Received for publication September 21, 2005.
Revised October 26, 2005.
Accepted for publication October 27, 2005.

DNA Repair Enzyme, O⁶-Methylguanine DNA Methyltransferase, Modulates Cytotoxicity of Camptothecin-Derived Topoisomerase I Inhibitors

Abstract

Two camptothecin-resistant cell lines, CPT30 and KB100, were established and characterized previously in our laboratory. Because enhanced sensitivity to 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) and decreased expression of O⁶-methylguanine-DNA methyltransferase (MGMT) protein were observed in these lines, we hypothesized that MGMT may be a determinant of cytotoxicity associated with camptothecin-derived DNA topoisomerase I inhibitors (CPTs). We used the Tet-On system to induce expression of MGMT in Chinese hamster ovary (CHO) cells and RNA interference to knockdown MGMT expression in human nasopharyngeal carcinoma HONE-1 cells in order to identify any correlations between MGMT expression and CPTs cytotoxicity. CHO-derived Tet-On inducible cells (S12+) showed MGMT overexpression and statistically significant more resistance to BCNU, camptothecin, 7-ethyl-10-hydrocamptothecin (SN38), and topotecan (TPT) than parental CHO cells (p < 0.05), but there was less resistance to CPTs than to BCNU. Knockdown of MGMT expression with siRNA in HONE-1 cells conferred increased sensitivity to BCNU and CPTs compared with mock control. Furthermore, alteration of MGMT expression coincides with CPT-induced cell death and poly (ADP-ribose) polymerase (PARP) cleavage. There were no differences in protein levels and catalytic activity of topoisomerase I between MGMT-proficient and MGMT-deficient cells from the Tet-On inducible and siRNA systems. Resistance to CPTs coincided with decreased amounts of protein linked-DNA breaks generated by CPTs in MGMT-proficient cells, and vice versa in MGMT-deficient cells. Our data indicate that MGMT can modulate cytotoxicity of CPT-derived Top I inhibitors.

Key words: Camptothecin-derived topoisomerase I inhibitors, Cell death, Cytotoxicity, MGMT, PARP, Protein-linked DNA breaks

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