5223-
8 (2013). 
24. Furukawa, T. et al. Major SNP (Q141K) variant of human ABC transporter ABCG2 
undergoes 
lysosomal and proteasomal degradations. Pharm Res 26, 469-79 (2009). 
25. Iwanaga, T., Kobayashi, D., Hirayama, M., Maeda, T. & Tamai, I. Involvement of 
uric acid transporter 
in increased renal clearance of the xanthine oxidase inhibitor oxypurinol induced by a 
uricosuric agent, 
benzbromarone. Drug Metab Dispos 33, 1791-5 (2005). 
26. Yue, T.F. & Gutman, A.B. Effect of Allopurinol (4-Hydroxypyrazolo-(3,4-
D)Pyrimidine) on Serum and 
Urinary Uric Acid in Primary and Secondary Gout. Am J Med 37, 885-98 (1964). 
27. Vlaming, M.L. et al. Abcc2 (Mrp2), Abcc3 (Mrp3), and Abcg2 (Bcrp1) are the main 
determinants for 
rapid elimination of methotrexate and its toxic metabolite 7-hydroxymethotrexate in vivo. 
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3350-9 (2009). 
28. Turnheim, K., Krivanek, P. & Oberbauer, R. Pharmacokinetics and 
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APPENDICES: 
 
Appendix 1. Allopurinol Informed Consent Form 
 
Appendix 2. Questionnaire: Effect of the Q141K BCRP variant on the pharmacokinetics 
and pharmacodynamics of allopurinol. 
 
Appendix 3. Subject Diary 
 
Appendix 4. Allopurinol Subject Recruitment Flyer 
 
Appendix 5. Allopuinol Insert 
 
Apendix 6.  Allopurinol AGC-9588 
 

Prot_SAP_000.pdf: 
 
The Effects of BCRP Q141K on Allopurinol Pharmacokinetics and 
Dynamics 
NCT02956278 
November 7, 2017 


 
 
 
PURPOSE: To investigate the potential association of polymorphic genetic variants 
ABCG2 with altered response to allopurinol, we will employ a genotype to phenotype 
strategy. Specifically, we will evaluate this hypothesis in Asians and Europeans, 
populations with rapidly growing incidence of hyperuricemia and gout that carry these 
genetic variants. To assess the effects of these variants on allopurinol response, we will 
measure serum uric acid and renal uric acid clearance as well as serum and urinary 
concentrations of allopurinol and its active metabolite, oxypurinol, to determine 
pharmacokinetic parameters. Other analyses may be done in the calculation of these 
parameters, such as serum creatinine and creatinine clearance.  
 
BACKGROUND: 
1. Gout is a major medical condition with a high prevalence in the U.S. and an 
increasing prevalence in global populations. Gout is an inflammatory arthritic disease 
caused by elevated serum uric acid (UA) levels. In humans, UA is the end-product of 
purine metabolism, and high UA levels are associated with gout and increased risk for 
cardiovascular events including stroke and myocardial infarction. Historically, because of 
protein-rich diets, gout was termed a “rich man’s disease” and confined to Western 
populations. However, with the global epidemic of metabolic syndrome, the prevalence 
of gout in other populations has increased dramatically1,2, e.g., certain populations in 
Asia, Polynesia and Africa have prevalence rates exceeding 10%3,4. 
2. Allopurinol (ALLO) is first line therapy for gout prevention with emerging uses 
in other diseases associated with metabolic syndrome. Discovered in the 1940s by 
researchers at Burroughs Wellcome ,including Nobel laureates Elion and Hitchings, 
ALLO has been first-line therapy for the treatment of hyperuricemia and gout for many 
years. ALLO is considered a pro-drug with weak pharmacologic effects; it is converted to 
an active metabolite, oxypurinol (OXY) via aldehyde oxidase, AOX1. OXY is a potent 
inhibitor of xanthine oxidase, the primary enzyme involved in the synthesis of UA. 
Though its major use is in the treatment of gout, ALLO is used for other diseases such as 
tumor lysis syndrome and kidney stones5-7, and is being repurposed for the treatment of 
diseases associated with metabolic syndrome, e.g., heart failure, stroke and cardiac 
shock8-11. Other xanthine oxidase inhibitors are also on the market or in clinical drug 
development. 
3. Recent data from our laboratory challenge the widely accepted mechanism of 
action of ALLO and OXY as its sole mechanism and suggest a major role for BCRP. 
In the first GWAS of ALLO response, we observed a single locus, in ABCG2, strongly 
associated with response to ALLO in 1300 patients of European ancestry. Previous 
genomewide association studies have shown ABCG2 and other UA transporters (e.g. 
GLUT9, URAT1) to be associated with high UA levels12-15. However, in our population 
of gout patients on ALLO, only ABCG2 (and not the other UA transporters) was 
significantly associated with drug response, i.e., ALLO-induced changes in UA levels. 
These results suggest that the gene may act on OXY directly. In fact, in exciting follow-
up studies in our laboratory, we found that both ALLO and OXY are excellent substrates 
of ABCG2 in HEK293 cells over-expressing the transporter suggesting that ABCG2 may 


