Developing a Multiepitope Vaccine Against Helicobacter pylori using the CTLA-4 Extracellular Domain

Helicobacter pylori is a pathogen that has been consistently linked to gastrointestinal cancer, particularly gastric cancer. Reports indicate that this pathogen is resistant to antibiotic treatments, necessitating the development of new treatment strategies. Here, to design a vaccine against H. pylori immunoinformatic tools were used. For this purpose, five virulence proteins including Flagellin B (FlaB), Flagellin A (FlaA), Urease subunit beta (UreB), CagA and Vacuolating cytotoxin autotransporter (VacA) were selected. Antigenicity and allergenicity were evaluated, and the epitopes with the highest scores were chosen. Linkers were used to connect the epitopes and an extracellular domain of CTLA-4 was positioned on the N-terminal. Biochemical features were predicted with the ProtParam server, and the second structure was predicted with the Prabi server, while the third structure was predicted with the Robetta, Alphafold and I-TASSER servers. VaxiJen and AllerTOP servers were used to evaluate the vaccine's antigenicity and allergenicity, respectively. After confirming the structure of the designed vaccine, molecular docking was performed with the TLR5 molecule using AutoDock Vina software. The SnapGene tool was utilized to in silico cloning of the vaccine in pET-3a vector. To evaluate the efficacy of the recombinant vaccine, the multi-epitope gene of H. pylori was cloned into the pCDNA3.1 vector, and its expression was analyzed in the spleen tissue of BALB/c mice using RT-qPCR. It has been shown in the results that the vaccine designed can bind to the TLR5 molecule on the surface of immune cells. Despite being an antigen, this vaccine didn't have any allergenic properties. Immunoinformatic is a promising tool for designing various drugs and vaccines. The results demonstrated that the expression levels of TNF-α and IFN-γ were significantly increased in the vaccine-treated group compared to the control group (P < 0.05). The elevated expression of pro-inflammatory cytokines indicates that the designed DNA vaccine successfully induced a cellular immune response against H. pylori. Keywords: H. pylori; Vaccine design; CTLA-4; Immunoinformatic; Gastric cancer