Abstract 831: Increased resistance of breast cancer cells to doxorubicin by clinically relevant doses of temsirolimus

Abstract

Abstract The phosphoinositide-3 kinase (PI3K) pathway is often activated in breast cancer due to frequent mutations in PIK3CA, loss of expression of PTEN or over-expression of receptor tyrosine kinases. This results in constitutive activation of this pathway and in turn resistance to anticancer treatments. One strategy to reverse this chemoresistance was to inhibit PI3K pathway by targeting key regulatory proteins such as PI3K, Akt or mTOR. Many inhibitors have been developed but clinical results obtained have been disappointing so far, probably due to the absence of information on the functional activities of the targeted kinases. We wanted to test how quantitative changes in enzymatic activities of PIK3 pathway would predict a decrease in chemoresistance to conventional therapeutic agents when combined with mTOR inhibitors. A prerequisite was to determine if the genomic mutations of PI3K pathway mentioned above were followed by quantitative changes in the corresponding enzymatic activities targeted by the specific inhibitors. We determined the enzymatic activities of the PI3K pathway in breast cancer cell lines with either mutation in PIK3CA (MCF-7 and T-47D cells), deletion of PTEN (MDA-MB-468 cells) or no alteration in the pathway (MDA-MB-231). Enzymatic activities were evaluated by quantifying the percentage of phosphorylated proteins. As expected, a genomic alteration of a PI3K pathway component resulted in activation of downstream kinases of the pathway (PDK1, mTORC2, Akt and mTORC1) but their level of activation varied depending on the cell line tested. Next, we treated those cell lines with clinically relevant, non-toxic doses (0.1nM or 1nM) of temsirolimus (mTORC1 inhibitor) and carried out a dose-response (from 0.1nM up to 10µM) of doxorubicin (anthracyclin). Temsirolimus induced the phosphorylation of Akt on both Ser473 and Thr308 in MCF-7 cells. This Akt activation remained much lower in MCF-7 than constitutive activation measured in MDA-MB-468 cells. As expected, doxorubicin, used at IC25 (66 nM) also induced the activation of Akt in MCF-7 cells. Addition of temsirolimus did not further increase doxorubicine toxicity. On the opposite, a marked increased resistance of cells to doxorubicin was measured in MCF-7 and in MDA-MB-468 cells treated with temsirolimus. Such induction of chemoresistance was not detected in MDA-MB-231 cells. Similar results were obtained with PI-103, a dual inhibitor of PI3K and mTORC1. Although PI3K pathway inhibitors have been used in clinical settings to restore the effects of conventional anti-cancer drugs, we found that some inhibitors of this pathway increased chemoresistance in breast cancer cells, depending on the altered enzymatic activities in PI3K pathway. Thus, in addition to identifying genomic alterations, kinases activities of the PI3K pathway should be quantified in breast cancer biopsies prior to contemplating the use of targeted drugs. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 831. doi:1538-7445.AM2012-831