Evaluation of cytotoxicity of bevacizumab on VEGF-enriched corneal endothelial cells

Purpose: To evaluate the cytotoxicity of varying doses of Bevacizumab on corneal endothelial cells in the presence of a range of concentrations of vascular endothelial growth factor (VEGF). Bevacizumab, a drug widely used in the treatment of neovascular glaucoma neutralizes all isoforms of VEGF and ameliorates neovascularization after intracameral administration. However, the safety of intracameral administration of Bevacizumab and dose–dependent toxicity on corneal endothelial cells has not been established. Methods: Bovine corneal endothelial (BCE) cells were treated with VEGF (50 ng/ml) and/or Bevacizumab (0.1–2 mg/ ml) for 72 h. Cell proliferation was measured with the water soluble tetrazolium salts (WST-1) assay. Morphological changes were recorded by bright-field microscopy of cells. Cytotoxicity in response to Bevacizumab was evaluated by trypan blue exclusion, as well as annexin V/propidium iodide (PI) staining. Results: Bevacizumab was not cytotoxic at the concentrations tested and the percentage of Bevacizumab-treated cells staining positively for both PI and Annexin V was less than 1%. The anti-proliferative effects of Bevacizumab on BCE cells were dose-dependent; a dose of 1.5 mg/ml or 2 mg/ml produced a 33% (p=0.005) or 47% (p=0.001) decrease in cell proliferation compared to controls. Similar results were obtained in cells treated with a combination of Bevacizumab and VEGF. VEGF (50 ng/ml) had no significant effect on cell proliferation compared to controls. Morphology of cells was unchanged after treatment with Bevacizumab and/or VEGF compared to controls. Conclusions: Bevacizumab was safe and not toxic to BCE cells at concentrations commonly used in clinical practice.