function directly in the pharmacokinetics of ALLO/OXY. These results are consistent 
with other pharmacogenomic studies that show that genetic variants in membrane 
transporters that affect pharmacokinetics are associated with altered drug response (e.g., 
SLCO1B1 and methotrexate and statin toxicities, and ABCG2 and rosuvastatin 
response)16-19. Because of its location on the apical membranes of intestinal and renal 
epithelia, the data suggest that ABCG2 functions in reducing the intestinal absorption and 
enhancing the renal secretion of the drugs as it does for several other drugs20-22. 
4. A common reduced function variant of ABCG2, Q141K, paradoxically leads to 
reduced response to ALLO. The ABCG2 locus identified in our GWAS included the 
nonsynonymous variant, Q141K (rs2231142), which is known to have reduced transport 
function due to reduced protein expression23,24. This variant had the lowest p-value of all 
ABCG2 nonsynonymous variants that were either genotyped or imputed. Interestingly, 
Q141K has also been associated, at genomewide level significance, with response to 
rosuvastatin19. Because our preliminary studies indicate that ABCG2 is an efflux 
transporter for OXY and ALLO, we expected that the variant allele would be associated 
with higher drug levels through increased absorption and reduced renal secretion 
mechanisms resulting in higher drug levels and improved response. The fact that we 
found that the variant was associated with a worse response suggests another mechanism. 
We propose that Q141K results in lower drug levels in the renal tubule, consistent with 
reduced renal secretion of the drugs. The lower levels of OXY in the renal tubule result in 
reduced inhibition of UA reabsorptive transporters through competitive inhibition 
mechanisms. In support of our hypothesis, we recently found that OXY and ALLO 
inhibit UA uptake via URAT1. Our data are consistent with previous studies showing that 
OXY is a substrate of URAT125. Also, reviewing data in the older literature we noted 
that ALLO treatment was associated with an increased renal clearance of UA in many 
patients after initiation of ALLO treatment26, supporting our hypothesis that ALLO 
and/or OXY inhibit UA reabsorptive transporters. Therefore we propose that ABCG2-
mediated secretion of OXY results in sufficiently high levels of the drug in the renal 
tubule to competitively inhibit uric acid reabsorptive transporters, e.g., URAT1. 
Competitive inhibition of uric acid reabsorption leads to increased UA excretion and 
therefore reduced serum levels of UA. Notably, ABCG2 can concentrate its substrates 
>20-fold20,27 and maximum plasma unbound concentrations of OXY and ALLO are as 
high as 100 µM. Therefore, estimated proximal tubule concentrations of OXY and ALLO 
could be in the mM range28. In contrast, lower tubule levels of OXY, which are achieved 
in individuals with the Q141K variant, result in less inhibition of UA reabsorption and 
therefore, poorer response. If our hypotheses are proven, our studies will lead to a new 
understanding of the mechanisms of action of ALLO and its active metabolite OXY. 
5. There is a major gap in our understanding of the genetic factors that determine 
response to OXY in all ethnic groups including under-represented populations. In 
contrast to the multiple GWAS studies identifying genetic risk factors for hyperuricemia 
and gout, there has been no published GWAS related to ALLO response. In preliminary 
studies using samples from individuals of European ancestry, we conducted the first 
GWAS of response to ALLO. We attempt to fill this gap by evaluating the effects of 
ABCG2 gene variants on the pharmacodynamics and pharmacokinetics of ALLO. This 
effort is a mechanistic genotype to phenotype pharmacogenetic study designed to 
supplement ongoing investigations of the contribution of genetic variability in drug 


