title: Characterizing clearance of helper adenovirus by a clinical rAAV1 manufacturing process.
abstract: Recombinant adeno-associated viral vectors (rAAV) are being developed as gene therapy delivery vehicles and as genetic vaccines, and some of the most scaleable manufacturing methods for rAAV use live adenovirus to induce production. One aspect of establishing safety of rAAV products is therefore demonstrating adequate and reliable clearance of this helper virus by the vector purification process. The ICH Q5A regulatory guidance on viral safety provides recommendations for process design and characterization of viral clearance for recombinant proteins, and these principles were adapted to a rAAV serotype 1 purification process for clinical vectors. Specific objectives were to achieve overall adenovirus clearance factors significantly greater than input levels by using orthogonal separation and inactivation methods, and to segregate adenovirus from downstream operations by positioning a robust clearance step early in the process. Analytical tools for process development and characterization addressed problematic in-process samples, and a viral clearance validation study was performed using adenovirus and two non-specific model viruses. Overall clearance factors determined were >23 LRV for adenovirus, 11 LRV for BVDV, and >23 LRV for AMuLV.
authors.affiliation: Targeted Genetic Corporation, Process Development, 1100 Olive Way, Suite 100, Seattle, WA 98101, USA. barb.thorne@targen.com
authors.forename: Barbara A
authors.initials: BA
authors.lastname: Thorne
authors.forename: Paulene
authors.initials: P
authors.lastname: Quigley
authors.forename: Gina
authors.initials: G
authors.lastname: Nichols
authors.forename: Christine
authors.initials: C
authors.lastname: Moore
authors.forename: Eric
authors.initials: E
authors.lastname: Pastor
authors.forename: David
authors.initials: D
authors.lastname: Price
authors.forename: Jon W
authors.initials: JW
authors.lastname: Ament
authors.forename: Ryan K
authors.initials: RK
authors.lastname: Takeya
authors.forename: Richard W
authors.initials: RW
authors.lastname: Peluso
mesh_terms: D000256:Adenoviridae / Q000302:isolation & purification* / Q000502:physiology; D036022:Capsid Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D002460:Cell Line; D000229:Dependovirus / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology*; D005822:Genetic Vectors / Q000096:biosynthesis* / Q000235:genetics; D006378:Helper Viruses / Q000302:isolation & purification* / Q000502:physiology; D006358:Hot Temperature; D006801:Humans; D038301:Virus Inactivation
chemical_list: D036022:Capsid Proteins