ACS DIVISION OF BIOLOGICAL CHEMISTRY

March 5th, 2014 by Bill, A., Hall, M. L., Borawski, J., Hodgson, C., Jenkins, J., Piechon, P., Popa, O., Rothwell, C., Tranter, P., Tria, S., Wagner, T., Whitehead, L., Gaither, L. A.

ANO1, a calcium-activated chloride channel, is highly expressed and amplified in human cancers and is a critical survival factor in these cancers. The ANO1 inhibitor CaCCinh-A01 decreases proliferation of ANO1-amplified cell lines, however, the mechanism of action remains elusive. We explored the mechanism behind CaCCinh-A01′s inhibitory effect on cell proliferation using a combined experimental and in silico approach. We show that inhibition of ANO1 function is not sufficient to diminish proliferation of ANO1-dependent cancer cells. We report that CaCCinh-A01 reduces ANO1 protein levels by facilitating endoplasmic reticulum-associated, proteasomal turnover of ANO1. Washout of CaCCinh-A01 rescued ANO1 protein levels and resumed cell proliferation. Proliferation of newly derived CaCCinh-A01 resistant cell pools was not affected by CaCCinh-A01 as compared to the parental cells. Consistently, CaCCinh-A01 failed to reduce ANO1 protein levels in these cells while ANO1-currents were still inhibited by CaCCinh-A01, indicating that CaCCinh-A01 inhibits cell proliferation by reducing ANO1 protein levels. Furthermore, we employed in silico methods to elucidate novel biological functions of ANO1 inhibitors. Specifically, we derived a pharmacophore model to describe inhibitors capable of promoting ANO1 degradation and report new inhibitors of ANO1-dependent cell proliferation. In summary our data demonstrate that inhibition of ANO1′s channel activity is not sufficient to inhibit ANO1-dependent cell proliferation indicating that ANO1′s role in cancer only partially depends on its function as a channel. Our results provide impetus for gaining a deeper understanding of ANO1 modulation in cells and introduce a new targeting approach for antitumor therapy in ANO1-amplified cancers.