gout and uric acid: from disease to transporters to therapy · gout: diagnosis and management of...

Jeffrey N. Miner, PhD Executive Director Biology Ardea Biosciences, Inc., a wholly-owned subsidiary of AstraZeneca PLC

Gout and Uric Acid: From Disease to Transporters to Therapy

Disclosures/Conflict of Interest

• Previous employee of Ardea Biosciences, Inc., a wholly-owned subsidiary of AstraZeneca PLC

• Member of ARTA Bioscience Advisory Board

• Funding for research was provided by Ardea Biosciences/AstraZeneca

• Editorial support was provided by Albert Balkiewicz, MSc, an employee of PAREXEL, and was funded by Ardea/AstraZeneca

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Proprietary & confidential © AstraZeneca 2014. For internal discussion only

History of Selective Uric Acid Reabsorption Inhibitor (SURI) Discovery and Development

FEUA=fractional excretion of uric acid.

• Ardea was a small biotech company conducting early trials in Oncology and Antivirals

• Phase 1 antiviral clinical trial - Treated patients were exhibiting unusual, very significant decreases

in serum uric acid levels • Higher FEUA – affecting uric acid excretion

• One metabolite was highly excreted to urine; this metabolite was purified

• It was a URAT1 inhibitor – a renal uric acid transporter important for maintaining uric acid levels in humans

• Lesinurad was approved in 2015 for the treatment of gout • Verinurad is a second generation compound in development

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Production and Excretion of Uric Acid (What Is Uric Acid?)

Fam AG. J Rheumatol. 2002;29:1350‒5. Keenan RT, et al. Chapter 94. In: Firestein GS, et al. (eds). Kelley's Textbook of Rheumatology (Ninth edition). Philadelphia: Elsevier Saunders, 2013. Rock KL, et al. Nat Rev Rheumatol. 2013;9:13‒23. So A, Thorens B. J Clin Invest. 2010;120:1791‒9. Terkeltaub R, Edwards NL. Chapter 1. In: Terkeltaub R, Edwards NL (eds). Gout: Diagnosis and Management of Gouty Arthritis and Hyperuricemia (Third edition). Oklahoma: Professional Communications, Inc, 2013.

N H

N H

N HN

O

O

Uric acid

Purine Hypoxanthine Xanthine

Xanthine oxidase

Xanthine oxidase

Overall uric acid excreted via gut: 30%

Urate reabsorbed

(90%)

Kidney Gut

Humans have high serum uric acid 4.0–6.0 mg/dL

Exogenous (dietary) purines

(~33%)

Endogenous purines (~66%)

Dietary purine

absorption

Urate excreted (10%)

Overall uric acid excreted

via kidney: 70%

N H

N

N HN

O

N H

N

N N

N H

N H

H N HN

O

O

O

4

Uricase mutations

Humans Have Higher sUA Levels Than Most Other Animals

• Great apes and humans lack uricase1,2

1. Oda M, et al. Mol Biol Evol. 2002;19:640–53. 2. Johnson RJ, et al. Semin Nephrol. 2005;25:3–8. 3. Fang J, Alderman MH. JAMA. 2000;283:2404‒10.

0.4 0.8 0.7

1.9 1.9

2.7

4.0

5.5

0.0

1.0

2.0

3.0

4.0

5.0

6.0

Lemur(n=16)

Langur(n=6)

Guenon(n=11)

Orangutan(n=7)

Gorilla(n=7)

Bonobo(n=10)

Americanwomen

Americanmen

Mea

n sU

A (m

g/dL

)

5

1Graph shows mean sUA levels of various primates obtained from the San Diego zoo. 2The values for mean sUA in American men and women were obtained from the NHANES study. 3NHANES. National Health and Nutrition Examination Survey.

Conversion of Tyrosine-365 (Y) to Phenylalanine-365 (F) in URAT1 Occurred Relatively Late in Evolution

Species AA 365: F/Y Urate transport Km (µM)

Rat Y 857*

Baboon F 79

Human F 122

Statistical significance compared to human URAT1 was determined using the unpaired Student’s t-test * P < 0.0001.

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Gain of High URAT1 Uric Acid Affinity Occurred at Same Time in Evolution as Loss of Uricase Activity

Tan PK, Farrar JE, Gaucher EA, Miner JN. Coevolution of URAT1 and Uricase during Primate Evolution: Implications for Serum Urate Homeostasis and Gout. Mol Biol Evol. 2016;33(9):2193−200.

