ACR Meeting November, 2012

Transcription

1 ACR Meeting November, 212

2 Arhalofenate is a Novel Dual-Acting Agent with Uricosuric and Anti-Inflammatory Properties Yun-Jung Choi, Vanina Larroca, Annette Lucman, Vic Vicena, Noe Abarca, Tim Rantz, Brian E. Lavan and Charles A. McWherter Metabolex Inc. Hayward, CA

3 Disclosures All Authors, Including Presenting Author, Have the Following Financial Relationships to Disclose: All authors are current employees of Metabolex, Inc.

4 Key Features of Gout: Hyperuricemia Causes Urate Crystal Deposits That Produce Gout Flares Mono sodium urate (MSU) crystal deposits Hyperuricemia Serum uric acid (sua) Therapeutic targets Inflammatory response IL-1β Joint erosion Painful gout flare 4

5 Arhalofenate Acid has Two Activities in a Single Molecule 1% Bioconversion Arhalofenate Arhalofenate acid Blocks renal tubular urate re-absorption URAT1, OAT4 and OAT1 Blocks MSU production of pro-il-1β mrna PPAR-γ trans-repression Gregoire et al, Mol. Endocrinol :

6 Renal Urate Transporters Implicated in the Re-absorption of Urate in Proximal Tubules Apical Side (Urine) URAT1 OAT1 OAT4 GLUT9b Urate Pyrazinoate Nicotinate Lactate Urate Thiazide diuretics Salicylates Glucose Urate GLUT9a Proximal Tubule Epithelial Cell Urate Basolateral Side (Blood)

7 [ 14 C] Urate Uptake (%Control) [ 14 C] Urate Uptake (%Control) [ 14 C] Urate Uptake (%Control) Arhalofenate Acid Blocks 14 C-Uric Acid Uptake by Human Renal Transporters 12 1 URAT OAT Arhalofenate Acid (M) Arhalofenate Acid (M) OAT1 Arhalofenate acid concentration in human urine was 116 ± 23 µm after dosing at 6 mg Arhalofenate Acid (M)

8 Comparison of Arhalofenate Acid with Other Uricosuric Agents IC 5 (μm) for 14 C-Uric Acid Uptake IC 5 (µm) URAT-1 OAT4 OAT1 Arhalofenate acid Lesinurad Probenecid Benzbromarone cdna encoding human renal transporters were transiently transfected into human embryonic kidney cells. Compound effects on 14 C-uric acid uptake was assessed in comparison to mock transfected parental cells 8

9 MSU Crystal Triggered Inflammation in Joints Arhalofenate selectively blocks the first step in IL-1β cascade MSU Crystals Rilonacept/ Canakinumab Neutrophil Colchicine NALP3 Inflammasome NF-κB genes Macrophage Arhalofenate Acid Inflammation/ Acute Flare Colchicine Joint Space 9 Interleukin 1b CXCL1/CXCL2 Prednisone/ NSAIDS Neutrophil Neutrophil Activation and Degranulation

10 Murine Air Pouch Model of MSU Induced Inflammation 6 day Air Pouch Established 24 hrs 24 hrs 3 min 4 hrs Dose 1 Dose 2 Dose 3 MSU Collect Exudate Drugs administered Arhalofenate 25 mg/kg Dexamethosone 2 mg/kg Colchicine 2 mg/kg Cytokines measured in exudate by ELISA assay IL-1b CXCL1 IL-6 MCP-1 CXCL2 Neutrophil, macrophage and total cell numbers in exudate measured by FACS analysis 1

11 Arhalofenate Potently Inhibits MSU Crystal Induced IL-1β Production in the Murine Air Pouch Model 15 + MSU I L 1 - b ( p g / m L ) 1 5 * ns No MSU Vehicle Arhalo Dexa Colchicine

