M200, an integrin α5β1 antibody, promotes apoptosis in proliferating endothelial cells

Abstract

3187 Background: The role of VEGF in mediating angiogenesis is well documented, but recent evidence indicates that integrins such as αvβ3 are also important. We identified α5 integrin using a proprietary gene array. The α5β1 integrin mediates the binding of endothelial cells to fibronectin in the extracellular matrix. The growth factors VEGF and bFGF also promote the recruitment of new blood vessels into expanding tumors. We assessed the relative roles of these factors in vitro and in vivo to elucidate the validity of inhibiting integrin α5β1. Methods: We measured affinity of a chimeric antibody, M200, against α5β1 integrin by BIAcore, and assessed the role of α5β1 using an in vitro angiogenesis model and in vivo in a laser-induced model of choroidal neovascularisation (CNV) in cynomolgus monkeys. We compared M200 to an anti-VEGF mAb for effects on endothelial cell survival and proliferation in vitro. Results: M200, and its derivative Fab F200, inhibit binding of fibronectin to α5β1. M200 and F200 have similar potencies by BIAcore (Kd∼0.3nM). Both M200 and F200 show significant inhibition both VEGF and bFGF dependent tube formation in an in vitro angiogenesis model, and of neovascularization in the cynomolgus CNV model. M200 inhibits HUVEC proliferation more effectively than an anti-VEGF mAb (0.40 nM vs. 45 nM) at both low and high VEGF concentrations (2ng-50 ng/ml). No additive affect was observed when cells were challenged with anti-VEGF and M200 together. Thus M200 exerts novel cytotoxic effects in addition to recapitulating the anti-proliferative properties of anti-VEGF. This difference in potency was found to be attributable to the unique ability of M200 to selectively induce apoptosis in proliferating, but not senescent, endothelial cells. Conclusions:Our data shows that M200 is a potent anti-angiogenic with significant activity in vitro and in vivo. Further evaluation of the mechanism in vitro suggests that signaling via α5β1 and VEGF occur through overlapping mechanisms, and that M200 exerts a more significant growth inhibition of HUVEC in vitro than anti-VEGF, independent of growth factor stimulation, and this may translate to better clinical efficacy Author Disclosure Employment or Leadership Consultant or Advisory Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Protein Design Labs, Inc. Protein Design Labs, Inc.