The substituted
phenazines XR11576 and
XR5944 were originally described as dual
topoisomerase-I/II
poisons. Subsequent reports, however, indicated that the association of their cytotoxicity with cellular topoisomerases was not clear. We set out to study this further using human tumour cell lines, PEO1
ovarian cancer, MDA-MB-231
breast cancer and variants with acquired resistance to
VP-16 and
XR11576: PEO1VPR, MB-231VPR, MB-231-11576R and
camptothecin: PEO1CamR. Cytotoxicity testing [3-(4,5-dimethylthiazol-2yl)-2,5-
diphenyl tetrazolium
bromide assay],
DNA-
protein crosslink formation, cell cycle analysis (flow cytometry) for
DNA content, apoptosis (flow cytometry) for
Annexin V and Western blotting for apoptotic factors. Cytotoxicity testing showed potent cytotoxicity with no cross-resistance to
XR11576 or
XR5944 in
VP-16 or
camptothecin-resistant lines. Importantly, we have shown for the first time that the activities of
XR5944 and
XR11576 are similar as MB-231-11576R cells were resistant to both agents and to a similar extent.
XR5944 showed the greatest, albeit slower, interaction with
DNA with high levels of
DNA-
protein crosslinks. Levels of apoptosis in XR5944-treated cells were significantly less than those in
VP-16 or
XR11576 treatments, suggestive of a more
cytostatic rather than cytotoxic mode of action. Interestingly,
XR5944 failed to give rise to a G2/M blockade, in contrast to
VP-16 or
XR11576.
XR5944 and
XR11576, in line with a dual
topoisomerase-I/II-directed mechanism of action, retain potent activity in tumour cells with acquired resistance to
VP-16 and
camptothecin. Although these agents appear to behave differently from each other according to experimental conditions, this study suggests a substantial overlap in their mechanism(s) of action.