Metformin as a Novel Therapy for Autosomal Dominant Polycystic Kidney Disease
Phase: PHASE2
Conditions: Polycystic Kidney, Autosomal Dominant
Sponsor: Kyongtae Ty Bae, M.D., Ph.D.

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Department of Defense Peer Reviewed Medical Research Program 
 
Investigational Medicinal Product 
Metformin 
 
 
 
 
 
CLINICAL PROTOCOL 
 
 
TAME-PKD: Trial of Administration of Metformin – Autosomal Dominant Polycystic Kidney 
Disease 
 
 
 
CONFIDENTIAL – PROPRIETARY INFORMATION 
 
 
 
 
 
 
 
 
 
 
Version 8 
Date: June 1, 2018 
 
 
 
 
 
 
 
 
 
 
 
 

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Table of Contents 
 
 
List of Abbreviations 
3 
Introduction 
5 
Hypothesis and Specific Aims 
8 
Aim 1 
9 
Aim 2 
9 
Known potential risks and benefits 
1. Foreseeable Risks 
2. Risk Management and Emergency Response 
3. Potential Benefits 
10 
10 
10 
12 
Overall Trial Design 
13 
1. Study Population/Eligibility 
13 
Table 1: Inclusion and Exclusion Criteria 
13 
2. Study Time Line 
14 
3. Recruitment Goals 
14 
4. Recruitment 
14 
5. Recruitment Methods 
15 
6. Study Procedures 
15 
7. Optional Genetic Samples 
19 
8. Retention 
9. Treatment Interruption 
10. Treatment Discontinuation 
19 
19 
20 
11. Outcome Measurements 
20 
12. Safety 
22 
13. Statistical Plan and Data Analysis 
24 
Biomarker Identification 
25 
Impact of the Study 
27 
Table 2: Schedule of Assessments for Screening, Baseline and     
Follow-up visits 
28 
Table 3: Metabolites and Metabolomic Biomarkers 
29 
MR Imaging Protocol for PKD Metformin Study 
TAME-PKD Statistical Analysis Plan 
30 
33 
References 
32 
 
 
 

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List of Abbreviations: 
ACC 
 
acetyl co-enzyme A carboxylase 
ACE-I  
angiotensin-converting-enzyme inhibitor 
ADPKD  
autosomal dominant polycystic kidney disease 
AKI 
 
acute kidney injury 
AMPK A 
AMP-activated protein kinase 
ARB 
 
angiotensin II receptor blocker 
BP 
 
blood pressure 
bid 
 
twice daily 
BUN 
 
blood urea nitrogen 
CBC 
 
complete blood count 
CDMRP 
Congressionally Directed Medical Research Programs 
CRHC-DC 
Center for Research on Health Care Data Center 
CKD 
 
chronic kidney disease 
CKDepi 
Chronic Kidney Disease Epidemiology Collaboration 
CFTR  
cystic fibrosis transmembrane conductance regulator 
CRISP  
Consortium for Radiologic Imaging Studies of PKD 
CTRC  
Clinical and Translational Research Center at Tufts 
DCC 
 
data coordinating center 
DCIAC  
Data Coordinating and Image Analysis Center 
DSMB  
data and safety monitoring board 
ESC 
 
evaluation of signed consent 
EDTA  
ethylenediaminetetraacetic acid 
eGFR  
estimated glomerular filtration rates 
EKG 
 
electrocardiogram 
ELISA  
enzyme-linked immunosorbent assays 
ESRD  
end-stage renal disease 
GCRC  
General Clinical Research Center at UMMC 
GSRS  
gastrointestinal symptoms rating scale 
HRQOL 
health-related quality of life 

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Ht 
 
height adjusted 
IAC 
 
Imaging Analysis Center 
IRB 
 
institutional review board 
ITT 
 
intention-to-treat 
KCV 
 
kidney cyst volume 
LCV 
 
liver cyst volume 
LDHA  
lactate dehydrogenase A 
MOP  
manual of procedures 
MRI 
 
magnetic resonance imaging 
mTOR  
mammalian target of rapamycin 
p70S6K 
p70 ribosomal protein 6 kinase 
PCC 
 
participating clinical center 
PCOS  
polycystic ovary syndrome 
PDK1  
pyruvate dehydrogenase kinase 1 
PI 
 
principal investigator 
PKD1  
polycystin 1 
PKD2  
polycystin 2 
PKDF  
Polycystic Kidney Disease Foundation 
PKM2  
pyruvate kinase M2 isoform 
p.o. 
 
orally 
RCT 
 
randomized clinical trial 
SAE 
 
serious adverse events 
SF-36  
Medical Outcomes Short Form 36 
TCA 
 
tricarboxylic acid  
TKV  
 
total kidney volume 
TLV 
 
total liver volume 
UMMC  
University of Maryland Medical Center 
 
 

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INTRODUCTION 
 
BACKGROUND  
 
A. ADPKD Background and Pathogenesis: Autosomal dominant polycystic kidney disease 
(ADPKD) affects 1:700 – 1:1,000 individuals, making it the most commonly inherited kidney 
disease. Over time, half of ADPKD patients will progress to end-stage renal disease (ESRD). To 
date, no targeted therapies have been shown to safely and effectively halt progression of ADPKD. 
ADPKD is caused by a mutation in the PKD1 or PKD2 genes, which encode polycystin 1 (PKD1) 
and polycystin 2 (PKD2), respectively, two proteins that together form a receptor channel complex. 
ADPKD can be considered, in large part, as a disease in which normally non-proliferating and 
absorptive cells of the kidney tubule take on a proliferative and largely secretory phenotype, 
manifesting as progressive cyst development. The pathological processes that produce enlarged cysts 
are thought to result from specific cellular abnormalities: 1) increased fluid secretion into the cyst 
lumen; and 2) inappropriately increased cell proliferation of the cyst epithelium (1).  
 
Grantham and colleagues demonstrated that the rate of fluid secretion into the lumen of cultured 
ADPKD cysts is proportional to the quantity of cystic fibrosis transmembrane conductance regulator 
(CFTR) Cl- channel present at the apical membranes of the cyst-lining epithelial cells (2). These data 
are consistent with the hypothesis that active fluid secretion plays a critical role in driving cyst 
growth. The evidence that CFTR acts as a significant contributor to cyst growth has led to preclinical 
explorations of the utility of CFTR-inhibitors in the treatment of ADPKD (3, 4). 
 
 A number of groups have also demonstrated increased proliferation rates in the cells of ADPKD 
cysts using cell lines derived from animal models (5). These observations suggest that secretion does 
not account for the entire process of cyst enlargement. Rather, the cyst lining cells have a hyper-
proliferative phenotype (6), which results in expansion of the cyst wall surface area required to 
enclose the increasing cyst volume. Intriguingly, it has been shown that mammalian target of 
rapamycin (mTOR) activity is elevated in murine models of PKD (7). mTOR, a serine/threonine 
kinase that regulates cell growth and proliferation, transcription, and protein synthesis, is inhibited by 
rapamycin (8, 9). Indeed, treatment with rapamycin (sirolimus) has been shown to improve renal 
cystic indices in mouse and rat models of ADPKD (10). 
 
 B. Prior Relevant Clinical Trials for ADPKD: Because of the promising results obtained in rodent 
ADPKD models, the mTOR inhibitors sirolimus and everolimus were recently tested in patients for 
the treatment of ADPKD (11, 12). However, these studies did not show significant benefit and there 
were dose-limiting side effects in the drug treatment groups that may have diminished their 
effectiveness. In the TEMPO study the vasopressin receptor 2 antagonist tolvaptan slowed the rate of 
GFR loss by 30% and decreased the growth of total kidney volume (TKV) over three years (2.8% per 
year vs. 5.5% per year in the placebo arm) (13). However, the tolvaptan group had an increased risk 
for hepatotoxicity, which led the FDA to issue a black box warning against its use in patients with 
liver disease and use for more than 30 days. In addition, there were tolerability problems caused by 
polyuria, which led to a significantly greater percentage of discontinuation of the study in the 
tolvaptan group (23% vs. 14% of placebo patients).  On April 24, 2018, the FDA approved Tolvaptan 
as a medication to slow kidney decline in some adults with ADPKD, specifically those at risk for 
rapid progression.  However, patients electing such treatment will be required to register in a FDA-
mandated Risk Evaluation and Mitigation Strategy (REMS) including frequent laboratory monitoring 
for liver toxicity.  There remains an unmet need for additional therapeutic options to reduce 
progression in ADPKD that would be safe and well-tolerated.   

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 C. Preclinical Data with Metformin: There is growing evidence that metformin, a drug widely 
used for the treatment of type 2 diabetes and polycystic ovary syndrome, may serve as a novel 
therapy for individuals in the early stages of ADPKD by activating the metabolic sensor AMP-
activated protein kinase (AMPK). AMPK is activated under conditions of metabolic and other 
cellular stresses. Through its actions on downstream 
mediators, AMPK activation during low energy states 
decreases cellular energy consumption while stimulating 
energy generating pathways. We have shown that AMPK 
phosphorylates and inhibits CFTR, thus suppressing 
epithelial fluid and electrolyte secretion (14, 15). Similarly, 
AMPK phosphorylates the tuberin protein, leading to 
indirect inhibition of the mTOR pathway. Thus, AMPK 
inhibits both CFTR (15) and mTOR (16), suggesting that 
targeted activation of this kinase by metformin may 
provide a therapeutic benefit in ADPKD (17). We have 
shown that metformin treatment of kidney epithelial cells 
leads to stimulation of AMPK and subsequent inhibition of 
both mTOR and CFTR activity. We have also shown that 
metformin slows cystogenesis in animal models of PKD, 
supporting the potential of this drug in ADPKD treatment 
(18) (see Fig. 1). 
 
D. Byproducts of Perturbed Metabolism as Biomarkers of PKD Progression and Therapeutic 
Efficacy: One of the major challenges in the field of PKD research is that monitoring disease 
severity, progression over time and the potential response to novel therapies is challenging because 
the disease is slowly progressive over decades with patients often remaining asymptomatic for 
extended periods. Moreover, besides serum creatinine, which is often a late marker of significant 
renal function decline, current monitoring methodologies involve expensive kidney imaging 
procedures like serial volumetric MRI measurements. Thus, the availability of biomarkers that are 
informative, reliable, inexpensive and widely available would be invaluable for the management of 
patients with ADPKD. This need for diagnostic and prognostic biomarkers will become increasingly 
important with new potential drug therapies for ADPKD on the horizon, such as metformin 
proposed in this application and others that are currently being tested in clinical trials. To date, 
there have been several recent studies investigating potential urinary biomarkers for ADPKD, 
including a recent multi-center trial that identified a negative correlation between peptides largely 
derived from collagen and other structural proteins and height-adjusted total kidney volume (and 
thus disease severity) (19) and another that performed NMR spectroscopic small molecule 
fingerprinting of the urine in a cross-section of ADPKD patients and healthy controls (20). However, 
major limitations of such biomarkers are their reliance on the monitoring of multiple simultaneous 
biomarkers (i.e., patterns) using sophisticated analytic equipment, their frequently lower sensitivity 
and specificity early in disease, and their ability to provide useful prognostic information. 
 
Recent work by Boletta and colleagues has suggested that the Pkd1 mutation in kidney cells alters 
energy metabolism and oxygen sensing by inducing aerobic glycolysis and inhibiting oxidative 
metabolism (the “Warburg effect” seen in many tumor cells (21)), which in turn has the deleterious 
effect of inhibiting AMPK activity in ADPKD (22). Of note, studies derived from the cancer literature 
suggest that metformin treatment may prove beneficial in cancer therapy, and one potential 

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mechanism behind its beneficial effect may be via reversal of the Warburg effect on metabolism. 
AMPK has been shown to negatively regulate the Warburg effect and tumor growth in vivo (23). 
Moreover, metformin decreased HIF-1α levels in a breast cancer model (24), exhibiting an 
antiproliferative, anti-Warburg effect, probably via AMPK activation. Metformin also inhibited cell 
growth and division by inhibiting expression of fatty acid synthase (25) and favoring fatty acid 
oxidation in colon cancer cells (26). In summary, based on the recent literature including our own 
preclinical work (18), we hypothesize that metformin will slow cyst growth in ADPKD and may 
improve dysregulated metabolism in the disease. Moreover, as metformin activates AMPK, 
targeting this fundamental ADPKD disease mechanism of dysregulated metabolism, we believe that 
the identification of metabolomic biomarkers will be particularly useful in the setting of metformin 
treatment in human patients. 
 
