Figure 6. Combination of 1, 9 PA and cetuximab induces apoptosis through downregulation of HIF-1α.

(A) Induction of PARP cleavage by the combination of 1, 9 PA and cetuximab. A431, HN5, and DiFi cells were untreated or treated with cetuximab (10 nM for A431 and HN5 cells and 2 nM for DiFi cells for 16 h), 1, 9 PA (10 µM or 40 µM added the last hour before cell lysis), or both in 0.5% FBS culture medium. Cell lysates were prepared and analyzed by Western blotting with the antibodies shown. (B) Increased induction of apoptosis by the combination of 1, 9 PA and cetuximab. A431 cells were treated as described in (A). Cell lysates were prepared and analyzed by apoptosis ELISA. The relative absorbance values are plotted. The p value was <0.01 when comparing the level of apoptosis by 1, 9 PA alone (10 or 40 µM) or cetuximab alone with that of apoptosis by combination of the 2 agents (note: only the p values comparing 10 µM 1, 9 PA alone and in combination with cetuximab are shown). (C) Dependence of induction of apoptosis by the combination of 1, 9 PA and cetuximab on HIF-1α downregulation. A431neo and A431/HIF-1α-ΔODD cells were treated as indicated, and the cell lysates were prepared and analyzed as described in (A).