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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