12 M C P - 1 ( p g / m L ) I L - 6 ( p g / m L ) C X C L - 1 ( p g / m L ) C X C L - 2 ( p g / m L ) Arhalofenate Also Blocks Production of Other NF-κB Inflammatory Mediators in the Murine Air Pouch Model 6 CXCL1 2 CXCL * ** 1 5 *** ** 1 MCP-1 5 IL * ** *** No MSU MSU Arhalo Dexa Colchicine

13 C e l l n u m b e r ( x 1 3 ) C e l l n u m b e r ( x 1 3 ) C e l l n u m b e r ( x 1 3 ) Arhalofenate Potently Inhibits MSU Crystal Induced Inflammatory Cell Production in the Murine Air Pouch Model Total Cells Neutrophils Macrophages (CD45+) (Ly-6B.2+) (F4/8+) ** ** *** *** No MSU MSU Arhalo Dexa Colchicine

14 A Model for Triggering of Gouty Inflammation via Macrophage Toll-Like Receptors Resolution FFA TLR2 TLR4 LPS MSU Crystals 14 NFκB genes: e.g. Pro-IL-1β NALP3 Inflammasome Activation Macrophage cf Liu-Bryan et al, Arth Rheum: 25: 52:2936 Martinon. et al, Nature 26: 44: 237 Interleukin 1b Interleukin 18 CXCL1 CXCL2/IL-8 IL-6 MCP-1 TNF-α + others TGF-β1 Gout attack Monocytes Mast cells Neutrophils

15 I L 1 - b ( p g / m L ) MSU and LPS Synergize in Producing IL-1β Culture 2 days 24 hours MSU or LPS or LPS + MSU Measure IL-1 b cf Petska. et al, Tox. Sci. 26: 92: 445 Giamarellos-Bourboulis, et al, Ann Rheum Dis 29: 68:273 15

16 I L 1 - b ( p g / m L ) I L 1 - b ( p g / m L ) Arhalofenate Acid Suppresses IL-1b in Isolated Mouse Macrophages Comparison to Dexamethasone and Colchicine Culture 2 days 2 hours 24 hours Compound Treatment MSU + LPS Measure Secreted IL-1 b Arhalofenate Acid Dexamethasone Colchicine I L 1 - b ( p g / m L ) MSU + LPS MSU + LPS +Arhalofenate Acid MSU + LPS + Dexamethasone MSU + LPS + Colchicine 16

17 Arhalofenate Acid Inhibits IL-1β Release in Isolated Macrophages by Reducing Levels of Intracellular Pro-IL-1β Culture 2 days 2 hours 24 hours I L 1 - b ( p g / m L ) Arhalofenate LPS or MSU or LPS + MSU Secreted IL-1β Measure Secreted and Intracellular IL-1 b MSU LPS Arhalo + + Intracellular Pro IL-1β

18 I L - 1 b m R N A F o l d C h a n g e I L 1 - b ( p g / m L ) Arhalofenate Acid Inhibits MSU Induced IL-1 b Release in Mouse Macrophages Through Suppression of Gene Expression Macrophage Culture 2 days 4 hours 2 hours 18 hours LPS Remove LPS Arhalofenate Acid MSU Measure secreted IL-1b and Pro-IL-1b mrna 5 mrna 3 Secreted 4 3 ** 2 ** MSU + MSU

19 Conclusions Arhalofenate acid inhibits uptake by renal uric acid transporters Arhalofenate potently blocks MSU crystal-induced inflammation in a mice IL-1b and other NF-κB-dependent inflammatory mediators Significantly reduces neutrophil influx Arhalofenate acid show potent anti-inflammatory action in macrophages Suppresses both IL-1b mrna and protein Arhalofenate has a unique dual mode of action to address the key needs of gout patients: Inhibits uric acid uptake Suppresses MSU driven inflammation 19

20 Acknowledgements In vivo Pharmacology Noe Abarca Vanina Barreiro Annette Lucman Vic Vicena Stanford Shared FACS Facility Marty Bigos Bianca Gomez In vitro Biology Yun-Jung Choi Brian Lavan Tim Rantz Chemistry Xin Chen Fang Ma 2