Kinetics of vascular normalisation by VEGFR2 blockade governs brain tumour response to radiation: role of angiopoietin-1 and matrix metalloproteinases

Background: A recent successful Phase III clinical trial using a VEGF-specific antibody suggests that antiangiogenic therapy must be combined with cytotoxic therapy for the treatment of solid tumors. However, there are no guidelines for optimal scheduling of these therapies. Specifically, Glioblastoma Multiforme is characterized by a dilated, tortuous, disorganized, and leaky vasculature and by high levels of VEGF. These abnormalities may impair the delivery of oxygen and therapeutics.

Methods: Using human glioma xenografts implanted under cranial windows in nude mice, we investigated the mechanism and kinetics of vascular changes induced by the anti-VEGFR2 antibody DC101, and their relation to tumor oxygenation and radiation response. We combined DC101 (given i.p. on day 0,3,6) with local gamma radiation (three consecutive days) according to 5 different treatment schedules. We evaluated tumor oxygenation (by pimonidazole staining), tumor vascular morphology (by in vivo multiphoton microscopy), vascular pericyte coverage and basement membrane morphology (by immunohistochemistry), and the role of Angiopoietin-1 upregulation (by cDNA microarray analysis, quantitative PCR, immunohistochemistry, and an anti-Tie2 antibody or peptide). All statistical tests were two-sided.

Results: VEGFR2 blockade created a „normalization window“ – a period, centered on day 5 after the start of DC101 treatment, during which radiation therapy delayed tumor growth for 22 days. (All other treatment schedules delayed growth for 17 days or less; P<.05.) This window was characterized by an increase in tumor oxygenation, which is a known radiation sensitizer (P<.0001 versus control mice). During the normalization window, but not before or after it, VEGFR2 blockade upregulated Ang-1 (P<.05 versus control mice), leading to an increase in the pericyte coverage of preexisting brain tumor vessels from 10% to 47% (day 5, P<.01). VEGFR2 blockade also degraded the pathologically thick basement membrane of tumor vessels (P<.00001 versus control mice) by increasing the activity of MMPs.

Conclusions: VEGFR2 blockade can temporarily normalize tumor vessel structure, leading to improved vascular function and enhanced response to radiation therapy. Thus, for optimal therapeutic response, the combination of antiangiogenic and radiation therapies must be carefully scheduled.