transporters to inter-individual variation in drug response. Comparing the variability in 
uric acid levels and renal clearance of ALLO in individuals with different ABCG2 
genotypes will provide information about the extent to which genetic factors contribute to 
inter-individual differences in the renal elimination and response to ALLO. Investigating 
these polymorphic variants, which occur at relatively high frequencies within Asians and 
Europeans, will provide vital information for the purpose of guiding antihyperuricemic 
drug regimens in these populations. 
 
NUMBER OF SUBJECTS: Up to 27 subjects will participate in the study. Subjects will 
be enrolled in a 1:1:1 ratio of reference BCRP, heterozygous for the BCRP Q141K 
variant, and homozygous for the BCRP Q141K variant. This number has been selected so 
that our study will be 80% powered to detect a 30% change in allopurinol clearance.  
 
GENDER OF SUBJECTS: Both males and females will be enrolled for this study. 
Because allopurinol has been shown to have detrimental effects on unborn fetuses, 
pregnant women will not be allowed to participate in the study. 
 
AGE OF SUBJECTS: Healthy volunteers over 18 years of age will be enrolled.  
 
RACIAL AND ETHNIC ORIGIN: Subjects of Asian and European descent will be 
enrolled in this study. These ethnicities are known to carry the genetic variant of interest 
for this study.  
 
INCLUSION CRITERIA: 
1. Subject self-identify racial background; identify themselves, their parents, and their 4 
grandparents as European or Asian; 
2. Subjects are generally healthy with approved (normal) laboratory values for: 
• CBC Panel: WBC, RBC, Hemoglobin, Hematocrit, MCV, MCH, MCHC, RDW, 
Platelet Count, MPV and Differential (Absolute and Percent - Neutrophils, 
Lymphocytes, Monocytes, Eosinophils, and Basophils)  
• HFP: Total Protein, Albumin, Globulin (calculated), Albumin/Globulin Ratio 
(calculated), Total Bilirubin, Direct Bilirubin, Indirect Bilirubin (calculated), 
Alkaline Phosphatase, AST, ALT 
• RFP: Albumin, BUN/Creatinine Ratio (calculated), Calcium, Carbon Dioxide, 
Chloride, Creatinine, Estimated Glomerular Filtration Rate (calculated), Glucose, 
Phosphate (as Phosphorus), Potassium, Sodium, Urea Nitrogen 
• Uric Acid 
3. Subjects with the ABCG2 genotype, homozygous, heterozygous or homozygous for 
the major allele of rs2231142 will be recruited; 
4. Subjects >18 years of age and <50 years of age. 
 
EXCLUSION CRITERIA: 
1. Patients with vascular disease; 
2. Patients with renal impairment (GFR < 60 ml/min); 
3. Patients taking medications/supplements that affect uric acid levels; 
4. Pregnant or lactating women; 


5. Prior history of any allergic reaction to allopurinol/testing positive for HLA-B*5801 
allele; 
6. Risk of urinary or gastric retention or narrow-angle glaucoma (by medical history 
examination); 
7. Impaired hepatic function (> 1.5 times the upper limit of normal); 
8. Evidence of anemia (hemoglobin < 10 g); 
9. Evidence or diagnosis of congestive heart failure; 
10. Smokers; 
11. Subjects with a mutation other than rs2231142 in the ABCG2 genotype; 
12. Subjects taking hormonal contraceptives or other hormonal medications; 
13. Evidence of recreational drug use as determined by questionnaire; 
 
VULNERABLE SUBJECTS: Students may be enrolled in this study. This study poses 
little to no risks for healthy students. Only students over 18 will be enrolled and informed 
consent will be obtained. 
 
