Effects of YC on the AMPK signaling pathway.

<p>(A) H1993 cells were treated for 24 h with the indicated concentrations of YC and the protein expressions were measured by Western blot analysis. (B) H1993 cells were treated for different short time points with 1 μM of YC and the protein expressions were measured by Western blot analysis. (C) H1993 cells were pretreated for 1 h with 20 μM of compound C, which is a well-known for AMPK inhibitor or 10 mM of metformin that is an AMPK activator. Then, 1 μM of YC was treated or co-treated for 2 h and the protein expressions were measured by Western blot analysis. (D) H358, H460, Calu-1, H1299, A549 and H1993 cells were treated for 2 h with 1 μM of YC and the protein expressions were measured by Western blot analysis.</p

Anti-proliferative effect of YC in combination with gefitinib or rapamycin in H1993 cells.

<p>The cells were treated with YC alone or in combination with gefitinib or rapamycin for 48 h. The cell proliferation was measured using a SRB assay. (A) YC in combination with gefitinib (B) YC in combination with rapamycin.</p

Inhibition of tumor growth by YC in H1993 xenograft model.

<p>(A) H1993 cells were injected s.c. into the flanks of nude mice and tumors, and were allowed to develop for 21 days. When they reached approximately 90 mm³, the indicated concentrations of YC were initiated. All study medications were given orally once daily for 21 days. Tumor sizes were monitored every 4~6 days. (B) The volume and weight of each tumor xenograft were measured at the termination of the experiment on day 21. (C) Animals in the control and treated groups were weighed every 4~6 days. The mean body weight of each group is presented.</p

Growth-inhibitory effects of YC in H1993 NSCLC cells.

<p>(A) The chemical structure of YC. (B) H1993 cells were treated with the indicated concentrations of YC and gefitinib for 72 h. Cell proliferation was measured by SRB assay. (C) H1993 cells were treated with various concentration of YC for the indicated times, and cell proliferation was determined with the SRB assay. (D) Morphological changes mediated by the treatment of YC for 24 h were observed under the phase-contrast microscope.</p

Effects of YC on the mTOR signaling pathway.

<p>(A) H1993 cells were treated for 24 h with indicated concentrations of YC and the expressions of mTORC1 signaling-related proteins were measured by Western blot analysis. (B) H1993 cells were treated for 24 h with indicated concentrations of YC and the expressions of mTORC2 signaling-related proteins were measured by Western blot analysis. (C) The interaction of mTOR and rictor have been analyzed by immunoprecipitation of rictor with the following immunoblots of the cell lysates (lower panel) and immunoprecipitates (upper panel) with the indicated antibodies. IP: immunoprecipitation. (D) H1993 cells were treated for 24 h with the indicated concentrations of YC and the F-actin expression was measured by immunocytochemistry.</p

Effects of YC on the expression of AMPK and proliferative biomarkers in H1993 xenograft tumors.

<p>The protein extracts were derived from tumor tissues of H1993 xenograft model. (A) The AMPK expressions were measured by Western blot analysis. (B) The p-AMPK expression was measured by immunohistochemistry. (C) The Ki-67 expression was measured by immunohistochemistry. (D) The PCNA expression was measured by immunohistochemistry.</p