ACTRIIA-Fc rebalances BMP and activin/TGF- signaling to attenuate - PowerPoint PPT Presentation

ACTRIIA-Fc rebalances BMP and activin/TGF-β signaling to attenuate experimental pulmonary hypertension Lai-Ming Yung, Ph.D 1 ; R. Scott Pearsall, Ph.D 2 ; Geoffrey Bocobo, B.S. 1 ; Dianne S. Sako 2 ; Teresa Dinter, B.S. 1 ; Ravindra Kumar, Ph.D 2 ; Paul B. Yu, M.D., Ph.D. 1 1 Division of Cardiovascular Medicine, Brigham & Women’s Hospital and Harvard Medical School, Boston, MA; 2 Acceleron Pharma, Cambridge, MA

Mutations of Heritable PAH syndromes implicate loss of BMP function in pulmonary vascular disease BMP BMP TGF TGF activin BMPRII ALK4 ALK1 TGFBRII Endoglin BMPRII BMPR2 HPAH ALK5 ALK2 ACTRIIA ALK7 ALK3 ALK1 ACVRL1 HHT2-HPAH ENG ENG HHT1 II II I I * SMAD4 SMAD4 JP-HT P P P P P SMAD P P P SMAD2/3 1/5/9 SMAD9 SMAD9 HPAH P P SMAD SMAD2/3 BMP9 GDF2 HHT5 1/5/9 SMAD 4 P P SMAD SMAD2/3 1/5/9 Nucleus SMAD4 SMAD4 CAGA BRE myogenic and vascular homeostasis fibrogenic differentiation

PAH is characterized by deficient BMP signaling and exaggerated TGFβ/activin signaling - Genetics implicates deficient BMP9- BMP BMP TGF TGF BMPRII-ALK1-ENG-SMAD1/5/9 axis activin BMPRII ALK4 ALK1 - Non-genetic forms of PAH exhibit TGFBRII Endoglin ALK5 ALK2 ACTRIIA ALK7 ALK3 deficient BMP, and exaggerated TGFβ/activin signaling II II I I * - Pattern recapitulated in multiple P P P P P SMAD P P P SMAD2/3 1/5/9 animal models of PH P P SMAD SMAD2/3 1/5/9 - Approved therapies appear to SMAD 4 improve this balance P P SMAD SMAD2/3 1/5/9 Nucleus Atkinson L, Circ 2002 SMAD4 SMAD4 Long L, Nat Med 2015 CAGA BRE Yndestad A, J Appl Physiol 2009 Yan Y, Int J Cardiol 2016 Yang J, Circ Res 2010 Yung LM, AJRCCM 2016 Ogo T, Circ 2013

What is the impact of selective activin/GDF blockade in pulmonary arterial hypertension? BMP2 BMP4 BMP6 BMP7 BMP12 BMP9 BMP10 GDF8 GDF11 activin A activin B activin AC TGFβ1 TGFβ3 TGFβ2 Hypothesis: Blockade of activin ligands attenuates pulmonary vascular remodeling by rebalancing SMAD1/5/8 vs. SMAD2/3 signaling ACTRIIA-Fc TGFBRII-Fc (ACE-011/Sotatercept) Yung LM et al. AJRCCM 2016 SMAD 1/5/9 SMAD 2/3 *Primary ligands inhibited vascular homeostasis myogenic and fibrogenic differentiation

Impact of ACTRIIA-Fc (15 mg/kg S.C. twice weekly x 4 weeks) prophylaxis in rats treated with MCT (60 mg/kg) Non-muscularized MPAP RV / LV+S Partially muscularized P < 0.001 P < 0.05 Completely muscularized P < 0.001 P < 0.05 0.7 50 100% 0.9 1.4 7.3 * 0.6 40 0.5 36.4 41.6 Weight Ratio 30 0.4 mm Hg 66.9 ** 0.3 20 62.3 0.2 57.9 10 0.1 25.8 ** 0 0 Vehicle ACTRIIA-Fc Sildenafil Vehicle ACTRIIA-Fc Sildenafil Vehicle ACTRIIA-Fc Sildenafil (10 mg/klg) (60 mg/day) * p ≤ 0.05 vs. control ** p ≤ 0.01 vs. control

