Shedding after in situ rAAV delivery Review of the literature and 8 - PowerPoint PPT Presentation

Shedding after in situ rAAV delivery Review of the literature and 8 years of experience in Nantes in large animal models Caroline LE GUINER, PhD PhD EMEA/ICH Workshop on viral/vector vector shedding shedding Caroline LE GUINER, EMEA/ICH Workshop on viral/ INSERM U649 (Philippe MOULLIER) – – Gene Gene Therapy Therapy Laboratory Laboratory INSERM U649 (Philippe MOULLIER) 10/30/07 10/30/07 Nantes - Nantes - FRANCE FRANCE Rotterdam, NL Rotterdam, NL

Animal Models in Nantes rAAV serotype serotype Maximal time course Mode of delivery Animal Models in Nantes rAAV Maximal time course Mode of delivery Intracerebral Intracerebral Spinal Muscular Atrophy Cat 1, 2, 5 8 months Spinal Muscular Atrophy Cat 1, 2, 5 8 months Mucopolysaccharidosis Type I Dog Type I Dog Mucopolysaccharidosis Nonhuman primate Nonhuman primate Intrathecal Intrathecal Spinal Muscular Atrophy Cat Spinal Muscular Atrophy Cat 1 1 3 months 3 months Subretinal Subretinal RPE65 deficient Dog RPE65 deficient Dog 2, 4, 5 2, 4, 5 6 years 6 years Normal dog Normal dog Nonhuman primate Nonhuman primate Intravitreal Intravitreal Normal dog 2 4 months Normal dog 2 4 months Isolated limb infusion Isolated limb infusion Nonhuman primate Nonhuman primate 1, 8 1, 8 2 years 2 years Intramuscular Intramuscular Nonhuman primate Nonhuman primate 1, 2, 8 1, 2, 8 5 years 5 years Intrathymic Intrathymic Nonhuman primate 8 1 month Nonhuman primate 8 1 month

Vector Vector Infectious Transgene Transgene Infectious Transgene Transgene particles particles mRNA mRNA product product genome genome + + + + >100 vg vg/ /cell cell >100 Mode of delivery: Target organ organ Target N/A N/A ( ≥ ( ≥ month month 8) 8) ( ≥ ( ≥ month month 6 6) ) ( ≥ ( ≥ month month 8) 8) Intracerebral + + + + Serum Serum nd nd nd nd (up to day (up to day 4) 4) (up to day (up to day 10) 10) Animal Model: + + Cerebrospinal Cerebrospinal - - nd nd nd nd Fluid Fluid ( ≥ ≥ month month 3) 3) ( Cat, Dog, Primate Urine Urine nd nd nd nd nd nd nd nd rAAV serotype: + + PBMC PBMC N/A N/A nd nd nd nd 1, 2, 5 (up to (up to month month 2) 2) + + Lymph nodes nodes Lymph N/A N/A nd nd nd nd rAAV dose (total): ( ≥ ≥ month month 3) 3) ( 1,4x10 10 vg to 4,6x10 11 vg Spleen Spleen N/A pending nd nd N/A pending nd nd + + Ciron et al., Liver Liver N/A N/A nd nd nd nd Annals of Neurol., 2006 ( ( ≥ ≥ month month 3) 3) Ciron et al., Manuscript in preparation Gonads Gonads N/A - nd nd N/A - nd nd Joussemet et al., Manuscript in preparation + + α - -transgene transgene α Humoral response response Humoral + + α - -capsid capsid α + ? + ? -transgene transgene α - α Cellular response response Cellular + ? + ? -capsid capsid α - α nd = not done

Vector Vector Infectious Transgene Transgene Infectious Transgene Transgene particles particles mRNA mRNA product product genome genome + + + + Mode of delivery: Target organ organ Target N/A N/A nd nd ( ≥ ( ≥ month month 3) 3) ( ( ≥ ≥ month month 3) 3) Intrathecal + + Serum Serum nd nd nd nd nd nd (up to (up to month month 2) 2) Animal Model: + + + + Cerebrospinal Cerebrospinal nd nd nd nd Fluid Fluid (up to day day 3) 3) (up to month month 3) 3) (up to (up to Cat Urine Urine nd nd nd nd nd nd nd nd rAAV serotype: + + PBMC PBMC N/A nd nd N/A nd nd 1 (up to (up to month month 2) 2) Lymph nodes nodes Lymph N/A N/A pending pending nd nd nd nd rAAV dose (total): 4,6x10 11 vg to 1,4x10 12 vg Spleen Spleen N/A pending nd nd N/A pending nd nd Liver Liver N/A N/A pending pending nd nd nd nd Joussemet et al., Manuscript in preparation Gonads Gonads N/A pending nd nd N/A pending nd nd α - -transgene transgene nd nd α Humoral Humoral response response α - -capsid capsid nd nd α α - -transgene transgene nd nd α Cellular response response Cellular α - -capsid capsid nd nd α nd = not done