METHODS AND PROCEDURES: 
 
Questionnaire: 
 
Effect of the Q141K BCRP variant on the pharmacokinetics and pharmacodynamics of 
Allopurinol – This questionnaire will be used to assess the subject’s inclusion or 
exclusion from the study. The questionnaire will ask ancestry, health history, 
medications, habits, and family history. This questionnaire will help determine whether 
the subject is eligible to participate in the cheek swab portion of the screening.  
 
Procedures, Timeline, and Biological Measures: 
 
Screening Procedures: Part 1: Once eligible subjects have expressed their interest to 
participate and have had their questions answered by clinical personnel, the potential 
subject will be asked to email or call the research staff at Open Medicine Institute, Inc. If 
the eligible subject is already in the presence of the research staff they may proceed with 
the visit at the moment. During this visit the research staff will perform a screening 
evaluation for genotyping to determine eligibility. If email correspondence is preferred a 
secure network will be used for all correspondence between potential subjects and 
research staff.  
 
Subjects will be asked to come to Open Medicine Institute, Inc in Mountain View, 
California, or other site determined by Kathleen M. Giacomini, Ph. D (P.I.), to complete 
a screening visit. The initial screening will include a review of the volunteer's medical 
history and history of drug use to identify any status that would determine eligibility. If 
the information on the health, drug use and ethnicity questionnaires are accurate, this will 
suffice for the inclusion criteria. Changes in medical or drug use history will be noted 
during the study. The purpose of these questionnaires is to characterize health status and 
ethnicity as accurately as possible (see Inclusion/Exclusion Criteria). If the volunteer is 
determined to be eligible to participate after the questionnaire, a cheek swab to determine 


the subject's eligibility based on their ABCG2 genotype will be performed. 
 
Part 2: If the genotype group of the subject is not yet full of enrolled subjects, the subject 
will then be asked to return to Open Medicine Institute, Inc. and an additional 
venipuncture of 20 ml will be drawn to screen for liver and renal insufficiency within 2 
weeks of scheduled study visit. Subject's DNA will also be extracted (either from blood 
or additional cheek swab) to test for HLA*B 5801 allele, which is associated with 
allopurinol sensitivity. Female subjects' blood samples will be used for pregnancy testing 
(see Inclusion/Exclusion Criteria). Remaining saliva or blood sample may be stored for 
future genome-wide genotyping/sequencing.  
 
The subject may not exercise, drink alcohol, or drink coffee. Strenuous exercise can 
affect uric acid excretion and alcohol and coffee, both caffeinated and decaffeinated, have 
been shown to affect uric acid levels. 
 
Study Day Procedures: Day 1: Subjects will be asked to come to the clinic after 8 hours 
of fasting to provide a 10 ml blood sample to determine baseline serum uric acid and 
creatinine concentrations. Female subjects will be asked to provide a urine sample (20 
mL) to ensure no change in pregnancy status. Subjects will be asked to empty their 
bladders and will then be given 300 mg of allopurinol. Blood samples collected at the 
following time points via an IV catheter: 0.5, 1, 1.5, 2, 3, 4, 5, 6, 8, 10, 24 hours. Subjects 
will be asked to collect all urine samples during their stay. The volume of the urine will 
be recorded and 20 ml of the sample stored at -20°C for analysis of uric acid, drug, and 
creatinine content. The remaining urine will then be stored with all other urine samples 
from the subject until the last time point. Subjects will be asked to return in the morning 
for their 24 hour blood draw and final urine sample. Subjects will be asked to turn in any 
urine from overnight. 20 ml of the 24-hour collective urine sample will be stored at -20°C 
for analysis of uric acid, drug, and creatinine.  
 
Day 3-4: Following the 24-hour time point, subjects will return at least two more times 
for blood and urine collection up to 72 hours post-allopurinol. In total, subjects will 
provide approximately 160 ml of blood. This is less than a typical blood donation of 
500 ml and is below minimal risk of 550 ml for healthy subjects. 
 
