iasoc - september 21-25, 2014 case study # 1 - september 21-25, 2014 ... a.v.k. (po) inhibit vitamin...

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IASOC - September 21-25, 2014 Aspects of cardiovascular drug discovery and development : development : Studies towards the Studies towards the discovery of Thrombin and TAFIa inhibitors 22/09/2014 1 22/09/2014 1 Case Study # 1 SYNTHESIS OF NEW TETRAHYDRO- PYRROLO-PYRAZINONE PYRROLO PYRAZINONE CONTAINING ORALLY BIOAVAILABLE THROMBIN BIOAVAILABLE THROMBIN INHIBITORS 22/09/2014 2 22/09/2014 2 COAGULATION CASCADE 22/09/2014 22/09/2014 THROMBIN (fIIa) Proteolytic Enzyme belonging to serine proteases family Cleaves soluble fibrinogen to fibrin and stabilizes clots by activation of factor XIII Amplifies its own formation by activation of several coagulation factors (factors V et VIII) Lowers its own formation by binding to thrombomodulin to ti t ti C (i ti ti f f t V t VIII ) activate protein C (inactivation of factors Vaet VIIIa) 22/09/2014 4 22/09/2014 4

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IASOC - September 21-25, 2014

Aspects of cardiovascular pdrug discovery and

development :development :

Studies towards theStudies towards the discovery of Thrombin and

TAFIa inhibitors

22/09/2014 122/09/2014 1

Case Study # 1y

SYNTHESIS OF NEW TETRAHYDRO-PYRROLO-PYRAZINONEPYRROLO PYRAZINONE CONTAINING ORALLY

BIOAVAILABLE THROMBINBIOAVAILABLE THROMBIN INHIBITORS

22/09/2014 222/09/2014 2

COAGULATION CASCADE

22/09/201422/09/2014

THROMBIN (fIIa)( )

Proteolytic Enzyme belonging to serine proteases familyy y g g p y

Cleaves soluble fibrinogen to fibrin and stabilizes clots by activationof factor XIII

Amplifies its own formation by activation of several coagulationfactors

(factors V et VIII)

Lowers its own formation by binding to thrombomodulin toti t t i C (i ti ti f f t V t VIII )activate protein C (inactivation of factors Va et VIIIa)

22/09/2014 422/09/2014 4

MARKETED ANTITHROMBOTICS

INDIRECT INHIBITORS OF THROMBIN

HEPARIN (iv/sc) Polysulfated polysaccharide

Potentiate the activity of AT III (neutralisation of thrombin and factor Xa)

Thrombopenia

LMWH (iv/sc) Chemical or Enzymatic Depolymerisation Heparin

Same activity compared to Heparin

Coumarin derivatives (ex: warfarin)

Same activity compared to Heparin

A.V.K. (po) Inhibit vitamin K- dpendent factors

Orally active

High bleeding risk

DDI

22/09/2014 522/09/2014 5

DDI

MARKETED ANTITHROMBOTICS

O

DIRECT INHIBITOR OF THROMBIN

N N

O

NO

N

N

NH

NH

NH

O

O

O

PRADAXA ( Dabigatran )B hi I lh iBoerhinger Ingelheim

Launched 2008

22/09/2014 622/09/2014 6

TARGET PRODUCT PROFILE

P t t d l ti I hibit f Th bi Potent and selective Inhibitor of Thrombin

Orally Active (good bioavailability)

Active on venous / arterial thrombosis models

Without Food /Drug Interaction

Once a Day Administration

No prodrug No prodrug

22/09/2014 722/09/2014 7

PEPTIDOMIMETIC CORES AS Phe-Pro ANALOG

N

O

NH2

O

N

O

O

NH2 OH

N

N

O

O

NH2 OH

OOH

O O

PFIZER ,NOVARTIS,

MERCK,ELI LILLY,

…. …

N

NNH2

O

N

OOH

SERVIER

22/09/2014 822/09/2014 8

( Tetrahedron Lett. 2002, 43, 3499-3501)

PEPTIDOMIMETIC INHIBITORS

N

O

NH2

O N N

N

N

NH

N

NH2O

NH

H

NH2

O H

S 34211S 34211

Direct and Reversible Thrombin InhibitorSelectiveSelectiveOrally active ( F=80 %)t1/2 (1h)

22/09/2014 922/09/2014 9

Pharmacological Profile of S 34840g

N

N

N

N

O

NH

ONH NH2O

BIOAVAILABILITY

IC50(nM)

SELECTIVITY

(nM)

COAGULATION H D R

BIOAVAILABILITY Dog Rat

t1/2 (h)

C(TT)2 (M)

3

100

34

> 33000

3 2

56

10

22/09/2014 1022/09/2014 10

CHEMICAL SYNTHESIS OF S 34840

CO2BzPhCH2OH

DMAP Boc O DIBAL(S)

(S)

NHO

CO2HNHO

CO2BzN

O

2

OO

DMAP Boc2O DIBALSOCl2(S)

(S)

