Binding Characteristics of the VEGF Arm of a Novel Bispecific Protein RO-101 in Comparison to Faricimab-svoa

Abstract

Purpose : The purpose of this study is to investigate the half maximal effective concentration (EC50) and half maximal inhibitory concentration (IC50) of the vascular endothelial growth factor (VEGF) arm of a novel bispecific protein, RO-101, in comparison to Faricimab-svoa (first bispecific monoclonal antibody used in neovascular eye diseases). RO-101 binds both VEGF-A and Angiopoietin 2, which are two key intermediates in the pathway for neovascularization. VEGF₁₆₅ and VEGF₁₂₁ are two intermediates in neovascularization that will be used to analyze the binding of RO-101 and Faricimab-svoa.

Methods : An Enzyme-linked Immunoassay (ELISA) was utilized to determine the EC50 of RO-101 to recombinant human VEGF₁₆₅ (rhVEGF₁₆₅) and VEGF₁₂₁ (rhVEGF₁₂₁) . The binding of rhVEGF₁₆₅ and rhVEGF₁₂₁ binding in the ELISA were detected by DkHumIgGFc-HRP. SgG1 or IgG1 were tested in duplicates starting at 10 nM; 1:3 serial dilutions. Corning 9017 medium binding plate was used and OD450 nm was measured by Victor 3 (PerkinElmer 1420 Multilabel Counter). In a second assay, an ELISA was used to determine the IC50 of RO-101 and Faricimab-svoa to VEGF₁₆₅and VEGF R2/KDR Receptor. Inhibition of rhVEGF₁₆₅ and rhKDR receptor binding was detected by streptavidin-HRP.

Results : The EC50 of RO-101 binding to rhVEGF₁₆₅ was found to be 0.0231 nM. Meanwhile, the EC50 of Faricimab-svoa binding to rhVEGF₁₆₅ was found to be 0.0228 nM. The EC50 of RO-101 binding to rhVEGF₁₂₁ was found to be 0.0086 nM and the EC50 of Faricimab-svoa binding to rhVEGF₁₂₁ was found to be 0.0113 nM (Figure 1). The IC50 of RO-101 to rhVEGF₁₆₅ was found to be at a lower concentration, 1.989 nM, than the IC50 of Faricimab-svoa, which was 6.471 nM (Figure 2).

Conclusions : The data suggests the EC50 for RO-101 to both VEGF₁₆₅ and VEGF₁₂₁ is similar to the EC50 of Faricimab-svoa when binding to both growth factors respectively. The IC50 data for the inhibition of rhVEGF₁₆₅ and rhKDR receptor binding was 3.25 times weaker when binding with Faricimab-svoa compared with RO-101. The IC50 data is suggestive that RO-101 could be effective at lower concentrations than Faricimab-svoa.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

 

Figure 1: Comparison of RO-101 binding to rhVEGF121 and rhVEGF165 vs Faricimab-svoa binding to rhVEGF121 and rhVEGF165.

 

Figure 2: Comparison of RO-101 inhibition of rhVEGF165+KDR vs Faricimab-svoa to rhVEGF+KDR vs control rhVEGF+KDR receptor.