Scientists decipher key factors in chronic hepatitis B virus infection, taking a step closer to a cure.

The journal Nature Microbiology published a research paper on hepatitis B virus (HBV) on the 10th. Scientists at Princeton University identified a group of proteins that decipher a key issue in the virus's life cycle, providing new directions for the treatment of chronic HBV infection.

Chronic hepatitis B virus infection is a common cause of liver disease, causing nearly 900,000 deaths each year, and is also the leading cause of liver cancer.

The first hepatitis B vaccine was introduced in the 1980s, and most children today are vaccinated at birth to avoid HBV infection. Although the vaccine can effectively prevent new infections, its help is limited for existing chronic infections. According to estimates by the World Health Organization, there are currently at least 257 million HBV carriers worldwide, with a particularly severe disease burden in Southeast Asia.

Current treatment methods include antiviral drugs targeting HBV reverse transcriptase, which are crucial for suppressing viral replication. However, unfortunately, they still cannot eradicate the virus within liver cells or achieve a true cure. Even some patients who continuously receive antiviral treatment still face the risk of developing liver cancer and cirrhosis. For this reason, chronic HBV infected individuals need more effective therapies.

Why is it so difficult to eliminate HBV from liver cells? This is closely related to the virus's life cycle, and it is necessary to first understand the genetic material of HBV. It is a very small enveloped virus, with genetic information encoded by DNA. When it first enters the host cell, the virus's genome appears in a form known as relaxed circular DNA (rcDNA), which has multiple gaps. To stably reside in the nucleus of liver cells, rcDNA undergoes a transformation through DNA repair, forming a covalently closed circular DNA (cccDNA) that exists long-term.

"The transition from rcDNA to cccDNA is crucial for the persistence of HBV. If we cannot understand this process, clinical treatments aimed at completely eradicating the infection will still be a long way off," said Professor Alexander Ploss, who is responsible for this research.

The transition from rcDNA to cccDNA requires repair at multiple sites, while the HBV genome itself only encodes four gene products. According to current understanding, none of these catalyze this repair step. Scientists naturally speculate that the host cell's DNA repair mechanisms are needed here. However, which components of the DNA repair mechanism HBV actually utilizes has been an unsolved problem for decades.

To answer this fundamental question, Professor Ploss and Dr. Lei Wei employed an "unconventional" method in this study, using yeast to decipher HBV's replication mechanism.

"Yeast has proven to be one of the best biological models for studying cellular replication mechanisms, with advantages such as simple genetic manipulation, rapid reproduction, and numerous research tools. More importantly, many molecular processes that occur in human cells are almost identical in yeast cells," explained Professor Ploss. "What is most noteworthy is that yeast cell extracts can support the transition of HBV from rcDNA to cccDNA."

Thus, the two researchers developed a set of experimental systems using yeast cells, screening dozens of cellular repair factors, and ultimately identified five core components in DNA lagging strand synthesis that are essential for HBV cccDNA formation, including proliferating cell nuclear antigen, replication factor C protein complex, DNA polymerase δ, flap endonuclease, and DNA ligase I.

The five repair factors found in yeast, as predicted by the researchers, have fundamentally the same mode of action in human cells. The five repair factors purified from human cells are sufficient to complete the repair of rcDNA; removing any one of these five core components prevents the successful formation of cccDNA. This means that targeting any one of these factors could potentially prevent HBV infection.

This hypothesis was soon validated in preliminary experiments in HBV-infected liver cells. For one of the core components, DNA polymerase δ, the researchers used the inhibitor aphidicolin to block the formation of cccDNA.

In the researchers' view, this discovery is "a great starting point that could ultimately answer decades of questions about how hepatitis B virus DNA produces a stable form." Professor Ploss said, "This knowledge will help develop more effective therapies to combat this terrible disease."

References

[1] Wei, L., Ploss, A. (2020) Core components of DNA lagging strand synthesis machinery are essential for hepatitis B virus cccDNA formation. Nature Microbiology. https://doi.org/10.1038/s41564-020-0678-0

[2] Researchers identify factors essential for chronic hepatitis B infection. Retrieved Mar. 10, 2020, from https://medicalxpress.com/news/2020-03-factors-essential-chronic-hepatitis-infection.html

[3] Baking, brewing…and virology: a fresh look under the hood of hepatitis B virus! Retrieved Mar. 10, 2020, from https://naturemicrobiologycommunity.nature.com/users/56573-alexander-ploss/posts/61401-baking-brewing-and-virology-a-fresh-look-under-the-hood-of-hepatitis-b-virus