Visit 
Timing 
Collection 
Screening Part 1 
Anytime 
Cheek Swab: ABCG2 genotype 
Questionnaire 
Screening Part 2 
Within two weeks of clinic visit 
Blood: CBC, HFP, RFP, UA 
If female - Pregnancy 
Cheek Swab: HLA*B 5801 
Allopurinol – Oral 
300 mg dose 
 
 
Before – Fast 8 hours 
Day 1— clinic visit 
 
*Must be started within 2 weeks of 
Day 0 (Placebo Phase) 
Blood: Baseline- Creatinine, UA 
During at 0.5, 1,1.5, 2, 3, 4, 5, 6, 8, 
10 hours (Creatinine, UA, 
ALLO/OXY_) 
Urine: Before – females 20 ml for 
pregnancy test. 


During- all urine during stay 
collected 
Follow-up 
As close as possible to 24 hours after 
oral dose 
Day 2 –  clinic visit  
 
Blood:, Creatinine, UA, 
ALLO/OXY 
Urine: Turn in urine that was 
collected overnight 
 
Follow-up 
Day 3 – clinic visit 
Blood: Creatinine, UA, 
ALLO/OXY 
 
Follow-up 
As close as possible to 72 hours after 
oral dose 
Day 4 – clinic visit 
Blood:, Creatinine, UA, 
ALLO/OXY 
 
 
 
 
 
 
 
 
 
 
 
ANALYTICAL METHODS: Measurement of allopurinol and oxypurinol will be 
performed by high performance liquid chromatography and mass spectrometry (LC-
MS/MS). Measurement of uric acid and its precursors, xanthine and hypoxanthine, will 
be measured via Amplex Red and Abcam colorimetric assays. 
 
Genotyping: All recruited subjects will sign informed consents and HIPAA forms. At 
screening, a blood sample (20 mL) or saliva sample will be obtained for DNA extraction. 
Serum will be stored for assays of endogenous compounds and drugs. The ABCG2 
variants will be genotyped at UCSF by a Taq Man Assay using primers and probes from 
ABI. The reaction will be run on an ABI 7900 HT. 
 
HLA genotyping and wellness panels will be analyzed by qualified personnel at OMI or 
other reputable company as approved by Kathy Giacomini or other study personnel. 
 
STATISTICAL ANALYSIS: 
Mechanistic pharmacokinetic modeling will be conducted to characterize the 
pharmacokinetics of allopurinol (ALLO) and oxypurinol (OXY). The mechanistic model 
will characterize details relevant for this study such as conversion mechanism from 
ALLO to OXY and estimation of fraction of metabolized drug; estimation of ALLO and 
OXY renal and secretory clearance; possible concentration-dependent saturation 
mechanism of secretory clearance and consequent longitudinal changes in OXY secretory 
CL and influence of pharmacogenomics (ABCG2) and demographic covariates 
(creatinine clearance, age, gender, race and body weight) on secretory and renal CL. By 
use of the physiological model, we will also attempt to estimate the drug concentration in 
the renal tubule; by knowing plasma and urinary concentration of OXY, and by 
estimating filtration and secretory clearance of OXY, we will be able to estimate the time 
profile of tubular concentrations of OXY. These tubular OXY concentrations represent 
concentration at the proposed site of action and they will further be linked with the 
pharmacodynamic response. A model describing concentration dependent inhibition of 


