PlGF blockade does not inhibit angiogenesis during primary tumor growth.

It has been recently reported that treatment with an anti-placenta growth factor (PlGF) antibody inhibits metastasis and primary tumor growth. Here we show that, although anti-PlGF treatment inhibited wound healing, extravasation of B16F10 cells, and growth of a tumor engineered to overexpress the PlGF receptor (VEGFR-1), neutralization of PlGF using four novel blocking antibodies had no significant effect on tumor angiogenesis in 15 models. Also, genetic ablation of the tyrosine kinase domain of VEGFR-1 in the host did not result in growth inhibition of the anti-VEGF-A sensitive or resistant tumors tested. Furthermore, combination of anti-PlGF with anti-VEGF-A antibodies did not result in greater antitumor efficacy than anti-VEGF-A monotherapy. In conclusion, our data argue against an important role of PlGF during primary tumor growth in most models and suggest that clinical evaluation of anti-PlGF antibodies may be challenging.

Anti-VEGF antibody therapy does not promote metastasis in genetically engineered mouse tumour models.

Resistance to anti-angiogenic therapy can occur via several potential mechanisms. Unexpectedly, recent studies showed that short-term inhibition of either VEGF or VEGFR enhanced tumour invasiveness and metastatic spread in preclinical models. In an effort to evaluate the translational relevance of these findings, we examined the consequences of long-term anti-VEGF monoclonal antibody therapy in several well-validated genetically engineered mouse tumour models of either neuroendocrine or epithelial origin. Anti-VEGF therapy decreased tumour burden and increased overall survival, either as a single agent or in combination with chemotherapy, in all four models examined. Importantly, neither short- nor long-term exposure to anti-VEGF therapy altered the incidence of metastasis in any of these autochthonous models, consistent with retrospective analyses of clinical trials. In contrast, we observed that sunitinib treatment recapitulated previously reported effects on tumour invasiveness and metastasis in a pancreatic neuroendocrine tumour (PNET) model. Consistent with these results, sunitinib treatment resulted in an up-regulation of the hypoxia marker GLUT1 in PNETs, whereas anti-VEGF did not. These results indicate that anti-VEGF mediates anti-tumour effects and therapeutic benefits without a paradoxical increase in metastasis. Moreover, these data underscore the concept that drugs targeting VEGF ligands and receptors may affect tumour metastasis in a context-dependent manner and are mechanistically distinct from one another.

Multimodal microvascular imaging reveals that selective inhibition of class I PI3K is sufficient to induce an antivascular response.

The phosphatidylinositol 3-kinase (PI3K) pathway is a central mediator of vascular endothelial growth factor (VEGF)-driven angiogenesis. The discovery of small molecule inhibitors that selectively target PI3K or PI3K and mammalian target of rapamycin (mTOR) provides an opportunity to pharmacologically determine the contribution of these key signaling nodes in VEGF-A-driven tumor angiogenesis in vivo. This study used an array of micro-vascular imaging techniques to monitor the antivascular effects of selective class I PI3K, mTOR, or dual PI3K/mTOR inhibitors in colorectal and prostate cancer xenograft models. Micro-computed tomography (micro-CT) angiography, dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI), vessel size index (VSI) MRI, and DCE ultrasound (DCE-U/S) were employed to quantitatively evaluate the vascular (structural and physiological) response to these inhibitors. GDC-0980, a dual PI3K/mTOR inhibitor, was found to reduce micro-CT angiography vascular density, while VSI MRI demonstrated a significant reduction in vessel density and an increase in mean vessel size, consistent with a loss of small functional vessels and a substantial antivascular response. DCE-MRI showed that GDC-0980 produces a strong functional response by decreasing the vascular permeability/perfusion-related parameter, K (trans). Interestingly, comparable antivascular effects were observed for both GDC-980 and GNE-490 (a selective class I PI3K inhibitor). In addition, mTOR-selective inhibitors did not affect vascular density, suggesting that PI3K inhibition is sufficient to generate structural changes, characteristic of a robust antivascular response. This study supports the use of noninvasive microvascular imaging techniques (DCE-MRI, VSI MRI, DCE-U/S) as pharmacodynamic assays to quantitatively measure the activity of PI3K and dual PI3K/mTOR inhibitors in vivo.

