Protective Effect of a bi-specific Fc-fusion protein on the barrier of hRPE cells

To evaluate the protective effects of IBI302, a bi-specific Fc-fusion protein that theoretically can bind vascular endothelial growth factor (VEGF), complement C3b and C4b in the cultured human retinal pigment epithelial (hRPE) cells. Primary hRPE cells were isolated and cultured to monolayer barrier. hRPE monolayers were divided into control group, VEGF-Trap group, complement receptor 1 (CR1) group, and IBI302 group. Identification of hRPE cells, barrier function, inflammation factors and immune response products were tested by immunofluorescent staining, transepithelial resistance (TER) and ELISA. IBI302 treatment significantly improved TER of hRPE cells after complement-activated oxidative stress compared with control group, VEGF-Trap group, and CR1 group. The maximum effect of IBI302 on protecting hRPE cell viability was observed at the concentration of 1μg/ml. Elevated expression of VEGF, CCL2, C3a, C5a and MAC were inhibited by IBI302. IBI302 could protect the barrier function of hRPE cells. IBI302 might be a potentially effective drug for the RPE barrier-associated ocular diseases.

A novel vascular endothelial growth factor-trap KP-VR2 with enhanced ligand blocking

Antiangiogenic therapies targeting vascular endothelial growth factor (VEGF)-A have been commonly used in clinics to treat cancers over the past decade. However, their clinical efficacy has been limited, with drawbacks including the acquisition of resistance and activation of compensatory pathways resulting from elevated circulating VEGF-B and placental growth factor (PlGF) levels. Thus, we developed a novel VEGF-Trap, KP-VR2, which can neutralize VEGF-A, VEGF-B, and PlGF to mediate these problems. KP-VR2 consists of two consecutive second Ig-like domains (D2s) of VEGF receptor 1 (VEGFR-1) fused to human IgG1 Fc. KP-VR2 showed more potent decoy activity than the current VEGF-Trap against VEGF and PlGF. Most importantly, two consecutive D2s of VEGFR-1 can generate two putative binding sites, resulting in a significant improvement in binding capacity. These advances resulted in stronger antitumor efficacy in implanted tumor models than aflibercept and bevacizumab. Overall, the results of this study highlight KP-VR2 as a promising therapeutic candidate for further clinical drug development.

Newcastle Disease Virus Expressing an Angiogenic Inhibitor Exerts an Enhanced Therapeutic Efficacy in Colon Cancer Model

Numerous studies demonstrate that the NDV-mediated gene therapy is a promising new approach for treatment of cancer. VEGF-Trap plays a vital role in anti-angiogenesis. Therefore, we hypothesize that a recombinant NDV (rNDV) expressing VEGF-Trap would be an ideal agent for the colon cancer therapy. In this study, VEGF-Trap gene was incorporated into the genome of rNDV (named rNDV-VEGF-Trap). rNDV-VEGF-Trap reduced cell growth ratio by 85.37% and migration ratio by 87.9% in EA.hy926 cells. In vivo studies, treatment with rNDV-VEGF-Trap reduced tumor volume of CT26-bearing mice by more than 3 folds and tumor weight by more than 4 folds. Immunohistochemistry analysis of CD34 showed rNDV-VEGF-Trap significantly decreased the number of vascular endothelial cells in the tumor tissues of the tumor-bearing mice. Moreover, Western blot analysis demonstrated that treatment with rNDV-VEGF-Trap significantly decreased the phosphorylation levels of AKT, ERK1/2 and STAT3 and increased the expression levels of P53, BAX and cleaved caspase-3 in the tumor tissue. In addition, to evaluate the toxicity of VEGF-Trap, serum chemistries were analyzed. The results showed that rNDV-VEGF-Trap caused insignificant changes of creatinine levels, alanine aminotransferase (ALT) and aspartate transaminase (AST). Futhermore, administration of rNDV-VEGF-Trap did not cause the diarrhoea，decreased appetite, weight decrease and haemorrhage of the experimental mice. These data suggest that rNDV-VEGF-Trap exhibits an enhanced inhibition of CT26-bearing mice by enhancing anti-angiogenesis and apoptosis. rNDV-VEGF-Trap is a potential candidate for carcinoma therapy especially for colon cancer.

Abstract WP328: Afilbercept, a Vegf-trap, Reduces Vascular Permeability and Stroke-induced Brain Swelling in Obese Mice

Introduction: Brain swelling is a pathological event of ischemic stroke that progresses the enlargement of the brain injury, especially in comorbid conditions. In our previous study, we reported that diabetic obese mice had disproportionately increased stroke-induced brain swelling and that VEGF signaling was involved in this pathophysiology. Aflibercept (VEGF-Trap), an FDA approved drug that targets vascular endothelial growth factor receptor (VEGFR), has been used to treat patients with age-related macular degeneration and metastatic colorectal cancer. This study addressed whether Aflibercept can be used to reduce brain swelling in obesity stroke. Methods: HUVECs were cultured and treated with rVEGF (50 ng/ml) in the presence or absence of VEGF-Trap (1 ug/ml). Permeability was assessed by the incubation with FITC-dextran. Tube formation was also assessed. Diet-induced obese mice were generated by feeding a high-fat diet (HFD) for 8 weeks for males and 12 weeks for females to achieve similar insulin resistance. Their body weight gain and glucose clearance by glucose tolerance test was evaluated. Obese mice were subjected to transient MCAO. VEGF-Trap (10 mg/kg) or IgG (Fc control) was injected intravenously at 3h post-MCAO. VEGFR2 and VEGF-A gene expression was measured by realtime RT-PCR. Infarct size, brainswelling and neurological score were determined. Results: VEGF-Trap reduced rVEGF-A-mediated permeability by 50.0±10.7% (p<0.01, n=7) and tube formation by 28.5±3.5% (p<0.001, n=6) in HUVEC cultures. Stroke-induced VEGFR2 and VEGF-A gene expression was reduced in the mice that received VEGF-trap (38.9±1.0% (VEGFR2) and 34.1±1.4% (VEGF-A), p<0.05, n=7/group). Compared to IgG control, treatment of VEGF-Trap reduced brain swelling (25.3±3.5 % (IgG, n=20) vs 17.3±1.7 % (VEGF-Trap, n=22), p<0.05) and neurological score without reaching a statistical significance (p=0.06). There was no difference in infarct size between the two groups (33.0±3.9 mm 3 (IgG) vs 40.0±4.8 mm 3 (VEGF-Trap), ns). Conclusion: The study demonstrates that targeting VEGF signaling effectively reduces stroke-induced brain swelling in obese mice. The observed benefit suggests the repurposing of VEGF-Trap as a potential therapy in treating stroke in obese subjects.