E. Preliminary Metabolomic Biomarker Data: We will 
measure activation of the AMPK pathway in urine as a 
potential readout of metformin effects in patients by 
quantitating the band densities on immunoblots 
corresponding to the ratios of phosphorylated to total 
AMPK, acetyl-CoA carboxylase (ACC), and P70 ribosomal 
S6 kinase (P70S6K). Regarding biomarkers, we will 
measure the following glycolytic enzymes: lactate 
dehydrogenase A (LDHA), pyruvate dehydrogenase kinase 
1 (PDK1), and pyruvate kinase M2 isoform (PKM2), as the 
levels of each of these enzymes appeared to positively 
correlate with disease severity in human patients and 
mice (22). We can detect these enzymes in the urine of 
control human subjects via immunoblots performed 
following methanol-chloroform precipitation (Fig. 2). For the studies proposed in this application, 
however, we will utilize more sensitive and quantitative commercially available enzyme-linked 
immunosorbent assays (ELISAs) to quantitate their levels in urine (normalized by creatinine 
concentration) to compare across patients and at various treatment times within patients, as 
described in Aim 2. In addition, we will use colorimetric enzymatic activity assay kits to quantitate 
levels of various key glycolytic and oxidative metabolites (lactate, pyruvate, succinate and cAMP). 
Interestingly, preliminary measurements of the lactate/pyruvate ratio and succinate levels in 
several fasting urine specimens obtained from a cross-section of ADPKD patients enrolled in the 
Baltimore PKD Research and Clinical Core Center suggest that the lactate/pyruvate ratio, indicative 
of enhanced glycolytic flux, may be elevated in ADPKD patients as compared with normal 
volunteers (albeit significant intersample variability exists) (Fig. 3A). Conversely, urine succinate 
levels (a marker of oxidative metabolism via the TCA cycle) appear to fall with rising serum 
creatinine (i.e., worsening renal function) (Fig. 3B). Thus, our preliminary data measuring 
metabolomic profiles in urine from ADPKD patients not only support the feasibility of these 
measurements in our hands, but are consistent with a metabolic reprogramming from 
mitochondrial oxidative respiration to aerobic glycolysis in ADPKD. 
 
 

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F. Synergy of this Study with Preclinical Work: We expect 
that our efforts may allow us to identify biomarkers 
whose levels can be used as indicators that the pathways 
targeted by metformin have been successfully modulated 
and may correlate with ADPKD disease severity and 
progression over time. Of note, one of the co-PIs of this 
proposal (Hallows) is also a partnering PI on a currently 
proposed Department of Defense Investigator-Initiated 
grant application to study metformin along with other 
AMPK activators in preclinical models, which include 
ADPKD 3-dimensional cell cultures and a progressive 
disease model in mice. In addition to screening mouse 
kidney tissue and urine samples for the same biomarkers 
to be assayed in this clinical study, a variety of additional 
glycolytic, TCA cycle, amino acid and nucleotide 
metabolites will be assayed by mass spectrometry in a 
targeted platform in collaboration with S. Pennathur at 
the Univ. of Michigan (see attached letter and biosketch). 
Any novel pharmacodynamic biomarker targets identified 
through the work in this preclinical study could also be 
interrogated in banked urine samples from this study on 
patients treated with the study drug over time to test 
whether new biomarker correlations with metformin 
therapy identified in animal models are also observed in 
ADPKD patients during experimental metformin therapy. 
Successful completion of this work could thus produce an 
additional justification to support bringing metformin and other new drugs to the later stages of 
drug development and reveal biomarkers that will be valuable in future larger clinical trials. 
 
G. Metformin Safety: Metformin has an extensive and long track record for safety. Originally 
described in 1922, the first clinical trial for metformin occurred in 1957 in patients with diabetes. 
This biguanide drug that decreases hyperglycemia has subsequently become a worldwide, first-line 
treatment for type 2 diabetes mellitus as well as a treatment for polycystic ovary syndrome. The 
most common side effect is gastrointestinal distress, but this can be mitigated by titration of the 
dose. Moreover, unlike other diabetic agents, it generally does not cause hypoglycemia (27, 28). 
Metformin, which is not metabolized, gets cleared by the kidneys. While metformin treatment may 
induce lactic acidosis in patients with severely decreased renal function, this risk is exceedingly rare 
for patients with an eGFR >30 ml/min (29, 30). 
 
 
 
HYPOTHESIS AND SPECIFIC  AIMS 
 
Based on our preclinical data as well as the literature, we hypothesize that metformin, an agent that is 
widely used to treat type 2 diabetes mellitus and polycystic ovary syndrome, may serve as a novel 
therapy for individuals with early stage ADPKD by activating the metabolic sensor AMPK. 
 
 

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Aim 1: We will perform a double-blind, multicenter parallel-group randomized clinical trial to assess 
tolerability and safety of treating ADPKD patients with early disease (eGFR ≥50 ml/min) with 
metformin. 
 
We hypothesize that metformin, an agent that is widely used to treat type 2 diabetes mellitus and 
polycystic ovary syndrome, may serve as a novel therapy for individuals with early stage autosomal 
dominant polycystic kidney disease (ADPKD) by activating the metabolic sensor AMPK. To test this 
hypothesis, we propose a pilot-level double-blind, phase 2, parallel-group, placebo-controlled 
multicenter clinical trial to evaluate the safety and tolerability of metformin in ADPKD patients over a 
two-year period. In addition, we will evaluate the effect of metformin therapy on the rate of change in 
height-adjusted total kidney volume (htTKV) and estimated glomerular filtration rate (eGFR) as well as 
on metabolomic biomarkers of disease severity and progression. We will enroll 96 adult patients, ages 
18-60 years with ADPKD, and eGFR ≥50 ml/min/1.73m2. We will restrict enrollment to individuals with 
eGFR ≥50 cc/min/1.73m2 in order to avoid excess risk of metformin adverse events that might be 
associated with more advanced renal impairment. In addition, interventions are more likely to be 
effective if applied early in ADPKD, before there is irreversible renal scarring, as is commonly seen with 
more advanced renal function impairment. It is anticipated that each clinical site (Tufts and UMMC) will 
recruit 48 participants (assumes 15% loss of information) who will be randomized in a 1:1 ratio to 
receive either placebo or metformin, titrated to 1000 mg twice daily, or maximal tolerated dose, over a 
6-week period. 
The primary outcome of the trial is the safety and tolerability of metformin, which will be 
evaluated at baseline, 1 and 3 months and every 3 months thereafter. Safety will be assessed through 
ascertainment of symptoms, laboratory measures (CBC, basic chemistries, liver function tests, 
hemoglobin A1C and plasma lactate levels), vital signs and physical exam. Other adverse and serious 
adverse event rates will also be collected. Tolerability will be assessed by a validated instrument, the 
Gastrointestinal Symptoms Rating Scale (GSRS), used to evaluate the most common anticipated side 
effects, which are GI-related symptoms. As well, at in-person visits, participants will be asked, “Can you 
tolerate this dose of the study drug for the rest of your life?” We will also examine rates of study drug 
`discontinuation, reasons for discontinuation, and adherence with study drug as assessed through pill 
counts. Efficacy endpoints include a) change in kidney, kidney cyst, liver and liver cyst volumes on MRI; 
b) change in glomerular filtration rate, estimated from serum creatinine, using the CKD-Epi equation 
(31); c) the incidence of kidney pain as reported on the TAME Pain Questionnaire; and d) the Medical 
Outcomes Short Form 36 (SF-36) questionnaire for measuring health-related quality of life (HRQOL). 
Urine will be collected from participants at each study visit to measure biomarkers that have been 
shown to correlate with ADPKD disease severity and response to metformin.  In addition, and with 
participant consent, optional blood, sera and urine samples will be collected for biobanking and DNA 
extraction and banking.  An independent Data and Safety Monitoring Board (DSMB) will monitor 
participants through interim analyses of adverse events. 
 
Aim 2: We will perform metabolomic and metabolite measurements on urine samples from our study 
patients both before and after initiation of the study drug to evaluate potentially relevant biomarkers 
that correlate with ADPKD disease severity and response to metformin. 
 
We hypothesize that metformin treatment will ameliorate ADPKD severity and progression in patients 
and that this beneficial effect occurs in large part through changes in the cellular bioenergetics and 
proliferative capacity of ADPKD kidney epithelial cells. Moreover, Boletta and colleagues recently 
identified a Warburg effect-like shift to excessive aerobic glycolysis, as evidenced by increased levels of 
certain key metabolic enzymes (e.g., lactate dehydrogenase A (LDHA), pyruvate dehydrogenase (PDH) 
kinase 1 (PDK1), and the pyruvate kinase M2 isoform (PKM2)), and a decrease in signaling through the 

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AMPK pathway that occurs in ADPKD cystic epithelial cells as compared with normal kidney epithelial 
cells (22). Metformin promotes cellular AMPK activation (55), which may confer beneficial effects in the 
treatment of diseases such as the metabolic syndrome, diabetes, and polycystic ovary syndrome (PCOS) 
(56). Our previously published data demonstrate that metformin, via AMPK activation, inhibits both 
epithelial cellular proliferation through the mTOR pathway and CFTR-dependent fluid secretion into PKD 
cysts (Fig. 1), resulting in decreased cystogenesis and disease severity in vivo in rapidly progressing 
mouse models of ADPKD (18). For the studies proposed in Aim 2, we will investigate candidate 
metabolic biomarkers (Table 3) in urine based on the work of Boletta and colleagues and others (22). 
We hypothesize that glycolytic pathway enzymes will be progressively increased and markers of AMPK 
pathway activation will be progressively reduced with worsening ADPKD severity in patients. We further 
hypothesize that treatment of patients with metformin will tend to reverse these effects, which are 
indicative of the metabolic disturbance in ADPKD, and potentially thereby represent a beneficial 
therapeutic effect. 
 
KNOWN POTENTIAL RISKS AND BENEFITS 
 
ADPKD is the most commonly inherited kidney disease with over half of the gene carriers developing end-
stage renal disease.   The risks a patient is willing to accept are based on his/her personal and/or familial 
relationship with ADPKD.  The risks of this research are well explained and documented in the consent 
process, and patients are encouraged to weigh the risks against any potential benefits.  
 
  
1. Foreseeable Risks 
Foreseeable risks of treatment with metformin include rare but serious adverse effects of lactic acidosis, 
hypoglycemia, leukocytoclastic vasculitis with pneumonitis, or Vitamin B12 deficiency induced 
neuropathy or megaloblastic anemia.   More commonly reported but less serious adverse effects include 
abdominal discomfort of bloating or dyspepsia, metallic taste, anorexia, nausea, soft stools or diarrhea, 
headache and myalgias.  Potential risks associated with phlebotomy are not greater than minimal and 
include bruising and bleeding.  Potential risks associated with MRI are also minimal, and include: mild 
physical discomfort from lying still in the scanning bed, claustrophobia; and (very rare) pain and injury 
from movement of implanted ferromagnetic material/devices.   Of note, we will not be utilizing 
gadolinium-based contrast agents in this trial.  As with all human subject research, there is also the 
potential for loss of privacy and confidentiality. 
 
2. Risk management and emergency response 
The research monitor, who is an endocrinologist with vast experience in the use of metformin in clinical 
care, will be available to the study team to discuss investigation and management (including whether or 
not to discontinue drug) for any side effect that the PI believes could be related to study drug.  In the 
event of potential serious adverse events that are believed to be study related, investigators will have a 
low threshold to discontinue the study drug until the situation is clarified. Drugs that interact with 
metformin will be carefully screened for and – where medically appropriate – we will recommend their 
providers to discontinue these medications prior to study enrollment. These drugs include nifedipine, 
furosemide, amiloride, ranitidine, triamterene digoxin, procainamide, quinidine, vancomycin, and 
trimethoprim.  Inability to discontinue one of these medications will be an exclusion criterion. Patients 
will be given written and verbal instruction about the signs and symptoms of the serious side effects of 
metformin and will be instructed to monitor for these and to alert the study team immediately should 
they experience one or more of these.  Precautions to reduce the likelihood of a serious adverse event 
will be taken as discussed in each specific SAE below.   

