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Responses to FDA Gene Therapy Letter : Adenovirus Vector Titer Measurements and RCA Levels

Responses to FDA Gene Therapy Letter : Adenovirus Vector Titer Measurements and RCA Levels. BRMAC July 13, 2001 Steven R. Bauer, Ph.D Division of Cellular and Gene Therapies CBER/FDA. Purpose. Updates Development of an adenovirus reference material

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Responses to FDA Gene Therapy Letter : Adenovirus Vector Titer Measurements and RCA Levels

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  1. Responses to FDA Gene Therapy Letter :Adenovirus Vector Titer Measurements and RCA Levels BRMAC July 13, 2001 Steven R. Bauer, Ph.D Division of Cellular and Gene Therapies CBER/FDA

  2. Purpose • Updates • Development of an adenovirus reference material • Change in recommendation on particle to infectious unit (IU) ratio • Change in recommendation on RCA limit • Discussion • Application of RCA recommendation

  3. Adenovirus Vector Final Product non-infectious particles IU: replication defective,infectious particle RCA: replication competent, infectious particle VP: all vector particles

  4. Characterization of Particles in Adenovirus Vector Products • VP: physical/chemical measurement • example : A260 after lysis of VP • IU: biological assay • examples: • measure plaques on a lawn of permissive cells • immunological detection of infected cells with fluorescent antibodies • RCA: biological assay • measure infection on cells that do not complement defective vector

  5. Impact of VP, IU, RCA: Safety, Efficacy, Production • Safety • replication competent virus • exposure to viral proteins • Efficacy • non-transducing particles • Production process

  6. Development of Adenovirus Reference Material • 1993: Adenoviral vectors used in CF Protocols • CF Foundation recommended vector reference standard • 1999: RAC Safety Symposium • RAC AdSAT calls for standards

  7. What Will Be Accomplished by Reference Material Development? • Produce more consistent, safer, adenoviral vectors • particle counts (10% inter-assay variability) • infectious units (over 30% inter-assay variability) • Allow comparability between preclinical and clinical studies • Develop regulatory policy • make recommendations based on “standardized” measurements

  8. Reference Material Development • Adenovirus Reference Material Working Group (ARMWG) • Partnership: Government/Industry/Academia • Williamsburg Bioprocessing Foundation • www.wilbio.com • www.fda.gov/cber/minutes/workshop-min.htm

  9. Adenovirus Reference Material Production Scheme Master Cell Bank ARMWG Meetings March 22, 2001 May 31, 2001 Ad5 wt virus Master Viral Seed Stock Production of purified formulated bulk virus Characterization, safety testing, provisional titer Vialing Particle and Infectivity Titer Determinations, Stability Repository and Distribution

  10. Purpose • Updates • Development of an adenovirus reference material • Change in recommendation on particle to infectious unit (IU) ratio • Change in recommendation on RCA limit • Discussion • Application of RCA recommendation

  11. Changes in Recommendations • ALARA • As Low As Reasonably Achievable • New recommendations based on review of information submitted in March 6 letter responses • Applied to all Adenovirus GT products

  12. Change in Recommendation on Particle to IU ratio • Previous Recommendation • Based on review of production lot data < 100 vp/iu • Current Recommendation • Based on review of production lot data from March 6 letter responses < 30 vp/iu (>3.3% iu)

  13. Change in Recommendation on RCA Limit • Previous recommendation • imprecise since based on infectious titer < 1RCA / 109 iu • Current recommendation • based on particle number • better precision < 1RCA / 3 x 1010 vp

  14. Purpose • Updates • Development of an adenovirus reference material • Change in particle to infectious unit (IU) ratio • Change in recommendation on RCA limit • Discussion • Application of RCA recommendation

  15. Application of RCA Recommendation • < 1RCA / 3 x 1010 vp • recommended for all adenovirus vector lots regardless of clinical use • Exposure risk at highest current doses • dose of 3 x 1013 vp • potential exposure up to 1000 RCA

  16. ALARA vs Risk Based • Shift from ALARA to Risk-Based recommendation? • What information do we have? • Literature, clinical experience with wild-type ad infection/reactivation • Clinical experience with gene transfer studies • Some notable adverse events of unknown cause but indications of innate immuniity • What information do we need?

  17. Wild-type Adenovirus in Bone Marrow Transplantation • Adenovirus, including types 2 and 5, is a significant cause of morbidity and mortality in BMT • Important to consider recipient immune status • Infection/Reactivation

  18. Lessons Learned from Patients with Primary or other Secondary Immunodeficiencies • Neonatal adenoviral pneumonia: Sporadic, severe, localized outbreaks • SCID population at high risk: Severe morbidity and mortality • DiGeoge syndrome: case reports of fatal hepatic necrosis • Solid organ transplant: • Infection (rejection?) of transplanted organ • Source: reactivation and donor • AIDS patients • Co-infection with other pathogens • Diversity of serotypes isolated

  19. Topics for Discussion • Should recommendations regarding acceptable levels of RCA in adenovirus gene transfer products be the same for all clinical uses? • ALARA vs risk-based recommendation for RCA exposure • severely immunosuppressed or immunocompromised patients • mildly immunosuppressed or immunocompromised patients • patients with genetic defects such as hemophilia, cystic fibrosis, or other. • What data should be used to set acceptable limits for RCA exposure? • Should RCA measurements be performed on ex-vivo transduced cells?

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