Figure 2

Metastatic resistance to mTOR inhibition. (a) Left panels, graphs showing the average and standard deviation of micro- and macro-metastases observed in the lungs of the DMSO- and sirolimus- or everolimus-treated orthoxenografts and 4T1 tumors, respectively. The results correspond to the last day of treatment, and micro- versus macro-metastases were defined using a 2 mm width threshold, and by examining at least three tissue levels separated by > 20 μM. Right panels, growth rates of the DMSO- and sirolimus- or everolimus-treated tumors. (b) Representative immunohistochemical results for pS6 at the invasive tumor fronts (magnifications; top right panels) and the lung metastases (right panels) of DMSO- or sirolimus/everolimus-treated mice. The middle panels show quantifications, which correspond to three tumors, three equal front areas, and three metastases in each case. (c) Representative immunohistochemical tumor results for FSCN1/Fscn1 in DMSO- or sirolimus/everolimus-treated mice; quantifications are shown in right panels. (d) Recovered pS6 signal with concurrent FSCN1 overexpression through adaptation to everolimus in MCF7 and HCC1937 cells. Days of treatment are shown. (e) Left panel, graph showing the quantification of colonies from untreated and everolimus-adapted cells (12 culture fields were analyzed). The one-tailed t-test P-values are shown. Representative images of cell cultures are shown in right panels. (f) Flow cytometry results showing the cell counts for CD49f/EpCAM and of CD44/CD24 positivity in untreated or everolimus-adapted MCF7 and HCC1937 cells, respectively.