80 %70 %98 %

N

CO2Bz

OH ON

CO2Bz

CH3O O NH

CO2BzMeOH APTS

TMSCN SnCl4

(S)(S)

(S)

(R/S) (R/S)(R/S)

98 %OHO

O CH3OO

HNC98 %85 %

NN

Cl

HCl gaz1)

NCl

O CO2Bz

ClCOCOCl (S)2)

55 %

N

Cl

N

Cl

+N

ClO CO2Bz

N NH2 NH

O CO2Bz

N(S)

+(S)84 %

N

N

Cl

N

N

NH

N

CO H(S)

H2 ; Pd/C

H2 ; Pd/CNaOH N

(S)85%N

HO CO2H

CH3

HO CO2H

(S)97 % 85%

N

NH

O

N

NH

NHBocN

NNH2

NHBoc

3

(S)HCl gaz

TBTU ; HOBT ; DIEA95 %O O H

N

82 %95 %

NH

O

N

ONH

NH2

N(S) ; 2 HCl

S 34840-1

SAR FROM S 34840

NN

NH

NN

OO

NH

NH2

22/09/2014 1322/09/2014 13

SUMMARY of SAR

N

NH

NN O CONHEt

HO CONH

S 35972S 35972

Potent and selective : IC 50 = 4nM , C(TT)2 = 0.24M Potent and selective : IC 50 4nM , C(TT)2 0.24M

Orally Active : F(Dog) = 85%

Without food interactionWithout food interaction

Orally active on venous and arterial models of thrombosis

Half-life = 4 h

22/09/2014 1422/09/2014 14

Half life = 4 h

Other scaffolds …..

IC50 (nM)

34

N

NH

O

N

N

N

NH

508

OO

NH

NH2

N

NH

O

N

ONH

N

NH2

5695

O H

N

N

NH

O

NH

N

375

OH

NH2

O

NO

NH

N

O

NH

810

ONH2

NN

NH

O

NH

N

22/09/2014 1522/09/2014 15

OH H

NH2

Metabolism Profile of S 35972

NO CONHEt

NH

ON

N

CONH

O CONHEt

S 35972

IC50 = 4 nMC(TT)2 = 0 24 MC(TT)2 = 0.24 M

t1/2 = 4 hF (Dog) = 85%

MF = 32%

22/09/2014 1622/09/2014 16

Metabolites of S 35972

N

NNN

O CONHEt(R/S) S 40240

IC 10 M

S 40243

IC 43 MO

H CONHOHIC50 = 10 nM

C(TT)2 = 0.22 Mt1/2 = 3h15

F(Dog) = 50%

IC50 = 43 nMC(TT)2 = 0.48 M

F(Dog) 50%MF = 65%

N

NNN

O CONHEtS 39733

IC50 = 1.8 nMO

NH

NCONH

O

50C(TT)2 = 0.11 M

22/09/2014 1722/09/2014 17

« Protected » S 35972

N

N

O

NH

NCONHF F

O CONHEt

OO

S 39858S 39858

IC50 = ndC(TT) = 0 19 MC(TT)2 = 0.19 M

F(Dog) = 18%MF = 67%

22/09/2014 1822/09/2014 18

Modification on RHS of S 35858od cat o o S o S 35858

N

NNH

NCONH

O CONHEt

O CONHF FO

S 39858IC50 = nd

C(TT) 0 19 MC(TT)2 = 0.19 MF = 18%

MF = 67% FN

NNN

S 40294

F F O

NH

NCONH

O

S 40294IC50 = 16 nM

C(TT)2 = 0.15 MF= 60%

MF 92%

22/09/2014 1922/09/2014 19

MF = 92%

Case Study # 2Case Study # 2

SYNTHESIS OF NEW TAFIaINHIBITORSINHIBITORS

( Thrombin Activable Fibrinolysis Inhibitor Activated )

22/09/2014 2022/09/2014 20

Role of TAFIa in Thrombosis

Lys Lys

IIa

Fibrinogen

TII

Fibrin

Plasmin

Lys

TmIIa

TAFIaFibrin degradation

productsPlasmin

Lys

TAFIa TAFIproducts

t-PA

plasminogenLys

Lys+

22/09/2014 2122/09/2014 21

Hit Compounds

Captopril inhibitor of TAFIaCaptopril inhibitor of TAFIa

V 811, V 812, M 326# 20 compounds

IC50 TAFIa (M)

# 20 compounds

N

OH

OHSH

OSHN

HOOC

O

SH

NH

SHSH

COOH

S 889725

S 991720

22/09/2014 2222/09/2014 22

Competitors

N

NH

N

N

SH

N

NH2(R)

NH2 COOH(S)

Pfizer (Phase I)

NCOOH(S)

Astra-Zeneca (Phase II)Pfizer (Phase I)UK-396082

Astra Zeneca (Phase II)AZD-9684

22/09/2014 2322/09/2014 23

Key Interactions in the Active Site

His 159

Zn2+

Glu 162

Zinc BindingGroup

His 288

Acid FunctionAmino Function

Arg 235 Arg 217

22/09/2014 2422/09/2014 24

Asp 348

Hit to Lead Exploration

COOHSH

O

NH

IC50 TAFIa (M)

