title: Replication-competent lentivirus analysis of clinical grade vector products.
abstract: Lentiviral vectors are now in clinical trials for a variety of inherited and acquired disorders. A challenge for moving any viral vector into the clinic is the ability to screen the vector product for the presence of replication-competent virus. Assay development for replication-competent lentivirus (RCL) is particularly challenging because recombination of vector packaging plasmids and cellular DNA leading to RCL has not been reported with the current viral vector systems. Therefore, the genomic structure of a RCL remains theoretical. In this report, we describe a highly sensitive RCL assay suitable for screening vector product and have screened large-scale vector supernatant, cells used in vector production, and cells transduced with clinical grade vector. We discuss the limitations and challenges of the current assay, and suggest modifications that may improve the suitability of this assay for screening US Food and Drug Administration (US FDA)-licensed products.
authors.affiliation: Department of Medical and Molecular Genetics, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA. kcornett@iupui.edu
authors.forename: Kenneth
authors.initials: K
authors.lastname: Cornetta
authors.forename: Jing
authors.initials: J
authors.lastname: Yao
authors.forename: Aparna
authors.initials: A
authors.lastname: Jasti
authors.forename: Sue
authors.initials: S
authors.lastname: Koop
authors.forename: Makhaila
authors.initials: M
authors.lastname: Douglas
authors.forename: David
authors.initials: D
authors.lastname: Hsu
authors.forename: Larry A
authors.initials: LA
authors.lastname: Couture
authors.forename: Troy
authors.initials: T
authors.lastname: Hawkins
authors.forename: Lisa
authors.initials: L
authors.lastname: Duffy
mesh_terms: D002460:Cell Line; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics / Q000302:isolation & purification* / Q000592:standards* / Q000627:therapeutic use; D057809:HEK293 Cells; D016655:HIV Core Protein p24 / Q000276:immunology; D006801:Humans; D016086:Lentivirus / Q000235:genetics / Q000302:isolation & purification*; D011786:Quality Control; D011995:Recombination, Genetic; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014779:Virus Replication
chemical_list: D016655:HIV Core Protein p24