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Lactic acidosis is a rare but life-threatening complication of metformin use; the incidence in a 
diabetic population is estimated at 0.03 per 1000 patient years of follow-up, and the incidence of fatal 
lactic acidosis is estimated at 0.015 per 1000 patient years.  The incidence increases with declining eGFR 
though and is believed to be extremely low at eGFR >30 mls/min/1.73m2.  Nonetheless, given the clear 
relationship of this life-threatening outcome to kidney function, we will be monitoring eGFR at least 
every 3 months (and more often during study drug titration) and if eGFR falls below 45 mls/min/1.73m2 
throughout the trial, the following changes will be made:  If GFR is between 30-44 mls/min/1.73m2, the 
metformin dose will be reduced by one-half and the frequency of bloodwork (including lactic acid level) 
will be increased to monthly bloodwork.  If eGFR is <30 mls/min/1.73m2, metformin will be 
discontinued.  Progressive decline in eGFR to <30 ml/min/1.73m2 over 2 years is unlikely in an ADPKD 
population with initial eGFR ≥50 mls/min/1.73m2, with the average rate of decline being 3-5 
ml/min/1.73m2/year. A more likely scenario in which GFR would decline to <30 ml/min/1.73m2 would be 
in the setting of AKI.  AKI is most likely during hospitalization and with exposure to IV contrast, and 
patients will be instructed about these associations and that they should discontinue metformin if they 
are admitted to a hospital or if they will be receiving iodinated contrast.  The metformin should not be 
resumed until eGFR has been confirmed to be >30 mls/min/1.73m2 (and eGFR is stable) after 
hospitalization or iodinated contrast exposure. The risk of lactic acidosis is increased with medical / 
surgical problems and this would be another reason to discontinue metformin during hospitalization.   
This approach to drug titration and temporary discontinuation is consistent with recently revised US FDA 
prescribing guidelines. (58)   
The onset of lactic acidosis is often subtle and accompanied only by nonspecific symptoms such as 
malaise, myalgias, increasing somnolence, dyspnea and nonspecific abdominal distress.  Study staff and 
patients will be educated to monitor for these symptoms.  Once a patient is stabilized on any dose level 
of metformin, GI symptoms, which are common during initiation of therapy, are unlikely to be drug 
related.  Later occurrence of GI symptoms could be caused by lactic acidosis and patients will be told to 
promptly communicate with the PCC should they experience such symptoms.   In the event of any of 
these symptoms, the study coordinator will inform the PI immediately, and metformin may be 
withdrawn (depending on symptom severity), per the PI’s discretion, until the situation is clarified – this 
may include measurement of serum electrolytes and lactate levels where appropriate.  If lactic acid 
levels are not elevated, then the drug will be resumed, with close-follow-up (within 3 days) by telephone 
to assess the patients’ symptoms. In the event lactic levels are >5 mmol/l or the patient describes more 
severe symptoms, the patient will be instructed to go to the closest Emergency Room immediately for 
further management.  Further management consists of supportive care and in severe cases, dialysis. 
Hypoglycemia is an infrequent complication of metformin therapy in patient populations with or 
without diabetes.  For example, there were no reported episodes of hypoglycemia in the Diabetes 
Prevention Trial in a patient population with prediabetes (59).  Under usual circumstances of use in 
populations with diabetes, it does not occur in patients receiving metformin alone but could occur if 
caloric intake was deficient, when strenuous exercise was not compensated by caloric supplementation 
or during alcohol intoxication.  Patients will be told to avoid the aforementioned circumstances that 
could precipitate hypoglycemia.  Patients will be closely monitored with serum glucose levels checked 
bi- weekly during titration and every 3 months thereafter in individuals with GFR >50 ml/min/1.73m2, 
and monthly for individuals with GFR 30-44 ml/min/1.73m2. Study drug will be discontinued at eGFR <30 
ml/min/1.73m2, as noted earlier. While there are established values of fasting blood glucose levels for 
defining abnormal glucose tolerance, the lower limit of normal for fasting glucose in individuals without 
diabetes is not as clear. For this reason, we will modify the current protocol to define hypoglycemia as a 
blood glucose <70 mg/dl if it is accompanied by symptoms and signs of hypoglycemia.  Staff and patients 
will be alerted to symptoms of hypoglycemia (E.g. shakiness, lightheadedness, tachycardia, blurred 
vision, weakness, diaphoresis, somnolence) and patients will be told to immediately drink a sugar 

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containing beverage (E.g. fruit juice or regular soda) and contact the PCC if one or more of these 
symptoms occur.   
 
A blood glucose >60 and  <70 mg/dl  noted on laboratory examination that is not accompanied 
by one or more of the described hypoglycemia symptoms will be considered an alert value that 
does not require modification of dosing of study medication or increased frequency of glucose 
monitoring,  
 
A blood glucose >60 and <70 mg/dl on laboratory examination that is accompanied by 
symptoms of hypoglycemia will prompt a discussion between the investigator and the research 
monitor.  Management will typically consist of modification of dosing of the study medication as 
well as an increase in frequency of glucose monitoring.  
 
A blood glucose <60 mg/dl) will be considered as defining hypoglycemia, independent of the 
presence or absence of symptoms.  The investigator must discuss with the research monitor but 
management will typically consist of reducing the dose or discontinuation of the study 
medication permanently. 
These recommendations are consistent with the Endocrine Society Clinical Practice Guideline for 
Evaluation and Management of Adult Hypoglycemic Disorders 2, which recommends “ . . .evaluation and 
management of hypoglycemia only in patients in whom Whipple’s triad—symptoms, signs, or both 
consistent with hypoglycemia, a low plasma glucose concentration, and resolution of those symptoms or 
signs after the plasma glucose concentration is raised—is documented.” (60) 
 
Vitamin B12 Deficiency Related Complications.  It is estimated that 30% of diabetics treated with 
metformin malabsorb Vitamin B12 (Vit B12) [Ting RZ Arch Intern Med 2006; 166:1975–1979]. 
Complications of Vitamin B12 deficiency – where severe and persistent – may include irreversible 
neuropathy and/or megaloblastic anemia.   To reduce the risk of these serious adverse events, Vitamin 
B12 levels will be checked prior to enrollment and Vitamin B12 deficiency will be an exclusion criterion.  
The risk of adverse effects from metformin-induced vitamin B12 malabsorption is extremely unlikely in 
this 2-year trial of metformin-naïve patients who are not Vitamin B12 deficient at baseline, as it takes 
twelve to fifteen years to totally deplete pre-existing Vitamin B12 stores in individuals with B12 
deficiency [Deller Q J Med 1962;31:71–88] .  Nonetheless we will measure Vitamin B12 levels annually in 
patients and if Vitamin B12 deficiency is found, the participant will be referred to appropriate medical 
care for B12 replenishment. Study drug will be continued provided that the patient receives B12 
replacement therapy with monitoring every three (3) months until the B12 value is >250 pg/ml.  
Leukocytoclastic vasculitis /pneumonitis was reported in 1 case. Patients will be instructed to alert the 
PCC immediately should they develop a rash.  Drug will be discontinued if rash is typical of vasculitis 
(palpable purpura), and patient will be referred for dermatology consultation where clinically 
appropriate; if the rash is not consistent with a drug-related leukocytoclastic vasculitis, the study drug 
may be restarted, with careful monitoring for symptoms.  The potential for pneumonitis will be 
considered if a patient presents with dyspnea or respiratory symptoms without fever or other upper 
respiratory infection symptoms, though other diagnoses are much more likely. 
 
GI symptoms are common, especially during drug titration, and usually resolve over time. Symptom 
burden can be reduced by titrating slowly and administering drug with food and these measures will be 
instituted to facilitate initiation, up titration and maintenance of therapy.  Patients will be followed very 
closely, every 2 weeks during drug titration phase, and will be assessed for GI symptoms burden using 
the Review of Symptoms checklist, the GSRS, a standardized and validated instrument, and the response 
to the question:  “Are you able to tolerate this dose of the study drug for the rest of your life?” Study drug 
will not be up-titrated in patients with persistent or distressing symptoms. 

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Risks associated with phlebotomy will be minimized by having skilled and certified phlebotomists 
performing all blood draws.  
 
Risks associated with MRI scans will be minimized by careful MRI safety screening using a 
standardized MRI safety checklist.   Specifically, those patients with implanted ferromagnetic devices 
which preclude safe MRI scanning will be excluded from participation and enrollment.  Likewise, any 
patient with severe claustrophobia or who has been otherwise unable to tolerate an MRI procedure will 
be ineligible.  This safety screening will be administered prior to each participants’ MRI acquisition 
during follow-up in this trial.  As current standard of care for ADPKD patients includes abdominal MRI, in 
most cases in this trial it will be relatively straightforward to ascertain an individual’s previous ability to 
safely tolerate such scanning.   
 
3) Potential Benefits 
Patients may receive direct benefits by being in this study.  These benefits include close follow-up of 
blood pressure and kidney disease, the latter assessed through eGFR and TKV.  Longitudinal information 
about eGFR and TKV may help to determine the rate of progression of ADPKD.  Repeated standardized 
measurement of kidney growth (size) by MRI is not normally performed in clinical practice.  The study 
will test treatments that are thought to play an important role in slowing kidney growth and loss of 
kidney function early in the disease.  The knowledge gained from this study will benefit all patients and 
families with PKD. 
 
OVERALL TRIAL DESIGN, AIM 1 
 
1. Study Population/Eligibility 
Study participants will include N=96 adults aged 18-60 with ADPKD as defined by Pei-Ravine criteria (32). 
Exclusion criteria are selected to identify participants for whom metformin therapy would pose potential 
excess risk, or for whom randomization to placebo control would be inappropriate or infeasible. 
Inclusion and exclusion criteria are described in Table 1 below.   
Concurrent use of tolvaptan will not be permitted in this trial, due to the lack of information regarding 
the safety of combined use of tolvaptan with metformin in ADPKD. Further, the large number of 
tolvaptan associated side effects will make it impossible to ascertain the tolerability and safety of 
metformin which is the primary endpoint of the present study.Also, among the specific aims of this 
randomized trial are to estimate the effects of metformin vs. placebo on change in kidney volume and 
renal function, measures which are affected (often acutely) by tolvaptan.   However, participant will be 
informed through the informed consent process and documentation that tolvaptan has been approved 
by the FDA for some patients with ADPKD, and that they should discuss with their treating nephrologist 
whether this clinical trial is appropriate for them, and that tolvaptan use will be prohibited during this 
clinical trial. 
 
 
Table 1 
Inclusion Criteria 
Rationale 
Autosomal Dominant Polycystic Kidney Disease 
as defined by Pei-Ravine criteria 
Standard clinical definition of this inherited renal 
disorder; distinguishes ADPKD from acquired 
cystic renal disease  
Age 18-60  
 
Fluent English-Speaking  
 
Instructions and patient-reported outcomes are 
in English language  

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Able to provide written informed consent  
Must have cognitive capacity to provide informed 
consent for participation  
Exclusion criterion  
Rationale  
Estimated GFR<50 cc/min/1.73m2 (estimated 
from serum creatinine using the CKD-Epi 
equation)  
Excess risk of metformin adverse drug reactions 
including lactic acidosis with more advanced 
renal impairment  
Diabetes (currently diagnosed, or fasting glucose 
>126 mg/dL, or non-fasting glucose > 200), or 
HbA1C>6.5 
Patients may require metformin or other oral 
hypoglycemic agent for clinical care 
Pregnancy or lactation or intending to become 
pregnant within the next three years 
Risk of metformin to fetus/infant 
Unstable or unclipped (>7mm) cerebral aneurysm High risk of acute intracranial hemorrhage, which 
is often fatal 
Implanted ferromagnetic object (e.g., pacemaker) 
or severe claustrophobia 
Contraindication to renal MRI 
Systemic disease (other than hypertension) likely 
to contribute to kidney disease (e.g., lupus) 
Exclude patients with renal diseases other than 
ADPKD 
Vitamin B12 deficiency 
Metformin may reduce B12 levels leading to 
worsening deficiency 
Active coronary artery disease, which will be 
defined as presence of stable or unstable angina 
Exclude patients expected to suffer acute 
cardiovascular events during 2-year follow-up 
Use of medications that may interact with 
metformin: 
nifedipine 
furosemide 
Cationic drugs (amiloride, ranitidine, triamterene 
digoxin, procainamide, quinidine, vancomycin, 
trimethoprim) 
Minimize drug-drug interactions that can increase 
potential toxicity of metformin. If patients are 
taking these drugs and wish to participate, they 
must have discontinued these drugs at least 2 
weeks prior to the baseline visit, based on 
supervision by their primary physicians. 
Current or Recent (within 2 weeks) use of 
tolvaptan (Jynarque or Samsca)  
Concurrent use of tolvaptan and metformin has 
not been previously examined, and its safety is 
uncertain.  The large number of tolvaptan 
associated side effects will make it impossible to 
ascertain the tolerability and safety of metformin 
which is the primary endpoint of the present 
study. Furthermore, the aims of this trial are to 
estimate the effects of metformin on change in 
kidney volume and renal function, measures 
which are affected by tolvaptan.   
Participation in a clinical trial with study 
medications 
Potential interaction between metformin and 
investigational drugs on primary and secondary 
outcomes 
Active military personnel 
DoD regulations regarding enhanced protections 
of active duty military personnel 
Any solid organ transplant 
 
History of non-compliance  
 
Allergy or intolerance to metformin 
 
Solitary kidney 
 
 

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Enrollment criteria note: We considered whether to define a minimal kidney size for eligibility in this 
trial. Although specific identification of patients with large kidney volumes may select those at greater 
risk for disease progression, requiring MRI scanning and TKV quantification as part of screening would 
significantly prolong and complicate the screening process, adding additional cost and making it less 
likely to reach our pre-specified accrual goals. Nevertheless, even without specific TKV eligibility criteria, 
we expect the vast majority of eligible participants to have significantly enlarged kidneys with volumes 
greater than 750 cc. Based on prior data from the CRISP study, participants had a mean baseline TKV of 
1060 ± 642 ml and a mean eGFR exceeding 99 ml/min/1.73 m2 (f Ref. (33) Table 1). 
 
2. Study Time Line 
The protocol and study procedures will be refined during the first 6 months of the award. At month 6, 
recruitment will begin and continue for 24 months. The last patient will be enrolled by 30 months. All 
study participants will complete the main protocol by Month 56. During the last 6 months, participants 
will complete close-out visits, and the data will be analyzed and prepared for publication. 
 
3. Recruitment Goals 
We propose that each clinical site (Tufts and UMMC) will enroll 48 participants over a two-year period. 
The recruitment goal of 48 participants per site assumes a 15% loss of information (dropout, lost to 
follow up). We anticipate that sites will be able to obtain and review serum creatinine measurements 
and clinic letters prior to the screening visit so as to minimize screen failures. We anticipate 10% screen 
failures per site (i.e., 5 screen failures/site). 
 
