Figure 4.

Amprenavir inhibits the expression of HIF-1α and VEGF. (A) U87MG cells were treated with 15 µM amprenavir (AMP) for various lengths of time, as indicated. Then cells were harvested, and Western blot analysis was performed. The membrane was probed for P-Akt (S473) and total Akt, and then was reprobed for β-actin (loading control). (B) U87MG cells were treated with amprenavir for 24 hours; thereafter, cells were harvested for RNA, and Northern blot analysis was performed for VEGF and 18S (loading control). (C) U87MG cells were treated with amprenavir (15 µM) for 24 hours, then cells were exposed to hypoxia (0.2% oxygen). Three hours later, cells were harvested, and Western blot analysis was performed for HIF-1α. Membranes were then reprobed for β-actin (loading control). (D and E) U251MG or NHA cells were treated with nelfinavir (NFV) or amprenavir. Thirty hours later, the cell culture medium was collected, and VEGF protein levels were determined by ELISA and normalized to the number of cells in each dish. *The comparison of ELISA values between these two groups (control and amprenavir-treated U251MG cells) was statistically significant at P < .01 (two-sided Student's t test). The comparison between control and NFV-treated U251MG cells was also statistically significant (P < .01).