Mechanism of cellular response to anticancer drugs: Involvement of ATM and topoisomerase

The first part of the dissertation involves the roles of ATM in histone acetylation-mediated gene activation. Induction of p21WAF1 by histone deacetylase inhibitors (FR901228 and trichostatin A) is defective in Ataxia telangiectacia (AT) cells. Caffeine and wortmannin attenuate p21 WAF1 induction in normal cells. It is also observed that ectopic expression of the wild-type ATM gene in an AT cell line restores p21WAF1 induction by the HDAC inhibitors. Besides the p21WAF1 promoter, activation of topoisomerase IIIalpha and SV40 promoters by the HDAC inhibitors are also decreased in the AT cell lines. Finally, despite the deficiency of induction of gene expression, the overall levels of H3 and H4 histone acetylation appear to be the same between AT and normal cells in response to HDAC inhibitor treatments. Taken together, the data indicate that ATM is involved in histone acetylation-mediated gene expression regulation.;The second part of the dissertation involves investigation of camptothecin (CPT) resistance in multiple myeloma (MM). In order to identify the resistance, the relationship between topoisomerase I (topo I) and CPT cytotoxicity in six MM cell lines as well as the effect of CPT combined with UCN-01 were investigated.;Among the six cell lines, three of them displayed relatively resistant to CPT. Despite the fact that topo I polypeptide levels and CPT-induced DNA-topo I cleavable complex levels varied among these cell lines, cytotoxicity correlated with topo I levels as well as DNA-topo I cleavable complex levels only in those cells possessing either very high or very low complex levels.;To identify potential approaches to overcome the resistance, CPT was combined with UCN-01 to treat two CPT resistant myeloma cell lines (8226/DOX40 and SKO-007). Cytotoxicity assays showed that UCN-O1 significantly enhanced CPT-induced cytotoxicity in both cell lines. Addition of UCN-01 to CPT specifically abrogated the S-phase checkpoint induced by CPT. Furthermore, CPT-induced DNA synthesis inhibition was partially abolished in 8226/DOX40. As a result, an increase of DNA double strand breaks (DSB) represented by gamma-H2AX was observed.