Impact of ACTRIIA-Fc (10 mg/kg weekly x 4 weeks) prophylaxis in rats treated with SUGEN5416 (200 mg/kg) and hypoxia Non-muscularized MPAP RV / LV+S Partially muscularized P < 0.001 P < 0.001 P < 0.001 P < 0.001 Completely muscularized 50 0.7 1.0 0.1 1.4* 100% 0.6 40 27.4 31.6 0.5 Weight Ratio 30 mm Hg 0.4 69.3 ** 0.3 20 72.5 67.4 0.2 10 0.1 29.3 ** 0 0 Vehicle ACTRIIA-Fc Sildenafil Vehicle ACTRIIA-Fc Sildenafil Vehicle ACTRIIA-Fc Sildenafil (10 mg/kg) (60 mg/day) * p ≤ 0.05 vs. control ** p ≤ 0.01 vs. control

Impact of ACTRIIA-Fc prophylaxis on PV remodeling in SU-Hx rats SUGEN-Hypoxia + SUGEN-Hypoxia + SUGEN-Hypoxia Sildenafil ACTRIIA-Fc Sugen hypoxia, lung histology (αSMA/elastin Staining)

Impact of ACTRIIA-Fc on progression of established PH in SU-Hx rats Normobaric hypoxia (FIO2 = 0.10) Echo RV function SU5416 (20 mg/kg S.C., weekly) Right heart catheterization Normoxia (3 weeks) SU-Hx (3 weeks) Fulton’s Index PV and RV Histology Sprague Dawley Rat ACTRIIA-Fc SU5416 SU-Hx Model Treatment n Control Vehicle 8 1 mg/kg twice weekly 7 ACTRIIA-Fc 3 mg/kg twice weekly 7 10 mg/kg twice weekly 8

ACTRIIA-Fc attenuates PH progression in SU-Hx rats Non -muscularized Partially muscularized Completely Muscularized 100 60 Percent of Total Vessels 18.5 31.9 31.2 42.3 RVSP (mmHg) 11.8 69.3** 0.5 p= 0.02 40 0.4 18.8 22.4 RV/(LV+S) p= 0.05 50 27.5 0.3 68.0 20 0.2 47.9 45.2 29.3** 28.8 0.1 (8) (7) (7) (8) (8) (7) (7) (8) 0 0 0.0 SU-Hx 1 3 10 SU-Hx 1 3 10 SU-Hx 1 3 10 ActRII-Fc (mg/kg twice weekly) ActRII-Fc (mg/kg twice weekly) ACTRIIA-Fc RAP-011 (mg/kg twice weekly) one-way ANOVA dose trend p = 0.03 (mg/kg twice weekly) one-way ANOVA dose trend p = 0.05 *one-way ANOVA trend p = 0.05

ACTRIIA-Fc attenuates intimal-medial remodeling in SU-Hx rats 100 SU-Hx 1 mg/kg % Fully Muscularized Vessels 80 60 p =0.05 40 20 (8) (7) (7) (8) 0 SU-Hx 1 3 10 ActRIIa-Fc (mg/kg twice weekly) one-way ANOVA trend p = 0.05 3 mg/kg 10 mg/kg 60 Medial wall thickness p =0.02 index (%) 40 20 vWF (8) (7) (7) (8) 0 SMA SU-Hx 1 3 10 DAPI 50 µm ActRII-Fc (mg/kg twice weekly) one-way ANOVA trend p = 0.01

ACTRIIA-Fc de-represses activin inhibition of BMP9 signaling BRE-Luciferase BRE-Luciferase Activity (RLU) Activity (RLU) 1 1 BMP9 - + + + - BMP9 - + + + - activin A - - + + + activin B - - + + + ACTRIIA-Fc - - - + - ACTRIIA-Fc - - - + - BMP Response Element (BRE-Luciferase) activity in A204 reporter cell line

