©Copyright 2013 Galapagos NV

GLPG0974, a selective FFA2 antagonist

A promising approach for the treatment of neutrophil-driven disorders

Johan Beetens, PhD, PharmD Development Project Leader, Head Fatty acid activation of G protein-coupled receptors: Basic and clinical perspectives (ASPET meeting, Boston) 22 April 2013

2

Presentation outline

• Drug target FFA2

biology and expression

• GLPG0974

pharmacology

initial Phase I clinical data

• Next steps: Proof-of-Concept

3

Free Fatty Acid Receptor 2 (FFA2)

• Ligands: short-chain fatty acids (SCFA; C1-C6)

acetate/propionate/butyrate

major source in human body: via fermentation by colonic bacteria

act also via other receptors and enzymes: FFA3, GPR109A, HDAC, PPARγ, COX,…

butyrate: energy fuel for enterocytes

Gαi

acetate, propionate, butyrate

Gαq

4

FFA2 Expression and biology

• FFA2 mainly expressed on

immune cells (neutrophils, monocytes, eosinophils)

colon mucosa (enterocytes, entero-endocrine cells)

adipocytes, pancreatic β-cells

• Biology data mainly derived from FFA2-/- KO mice

role in neutrophil-driven inflammation (a.o. IBD) Maslowski, 2009; Sina, 2009; Vinolo, 2011

role in metabolism: obesity, dyslipidemia, diabetes Ge, 2008; Bjursell, 2011; Tolhurst, 2012

5

Neutrophil migration and activation

calcium flux

L-selectin shedding

CD11b activation

chemotaxis

ROS production

MPO release

phagocytosis

Maslowski, 2009; Sina, 2009; Vinolo, 2011

SCFA effects via FFA2

Sherwood, 2004

6

Presentation outline

• Drug target FFA2

biology and expression

• GLPG0974 data

pharmacology

initial Phase I clinical data

• Next steps: Proof-of-Concept

7

FFA2 and candidate drug GLPG0974 Discovery Cascade

Target & Chemistry

• Target discovery • Medicinal Chemistry

Primary

• Ca2+flux assay • FFA2 [35S]-GTPγS binding assay • Human neutrophil migration

ADME & PK

• Solubility, Microsomal stability • Early rat PK

ADME-T & PK

• Advanced ADME-T assays • Receptor selectivitity profile

Ex vivo

• Whole blood assay • PK 2nd species

• Confirmed compound attributes

high potency in a neutrophil biology-dependent assay

high selectivity for FFA2

PK profile in rat and monkey supporting prediction in human to cover IC50/IC90 for 24 hrs

8

GLPG0974 Acetate-stimulated target assays

FFA2 GTPγS binding assay (human FFA2 in HEK293 membranes)

Intracellular Ca++ mobilization assay (human FFA2 in HEK293 cells)

IC50: 11 ± 2 nM IC50: 75 ± 13 nM

GLPG0974 potent and selective antagonist of FFA2 Not active on FFA1 or FFA3 or other receptors

-20

0

20

40

60

80

100

-10 -9 -8 -7 -6 -5

% o

f con

trol

GLPG0974 (log [M])

-20

0

20

40

60

80

100

-10 -9 -8 -7 -6

% o

f con

trol

GLPG0974 (log [M])

9

GLPG0974 Acetate-induced neutrophil migration

compounds in plasma

polycarbonate membrane (5µM) Medium + acetate

neutrophils (12-15µM)

Human neutrophils

GLPG0974 antagonize human and monkey, but not rodent, canine or rabbit FFA2

0

50

100

150

-10 -9 -8 -7 -6 -5

% o

f con

trol

GLPG0974 (log [M])

Species IC50 (nM)

Human 43 ± 10

Cynomolgus monkey 70 ± 10

Rat not active

Mouse not active

Dog not active

Rabbit not active

10

FFA2 in animal IBD models Experience in FFA2-/- KO mouse

Model Maslowski et al. 2009 Sina et al. 2009

Acute DSS colitis Exaggerated inflammation ↑ Histology score ↑ MPO in colon

Mortality due to sepsis ↓ PMN infiltration in colon ↓ MPO & TNFα in colon ↑ TNFα & KC in serum

Chronic DSS colitis

Exaggerated inflammation ↑ Mortality in the 1st week ↑ Histology score ↑ MPO in colon on D42 ↓ effect of oral acetate

Reduced inflammation ↓ PMN infiltration in colon ↓ MPO & TNFα in colon

11

GLPG0974 Monkey model for IBD

• Cotton-top tamarin IBD model spontaneous acute and chronic colitis

close similarities to human ulcerative colitis

good response to marketed therapies

but

listed as endangered species, only breeding programs allowed

model no longer available

• Invest in other monkey model of inflammation?

12

Neutrophil migration biomarker Activation of CD11b

• CD11b/CD18 (Mac-1) interaction with ICAM-1 mediates firm adhesion

• Conformational change of CD11b (CD11b[AE]) needed for ICAM-1 interaction

total CD11b expression does not per se correlate with adhesion capacity of Mac-1

CD11b[AE] more relevant marker of neutrophil activation in blood than total CD11b

Diamond, 1993; Orr, 2007; Zarbock, 2009;

Sherwood, 2004

13

GLPG0974 CD11b[AE] as biomarker in human whole blood

In vitro inhibition of acetate-induced CD11b[AE]

GLPG0974 inhibits acetate-induced CD11b[AE] expression, not IL8-, fMLP-, TNFα or LPS-induced expression

IC50: 483 ± 72 nM

14

GLPG0974 single dose FiH study Robust effect on biomarker

Ex vivo inhibition of acetate-induced CD11b[AE]

-20

0

20

40

60

80

100

0 1 2 3 4 5 6 7 8

% I

nh

ibit

ion

Time post dose (h)

Placebo

30 mg

90 mg

250 mg

Safety: safe & well tolerated up to 250 mg SD PK: good oral bioavailability and dose-proportional exposure PD: dose-dependent response, apparent PK/PD relationship

15

GLPG0974 Multiple Ascending Doses Challenges and questions

• Prolonged (up to 24 h) and sustained effect (up to 14 days) on biomarker activity?

