Fabien Gautier

It is still unclear whether the BH3-only protein Puma (p53 up-regulated modulator of apoptosis) can prime cells to death and render antiapoptotic BH3-binding Bcl-2 homologues necessary for survival through its ability to directly interact with proapoptotic Bax and activate it. In this study, we provide further evidence, using cell-free assays, that the BH3 domain of Puma binds Bax at an activation site that comprises the first helix of Bax. We also show that, in yeast, Puma interacts with Bax and triggers its killing activity when Bcl-2 homologues are absent but not when Bcl-xL is expressed. Finally, endogenous Puma is involved in the apoptotic response of human colorectal cancer cells to the Bcl-2/Bcl-xL inhibitor ABT-737, even in conditions where the expression of Mcl-1 is down-regulated. Thus, Puma is competent to trigger Bax activity by itself, thereby promoting cellular dependence on prosurvival Bcl-2 family members.

A functional imbalance between proapoptotic Bax and antiapoptotic Bcl-2 is likely to participate in the resistance of cancer cells to therapy. We show here that ethyl 2-amino-6-bromo-4-(1-cyano-2-ethoxy-2-oxoethyl)-4H-chromene-3-carboxylate (HA14-1), a small organic compound recently proposed to function as an inhibitor of Bcl-2, increases the sensitivity of human glioblastoma cells to radiotherapy and chemotherapy. This sensitizing effect is lost if Bcl-2 expression, but not Bcl-xL expression, is knocked down or if cells only express a mutant of Bax that does not interact with Bcl-2. This points to a specific Bcl-2 inhibitory function of HA14-1 and implies that it selectively involves hindrance of Bcl-2 binding to Bax, which HA14-1 inhibits in cell-free assays and in cells in receipt of an apoptotic stimulation. Moreover, HA14-1, in combination with a cytotoxic treatment, slows down the growth of glioblastoma in vivo. Thus, the inhibition of Bcl-2 achieved by HA14-1 might improve treatment outcome.