Slide 1: The Development of CRISPR Based Medicines for the Treatment of Hematological Diseases
The Development of CRISPR Based Medicines for the Treatment of Hematological Diseases Grant Welstead, PhD Editas Medicine editasmedicine.com © 2017 Editas Medicine 1
Slide 2: Medicines that Aim to Repair Any Broken Gene
Medicines that Aim to Repair Any Broken Gene Potential to create the next major category of transformative medicines © 2017 Editas Medicine 2
Slide 3: Pipeline Strategy to Enable Successful Medicines
Pipeline Strategy to Enable Successful Medicines Medical Need Eye ▪ Severe diseases where current ▪ Leber Congenital Amaurosis 10 ▪ Ocular HSV treatments, if any, are poor ▪ Additional ocular indications ▪ Potential for durable therapies to provide unique benefit Lung ▪ Cystic Fibrosis Biology & Cli
Slide 4: CRISPR Unlocking the Promise of Cell Therapy
CRISPR Unlocking the Promise of Cell Therapy Gene Editing CD34+ HSC Bone Marrow Erythroid Myeloid Lymphoid Hematopoietic stem cells have the potential to yield multiple medicines for blood diseases including sickle cell disease and beta thalassemia T cells are therapeutic platform for cancer, autoim
Slide 5: CRISPR is a RNA-Guided Nuclease
CRISPR is a RNA-Guided Nuclease Guide Sequence Cut Cas9 DNA Sites ~100 nt PAM Guide Sequence DNA 42 nt Cpf1 PAM Cut Sites Nuclease Guide RNA Editing machinery can be engineered to target nearly any genomic location © 2017 Editas Medicine 5
Slide 6: Broad Toolkit of CRISPR Nucleases
Broad Toolkit of CRISPR Nucleases Cas9 AsCpf1 LbCpf1 MULTIPLE EDITING SpCas9 SaCas9 SYSTEMS AsCpf1 LbCpf1 ADVANCED FORMS FOR PAM PAM FLEXIBLE SpCas9 SaCas9 Cpf1 TARGETING PAM PAM ADVANCED SpCas9 eS SaCas9 eS FORMS WITH INCREASED SPECIFICITY HiFi HiFi AsCpf1: Acidaminococcus species Cpf1; LbCpf1: Lac
Slide 7: CRISPR Flexibility Addresses Diverse Mutations
CRISPR Flexibility Addresses Diverse Mutations Cut and Revise Cut and Remove Cut and Replace Cut and Insert Mutation Mutation Mutation PAM Insertion or Deletion Copy Insertion Deletion e.g., Engineered T cells e.g., LCA10 e.g., Hemoglobin Beta e.g., Safe harbor Non-homologous end joining Homology-di
Slide 8: Platform to Create CRISPR Medicines
Platform to Create CRISPR Medicines Achieve Right Repair Efficiently Edit Wide Range of Mutations ▪ Engineer and deliver correct template ▪ Expand opportunity with broader PAM recognition ▪ Use cut orientation to drive template utilization ▪ Enhance safety and efficacy with modifications to nuclease
Slide 9: Scalable, Consistent Engineered Cell Therapies
Scalable, Consistent Engineered Cell Therapies Electroporation + RNA Guided RNA Ribonucleoprotein Patient Engineered Endonuclease Guide Particle (RNP) Cell Patient Cell Optimized Delivery of RNP to Primary T cells Via Electroporation © 2017 Editas Medicine 9
Slide 10: Proprietary Approaches to Guide RNAs
Proprietary Approaches to Guide RNAs 5’ Single gRNA Frequency of Error Targeting Sequence High Low Heterogeneous product 3’ Length Change (full-length, truncated, errors) (Bases) 5’ 3’ Position (5’ 3’) 5’ 3’ Covalently-Coupled Dual gRNA Targeting Sequence Length Change Well-defined product (Bases)
Slide 11: Orthogonal Specificity Approaches for Best gRNAs
Orthogonal Specificity Approaches for Best gRNAs Proprietary Testing of in silico On-Target Prediction of Cutting (Cas9, Cpf1 Cutting Sites WT, nickase) Targeted Panels Unbiased for Detection of Detection of Sites from Biased Off-Target Cuts & Unbiased and Genomic Screens Alterations (e.g., GUIDE-Se