4. Recruitment 
Although recruitment is always challenging, ADPKD patients are highly motivated to participate in 
research that may benefit either themselves or their children. In fact, there was a marked over-
representation of ADPKD patients in the MDRD (Modification of Diet in Renal Disease) Study, at 25% of 
the study population, as compared with the prevalence of ADPKD among all causes of ESRD at 10% (34). 
Potential study participants will be identified from among several sources: 
a. PKD specialty clinics at UMMC and Tufts, staffed by site PIs Drs. Terry Watnick and Ronald 
Perrone, respectively, and identified after review of their medical records performed under an 
IRB-approved HIPAA waiver. Dr. Watnick has been in charge of an inherited renal disease clinic 
for more than 15 years. This clinic draws patients from the entire mid-Atlantic region including 
Maryland, Virginia, Washington DC, southeastern Pennsylvania and Delaware. Her site was one 
of the highest recruiting sites in the Otsuka-sponsored tolvaptan (TEMPO) study. Similarly, Dr. 
Perrone has a well-established clinic and he has successfully recruited participants for a number 
of large studies including TEMPO 3:4 and HALT PKD, and several smaller studies including 
TEMPO 271, TEMPO 285, TEMPO 290, and TEMPO 291. Dr. Perrone’s clinic draws patients from 
a wide geographic distribution including international referrals, but the majority of patients are 
from the northeastern U.S., particularly New England and the greater Boston area. Both UMMC 
and Tufts serve as tertiary referral centers in their respective geographic regions and therefore 
have a large population from which to recruit. Of note, both sites are currently recruiting for the 
REPRISE study, which evaluates tolvaptan in PKD patients with eGFRs 25-65.  
b. PKD patient registries maintained at UMMC and Tufts of patients who have expressed interest 
in participation in interventional clinical trials. Drs. Watnick and Seliger maintain registries of 
well-characterized patients who are enthusiastic about participation in clinical trials. Any patient 
seen at UMMC is asked for permission to be contacted about participation in clinical studies. 
The UMMC registry is about to expand to an online format that should make recruitment easier.  
Dr. Perrone established an IRB-approved PKD Research Registry (IRB protocol #8891) in 3/2006; 
any patient with ADPKD seen at Tufts is asked for permission to be included in this research 

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registry and informed consent is obtained. By signing the informed consent, Dr. Perrone and his 
research team are given permission to contact potential subjects about participation in clinical 
studies and to store clinical data elements. As of 9/23/2014, 334 subjects have been consented 
to participate in this database. 
c. PKD patients from the greater Baltimore-Washington and Boston metropolitan regions who 
respond to recruitment materials distributed through printed media or to study descriptions on 
clinicaltrials.gov. Potentially eligible patients who express interest in participation will be 
referred to the study coordinator and/or site investigators, who will discuss details of the study 
protocol with them. 
d. The Polycystic Kidney Disease Foundation (PKDF) has played an important role in recruiting 
participants for many clinical trials. The mission of the foundation is to “promote programs of 
research, advocacy, education, support and awareness in order to discover treatments and a 
cure for PKD”. They maintain a tab on their website (http://www.pkdcure.org/research/clinical-
trials) that lists all clinical trials that are recruiting participants. We propose to list this clinical 
trial on the PKDF website if funded. We also note that Investigator Dr. Perrone is a past Chair of 
the Scientific Advisory Committee and Investigator Dr. Watnick is the current chair; both have 
been active members of the PKDF Scientific Advisory Committee for many years. They have 
been visible and dedicated members of their local chapters and the national organization. They 
personally know many of the members and their families and have given numerous educational 
talks and presentations at annual meetings and other events. Their active involvement is a key 
strength that will increase the success of recruiting members to this study. 
e. Referring Community Nephrologists: The Baltimore PKD Clinical Translational Core has 
established a network of community nephrologists/practices who have actively referred 
participants for the PKD Core study. We have carefully nurtured these collaborations and we 
anticipate that these nephrologists will continue to refer participants for this study. Similarly, Dr. 
Perrone works closely with referring physicians throughout New England and will communicate 
the existence of this study to this network. 
 
5. Recruitment Methods 
We will use a multifaceted approach to recruitment including 1) communicating directly with patients 
when they come for clinic visits; 2) mailing invitations to PKD patients identified by collaborating 
physicians; 3) disseminating  information about the study on clinicaltrials.gov and the PKD foundation 
web site (see above); 4) posting information on the Baltimore PKD Center website, once such activity 
has been IRB approved; and 5) posting information on  the Tufts Medical Center websites, including the 
research portal and the Division of Nephrology. 
 
6. Study Procedures 
All study procedures will take place at the General Clinical Research Center (GCRC) at UMMC and at the 
Clinical and Translational Research Center (CTRC through the Tufts Clinical and Translational Science 
Institute) at Tufts Medical Center. 
 
a. Patient Enrollment and Randomization: The data entry process will begin during the online 
patient enrollment into the study. There will be an Eligibility form, which will generate a 
participant ID. Screening information will then be completed to assess essential inclusion and 
exclusion criteria detailed in the protocol. Any information collected that identifies the patient 
will remain at the clinical center and will not be sent to the data coordinating center (DCC). Once 
the inclusion criteria have been met, the clinical center will submit the randomization form for 
the subject, and the Web-based system will return the participant’s drug card assignment in a 

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double-blind fashion. New subjects enrolled will be given IDs that include a clinical center 
number but will not be sequential to avoid case confusion. 
b. Screening Visit (S): After written informed consent is provided, a complete medical history will 
be obtained, including documentation of comorbid conditions, concomitant medications, and 
PKD related history. A complete physical examination including vital signs, height and weight will 
be conducted. Screening labs will include a CBC, renal panel (electrolytes, BUN –including a 
fasting glucose, and creatinine), hemoglobin A1C, EKG and a urine pregnancy test for women of 
child-bearing potential. Laboratory studies will be performed at either UMMC or Tufts. Patients 
with an eGFR <50cc/min/1.73m2 or any of the other criteria listed in Table 1 will be excluded 
from enrollment into the trial and will exit the study prior to the baseline visit. Since long-term 
metformin treatment can result in low Vitamin B12 levels (35), this will be measured and, if 
deficiency is found, will be repleted by the patient’s primary care physician as part of standard 
medical care prior to randomization. Confirmation of normalization of B12 levels will be 
required prior to the baseline visit. Participants who meet the entry criteria will proceed to the 
baseline visit. 
c. Baseline Visit (B; within 8 weeks of Screening Visit): The baseline visit will include a review of 
interval medical history family history, liver history and life-style history and a full set of baseline 
laboratory studies including CBC, renal panel, liver function tests, fasting glucose, hemoglobin 
A1C, lactate level, lipid profile and a urine pregnancy test in women of child bearing potential. 
Urine will also be collected for biomarker studies to be performed at the University of Southern 
California. With specific patient consent, optional plasma, sera and urine will be collected and 
frozen at -80°C and shipped or transferred to the UMBiobank at the University of Maryland for 
long-term banking for future analyses. Optional blood may also be collected in two EDTA-coated  
tubes and shipped or transferred to the UMBiobank at University of Maryland for DNA isolation 
and banking for potential future genotyping or other analyses. An unenhanced abdominal MRI 
study will be obtained at the participating clinical center (either UMMC or Tufts) and, after de-
identification, uploaded to the University of Pittsburgh Imaging center via Pitt.BOX, a secure 
file sharing system, which is supported by the Tufts Medical Center Information Services 
department  and the University of Maryland Medical Center Information Services 
department and is approved by all of the institutions for sharing data with external 
collaborators.  In the event Pitt.Box is not available, the de-identified images can be 
recorded onto CDs and FedEx’d to the University of Pittsburgh imaging center.   In addition, 
each participant’s scans will be reviewed at their respective clinical center by a certified 
radiologist for any unexpected findings.    Volumetric measurements of the kidney, kidney cysts, 
liver, and liver cysts will be performed on the MR images.  Baseline questionnaires, which 
include the Review of Symptoms, the TAME-PKD pain questionnaire, the Gastrointestinal 
Symptom Rating Scale (GSRS) and the Short Form-36 questionnaire will be administered. After 
these data are collected, participants will be randomized in blinded fashion to metformin or 
matching placebo, in a 1:1 ratio, as described below. 
d. Supply of Study Drug: Study drugs will be dispensed from the Tufts or UMMC investigational 
drug pharmacy. Supplies will be dispensed at each visit to ensure an adequate number of pills 
until the next in-person follow-up visit. In order to save costs on drug dispensing and placebo 
manufacturing, as well as to simplify the drug administration, we will dispense only 500-mg 
capsules of metformin/placebo. The University of Pittsburgh Investigational Drug Service will 
encapsulate and bottle the placebo and metformin so that they appear identical then dispense 
to the sites. 
e. Drug Titration Protocol and Study Drug Discontinuation: Participants will be started on 500 mg 
of Metformin (or matched placebo) once daily for two weeks, with the following scheduled dose 
titrations: 
 
Increase to 500 mg twice daily at the beginning of week 3 

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 
Increase to 1000 mg qAM, 500 mg qPM at the beginning of week 5 
 
Increase to 1000 mg twice daily at the beginning of week 7 
This gradual titration regimen was chosen in an attempt to minimize gastrointestinal 
side effects. Laboratory studies, including a CBC and renal panel, will be obtained at the PCC or 
local Quest lab, whichever is most convenient for the patient, at weeks 2, 4, and 6 per the 
schedule in Table 2. An in-person visit with an extended laboratory evaluation will take place in 
the middle of the titration period at week 4. A member of the study team will contact the 
participant by phone at 2 weeks and 6 weeks during the titration period to assess tolerability 
and to up- or down-titrate the dose, depending on tolerability. Tolerability will be assessed by 
asking a standardized question “Can you tolerate this dose of the study drug for the rest of your 
life?” and responses to the Review of Symptoms checklist and the GSRS.  Up-titration will occur 
if the participant’s eGFR within the past 2 weeks is ≥50 cc/min/1.73m2, and if the person 
answers ”yes” to the standardized question re: tolerability of the current dose and has few, if 
any, adverse symptoms based on responses to the review of symptoms and GSRS. If a 
participant is unable to tolerate a dose of the study drug, and after discussing the participant’s 
responses with the study doctor, the coordinator will then instruct the participant whether to 
adjust the dose of the study drug or placebo.   If the patient is unable to tolerate a specific dose, 
then he/she will be down-titrated progressively to the highest tolerated dosage; this dose will 
be continued for that participant for the remainder of the trial. 
Although metformin is considered to be a Category B drug during pregnancy and has 
been used safely in pregnant women with polycystic ovary syndrome (36), to ensure maximal 
participant safety and to avoid confounding effects of pregnancy on study outcomes, pregnancy 
will be considered an exclusion criterion for this study and female participants of child-bearing 
potential will be required to use contraception during their participation. Furthermore, urine 
pregnancy tests will be administered during each in-person visit. In the unlikely event that a 
pregnancy is detected, the study drug will be stopped immediately. 
Participants will be instructed at each visit to avoid medications that may raise the level 
of metformin, including cationic drugs such as amiloride, digoxin, morphine, procainamide, 
quinidine, quinine, ranitidine, triamterene, trimethoprim, or vancomycin. Nifedipine and 
furosemide have also been shown to elevate the levels of metformin (see package insert).  
Although the risk of lactic acidosis in individuals with GFR >45 is small (29), participants will be 
instructed to contact the study coordinator and hold metformin before any surgical procedure 
that requires restricted intake of food or fluids or if they should require iodinated contrast. 
Participants will also be warned to avoid excessive alcohol intake since alcohol is known to 
increase exponentially the effects of metformin on lactate metabolism. 
f. Drug Adjustment in Setting of low eGFR: The study drug dose will be reduced to 500 mg twice 
daily (or maximally tolerated dose less than 500 mg twice daily) if and when eGFR falls to 
between 30-44 ml/min, and patients will be monitored more closely and have laboratory tests, 
including lactic acid levels, checked every month. It is conceivable that a participant’s eGFR may 
fall below 30 ml/min during the study period of two years. Progression rate reported in the 
TEMPO control population, which had a mean eGFR of 82 ml/min/1.73 m2 at enrollment, was 
3.7 ml/min/1.73m2 per year (13), making it unlikely that disease progression would lead to 
eGFRs below 30 during the two-year study duration; episodes of AKI rather than disease 
progression would be a more likely cause of reduced eGFR. If a patient experiences AKI or is 
found to have eGFR <30 ml/min on a lab test taken outside of the study, the participant will be 
told to contact the PCC, and monthly lab-only visits will be arranged at the PCC or local Quest 
lab (location as per patient preference). If a participant’s eGFR remains below 30 
ml/min/1.73m2 (confirmed on a repeat value), the study drug will be discontinued; however, 
the participant will continue to be followed per the protocol consistent with intention-to-treat 

