Fewer chromosome aberrations and earlier apoptosis induced by DNA synthesis inhibitors, a topoisomerase II inhibitor or alkylating agents in human cells with normal compared with mutant p53

Abstract

The human lymphoblastoid cell lines TK6 (normal p53) and WI-L2-NS or WTK1 (mutant p53) differ in sensitivity to killing and induction of gene mutations and chromosome aberrations by ionizing radiation. This may be related to decreased apoptosis in the cells with mutated p53, such that more damaged cells survive. We compared the response of the two cell types to various chemicals. First, to ensure that the thymidine kinase deficiency does not increase the sensitivity of TK6 tk+/- cells to mutagens, we demonstrated that they were not hypersensitive to aberration induction by altered DNA precursor pools or DNA synthesis inhibition, by aphidicolin (APC), methotrexate, hydroxyurea (HU), cytosine arabinoside and thymidine. TK6 cells were then compared with WI-L2-NS or WTK1 cells. With APC, HU, methyl methanesulfonate (MMS), ethyl nitrosourea (ENU) and etoposide (etop), TK6 cells had more apoptosis in the first two days after treatment. Fewer aberrations were seen in normal p53 TK6 cells than the mutant p53 WI-L2-NS cells, ranging from very little difference between the two cell types with MMS to very large differences with ENU and etop. For MMS and ENU we followed cultures for several days, and found that WI-L2-NS cells underwent delayed apoptosis 3 to 5 days after treatment, in parallel with published observations with ionizing radiation. WI-L2-NS cells also had a delayed increase in aberrations (up to 5 days post-treatment) when no aberrations remained in TK6 cells. Colony forming efficiency was measured for APC, MMS and ENU, and was greater in the p53 mutant cells. Our results show that normal p53 function is required for rapid and efficient apoptosis in these lymphoblastoid cells with DNA synthesis inhibitors, alkylating agents and a topoisomerase II inhibitor, and support the hypothesis that induced levels of aberrations are higher in p53 mutant cells because of a failure to remove damaged cells by apoptosis.