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How did we know to examine the evolution of amino acid 365? - Mapping inhibitor binding site on URAT1

Conclusions

• Multiple proteins changed function to accommodate higher levels of uric acid

• There was selective pressure for this advance, and gout in humans was an unintended consequence

• Many hypotheses exist, though empirical data favors the notion that uric acid enhances the ability to store energy (fat) when living in a limited resource environment

Aegyptopithecus

The Missing Link to Gout?

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Disadvantages of Higher sUA: Prevalence of Comorbidities Rises With Increasing sUA

31.9

42.7

49.2

52.5

58.2

66.4

74.0

85.9

16.9

22.7

30.2

34.4

41.4

50.0

67.2

66.0

16.9

24.1

34.0

41.2

46.3

52.9

56.6

65.3

0

20

40

60

80

100

<4.0 4.0–4.9 5.0–5.9 6.0–6.9 7.0–7.9 8.0–8.9 9.0–9.9 ≥10.0

Prev

alen

ce o

f com

orbi

ditie

s (%

)

sUA (mg/dL)

CKD stage ≥2 HypertensionObesity

Figure indicates prevalence of comorbidities according to sUA level in participants with and without hyperuricaemia from the NHANES 2007–2008. CKD, chronic kidney disease; NHANES, National Health and Nutrition Examination Survey.

9 Zhu Y, et al. Am J Med. 2012;125:679–87.

Asymptomatic Hyperuricemia: MSU Crystal Accumulation/Deposition

• MSU crystal deposition occurs prior to clinical manifestations of gout1

• Deposition is greater in patients with asymptomatic hyperuricemia than with normal uric acid levels2

DECT, dual energy computed tomography; MSU, monosodium urate; MTP, metatarsophalangeal.

DECT scan showing MSU crystal deposition (green) surrounding the right first MTP and along left midfoot in a patient with asymptomatic hyperuricemia3

10 10

1. Terkeltaub R, Edwards NL. Chapter 3. In: Terkeltaub R, Edwards NL (eds). Gout: Diagnosis and Management of Gouty Arthritis and Hyperuricemia. (Third edition). Oklahoma: Professional Communications, Inc, 2013.

2. Pineda C, et al. Arthritis Res Ther. 2011;13:R4. 3. Nicolaou S, et al. AJR Am J Roentgenol. 2012;199(5 Suppl):S78‒86.

Gout – A Urate Crystal Deposition Disease

Taylor JW, Grainger R. Chapter 9. In: Terkeltaub R (ed). Gout and Other Crystal Arthropathies (First edition). Philadelphia: Elsevier Saunders, 2012. 11

Inefficient Renal Excretion of Uric Acid (Low FEUA) Is a Characteristic of Gout Patients1

Frac

tiona

l Exc

retio

nof

Uric

Aci

d (F

EUA,

%)

Healthy Gout Patients0

5

10

15

P<0.001

• Data from Phase 1 clinical trials

Choi et al. Ann Intern Med. 2005,143:499−516.

FEUA, Fractional excretion of uric acid.

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Distal Tubule

Proximal Tubule

Vasa Recta

Glomerulus

Renal Transport and Excretion of Urate

1. Bobulescu IA et al. Adv Chronic Kidney Dis. 2012;19:358-371. 2. Pittman JR et al. Am Fam Physician. 1999;59:1799-1806. 3. Ichida K et al. Nat Commun. 2012;764:doi:10.1038ncomms1756. 13

Renal Transport and Excretion of Urate

• Urate is completely filtered through the glomerulus into the proximal tubule1,2

Distal Tubule

Glomerulus

Vasa Recta

Urate

Filtration

Proximal Tubule

1. Bobulescu IA et al. Adv Chronic Kidney Dis. 2012;19(6):358−371. 2. Pittman JR et al. Am Fam Physician. 1999;59(7):1799−1806. 14

Distal Tubule

Proximal Tubule

Reabsorption

Secretion

Vasa Recta

ABCG2

urate

Lumen Blood URAT1

(SLC22A12)

urate

GLUT9 (SLC2A9)

SECRETION

REABSORPTION

OAT4 (SLC22A11)

OAT2 (SLC22A7)

NPT1 (SLC17A1)

• URAT1 is a major transporter in the tubule

system responsible for uric acid reabsorption and maintaining overall uric acid homeostasis3,4

