Targeting NRP1 with EG00229 induces neurovascular permeability

Abstract

AbstractNRP1 is a therapeutic target to inhibit vascular endothelial growth factor (VEGF)-induced blood vessel dysfunction. The small molecule EG00229 inhibits VEGF binding to NRP1 and reduces pathological blood vessel growth. However, it is unknown whether it could be used to reduce VEGF164-induced vascular leakage, which often exacerbates ischemic diseases due to VEGF upregulation. Here, we show that EG00229 increased rather than inhibited vascular leakage in perfused retinal explants and across primary brain EC monolayers, either when added alone or together with VEGF164. EG00229-induced vascular leakage was not an off-target effect, because it required its intended target, endothelial NRP1 expression and NRP1’s VEGF164 binding pocket, but was independent of VEGFR1 and VEGFR2. Moreover, EG00229 activated molecular events typical of VEGF164-induced paracellular permeability, including p38 MAP kinase (p38) and SRC family kinase (SFK) phosphorylation as well as endothelial junction rearrangement. Investigating EG00229-induced signalling therefore helps elucidate NRP1-dependent mechanisms of paracellular permeability induction and, in the future, might also help identify new approaches to modulate the neurovascular barrier.