Delivery of ALX-0171 by inhalation greatly reduces disease burden in a neonatal lamb RSV infection model 9th RSV Symposium Stellenbosch –South Africa 13th November 2014 Laurent Detalle Nanobodies® Inspired by nature Disclosure Laurent Detalle • employee at Ablynx Study performed at Iowa State University • paid for by Ablynx • Ablynx as study monitor www.ablynx.com 2 Ablynx – Facts and figures Corporate • Drug discovery and development company in Ghent, Belgium • >300 employees Technology • Pioneer in next generation biologics – Nanobodies® • >500 granted and pending patents Products • >30 programmes – six in clinical development • Three clinical proof-of-concepts (POC) • >900 healthy volunteers and patients treated with Nanobodies Partners • AbbVie, Boehringer Ingelheim, Eddingpharm, Merck & Co, Merck Serono and Novartis www.ablynx.com 3 Nanobodies – derived from heavy-chain only antibodies Camelid heavy-chain only antibodies are stable and fully functional Nanobodies represent the next generation of antibody-derived biologics VH VHH CH1 VHH VL CL CH2 CH3 Conventional antibodies www.ablynx.com 12-15kDa Ablynx’s Nanobody CH2 CH3 Heavy chain only antibodies • small • robust • sequence homology comparable to humanised/human mAbs • easily linked together • nano- to picomolar affinities • intractable targets • multiple administration routes • manufacturing in microbial cells 4 ALX-0171 – designed to treat infant RSV infection First-in-class treatment of RSV infection Being developed to treat hospitalised and out-patient infants anti-RSV Nanobody anti-RSV Nanobody Trivalent Nanobody: demonstrated in vitro efficacy against A and B serotypes ALX-0171 42kD anti-RSV Nanobody Pulmonary delivery: fast onset of action and high concentration at infection site In vivo efficacy demonstrated in cotton rat model and neonatal lambs Three phase I studies in man with inhaled ALX-0171 successfully completed First-in-infant phase IIa study expected to start in Nov/Dec 2014 www.ablynx.com 5 ALX-0171 – translational strategy Bridging the translational gap between adults and infants Requirement for a fit-for-purpose model ALX-0171 Efficacy/safety • ALX-0171 delivery Preclinical In vitro studies Preclinical In vivo studies ALX-0171 characterisation Efficacy in cotton rats Safety and PK in adult/juvenile rats Stability/ nebulisation studies Intranasal/whole body delivery www.ablynx.com Clinical In vivo studies Safety and PK in Human adults Oral inhalation Breath actuation Preclinical In vivo studies Efficacy, Safety and PK in Neonatal setting with relevant administration route Nasal inhalation with facemask 6 Neonatal lambs as model system for safety and efficacy testing of inhaled ALX-0171 upon RSV infection Safety Age/lung development • 2-5 days old • Colostrumdeprived • Alveolarisation starts preterm1 • Immature immune system Lung deposition/ Pharmacokinetics Size • Allows relevant administration route (facemask) • Enables multiple assessments • Similar breathing patterns Respiratory tract anatomy • Size of nasal cavity and airways2 • Organisation of local lymphoid tissue2 • Airway branching pattern2 • Distributon of epithelial cells, mast cells and airway smooth muscle3 • ... Efficacy RSV disease • Lower respiratory tract infection • Develop mild clinical symptoms4 • Histologic lesions4 • Enhanced disease postvaccination5 Similar characteristics to human infants 1ATS 4Derscheid, 2Scheerlinck, 5Derscheid, American journal of respiratory and critical care medicine, 2004. 170(3): p. 319-43 J.P., et al., Trends in biotechnology, 2008. 26(5): p. 259-66. 3Meeusen, E.L., et al., Drug Discovery Today, 2009. 