O S 43970 30

NH

COOHNH2

NH

SHSH

COOH> 30

NH

COOHSH

OS 991720

N

S 46321> 30

22/09/2014 2522/09/2014 25

30

S 43621-1 Binding Mode

22/09/2014 2622/09/2014 26

Hit to Lead Optimisation

IC50 TAFIa (M)

n

NH

COOHSH

O

NH2

SHNH2 n

COOH

S 43970

22/09/2014 2722/09/2014 27

Lead Optimisation

IC50 TAFIa (M)

SHNH2 COOH

SH

NH2 COOH

S 457080.014

S 449000.17

SH

COOHNH2

S 453771 65

22/09/2014 2822/09/2014 28

1.65

S 44900-1 Binding Mode g

22/09/2014 2922/09/2014 29

S 44900-1 / S 45708-1Enantiomers SeparationEnantiomers Separation

SH

S 456462.03

IC50 TAFIa (M)

NH2 COOH

S 456470.081

S 449000 17

Syn-rac

0.17

SHNH

S 469115

NH2 COOH

S 469130 005

S 457080 014

22/09/2014 3022/09/2014 30

0.0050.014

Lead Optimisation : SAR Results

SHNH2 COOH2 COOH

Thiol function replacementsS 46913

Basic side chain modifications

• N methyl amine

Thiol function replacements

• Carboxylic Acid

• Hydroxamic Acid• N-methyl amine

• N,N-dimethyl amine

• Cyclic amine

Hydroxamic Acid

• Retro-hydroxamic Acid

• Triazoley

• Pyridine

IC > M

22/09/2014 3122/09/2014 31

IC50 > M

Stereoselective synthesis of S 46913

OH

tBuOH Pyridine tBuOH - NaH Cat*CO2tBu

O

COCl CO2tButBuO2CClOC

O

90%93%

tBuOH - Pyridine Toluène Toluène

90 %

OCO2tBu

H

OCO2tBu

OH

OCO2tBu

Br

OHCO2tBu

Br

H

(+)DIPCl NaBH(OAc)3 THF

82%ee > 99%

Ph3P - NBS CH2Cl2

87%de > 99%

43%

O

Boc2NH - Cs2CO3 - DMFOH

CO2tBuHO

SOO

CO2tBuHS

CO2tBuO

H MsCl-THF-TEA AcSK-Isopropanol

ee 99%

2 2 3

NBoc2NBoc2

2

NBoc255 % 87% 28%

SCO2H

OH

SHCO2HHHCl/Dioxane 4M

76%

HCl aq 4M

95%

22/09/2014 3222/09/2014 32

NH2NH2

76%HCl

95%HCl

ACKNOWLEDGEMENTS

CHEMISTRY PHARMACOLOGY

Guillaume DE NANTEUIL

Bernard PORTEVIN

Tony VERBEUREN

Alain RUPIN

Alain BENOIST

Danièle HENO

Sophie CORNET

Marie-Odile VALLEZ

Philippe MENNECIER

Isabelle RICHARD

Camille POINDRON

Aurélie MOTHE

Christian MERIAUX

A F i GUILLOUZIC

Christine MAUCLAIR

Anne Françoise GUILLOUZIC

Nicolas BOYER

22/09/2014 3322/09/2014 33

Milestones of our companyp y

Fi i i h

Servier crosses the threshold of  75% of its 

turnover made worldwide

First affiliates abroad : SpainBrazil

Portugal

First operations with China and Russia

worldwide

Portugal

Fi t ffili t i A i

Construction of Arklow production plant 

3 new chemical entities : PROTELOS

PROCORALAN VALDOXAN

Doctor Servier founded“ LES LABORATOIRES SERVIER”

in Orléans (FRANCE)

First affiliates in Asia :Hong Kong,Malaysia,Singapore

(Ireland) 21.000 employees

in Orléans (FRANCE)9 employees

22/09/2014 34

Servier : Research Orientations

Epigeneticsp gAngiogenesisProliferation / Differention PathwaysCell metabolismApoptosis

Coronary Arterial DiseaseHeart Failure

Venous DiseaseIschemia p p

Immunotherapy Protein degradationONCOLOGY

Arterial HypertensionCARDIOVASCULAR

METABOLISMType II Diabetes Mellitus

Depression / AnxietySchizophrenia

Cognitive ImpairmentNeurodegenerative diseases

(Alzheimer, Parkinson)CNS

RHUMATOLOGYRHUMATOLOGYOsteoporosisOsteoarthritis

Inflammatory & auto-immune diseasesMuscular pathologies

22/09/2014 35

uscu a pat o og es

Main marketed productsp

CARDIOVASCULARCoversyl Hyperium

Coveram Artex

ONCOLOGYMuphoran

CNSValdoxanSt bl

Preterax ProcoralanFludex Vastarel

METABOLISMDiamicron

Di i MR

StablonTrivastal

Diamicron MRRHUMATOLOGY

Protelos

22/09/2014 3622/09/2014 36