932. Combined Treatment with Tarceva, an EGFR Inhibitor with Ad-mda7 Synergizes To Promote Anti-Tumor Activity Against Human Cancer Cells
Authors: Rajagopal Ramesh, Satoshi Inoue, Sunil Chada
Journal: Molecular Therapy (2006)

The epidermal growth factor receptor (EGFR) is highly expressed in many human cancers and plays an important role in EGF-mediated cell signaling. The binding of epidermal growth factor (EGF) to EGFR results in molecular signaling that favors cell survival. Preclinical studies have shown targeting of the extraceullar domain of EGFR using antibodies such as Cetuximab or the intracellular kinase domain using small molecule inhibitors such as Tarceva results in tumor growth inhibition. Based on these studies several clinical trials have been conducted testing the antitumor activity of these drugs against human cancers. However, in majority of these studies treatment with drug alone has shown limited clinical response. These studies indicate that additional treatment modalities may need to be incorporated to achieve an effective therapeutic response. We have previously shown the melanoma differentiation associated gene-7 (MDA-7) protein exerts a potent antitumor activity in vitro and in vivo. Mda-7 gene transfer via Ad-mda7 inhibits tumor growth and induces apoptosis. In the present study we evaluated the therapeutic effects of Tarceva in combination with Ad-mda7 using human lung and prostate cancers as a model. Treatment of lung (H1299) and prostate (LNCaP) tumor cells with Ad-mda7 (1000–2000 vp/cell) or Tarceva (2.5 or 5 uM) showed inhibition of tumor cell growth in vitro. However, treatment of tumor cells with Ad-mda7 plus Tarceva resulted in enhanced growth (P<0.05) inhibition as determined by cell viability assay compared to cells treated with Ad-luc plus Tarceva. Furthermore, combination treatment with Ad-mda7 plus Tarceva induced an additive to synergisitic growth inhibition in both H1299 and LNCaP tumor cells compared to Ad-luc plus Tarceva treatment. That the growth inhibition observed in Ad-mda7 plus Tarceva treatment wasdue to increased Ad-mda7 transduction was eliminated by conducting transduction efficiency studies using Ad-GFP. No significant increase in the number of GFP-positive cells was observed when treated with Ad-GFP plus Tarceva compared to cells treated with Ad-GFP alone. Molecular analysis revealed inhibition of phosphorylation of EGFR in tumor cells treated with Tarceva, Ad-luc plus Tarceva and Ad-mda7 plus Tarceva. However, the inhibition of EGFR phosphorylation was greater in Ad-mda7 plus Tarceva-treated cells. These results show that Tarceva and Ad-mda7 act in concert to inhibit EGFR activity. Ad-mda7-mediated inhibition of EGFR phosphorylation occurs by inhibiting Src kinase activity and the Src-specific phosphorylation site in kinase domain of EGFR. Correlating with enhanced growth inhibition was the increased detection of caspase-3 activation in Ad-mda7 pus Tarceva-treated cells compared to Ad-mda7- or Ad-luc plus Tarceva-treated cells. No significant change in Akt or p44/42 MAPK, targets for Tarceva was observed in any of the treatment groups indicating alternate molecular signaling pathways being inhibited. In conclusion, Ad- mda7 plus Tarceva treatment significantly increases the antitumor activity against human lung and prostate cancer cells in vitro.