Fox Chase Cancer Center

Avid Therapeutics, Inc.

Christoph Seeger

Ju-Tao Guo

Robert W King

Qing Zhu

Christopher S Barker

Cells and cell lines which replicate HCV of non-hepatic human and non human origin are disclosed. Also provided are methods of using such cells and cell lines to identify anti-HCV agents for the treatment of HCV infection.

Replication of hepatitis C virus in non-hepatic epithelial and mouse hepatic cells

Novel cell lines and methods are provided for inducibly expressing the Hepatitis B virus (HBV) genome within cultured cells. Cells are stably transformed with an HBV genome under control of an operator/promoter target sequence which can activate expression of the HBV genome when bound by a cognate transactivator protein. The cell lines are also stably transformed with a gene that encodes and expresses a suitable transactivator protein. Cells may be maintained under non-inducing conditions whereby the transactivator protein is unable to interact with the operator/promoter target sequence, so the HBV genome is not expressed. Under appropriate inducing conditions, the transactivator protein is enabled to interact with the operator/promoter target sequence, thereby activating expression of the HBV genome. The inducible system provides an effective and rapid cell-based assay to screen compounds for anti-hepadnaviral activity.

Cultured cell line that inducibly expresses the hepatitis B virus

An enzymatically active composition of matter, comprising an extracellular combination of a substantially virus-free hepadnavirus polymerase gene product and an RNA template for hepadnavirus minus strand DNA synthesis, is provided. The composition possesses a DNA priming activity and a reverse transcriptase activity. The hepadnavirus polymerase gene product is produced by in vitro expression of a hepadnavirus pol gene. The enzymatically active composition of matter is used in rapid in vitro assays to screen potential anti-hepadnaviral agents. Assays for various functions of the hepadnavirus DNA polymerase are provided, including: (1) binding of the polymerase gene product to hepadnavirus pre-genomic RNA; (2) DNA priming activity of the polymerase gene product; and (3) reverse transcriptase activity of the polymerase gene product.

Hepadnavirus polymerase gene product having RNA-dependent DNA priming

An enzymatically active, substantially virus-free hepadnavirus polymerase gene product is provided, which possesses a DNA priming function and reverse transcriptase activity, both of which are dependent on the presence of an RNA template and magnesium ions. The hepadnavirus polymerase gene product is produced by in vitro transcription and translation of a hepadnavirus pol gene. The enzymatically active hepadnavirus polymerase gene product is used in rapid in vitro assays to screen potential anti-hepadnaviral agents. An assay for the DNA priming activity of the gene product is provided, wherein covalent attachment of labelled dGTP to the gene product is quantitatively measured. An assay for the reverse transcriptase activity of the gene product is provided, wherein incorporation of nucleotide triphosphates into the nascent DNA minus strand is measured, taking advantage of the covalent attachment of said nascent strand to the polymerase gene product.

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Melrose Park, PA, United States of America

Christoph Seeger