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. Author manuscript; available in PMC: 2014 Jul 1.
Published in final edited form as: Cancer Discov. 2013 Apr 25;3(7):761–769. doi: 10.1158/2159-8290.CD-13-0103

Figure 3.

Figure 3

PIK3CA mutation enhances sensitivity to PI3K pathway inhibition. (A) HNSCC cells containing endogenous PIK3CA mutation (H1047R) (CAL-33; Detroit 562) and cells containing WT PIK3CA [SCC-9, PE/CA-PJ34 (clone C12)] were treated with a PI3K/mTOR inhibitor, BEZ-235, followed by growth determinations at 48 hrs (n=4). Experiments were repeated 3 times with similar results. (B) BEZ-235 inhibited growth of CAL-33 xenografts [with endogenous PIK3CA(H1047R) mutation]. CAL-33 cells (0.5 ×106 cells) were inoculated into the flanks of nude mice. Treatment was started when the tumors became palpable 8 days after tumor cell inoculation. BEZ-235 (25mg/kg/day, n=9) or vehicle (n=10) was given by oral gavage. (C) Sanger sequencing results showing PIK3CA(E542K) mutation in the HPV-HNSCC patient tumor that were implanted the flanks of NOD SCIDγ mice. (D) PIK3CA-mutated tumorgrafts are sensitive to BEZ-235 treatment. HNSCC patient tumorgrafts were implanted into the flanks of NOD SCIDγ mice and treatment was started when tumors became palpable. BEZ-235 (25mg/kg) or vehicle control was given daily by oral gavage. Mice were given vehicle (n=7) or BEZ-235 (n=5) when tumors became palpable. Treatment with BEZ-235 significantly reduced the tumor size when compared to vehicle control (P<0.0001). (E) Western blots showing the effects of BEZ-235 (vs. vehicle control) on expression of PI3K signaling components in the PIK3CA-mutated tumorgrafts. Tumors were harvested for Western blotting at the end of the experiment on day 22. Densitometry values of band intensity are shown below each band (normalized to total beta-tubulin level). Phospho-AKT (S473) and phospho-S6 (S235/236) levels were significantly reduced upon BEZ-235 treatment when compared to the vehicle-treated tumors (P=0.0124 and P<0.0001, respectively). (F) HNSCC patient tumorgrafts from a WT PIK3CA tumor expressing low levels of pAKT were implanted into the flanks of NOD SCIDγ mice and treatment was started when tumors became palpable. BEZ-235 (25mg/kg) (n=6) or vehicle control (n=6) was given daily by oral gavage. Treatment with BEZ-235 failed to significantly reduce the tumor size when compared to vehicle control (P=0.300).