Vector Vector Infectious Transgene Transgene Infectious Transgene Transgene particles particles mRNA mRNA product product genome genome + + 44 vg vg/ /cell cell 44 Mode of delivery: Target organ organ Target N/A N/A nd nd ( ≥ ( ≥ y year ear 1,5) 1,5) ( ≥ ( ≥ y year ear 6) 6) Subretinal/Intravitreal + + Serum Serum nd nd nd nd nd nd (up to day (up to day 14) 14) Animal Model: Cerebrospinal Cerebrospinal - - nd nd nd nd nd nd Fluid Fluid Dog, Primate Urine Urine nd nd nd nd nd nd nd nd rAAV serotype: + + PBMC PBMC N/A N/A nd nd nd nd 2, 4, 5 ( ≥ ≥ month month 3 3) ) ( + + Lymph nodes nodes Lymph N/A N/A nd nd nd nd rAAV dose (total): ( ≥ ≥ month month 3) 3) ( 1,6x10 10 vg to 5x10 11 vg Spleen Spleen N/A nd nd nd N/A nd nd nd Provost et al., - Liver - Liver N/A nd nd N/A nd nd Mol. Ther., 2005 + unpub. data Stieger et al., Mol. Ther., 2006 - Gonads - Gonads N/A nd nd N/A nd nd Le Meur et al., Gene Ther., 2006 + + -transgene transgene α - α Humoral Humoral response response - - -capsid capsid α - α α - -transgene transgene nd nd α Cellular response Cellular response α - -capsid capsid nd nd α nd = not done

Vector Vector Infectious Infectious Transgene Transgene Transgene Transgene particles mRNA product particles mRNA product genome genome + + + + 1 to 20 vg 1 to 20 vg/ /cell cell Mode of delivery: Target organ organ Target N/A N/A ( ≥ ( ≥ year year 2) 2) ( ≥ ( ≥ year year 2) 2) ( ≥ ( ≥ year year 2) 2) Isolated limb infusion + + + + + + Serum Serum nd nd (up to day day 7) 7) (up to month month 3) 3) ( ≥ ≥ year year 2) 2) (up to (up to ( Animal Model: Cerebrospinal Cerebrospinal nd nd nd nd nd nd nd nd Fluid Fluid Primate Urine Urine nd nd nd nd nd nd nd nd rAAV serotype: + + PBMC PBMC N/A N/A pending pending nd nd 1, 8 ( ≥ ( ≥ month month 1 1) ) + + Lymph nodes Lymph nodes N/A pending nd N/A pending nd ( ≥ ≥ year year 2) 2) rAAV dose (total): ( + + + + 6,7x10 12 vg to 1,2x10 13 vg Spleen Spleen N/A N/A pending pending ( ( ≥ ≥ year year 2 2) ) ( ≥ ( ≥ year year 1) 1) + + + + Liver Liver N/A N/A nd nd Toromanoff et al., ( ≥ ≥ year year 2) 2) ( ≥ ≥ year year 2) 2) ( ( Manuscript in preparation + + Gonads Gonads N/A nd nd N/A nd nd (up to month month 3) 3) (up to α - -transgene transgene pending pending α Humoral response response Humoral +/- - +/ α - -capsid capsid α -transgene transgene pending α - pending α Cellular response response Cellular -capsid capsid pending α - pending α nd = not done

Vector Vector Infectious Infectious Transgene Transgene Transgene Transgene particles mRNA product particles mRNA product genome genome + + + + 5 to 50 vg 5 to 50 vg/ /cell cell Mode of delivery: Target organ organ Target N/A N/A ( ≥ ( ≥ year year 3) 3) ( ≥ ≥ year year 3) 3) ( ≥ ≥ year year 5) 5) ( ( Intramuscular + + + + + + Serum Serum nd nd (up to day (up to day 7) 7) (up to month (up to month 1,5) 1,5) ( ≥ ( ≥ y year ear 5 5) ) Animal Model: Cerebrospinal Cerebrospinal nd nd nd nd nd nd nd nd Fluid Fluid Primate + + Urine Urine nd nd nd nd nd nd (up to day day 6) 6) rAAV serotype: (up to + + 1, 2, 8 PBMC PBMC N/A N/A pending pending nd nd (up to month month 15) 15) (up to + + + + rAAV dose (total): Lymph nodes nodes Lymph N/A N/A nd nd ( ≥ ( ≥ year year 3) 3) ( ≥ ( ≥ day day 20) 20) 4x10 11 vg to 1,4x10 13 vg + + + + Spleen Spleen N/A pending N/A pending ( ≥ ( ≥ year year 3) 3) ( ≥ ( ≥ year year 2) 2) Favre et al., + + + + Mol. Ther., 2001 Liver Liver N/A N/A nd nd ( ≥ ≥ year year 3) 3) ( ≥ ≥ year year 3) 3) ( ( Chenuaud et al., Unpublished data + + Toromanoff et al., Gonads Gonads N/A N/A nd nd nd nd Manuscript in preparation (up to month month 3) 3) (up to + + -transgene transgene α - α Humoral response Humoral response + + α - -capsid capsid α + + α - -transgene transgene α Cellular response response Cellular + + α - -capsid capsid α nd = not done

One single IM injection of rAAV1, rAAV2, rAAV8 in primate results in widespread detection of vector genome in lymph nodes for years Positive lymph nodes