A reverse vaccinomics approach for the designing of novel immunogenic multi-epitope vaccine construct against gas gangrene and related colorectal cancer for Clostridium septicum DSM 7534.

PURPOSE: Costridium septicum, the causative agent of clostridial myonecrosis or gas gangrene, has been linked with colorectal malignancies, particularly tumors of the ascending colon. Due to the pathogen's resistance and the limited understanding of its genomic structure, effective therapeutic strategies are needed. This study focuses on developing a vaccine to address this issue.

METHOD: In this study, we employed both subtractive proteomics and reverse vaccinology to identify potential therapeutics against C. septicum. Through subtractive proteomics, we pinpointed antigenic membrane proteins that harbor virulence factors, resistance factors, and essential components for this pathogenic bacterium. These identified membrane proteins were then used to design a chimeric subunit vaccine utilizing reverse vaccinology techniques. We utilized the most effective tools and servers to identify MHC class I, II, and B-cell epitopes. These epitopes were further evaluated computationally to assess their immunogenicity, antigenicity, allergenicity, conservancy, and toxicity. The selected immunogenic epitopes were incorporated into a chimeric subunit vaccine, complemented with immune-modulating adjuvants, linkers, and the PADRE (Pan HLA-DR epitope) amino acid sequence. The resultant vaccine construct was then shortlisyed based on their interaction with MHC molecules and the TLR4/MD2 complex via docking and molecular dynamics simulations.

RESULTS: Our analysis identified the Flagellar biosynthetic protein (FliR; ID: A0A386PD61) as a promising vaccine target based on systemic screening of proteome of C. septicum using subtractive genome analysis. Employing a Reverse Vaccinology approach, we identified 11 MHC-I, 19 MHC-II, and 33B-cell epitopes. Among the presented multi-epitope vaccines, the chimeric V1 had the strongest interaction with TLR4. Moreover, the immune simulation response revealed that the chimeric protein had the strongest ability to stimulate the immune system. In addition, molecular docking and molecular dynamic simulation indicated the proper and stable interactions between the chimeric cell-wall-associated transporter and TLR4 and HLAs, simulation, and in silico expression.

CONCLUSION: This study advances our understanding of preventive strategies against C. septicum by proposing a vaccine framework capable of inducing an immunogenic response. However, the effectiveness of this vaccine in preventing infection and potential colorectal cancer development requires validation through immunoassays (both in vitro and in vivo) and site-directed mutagenesis studies.