title: In vivo selection of human hematopoietic cells in a xenograft model using combined pharmacologic and genetic manipulations.
abstract: Strategies that increase the ability of human hematopoietic stem and progenitor cells to repair alkylator-induced DNA damage may prevent the severe hematopoietic toxicity in patients with cancer undergoing high-dose alkylator therapy. In the context of genetic diseases, this approach may allow for selection of small numbers of cells that would not otherwise have a favorable growth advantage. No studies have tested this approach in vivo using human hematopoietic stem and progenitor cells. Human CD34(+) cells were transduced with a bicistronic oncoretrovirus vector that coexpresses a mutant form of O(6)-methylguanine DNA methyltransferase (MGMT(P140K)) and the enhanced green fluorescent protein (EGFP) and transplanted into nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. Mice were either not treated or treated with O(6)-benzylguanine (6BG) and 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). At 8-weeks postinjection, a 2- to 8-fold increase in the percentage of human CD45(+)EGFP(+) cells in 6BG/BCNU-treated versus nontreated mice was observed in the bone marrow and was associated with increased MGMT(P140K)-repair activity. Functionally, 6BG/BCNU-treated mice demonstrated multilineage differentiation in vivo, although some skewing in the maturation of myeloid and B cells was observed in mice transplanted with granulocyte-colony stimulating factor (G-CSF)-mobilized peripheral blood compared to umbilical cord blood. Expansion of human cells in 6BG/BCNU-treated mice was observed in the majority of mice previously transplanted with transduced umbilical cord blood cells. In addition, a significant increase in the number of EGFP(+) progenitor colonies in treated versus nontreated mice were observed in highly engrafted mice indicating that selection and maintenance of human progenitor cells can be accomplished by expression of MGMT(P140K) and treatment with 6BG/BCNU.
authors.affiliation: Herman B Wells Center for Pediatric Research, Department of Pediatrics, The Riley Hospital for Children, Indianapolis, IN 46202, USA. kpollok@iupui.edu
authors.forename: Karen E
authors.initials: KE
authors.lastname: Pollok
authors.forename: Jennifer R
authors.initials: JR
authors.lastname: Hartwell
authors.forename: Annemarije
authors.initials: A
authors.lastname: Braber
authors.forename: Ryan J
authors.initials: RJ
authors.lastname: Cooper
authors.forename: Michael
authors.initials: M
authors.lastname: Jansen
authors.forename: Susanne
authors.initials: S
authors.lastname: Ragg
authors.forename: Barbara J
authors.initials: BJ
authors.lastname: Bailey
authors.forename: Leonard C
authors.initials: LC
authors.lastname: Erickson
authors.forename: Emiko L
authors.initials: EL
authors.lastname: Kreklau
authors.forename: David A
authors.initials: DA
authors.lastname: Williams
mesh_terms: D000818:Animals; D018952:Antigens, CD34; D018906:Antineoplastic Agents, Alkylating / Q000009:adverse effects*; D002330:Carmustine / Q000009:adverse effects*; D002454:Cell Differentiation; D002455:Cell Division; D004249:DNA Damage; D015254:DNA Modification Methylases / Q000235:genetics*; D004260:DNA Repair; D005260:Female; D015316:Genetic Therapy / Q000379:methods; D049452:Green Fluorescent Proteins; D006412:Hematopoietic Stem Cells / Q000276:immunology*; D008164:Luminescent Proteins / Q000235:genetics; D008297:Male; D051379:Mice; D016513:Mice, SCID; D009369:Neoplasms / Q000188:drug therapy; D012641:Selection, Genetic; D014161:Transduction, Genetic; D014183:Transplantation, Heterologous
chemical_list: D018952:Antigens, CD34; D018906:Antineoplastic Agents, Alkylating; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D015254:DNA Modification Methylases; D002330:Carmustine