6(4): p. 101-106 www.ablynx.com R.J. and M.R. Ackermann, Viruses, 2012. 4(10): p. 2359-78. R.J., et al. PloS one, 2013. 8(12): p. e81472. 7 Lamb study design Objectives • Assess efficacy/safety in neonatal setting Nebulisation of RSV M37 strain or culture medium Necropsy Onset of RSV infection - therapeutic setting starting at viral peak • Pharmacokinetics in target tissue - epithelial lining fluid (ELF) Day -1 0 1 2 3 4 5 6 7 8 5-6 animals/group Note: Pilot study was performed with similar schedule Treatment ALX-0171 or formulation buffer Study design Group RSV status Treatment Nebuliser filling dose Time taken for dose to be delivered Endpoints 1 Mockinfected Vehicle - - 2 Mockinfected ALX-0171 11 mg (Low dose) ~2 minutes • • • • • Viral titers (qPCR, FFU) Viral antigen expression in lungs (IHC) Gross lung viral lesions Histopathology General health status 3 RSV Vehicle - - 4 RSV ALX-0171 11 mg (Low dose) ~2 minutes • ELF Ctrough ALX-0171 levels www.ablynx.com 8 Nebulisation and PK Nebuliser • Mesh nebuliser • MMAD: 3.27 ± 0.13 μm • 2L/min airflow A L X - 0 1 7 1 c o n c e n t r a t io n s i n A L X - 0 1 7 1 c o n c e n t r a t io n s lu n g s 1000 100 L o w d o s e A L X -0 1 7 1 H ig h d o s e A L X - 0 1 7 1 100 IC 9 0 LLO Q [ A L X - 0 1 7 1 ] E L F ( µ g /m L ) 10 1 0 .1 IC 9 0 o L d A L o w V S R s X e V V S R e h -0 1 ic 7 le 1 7 e 1 s -0 o d X L o w A L N o m in a l t im e ( h o u r s p o s t f ir s t d o s e ) 80 le 60 ic 40 h 20 e 0 1 0 .0 1 10 V [ A L X -0 1 7 1 ] p la s m a ( n g /m L ) in p la s m a f r o m p ilo t s t u d y ALX-0171 concentrations in ELF were far above the IC90 following 3 daily inhalations and 2-3 log higher than systemic concentrations www.ablynx.com 9 Improved health status in RSV infected lambs General health status of lambs was scored on a daily basis G e n e r a l i ll n e s s s c o r e Score General illness score 0 No clinical signs 1 Reluctant to move 2 Reluctant to move, head down, depressed, not interested in eating 3 Down, unwilling to get up or difficulty standing, not eating 4 Down and should be euthanised, probably cannot eat % la m b s w it h s c o r e  1 100 -1 M o c k V e h ic le M o c k A L X -0 1 7 1 80 R S V V e h ic le R S V A L X -0 1 7 1 60 40 20 0 1 2 3 4 5 6 www.ablynx.com 6 5 4 3 2 1 y a D D a y 8 6 4 a D D a y y 3 1 0 y Poster number 120 n e o n a ta l la m b s a Treatment with ALX-0171 treatment improved the health status V ir a l k in e t ic s i n D Decline in general health status was observed in all lambs starting on day 3 V ir a l t it e r s in B A L F ( L o g 1 0 F F U /m L ) (  s e ) D a y s p o s t in f e c tio n 10 Antiviral effect of ALX-0171 ALX-0171 decreases viral load in the lungs of infected lambs Viral Titers V ir a l t it e r s  4 .1 L o g 6 4 2 DL 1 1 7 le -0 ic X h L e A Viral RNA V ir a l R N A in B A L F S R R S V V A V L e X h -0 ic 1 7 le 1 1 7 1 -0 X L A 1 -0 X L A V S R 0 le 1 7 le h e V R S V A L e X h -0 ic 1 ic 7 le 1 0 2 ic 2 h 4 4 e 6  0 .4 7 L o g 6 V 8 L o g 1 0 M 3 7 R N A c o p ie s /m g lu n g (  s e ) V ir a l R N A in lu n g t is s u e  0 .5 5 L o g V L o g 1 0 M 3 7 R N A c o p ie s /m L (B A L ) (  s e ) R R S S V V A V L e X h -0 ic 1 7 le 1 0 V L o g 1 0 F F U /m L B A L ( R t C d lo b e ) (  s e ) • 4 log10 on cultivatable virus • 0.5 log10 on viral RNA copies (>1 Log10 in pilot study) www.ablynx.com 11 Antiviral effect of ALX-0171 ALX-0171 decreases viral load in the lungs of infected lambs RSV Vehicle 10 5 1 7 1 X L A V S R R S V A L V X e h -0 -0 1 ic 7 le 1 0 X h -0 ic 1 7 le 1 1 7 1 -0 X 1 -0 X L 15 0 le 1 7 le h e V L e A S R R S V V A V L e V A R S V S R 20 Daily inhalation of