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Scripps Research scientists identify key interaction in Hepatitis C virus
JUPITER, Fla. – Scientists at the Florida campus of The Scripps Research Institute have discovered a molecular interaction between a structural hepatitis C virus protein (HCV) and a protein critical to viral replication.
This new finding strongly suggests a novel method of inhibiting the production of the virus and a potential new therapeutic target for hepatitis C drug development.
The study was published in the January 2010 issue of the Journal of General Virology.
Hepatitis C virus infects between 130 and 170 million people worldwide. Hepatitis C is an infectious disease affecting the liver, caused by the hepatitis C virus (HCV). The infection is often asymptomatic, but once established, chronic infection can progress to scarring of the liver (fibrosis), and advanced scarring (cirrhosis) which is generally apparent after many years. In some cases, those with cirrhosis will go on to develop liver failure or other complications of cirrhosis, including liver cancer or life threatening esophageal varices and gastric varices.
Because current HCV treatments are only partially effective, a number of alternative molecular mechanisms are actively being pursued as possible drug targets.
These new data underline the essential role of the viral protein known as "core" as a primary organizer of the infectious HCV particle assembly and support a new molecular understanding of the formation of the viral particle itself.
"While our finding that the HCV core interacts with the non-structural helicase protein was not totally unexpected, this had not really been confirmed until this study," Scripps Florida Professor Donny Strosberg, who led the study, says in a statement. "But the most exciting part is that small molecule inhibitors of dimerization (the joining of two identical subunits) of core actually inhibit interaction between core and helicase, thus possibly preventing production of an infectious viral particle."
One of the critical problems of finding inhibitors for the hepatitis C virus is that it mutates at such prodigious rates. An RNA virus such as hepatitis C can mutate at a rate estimated as high as one million times that of DNA viruses such as the herpes virus.
Looking closely at the core interaction with itself, Strosberg developed several novel quantitative assays or tests for monitoring these protein-protein interactions with the specific goal of identifying inhibitors of the core dimerization, which would block virus production.
"People have been dreaming about inhibiting protein-protein interactions, as a new El Dorado for finding novel drug targets," says Strosberg, "but few conclusive studies have emerged, except in the virus-host area."
Core is particularly important in the assembly of the hepatitis C nucleocapsid or capsid, an essential step in the formation of infectious viral particles; the nucleocapsid is the virus genome protected by a protein coat.
"In one sense, the ongoing issue with hepatitis C is that there are still so very few drugs to treat the virus and very few tools to study it," Strosberg points out. "We set out to develop new tools and to identify a new target – core, the capsid protein. By targeting the interactions of core with itself or other proteins, we could reduce the problem of rapid mutation not only because the core protein mutates significantly less, but also because mutations that would affect the interface between core and itself or other proteins would often be more likely to deactivate the virus, in contrast to mutations in viral enzymes which often lead to increased resistance to drugs."
Last year, Strosberg developed a novel quantitative test for monitoring these protein-protein interactions with the specific goal of identifying inhibitors of the core dimerization, which would block virus production. Strosberg and his colleagues uncovered peptides derived from the core protein of hepatitis C that inhibit not only dimerization of the core protein, but also production of the actual virus.
That earlier study led to the discovery of non-peptidic small organic molecules that strongly inhibited HCV production, one of which, SL201, was used in the new study.
In the new study, Strosberg and his colleagues focused on non-structural proteins that provide functions relating to HCV production, in particular NS3 helicase. The scientists' findings support a growing body of evidence that this protein participates in the assembly and production of infectious viral particles. The interaction of the core protein with this non-structural protein also confirms core as a key organizer of virus assembly and suggests it acts to facilitate the packaging and integration of the newly synthesized viral RNA.
The new research, however, led to the discovery of two peptides that inhibited HCV production by 68 percent and 63 percent, respectively; a third related peptide showed 50 percent inhibition. When added to HCV-infected cells, each of the three peptides blocked release but not replication of infectious virus; viral RNA levels were reduced by seven fold. Strosberg notes that the efficacy of small molecules like these can often be improved over initial levels.
"After we'd finished our work, we learned that Frank Chisari—one of the leading experts on HCV who works at Scripps Research in La Jolla—had been looking at similar peptides using a very different approach," notes Strosberg. "One of his peptides was the same as ours—it also inhibited virus production. It's an incredible coincidence and a confirmation of our study."
The first author of the study, "Dimerization-Driven Interaction of Hepatitis C Virus Core Protein with Ns3 Helicase," is Guillaume Mousseau of Scripps Research. Additional authors include Smitha Kota, S. and Virginia Takahashi of Scripps Research, and David Frick of the University of Wisconsin, Milwaukee.