Bioinformatic designing for producing vaccine peptide of human vascular endothelial growth factor (VEGF-A), and evaluation of polyclonal antibodies in mice

Abstract

Background: It is stated that in the absence of angiogenesis, the tumoral tissue will not grow beyond 2 mm3. Vascular endothelial growth factor (VEGF) plays a pivotal role in angiogenesis and blockade of this phenomenon could be applied as a novel strategy for immunotherapy of cancer. Methods: Peptide sequences of VEGF-A isoforms were derived from protein databases and aligned. Immunodominant epitopes were determined and the selected one was rechecked for dissimilarity with other human proteins. The selected conserved peptide sequence was synthesized and conjugated with Keyhole limpet hemocyanin (KLH). Then, it was applied for immunization of mice. The polyclonal anti-VEGF antibody titer was measured using an indirect peptide-enzyme-linked immunosorbent assay (ELISA) with a Bovine serum albumin (BSA)-conjugated peptide. Findings: According to bioinformatic findings, the selected 41-aminoacid sequence did not show any similarity with other human proteins and revealed enough antigenicity to stimulate anti-tumor specific responses. A substantial increase of specific antibody titer was observed in vaccinated mice. Sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE) analysis of the BSA-conjugated peptide showed efficient coupling of the molecules. Optimization steps in ELISA procedures revealed that coating of microtiter plates with BSA-conjugated antigen provided more reproducible outcome than unconjugated peptide. Conclusion: Our results reinforce the potential of KLH-conjugated peptides for immunization and production of specific polyclonal antibodies against VEGF-A. Production of high-titer antibodies against this autoantigen indicates that the designed peptide-vaccine could be used as a potential immunogen for stimulation of humoral immune system in animal model. © 2016, Isfahan University of Medical Sciences(IUMS). All rights reserved.