Researchers at Rockefeller University and Washington University School of Medicine in St. Louis have identified mutations in a protein of certain strains of hepatitis C virus (HCV) that allow these strains to replicate more vigorously in human cell culture. The finding allows scientists to improve an essential tool for studying the virus and suggests a starting point for the design of effective vaccines.
"This is a major breakthrough for genetic studies on the virus and for designing ways to screen for effective drugs," says Rockefeller Professor Charles M. Rice, Ph.D., who led the study. "This could really speed up the drug discovery process." Rice, a prominent figure in HCV research, reported the study in the December 8 issue of Science along with Keril J. Blight, Ph.D., and Alexander A. Kolykhalov, Ph.D., both of Washington University School of Medicine.
Like all viruses, HCV cannot replicate by itself; instead it takes over the machinery of the host cell to make copies of itself. Much about the life cycle of HCV remains poorly understood, but researchers think that in humans the virus enters a liver cell and delivers RNA and its proteins into the cell cytoplasm. The RNA is separated from the protein, copied and then joined with new protein components before being released from the liver cell to infect a large number of new cells.
In order to clarify this process and conduct studies of drugs and vaccines, researchers need a method for growing the virus reliably in cell culture. In 1999, German scientists developed a new HCV culture system, but it was limited in how efficiently the viral RNA was able to start replicating in the host cells. The new culture system developed by Rice and his colleagues greatly improves on the method, producing a 10,000-fold increase in the number of cells suitable for study.
Researchers know that hepatitis C has genetic variations that result in different structures of the viral proteins, but they do not understand yet how these variations determine the virus's effect on the liver cells of the person carrying it. The greatly increased availability of HCV-infected cells will help scientists such as Rice conduct genetic analyses of the virus in the search for ways to slow or halt it.
The robust HCV strains tend to have adaptive mutations in a specific region of a protein called NS5A. "When mutations in one area greatly affect the virusís resistance and robustness, it suggests that NS5A plays an essential role in HCV replication in cell cultures," Rice says.
The availability of more cells in which the virus replicates reliably should speed the search for effective drugs against HCV through a process called "high-throughput screening," a technique using miniaturized, automated technology to test a large number of compounds rapidly. The identification of a protein region critical to replication gives researchers a more precise target toward which they can direct the compounds.
The research was conducted at Washington University School of Medicine before Rice joined the Rockefeller faculty in September 2000. Currently, Rice is the Maurice R. and Corinne P. Greenberg Professor and head of the Laboratory of Virology and Infectious Disease at Rockefeller. He also is the scientific and executive director of the Center for the Study of Hepatitis C, a newly established collaboration among Rockefeller University, NewYork-Presbyterian Hospital, and Weill Medical College of Cornell University. (Rice also still has an appointment at Washington University School of Medicine as professor of molecular microbiology.)
Some four million people in the United States are infected with HCV, and about 30,000 new acute infections occur every year. HCV is responsible for 8,000 to 10,000 deaths per year in the United States. Liver failure due to hepatitis C is the leading cause of liver transplants in the United States, and about 25 percent of liver cancer cases in the country are associated with HCV. Although about 85 percent of those who are infected develop chronic infection, the virus usually remains undetected for years, or even decades, until it causes advanced liver disease.
The research reported in Science was funded in part by grants from the National Institutes of Health and the Greenberg Medical Foundation.