Anti-angiogenic activity of rapamycin in rapamycin-sensitive and -resistant tumor xenograft models

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Antitumor activity of rapamycin is variable and tumor type specific. The factors contributing to this differential activity are not known. It is generally believed that rapamycin inhibits tumor growth by a variety of mechanisms including a direct effect on tumor cell proliferation by inhibiting the mTOR/raptor signaling, inhibition of angiogenesis by decreasing the production of VEGF from tumor cells and possibly by blocking AKT/mTOR signaling that is activated in response to VEGF in endothelial cells. To gain further insights into the antitumor/anti-angiogenic activity of rapamycin, we investigated the response in sensitive and resistant tumor xenograft models including glioma (U-87 MG), prostate (PC-3), colon (HCT 116) and non-small cell lung (NCI-H460) cancer. In sensitive xenograft models PC-3 and U-87 MG, Rapamycin showed profound antitumor activity with a T/C ratio of 27% and 1.2%, respectively. In a refractory HCT 116 model, a moderate inhibition of tumor growth was observed with a T/C ratio of 51%. Response of tumor biomarkers was measured using western blots, Meso Scale Discovery platform and immunohistochemistry. There was a robust inhibition of tumor phospho-S6 (pS6) in all of the models studied. However, inhibition of phospho-AKT (pAKT) was observed only in sensitive models. Tumor vasculature was measured using a real-time quantitative high frequency micro-ultrasound in combination with a VEGFR-2 targeted contrast-enhanced agent. This method not only allowed the visualization of VEGFR-2 expression in vivo, but also measured the tumor vasculature in real-time. Chronic treatment of tumor bearing mice with rapamycin significantly inhibited the VEGFR-2 positive tumor vasculature but the magnitude of the inhibition was tumor type dependent. Micro-ultrasound data correlated with CD31-positive blood vessel counts in the tumor sections. Further investigation of tumor sections with immunohistochemial methods revealed that rapamycin significantly inhibited pS6 and pAKT in tumor cells but did not effect the expression of total S6 and AKT proteins. Together these data suggests that chronic treatment of rapamycin exerts profound effect on tumor cells by inhibiting mTOR signaling molecules such as pS6 and pAKT and also on the VEGFR-2 expressing vasculature.