Triple-negative breast cancers (TNBC) represent the 10-17% of all diagnosed breast cancers (BC) and are characterized by the absence of ER/PgR expression, HER2 amplification and often show a basal-like phenotype. TNBC are often diagnosed in patients with BRCA1 germline mutation and unfortunately treatment options are still limited. The mTOR (Mammalian Target Of Rapamycin) pathway seems to play an important role in BC pathogenesis and it is possible to target this pathway by inhibitors such as rapamycin. In human BC cross talk between ER/PgR receptors signaling and the mTOR pathway is believed to be responsible for resistance to hormone therapy probably due to a down regulation of hormone receptors. Based on these evidences we have hypothesized that the inhibitors of mTOR pathway may lead to the up-regulation of ER, PgR and HER2 in TNBC cell lines.
For this study we used TNBC cells (MDA-MB-231 and BT20) cultured in DMEM:F12 (Dulbecco's Modified Eagle's Medium) with 10% bovine serum (FBS), 100 U/mL penicillin (1%) and 50 mg/mL of streptomycin under standard conditions (37°C in an atmosphere composed of 16% O2, 79% N2 and 5% CO2).
Cells were initially treated with Rapamycin (1, 5, 10, 15 microM) for 24, 48, and 72h in order to verify if the drug determines a blockade of cell proliferation. Before drug administration, the cells were subjected to serum starvation by eliminating serum from culture medium. The evaluation of cell viability following the administration of the drug was carried out using the MTT assay. Real time PCR analyses were carried out in order to evaluate gene expression modifications of ER, PgR and HER2 receptors, through Taqman probe chemistry.
The preliminary cell viability experiments conducted on different TNBC cell lines (MDA-MB-231 and BT20), showed no significant cytotoxic effects by increasing Rapamycin concentrations (1, 5 and 10 microM) after 72h treatment, except for the higher concentration (15 microM) for which a cytotoxic effect was observed.
The following qPCR approach highlighted significant variations in estrogen and progesterone receptor gene expression for the TNBC cell lines after 24, 48, 72h with 1, 5, 10 microM Rapamycin. In particular their expression level resulted up-regulated. Unlikely no association between mTOR inhibition and HER2 expression level were identified, suggesting no effects of mTOR inhibition on HER2 expression.
CONCLUSION AND FUTURE PERSPECTIVES:
To these preliminary results suggest that the mTOR inhibition leads to re-expression of hormone receptors. This finding supports a potential clinical application of mTOR inhibition in TNBC. The perspective of phenotype change upon rapamycin treatment prompts new therapeutic scenarios. However, further investigations are needed to explain the biological mechanisms driving these changes.
Citation Format: Daniela Massihnia, Giuseppe Bronte, Marta Castiglia, Nadia Barraco, Antonina Cangemi, Alessandro Perez, Daniele Fanale, Gianni Pantuso, Salvatore Vieni, Valentina Calò, Christian D. Rolfo, Viviana Bazan, Antonio Russo. Role of mTOR inhibition in triple-negative breast cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1855.