Methods for producing cells having a phenotype of a primary human hepatocytes and compositions
AssigneeThe Governors of the University of Alberta
InventorsD. Lorne Tyrrell, Hendrikje Geesje Steenbergen, Michael A. Joyce
Life of this patent
Estimating the term…
Abstract
The present disclosure provides methods and compositions relating to in vitro cultures of human hepatocyte cell lines which exhibit a primary human hepatocyte phenotype. Such cell lines are susceptible to infection by a hepatotrophic virus, such as HCV or HBV, and support both viral replication and high levels of viral particle production. Such in vitro cultures find use in production and study of hepatotrophic virus, as well as methods of screening (e.g., for antiviral drugs, assessing drug metabolism), and study of primary human hepatocytes.
Claims25
A method of producing a cell culture comprising cells having a primary human hepatocyte phenotype, the method comprising:
- culturing a human hepatocellular carcinoma (hHCC) cell line in a culture medium comprising human serum for more than 11 days,
- wherein said culturing is without subculturing after 10 days of culturing in the culture medium comprising human serum,
- wherein said culturing does not comprise transforming the hHCC cell line, and
- wherein said culturing induces differentiation of the hHCC cell line into a cell having a primary human hepatocyte phenotype.
A method of producing viral particles, the method comprising:
- incubating a cell culture comprising a human hepatocellular carcinoma (hHCC) cell line in a culture medium comprising human serum for more than 11 days,
- wherein said incubating is without subculturing after 10 days of culturing in the culture medium comprising human serum,
- wherein said incubating does not comprise transforming the hHCC cell line,
- wherein said incubating induces differentiation of the hHCC cell line into a cell having a primary human hepatocyte phenotype;
- introducing a genome of a hepatotrophic virus into the cell having primary human hepatocyte phenotype;
- and
- maintaining the cell culture under conditions suitable for production of viral particles.
A method for screening a candidate agent for antiviral activity, the method comprising:
- incubating a cell culture comprising a human hepatocellular carcinoma (hHCC) cell line in a culture medium comprising human serum for more than 11 days, wherein said incubating is without subculturing after 10 days of culturing in the culture medium comprising human serum wherein said incubating does not comprise transforming the hHCC cell line, wherein said incubating induces differentiation of the hHCC cell line into a cell having a primary human hepatocyte phenotype;
- introducing a genome of a hepatotrophic virus into the cell having primary human hepatocyte phenotype;
- contacting the cell culture with a candidate antiviral agent;
- maintaining the cell culture under conditions suitable for viral replication;
- and
- detecting the presence or absence of an effect of the candidate agent upon viral replication;
- wherein a decrease in viral particle production in the presence of the candidate agent as compared to the absence of the candidate agent indicates the candidate agent have antiviral activity.
A method for screening a sample suspected of containing an antibody for antiviral activity, the method comprising:
- incubating a cell culture comprising a human hepatocellular carcinoma (hHCC) cell line in a culture medium comprising human serum for more than 11 days, wherein said incubating is without subculturing after 10 days of culturing in the culture medium comprising human serum wherein said incubating does not comprise transforming the hHCC cell line, wherein said incubating induces differentiation of the hHCC cell line into a cell having a primary human hepatocyte phenotype;
- introducing a genome of a hepatotrophic virus into the cell having primary human hepatocyte phenotype;
- contacting the cell culture with a sample of suspected of containing an antibody;
- maintaining the cell culture under conditions suitable for viral replication;
- and
- detecting the presence or absence of an effect of the sample upon viral replication;
- wherein a decrease in viral particle production in the presence of the sample as compared to the absence of the sample indicates the sample contains an antibody having antiviral activity.
Dependencies are inferred from the claim text.