Ang1 and Ang2 are endothelial-secreted proteins with a complex relationship and potentially competing overall effects on tumor angiogenesis. Ang2 is most commonly described as a molecule that destabilizes vascular networks, supporting neoangiogenesis [13] and [14]. Ang1 binds to the Tie2 receptor to promote vascular maturation, inhibiting angiogenesis. Ang2 is an antagonist of Ang1 signaling through Tie2. Thus, one of the key questions in the Ang field is whether, in RCC, Ang1 inhibition undermines or augments

effects of Ang2 inhibition. In previous studies, the Ang2-specific inhibitor L1-7, Ang2-CovX bodies, and the Ang2 antibody 3.19.3 slowed the growth of colon and lung cancer xenografts and accentuated the activity of VEGF pathway inhibitors [10], [15] and [16]. The dual Ang1/2 inhibitor, trebananib (AMG386), was found to have more activity

than Ang2-specific inhibitors E7080 cell line alone in colon cancer models [9]. Falcón et al. described similar findings in a colon cancer model and showed that Ang1 inhibition augmented the effect of Ang2 inhibition by preventing vascular normalization seen with the Ang2 inhibitor [13]. RCC is typified by Von Hippel–Lindau (VHL) loss leading to exquisite dependency on the VEGF-driven ERK inhibitor cost angiogenesis. As a consequence, RCC exposure to VEGF pathway inhibitors has been shown to result in “vascular infarction” rather than vascular normalization. Given this distinct biology, we sought to determine the relative effects on tumor growth and perfusion of Ang1, Ang2, and dual Ang1/2 inhibition alone and in combination with VEGF pathway inhibitors in a mouse model of RCC. Another key question related directly to the clinical development of Ang inhibitors is how to select the patients most likely

to benefit from this treatment. Currently, there is little data to guide optimal patient selection and determine the optimal treatment setting. To explore the possibility that Ang2 may be a useful surrogate or predictive marker of activity in RCC, we measured Ang2 plasma levels in patients with RCC either at presentation or during the course of VEGFR-targeted therapy. Taken together, these data inform the continued exploration of Ang2 inhibitors such as trebananib in patients with RCC or other cancers. Frozen tumor specimens for of several human tumor types and non-malignant renal tissues, including non-malignant kidney tissue (cortex and medulla from non-oncology patients), clear cell RCC (ccRCC) tissue, and other non-renal tumor tissue including bladder, lymphoma, lung (adeno), lung (squamous), laryngeal, ovarian, prostate, gastric, breast, colorectal, and pancreatic tumors, were obtained. Total RNA was obtained either directly from a vendor (Ardais Corporation, Lexington, MA) or extracted from frozen tissue samples (Zoion Diagnostics Inc, Shrewsbury, MA) with the Qiagen RNeasy Mini Kit.