UA re-absorption from the renal tubule by OXY will be implemented. We will examine 
linear and non-linear (Emax type of model) relationships between tubular OXY and 
inhibition of UA reabsorption. In the pharmacokinetic modeling, we expect to see a 
significant influence of creatinine clearance on renal clearance and a significant influence 
of ABCG2 genotype on secretory CL and oral bioavailability, as described below. In the 
pharmacokinetic-pharmacodynamic analysis we expect that individuals with higher 
tubular concentrations of OXY will show the better response in terms of plasma UA 
lowering resulting in higher urine UA concentrations. Consequently, higher tubular 
concentrations will be mostly expected in individuals with high secretory clearance, e.g. 
individuals with ABCG2 141Q/Q, however the modeling may also account for other 
factors contributing to high tubular concentration, such as higher filtration rates and other 
factors, therefore providing more granularity and better mechanistic understanding in all 
processes leading to overall UA physiology. Although this specific PKPD analysis will 
be focused on the mechanism and role of ABCG2 in UA response to ALLO and OXY, 
the established mechanistic framework can be used to test the role of potential new 
pharmacogenetic variants. Longitudinal PKPD modeling using nonlinear mixed effects is 
a method carrying the most power to separate the signal from the noise; therefore 
modeling analysis will be associated with at least 80% power to detect differences in 
primary endpoints (net secretory clearance and change in UA) as described above. In 
addition to confirming the result of the primary analysis, this analysis will establish a 
mechanistic representation of the physiology underlying UA metabolism, therefore 
providing strong support and evidence to our mechanistic hypothesis. Pharmacokinetic 
parameters including renal clearance, and net reabsorptive or secretory clearance of UA, 
OXY and ALLO will be calculated from serum and plasma drug levels by standard 
pharmacokinetic methods. To assess net secretory or reabsorptive clearance of the three 
compounds, we will subtract fu*GFR (where fu is the unbound fraction of ALLO, OXY 
or UA and GFR is the glomerular filtration rate), as measured or estimated from serum 
and urine creatinine values, from renal clearance of ALLO, OXY and UA, directly 
measured from plasma and urine values. Previously we have used these methods to 
calculate net secretion (or reabsorption) of several drugs. For pharmacodynamics, 
differences in DSUA between the genotype groups will be evaluated first with an 
analysis of variance. If DSUA is not normally distributed, we will evaluate the 
differences between the genotype groups using a non-parametric Kruskal-Wallis test. We 
expect individuals with ABCG2 141K/K and 141Q/K will have a reduced response to 
ALLO (smaller DSUA) than individuals with ABCG2 141Q/Q. In addition, individuals 
with ABCG2 141K/K will have reduced renal clearance of ALLO and OXY in 
comparison to other genotypes since we propose that ABCG2 functions as a secretory 
transporter for ALLO/OXY in the kidney. We will also determine whether the ABCG2 
141K carriers have a reduced renal clearance. 
 
DATA STORAGE AND CONFIDENTIALITY:  
All personal and medical data will be considered confidential. Subjects will be assigned a 
unique number that will code for all personal data collected. This code will be used for all 
stored samples. All electronic study data will be stored on a password-protected computer 
on a secure network in the PI's laboratory, and backup files of the data will be stored on a 
hard disk which will be kept in a locked file cabinet or office. 


 
Participation in research may involve a loss of privacy, but information about subjects 
will be handled as confidentially as possible and in compliance with HIPAA regulations. 
All information gathered in the study will be used collectively with information from 
other participants. All personal and medical data will be considered confidential. Upon 
the subjects enrollment in the study the researchers will assign them a number that will 
code for all of the personal data collected. Dr. Giacomini, her research associates, and 
team members will have access to information about each subject. Research staff, UCSF 
Committee on Human Research, and other University of California personnel may also 
review or receive information about the subject for quality assurance purposes (i.e. to 
review an adverse drug response). The subject's names or any identifying information 
will not be used in any published reports about this study, and data will be submitted and 
reported in consolidated, coded form. Furthermore, coded data obtained in the study will 
be deposited into the Pharmacogenetics Research Network Database (PharmGKB) on the 
World Wide Web. Researchers for further studies may access information on this 
database. However, the database will NOT include any information pertaining to the 
identity of the samples. Information identifying the subjects will not be available on the 
database. 
 
For subjects enrolled at Open Medicine Insitute (OMI), access to patients’ records will be 
limited to access by the study and clinic staff.  OMI and site staff will go over the 
questionnaire with the volunteer to determine initial inclusion/exclusion.  Each 
participating patient will be assigned a unique identification number.  All data collected 
on each subject will be labeled with the corresponding study ID number. Patients will be 
enrolled in the study by providing signed written informed consent or by completing the 
existing web-based consent process system 
 
Confidentiality of all records and materials will be maintained to the fullest extent 
possible.  Written informed consent forms will be kept in locked files at participating 
sites or, if completed online, password protected. 
 
All data generated by OMI will be stored on OMI's state of the art data center in Fremont, 
CA. This center runs a VMware distributed architecture with a large EMC2 VNX data 
store with their own proprietary OpenMedNet layer. This ensures data integrity and 
security. 
 
