Silent mutations in coding regions of Hepatitis C virus affect patterns of HCV RNA structures and attenuate viral replication and pathogenesis

This is the source code for the following paper, published in Genome Biology, 2025 "Silent mutations in coding regions of Hepatitis C virus affect patterns of HCV RNA structures and attenuate viral replication and pathogenesis" Authors: Roba Dabour, Shaked Bergman, Zohar Zafrir, Ateret Davidovitch, Michal Werbner, Meital Gal-Tanamy, Tamir Tuller Abstract Vaccines based on live attenuated viruses are the most effective strategy for controlling infections, since they elicit long-lasting natural and effective immune response, but entail challenges as safety and virulence. Hepatitis C Virus (HCV) is a major global health problem, causing liver diseases and liver cancer, with millions infected each year and hundreds of thousands of annual fatalities; but no vaccine is currently available for the virus. Here we present a novel computational approach for the accurate prediction of virus attenuation. We rationally designed viral variants by inserting a large number of synonymous mutations to disrupt the viral RNA’s secondary structure and regulatory sequences important for the viral life cycle. By measuring RNA levels and virus spread in an HCV infection model, we showed that these variants have lower viral fitness relative to the wild-type virus, with gradient of attenuation in concordance with the prediction model. Deep sequencing of replicating viruses demonstrated genomic stability of the attenuated variant. Differential expression analysis and evaluation of cancer-related phenotypes revealed that the variants have a lower pathogenic influence on the host cells, compared to the WT virus. These rationally designed variants may be further considered as a promising direction for a viable HCV vaccine. Importantly, the computational approach described here is based on the most fundamental viral regulatory motifs and therefore may be applied for almost all viruses as a new strategy for vaccine development.