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2. Some words about RIOCIGUAT First in class New target Direct sGC stimulator Sensitises sGC to NO Vasodilatation Orally treatment of PAH Improves quality of life Currently in clinical trial, phase III 2 3. PLAN Pathology Treatments sGC Discovery and development of Riociguat Clinical trials 3 4. Pulmonary Arterial Hypertension WHO classification Orphan disease, 30-50 cases per million Poor prognosis Symptoms: breathlessness, chest tightness, fatigue Vascular proliferation and remodelling Progressive increase in PVR Right ventricular failure, hypertrophyand death Imbalance of vascular effectorsN ENGL J MED 351;16 OCTOBER 14, 2004 PULMONARY ARTERIAL HYPERTENSION .FARBER ET AL 4 5. Diagnostic of PAH Echocardiography 6-MWT and right heart catheterisation 4 classes: class I is the least severe and class IV the mostadvanced 5 6. 6-Minutes Walk TestThe patient walks as faras possible in 6 minutesSafeHighly reproductibleInexpensive equipmentUsed as primary endpoint in clinical trials ATS. Am J Crit CARE MED 20026 7. PAH associated with HIV 0,5% of HIV-patients (6 to 12 times as high as the generalpopulation) related to the duration of HIV infection, due toconcomitant infections Mechanism unclear HIV-1 GP120 may stimulate theproduction of ET by macrophages seems to be independent of thedegree of immunosuppressionN ENGL J MED 351;16 OCTOBER 14, 2004 PULMONARY ARTERIAL HYPERTENSION .FARBER ET AL 7 8. PAH associated with others conditions Scleroderma = systemic sclerosis : prevalence of PAH isup to 16%PAH is the leading cause of deathVery poor prognosis Human herpesvirus N ENGL J MED 351;16 OCTOBER 14, 2004 PULMONARY ARTERIAL HYPERTENSION .FARBER ET AL8 9. ACTUAL TREATMENTS Prostacyclin Endothelin NO PDE59 10. N ENGL J MED 351;14 SEPTEMBER 30, 2004 TREATMENTOF PAH HUMBERT ET AL10 11. Prostacyclin therapy PGI2 is producted by metabolisation of arachidonic acid Induces relaxation Stimulating AMPc productionN ENGL J MED 351;14 SEPTEMBER 30, 2004 TREATMENT OF PAH HUMBERT ET AL11 12. Epoprostenol Flolan Intravenous infusion Improvement in 6-MWT Short half-life (3 min) Iloprost Ventavis Delivered by inhaler Short duration of action N ENGL J MED 351;14 SEPTEMBER 30, 2004 TREATMENT OF PAH HUMBERT ET AL 12 13. 13 14. Endothelins actions 14 15. Antagonists of ET1-R:Bosentan Tracleer Oral treatment Improvements in 6-MWT No dose-response effect Hepatotoxicity Sitaxsentan and Ambrisentan: selective of Eta TezosentanN ENGL J MED 351;14 SEPTEMBER 30, 2004 TREATMENT OF PAH HUMBERT ET AL15 16. 16 17. Nitric Oxide Vasodilatator Synthetised by NO synthase in endothelial and epithelialcells Inhaled treatments with NO are unsuitable as long termtherapies for PH due to: short life, development oftolerance, non specific interactions with biomolecules Toxicity with high dose CHEM MED CHEM 2009 4, 853-865 DISCOVERY OF RIOCIGUAT MITTENDORF ET AL 17 18. PDE5 effects+ 18 19. Inhibition of PDE5: Sildnafil Revatio Predominant PDE isoform in the lung that metabolizescGMP Pulmonary vasodilator effect Oral treatment Increase of cGMP in smooth muscle Improved exercise capacityand pulmonary hemodynamics No evidence of dose-response relationshipN ENGL J MED 353;20 NOVEMBER 17, 2005 SILDENAFIL CITRATE THERAPY FOR PAH GALIE ET AL 19 20. Soluble guanylate cyclase Heterodimer: larger subunit and a smaller haem-binding subunit Prosthetic haem moiety on the haem binding domain witha reduced Fe2+sGC GTP cGMP Potentiated by NONATURE REVIEWS 1028 NRD 2038 SEPTEMBER 2006 NO-INDEPENDENT STIMULATORS AND ACTIVATORS OF SOLUBLE SGC EVGENOV ET AL 20 21. +NOsGCGTP cGmP 21 22. Model of the haem binding domainof human sGC -subunitNATURE REVIEWS 1028 NRD 2038 SEPTEMBER 2006 NO-INDEPENDENT