ALX-0171 markedly reduced viral titres and lung lesions in RSV-infected lambs V A L e X h -0 ic 1 ic 7 le 1 0 2 ic 2 25 Decrease in virus load is reflected by a strong decrease in viral antigen expression 4 h 4 L o g 1 0 M 3 7 R N A c o p ie s /m g lu n g (  s e ) 6  0 .4 7 L o g 6 80 le V S S R R V ir a l R N A in lu n g t is s u e  0 .5 5 L o g V L o g 1 0 M 3 7 R N A c o p ie s /m L (B A L ) (  s e ) V ir a l R N A in B A L F A lv e o li 100 ic -0 X L A Viral RNA B r o n c h i/b r o n c h io le s 120 h 1 1 7 le ic h e V A V L e X h -0 ic 1 7 le 1 0 140 e DL IH C s c o r e s V ir a l a n t i g e n e x p r e s s io n V 2 M e a n n u m b e r o f a f fe c te d 4 b r o n c h i/b r o n c h io le s o r a lv e o li p e r fie ld (  s e )  4 .1 L o g 8 RSV ALX-0171 Viral Titers V ir a l t it e r s 6 V L o g 1 0 F F U /m L B A L ( R t C d lo b e ) (  s e ) • 4 log10 on cultivatable virus • 0.5 log10 on viral RNA copies (>1 Log10 in pilot study) www.ablynx.com 12 Effect of ALX-0171 on viral lung lesions Plum red RSV lesions seen in lungs of RSV infected lambs on day 6 post-infection • present on all lung lobes assessed R ig h t C ra n ia l R ig h t M id d le 50 R ig h t C a u d a l A cc e s so ry 40 L e ft C r a n ia l 30 L e ft M id d le L e ft C a u d a l 20 10 -0 X L A S R R S V V A V L e X h -0 ic 1 7 le 7 1 ic h e V 1 1 0 le M e a n % in v o lv m e n t /lo b e (  s e ) G r o s s v ir a l le s io n s Daily inhalation of ALX-0171 markedly reduced gross lung viral lesions www.ablynx.com 13 Histological scoring of lamb lung Alveolar consolidation was scored Observed lesions inflammatory infiltrate in bronchiolar lumen, alveolar spaces, and alveolar septa Bronchioles and alveolar spaces variably filled with degenerate neutrophils and sloughed epithelial cells with occasional large epithelial syncytial cells. 0% consolidation = 0 1%-9% consolidation = 1 10%-39% consolidation = 2 40%-69% consolidation = 3 70%-100% consolidation = 4 2 1 1 7 L A V S R R S V A L V X e h -0 1 ic 7 -0 X h e V Degenerate/Necrotic epithelial cells le 1 0 ic Syncytial cell 3 1 Accumulation of degenerate neutrophils 4 le alveolar septa mildly to moderately thickened by hyperplasia of type II cells H is to lo g ic a l c o n s o lid a t io n s c o r e alveolar spaces variably filled with degenerate neutrophils and sloughed epithelial cells Daily inhalation of ALX-0171 markedly reduced microscopic lung viral lesions www.ablynx.com 14 ALX-0171 – Conclusions Neonatal lambs proves to be a relevant model for treatment of RSV infection Administrations of nebulized ALX-0171 resulted in effective target lung concentrations and low systemic exposure ALX-0171 administration was well tolerated with no adverse events noted in a neonatal setting ALX-0171 decreased RSV disease burden in neonatal lambs when treatment started at peak of viral load and when symptoms were apparent • Beneficial influence shown on: - general health status - viral titers and lung viral antigen expression - gross and microscopic lung viral lesions First-in-infant study expected to start in Nov/Dec 2014 using a customdeveloped infant inhalation device based on a vibrating mesh www.ablynx.com 15 Acknowledgements Ablynx, Gent, Belgium • • • • • • • • • • • • • • • Thomas Stöhr Kjell Mortier Linde Duprez Lieselot Bontinck Massimiliano Germani Judith Baumeister Sandy Jacobs Donata Thuy Toon Wauman Gregory Daelman Koen Allosery Erik Depla Catelijne Stortelers Stephanie Staelens Francis Descamps www.ablynx.com Iowa State University • • • • • Mark R Ackermann Jack M Gallup Albert Van Geelen Alejandro Larios-Mora Shannon Jones Hostetter IWT, Belgium • Grant 130562 16