Abstract
Abstract Background. In RCC temsirolimus has been approved for first line treatment in advanced disease whereas everolimus has been approved after failure of sunitinib or sorafenib treatment. The aim of our study was to compare everolimus and temsirolimus activity in renal and hepatic cancer cell lines characterized for primary or acquired resistance to sunitinib and sorafenib. Materials and Methods: Parental SK-HEP1 hepatocellular, CAKI1 and 786-0 renal carcinoma cells were exposed to stepwise increasing doses of sunitinib and sorafenib for >6 months until acquisition of resistance. Antiproliferative effects of rapalogues were assessed using MTT assay. mRNA levels of selected genes were evaluated in sensitive and resistant cell lines using q-RT-PCR. Protein phoshorylation was assessed by western-blot. Results. Our panel of HCC and RCC cell lines was characterized for the expression of AKT, Bcl2, PTEN, Cyclin D1 and p27 as well as for the mutation state of KRAS and BRAF that are known to be possible predicting factors of sensitivity to mTOR inhibitors. The HCC cell lines SK-HEP1 and its sunitinib- and sorafenib-resistant counterparts, SK-Suni and SK-Sora, carrying a mutated BRAF gene, displayed high BCL2, PTEN and CCND1 gene expression. Constitutive activation of AKT was found in all sunitinib and sorafenib-resistant cell lines. Rapamycin, everolimus and temsirolimus at concentrations that ranged from 1 to 10µM displayed only low antiproliferative effects (15-20% growth inhibition). In contrast, at 20µM concentration the three rapalogues displayed cytotoxic effects in all cell lines. At this concentration everolimus was more cytotoxic than temsirolimus with a growth inhibition of 80-85% and 55-60%, respectively; both drugs were more active than equimolar rapamycin. Interestingly, SK-Sora and CAKI-Suni cells were more sensitive to the different rapalogues than their parental counterparts. In 786-0, 786-Suni and 786-Sora cells, 72h exposure to rapalogues induced cell cycle modifications with an increase in the G1-phase and a decrease in the S-phase. Exposure of these cells to 1µM rapalogues for 5 and 24h resulted in strong inhibition of phosphorylation of the S6 and P70S6K proteins, two well-established mTORC1 downstream targets. Exposure to 1 µM everolimus also decreased 4EBP-1 phosphorylation (p4EBP-1) whereas temsirolimus and rapamycin effects on p4EBP-1 were less potent. In response to 5h rapalogues exposure, phospho AKT (Ser473) was transiently activated. After 24 hours, AKT activation was inhibited in presence of everolimus and temsirolimus but not rapamycin. Conclusion. Everolimus displayed potent antiproliferative effects that were equal or superior to those observed with temsirolimus and rapamycin in HCC and RCC cancer cell lines with acquired resistance to sunitinib and sorafenib. 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 1805. doi:1538-7445.AM2012-1805
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