Down-regulation of DNA repair in human CD34(+) progenitor cells corresponds to increased drug sensitivity and apoptotic response.
Although DNA repair processes have been shown to considerably modulate the cytotoxic effects of alkylating agents, little information is available on the role of these mechanisms in chemotherapy-induced myelosuppression. Therefore, we have analyzed in detail the DNA repair capacity of primary human hematopoietic cells from cord blood (CB) or bone marrow (BM) by 2 functional assays, the immunocytologic assay (ICA) and single-cell gel electrophoresis (comet assay). Besides substantial interindividual differences, we consistently observed significantly lower repair capacity of CD34(+) cells in comparison to CD34(-), CD19(+), or CD33(+) cells of the same donor. After exposure to the alkylating agent ethylnitrosourea (EtNU), the comet assay displayed on average twice as many DNA single-strand breaks (SSBs) in CD34(+) cells and a tripled half-life of these lesions in comparison to corresponding CD34(-) cells. Similarly, reduced SSB repair activity in CD34(+) cells was detected following melphalan or cisplatin application. When specific antibodies were used to monitor DNA reaction products of these drugs, adduct levels were significantly higher and lesions persisted longer in the CD34(+) fraction. To assess the contribution of individual pathways to overall DNA repair, modulators blocking defined steps in repair processes were coapplied with alkylating drugs. Similar "modulation pattern" in corresponding CD34(+) and CD34(-) cell fractions indicated a generalized reduction in DNA repair capacity of CD34(+) cells, rather than deficiencies in a specific pathway. Because CD34(+) cells also displayed higher frequencies of apoptosis in response to melphalan or cisplatin, these findings may help to explain the myelosuppression after exposure to alkylating agents.
Buschfort-Papewalis
Claudia
C
Institute of Cell Biology and Department of Internal Medicine, Center for Cancer Research and Cancer Treatment, University of Essen Medical School, Essen, Germany.
Moritz
Thomas
T
Liedert
Bernd
B
Thomale
Jürgen
J
D018952:Antigens, CD34 / Q000097:blood*; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology; D017209:Apoptosis / Q000235:genetics*; D002945:Cisplatin / Q000494:pharmacology; D020552:Comet Assay; D018736:DNA Adducts; D004249:DNA Damage; D004260:DNA Repair; D015536:Down-Regulation; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D006412:Hematopoietic Stem Cells / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D007700:Kinetics; D008558:Melphalan / Q000494:pharmacology
D018952:Antigens, CD34; D018906:Antineoplastic Agents, Alkylating; D018736:DNA Adducts; D002945:Cisplatin; D008558:Melphalan