Therapeutic doses of Tamoxifen only partially inhibit the non-genomic effects of Estrogen: Evidence for additive anti-proliferative effect of Tamoxifen with MEK inhibitor

Abstract

Aims: Data suggests that non-genomic effects of Estradiol (E2) in estrogen-sensitive carcinoma cell lines result in the activation of the MAP-Kinase (MAPK) pathway (p44/42). MAPK activation is a potent proliferative stimulus and E2 activation of this cascade seems to occur rapidly at the plasma membrane. We investigated the interaction of the SERM Tamoxifen (Tam) and a MAPK inhibitor (MEK inhibitor PD98059) on the non-genomic effects of E2. Methods: MCF-7 cells were maintained in DMEM with 10% FCS. For proliferation experiments, cells were cultured into 96-well plates using seeding densities of 5000 cells /cm3. After incubation, the culture media were replaced with media containing vehicle: E2 (10–9 & 10–8 M) or EGF (100ng/ml). Defined number of wells contained Tam (10–8 M) and/or PD98059 (23µM) in addition to the vehicle. Proliferation was analyzed after 48 & 96 hrs using MTS assay. Western blots were performed on untreated or treated (E2, EGF, Tam or PD98059) cells. Defined number of wells contained Tam and/or PD98059 in addition to treatment. Cell extracts were analyzed with SDS/PAGE, transferred to nitrocellulose filters and probed with Abs specific for p44/p42 MAPKs. Results: data reconfirms the proliferative effects of E2 and EGF. Tam at therapeutic doses weakly reduces the proliferative effects of E2 and partially reduces MAPK activity. PD98059 highly and significantly reduces proliferation rates and MAPK activity. The effects of Tam and MAPK inhibition seem to be additive. Conclusion: experimental data suggests a higher anti-proliferative efficacy in the combination of Tam at therapeutic doses with MAPK inhibitors.