RISK CATEGORY: Minimal 
 
POTENTIAL RISKS:  
 
1. Allopurinol: Allopurinol is a xanthine oxidase inhibitor used to treat gout and 
hyperuricemia that can be dosed from 100 mg to 800 mg daily. The most frequent 
side effects are skin rash (1.5%), diarrhea (1.3%), nausea (1.2%), and acute gout 
attacks. Furthermore, these side effects occur in less than 1% of those patients 
dosed and at a dose of 300 mg/day, we do not expect to see any discomfort in the 
volunteers. 


a. Common but less serious: 
• Diarrhea 
• Drowsiness 
• Headache 
• Change in taste 
• Muscle pain 
b. Rare but serious 
• Renal Failure 
• Stevens-Johnson Syndrome* 
 
* Although some patients develop Stevens-Johnson syndrome on allopurinol, subject’s 
side effects will be monitored closely and dosing will be discontinued with any signs of 
skin rash. Stevens-Johnson Syndrome is also most likely to occur with repeat dosing of 
allopurinol and subjects will only take a single dose.  
 
2. Blood loss and Venipuncture: Withdrawal of blood from a vein may cause pain, 
bruising, and in rare cases infection at the site of needle puncture. Some people 
may faint when blood is drawn. All subjects will provide approximately 160 mL 
of blood. Loss of 160 mL of blood is not harmful and will be replenished by the 
body within 1-2 weeks. 
 
3. Questionnaire: Some of the questions may make the participants feel 
uncomfortable. All participants will be informed that they have the right to 
decline to answer any questions. 
 
4. Genetic Testing: Participation in this study may involve a loss of privacy. All 
DNA testing will be performed using blood/saliva samples tagged with a unique 
identification number, to minimize the risk of loss of confidentiality. Only the 
research staff will have access to the samples and records and the key to the code. 
Information from this study used for scientific publication will not contain any 
identifying information. Subjects may withdraw consent for DNA storage at any 
time during or after the study by writing a letter to REDACTED 
 
5. Inconvenience of Study Visit: Subject may experience an inconvenience by 
having to attend multiple study visits while study procedures and questionnaire 
data are being collected. 
 
PROTECTION AGAINST RISKS: 
To minimize the risks/discomforts to subjects, subjects will be excluded from 
participation if: 
1. Under 18 years old 
2. Patients with vascular disease 
3. Patients with renal impairment (GFR < 60 ml/min) 
4. Patients taking medications that affect uric acid levels 
5. Pregnant or lactating women 
6. Prior history of any allergic reaction to allopurinol 


7. Risk of urinary or gastric retention or narrow-angle glaucoma (by medical history 
examination) 
8. Impaired hepatic function (> 1.5 times the upper limit of normal) 
9. Evidence of anemia (hemoglobin < 10 g) 
10. Evidence or diagnosis of congestive heart failure 
11. Evidence of recreational drug use 
 
To further minimize risks, OMI is equipped with emergency medical tools such as ACLS 
trained personnel, crash carts, and emergency drugs and supplies to stabilize any subject 
until emergency personnel arrive after 911 is called. El Camino Hospital is directly across 
the street in case of emergencies as well. 
 
BENEFITS TO SUBJECTS: 
The benefit of participating in this studies is that you are able to add to our knowledge 
about how the pharmacokinetic and pharmacodynamic mechanisms by which Q141K 
(rs2231142) associates with uric acid lowering in gout patients on allopurinol with 
relatively little inconvenience to you. While these types of studies are unlikely to offer 
direct benefit to your disease at the time of your involvement, the information gained 
may be used to develop new therapies to improve survival or quality of life for you or 
other patients like you in the future. 
 
ALTERNATIVES TO PARTICIPATION: 
The subject can choose not to participate in this study.  
 