1. Bobulescu IA et al. Adv Chronic Kidney Dis. 2012;19:358-371. 2. Pittman JR et al. Am Fam Physician. 1999;59:1799-1806. 3. Crittenden DB, et al. Bull NYU Hosp Jt Dis. 2011;69:257-263. 4. Enomoto A et al. Clin Exp Nephrol. 2005;9:195-205. 15

Renal Transport and Excretion of Urate

Excretion

Distal Tubule

Glomerulus

Vasa Recta

Proximal Tubule

Renal Transport and Excretion of Urate

• Uric acid that is not reabsorbed by the kidneys is excreted in the urine – Of uric acid excreted,

70% is renal and 30% is intestinal

Reabsorption

Secretion

So and Thorens. J Clin Invest. 2010;120:1791. 16

Glomerulus

Distal Tubule

Proximal Tubule

Vasa Recta

Filtration

Renal Transport and Excretion of Urate

• Normal uric acid excretion is approximately 10% of the filtered load1

Urate

Reabsorption Secretion

Excretion

NORMAL URIC ACID EXCRETION

Richette P, Bardin T. Lancet. 2010;375:318−328.

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Excretion

Distal Tubule

Glomerulus

Proximal Tubule

Reabsorption

Secretion

Gout: Renal Transport and Excretion of Urate

• In patients with gout, increased reabsorption (via URAT-1, etc) occurs, resulting in under-excretion of uric acid (~5% of filtered load)1

ABCG2

urate

Lumen Blood URAT1

(SLC22A12)

urate

GLUT9 (SLC2A9)

SECRETION

REABSORPTION

OAT4 (SLC22A11)

OAT2 (SLC22A7)

NPT1 (SLC17A1)

Richette P, Bardin T. Lancet. 2010;375:318−328. 18

Glomerulus

Distal Tubule

Proximal Tubule

Vasa Recta

Filtration

Renal Transport and Excretion of Urate

• Inhibiting the mechanisms associated with reabsorption can increase uric acid excretion1

Urate

Reabsorption Secretion

Excretion

GOUT PATIENT URIC ACID EXCRETION

URAT-1 inhibition

Probenecid Benzbromarone

Lesinurad Verinurad

Arhalofenate

Terkeltaub R et al. Arthritis Res Ther. 2006;8(Suppl 1):S4. 19

Verinurad (RDEA3170) A Highly Potent ULT in Phase II • Potent uric acid lowering potential • Potential for substantial uric acid lowering with low-dose

combinations compared to high dose (up-titrated xanthine oxidase inhibitor)

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Verinurad

URAT1 Phenylalanine 365 Is Crucial for High Affinity Verinurad Interaction

Human URAT1 Rat URAT1 (rRST)

hURAT1-rRST TM7 rRST-hURAT1 TM7

hURAT1-F365Y rRST-Y365F

0.025 34.0

Verinurad IC50 (µM)

2.3 4.3 2.7 F

Y

TM 1 2 3 4 5 6 7 8 9 10 11 12

Loops EC loop 1 IC loop 3

4.5

Y365F is a gain of function mutation (10 X increased affinity)

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[Verinurad]

Human URAT1 Residues 35, 365, and 481 Interact to Enhance Affinity for Verinurad

verinurad IC50 (µM, mean ± SEM)0 5 10 15

human URAT1r-N35S/Y365F/M481I

r-Y365F/M481Ir-N35S/M481Ir-N35S/Y365F

r-M481Ir-Y365Fr-N35S

rat URAT1

40 45

IC50 (µM)41102.911

0.474.430.590.140.025

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O

O OH

Br

Br

benzbromarone

O OH

SO O

N

probenecid sulfinpyrazone

N

N

S OHO

Verinurad (RDEA3170)

NNN

SOH

OBr

lesinurad

URAT1 Inhibitors

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URAT1 Binding Assay Demonstrates Competitive Binding by Inhibitors

• Bind 3H-verinurad to membranes from HEK-293T cells expressing URAT1

• 3H-verinurad binding to URAT1 is… - Displaced by URAT1 inhibitors, at potencies similar to the inhibition

of URAT1 transport activity

- Specific and high affinity

1. Tan et al. [abstract]. Arthritis Rheum. 2011; 63 (Suppl10):1592; 2. Tan et al. [Abstract]. J Am Soc Nephrol .2011; 22:754A; 3. Miner and Tan. Ann Rheum Dis. 2012;71(Suppl 3):446.