Targeting the PI3K pathway in the brain--efficacy of a PI3K inhibitor optimized to cross the blood-brain barrier.

PURPOSE: Glioblastoma (GBM), the most common primary brain tumor in adults, presents a high frequency of alteration in the PI3K pathway. Our objectives were to identify a dual PI3K/mTOR inhibitor optimized to cross the blood-brain barrier (BBB) and characterize its brain penetration, pathway modulation in the brain and efficacy in orthotopic xenograft models of GBM. EXPERIMENTAL DESIGN: Physicochemical properties of PI3K inhibitors were optimized using in silico tools, leading to the identification of GNE-317. This compound was tested in cells overexpressing P-glycoprotein (P-gp) or breast cancer resistance protein (BCRP). Following administration to mice, GNE-317 plasma and brain concentrations were determined, and phosphorylated biomarkers (pAkt, p4EBP1, and pS6) were measured to assess PI3K pathway suppression in the brain. GNE-317 efficacy was evaluated in the U87, GS2, and GBM10 orthotopic models of GBM. RESULTS: GNE-317 was identified as having physicochemical properties predictive of low efflux by P-gp and BCRP. Studies in transfected MDCK cells showed that GNE-317 was not a substrate of either transporter. GNE-317 markedly inhibited the PI3K pathway in mouse brain, causing 40% to 90% suppression of the pAkt and pS6 signals up to 6-hour postdose. GNE-317 was efficacious in the U87, GS2, and GBM10 orthotopic models, achieving tumor growth inhibition of 90% and 50%, and survival benefit, respectively. CONCLUSIONS: These results indicated that specific optimization of PI3K inhibitors to cross the BBB led to potent suppression of the PI3K pathway in healthy brain. The efficacy of GNE-317 in 3 intracranial models of GBM suggested that this compound could be effective in the treatment of GBM.

Quantifying antivascular effects of monoclonal antibodies to vascular endothelial growth factor: insights from imaging.

PURPOSE: Little is known concerning the onset, duration, and magnitude of direct therapeutic effects of anti-vascular endothelial growth factor (VEGF) therapies. Such knowledge would help guide the rational development of targeted therapeutics from bench to bedside and optimize use of imaging technologies that quantify tumor function in early-phase clinical trials. EXPERIMENTAL DESIGN: Preclinical studies were done using ex vivo microcomputed tomography and in vivo ultrasound imaging to characterize tumor vasculature in a human HM-7 colorectal xenograft model treated with the anti-VEGF antibody G6-31. Clinical evaluation was by quantitative magnetic resonance imaging in 10 patients with metastatic colorectal cancer treated with bevacizumab. RESULTS: Microcomputed tomography experiments showed reduction in perfused vessels within 24 to 48 h of G6-31 drug administration (P

Blocking neuropilin-2 function inhibits tumor cell metastasis.

Metastasis, which commonly uses lymphatics, accounts for much of the mortality associated with cancer. The vascular endothelial growth factor (VEGF)-C coreceptor, neuropilin-2 (Nrp2), modulates but is not necessary for developmental lymphangiogenesis, and its significance for metastasis is unknown. An antibody to Nrp2 that blocks VEGFC binding disrupts VEGFC-induced lymphatic endothelial cell migration, but not proliferation, in part independently of VEGF receptor activation. It does not affect established lymphatics in normal adult mice but reduces tumoral lymphangiogenesis and, importantly, functional lymphatics associated with tumors. It also reduces metastasis to sentinel lymph nodes and distant organs, apparently by delaying the departure of tumor cells from the primary tumor. Our results demonstrate that Nrp2, which was originally identified as an axon-guidance receptor, is an attractive target for modulating metastasis.