Role of heparin‐binding mediators in the resistance to anti‐VEGF therapies in diabetic retinopathy

Abstract

Aims/Purpose: Anti‐vascular endothelial growth factor (VEGF) drugs represent the standard of care for diabetic retinopathy (DR); however, lack of response occurs in approximately 50% of patients. Aim of our work is the identification of the mediators responsible for anti‐VEGF drug resistance.Methods: Vitreous samples were collected from anti‐VEGF non‐responder (non‐RES) patients; non‐RES heat‐inactivated (HI) vitreous were used as control. To develop a new orthotopic model of DR, non‐RES or HI vitreous were injected intravitreally in mice and after 48 h retinas were explanted and analysed. The composition of non‐RES vitreous was assessed by an angio‐inflammatory protein array. The effect of the inhibition of the bioactive heparin‐binding mediators (HBMs) present in non‐RES vitreous was evaluated in the endothelial spheroid sprouting assay.Results: Mouse orthotopic intravitreal injection of non‐RES vitreous upregulated the expression of pro‐angiogenic/pro‐inflammatory mediators; moreover, it induced alterations of retinal vessels, accumulation of inflammatory infiltrate, and activation of astrocyte gliosis. Notably, HI vitreous was ineffective, pointing to a proteinaceous nature of the non‐RES vitreal mediators acting on mouse retinae. In this context, the blockade of vitreal HBMs mediated by the biotechnological heparin‐like molecule K5‐N, OS (H) or by the pentapeptide Boc2, completely abrogated the activity of non‐RES samples. It is worth noticing that the potency of anti‐HBP compounds was significantly higher than the anti‐VEGF ranibizumab, which was partially effective only at the clinically relevant and saturating concentration of 10 μM, with no further improvement at a double dose.Conclusions: Our data show that VEGF is only part of a complex machinery regulating angiogenesis, inflammation and oedema formation during DR and that drug resistance is due to the activation of compensatory mechanisms mediated by HBMs besides VEGF.