طراحي بيوانفورماتيكي و ارزيابي آزمايشگاهي يك واكسن پپتيدي با استفاده از آنتي¬ژن ساركوما NY-SAR-35 همراه با ادجوانت ميكروبي براي درمان سرطان پستان

Bioinformatics design and experimental eva‎luation of a peptide vaccine using sarcoma antigen NY-SAR-35 along with a microbial adjuvant for breast cancer treatment

زيست فناوري

Breast cancer is the most common type of cancer among women, resulting from uncontrolled growth, proliferation, and invasion of cells. Designing a cancer vaccine has become one of the main focuses of research in medical biotechnology and immunology. This study aimed to design a peptide vaccine based on epitopes derived from the cancer/testis antigen NY-SAR-35 against breast cancer. To achieve this, the sequence of the NY-SAR-35 was obtained from UniProt. T-cell epitopes were predicted using IEDB and eva‎luated for antigenicity, immunogenicity, cytotoxicity, stability, and ability to induce IFN-γ and IL-4. The best epitopes were selected to construct of the final vaccine. The selected epitopes were linked using linkers. The binding affinity of the final construct to MHC molecules and their stability were assessed using docking and molecular dynamics methods. Additionally, the ability of the final construct to stimulate an immune response was eva‎luated using C-ImmSim. The designed vaccine was synthesized after bioinformatics validation. For immunological eva‎luation, BALB/c mice were used and divided into four groups: the first group (PBS) received PBS, the second group (Adjuvant) received Freundʹs adjuvant, the third group (Peptide) received peptide, and the fourth group (Adj+pep) received peptide with Freundʹs adjuvant. Two weeks after the second and third injections, blood was collected and splenocyte cultures were prepared from the mice. To measure IgG antibody levels in the serum and IFN-γ, IL-4, and granzyme B levels in the culture supernatant, ELISA was used. As a final step, the potential effect of the vaccine on splenocyte proliferation was assessed using MTT. Bioinformatics results showed that HRAMRVAHL and FVCYYLSYY had the highest antigenicity and were linked together by AAY to construct the final construct. This final construct was antigenic, non-allergenic, non-toxic, stable, and capable of stimulating IFN-γ and IL-4. Docking and molecular dynamics results indicated that the vaccine had a high affinity for mouse and human MHC molecules and that its binding to most MHC molecules increased their stability. C-ImmSim results showed that the vaccine induced a cellular immune response. Laboratory results indicated that the peptide vaccine with Freundʹs adjuvant significantly increased levels of IgG, IFN-γ, IL-4, and granzyme B, meaning the vaccine stimulated both humoral and cellular immune responses. Examination of splenocyte proliferation showed that the vaccine had a remarkable effect on increasing these cells. Based on the results, the designed vaccine could be a promising candidate against breast cancer.

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