title: Highly efficient transduction of repopulating bone marrow cells using rapidly concentrated polymer-complexed retrovirus.
abstract: Using the cationic polymer, Polybrene, and the anionic polymer, chondroitin sulfate C, we concentrated recombinant retrovirus pseudotyped with an ecotropic envelope, which is susceptible to inactivation by high-speed concentration methods. To evaluate gene marking, murine bone marrow was harvested from C3H mice, transduced with polymer-concentrated GFP virus, and transplanted into lethally irradiated recipients. Total gene marking in mice averaged 30-35% at 8 weeks post-transplant and transgene expression remained stable for over 16 weeks. Using the polymer concentration method, a second retroviral vector encoding the drug resistant variant of dihydrofolate reductase (L22Y-DHFR) was concentrated and tested. Approximately 40% of transduced murine bone marrow progenitor cells were protected against trimetrexate concentrations that completely eliminated the growth of non-modified cells. These results show that anionic and cationic polymers can be combined to rapidly concentrate viruses that are normally difficult to concentrate, and the concentrated virus efficiently transduces hematopoietic stem cells.
authors.affiliation: Department of Pediatrics, Division of Hematology/Oncology and Bone Marrow Transplantation, Emory University School of Medicine, Atlanta, GA 30322, USA.
authors.forename: Douglas W
authors.initials: DW
authors.lastname: McMillin
authors.forename: Natalia
authors.initials: N
authors.lastname: Landázuri
authors.forename: Bagirath
authors.initials: B
authors.lastname: Gangadharan
authors.forename: Becker
authors.initials: B
authors.lastname: Hewes
authors.forename: David R
authors.initials: DR
authors.lastname: Archer
authors.forename: H Trent
authors.initials: HT
authors.lastname: Spencer
authors.forename: Joseph M
authors.initials: JM
authors.lastname: Le Doux
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology* / Q000378:metabolism* / Q000821:virology; D002460:Cell Line; D002809:Chondroitin Sulfates / Q000737:chemistry; D006583:Hexadimethrine Bromide / Q000737:chemistry; D051379:Mice; D011108:Polymers / Q000737:chemistry*; D012190:Retroviridae / Q000737:chemistry* / Q000235:genetics* / Q000502:physiology; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics / Q000378:metabolism; D014161:Transduction, Genetic / Q000295:instrumentation / Q000379:methods*; D019076:Transgenes / Q000235:genetics
chemical_list: D011108:Polymers; D006583:Hexadimethrine Bromide; D002809:Chondroitin Sulfates; D013762:Tetrahydrofolate Dehydrogenase