Methods section. Drug treatment induced apoptosis in a dose- dependent manner in all ES cell lines, mainly inducing late apoptosis, with few cells undergoing necrosis. Data presented as the average of 4–6 independent replicates. (TIF) Figure S5 Effects of CAV1 or clathrin inhibition on an IGF-1-independent ES cell line. Ewing’s sarcoma TTC466 cells were grown until they reached 30–40% confluence, treated with MCD/CPMZ for 72 hours, and then proliferation was measured with the MTT assay, as described in the Materials and Methods section. A) Characterization of the IGF1 proliferation-driven signaling dependence of the TTC466 cell line, as already published by Martins el al. Cancer Res. 2008; 68(15):6260–70. B) CPMZ or MCD treatment. CAV1 or clathrin inhibition hardly affected ES cell line proliferation. 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PLoS ONE | www.plosone.org 11 May 2011 | Volume 6 | Issue 5 | e19846