STIMULATORS AND ACTIVATORS OF SOLUBLE SGC EVGENOV ET AL 22 23. Structure of sGCCleavage of His NHis N 2+ the haem- histidine bond N2+N 2+NN Fe NN Fe N Fe NN NO N NOPenta coordinated Hexa coordinated Penta coordinatedhistidyl haem complexhistidine-haem-NO nitrosyl-haem intermediate complex Abolition of activation if remove, oxydise or inhibit Changes in the redox state leads to the formation of an NOinsensitive form of sGC NATURE REVIEWS 1028 NRD 2038 SEPTEMBER 2006 NO-INDEPENDENT STIMULATORS AND ACTIVATORS OF SOLUBLE SGC EVGENOV ET AL23 24. NO-sGC-cGMP Signal Transduction PathwayNATURE REVIEWS 1028 NRD 2038SEPTEMBER 2006 NO-INDEPENDENT STIMULATORS ANDACTIVATORS OF SOLUBLE SGC EVGENOV ET AL 24 25. Differences between systemic and pulmonary circulationIn response to hypoxia : Pulmonary arterial pressure Systemic pressure Pressure increases in PAs and decreases in renal arteries Mitochondria alter the production of ROS (O2;ONOO), causesthe PAs constriction : oxidation Therapies can target the redox 25 EUR RESPIR J 2009; 33: 717-721 SGC STIMULATORS AS A POTENTIAL THERAPY FOR PAH E.D. MICHELAKIS 26. Two novel drug class sGC activators dont modulate NO signalling at all butactivate the NO unresponsive haem oxidized or haem freeenzymeNO and haem independent By binding the unoccupied haem binding pocket Or by replacing the weakly bound oxidized haem BAY 58-2667 = Cinaciguat, intravenous form, currently inclinical trialNATURE REVIEWS 1028 NRD 2038 SEPTEMBER 2006 NO-INDEPENDENT STIMULATORS AND ACTIVATORS OF SOLUBLE SGC EVGENOV ET AL 26 27. NATURE REVIEWS 1028 NRD 2038 SEPTEMBER 2006 NO-INDEPENDENT STIMULATORS AND ACTIVATORS OF SOLUBLE SGC EVGENOV ET AL 27 28. Two novel drug class BAY 63-2521: Riociguat sGC stimulators stimulate sGC directly and enhance the sensitivity of the reduced enzyme to low levels of bioavailable NONO-independent but haem dependent Stabilization of the nitrosyl-haem complex Crucial dependency of Fe2+ NATURE REVIEWS 1028 NRD 2038 SEPTEMBER 2006 NO-INDEPENDENT STIMULATORS AND ACTIVATORS OF SOLUBLE SGC EVGENOV ET AL28 29. NATURE REVIEWS 1028 NRD 2038 SEPTEMBER 2006 NO-INDEPENDENT STIMULATORS AND ACTIVATORS OF SOLUBLE SGC EVGENOV ET AL 29 30. DISCOVERY AND OPTIMIZATION30 31. 1994: HTS which induce an increase of NO synthesis andstimulate sGC Led to 5-Substituted-2-furaldehyde-hydrazone sGC stimulator Increase when exposed to light, which is unwanted Dec 1994: Ko and co-workers describedindazole derivative YC-1 CHEM MED CHEM 2009 4, 853-865 DISCOVERY OF RIOCIGUAT MITTENDORF ET AL 31 32. Chemical optimization Based on YC-1 Similar mode of action but (2) and (3) are more potent and more specificof PDEActivity worn by 1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine32CHEM MED CHEM 2009 4, 853-865 DISCOVERY OF RIOCIGUAT MITTENDORF ET AL 33. In vitro assays and isolated organs cGMP formation in a sGC overexpressingChinese hamster ovarian cell line Iinhibition of phenylephrine induced contractions of rabbitaortic rings33CHEM MED CHEM 2009 4, 853-865 DISCOVERY OF RIOCIGUAT MITTENDORF ET AL 34. Activity of (2) and (3) (2): significant decrease of pulmonary arterialpressure, reversal in right ventricular hypertrophy, butinduction of CYP450 (3): no relevant CYP interactionbecause of the more polarmorpholine substituent,unfavorable PK profile 34 CHEM MED CHEM 2009 4, 853-865 DISCOVERY OF RIOCIGUAT MITTENDORF ET AL 35. Actions on PDE YC-1: Inhibits PDE 5 with IC-50 of 10M. BAY 41-2272 (2): No relevant inhibition of PDE1,2,5,9 up to 10mM. BAY 41-8543 (3): No PDE inhibition atpharmacologically relevant concentration.NATURE REVIEWS 1028 NRD 2038 SEPTEMBER 2006 NO-INDEPENDENT STIMULATORS AND ACTIVATORS OF SOLUBLE SGC EVGENOV ET ALCHEM MED CHEM 2009 4, 853-865 