Comparison of ACTRIIA-Fc to approved therapies in PH models Monocrotaline - Prevention SUGEN-Hypoxia - Prevention SUGEN-Hypoxia – Therapeutic % % Reduction % % Reduction % % Reduction Reduction in RVH Reduction in RVH Reduction in RVH Agent in mPAP RV/(LV+S) Agent in mPAP RV/(LV+S) Agent in RVSP RV/(LV+S) 21 30 36 5 31 Bosentan 1 Macitentan 3 10 19 Sildenafil 1 24 18 22 10 Sildenafil 4 Sildenafil 4 Riociguat 1 10 20 Beraprost NP 2 25 5 28 Beraprost NP 2 27 5 32 Tadalafil + 28 28 Macitentan 2 ACTRIIA-Fc 4 51 ACTRIIA-Fc 3 33 ACTIIA-Fc 4 56 47 54 30 1. Clozel etal Exp. Biol and Med (2006) 231: 967-973; Bosentan 300 mg/kg/d 1. Lang et al. PLoS One. 2012;7(8):e43433 2. Akagi et al J Cardiovasc Pharmacol 2016; 67; 290-298; Beraprost NP 150 μg/kg Sildenafil 50 mg/kg/d; Riociguat 10 mg/kg/d 3. Shinohara et al Am J Physiol Lung Cell Mol Physiol 2015; Macitentan 30 mg/kg/d 2 Boucherat et al. Sci Rep. 2017 Jul 3;7(1):4546 4. RAP-011 and Sildenafil (60 mg/kg/d) were tested in same study at CorDynamics Tadalafil 10 mg/kg/d; Macitentan 30 mg/kg/d 5. Right ventricular systolic pressure 3. Current study; 10 mg/kg twice weekly

Summary 1. ACTRIIA-Fc is a potentially mechanism-targeted , non-vasodilator PAH therapy that potently inhibits neo-intimal and medial remodeling. 2. ACTRIIA-Fc inhibits signaling of activins/GDFs and may augment BMPs; multiple mechanisms of action and cellular targets being considered. 3. Sotatercept has favorable pharmacokinetics , dosed SC every three weeks. 3. Human clinical experience includes nearly 400 patients across 13 trials . 4. Well tolerated at 0.3, 0.7, and 1.5 mg/kg in human subjects, corresponding to human equivalent doses of 1.8, 4.2 and 10 mg/kg in rats, overlapping with efficacious doses in rat models. 6. Phase 2 studies for PAH are planned.

Acknowledgements Yu laboratory – BWH Cardiology Lai-Ming Yung, PhD Peiran Yang, PhD Ivana Nikolic, MD Geoff Bocobo, BS Teresa Dinter, BS Lai-Ming Brian Peiran Geoff Teresa Megan McNeil, BS Yung, PhD Yang, PhD Bocobo, BS Dinter, BS Luca Troncone, PhD Acceleron Pharma Funding Ravindra Kumar, Ph.D NHLBI R01-HL131910 R. Scott Pearsall, Ph.D NHLBI R42-HL132742 Dianne S. Sako, B.S. B-BIC DRIVE GRANT Fondation Leducq

ACTRIIA-Fc fusion protein (RAP-011/Sotatercept) is an activin/GDF ligand trap BMP BMP TGF TGF BMP BMP activin BMPRII ALK4 ALK1 BMP BMP TGFBRII Endoglin ALK5 ALK2 ACTRIIA ALK7 ALK3 II II I I * P P P P P SMAD P P P SMAD2/3 1/5/9 P P SMAD SMAD2/3 1/5/9 ACTRIIA-Fc SMAD 4 (RAP-011/Sotatercept) P P SMAD SMAD2/3 1/5/9 Nucleus traps activin A, activin B, SMAD4 SMAD4 BRE CAGA - + GDF8, GDF11 ACTRIIA-Fc