Abstract
Abstract Background: Aurora kinases are promising targets for cancer chemotherapy. Many reports have been published that some cancers overexpress Aurora kinases, which correlates with tumor grade and poor prognosis. Hence Aurora kinases were considered key genes for cancerogenesis and progression. Many Aurora kinase inhibitors(AKI,s) are in various phases of pre-clinical and clinical development and promising anticancer effects have been reported. Tumor resistance remains one of the major problems in chemotherapy. Objectives: Our work is mainly aimed at identification of potential resistance mechanisms towards CYC116 (new small molecule pan Aurora kinase inhibitor) and ZM447439. The main aims include generation and selection of resistant tumor cell clones, characterization of resistance, cross resistance with other AKI,s and multidrug resistance. Further characterization of the resistant clones included cell cycle analysis, expression and DNA sequencing of Aurora kinases, cellular target inhibition, and expression of common drug transporters.Gene expression analysis of the resistant clones was carried out to reveal differential gene expression profiles. Results: The isogenic pair HCT116 cell lines (p53+/+ and p53-/-) were used to study CYC116 and ZM447439 induced resistance. HCT116p53+/+ (called HCT116) and HCT116p53-/- resistant clones were generated. HCT116:CYC116, HCT116p53-/-:CYC116, HCT116:ZM447439, HCT116p53-/-:ZM447439 clones were 9-82 fold, 36-64 fold, 18 – >83 fold, and 33-39 fold resistant respectively to selecting agents. They were also cross resistant to other AKI,s. 13 approved anticancer drugs were used to determine multidrug resistant phenotype. All clones were highly resistant to etoposide(5-53 fold). As expected, ZM447439 clones contained mutated Aurora B; three novel mutation sites were identified. No mutations of Aurora A or B were identified in CYC116 resistant clones. Instead these became stably tetraploid. Aurora kinases expression were similar to that of parent cells, however no inhibition of phospho Histone H3 (Aurora B target) was observed at effective CYC116 and ZM447439 concentrations. Flowcytometry studies revealed no overexpression of PgP or MRP1 transporters. Microarray gene expression analysis showed large number of genes which were differentially expressed in the resistant clones in comparison with the parent cell lines. A number of these were validated by qRT-PCR. Further analysis will be carried out to identify gene expression signature of CYC116 induced resistance. Using the developed CYC116 resistant clones we have identified chemotherapeutic agents which are highly effective in the induced phenotype. Our results can assist the further development of CYC116, a pan Aurora inhibitor which cross-reacts with VEGFR2, has potent anti-mitotic and anti-angiogenic activity in vivo and is tested in Ph I clinical trial. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 735. doi:10.1158/1538-7445.AM2011-735
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