Investigating the Immunogenic Properties of a Mutagenized NS3/4A-Based HCV Genotype 3a DNA Vaccine

Chronic hepatitis C virus (HCV) infection poses a major health risk worldwide, with patients susceptible to liver cirrhosis and hepatocellular carcinoma. This study focuses on the development of effective therapeutic strategies for HCV infection through the investigation of immunogenic properties of a DNA construct based on the NS3/4A gene of HCV genotype (g)3a. Gene expression of the mutagenized (mut) NS3/4A target genes was assessed through reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis. Additionally, bioinformatics tools were employed to evaluate the impact of the mut-NS3/4A-based DNA vaccine. Analysis revealed increased mut-NS3/4A mRNA levels and target protein abundance compared with the native sequence. Elevated mut-NS3/NS4A levels could result from increased RNA stability and proper protein folding. Physicochemical analyses of the protein demonstrated favorable attributes such as thermostability and solubility. Three-dimensional mut-NS3/4A protein modeling confirmed its high stability and agreement with known protein structures. Additionally, potential immunogenic regions of both T and B cell epitopes were discovered based on peptide binding to major histocompatibility complex molecules of Asian origin. Importantly, these epitopes exhibited nonallergenic and nontoxic characteristics. These findings highlight the potential of the NS3/4A-based DNA construct as a promising candidate for an HCVg3a vaccine tailored for the Asian population, providing valuable insights for future immunotherapeutic approaches.