Recent advances toward understanding the molecular mechanisms regulating cancer initiation and progression provide new insights into the therapeutic value of targeting tumor vascularity by interfering with angiogenic signaling pathways. The functional contribution of key angiogenic factors toward increased vascularity characterizing metastatic tumors and their therapeutic exploitation is considered in three major urologic malignancies, renal, bladder, and prostate cancer. With the realization that the success of the therapeutic efficacy of the various anti-angiogenic approaches for the treatment of urologic tumors has yet to be proven clinically, the challenge remains to select critical angiogenesis pathways that can be targeted for an individual tumor. Here we discuss the major mechanisms that support formation of vasculature in renal, bladder, and prostate tumors and the current results of targeting of specific molecules/regulators for therapeutic intervention against metastastic disease. Keywordsvascularity; tumor growth; apoptosis; VEGF; bladder cancer; renal cancer; prostate cancer In 2007, there will be an estimated 346,440 new cases diagnosed with urologic cancer in the United States and 54,360 Americans will die from a urologic malignancy (SEER Cancer Statistics Review, http://cancernet.nci.nih.gov/statistics). This mortality rate is alarmingly high as it translates to one individual dying every 9 min in the US due to a urologic tumor and thus a significant health issue.Angiogenesis is an essential process in normal physiological functions such as ovarian cycle in female reproductive system [Kaczmarek et al., 2005] and a contributing factor in disease states such as chronic inflammation, arthritis, cancer, and macular degeneration . During the development of the embryo, mesoderm differentiates into angioblasts; these endothelial cells, not yet organized into a lumen, form primitive vessels toward development of blood vessel network, via vasculogenesis. In the adult, new blood vessels form from preexisting vasculature, via angiogenesis [Risau, 1997], while malignant conditions induce a hypercoagulable state in their hosts [Nash et al., 2001]. By early 1960s it was evident that tumors could elaborate diffusible substances that induce angiogenesis from the host vasculature [Algira et al., 1945;Greenblatt and Shubick, 1968]. The increased tumor vascularity was originally believed to be vasodilation of the host endothelium in response to metabolic waste products from within the tumor . A decade later Dr. Folkman's pioneering work identified angiogenesis as a required phenomenon for tumor growth and metastasis, first defining the potential therapeutic value of agents targeting this process , 1971]. Tumor blood vessels exhibit characteristic markers which are not present in normal angiogenic tissues . After enduring the circulation "journey," metastatic cancer cells can escape out of the endothelial vasculature and in the target tissue via extravasation. How do the metastastic cells signal activa...

Purpose: The effect of ZD6126 on tumor oxygen tension and tumor growth delay in combination with ionizing radiation was examined in the human U87 glioblastoma tumor model. Resistance to ZD6126 treatment was investigated with the nitric oxide synthase inhibitor, l-NG-nitroarginine methyl ester (hydrochloride; l-NAME/active form, l-NNA). Methods: U87 human xenografts were grown in athymic nude mice. ZD6126 was given with or without l-NNA. Tumor oxygen tension was measured using the Oxford Oxylite (Oxford, England) fiberoptic probe system. Tumor volume was determined by direct measurement with calipers and calculated by the formula [(smallest diameter2 × widest diameter)/2]. Results: Multiple doses of ZD6126 treatment (three doses) had a significant effect on tumor growth delay, reducing the average daily tumor growth rate from 29% to 16%. When given 1 hour before radiation, ZD6126 caused an acute increase in hypoxia in U87 tumors, and reduced tumor growth delay compared with that of radiation alone. The combination of ZD6126 given after radiation, either as a single dose or in multiple doses, had greater or similar antitumor activity compared with radiation alone. Twenty-four hours after administration, a single dose of ZD6126 induced little (10 ± 8%) necrosis in U87 xenografts. l-NNA, when given in combination with ZD6126, significantly enhanced the effectiveness of ZD6126 in inducing tumor necrosis. Conclusions: Our observation that ZD6126-induced tumor hypoxia can decrease radiation response when ZD6126 is given prior to radiation indicates the importance of scheduling. Our findings suggest that the optimal therapeutic benefit of ZD6126 plus radiation in human glioblastoma may require multiple dosing in combination with a nitric oxide synthase inhibitor, to be scheduled following radiotherapy.

Emergence of therapeutic resistance to angiogenesis inhibitors, the mechanisms of which are poorly understood, remains a major obstacle in treatment of NSCLC patients. Previously we reported that mechanisms governing resistance to anti-angiogenic therapy may involve both tumor and stromal cells in the tumor microenvironment. In this study we investigated potential mechanisms of resistance to the multi-tyrosine kinase inhibitors cediranib (AZD2171, Recentin®) and vandetanib (ZD6474, Zactima®) using NSCLC xenografts treated either for 2 weeks (sensitive tumors) or until resistance occurred. Quantification of TUNEL+ staining using laser scanning cytometry (LSC) showed increased apoptosis in H1975 xenografts sensitive to cediranib (p<0.01) and vandetanib (p<0.05) when compared with controls, whereas no changes were noticed at time of resistance. Microvessel density (MVD) was significantly increased in resistant H1975 xenografts compared with controls (p<0.05) and sensitive tumors (p<0.01), whereas in A549 model, vandetanib-resistance was associated with an angiogenic independent phenotype. To investigate stromal mechanisms of Vascular Endothelial Growth Factor Receptor (VEGFR) TKI resistance, we characterized the stromal angiogenic gene expression profiles of H1975 sensitive and resistant tumors using a mouse-specific gene expression array (mouseWG-6 v2 Expression BeadChip, Illumina®). Differentially modulated genes were selected based on a p<0.005 of the univariate t-test and at least a 1.5 fold-change in expression and cross-referenced to defined list(s) of angiogenesis-related genes. Stromal Hgf (hepatocyte growth factor) was up-regulated in VEGFR TKI-resistant xenografts compared to sensitive tumors. Hgf up-regulation was confirmed at the protein level using immunofluorescent staining and confocal microscopy. HGF protein levels were strongly decreased after 2 weeks of treatment with cediranib and vandetanib (p<0.01), whereas a significant increase in HGF was observed in resistant xenografts (p<0.01). To assess whether HGF upregulation contributes to tumor resistance to TKIs, we implanted HGF-overexpressing and vector control HCC827 NSCLC cells into nude mice. In HCC827-vector control xenografts, cediranib inhibited tumor growth by 93%, whereas a 60% of growth inhibition was observed in HGF-overexpressing tumors. These data agree favorably with our previous analysis of clinical specimens from patients with stage IIIB/IV NSCLC that identified HGF as a predictive marker of resistance to vandetanib treatment alone when compared to chemotherapy or the combination of chemotherapy and vandetanib (p=0.033). Our results suggest that HGF up-regulation may resistance to VEGFR pathway inhibition and that the HGF/MET axis may represent a crucial target for NSCLCs that are resistant to anti-angiogenic therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 376.

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