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(ITT) principles. Resolution of the cause of AKI and improvement of eGFR to ≥30 ml/min/1.73 m2 
will permit resumption of the study medication, with dose adjustment if eGFR is 
<45mL/min/1.73m2. 
g. In-person Follow-up Visits (F): In person follow-up visits will occur at the PCC at Months 1, 3, 6, 
9, 12, 15, 18, 21, and 24. These visits will consist of an interval medical history, a review of 
symptoms, pain evaluation, reconciliation of concomitant medications, adverse event history, 
and a symptom-directed physical examination.  Laboratory evaluation will be similar to the 
baseline visit, including CBC, renal panel, liver function tests, hemoglobin A1C, lactate level and 
a urine pregnancy test for women of child-bearing potential. Laboratory tests will be performed 
at the UMMC or Tufts clinical laboratories. Serum creatinine is measured at both institutions 
and at Quest Labs by using standardized IDMS-traceable methods (37). The TAME Pain 
Questionnaire, GSRS and SF-36 will be administered at each of these visits. Pill counts will be 
conducted to assess adherence to the medication regimen.  Repeat Vitamin B12 will be 
measured at the 12- and 24-month visits. Urine will be collected for biomarkers.  Optional 
plasma, sera and urine will be collected for long-term banking. MRIs will be performed at 
months 6, 12, 18 and 24.  A complete physical examination will be performed at month 24.   
h. Laboratory-only Visits (L): Kidney function will be monitored on a monthly basis during the first 6 
months of the study protocol. In order to avoid frequent in-person visits to either UMMC or 
Tufts, safety labs consisting of a CBC and a renal panel can be drawn at a local Quest lab 
convenient to the participant at and months 2, 4 and 5. Quest uses standardized IDMS-traceable 
methodologies. If a participant’s eGFR falls below 45 ml/min/1.73m2, then additional 
Laboratory-only visits will be continued on a monthly basis during those months where there is 
no in-Person Follow up visit scheduled.   
i. 
Close-out Visit: Study drug will be discontinued at the Month 24 visit. Participants will return for 
a final Close-out visit within 2 months after stopping the study medication. This visit will be 
similar to the in-person follow-up visits and will include the review of systems, labwork, GSRS, 
Tame Pain Questionnaire, SF-36, urine sample for biomarkers and an EKG.  Optional blood, sera 
and urine will be also be collected for transfer to the biorepository. 
j. 
Blood Pressure goals: Blood pressure (BP) management will not be a component of the 
randomized intervention for this pilot-level metformin trial; however, BP control may impact on 
outcomes, making it important to manage BP uniformly in all study participants. Every effort will 
be made to ensure BP is below 130/80.  In the event BP is not consistently at goal (<130/80) the 
study team will communicate with the participant’s treating nephrologist to recommend 
appropriate adjustments or additional anti-hypertensive therapy. Unless contra-indicated by 
drug intolerance (e.g., hyperkalemia, h/o angioedema), agents that target the renin-angiotensin 
system (ACE inhibitors or angiotensin receptor blockers) will be recommended for first-line anti-
hypertensive therapy. 
 
 
 
 

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7. Optional Genetic Samples 
An optional blood sample will be collected at the baseline visit in two EDTA-coated (purple-top) tubes 
and shipped or transferred to the UMBiobank at University of Maryland for DNA isolation and banking 
for potential future genotyping or other analyses.  Patients will be able to opt out of the DNA collection 
but still participate in this clinical trial.  As stated in the consent document, participants will have the 
option to withdraw their samples from the biorepository at any time by submitting a written request.  
However measurements already performed on biorepository samples will not be removed from the 
study database.   Samples will be tracked by the University of Pittsburgh at the Data Coordinating Center 
using standard tracking software.  This DNA may be tested for changes in the genes that cause ADPKD, 
or in other genes that can influence the progression of ADPKD or regulate energy metabolism.  Patients 
will not be provided with the results of these genetic tests as they are research related and are not done 
as part of routine medical care.   
 
8. Retention 
Retention of participants is vital to the success of any study. We will utilize decades of experience at 
both Tufts and UMMC in conducting studies to ensure retention of participants. We have attempted to 
minimize participant burden by using local Quest laboratory facilities to monitor safety labs between in-
person visits. We will aim to schedule participants at convenient times so as to minimize interruption of 
their work schedule. In addition, we have planned to pay participants a stipend for gas and parking costs 
in order to keep the economic burden to a minimum.  In the event a patient relocates from one clinical 
site to the other, the data center will be able to accommodate the participant’s retention in the study. 
Patients unlikely to understand the importance of maintaining follow-up for the entire duration of 
the 2-year study, following strict adherence to study medications, or that are unwilling or unable to 
make the required visits during the screening and baseline periods, will not be enrolled. We have 
generally found that patient retention at Tufts and UMMC in other PKD clinical trials has been excellent. 
Patients highly valued the unique opportunity they had to be evaluated by and ask questions of 
internationally recognized experts in ADPKD at each study visit. Contact between patients and study 
staff, particularly these clinicians, is frequent in the proposed study, and we believe this will similarly 
promote high rates of study retention. 
 
9. Treatment Interruption 
During the 2-year long trial, it is expected that subjects may have one or more treatment interruptions 
during the study period.   If a subject’s study drug is interrupted for safety reasons, laboratory 
abnormalities, use of a prohibited concomitant medication or recommendation of the PCP, PI or safety 
monitor, the subject’s study medication should be resumed as early as the situation allows and with 
Research Monitor approval, if required.  Any study drug interruption of <7 days will be recorded as 
missed doses rather than as a temporary interruption of study drug.  The subject should immediately 
inform the investigator of any missed doses reaching or expected to reach 2 consecutive days or more so 
that the investigator or coordinator can continue to monitor the subject’s treatments.  If the subject has 
a study drug interruption >7 consecutive days, the subject may be asked to complete a subset of the 
screening labs to re-evaluate safety before resuming the study drug, as determined by clinical center PI 
based on the clinical status of the participant and reasons for study drug interruption.   The PI may 
consult with the Study Research Monitor on this to ensure appropriate safety of the participant.  
 
 

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10.  Treatment Discontinuation 
If a subject stops the study drug for medical or personal reasons, they will be asked to continue their 
participation in the study until its conclusion at 26 months.  If the subject is unwilling or unable to 
continue in the study, a Close-Out visit should be scheduled within two months to obtain the same 
measurements that would have been collected at the end of the study.   
Patients choosing to initiate treatment with the (non-study) medication tolvaptan after enrollment will 
be withdrawn from the study permanently (early completion as per protocol) and will not continue with 
follow-up study visits.  The rationale for this discontinuation from follow-up is the expected extreme 
difficulty of performing regularly scheduled study visits while a participant also requires differently 
scheduled safety laboratory testing for the clinical use of tolvaptan under the REMS program.  Further, 
the large number of tolvaptan associated side effects will make it impossible to ascertain the tolerability 
and safety of metformin which is the primary endpoint of the present study. 
 
11.  Outcome Measurements 
The primary outcome is the safety and tolerability of using metformin in ADPKD patients with baseline 
eGFR ≥50 ml/min. 
a. Assessment of Tolerability: Tolerability will be assessed using two standardized patient-reported 
instruments that will be administered at baseline, at each telephone visit during titration, and at 
every PCC visit. The most anticipated adverse effects of metformin are gastrointestinal (GI)-
related. Accordingly, we will use the GSRS instrument to assess GI symptom burden. A widely 
used and validated 15-item questionnaire, the GSRS instrument has been used to compare the 
GI tolerability of different mycophenolate preparations in renal transplant recipients (39). The 
change in GSRS during follow-up will be compared between metformin and control groups, as 
described in “Statistical Plan”. The second assessment of tolerability will be based on responses 
(yes or no) to the following question “Can you tolerate this dose of the study drug for the rest of 
your life?” As described in Section IIIB.7e Drug titration, a negative response will lead to down-
titration of drug. We will assess the maximally tolerated dose of metformin and also analyze the 
rate of discontinuation of study drug and the reasons for discontinuation by study arm. 
b. Safety Outcomes: At each study visit (telephone or PCC), patients will be asked if they had an 
interim hospitalization or medical encounter and if so, the nature of the event. The patient will 
sign a release of medical information form at the baseline visit and this will be used to obtain 
discharge summaries or other clinical documentation to determine the nature and severity of 
adverse events. Serious adverse events (SAE) will be defined as an undesirable experience 
occurring from the time a participant signs the informed consent (at the screening visit) until the 
end of the study, meeting 1 or more of the criteria of: 1) Resulting in death, 2) Non-elective 
hospitalization, 3) Life threatening (if patient continued on study drug would result in death), 4) 
Persistent or permanent harm or disability, 5) Exceeding the nature, severity or frequency of risk 
described in the protocol or 6) resulting in Congenital anomaly. Reporting requirements to the 
DCC and IRBs are described in part 12 SAFETY, Section C. A symptoms checklist (which lists the 
more common or concerning side-effects of the study drug, but allows for free text entry of 
other symptoms) will be collected at baseline and at each subsequent PCC visit. Rates of AEs 
(based on the symptoms checklist) and SAEs will be compared across arms. Hypoglycemia is a 
potential complication of metformin use in a non-diabetic population. The number of 
hypoglycemia events (based on lab results) will be compared across arms. Values for other 
electrolytes, eGFR and liver function tests that are being drawn per the Study Schedule Table 2 
will also be compared across arms. 
c. Adherence: Adherence with study medication will be assessed through pill counts, which will be 
performed by the study coordinator at each study visit. Patients that miss their in-person visit 

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and thus do not refill their study drug will be contacted in a timely fashion by the study 
coordinator, who will have a record of the missed visit. Adherence with study visits will be 
assessed by tracking the number of missed study visits.   During the drug titration telephone 
visits and the F1 in-person visit, the missed study visit window will be +/- 1 day; for subsequent 
visits a missed study visit will be documented as +/- 6 days.   Practical measures to minimize 
inconvenience (parking, stipends if possible), maintaining communication with referring 
physicians, and other means of maintaining direct communication with the patient (follow-up 
and thank you cards after visits, birthday and holiday cards, small gifts) should also promote 
adherence. Finally, regular and frequent follow-up visits should also aid in the retention of study 
patients. 
d. Patient-reported Outcomes: Metformin is hypothesized to reduce TKV growth, which may 
manifest as a difference in pain and other symptoms relating to organ enlargement. The Medical 
Outcomes Short Form 36 (SF-36) questionnaire is the most widely used instrument for 
measuring health-related quality of life (HRQOL), though scores have not been shown to 
discriminate among ADPKD patients with larger vs. smaller kidneys at eGFR >60 ml/min (40). 
The TAME Pain Questionnaire is a modified version of the Wisconsin Brief Pain Questionnaire 
(41) that measures the frequency and intensity of pain and symptoms relating to organ 
enlargement as well as their impact on everyday living. It was validated in a clinical population 
of ADPKD patients from a single center observational study and subsequently adapted for use in 
the HALT Study (42). The frequency of back pain and symptoms related to abdominal distension 
correlated with htTKV in a cross-sectional analysis of baseline questionnaire results in the HALT-
PKD Study A cohort, which is a population with similar eGFR range as expected in the proposed 
study (43). GI symptoms from metformin may affect HRQOL, and thus we will also examine this 
at every visit. The GSRS (a widely-used, validated 15-item questionnaire, described earlier) and 
the SF-36 (as a generic measure of HRQOL) will be administered by study staff (39, 44). The 
questionnaires will be administered at baseline and at in-person follow-up visits.  In addition, on 
telephone visits during drug titration, the GSRS will be asked to evaluate common 
gastrointestinal symptoms. Each question is rated on a seven-point Likert scale, from no 
discomfort to very severe discomfort. For the GSRS the responses on the 15 items are reduced 
to average scores on 5 specific scales (diarrhea, reflux, constipation, abdominal pain, 
indigestion), identified from factor analysis. Among U.S. patients with gastro-esophageal reflux, 
internal reliability of GSRS scores on the outcomes that may be affected by metformin use was 
moderate to high (Cornbach’s alpha: 0.61 to 0.83), and the instrument displayed good construct 
validity (45). During follow-up GSRS scores will be compared between metformin and control 
groups, as described in “Statistical Plan” below. 
e. Death, ESRD and hospitalizations: At each death, if any, study coordinators will request release 
of information from next of kin. Supporting materials will include hospital death summary, 
death certificate, and ESRD Death Notification Form (HCFA Form 2746; the latter if the patient is 
ESRD). ESRD is unlikely in patients with a starting eGFR of ≥ 50 ml/min, but we will ascertain if 
this occurs through communication with the patient at telephone and/or in-center PCC visits. 
Patients will be asked at each in-person or telephone-based study visit about interim medical 
encounters and specifically about hospitalizations. Discharge summaries will be requested to 
determine reasons for hospitalization; length of stay will be recorded. After receiving supporting 
information, the event will be classified by the study team on the appropriate forms 
(Hospitalization, Death and ESRD Forms) and submitted to the DCC. Such information will be 
communicated through the study portal, as was done for the HALT PKD study (46). 
f. Biomarker & Biobank: For biomarker analyses, urine samples (30cc) taken at the clinical sites will 
be divided into 10 –ml aliquots before freezing at -80°C.  One 10 ml urine sample will be sent to 
Dr. Kenneth Hallows at the Keck School of Medicine of USC.  The remaining optional urine 

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samples along with the optional blood samples (20 cc), which will be centrifuged before dividing 
the plasma into 1-ml aliquots and freezing at -80°C, will be transferred or overnight shipped on 
dry ice to the UMMC Biorepository for long-term storage at -80°C. At the Maryland Clinical 
Center, this aliquoting and freezing will be performed directly by the UMMC Biorepository.  
Presently we have no plans for mutation detection, but for patients who also gave consent to 
have their DNA stored for potential future analyses, the UMMC Biorepository will also isolate 
the DNA from blood taken at the baseline visit, according to their standard protocols.   
g. MR Imaging and Volumetric Measurements: The Image Analysis Center (IAC) has been 
established to handle the imaging data and to manage all imaging-related issues for the CRISP 
and HALT studies. The MR Imaging and volumetric measurements established in the CRISP study 
will be employed in the present study, as results from the CRISP study indicate that this MR 
protocol will provide an effective means for longitudinal evaluation of kidney, kidney cyst, liver, 
and liver cyst volumes in ADPKD patients (47). MR images will be obtained at each clinical center 
and transferred to the IAC at the University of Pittsburgh. Prior to scanning participants, the MR 
technologist will be trained or will have experience in scanning PKD participants according to the 
MR study protocol. MR Imaging begins with placing a phased-array surface coil with its center 
over the inferior costal margin, estimated as the upper margin of the kidney, in each participant. 
The field of view will be maintained between 30 and 35 cm. The kidneys will be imaged at 3 mm 
fixed slice thickness in the coronal plane using the T2-weighted single-shot fast spin-echo 
sequence with fat saturation as well as three-dimensional spoiled gradient interpolated T1-
weighted images without fat saturation and during breath-hold. After the acquisition of the 
images, participant names and identifiers will be removed from the images and replaced with 
study numbers and accession numbers for participant confidentiality prior to image 
transmission. The transmitted images will be reviewed by IAC to generate quality control reports 
for the clinical centers. Images determined to be inadequate for measurement must be 
reacquired within 30 days. 
 