SUBJECT IDENTIFICATION AND RECRUITMENT:  
 
Subjects will be recruited from Asian and European populations within reasonable 
driving distance of Open Medicine Institution (OMI), with the initial focus on Asian 
populations. OMI will also reach out to locations where Asian and European descent 
populations are prevalent to request permission to hand out or hang an IRB approved 
flyer. We will also post a booth with the flyer at other local sites. Interested individuals 
may contact OMI directly through the information on the flyer to begin the screening 
process. If research personnel are available in person the interested individual may also 
talk directly to the research personnel at that time in a private location. 
 
PROCESS OF CONSENT: 
 
At OMI, all research intervention will be conducted in a private room to maintain 
privacy. All subjects are required to sign an Informed Consent Form prior to participation 
in compliance with 21CFR50 Subpart B.  Open Medicine Institute (OMI) will receive 
approval of the Informed Consent Form from the Western Institutional Review Board 
(WIRB) as well as continuing review approval, as necessary, in compliance with 
21CFR812 Subpart D. 
All subjects must be provided a copy of the Informed Consent and the California 
Experimental Bill of Rights but the bill of rights and consent may be done either using 
the electronic informed consent on https://www.OpenMedNet.org or using written 


informed consent.  If the subject signs the electronic consent form this form is always 
available for view by the subject, or the subject may print it out, so a signed written copy 
of the informed consent is not necessary.  If the subject signs a written copy the subject 
must receive a copy of the signed and dated consent document.  The original copy of the 
consent form will be retained at OMI in a secure, locked location.  Each subject may be 
enrolled into the study only once. 
Subjects must be explained the purpose of the study in terms understandable to them with 
sufficient information for them to make an informed decision regarding their 
participation.  They must be informed and understand that participation is always 
voluntary and that participation in the study will not impact the clinical care that they 
currently receive.  The consent form will specify whom to contact if there are injuries as 
a result of the study and whom to contact with general questions about the study.  The 
consent form will notify subjects that blood, urine and DNA samples will be collected 
and will be used for research purposes and stored for the purpose of conducting future 
research by UCSF, including genetic DNA sequencing. 
 
SUBJECT CAPACITY:  
Subjects will only be enrolled if they are able to give informed consent themselves.  
 
SUBJECT COMPREHENSION:  
Informed consent will be obtained using the form attached to this protocol. Subjects will 
be briefed on all procedures in the trial in plain speech and will have the opportunity to 
have all of their questions answered. Subjects will indicate that they understand the risks 
and benefits and agree to participate in the study by signing the informed consent form.  
 
DOCUMENTATION OF CONSENT:  
Signed consent forms will be retained on a secure server along with the subject 
information as previously described.  
 
COSTS TO THE SUBJECT:  
There will be no cost to the subject other than the time it takes to participate in this study 
(approximately 14 hours).  
 
PAYMENT FOR PARTICIPATION:  
If subjects agree to be screened, they will receive a $10 payment for the initial screening 
visit (cheek swab). If they qualify and decide to continue onto the secondary screening, 
they will receive an additional $15. If they are enrolled into the remaining phases of the 
study, they will be eligible to receive an additional $325 payment to offset their time and 
inconvenience. Only if they complete everything and comply with all aspects of the study 
will they be paid the entire amount of $325 (in addition to the $25 received for the 
screening for a total of $600). If they fail to comply any portion of the study visit or do 
not complete the entire visit then they will only receive a prorated payment based on the 
time of participation (approximately $10.0 /per hour). Payments will be paid in the form 
of a check or debit card at the completion of the study, which they should receive four to 
six weeks after the last visit. Screening visit may be paid in cash or check. Subjects must 


provide the researchers their Social Security number and mailing address so the check or 
debit card can be processed. Transportation and Parking fees may be provided if 
requested in advance. According to law, subjects will have to pay taxes on payment for 
research participation in excess of $600 per calendar year. 
 
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APPENDICES: 
 
Appendix 1. Allopurinol Informed Consent Form 
 
Appendix 2. Questionnaire: Effect of the Q141K BCRP variant on the pharmacokinetics 
and pharmacodynamics of allopurinol. 
 
Appendix 3. Subject Diary 
 
Appendix 4. Allopurinol Subject Recruitment Flyer 
 
Appendix 5. Allopuinol Insert 
 
Apendix 6.  Allopurinol AGC-9588