title: Chemoprotective gene transfer II: multilineage in vivo protection of haemopoiesis against the effects of an antitumour agent by expression of a mutant human O6-alkylguanine-DNA alkyltransferase.
abstract: Murine bone marrow cells were transduced ex vivo with a retrovirus encoding an O6-benzylguanine (O6-beG) insensitive, double mutant form of the human DNA repair protein O6-alkylguanine-DNA alkyltransferase (hATPA/GA). In animals reconstituted with the transduced bone marrow, about 50% of cells in the multipotent spleen colony-forming cells (CFU-S) and lineage restricted granulocyte-macrophage (GM-CFC) haemopoietic progenitor populations were found to be carrying the transgene and this correlated with the frequency of bone marrow cells and spleen colonies which stained positive for hATPA/GA by immunocyto-chemistry. Expression of hATPA/GA was associated with significant in vivo protection of both CFU-S (P = 0.001) and GM-CFC (P < 0.024) against the toxicity of the antitumour methylating agent, temozolomide, given in combination with O6-beG. Expression of hATPA/GA also led to a reduction in the frequency of combined O6-beG/temozolomide-induced micronuclei seen in polychromatic erythrocytes (P < 0.003). This study is the first to demonstrate in vivo protection of multipotent haemopoietic progenitors against the toxic and clastogenic effects of an O6-alkylating agent in the presence of O6-beG. It also represents the first report of reduced clastogenesis as a consequence of expression of an O6-beG-resistant ATase. In the accompanying article we report hATPA/GA-mediated resistance of human CD34+ haemopoietic progenitors to combined O6-beG/O6-alkylating agent toxicity. Together these two reports suggest that a gene therapy strategy whereby protection of normal haemopoietic tissue may be combined with O6-beG-mediated tumour sensitisation may be efficacious in achieving an increase in therapeutic index.
authors.affiliation: CRC Sections of Genome Damage and Repair, Paterson Institute for Cancer Research, Manchester, UK.
authors.forename: N
authors.initials: N
authors.lastname: Chinnasamy
authors.forename: J A
authors.initials: JA
authors.lastname: Rafferty
authors.forename: I
authors.initials: I
authors.lastname: Hickson
authors.forename: L S
authors.initials: LS
authors.lastname: Lashford
authors.forename: S J
authors.initials: SJ
authors.lastname: Longhurst
authors.forename: N
authors.initials: N
authors.lastname: Thatcher
authors.forename: G P
authors.initials: GP
authors.lastname: Margison
authors.forename: T M
authors.initials: TM
authors.lastname: Dexter
authors.forename: L J
authors.initials: LJ
authors.lastname: Fairbairn
mesh_terms: D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000633:toxicity*; D001854:Bone Marrow Cells; D002470:Cell Survival; D003606:Dacarbazine / Q000031:analogs & derivatives* / Q000633:toxicity; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012190:Retroviridae; D013154:Spleen / Q000166:cytology; D013234:Stem Cells; D000077204:Temozolomide
chemical_list: D018906:Antineoplastic Agents, Alkylating; D003606:Dacarbazine; D019853:O(6)-Methylguanine-DNA Methyltransferase; D000077204:Temozolomide