Retrovirus-mediated expression of a DNA repair protein in bone marrow protects hematopoietic cells from nitrosourea-induced toxicity in vitro and in vivo.
Severe and delayed myelosuppression is a major side effect encountered with the clinical use of nitrosourea-type chemotherapeutic drugs. The DNA repair protein O6-methylguanine DNA methyltransferase (MGMT) has been shown to repair nitrosourea-induced DNA damage. We therefore investigated the effect of expressing MGMT in hematopoietic cells (via retrovirus-mediated gene transfer) on nitrosourea-induced toxicity. A retroviral vector (N2/ZipPGK-MGMT) expressing the human MGMT cDNA from the phosphoglycerate kinase promoter was constructed. Infection of murine bone marrow with the N2/ZipPGK-MGMT retrovirus significantly increased the survival of murine bone marrow-committed progenitor cells following in vitro exposure to N-N'-bis(2-chloroethyl)-N-nitrosourea (BCNU, carmustine). MGMT gene transfer also protected murine hematopoietic cells in vivo in a murine model of BCNU-induced myelosuppression. The infusion of 4-6 x 10(6) N2/ZipPGK-MGMT-transduced bone marrow cells into mice every 2 weeks significantly increased peripheral leukocyte counts, platelet counts, and hematocrits compared to infusions of mock-infected bone marrow cells. In addition, bone marrow-committed progenitor cells from some recipient animals demonstrated increased resistance to BCNU in vitro when analyzed 2.5 months after initial treatment. The integration of the N2/ZipPGK-MGMT provirus in the spleen DNA from these animals correlated with committed progenitor cell resistance to BCNU. These data suggest that MGMT expression in hematopoietic progenitor and precursor cells protects against nitrosourea-induced toxicity and that gene transfer may prove useful in attempts to reduce nitrosourea-induced myelosuppression in the clinical setting.
Moritz
T
T
Herman B Wells Center for Pediatric Research, James Whitcomb Riley Hospital for Children, Indiana University School of Medicine, Indianapolis 46202-5225, USA.
Mackay
W
W
Glassner
B J
BJ
Williams
D A
DA
Samson
L
L
D016475:3T3 Cells; D000818:Animals; D015153:Blotting, Western; D001854:Bone Marrow Cells; D002330:Carmustine / Q000633:toxicity*; D002478:Cells, Cultured; D004247:DNA / Q000032:analysis; D004260:DNA Repair; D004305:Dose-Response Relationship, Drug; D004351:Drug Resistance; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology*; D006801:Humans; D008780:Methyltransferases / Q000096:biosynthesis / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D019853:O(6)-Methylguanine-DNA Methyltransferase; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D012190:Retroviridae; D014162:Transfection
D011994:Recombinant Proteins; D004247:DNA; D008780:Methyltransferases; D019853:O(6)-Methylguanine-DNA Methyltransferase; D002330:Carmustine