Repair of O6-alkylguanines in the nuclear DNA of human lymphocytes and leukaemic cells : analysis at the single-cell level

• Published in: British journal of cancer Vol. 69; no. 4; pp. 698 - 705
• Main Authors: THOMALE, J, SEILER, F, MÜLLER, M. R, SEEBER, S, RAJEWSKY, M. F
• Format: Journal Article
• Language: English
• Published: Basingstoke Nature Publishing Group 01.04.1994
• Subjects: Alkylating Agents - pharmacology; Antibodies, Monoclonal; Biological and medical sciences; Blast Crisis - genetics; Blast Crisis - metabolism; DNA Repair - physiology; DNA, Neoplasm - metabolism; Drug Resistance - physiology; Guanine Nucleotides - metabolism; Hematologic and hematopoietic diseases; Humans; Leukemia, Myelogenous, Chronic, BCR-ABL Positive - genetics; Leukemia, Myelogenous, Chronic, BCR-ABL Positive - metabolism; Leukemias. Malignant lymphomas. Malignant reticulosis. Myelofibrosis; Lymphocytes - drug effects; Lymphocytes - metabolism; Medical sciences; Methyltransferases - metabolism; Microscopy, Fluorescence; Nitrosourea Compounds - toxicity; O-Methylguanine-DNA Methyltransferase; Precursor Cell Lymphoblastic Leukemia-Lymphoma - genetics; Precursor Cell Lymphoblastic Leukemia-Lymphoma - metabolism; Human; Chronic; Chronic lymphocytic leukemia; Lymphoproliferative syndrome; Acute; DNA; Malignant hemopathy; Repair; Lymphocyte; Alkylation; Guanine; Acute myelocytic leukemia
• ISSN: 0007-0920; 1532-1827
• DOI: 10.1038/bjc.1994.132

Inter-individual and cell-cell variability of repair of O6-alkylguanines (O6-AlkGua) in nuclear DNA was studied at the single-cell level in peripheral lymphocytes from healthy donors and in leukaemic cells isolated from patients with chronic lymphatic leukaemia (CLL) or acute myeloid leukaemia (AML). Cells were pulse exposed to N-ethyl- or N-(n-)butyl-N-nitrosourea in vitro, and O6-AlkGua residues in DNA were quantified using an anti-(O6-AlkGua) monoclonal antibody and electronically intensified fluorescence. The kinetics of O6-AlkGua elimination revealed considerable inter-individual differences in O6-ethylguanine (O6-EtGua) half-life (t1/2) values in DNA, ranging from 1.5 to 4.5 h (five AML patients), from 0.8 to 2.8 h (five CLL patients) and from 1.2 to 7.3 h (five healthy donors). The elimination from DNA of equimolar amounts of O6-butylguanine was generally 3-5 times slower in comparison with O6-EtGua. The t1/2 values of individual samples varied in parallel for both DNA alkylation products. Upon preincubation with O6-benzylguanine, the activity of the DNA repair protein O6-alkylguanine-DNA alkyltransferase (AT) in both lymphocytes and leukaemic blasts was reduced to < or = 1%. However, while the rate of O6-EtGua elimination from DNA was decelerated it was not abolished, suggesting the possible involvement of additional repair systems that might be co-regulated with AT. Within individual samples, no major cell subpopulations were observed whose repair kinetics would differ significantly from the remaining cells.