Slide 12: Lead Finding and Specificity to Select gRNA
Lead Finding and Specificity to Select gRNA In silico selection eliminates 50%+ of gRNAs Example in silico filtering In silico Selection Gene Total Tested gRNAs gRNAs Count of Genomic Loci 1 198 100 2 100 37 3 181 95 4 147 72 5 283 90 © 2017 Editas Medicine 12
Slide 13: Scale Up Powers Lead Finding & Optimization
Scale Up Powers Lead Finding & Optimization Target: 1 2 3 4 5 Exon: Primary screening of 5 targets with 2 enzymes in primary human T cells demonstrating high activity and reproducibility ▪ Fully tracked and automated process ▪ Performed in primary T cells ▪ RNP and target agnostic (any variant ▪ Syn
Slide 14: Gene Editing for Next-Gen CARs/TCRs
Gene Editing for Next-Gen CARs/TCRs Juno Therapeutics collaboration expands and accelerates ex vivo products ▪ CAR and TCR engineered T cell therapies have the potential to be transformative additions to immuno-oncology landscape ▪ Alliance with Juno seeks to address key goals for engineered T cells
Slide 15: Gene Editing for Next-Gen CARs/TCRs: Targeting T cell checkpoint pathways PD-1
Gene Editing for Next-Gen CARs/TCRs Targeting of T cell checkpoint pathways => PD-1 ▪ T cell reactivity against “self” is controlled by a series of checkpoint pathways downstream of cell surface receptors such as PD-1 and CTLA-4. ▪ In cancer patients, T cells recognize tumor cells that express neo a
Slide 16: Can We Target PD-1 in Engineered T cells By Gene Editing?
Gene Editing for Next-Gen CARs/TCRs Targeting of T cell checkpoint pathways – PD-1 Can We Target PD-1 in Engineered T cells By Gene Editing? Image source: smartpatients.com © 2017 Editas Medicine 16
Slide 17: Identification of Robust Guide RNA Leads
Identification of Robust Guide RNA Leads 100 % P D - 1 N e g a t iv e 80 60 ▪ S. pyogenes Cas9 RNPs in primary human T cells to knock-out PD-1 40 ▪ Several gRNAs performed well as assessed by FACS 20 ▪ Sequencing confirmed indels 0 ▪ gRNAs analyzed by GUIDE-Seq to identify off targets 0 1 2 3 4 9 1
Slide 18: Control and Specificity to Drive Precision
Control and Specificity to Drive Precision GUIDE-Seq Read Count Reads Guide RNA On-Target Off-Target 1 Off-Target 2 Off-Target 3 ▪ GUIDE-Seq drives empirical demonstration of selectivity of product candidates ▪ Off-targets identified by GUIDE-Seq would not be accurately predicted by in silico method
Slide 19: GUIDE-Seq Confirmation by Targeted Sequencing
GUIDE-Seq Confirmation by Targeted Sequencing Rank order confirmed 100 Transfection Off-target +rank guide order is the Mock sametransfection (no guide) as Guide-Seq run Reads Log10 % INDELS 10 1 Guide RNA Note: background 0.1 sensitivity is ~0.15- 0.1% On-Target Off-Target 1 Off-Target 2 Off-Target
Slide 20: High Efficiency Editing in CAR-T Cells
High Efficiency Editing in CAR-T Cells Gene Disruption in T cells ± CD19 CAR 100 Highly efficient gene knockout with >90% PD-1 KO in T cells also carrying Juno CAR % Insertions & Deletions 80 Guides with no detected off-target edits 60 using multiple orthogonal methodologies 40 Success on many targe
Slide 21: Summary
Summary Robust guide screening platform in T cells for multiple enzymes Highly efficient gene knockout for many T cells targets with >90% KO efficiency Guides with no detected off-target edits using multiple orthogonal methodologies Building internal manufacturing expertise for multiple ex vivo prog
Slide 22: Thank You
Thank You © 2017 © Editas Medicine 22