[compound] log M

% 3

H-RD

EA31

70 b

ound

-10 -8 -6 -4 -20

20406080

100120

RDEA3170 (unlabeled)benzbromaroneprobenecidsalicylate

%

30

% 3 H

-Ver

inur

ad b

ound

Verinurad (unlabeled)

Differential Importance of Individual URAT1 Residues Is Due to Alterations in Binding Affinity

28

verinurad

verinurad

verin

urad

Phe-365 and Other Residues That Interact With Inhibitors Project Within the Transporter Channel

Phe-365

Ile-481

Ser-35

Images of human OAT1 3-dimensional model1 with the cytoplasmic surface facing the viewer – equivalent URAT1 residues highlighted

1. Perry et al. J Biol Chem. 2006;281:38071-38079. 28

Conclusions

• Multiple uricosuric agents compete for direct interaction at a single site on URAT1

• Binding pocket is likely centrally located within the core of the transporter

• Amino acids within the binding pocket have differing effects on the affinity of different uricosuric agents

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Verinurad lowers serum uric acid (sUA) by increasing fractional excretion of uric acid (FEUA) in humans

Time post dosing (hr)

% c

hang

e in

sUA

(mea

n±

SEM

)

0 6 12 18 24-80

-60

-40

-20

0

Weighted average plasma verinurad (log nM)

FEU

A (%

)

-1 0 1 2 3

0

10

20

30

40

25

A single 40 mg dose of verinurad lowered baseline sUA levels by up to 60% for

a sustained time period

Verinurad increased the FEUA in a dose-dependent manner, with a half-maximal effective plasma

concentration of 0.022 µM (22 nM)

Experiments were performed in healthy human volunteers.

Study 204 (Phase 2a, US): PK/PD Study of Verinurad in Combination With Febuxostat

FBX, febuxostat; PD, pharmacodynamic; PK, pharmacokinetic; QD, once daily; ULT, urate-lowering therapy.

Cohort 1 (verinurad 10 mg): PK data available for 13 subjects; PD data available for 10–11 subjects Cohort 2 (verinurad 15 mg): PK/PD data available for 12 subjects Cohort 3 (verinurad 5 mg): PK/PD data available for 12 subjects Cohort 4 (verinurad 2.5 mg): PK/PD data available for 8–9 subjects Cohort 5 (verinurad 10, 15 or 20 mg): Adaptive design (n=11)

POPULATION Gout subjects with sUA ≥8.0 mg/dL, who are either ULT-naïve or have previously received ULT, N=64

PRIMARY OBJECTIVES

• To assess the PK and PD effect of verinurad + febuxostat • To evaluate the safety and tolerability of verinurad + febuxostat

≥14 days Day 1–Day 7 Day 8–Day 14 Day 15–Day 21 Day 22−Day 28

Colchicine 0.6 mg QD

Cohorts 1–4

Washout period*

RAN

DO

MIZ

E FBX 40 mg

verinurad + FBX 40 mg

verinurad + FBX 80 mg

FBX 80 mg

FBX 80 mg

verinurad + FBX 80 mg

verinurad + FBX 40 mg

FBX 40 mg

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*Washout period of at least 14 days for subjects who have previously received ULT.

Study 204 (Phase 2a, US): sUA Lowering With Verinurad + Febuxostat

• There was a consistent dose trend between 40 mg and 80 mg febuxostat in the range of 2.5–10 mg verinurad

FBX, febuxostat; sCr, serum creatinine.

Maximum sUA Percentage Change

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Study 204 (Phase 2a, US): Urinary Urate Excretion With Verinurad + Febuxostat

FBX, febuxostat; sCr, serum creatinine; VERU, verinurad.

Urinary Urate Excretion

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• Urinary urate excretion with verinurad + febuxostat was comparable to or lower than baseline at all time points

• No sCr elevations ≥1.5 x baseline were observed

Summary

• Inhibition of URAT1 by verinurad is dependent on high affinity interaction with F365, S35 and I481

• Verinurad, a SURI that inhibits URAT1, directly improves poor fractional excretion of uric acid in gout patients

• A medical need exists for effective and well tolerated URAT1 inhibitors for combination therapy

• Verinurad is a potential new therapy for hyperuricemia associated with gout used in combination with a xanthine oxidase inhibitor (e.g., allopurinol, febuxostat)

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