Infections with hepatitis C virus (HCV) are persistent and will severely damage and ultimately destroy the liver without treatment. HCV infections are spread through unsterilized needles; in the UK and other developed countries, HCV typically targets injecting drug users, but it is also widely distributed worldwide with an estimated 160 million people infected with the virus and at risk of developing life-threatening complications of infection. While effective treatments for HCV have been recently developed, they are expensive and their use will remain cost-limited for many years. Importantly, such treatments will not stem the spread of HCV and the large numbers of new infections recorded every year; even those who have been effectively treated remain prone to re-infection. HCV also poses a much greater medical problem worldwide where access to treatment will remain limited in much the same way as HIV therapy. HCV-related deaths worldwide exceed 350,000 per year, of the same order to those attributed to AIDS, TB and malaria.
An effective vaccine for HCV would induce life-long immunity to HCV and prevent further spread of the virus to those currently at risk for infection. There is a further tantalising possibility that therapeutic immunisation with such a vaccine may be able to control and clear infection in those already infected with HCV. The development of such vaccines would therefore have profound effects on global health.
The development of HCV vaccine has been hampered by a lack of animal models with which to test different type of vaccine. It has also proven exceptionally difficult to develop vaccines that induce long-lived protective immunity - even clearance of natural infections does not protect from subsequent re-infection, quite unlike the situation of poliovirus, measles and hepatitis A virus (as examples) for which effective vaccines have been developed.
An important recent development in ongoing attempts to develop an effective vaccine for HCV is the is the very recent discovery of a virus called rodent hepacivirus (RHV). RHV is closely related virus to HCV that infects rats and causes the same pattern of liver disease and frequency of persistence as HCV in humans. In our planned project, we will develop methods to attenuate the replication of RHV though modification of its CpG and UpA dinucleotide composition. Previous data from influenza A virus (Gaunt et al., 2016) indicates that incorporating additional CpGs can greatly enhance the cellular innate and inflammatory responses proteins expressed in infected cells and may potentiate the downstream adaptive immune responses. Infection with such attenuated viruses may induce strong immune responses that are protective from challenge with wild type virus. The attenuated vaccine may potentially also create a sufficiently powerful immune response to cure rats chronically infected with wild type virus.
The DPhil will provide experience and training in a wide range of laboratory molecular virology methods, gene expression technology, whole host studies, bioinformatic analysis methods. The work is at the forefront of our growing understanding of the remarkably complex regulatory systems underlying gene expression and how these can be manipulated for biotechnology purposes.
The DPhil will provide experience and training in a wide range of laboratory molecular virology methods, in vivo studies and in immunology. These include:
The project will take place alongside groups of scientist working the supervisors’ groups investigating other aspects of vaccine design and HCV biology / immunology.
Project reference number: 838
|Professor Peter Simmonds||Experimental Medicine Division||Oxford University, Peter Medawar Building||GBRfirstname.lastname@example.org|
|Professor Ellie (Eleanor) Barnes||Experimental Medicine Division||Oxford University, Peter Medawar Building||GBRemail@example.com|
Most RNA viruses infecting mammals and other vertebrates show profound suppression of CpG and UpA dinucleotide frequencies. To investigate this functionally, mutants of the picornavirus, echovirus 7 (E7), were constructed with altered CpG and UpA compositions in two 1.1-1.3 Kbase regions. Those with increased frequencies of CpG and UpA showed impaired replication kinetics and higher RNA/infectivity ratios compared with wild-type virus. Remarkably, mutants with CpGs and UpAs removed showed enhanced replication, larger plaques and rapidly outcompeted wild-type virus on co-infections. Luciferase-expressing E7 sub-genomic replicons with CpGs and UpAs removed from the reporter gene showed 100-fold greater luminescence. E7 and mutants were equivalently sensitive to exogenously added interferon-β, showed no evidence for differential recognition by ADAR1 or pattern recognition receptors RIG-I, MDA5 or PKR. However, kinase inhibitors roscovitine and C16 partially or entirely reversed the attenuated phenotype of high CpG and UpA mutants, potentially through inhibition of currently uncharacterized pattern recognition receptors that respond to RNA composition. Generating viruses with enhanced replication kinetics has applications in vaccine production and reporter gene construction. More fundamentally, the findings introduce a new evolutionary paradigm where dinucleotide composition of viral genomes is subjected to selection pressures independently of coding capacity and profoundly influences host-pathogen interactions. Hide abstract
Previously, we demonstrated that frequencies of CpG and UpA dinucleotides profoundly influence the replication ability of echovirus 7 (Tulloch et al., 2014). Here, we show that that influenza A virus (IAV) with maximised frequencies of these dinucleotides in segment 5 showed comparable attenuation in cell culture compared to unmodified virus and a permuted control (CDLR). Attenuation was also manifested in vivo, with 10-100 fold reduced viral loads in lungs of mice infected with 200PFU of CpG-high and UpA-high mutants. However, both induced powerful inflammatory cytokine and adaptive (T cell and neutralising antibody) responses disproportionate to their replication. CpG-high infected mice also showed markedly reduced clinical severity, minimal weight loss and reduced immmunopathology in lung, yet sterilising immunity to lethal dose WT challenge was achieved after low dose (20PFU) pre-immunisation with this mutant. Increasing CpG dinucleotide frequencies represents a generic and potentially highly effective method for generating safe, highly immunoreactive vaccines. Hide abstract
Chronic hepatitis C virus infection is now curable by antiviral therapy but the global burden of liver disease is unlikely to diminish without a vaccine to prevent transmission. The objective of HCV vaccination is not to induce sterilizing immunity, but instead to prevent persistent infection. One vaccine that incorporates only non-structural HCV proteins is now in phase I/II efficacy trials to test the novel concept that T cell priming alone is sufficient for protection. Evidence also suggests that antibodies contribute to infection resolution. Vaccines comprised of recombinant envelope glycoproteins targeted by neutralizing antibodies have been assessed in humans for immunogenicity. Here, we discuss current concepts in protective immunity and divergent approaches to vaccination against a highly mutable RNA virus. Hide abstract
Recent advances in sequencing technologies have greatly enhanced our abilities to identify novel microbial sequences. Thus, our understanding of the global virome and the virome of specific host species in particular is rapidly expanding. Identification of animal viruses is important for understanding animal disease, the origin and evolution of human viruses, as well as zoonotic reservoirs for emerging infections. Although the human hepacivirus, hepatitis C virus (HCV), was identified 25years ago, its origin has remained elusive. In 2011, the first HCV homolog was reported in dogs but subsequent studies showed the virus to be widely distributed in horses. This indicated a wider hepacivirus host range and paved the way for identification of rodent, bat and non-human primate hepaciviruses. The equine non-primate hepacivirus (NPHV) remains the closest relative of HCV and is so far the best characterized. Identification and characterization of novel hepaciviruses may in addition lead to development of tractable animal models to study HCV persistence, immune responses and pathogenesis. This could be particular important, given the current shortage of immunocompetent models for robust HCV infection. Much remains to be learned on the novel hepaciviruses, including their association with disease, and thereby how relevant they will become as HCV model systems and for studies of animal disease. This review discusses how virome analysis led to identification of novel hepaci- and pegiviruses, their genetic relationship and characterization and the potential use of animal hepaciviruses as models to study hepaciviral infection, immunity and pathogenesis. This article forms part of a symposium in Antiviral Research on "Hepatitis C: Next steps toward global eradication." Hide abstract