Individual whole-kidney volumes (TKV) will be measured from T1-weighted images by means of 
the stereology method, while T2-weighted images are reviewed simultaneously. The areas of 
the whole kidney in each image are calculated from the collection of points, and volume 
measurements are made from a set of contiguous images by summing the products of the areas 
measured and the slice thickness. The volume of kidney cysts (KCV) in each kidney will be 
measured using a region-based thresholding method. Cysts are brighter than the renal 
parenchyma in T2-weight images. Therefore, they could be measured by summing voxels with 
intensity values greater than those of the background renal parenchyma. On each renal MR 
image slice, a binary signal-intensity map will be generated. This will be done by determining a 
threshold signal intensity that visually distinguishes the cyst and renal parenchymal regions. In 
the binary map, cysts that are brighter than renal parenchyma are represented as white regions, 
whereas the background renal parenchyma will be designated as black regions. By summing the 
pixels of white regions, the cystic area will be measured in each slice. The total cyst volume will 
be calculated from each set of contiguous images. Similarly, the total liver volume (TLV) and liver 
cyst volume (LCV) will be measured using the region-based thresholding method. Each 
volumetric measurement will be made by a trained analyst at the IAC and will be reviewed by a 
radiologist for quality control. 
 
12. Safety 
a. Data and Safety Monitoring Board (DSMB): An independent data and safety monitoring board is 
being appointed by the steering committee. The composition of this committee is described 
below: 

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Member 
Expertise  
Institution 
Mary Korytkowski 
Endocrinology 
University of Pittsburgh 
Mark Unruh, MD 
Nephrology 
University of New Mexico 
Silvio Inzucchi, MD 
Endocrinology/Diabetology Yale University 
Ben D. Cowley, MD 
Nephrology/PKD 
University of Oklahoma 
Kenneth Wilkins, PhD 
Biostatistician 
NIH/NIDDK 
 
The DSMB will convene to monitor safety and adverse events every 3 months at the beginning 
of the study until the first 10 subjects have reached the three-month mark, and then every 6 
months thereafter.   Consistent with the phase II design of this study, there will be no planned 
interim analysis of efficacy as the study is not powered for efficacy. Unanticipated serious 
adverse events, related to study medication will be carefully scrutinized. If significant lactic 
acidosis occurs in any participant, Dr. Korytkowski, chair of the DSMB and the DoD Research 
Monitor, will convene the DSMB urgently to determine whether the study will continue.  The 
DSMB will report to the sponsor (DoD) and to the Steering Committee, including each site PI, 
who will have responsibility for submitting reports to their institutions’ IRB for review and 
acknowledgement 
 
DoD Research Monitor:  Mary Korytkowski, MD is Professor of Medicine and Interim Chief, 
Division of Endocrinology.   Dr. Korytkowski is an outstanding clinical diabetologist and is a 
Professor of Medicine at the University of Pittsburgh. She is the Director of Diabetes QI (Quality 
Improvement) at UPMC. Her QI program has received support from the Department of 
Defense, which has adopted several of the protocols developed in Pittsburgh for use in military 
hospitals. Dr. Korytkowski has significant expertise in the management of diabetes and the use 
of metformin. Dr. Korytkowski will serve as the DoD Research monitor and as such, will be 
responsible for monitoring and coordinating all patient safety, data and security measures at all 
sites.  Her role includes: discussing the research protocol with the investigators; the authority 
to stop a research protocol in progress; to remove individual human subjects from a research 
protocol, and to take whatever steps are necessary to protect the safety and well-being of 
human subjects until the IRB can assess the monitor’s report; and the responsibility to 
promptly report observations and findings to the IRB or other designated official and the HRPO. 
 
b. Safety Monitoring: Study visits and safety laboratory monitoring will occur at an interval of every 
3 months or more often (as during titration) throughout this trial, and the frequency will be 
increased to every month for individuals whose GFR falls below 45 ml/min/1.73m2. Lactic 
acidosis is a rare but life-threatening complication of metformin use; the incidence in a diabetic 
population is estimated at 0.03 per 1000 patient years of follow-up. The incidence in non-
diabetics is unknown, but we believe it is lower because diabetes, reduced kidney function, and 
older age predispose to lactic acidosis. The incidence of lactic acidosis increases with declining 
eGFR but is believed to be low at eGFR >30 ml/min/1.73m2, and European guidelines have 
recommended continuation of metformin unless renal function declines below this level (48). 
Similarly, in the US, prescribing guidelines were recently (April 2016) revised by the Food and 
Drug Administration; these guidelines recommend discontinuation of metformin if the eGFR falls 
below 30 ml/min/1.73m2, and to “assess the benefits and risks of continued treatment” for 
patients with eGFR 30-44 ml/min/1.73m2 with regular monitoring of eGFR for these patients 
(58). To provide a high level of protection against the risk of lactic acidosis for participants in this 
study, we have developed the following monitoring and dose adjustment procedures:  

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1) Study drug dose will be reduced to no more than 1000 mg/day, or ½ of the full dose 
(if patient was tolerating a full dose) when eGFR is between 30-44 ml/min/1.73m2, and 
patients will be monitored more closely with monthly blood tests.  
2) The study drug will be discontinued when eGFR is <30 ml/min/1.73m2, although 
patients will continue to participate in follow-up measurements according to intention-
to-treat principles.  
As mentioned in Section IIIB.7.f, progressive decline in eGFR to <30 ml/min over 2 years is 
unlikely in an ADPKD population starting with eGFR ≥50 ml/min, and a more likely risk for lactic 
acidosis is acute kidney injury (AKI). Patients will be instructed about this risk and that they 
should alert their provider to the fact that they are in this study if they are admitted to the 
hospital and that the study drug should be discontinued during hospitalization. The risk of 
metformin-induced lactic acidosis is well known, but to ensure metformin is not continued when 
patients have or are at risk for AKI (as is common during hospitalizations), patients will be 
instructed to contact the PCC if they are admitted, and the PCC staff will communicate this to 
the hospital provider. 
Hypoglycemia is another potential serious risk of metformin use in non-diabetics and this will be 
monitored by symptoms and with lab work, which will be performed every 2 weeks during 
titration and every 3 months thereafter, or for individuals with eGFR <45/min/1.73m2, monthly. 
Patients will be instructed about symptoms associated with hypoglycemia and appropriate 
intervention including prompt ingestion of sugar-containing food or liquids. 
c. SAE Reporting Requirements: All SAEs must be reported within 24 hours of study personnel 
learning of the event to the local PI and to the DCC via data-entry of SAE Report Form 60. The 
DCC will notify the DSMB of SAEs that are drug-related and unanticipated within 5 business days 
of initial knowledge of the event and all PIs within 5 business days (annually if anticipated). Each 
site PI will be responsible for informing their IRB of study-related unanticipated SAEs according 
to their local reporting requirements. Lactic acidosis is an anticipated event but will fall under 
the same guidelines for reporting as unanticipated events, given the life-threatening nature of 
the event and the potential for further events in active participants. The DCC will prepare 
reports of such events for the DSMB at least annually. PIs at the clinical centers are responsible 
for fulfilling local IRB reporting requirements for anticipated SAEs, which may vary by center. 
 
Statistical Plan and Data Analysis for Aim 1 – See detailed addendum p.33-37: TAME –PKD Statistical Analysis Plan  
All primary and secondary outcomes will be described using sample means or sample proportions along 
with 95% confidence intervals, depending on the nature of the outcomes. Volume outcomes from MRI 
(i.e. total kidney, kidney cyst, liver, liver cyst) that are known to be highly skewed will be assessed to see 
if suitable transformations are needed (i.e., natural log). Participant demographics and baseline clinical 
characteristics will be compared between study arms (Metformin versus Placebo) using two-sample t-
tests or chi-squared tests of independence. All primary outcomes will be analyzed under intent-to-treat. 
Every effort will be made to minimize the amount of participant dropout and missing data. We will 
record specific reasons for attrition as well as implement various data verification measures for missing 
data. Ultimately, we will compare participants who dropped out with those who completed follow-up as 
a way to investigate the relationship between attrition and key variables. We will also assess the 
mechanism and degree of missing data (i.e. missing at random) using established techniques by Little 
and Rubin (50). If missing data are substantial, we will compare various multiple imputation techniques 
to see how robust the overall inferences are. 
The primary objective of this phase II, multicenter clinical trial is to evaluate the safety and 
tolerability of metformin versus placebo with a secondary emphasis on efficacy. The safety outcome is 
defined by the occurrence of adverse events and serious adverse events (as defined in Section IIIB.9.b) 
throughout the study. We will calculate cumulative incidence between study arms and use logistic 

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regression to assess the association between study arms and the number of participants affected across 
various SAEs. 
Tolerability will be defined by the GSRS scale, the rate of discontinuation of study drug, and the 
maximal dose that patients are using at the end of the 2-year study, as discussed in Section IIIB.9.a. 
Using a linear mixed model, we will compare the change in overall GSRS score as a function of time 
(months since baseline), the interaction between time and study arm, and clinical site. We will also 
include a random intercept to allow for participant-level variability of baseline GSRS. Of primary interest 
is whether the interaction terms are significant, which would indicate the change over time differs 
between the metformin and placebo arms. Finally, tolerability will also be assessed by comparing drug 
discontinuation rates between study arms. Chi-squared tests of independence will be utilized to 
compare discontinuation rates as well as adherence (measured by pill counts) between study arms. In 
addition, we will also calculate Kaplan-Meier curves to compare time-to-drug discontinuation between 
arms. 
 
Secondary outcomes: The annual rate of change in height-adjusted total kidney volume (htTKV) will be 
compared between Metformin and Placebo arms to give an indication of the efficacy of metformin. A 
Laird and Ware linear mixed model (51) will be fit with the natural log of htTKV (LnTKV) as a function of 
time, the interaction between time and study arm, and clinical site. In order to account for the 
participant-level variabilities of baseline LnTKV as well as rate of change, the model intercept and slope 
will be allowed to vary if needed. A significant interaction between time and study arm could indicate a 
slowing of PKD progression due to metformin. This same model will be used to evaluate secondary 
efficacy outcomes such as kidney cyst volume (KCV), total liver volume (TLV), and liver cyst volume 
(LCV). Renal function, as measured by eGFR, will be compared between metformin and placebo using a 
similar Laird & Ware model as above. The TAME-PKD Pain questionnaire has over 50 questions. A priori 
we identify the following items of the TAME PKD Pain questionnaire as being most likely to differ across 
study arms after 2 years of study drug treatment: back pain frequency, impact of pain on physical 
activity and sleep, and abdominal distension symptoms. The response to each of these questions is a 5-
point Likert scale. The sample size is small in the present study and we are likely to have low frequencies 
in each of the 5 categories. Thus, we will group the lowest 3 levels together with the top 2 levels when 
comparing these items across study arms. 
Power and Sample Size: The proposed study is a phase II trial assessing safety and tolerability.  
We anticipate enrolling 48 participants per study arm with 15% attrition, resulting in an effective sample 
size of 40 per arm. We are not powered to detect meaningful differences in efficacy, but instead focus 
on 95% confidence interval width (two times the margin-of-error) for relevant point estimates within 
each study arm. For the primary safety outcome of SAEs, we will have the ability to estimate a CI width 
no larger than 0.16, assuming the proportion of participants who experience at least one SAE is no more 
than 6% (similar to the highest rates in the HALT trial (Schrier et al., 2014)). For the primary tolerability 
outcome of GSRS, we will have the ability to estimate a CI width no greater than 21, assuming standard 
deviations of the GSRS are below 1.3. With regard to the tolerability outcome of drug discontinuation, 
our anticipated sample size will allow the ability to estimate a CI width no larger than 0.25, assuming the 
rate of drug discontinuation is 15% (similar to TEMPO (Torres et al., 2012)). For the efficacy outcome of 
TKV, we assume the participants taking placebo will see a 5.51% annual increase in TKV (as seen in 
TEMPO (Torres et al., 2012)) with standard deviations for the slope and error of 0.041 and 0.044 (on the 
natural log scale), respectively.  With our anticipated sample size, we will be able to estimate a margin-
of-error of 1.5 percentage points for 95% intervals. 
 
