Selective IκB kinase β inhibitor IMD-0354 is a potent antineovascular compound in model of corneal neovascularisation in rats

Abstract

Purpose : IMD0354 is a known specific NFκB inhibitor, which acts by inhibiting the IKK-induced phosphorylation of IkBα, and results in reduced NFκB activation. There was several publications regarding IMD0354 potent antinflamatory effect in several animal models, but it’s potency as anti-angiogenic agent, has not been thoroughly investigated. Here we use the model of inflammatory induced corneal neovascularisation in rats to evaluate IMD0354 immune modulated effect on corneal neovascularisation in rats as well as to study the direct effects on HUVEC cell culture in vitro.

Methods : The surgical procedure consisted in placement of two intrastromal corneal sutures (10-0 nylon) at a distance of 1.5 mm from the temporal limbus in 10 weeks old Wistar rats. All procedures involving animals were performed in accordance with the ARVO resolution on the use of animals in research. IMD-0354 30 mg/kg was suspended in 200 µl of 0.5% carboxymethylcellulose (CMC) vehicle immediately before use. Animals were administered with CMC or IMD0354 via IP injection every 48 hours. Formation of new vessels in cornea, limbal vessel dilation and inflammatory cells infiltration were studied for 4 days in-vivo.
Ex-vivo analysis included: qPCR, IHC and WB analysis of VEGFa, CXCL5, CCL2, CXCR2, CD45 in corneal tissue. In-vitro study of IMD0354 at 10 ng/ml in HUVEC cells evaluated in cell migration and Geltrix tube formation assays, as well as expression profiling for VEGF, CXCL5 and CCL2, using IF and WB analysis.

Results : IMD0354 treatment have significantly reduced inflammatory corneal cell infiltration (p<0.05) and vascular dilation (p<0.05) in vivo. qPCR data, supported with IHF and WB observations indicated reduced expression of VEGF (p<0.01), CXCL5 (p<0.01), CCl2 (p<0.01), CXCR2 (p<0.05) and CD45 in the corneal tissue.
In-vitro experiments demonstrated a direct inhibitory effect of IMD0354 on cell migration (p<0.01) and tube formation (p<0.01) assays. IHF and WB findings supported reduced expression of VEGFa, CXCL5 and CCL2 in-vitro.

Conclusions : IMD0354 have immune system modulated and direct inhibitory effect on corneal neovascularisation in rats by reduction of NFkB downstream signaling and endothelial cells mobility in-vivo and in-vitro.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.