The roles of the pre-genomic RNA in Hepatitis B Virus nucleocapsid assembly.

Lead Research Organisation: University of Leeds
Department Name: Astbury Centre

Abstract

Hepatitis B virus is a common infection in humans with over 2 billion people worldwide having been exposed to the virus. The majority of people recover fully from such infections but a significant minority do not, becoming carriers of the virus, a condition linked to later lethal cirrhosis and liver cancer. It is estimated that ~1 million people die every year as a result of HBV infection. In advanced western countries there is an effective vaccine, but this is not sufficient in many parts of the world where over 8% of the population may be infected carriers. As a result there is a need to devise novel anti-viral drugs that would inhibit HBV infections, and in particular render the virus visible to the immune system so that it can be cleared from patients. This is the goal of our application, which is based on a recent discovery we made relating to the way that such viruses having RNA as their genomic nucleic acid, assemble their viral particles. We have discovered that they encode a hidden instruction manual in the sequence and structure of these RNAs that ensures rapid and efficient assembly of infectious virions. HBV is a slightly unusual virus because it assembles initially around an RNA pregenomic RNA but later converts that to a double-stranded DNA version within the viral particle. In extensive preliminary experiments we have shown that HBV may well contain the same sort of instruction manual for its initial assembly. It should therefore be possible to target this process using drugs. We are asking for a two year pump-priming grant that will allow us to investigate this mechanism more comprehensively and begin to screen for the types of ligands (drugs) that could block assembly. In other viral systems, such drugs lead to partial assembly, that is they create non-infectious but highly immunogenic versions of the virus that are readily seen by the immune system. The information we gain here will be invaluable in future discussions with drug companies about developing novel anti-HBV therapy.

Technical Summary

This is a two year pump-priming grant to enable us to explore the roles of the HBV pre-genome in assembly of the nucleocapsid. Our objectives are to: define precisely the recognition motif in the RNA packaging signals and their role(s) in binding either the CP or the polymerase, using a combination of bioinformatics, smFCS & cryo-EM, and the consequences of those interactions for efficient assembly of the T=4 HBV nucleocapsid. The role(s) of the polymerase-epsilon complex as well as phosphorylation of the CP in RNA binding will also be examined. In addition we will screen, using mass spectrometry, small numbers of known drug ligands likely to bind nucleic acids for binding to PSs. This work will lay the foundations of a novel anti-viral strategy for treatment of HBV infection.

Planned Impact

The World Health Organisation estimates that over 2 billion people have been infected by Hepatitis B virus (HBV), with >350 million becoming chronic carriers, leading to ~1 million HBV-related deaths annually, including the slow and expensive to treat cirrhosis and cancer of the liver. Estimates suggest that chronic HBV costs the NHS between £26-375M pa. These figures are despite the fact that there is an extremely safe vaccine for this virus, as well as a number of effective anti-viral drugs. Most infected people, however, live in the developing world, particularly in Asia and Africa, where the incidence of infection is >8%. This level of infection requires continuing efforts to understand the basic biology of the virus in order to identify novel routes of anti-viral intervention. This is the goal of this grant application. The Universities of Leeds, York and Helsinki have patented applications of the assembly mechanism operating in HBV and other ssRNA viruses, so that drugs developed to block this process should generate income for these institutions. This work is thus of worldwide importance encompassing people who are potentially infected, chronic carriers of the virus, and healthcare workers and administrators who have to deal with this health burden.

Publications

10 25 50
 
Description • In situ Structural & Mechanistic Interrogation of dsDNA synthesis in Hepatitis B Virus
Amount £291,000 (GBP)
Funding ID MRF-044-0002-RG-PATEL 
Organisation Medical Research Council (MRC) 
Department Medical Research Foundation
Sector Charity/Non Profit
Country United Kingdom
Start 05/2018 
End 04/2021
 
Title RNA-binding ligand assay 
Description Immobilised ligands were prtobed with a fluorescent oligo encompoassing the viral drug target. 
Type Of Material Technology assay or reagent 
Year Produced 2018 
Provided To Others? Yes  
Impact There is now a paptenmt application from NIH 
 
Title Cryo-EM model of HBV VLP 
Description Cryo-EM public database of structures 
Type Of Material Database/Collection of data 
Year Produced 2017 
Provided To Others? Yes  
Impact None as yet 
 
Description Interviewed for website press release 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Supporters
Results and Impact The outcomes were reported via Press Release
Year(s) Of Engagement Activity 2017