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Background: Resistance to taxane-based therapy in breast cancer patients is a major clinical problem that may be addressed through insight of the genomic alterations leading to taxane resistance in breast cancer cells. In the current study we used whole exome sequencing to discover somatic genomic alterations, evolving across evolutionary stages during the acquisition of docetaxel resistance in breast cancer cell lines. Results: Two human breast cancer in vitro models (MCF-7 and MDA-MB-231) of the step-wise acquisition of docetaxel resistance were developed by exposing cells to 18 gradually increasing concentrations of docetaxel. Whole exome sequencing performed at five successive stages during this process was used to identify single point mutational events, insertions/deletions and copy number alterations associated with the acquisition of docetaxel resistance. Acquired coding variation undergoing positive selection and harboring characteristics likely to be functional were further prioritized using network-based approaches. A number of genomic changes were found to be undergoing evolutionary selection, some of which were likely to be functional. Of the five stages of progression toward resistance, most resistance relevant genomic variation appeared to arise midway towards fully resistant cells corresponding to passage 31 (5 nM docetaxel) for MDA-MB-231 and passage 16 (1.2 nM docetaxel) for MCF-7, and where the cells also exhibited a period of reduced growth rate or arrest, respectively. MCF-7 cell acquired several copy number gains on chromosome 7, including ABC transporter genes, including ABCB1 and ABCB4, as well as DMTF1, CLDN12, CROT, and SRI. For MDA-MB-231 numerous copy number losses on chromosome X involving more than 30 genes was observed. Of these genes, CASK, POLA1, PRDX4, MED14 and PIGA were highly prioritized by the applied network-based gene ranking approach. At higher docetaxel concentration MCF-7 subclones exhibited a copy number loss in E, Background: Resistance to taxane-based therapy in breast cancer patients is a major clinical problem that may be addressed through insight of the genomic alterations leading to taxane resistance in breast cancer cells. In the current study we used whole exome sequencing to discover somatic genomic alterations, evolving across evolutionary stages during the acquisition of docetaxel resistance in breast cancer cell lines. Results: Two human breast cancer in vitro models (MCF-7 and MDA-MB-231) of the step-wise acquisition of docetaxel resistance were developed by exposing cells to 18 gradually increasing concentrations of docetaxel. Whole exome sequencing performed at five successive stages during this process was used to identify single point mutational events, insertions/deletions and copy number alterations associated with the acquisition of docetaxel resistance. Acquired coding variation undergoing positive selection and harboring characteristics likely to be functional were further prioritized using network-based approaches. A number of genomic changes were found to be undergoing evolutionary selection, some of which were likely to be functional. Of the five stages of progression toward resistance, most resistance relevant genomic variation appeared to arise midway towards fully resistant cells corresponding to passage 31 (5 nM docetaxel) for MDA-MB-231 and passage 16 (1.2 nM docetaxel) for MCF-7, and where the cells also exhibited a period of reduced growth rate or arrest, respectively. MCF-7 cell acquired several copy number gains on chromosome 7, including ABC transporter genes, including ABCB1 and ABCB4, as well as DMTF1, CLDN12, CROT, and SRI. For MDA-MB-231 numerous copy number losses on chromosome X involving more than 30 genes was observed. Of these genes, CASK, POLA1, PRDX4, MED14 and PIGA were highly prioritized by the applied network-based gene ranking approach. At higher docetaxel concentration MCF-7 subclones exhibited a copy number loss in E

BACKGROUND: Studies in taxane and/or anthracycline refractory metastatic breast cancer (mBC) patients have shown approximately 30 % response rates to irinotecan. Hence, a significant number of patients will experience irinotecan-induced side effects without obtaining any benefit. The aim of this study was to lay the groundwork for development of predictive biomarkers for irinotecan treatment in BC.METHODS: We established BC cell lines with acquired or de novo resistance to SN-38, by exposing the human BC cell lines MCF-7 and MDA-MB-231 to either stepwise increasing concentrations over 6 months or an initial high dose of SN-38 (the active metabolite of irinotecan), respectively. The resistant cell lines were analyzed for cross-resistance to other anti-cancer drugs, global gene expression, growth rates, TOP1 and TOP2A gene copy numbers and protein expression, and inhibition of the breast cancer resistance protein (ABCG2/BCRP) drug efflux pump.RESULTS: We found that the resistant cell lines showed 7-100 fold increased resistance to SN-38 but remained sensitive to docetaxel and the non-camptothecin Top1 inhibitor LMP400. The resistant cell lines were characterized by Top1 down-regulation, changed isoelectric points of Top1 and reduced growth rates. The gene and protein expression of ABCG2/BCRP was up-regulated in the resistant sub-lines and functional assays revealed BCRP as a key mediator of SN-38 resistance.CONCLUSIONS: Based on our preclinical results, we suggest analyzing the predictive value of the BCRP in breast cancer patients scheduled for irinotecan treatment. Moreover, LMP400 should be tested in a clinical setting in breast cancer patients with resistance to irinotecan.