KNK437 Inhibits Replication and Transcription of the Hepatitis B Virus.

During replication of the hepatitis B virus (HBV) in liver cells, the reverse transcription of pre-genomic RNA (pgRNA) is initiated by protein priming at an RNA packaging signal ε located near the 5' end of pgRNA. Heat-shock proteins (Hsps) such as Hsc70, Hsp40, and Hsp90 have been reported to be involved in the reconstitution of HBV polymerase (P protein) and E. The P - E complex initiates the reverse transcription and assembly of nucleocapsids. Hence, blockade of P - ε interactions is an attractive target for drug intervention. We explored the influence of the Hsp inhibitor KNK437 on replication and transcription of the HBV. Three working models were applied: HepG2. 2. 15 cell line; Huh7 cells transfected transiently with the 1. 05 X HBV (pCH9-3091) plasmid; Huh7 cells transfected transiently with the 1. 3 X HBV (pGEM-1. 3 X HBV) plasmid. Cytotoxic effects of KNK437 were detected by the CCK-8 method. Levels of hepatitis B surface antigen (HBsAg) and hepatitis B viral protein (HBeAg) in the media secreted from cells were measured using an ELISA. Intracellular HBV DNAs within nucleocapsids were measured by quantitative polymerase chain reaction (qPCR), and intracellular HBV RNAs by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Transcription of Hsps in cells was determined by qRT-PCR. Data suggested that KNK437 reduced the extracellular secretion of HBsAg and HBeAg in most cases; it downregulated expression of intracellular HBV DNAs within nucleocapsids and RNA transcripts. The lowest rate of viral DNAs in KNK437-treated hepatocytes for all experimental groups was ~1. 5%o (control, 100%), whereas that for RNAs was ~30%. Western blotting revealed KNK437 to inhibit intracellular core expression in HepG2. 2. 15. As a general inhibitor, KNK437 suppressed transcription of hsp70, hsp90b, and hsp4o. These data suggest that KNK437 may be a potent anti-HBV inhibitor for future therapy against chronic hepatitis.