title: Correction of murine hemophilia A following nonmyeloablative transplantation of hematopoietic stem cells engineered to encode an enhanced human factor VIII variant using a safety-augmented retroviral vector.
abstract: Insertional mutagenesis by retroviral vectors is a major impediment to the clinical application of hematopoietic stem cell gene transfer for the treatment of hematologic disorders. We recently developed an insulated self-inactivating gammaretroviral vector, RMSinOFB, which uses a novel enhancer-blocking element that significantly decreases genotoxicity of retroviral integration. In this study, we used the RMSinOFB vector to evaluate the efficacy of a newly bioengineered factor VIII (fVIII) variant (efVIII)--containing a combination of A1 domain point mutations (L303E/F309S) and an extended partial B domain for improved secretion plus A2 domain mutations (R484A/R489A/P492A) for reduced immunogenicity--toward successful treatment of murine hemophilia A. In cell lines, efVIII was secreted at up to 6-fold higher levels than an L303E/F309S A1 domain-only fVIII variant (sfVIIIDeltaB). Most important, when compared with a conventional gammaretroviral vector expressing sfVIIIDeltaB, lower doses of RMSin-efVIII-OFB-transduced hematopoietic stem cells were needed to generate comparable curative fVIII levels in hemophilia A BALB/c mice after reduced-intensity total body irradiation or nonmyeloablative chemotherapy conditioning regimens. These data suggest that the safety-augmented RMSin-efVIII-OFB platform represents an encouraging step in the development of a clinically appropriate gene addition therapy for hemophilia A.
authors.affiliation: Department of Anatomy and Regenerative Biology, The George Washington University Medical Center, 2300 I Street NW, Washington, DC, USA.
authors.forename: Ali
authors.initials: A
authors.lastname: Ramezani
authors.forename: Robert G
authors.initials: RG
authors.lastname: Hawley
mesh_terms: D000818:Animals; D005169:Factor VIII / Q000008:administration & dosage* / Q000235:genetics / Q000378:metabolism; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000378:metabolism; D006467:Hemophilia A / Q000628:therapy*; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D009154:Mutation; D015202:Protein Engineering; D012190:Retroviridae / Q000235:genetics; D014161:Transduction, Genetic; D019172:Transplantation Conditioning
chemical_list: C078147:F8 protein, human; D005169:Factor VIII