• Effect of multiple dosing on exposure?

• Conflicting data on metabolic function (FFA2-/- KO mice)

improved (high-fat diet) or no effect (normal diet) on glucose tolerance, protected from obesity and dislipidemia (HF diet) (Bjursell, 2011)

impaired glucose tolerance, reduced insulin & GLP-1 secretion (Tolhurst, 2012)

effects on oral glucose tolerance test?

• Study design

32 healthy volunteers, 4 dose levels, per level 6 active & 2 placebo

safety, tolerability, PK, PD

16

Pharmacokinetics (Day 13) Pharmacodynamics (Day 13) Acetate-induced CD11b[AE] up regulation

Dose-proportional increase in exposure; 24 h inhibition at 200 mg BID

1

10

100

1000

10000

0 4 8 12 16 20 24

GLP

G09

74 (

ng/m

L)

Hours post dose

-60

-40

-20

0

20

40

60

80

100

0 4 8 12 16 20 24%

inhi

biti

on v

s pr

edos

e D

1 Hours post dose

GLPG0974 Multiple Ascending Doses Results

17

GLPG0974 Multiple Ascending Doses Conclusion

• Well tolerated and safe

no relevant treatment-emergent adverse events

no relevant effects on lab safety parameters, ECGs and vital signs

OGTT: no clinically relevant changes

• Good pharmacokinetics

good oral bioavailability and dose-proportional exposure

terminal half life of 6 h

• Robust pharmacodynamic signal

dose-dependent inhibition during 24 h, sustained over 14 days

18

Presentation outline

• Drug target FFA2

biology and expression

• GLPG0974 data

pharmacology

initial Phase I clinical data

• Next steps: Proof-of-Concept

19

• Inflammatory Bowel Diseases (IBD)

ulcerative colitis

Crohn’s disease

• Periodontitis

• Chronic Obstructive Pulmonary Disease (COPD)

• Vasculitis

GLPG0974: Proof-of-Concept study PoC and therapeutic opportunities

20

control UCR UCNR before Rx after Rx before Rx after Rx

FFA2 in ulcerative colitis (UC) FFA2 expression in colon biopsies (mRNA)

Arijs et al., (2011)

FFA2 is upregulated in colon tissue from UC patients and normalizes after successful treatment with anti-TNF mAB

Rx: infliximab, 4-6 weeks UCR: responders UCNR: non-responders

21

Cummings, 1987

SCFAs levels are high in colon

Blood ~100 μM • Acetate 100-150 μM • Propionate 4-5 μM • Butyrate 1-3 μM

Gut lumen ~100 mM

Wolever, 1997

22

GLPG0974: Proof-of-Concept study Questions and considerations

• Role of neutrophils in IBD

inhibition of neutrophil migration in IBD beneficial?

consistent with lowering fecal calprotectin

• Conflicting data in mouse IBD models (FFA2-/- KO)

increased inflammation and disease exacerbation (Maslowski, 2009)

decreased inflammation and PMN influx in colonic tissue (Sina, 2009)

• GLPG0974 in IBD

concept new mode of action

23

GLPG0974: Proof-of-Concept study Feedback Key Opinion Leaders

• In general, neutrophils deleterious in IBD

• Neutrophils more prominent in UC than Crohn’s

neutrophil predominant cell type in UC

• KoLs: “evaluate GLPG0974 in IBD patients”

mouse models not predictive for disease

new MoA: hard to predict outcome

• Monitoring: opportunistic infections, UC flare incidence

UC provides good opportunity for Proof-of-Concept

24

GLPG0974: conclusions and prospects • Potent and very selective FFA2 antagonist

only active on human and monkey receptor

• Inhibits FFA2-induced activation and migration of neutrophils

• Good PK/PD profile, safe and well-tolerated in volunteers

• First FFA2 antagonist to be evaluated in patients (IBD)

MoA clearly differentiates from existing therapies and new compounds currently in pipeline for IBD

Four-week PoC study ongoing in UC patients in Belgium and Eastern European Union

25

Acknowledgements Chemistry Laurent Sanière Mathieu Pizzonero Marielle Babel Jean-Michel Lefrançois Fatoumata Djata Sabrina Kopiejewski Christelle L’ebraly Murielle Manioc Stephane Beaumont Elsa De Lemos Benedetta Crescenzi François Nique Natacha Bienvenu Hélène Jary Laetitia Peltier Florilène Soulas Pierre Deprez Stephen Fletcher

Biology/ADME Sonia Dupont Roland Blanque René Galien Thierry Christophe Katja Conrath Luc Nelles David Craeye Celine Cottereaux Corinne Saccomani Steve De Vos Nicolas Triballeau Philippe Delerive Nick Vandeghinste Emanuelle Wakselman Ellen van der Aar Reginald Brys Dennis Polancec Katarina Oreskovic

Development Gerben van ‘t Klooster Florence Namour Giocondo Lorenzon Christine Guerin Frédéric Vanhoutte Marc De Weer Nicolas Sabourault Annegret Van der AA Tim Van Kaem Bart Franken Luc Meuleneers Piet Wigerinck

External experts Paul Rutgeerts Severine Vermeire Walter Reinisch Brian Feagan Daniel Podolsky Axel Dignass Tim Orchard Martine Devos Graeme Milligan