Nelfinavir Down-regulates Hypoxia-Inducible Factor 1α and VEGF Expression and Increases Tumor Oxygenation: Implications for Radiotherapy

Abstract

The phosphatidylinositol 3-kinase (PI3K)/Akt pathway can increase vascular endothelial growth factor (VEGF) and hypoxia-inducible factor 1A (HIF-1A) expression. We examined the effect of nelfinavir, an HIV protease inhibitor that inhibits Akt signaling, on VEGF and HIF-1A expression and on angiogenesis, tumor oxygenation, and radiosensitization. Nelfinavir decreases VEGF expression under normoxia via the transcription factor Sp1, which regulates the proximal core VEGF promoter. Nelfinavir decreased Sp1 phosphorylation and decreased Sp1 binding to a probe corresponding to the proximal VEGF promoter in a gel shift assay. Nelfinavir also decreased the hypoxic induction of HIF-1A, which also regulates the VEGF promoter, most likely by decreasing its translation. The effect of nelfinavir on VEGF expression had the functional consequence of decreasing angiogenesis in an in vivo Matrigel plug assay. To determine the effect this might have on tumor radiosensitization, we did tumor regrowth assays with xenografts in nude mice. The combination of nelfinavir and radiation increased time to regrowth compared with radiation alone whereas nelfinavir alone had little effect on tumor regrowth. This radiosensitizing effect was greater than suggested by in vitro clonogenic survival assays. One possible explanation for the discordance is that nelfinavir has an effect on tumor oxygenation. Therefore, we examined this with the hypoxia marker EF5 and found that nelfinavir leads to increased oxygenation within tumor xenografts. Our results suggest that nelfinavir decreases HIF-1A/VEGF expression and tumor hypoxia, which could play a role in its in vivo radiosensitizing effect. These data support the use of nelfinavir in combination with radiation in future clinical trials.

Vascular Normalization as a Therapeutic Strategy for Malignant and Nonmalignant Disease

“…As discussed below, many studies testing the effects of antiangiogenic drugs have shown a presence of a "normalization window"-a time period beginning with the appearance of a normalized vascular phenotype (typically within 1 -2 days of starting treatment), and ending when features of normalization are lost Winkler et al 2004;Jain 2005b;Batchelor et al 2007;Kamoun et al 2009 Xian et al 2006;McCarty et al 2007;Mazzone et al 2009. c This is controversial given recent reports of increased metastasis after antiangiogenic therapy in certain animal models (Ebos et al 2009;Paez-Ribes et al 2009 Gorski et al 1999;Hansen-Algenstaedt et al 2000;Lee et al 2000;Kozin et al 2001;Ader et al 2003;Winkler et al 2004;Ansiaux et al 2005Ansiaux et al , 2006Pore et al 2006a;Dings et al 2007;Kashiwagi et al 2008;Batra et al 2009;Cerniglia et al 2009;Tsukada et al 2009;McGee et al 2010. g Teicher et al 1995a,b;Wildiers et al 2003;Segers et al 2006;Dickson et al 2007a,b,c;Bhattacharya et al 2008Bhattacharya et al , 2009Zhou et al 2008;Juan et al 2009;Zhou and Gallo 2009;J Liu, S Liao, B Diop-Frimpong, et al, unpubl. data. section, we discuss the key molecules implicated in tumor vessel biology with a focus on their role in regulating vascular normalization.…”

“…As a consequence, inhibitors of oncogene activity can indirectly normalize tumor vessels, potentially enhancing not only the action of chemotherapy and radiotherapy, but also their own antitumor cell activity. Investigators have reported various aspects of normalization in response to inhibition of several oncogenes including HER-2 (Izumi et al 2002), the PI3K-AKT-mTOR axis (Pore et al 2006a;Schnell et al 2008;Xue et al 2008;Qayum et al 2009), Ras (Cohen-Jonathan et al 2001Delmas et al 2003;Qayum et al 2009), and the epidermal growth factor receptor (EGFR) (Cerniglia et al 2009;Qayum et al 2009;Johns et al 2010). In the case of HER-2, we have shown that treatment of leptomeningeal HER-2 positive breast cancer with trastuzumab (a monoclonal antibody directed against HER-2) not only inhibits tumor cell proliferation, but also normalizes vessels.…”

Ritonavir inhibits HIF-1α-mediated VEGF expression in retinal pigment epithelial cells in vitro

“…Our results corroborated with Pore et al, who demonstrated that nelfinavir, another HIV protease inhibitor, interfered with HIF-1a protein translation in carcinoma cells. 38 An active PI3K/Akt pathway is required for hypoxia-induced expression of HIF-1a protein translation. 39,40 Hence, a preliminary study was performed to analyze the levels of Akt in RPE cells post treatment with ritonavir.…”

Z-360, a Novel Cholecystokinin-2/Gastrin Receptor Antagonist, Inhibits Gemcitabine-Induced Expression of the Vascular Endothelial Growth Factor Gene in Human Pancreatic Cancer Cells

“…27) HIF-alpha activation is mediated by its stabilization via the Akt/PI3kinase signaling pathway. 28) Akt/PI3kinase is a major kinase involved in the down-stream signaling pathway of the CCK-2/gastrin receptor. 29,30) Thus, gastrin is a likely mediator of HIF-alpha activation or VEGF expression.…”