BIOMARKER IDENTIFICATION, AIM 2 
 

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1. Rationale: We hypothesize that metformin treatment will ameliorate ADPKD severity and 
progression in patients and that this beneficial effect occurs in large part through changes in the 
cellular bioenergetics and proliferative capacity of ADPKD kidney epithelial cells. Moreover, Boletta 
and colleagues recently identified a Warburg effect-like shift to excessive aerobic glycolysis, as 
evidenced by increased levels of certain key metabolic enzymes (e.g., lactate dehydrogenase A 
(LDHA), pyruvate dehydrogenase (PDH) kinase 1 (PDK1), and the pyruvate kinase M2 isoform 
(PKM2)), and a decrease in signaling through the AMPK pathway that occurs in ADPKD cystic 
epithelial cells as compared with normal kidney epithelial cells (22). Metformin promotes cellular 
AMPK activation (55), which may confer beneficial effects in the treatment of diseases such as the 
metabolic syndrome, diabetes, and polycystic ovary syndrome (PCOS) (56). Our previously published 
data demonstrate that metformin, via AMPK activation, inhibits both epithelial cellular proliferation 
through the mTOR pathway and CFTR-dependent fluid secretion into PKD cysts (Fig. 1), resulting in 
decreased cystogenesis and disease severity in vivo in rapidly progressing mouse models of ADPKD 
(18). For the studies proposed in Aim 2, we will investigate candidate metabolic biomarkers (Table 3) 
in urine based on the work of Boletta and colleagues and others (22). We hypothesize that glycolytic 
pathway enzymes will be progressively increased and markers of AMPK pathway activation will be 
progressively reduced with worsening ADPKD severity in patients. We further hypothesize that 
treatment of patients with metformin will tend to reverse these effects, which are indicative of the 
metabolic disturbance in ADPKD, and potentially thereby represent a beneficial therapeutic effect. 
2. Methods and Study Design: We will measure key metabolites that are markers of glycolytic vs. 
oxidative metabolism and cAMP by enzymatic activity assays (Table 3A), candidate glycolytic 
pathway enzymes previously reported to be elevated in ADPKD (22) by ELISA assays (Table 3B), and 
known markers of AMPK pathway activation by immunoblotting (Table 3C). Assays for the various 
analytes will be performed as described in Table 3 (legend) at the various time points throughout 
the study specified in Table 2 (baseline at 0 mos., and then 1, 3, 6, 9, 12, 15, 18, 21, and 24 mos. 
after treatment initiation) for each of the patients (~40 in the metformin treatment group and ~40 
in the placebo control group). Enzymatic assays will be fit to a standard curve at an appropriate 
dilution so that the levels fall within the linear range of the standard fit curve. We will analyze 
changes in the mean levels of each of the biomarkers over time for each patient and correlate these 
with metformin treatment status and disease severity/progression over time, as measured by htTKV 
measurements performed every 6 months and eGFR measured at each visit. In addition, in the 
cross-section of all of our patients, stratified by treatment group, we will correlate levels of each 
biomarker with htTKV and eGFR. 
3. Statistical Plan and Data Analysis for Aim 2: All variables will be described using sample means or 
sample proportions along with 95% confidence intervals, depending on the nature of the outcomes. 
The primary goal of Aim 2 is to evaluate potential biomarkers that correlate with ADPKD disease 
severity and response to metformin. Our focus will be on both prognostic as well as predictive 
biomarkers of total kidney volume and eGFR. The analysis for prognostic biomarkers will be 
restricted to participants taking placebo and will utilize a Laird & Ware linear mixed model with the 
following predictors: time (months since baseline), a time-dependent covariate for the biomarker of 
interest, their interaction, and random effects for the intercept and slope. Apart from the ability to 
see whether a particular biomarker at baseline is related to disease severity and progression, we will 
also be able to estimate cross-sectional and longitudinal effects on TKV and eGFR among untreated 
participants. 
The analysis for predictive biomarkers will utilize the same mixed model with the following 
predictors: time, time-by-study arm interaction, baseline biomarker of interest, time-by-biomarker 
interaction, and the three-way interaction of these predictors. This model will give us the ability to 
identify subgroups of participants who may or may not benefit from metformin therapy. 

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4. Expected Results and Future Directions: In general, we will consider a particular biomarker to be 
predictive of ADPKD disease severity if it positively correlates with htTKV and negatively correlates 
with eGFR, our surrogate disease markers. Based on the reported Warburg effect on metabolism in 
ADPKD (22), we expect to see higher levels of the glycolytic enzymes LDHA, PKM2 and PDK1, 
indicative of a reliance on aerobic glycolysis, to correlate with ADPKD disease severity and/or 
progression in patients’ urine samples. In addition, we predict that the lactate/pyruvate ratio will 
positively correlate and the succinate levels will negatively correlate with disease severity. We also 
predict that metformin will: (1) reduce the lactate/pyruvate ratio and increase succinate levels, 
which would suggest a shift from aerobic glycolysis to mitochondrial oxidative metabolism, and (2) 
decrease cAMP levels in urine, as it has been reported that metformin, independently of AMPK 
activation, inhibits cAMP production in cells (57). Elevated cAMP signaling is a well-established 
component of ADPKD pathogenesis that drives cystogenesis via enhanced CFTR-dependent fluid 
secretion into cysts (2). We also predict that treatment with metformin will tend to increase markers 
of AMPK pathway activation (pAMPK/AMPK ratio and pACC/ACC ratio) and decrease the 
pP70S6K/P70S6K ratio, a downstream target of mTOR. Finally, we predict that metformin-induced 
changes in these markers may be associated with a potentially beneficial response to metformin 
treatment on disease progression. 
 
IMPACTS OF THE STUDY 
 
Potential Short-term and Long-term Impacts of this Study: To test the potential of metformin as a novel 
pharmaceutical intervention to delay the progression of early ADPKD, we propose a double-blind, phase 
2, multicenter parallel-group clinical trial that will evaluate the safety and tolerability of metformin in 
ADPKD patients with eGFR ≥50 ml/min over a two-year period. We will also evaluate the effect of the 
drug on TKV and eGFR. Exploratory analysis of urinary metabolomics and AMPK signaling pathway 
biomarkers related to the predicted effect of metformin will also be assessed. We expect that this study 
will demonstrate the safety and tolerability of metformin in the setting of treatment of ADPKD patients 
with only mildly decreased renal function. If this short-term expectation is realized, then a potential 
long-term impact for this study will be the planning and execution of a much larger phase III clinical trial 
that will be sufficiently powered to detect potential efficacy of metformin on disease progression. 
 One major advantage of this drug over other drugs that have been tested recently for ADPKD 
therapy is that metformin has a very long track record of clinical use and safety and tolerability, which is 
critical in this population because for any drug to be useful in slowing disease progression, it will likely 
need to be taken over decades, if not for the rest of these patients’ lives. Of note, despite its primary 
usage in diabetic patients, metformin does not appear to cause clinically relevant hypoglycemia in non-
diabetic subjects. Another advantage is that the drug is generic and very inexpensive, so there will likely 
be a potential for wide distribution to patients at relatively minimal costs to the healthcare system. A 
third advantage is that the drug is FDA-approved for other indications, so its potential off-label use will 
be more quickly translated to the clinic. Another short-term benefit is that we expect to identify and 
confirm the importance of urinary metabolomic biomarkers in ADPKD, which may reflect disease 
responsiveness to metformin. In the longer term, we expect that the identification of such markers 
could play a key role in ADPKD disease management as they could provide a relatively inexpensive and 
non-invasive tool to monitor disease severity, progression potential and response to metformin 
treatment and potentially other therapies that may be used alone or in combination with metformin. 
 
 
 
 
 

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Table 2: Schedule of Assessments for Screening, Baseline, and Follow-up Visits
 
 

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MR Imaging protocol for PKD Metformin study 
 
Before each study, the MR scanner will be adjusted for proper shimming.   
1. Breath-holding instruction will be provided, and the subject will be coached prior to MR scanning. 
Administration of oxygen via nasal cannula may help improve the breath-hold capacity, particularly 
for subjects with limited breath-hold capacity.  
 
2. Subject will be placed supine on the MR table with his or her arms to the side.  
 
3. A phased-array surface coil will be positioned with its center over the inferior costal margin, i.e. over 
the expected location of the kidneys.  
 
4. Scout scan to locate the scan range of the entire kidneys. A stack of axial images to cover the most 
anterocaudal and posterocranial aspects of the kidneys is highly recommended.  
 
5. The field-of-view (FOV) should be kept as small as possible (30-35 cm) without producing wrap-
around artifacts. 
 
6. (Entire Abdomen covering kidney and liver) Breath-hold, coronal T2 scan (SSFSE/HASTE with fat sat) 
at 9mm fixed slice thickness, usually achievable in a single breath-hold. Please make sure both 
kidneys and liver are imaged completely without missing any anterior or posterior portions.  
 
7. (Kidney only) Breath-hold coronal T2 scan (SSFSE/HASTE with fat sat) at 3mm fixed slice thickness, 
which may require 1-4 breath-holds depending on the kidney size. Use as few breath-holds as 
possible. The first scan should cover the posterior aspect of the kidney. Neighboring image groups 
should be overlapped by a single 3mm slice. To determine correct table position choose the “shift-
mean (starting point in GE)” of the second scan for example: the first shift-mean = -60mm, the 
number of slices in the first set =23, (23-1)x3=66mm, new shift mean =-60+66=6mm.  
 
8. (Kidney only) Breath-hold coronal T1 scan (3D VIBE/FMPSPGR/LAVA without fat sat) at 3mm fixed 
slice thickness (acquisition will be performed at 6mm thickness and then the slice will be 
interpolated at 3mm, i.e., in GE, ZIP =1 in the slice direction). Keep the flip angle ≤15o. To improve 
SNR, keep the Bandwidth low (62 kHz or 42 kHz) and/or increase the number of phase-encoding 
steps (be aware, the acquisition time will increase). In GE LAVA sequence, turning off “optimize flip 
for CNR” will allow to change the flip angle or bandwidth. Do NOT use parallel imaging (no SENSE, 
ASSET, iPAT or GRAPPA).  
 
9. (Entire Abdomen covering kidney and liver) Breath-hold coronal 2D true-FISP (FIESTA) without fat 
sat at 3mm fixed slice thickness, which may require 2-3 breath-holds depending for the size of the 
kidney and liver. Use as few breath-holds as possible. The first scan should cover the posterior 
aspect of the kidney. The last scan covers the anterior liver. No incomplete coverage or wrap of 
anterior liver onto posterior kidney. Neighboring image groups should be overlapped by a single 
3mm slice. To determine correct table position choose the “shift-mean (starting point in GE)” of the 
second scan for example: the first shift-mean = -60mm, the number of slices in the first set =23, (23-
1)x3=66mm, new shift mean =-60+66=6mm.  
 

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10. (For Liver and Liver cyst volume)  Breath-hold coronal T2 scan (SSFSE/HASTE) without fat sat of the 
kidneys with adjusted slice thickness, 3-6 mm, i.e. the slice thickness best attainable with a single 
breath-hold (The adjusted slice thickness may not remain the same in a follow-up MR scan if there is 
a change in the subject's breath-hold capacity or kidney size.) Repeat the scan over the liver with the 
same slice thickness. This scan and the scan for the kidney should share one overlapping liver slice 
(i.e., the most posterior slice of the liver scan should be identical to the most anterior slice imaging 
the liver in the kidney scan. If more than two scans are required to cover the anterior liver, again the 
neighboring scans should be overlapped by one slice. 
 
 
After Image acquisition, images will be transferred to the Coordinating Center at University of Pittsburgh 
via CD-ROM.  At each clinical site, a staff radiologist with expertise in abdominal MRI will review each 
scan for unexpected findings.  In the event of such findings, the site investigators will notify the 
participant and –with the participant’s permission – the participant’s physician.    
 
 
 
 
 
 
 
 

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TAME –PKD Statistical Analysis Plan 
Study Design & Objectives. The design of the proposed study is a double-blind, multicenter parallel-
group randomized clinical trial. The scientific aims are as follows: to assess tolerability and safety of 
treating ADPKD patients with early disease (eGFR ≥50 ml/min) with metformin (Aim 1) and to evaluate 
potentially relevant biomarkers that correlate with ADPKD disease severity and response to metformin. 
This study will be conducted at 2 institutions in the United States (Tufts University and University of 
Maryland), and we will enroll adult patients, ages 18-60 years with ADPKD, and eGFR ≥50 
ml/min/1.73m2.  
 
Aim 1: We hypothesize that metformin, an agent that is widely used to treat type 2 diabetes mellitus 
and polycystic ovary syndrome, may serve as a novel therapy for individuals with early stage autosomal 
dominant polycystic kidney disease (ADPKD) by activating the metabolic sensor AMPK. To test this 
hypothesis, we propose a pilot-level double-blind, phase 2, parallel-group, placebo-controlled 
multicenter clinical trial to evaluate the safety and tolerability of metformin in ADPKD patients over a 
two-year period. In addition, we will evaluate the effect of metformin therapy on the rate of change in 
height-adjusted total kidney volume (htTKV) and estimated glomerular filtration rate (eGFR) as well as 
on metabolomic biomarkers of disease severity and progression.  
 
