Scientists Replicate Hepatitis C Virus in Laboratory
New In Vitro Model System Will Allow Study of Therapeutics and Virus Life Cycle
For the first time, scientists have replicated hepatitis C virus (HCV) in the
laboratory. The ability to replicate HCV in cell culture will allow researchers
to better study the life cycle and biology of this virus and to test potential
antiviral compounds, which may lead to new therapies for the liver disease that
results from infection with HCV. Scientists at the National Institute of
Diabetes and Digestive and Kidney Diseases (NIDDK), one of the National
Institutes of Health (NIH), conducted the study, which appears in the Feb. 15,
2005 issue of Proceedings of the National Academy of Sciences (PNAS).

“Until recently, research on this infectious disease has suffered from the lack
of a robust in vitro model system,” says T. Jake Liang, M.D., Chief of the Liver
Diseases Branch of the NIDDK and co-author of the study. “Our model system
produced viral particles that have all the properties of the whole virus. This
evidence together with an analysis of the replicated viral RNA supports a
conclusion of viral replication and production.”

The NIDDK group used a strain of HCV that would have applications to the
greatest number of people — genotype 1, the major type of HCV of human
infections worldwide and the type most resistant to current therapies. They
constructed an HCV replica using a DNA copy of the original HCV single-strand
RNA genome. They placed the DNA copy between two ribozymes, RNA molecules that
have enzymatic function and can cleave RNA sequence at specific locations. These
two ribozymes were designed to generate the correct ends of the HCV genome and
to act as start and stop buttons to gene activity. The construct was “naked,”
meaning that it contained only nucleic acids, the genetic material of the virus,
and did not have the HCV viral envelope, a protective shell of lipids and
proteins that surrounds the viral RNA in fully-formed HCV. The naked HCV
construct was then placed into human liver cells in a cell culture medium.

The NIDDK scientists found evidence of HCV proteins and HCV RNA within the human
liver cells in the culture. Electron microscopy showed evidence of high levels
of viral particles resembling fully-formed HCV outside of the human liver cells
in the culture medium. The researchers believe that the HCV construct contained
within the human liver cells behaved like a true HCV infection by producing
fully formed copies of the virus and releasing them from the host cell into the
culture medium. Further testing is needed before the researchers can determine
if the viral particles produced in this system are in fact infectious. Also,
this system only represents the tail end of the viral life cycle — viral
replication, assembly and release from host cells. Another HCV model system is
needed to show the beginning stages of the viral life cycle — viral entry into
host cells and viral activity in the host cell before replication.

“With this cell-based system, we can screen compounds with a cell-based assay to
look for inhibitors of virus replication,” says Liang. “We can also apply this
technique to develop model systems for other similar viruses.”

HCV is a small, enveloped, single-stranded RNA virus in the family Flaviviridae.
HCV is a major cause of liver disease in the United States and the world. One in
a series of hepatitis viruses, HCV accounts for about 15 percent of acute
hepatitis cases, 60 to 70 percent of chronic hepatitis cases, and up to 50
percent of cases of cirrhosis, end-stage liver disease, and liver cancer. Almost
4 million Americans, or 1.8 percent of the U.S. population, have antibodies to
HCV indicating ongoing or previous infection with the virus. Approximately
10,000 to 12,000 deaths each year in the United States are due to HCV.

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Heller, Theo; Jonathan Auerbach; Tarice Williams; Tzivia Rachel Moreen; Allison
Jazwinski; Brian Cruz; Neha Jeurkar; Ronda Sapp; Guangxiang Luo; and T. Jake
Liang. “An in vitro model of hepatitis C virion production.” Proceedings of the
National Academy of Sciences, Vol. 102, No. 7, pp. 2579-2583.
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Sandra Tara Balduf (Ane)

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