Slide 1: Towards the development of a therapeutic RNA editing platform
Towards the development of a therapeutic RNA editing platform Chikdu Shivalila Presented at Deaminet 2022 January 24, 2022 Chikdu Shivalila, P Monian, G Lu, C Acker, B Bhattarai, D Boulay, K Bussow, M Byrne, A Chatterjee, D Chew, O Chivatakarn, J Desai, F Favaloro, J Godfrey, I Harding, A Hoss, N Iw
Slide 2: Forward-looking statements
Forward-looking statements This document contains forward-looking statements. All statements other than statements of historical facts contained in this document, including statements regarding possible or assumed future results of operations, preclinical and clinical studies, business strategies, r
Slide 3: Innovating stereopure backbone chemistry modifications
Innovating stereopure backbone chemistry modifications PRISM backbone linkages PO PS PN O B O B O B O O O O O R O (Sp) O R O (Rp) O R -O P -S P P O B O B N O B O O O O R O R O R … … … … … … Phosphoryl guanidine Chirality Chirality Chirality x-ray structure None PS backbone Rp PN backbone Rp PS backb
Slide 4: RNA editing with AIMers: A-to-I editing oligonucleotides
RNA editing with AIMers: A-to-I editing oligonucleotides Free-uptake of chemically modified oligonucleotides • First publication (1995) ✓ Learnings from using oligonucleotide to biological concepts edit RNA with AIMer: endogenous ADAR1 ✓ Applied to ASO structural concepts Wave’s A-to-I • Wave goal:
Slide 5: GalNAc-AIMers enable durable and specific editing out to day 50 in liver of NHPs
GalNAc-AIMers enable durable and specific editing out to day 50 in liver of NHPs AIMers detected in liver Substantial and durable ADAR editing with ACTB of NHP at day 50 editing in NHP liver in vivo AIMer is highly specific pre-dose RNA editing within full transcriptome (primary human hepatocytes) G
Slide 6: Efficient and durable editing in mouse CNS with unconjugated AIMer
Efficient and durable editing in mouse CNS with unconjugated AIMer Peak editing observed 4-weeks post-single ICV dose across tissues Cortex Hippocampus Striatum Brain stem Cerebellum Spinal cord 80 80 80 80 80 80 **** ** ** % UGP2 editing 60 60 60 60 60 60 ** * **** ** *** * *** * *** *** * *** ***
Slide 7: Productive editing beyond liver and CNS with unconjugated AIMers
Productive editing beyond liver and CNS with unconjugated AIMers Durable, dose-dependent editing in mice post-IVT 80 injection of unconjugated UGP2 AIMer %UGP2 mRNA editing 60 Ophthalmology AIMers 40 20 0 PBS 10 ug 50 ug PBS 10 ug 50 ug 1 week 4 weeks Editing in NHP 1-week post-single SC dose Editin
Slide 8: An ADAR editing approach to correct Alpha-1 antitrypsin deficiency (AATD)
An ADAR editing approach to correct Alpha-1 antitrypsin deficiency (AATD) Inverse relationship between circulating AAT levels and disease risk Objectives Low High • Recruit endogenous ADAR enzyme to edit SERPINA1 Z mRNA • Restore circulating M-AAT protein to Serum Risk of expected therapeutic thresh
Slide 9: Optimized AIMers achieve ~50% mRNA editing and restore AAT protein in mouse model
Optimized AIMers achieve ~50% mRNA editing and restore AAT protein well above therapeutic threshold in mouse model SERPINA1 RNA editing huADAR mouse AAT protein concentration in serum (3x5 mg/kg, SC) (3x10 mg/kg, SC) 1000 M-AAT (wild-type) 4-fold increase in 1000 total AAT * Z-AAT (mutant) Serum lev
Slide 10: Durable restoration of functional, M-AAT protein with ADAR editing
Durable restoration of functional, M-AAT protein with ADAR editing Human AAT serum concentration ≥3-fold Restored wild-type M-AAT detected over higher over 30 days post-last dose 30 days post-last dose 800 800 M-AAT production *** *** Elevated Z-AAT at Day 35 Serum AAT mg/mL Serum AAT mg/mL 600 11 m
Slide 11: ADAR editing is highly specific; no bystander editing observed on SERPINA1 transcript
ADAR editing is highly specific; no bystander editing observed on SERPINA1 transcript RNA editing only detected at PiZ mutation site in SERPINA1 transcript (mouse liver) RNA editing within transcriptome (mouse liver) C 0% SERPINA1 PBS (PiZ mutation site) T 100% Coverage Coverage C 48.2% SA1-4 AIMer
Slide 12: Apply AIMers to modify protein-protein interactions
Apply AIMers to modify protein-protein interactions KEAP1 editing Percentage editing (293T cells) KEAP1 Amino acid Control10 substitution NRF2 Tyr Cys AIMer 1 9 NRF2 is Arg Gly AIMer 2 8 degraded by proteasome Tyr Cys AIMer 3 7 Transcription is repressed Asn Asp AIMer 4 6 Basal conditions KEAP1 Edit
Slide 13: ADAR editing activates multiple genes, confirming disrupted protein-protein interaction in vitro
ADAR editing activates multiple genes, confirming disrupted protein-protein interaction in vitro ADAR editing of either KEAP1 or NRF2 directs gene activation Control 10 NQO1 Control Control 10 SLC7a11 Control9 Control Control9 AIMer 18 AIMer 18 AIMer 27 AIMer 27 KEAP1 AIMer 36 AIMer 36 AIMer 45 AIMe
Slide 14: Summary
Summary • AIMers represent Wave’s therapeutic RNA editing platform that leverages endogenous ADAR proteins – Achieve potent and specific editing – Support durable activity – Amenable to multiple routes of administration – Active in animals as GalNAc-conjugates or unconjugated • AIMers restore expres