Analysis of baseline CTC enumeration Of the 27 patients enrolled in the

It cannot be excluded that additional changes in other cellular signalling path ways parallel or downstream of the EGFR may be mutated in these models as well. It is relevant to note that while EGF induced cell proliferation in MCF7 EGFR cells was ER independent and tamoxifen insensitive, the majority of E2 induced transcriptional buy ARQ 197 changes in MCF7 EGFR cells remained sensitive to tamoxifen after EGF stimulation. These data clearly indicate that during E2 and EGF co exposure, cell proliferation and E2 induced transcrip tion are controlled by different signalling pathways. The parental MCF7 and MCF7 EGFR cells showed a small increase in MAPK1 3 activation after E2 stimulation which seems consistent with the results of Migliaccio et al.<br><br> AZD0530 379231-04-6 who observed a 2 3 fold MAPK1 3 activation in MCF7 cells several minutes after estradiol exposure by measuring radiolabelled phosphate incorporation in a MAPK substrate. However, the increase MAPK1 3 activa tion by EGF in our MCF7 and MCF7 EGFR cells is much bigger than the activation by estradiol. In tamoxifen resistant breast tumour cells an agonistic effect was observed by tamoxifen both at the level of ER mediated transcription and cell proliferation. It has been suggested that these effects of tamoxifen depend on the phosphorylation of ER by MAPK1 3. However, not all groups find agonistic effects of tamoxifen on transcription and or proliferation after increased MAPK activation and ER serine 118 phosphorylation. Similarly, in our MCF7 EGFR model also no agonistic effects of tamoxifen were observed on proliferation and transcription.<br><br> This is not surprising as the proliferation supplier Alvocidib of MCF7 EGFR cells after EGF stimulation is already high, and any possible additional agonistic effects of tamoxifen may therefore not become manifest. However, it may also be the result of other cell types being used in the previous studies compared to our present cell lines. The lack of an agonistic effect of tamoxifen on transcription after EGFR activation actually suggests that no agonistic effects of tamoxifen are induced in our MCF7 EGFR cells by enhanced EGFR signalling. EGFR activation in MCF7 EGFR cells caused strong downstream activation of both the MAPK and Akt signalling cascades. Using specific inhibitors we demon strated that the MEK MAPK pathway is not dominant in EGFR driven proliferation.<br><br> Recently, using insertion mutagenesis in an estrogen dependent breast carcinoma cell line, a panel of 7 candidate breast cancer anti estrogen resistant genes were identified that directly underlie estrogen independence leading to tamoxifen resistance, including both EGFR, AKT1, and AKT2. Importantly, the mRNA levels of these latter candidates in breast cancer material were significantly correlated with progression or metastasis free survival. These data support our findings about the importance of the PI3K Akt in the EGFR signalling leading to estrogen independent proliferation and tamoxifen insensitivity. The remaining question is which of the downstream targets of AKT are ultimately responsible for EGF induced prolifer ation in MCF7 EGFR cells.