Therapeutic Genome Editing with CRISPR-Cas and other programmable gene scissors

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1 Brussels, April 12 th 2018 Therapeutic Genome Editing with CRISPR-Cas and other programmable gene scissors Toni Cathomen

2 Gene Therapy returns to Center Stage Engineering the Genome Jameson Golliday (source: The Atlantic) Emily Whitehead ( 2

3 Gene Therapy & Genome Editing Novel Tools in Cell and Gene Therapy Gene Therapy CRISPR-Cas Genome Editing Gene Scissors 3

4 Our Genome Genome à Chromosome à Gene Characteristics Cell Disease 4

5 Therapeutic Applications Gene Therapy Genome Editing Hereditary Disorders Infectious Disorders Cancer 5

6 Clinical Translation of Gene Therapy Some Definitions Transfer ex vivo Genome Editing Gene Scissors (CRISPR-Cas, TALEN, ) Transfer in vivo (Blood) Stem Cells Immune T Cells Liver Eye 6

7 Gene Scissors What is a Gene Scissor (Designer Nuclease)? CRISPR seek cut Cas 7

8 Designer Nucleases prominent classes of engineered nucleases 2 moieties Zinc-finger nucleases TALE nucleases (TALEN) CRISPR-Cas nucleases DNA recognition - seek DNA cleavage - cut ~24 bp ~36 bp ~22 bp From: Éva Scheuring- Vanamee, Mount Sinai From: Stoddard et al., 2012 Science From: Nishimasu et al., 2014 Cell 8

9 Genome Surgery How to employ Gene Scissors Gene Correction Gene Knockout Donor (A functional ) Gene A mutated Gene A functional Gene A functional Gene A KO 9

10 clinical genome editing trials fast increasing number of clinical gene editing trials (June 2017) Mechanism NHEJ Knockout HDR correction Application ex vivo in vivo in vivo Nuclease type Disease entity Locus Edited cells country Trial number HIV infection CCR5 CD4 + T cells USA e.g. NCT HIV infection CCR5 CD34 + cells USA NCT TALEN Leukemia (B-ALL) TRAC, CD52 CAR T cells UK, F, USA NCT CRISPR/Cas9 TALEN & CRISPR/Cas9 Leukemia (B-ALL) TRAC, B2M CAR T cells PRC NCT EBV-associated tumor-specific T PDCD1 malignancies cells PRC NCT Solid tumors PDCD1 tumor-specific T e.g. PRC cells NCT HIV infection CCR5 CD34 + cells PRC NCT Cervical pre-cancerious lesions HPV E7 Epithelial cells PRC NCT Cervical epithelial neoplasia HPV E6/E7 Epithelial cells PRC NCT Hemophilia B ALB, ins FIX Hepatocytes USA NCT Mucopolysaccharidosis I ALB, ins IDUA Hepatocytes USA NCT Mucopolysaccharidosis II ALB, ins IDS Hepatocytes USA NCT

11 key parameters for clinical translation Effective (Activity) & Precise (Specificity) Develop designer nucleases with: Ø High on-target activity Ø Low off-target activity Specificity profiles 11 Joung et al., Nat Biotechnol 2015

12 clinical genome editing trials Knocking out CCR5 in blood stem cells Mechanism NHEJ knockout HDR correction Application ex vivo in vivo in vivo Nuclease type Disease entity Locus Edited cells country Trial number HIV infection CCR5 CD4 + T cells USA e.g. NCT HIV infection CCR5 CD34 + cells USA NCT TALEN Leukemia (B-ALL) TRAC, CD52 CAR T cells UK, F, USA NCT CRISPR/Cas9 TALEN & CRISPR/Cas9 Leukemia (B-ALL) TRAC, B2M CAR T cells PRC NCT EBV-associated tumor-specific T PDCD1 malignancies cells PRC NCT Solid tumors PDCD1 tumor-specific T e.g. PRC cells NCT HIV infection CCR5 CD34 + cells PRC NCT Cervical pre-cancerious lesions HPV E7 Epithelial cells PRC NCT Cervical epithelial neoplasia HPV E6/E7 Epithelial cells PRC NCT Hemophilia B ALB, ins FIX Hepatocytes USA NCT Mucopolysaccharidosis I ALB, ins IDUA Hepatocytes USA NCT Mucopolysaccharidosis II ALB, ins IDS Hepatocytes USA NCT

13 Steve Crohn Towards a Cure for HIV Therapeutic Knockout of a Gene Timothy Brown HIV X Donor HIV-Patient cured HIV-Patient CD4 CCR5 CD4 32 Stammzellen CD4 CCR5 Blood stem cells CD4 infected resistant HIV resistant HIV + resistant 13

14 clinical genome editing trials as of June 2017 Mechanism NHEJ HDR Application ex vivo in vivo in vivo Nuclease type Disease entity Locus Edited cells country Trial number HIV infection CCR5 CD4 + T cells USA e.g. NCT HIV infection CCR5 CD34 + cells USA NCT TALEN Leukemia (B-ALL) TRAC, CD52 CAR T cells UK, F, USA NCT CRISPR/Cas9 TALEN & CRISPR/Cas9 Leukemia (B-ALL) TRAC, B2M CAR T cells PRC NCT EBV-associated tumor-specific T PDCD1 malignancies cells PRC NCT Solid tumors PDCD1 tumor-specific T e.g. PRC cells NCT HIV infection CCR5 CD34 + cells PRC NCT Cervical pre-cancerious lesions HPV E7 Epithelial cells PRC NCT Cervical epithelial neoplasia HPV E6/E7 Epithelial cells PRC NCT Hemophilia B ALB, ins FIX Hepatocytes USA NCT Mucopolysaccharidosis I ALB, ins IDUA Hepatocytes USA NCT Mucopolysaccharidosis II ALB, ins IDS Hepatocytes USA NCT

15 In Vivo Genome Editing for Hemophilia 15 K. High et al, Blood 2016

16 Investments Biotechnology 5 billion US$ until 2021 Platform Companies Disease Targets Sangamo BioSciences HIV, Hemophilia, Lysosomal Storage Disorders TALEN Cellectis S.A. Cancer (CAR T cells) CRISPR-Cas Crispr Therapeutics Editas Medicine Intellia Therapeutics Hemoglobinopathies (Thalassemia, Sickle Cell Disease) Cancer (CAR T cells), inherited blindness, Cystic Fibrosis, Duchenne Muscle Dystrophy Hepatitis-B-Virus, Alpha-1-Antitrypsin-Insufficency, hereditary metabolic disorders, cancer (CAR T cells) MegaTAL Bluebird Bio cancer (CAR T cells) (as found on company websites) 16

17 Benefits / Risks Benefits To CURE life-threating disorders Risks Long-term effects? Gene Scissors, Vectors Single shot Costs? 17