DISCOVERY OF RIOCIGUAT MITTENDORF ET AL35 36. SAR36 37. Activity worn by 1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine R3-R1: NH2: diamino analogues display a slightly more potentrelaxation of rabbit aorta and positive impact of the aminogroup on oral exposure. R2: pyrimidin C5 : No clear SAR demonstrated on CYP 1A2 et3A4, wide range of polars substituants and lipophilic groups. CHEM MED CHEM 2009 4, 853-865 DISCOVERY OF RIOCIGUAT MITTENDORF ET AL 37 38. Pyrimidine C5: most fruitful position38CHEM MED CHEM 2009 4, 853-865 DISCOVERY OF RIOCIGUAT MITTENDORF ET AL 39. (12) and (20) are derivated from(3) because of its no relevantCYP interactionCHEM MED CHEM 2009 4, 853-865 DISCOVERY OFRIOCIGUAT MITTENDORF ET AL39 40. (20): RiociguatResults for female Beagle dogs: R2 = N-mthylcarbamate Freatest PK profile Selected as a drug development candidate 40 CHEM MED CHEM 2009 4, 853-865 DISCOVERY OF RIOCIGUAT MITTENDORF ET AL 41. CYP and PDE profile No significant effects on CYP 1A2, 2B6, 2C19, 3A4 Not dose-dependent BAY 63-2521: IC-50 values for PDE 2,3,4,5,8,9are > 10M.PATENT US 2006/0052397A1 Mar. 9,2006 CARBAMATE-SUBSTITUTED PYRAZOLOPYRIDINES 41 42. SYNTHESE 42 43. F F F FCNTFANH3, MeOH OTFA + H2N N 3-dimethylamine N N N N HNN N N CO2 Etacroleine NH2 condensation aminolyse CO2 Etcyclo- CO2 EtCONH2 F F condensationTFAA, PyNaOMe, MeOH N N N N N NPinnerdeshydratation CN NH2 HN F F F N NC CN NaOMe, DMFN N NNi Raney N+N N 110C N NNNPh NH2cyclo- Nreduction malonitrileNN HNN additionNH2 NH2 H2N H2N NNPhNH2 Methyl-4,6-diamino-2-(1-(2-fluorobenzyl)-1H-pyrazolo [3,4-b]pyrydin-3-yl)-pyrimidin-(-ylmethylcarbamate) carbamatedeprotonation methylation CHEM MED CHEM 2009 4, 853-865 DISCOVERY OF RIOCIGUAT MITTENDORF ET AL 43 44. In vivo experiments Models of PAH Mice exposed to chronic hypoxia (10% O2) in a ventilated chamber Rats with 60 mg.kg-1 monocrotaline subcutaneously injection EUR RESPIR J 2008; 32: 881-891 EXPRESSION AND FUNCTION OF SGC IN PAH SCHERMULY ET AL44 45. Mice in chronic hypoxiaRight ventricular systolic pressure Right ventricular systolic pressureSystemic arterial pressure Cardiac frequency45EUR RESPIR J 2008; 32: 881-891 EXPRESSION AND FUNCTION OF SGC IN PAH SCHERMULY ET AL 46. Hypertensive rats Dose dependent blood pressure decrease Single dose 46 47. CLINICAL TRIALS47 48. Clinical trial, phase I 58 healthy male volunteers Single oral dose 0,25 to 5mg or placebo No serious adverse events Effect increased dose dependently at doses of 1mg to 5mg Mean arterial and diastolic pressures were decreased atdoses of 1mg and 5mg Systolic pressure not significantly affected 48 49. Pharmacokinetics (1) n=15 persons Riociguat plasma concentration Riociguat plasma concentrationsfollowing a single oral dose of 1mgand 2,5mg Changes in PVR from baseline (dosedependent increase whith pronouncedinterindividual variability)EUR. RESPIR.J.2009; 33: 785-792 FIRST ACUTE HAEMODYNAMIC STUDY OF sGC 49STIMULATOR RIOCIGUAT IN PH F.GRIMMINGER AND COWORKERS 50. Pharmacokinetics (2) n=5 in 1mg dose group and n=10 in 2,5mg dose group Vz/f: apparent volume of distribution during terminal phase after oral administration CL/f: total body clearance of drug from plasma calculated after oral administration Peak concentration after 0.25-1.5h Half-life of 10-12h Dose proportionnality for the both doses (AUC and Cmax): factor dosehad no influence on either parameters No hepatotoxicity, no abnormalities in laboratory values 50EUR. RESPIR.J.2009; 33: 785-792 FIRST ACUTE HAEMODYNAMIC STUDY OF sGC STIMULATOR RIOCIGUAT IN PH F.GRIMMINGER AND COWORKERS 51. Clinical trial, phase II During 12 weeks, 3 times a day, orally 75 patients with PAH disease PVR>300 dyn.s.cm-1 Evaluation of safety and tolerability: dose