Aim 2: We hypothesize that metformin treatment will ameliorate ADPKD severity and progression in 
patients and that this beneficial effect occurs in large part through changes in the cellular bioenergetics 
and proliferative capacity of ADPKD kidney epithelial cells. We will investigate candidate metabolic 
biomarkers in urine based on the work of Boletta and colleagues and others (22). We hypothesize that 
glycolytic pathway enzymes will be progressively increased and markers of AMPK pathway activation 
will be progressively reduced with worsening ADPKD severity in patients. We further hypothesize that 
treatment of patients with metformin will tend to reverse these effects, which are indicative of the 
metabolic disturbance in ADPKD, and potentially thereby represent a beneficial therapeutic effect. 
 
Sample Size Calculation. We anticipate enrolling 48 participants per study arm with 15% attrition, 
resulting in an effective sample size of 40 per arm. We are not powered to detect meaningful 
differences in efficacy, but instead focus on a 95% confidence interval width (two times the margin-of-
error) for relevant point estimates within each study arm. For the primary safety outcome of SAEs, we 
will have the ability to estimate a CI width no larger than 0.16, assuming the proportion of participants 
who experience at least one SAE is no more than 6% (similar to the highest rates in the HALT trial 
(Schrier et al., 2014)). For the primary tolerability outcome of GSRS, we will have the ability to estimate 
a CI width no greater than 21, assuming standard deviations of the GSRS are below 1.3. With regard to 
the tolerability outcome of drug discontinuation, our anticipated sample size will allow the ability to 
estimate a CI width no larger than 0.25, assuming the rate of drug discontinuation is 15% (similar to 

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TEMPO (Torres et al., 2012)). For the efficacy outcome of TKV, we assume the participants taking 
placebo will see a 5.51% annual increase in TKV (as seen in TEMPO (Torres et al., 2012)) with standard 
deviations for the slope and error of 0.041 and 0.044 (on the natural log scale), respectively.  With our 
anticipated sample size, we will be able to estimate a margin-of-error of 1.5 percentage points for 95% 
intervals.  
 
Interim & Final Analyses.  We will not have any planned interim looks for stopping for efficacy, although 
safety and tolerability data will be monitored by the study DSMB at each meeting. The final analyses will 
be conducted once study follow-up is complete, after all data is cleaned, and once the study database is 
locked. We anticipate database lock will happen in September of 2020 with completion of primary 
analyses occurring in October of the same year. 
 
Hypotheses. The primary study hypotheses are:  
1) compared to placebo, participants taking metformin (titrated to 1000 mg twice daily or 
maximal tolerated dose) will have a better safety profile over 24 months  
2) compared to placebo, participants taking metformin will have better tolerability, defined by 
change in the Gastrointestinal Symptom Rating Scale (GSRS) over 24 months.  
The secondary hypotheses are:  
3) compared to placebo, participants taking metformin will have a smaller annual % increase in 
height-adjusted total kidney volume (htTKV) over 24 months 
4) compared to placebo, participants taking metformin will have a slower annual rate of decline 
in estimated glomerular filtration rates (eGFR) over 24 months. 
 
Analysis Sets. The full analysis set will be based on an intention-to-treat (ITT) analysis with a per-
protocol (PP) sensitivity analyses. The ITT analysis set will comprise all participants who have been 
randomized to either study arm, regardless of length of follow-up or actual intervention received. The 
PP analysis set will include participants who have “adhered” to study intervention, in this case defined 
as all participants who have successfully completed drug titration (completed at least F3 visit) with a 
maximum tolerate dose. The safety analysis set will be a subset of the ITT set and include all participants 
who have at least received and taken metformin or placebo.  
 
Endpoints & Covariates. The primary outcomes are the safety and tolerability of using metformin in 
ADPKD patients with baseline eGFR ≥50 ml/min. 

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a. Assessment of Tolerability: Tolerability will be assessed using two standardized patient-reported 
instruments that will be administered at baseline, at each telephone visit during titration, and at 
every PCC visit. The most anticipated adverse effects of metformin are gastrointestinal (GI)-
related. Accordingly, we will use the GSRS instrument to assess GI symptom burden. A widely 
used and validated 15-item questionnaire, the GSRS instrument has been used to compare the 
GI tolerability of different mycophenolate preparations in renal transplant recipients (39). The 
second assessment of tolerability will be based on responses (yes or no) to the following 
question “Can you tolerate this dose of the study drug for the rest of your life?” As described in 
Section 6e (Drug titration), a negative response will lead to down-titration of drug.  
b. Safety Outcomes: At each study visit (telephone or PCC), patients will be asked if they had an 
interim hospitalization or medical encounter and if so, the nature of the event. The patient will 
sign a release of medical information form at the baseline visit and this will be used to obtain 
discharge summaries or other clinical documentation to determine the nature and severity of 
adverse events. Serious adverse events (SAE) will be defined as an undesirable experience 
occurring from the time a participant signs the informed consent (at the screening visit) until the 
end of the study, meeting 1 or more of the criteria of: 1) Resulting in death, 2) Non-elective 
hospitalization, 3) Life threatening (if patient continued on study drug would result in death), 4) 
Persistent or permanent harm or disability, 5) Exceeding the nature, severity or frequency of risk 
described in the protocol or 6) resulting in Congenital anomaly. Reporting requirements to the 
DCC and IRBs are described in Section 12 (Safety). A symptoms checklist (which lists the more 
common or concerning side-effects of the study drug, but allows for free text entry of other 
symptoms) will be collected at baseline and at each subsequent PCC visit. Hypoglycemia is a 
potential complication of metformin use in a non-diabetic population.  
c. Adherence: Adherence with study medication will be assessed through pill counts, which will be 
performed by the study coordinator at each study visit. Patients that miss their in-person visit 
and thus do not refill their study drug will be contacted in a timely fashion by the study 
coordinator, who will have a record of the missed visit. Adherence with study visits will be 
assessed by tracking the number of missed study visits.   During the drug titration telephone 
visits and the F1 in-person visit, the missed study visit window will be +/- 1 day; for subsequent 
visits a missed study visit will be documented as +/- 6 days.   Practical measures to minimize 
inconvenience (parking, stipends if possible), maintaining communication with referring 
physicians, and other means of maintaining direct communication with the patient (follow-up 
and thank you cards after visits, birthday and holiday cards, small gifts) should also promote 
adherence. Finally, regular and frequent follow-up visits should also aid in the retention of study 
patients. 
d. Patient-reported Outcomes: The Medical Outcomes Short Form 36 (SF-36) questionnaire is the 
most widely used instrument for measuring health-related quality of life (HRQOL), though scores 
have not been shown to discriminate among ADPKD patients with larger vs. smaller kidneys at 
eGFR >60 ml/min (40). The TAME Pain Questionnaire is a modified version of the Wisconsin 
Brief Pain Questionnaire (41) that measures the frequency and intensity of pain and symptoms 
relating to organ enlargement as well as their impact on everyday living. It was validated in a 
clinical population of ADPKD patients from a single center observational study and subsequently 
adapted for use in the HALT Study (42). The frequency of back pain and symptoms related to 
abdominal distension correlated with htTKV in a cross-sectional analysis of baseline 
questionnaire results in the HALT-PKD Study A cohort, which is a population with similar eGFR 
range as expected in the proposed study (43). GI symptoms from metformin may affect HRQOL, 
and thus we will also examine this at every visit. The GSRS (a widely-used, validated 15-item 
questionnaire, described earlier) and the SF-36 (as a generic measure of HRQOL) will be 

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administered by study staff (39, 44). The questionnaires will be administered at baseline and at 
in-person follow-up visits.  In addition, on telephone visits during drug titration, the GSRS will be 
asked to evaluate common gastrointestinal symptoms. Each question is rated on a seven-point 
Likert scale, from no discomfort to very severe discomfort. For the GSRS the responses on the 15 
items are reduced to average scores on 5 specific scales (diarrhea, reflux, constipation, 
abdominal pain, indigestion), identified from factor analysis. Among U.S. patients with gastro-
esophageal reflux, internal reliability of GSRS scores on the outcomes that may be affected by 
metformin use was moderate to high (Cronbach’s alpha: 0.61 to 0.83), and the instrument 
displayed good construct validity (45).  
  
 Handling of Missing Values. As a preventive measure, we will make every attempt to document all 
reasons for missing data. In addition, baseline characteristics will be compared between participants 
who do and do not withdraw from the study as a way to assess the impact of missing information and 
attrition. We will also compare the rates of lost-to-follow-up (LTF) between study arms. 
 
Our general assumption will be that missing data is of the Missing at Random (MAR) mechanism. In this 
case, the use of linear mixed models for the primary analysis will be sufficient in reducing the impact 
that missing data has on biasing the primary results. Additionally, we will conduct a sensitivity analysis 
using multiple imputation via chained equations (MICE) to see how robust the overall inferences are.  
 
There is the potential to have non-ignorable missingness (NMAR) on key secondary outcomes of htTKV 
and eGFR if there is substantial LTF/dropout or death. We will assess this by using shared parameter 
models to jointly estimate annual rate of change and time-to-event. 
 
Statistical Analyses. All primary and secondary outcomes will be described using sample means or 
sample proportions along with 95% confidence intervals, depending on the nature of the outcomes. 
Volume outcomes from MRI (i.e. total kidney, kidney cyst, liver, liver cyst) that are known to be highly 
skewed will be assessed to see if suitable transformations are needed (i.e., natural log). Participant 
demographics and baseline clinical characteristics will be compared between study arms (Metformin 
versus Placebo) using two-sample t-tests or chi-squared tests of independence. All primary outcomes 
will be analyzed under intent-to-treat. 
 
The primary objective of this phase II, multicenter clinical trial is to evaluate the safety and tolerability of 
metformin versus placebo with a secondary emphasis on efficacy. The safety outcome is defined by the 
occurrence of adverse events and serious adverse events (as defined in Section 11b) throughout the 
study. We will calculate cumulative incidence between study arms and use logistic regression to assess 
the association between study arms and the proportion of participants affected across various SAEs. 

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Tolerability will be defined by the GSRS scale. Using a linear mixed model, we will compare the change in 
overall GSRS score as a function of time (months since baseline), the interaction between time and study 
arm, and clinical site. We will also include a random intercept to allow for participant-level variability of 
baseline GSRS. Of primary interest is whether the interaction terms are significant, which would indicate 
the change over time differs between the metformin and placebo arms. We will compare drug 
discontinuation rates between study arms using Chi-squared tests of independence. Adherence (defined 
as >=80% of proportion of pills taken) will be compared between study arms using Chi-squared tests of 
independence. In addition, we will also utilize Kaplan-Meier curves, along with logrank tests for 
significance, to compare time-to-drug discontinuation between arms. 
 
Secondary analyses: The annual rate of change in height-adjusted total kidney volume (htTKV) will be 
compared between Metformin and Placebo arms to give an indication of the efficacy of metformin. A 
Laird and Ware linear mixed model (51) will be fit with the natural log of htTKV (LnTKV) as a function of 
time, the interaction between time and study arm, and clinical site. In order to account for the 
participant-level variabilities of baseline LnTKV as well as rate of change, the model intercept and slope 
will be allowed to vary if needed. A significant interaction between time and study arm could indicate a 
slowing of PKD progression due to metformin. This same model will be used to evaluate secondary 
efficacy outcomes such as kidney cyst volume (KCV), total liver volume (TLV), and liver cyst volume 
(LCV). Renal function, as measured by eGFR, will be compared between metformin and placebo using a 
similar Laird & Ware model as above. The TAME-PKD Pain questionnaire has over 50 questions. A priori 
we identify the following items of the TAME PKD Pain questionnaire as being most likely to differ across 
study arms after 2 years of study drug treatment: back pain frequency, impact of pain on physical 
activity and sleep, and abdominal distension symptoms. The response to each of these questions is a 5-
point Likert scale. The sample size is small in the present study and we are likely to have low frequencies 
in each of the 5 categories. Thus, we will group the lowest 3 levels together with the top 2 levels when 
comparing these items across study arms. 
 
Exploratory Aim 2: The primary goal is to evaluate potential biomarkers that correlate with ADPKD 
disease severity and response to metformin. Our focus will be on both prognostic as well as predictive 
biomarkers of total kidney volume and eGFR. The analysis for prognostic biomarkers will be restricted to 
participants taking placebo and will utilize a Laird & Ware linear mixed model with the following 
predictors: time (months since baseline), a time-dependent covariate for the biomarker of interest, their 
interaction, and random effects for the intercept and slope. Apart from the ability to see whether a 
particular biomarker at baseline is related to disease severity and progression, we will also be able to 
estimate cross-sectional and longitudinal effects on TKV and eGFR among untreated participants. The 
analysis for predictive biomarkers will utilize the same mixed model with the following predictors: time, 
time-by-study arm interaction, baseline biomarker of interest, time-by-biomarker interaction, and the 
three-way interaction of these predictors. This model will give us the ability to identify subgroups of 
participants who may or may not benefit from metformin therapy. 

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