Molecular analysis of DNA damage induced by a novel trinuclear platinum complex (BBR 3464)

Colella, Gennaro Giovanni Domenico (2001). Molecular analysis of DNA damage induced by a novel trinuclear platinum complex (BBR 3464). PhD thesis The Open University.



We evaluated the activity of the trinuclear platinum complex BBR 3464 in two human ovarian carcinoma cell lines (OAW42, A2780) and in their cisplatin-resistant counterparts (OAW42Mer, A2780cp8). An increased cytotoxic potency of BBR 3464 compared to cisplatin was generally observed, an a collateral sensitivity or a very modest cross-resistance to BBR 3464 was found in OAW42Mer and A2780cp8 cell lines, respectively. Loss of mismatch repair proteins (hMLHl, hPMS2) or overexpression of nucleotide excision repair proteins (ERCC1) was not detrimental for the cellular sensitivity to the trinuclear platinum complex. BBR 3464 intracellular accumulation and DNA-bound platinum were consistently higher than those observed with cisplatin. After exposure to BBR 3464 and cisplatin of purified DNA or intact cells, a similar sequence preference of DNA damage was observed. Conversely, interesting differences in the kinetics of formation and removal of DNA lesions at the single-gene (N-ras) level were observed between the two drugs.

The interference exerted by BBR 3464 with cell cycle progression and its ability to induce apoptosis were evaluated in OAW42 and OAW42Mer cell lines. Flow cytometric experiments indicated that in the two cell lines BBR 3464 was able to induce a persistent G2M block whereas cisplatin caused an initial accumulation of cells in the S phase followed by an increase in the G2M cell fraction. Exposure to IC50 drug concentrations induced apoptosis in both cell lines. However, the percentage of cells with an apoptotic nuclear morphology was slightly higher after cisplatin than BBR 3464 treatment in OAW42 cells, whereas the opposite pattern was observed in OAW42Mer cells. Degradation of the nuclear lamin B was detected in OAW42 cells after exposure to each drug whereas in OAW42Mer cells the cleavage was only appreciable after BBR 3464 exposure. In OAW42 cells the mitochondrial membrane potential (ɅѰmt) was not affected by the two drugs, whereas in the OAW42Mer cell line a marked (ɅѰmt) reduction was observed only after exposure to BBR 3464.

Overall, the results would suggest that the collateral sensitivity to BBR 3464 observed in the OAW42Mer cell line might be attributable to the ability of this drug to modify DNA differently from that of cisplatin and, as a consequence, to induce different cellular responses to DNA damage such as the triggering of specific apoptotic pathways.

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