Identification of disubstituted sulfonamide compounds as specific inhibitors of hepatitis B virus covalently closed circular DNA formation.
Hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) plays a central role in viral infection and persistence and is the basis for viral rebound after the cessation of therapy, as well as the elusiveness of a cure even after extended treatment. Therefore, there is an urgent need for the development of novel therapeutic agents that directly target cccDNA formation and maintenance. By employing an innovative cell-based cccDNA assay in which secreted HBV e antigen is a cccDNA-dependent surrogate, we screened an in-house small-molecule library consisting of 85,000 drug-like compounds. Two structurally related disubstituted sulfonamides (DSS), termed CCC-0975 and CCC-0346, emerged and were confirmed as inhibitors of cccDNA production, with low micromolar 50% effective concentrations (EC(50)s) in cell culture. Further mechanistic studies demonstrated that DSS compound treatment neither directly inhibited HBV DNA replication in cell culture nor reduced viral polymerase activity in the in vitro endogenous polymerase assay but synchronously reduced the levels of HBV cccDNA and its putative precursor, deproteinized relaxed circular DNA (DP-rcDNA). However, DSS compounds did not promote the intracellular decay of HBV DP-rcDNA and cccDNA, suggesting that the compounds interfere primarily with rcDNA conversion into cccDNA. In addition, we demonstrated that CCC-0975 was able to reduce cccDNA biosynthesis in duck HBV-infected primary duck hepatocytes. This is the first attempt, to our knowledge, to identify small molecules that target cccDNA formation, and DSS compounds thus potentially serve as proof-of-concept drug candidates for development into therapeutics to eliminate cccDNA from chronic HBV infection.
Institute for Biotechnology and Virology Research, Department of Microbiology and Immunology, Drexel University College of Medicine, Doylestown, Pennsylvania, USA.
Dawei
D
Cai
Courtney
C
Mills
Wenquan
W
Yu
Ran
R
Yan
Carol E
CE
Aldrich
Jeffry R
JR
Saputelli
William S
WS
Mason
Xiaodong
X
Xu
Ju-Tao
JT
Guo
Timothy M
TM
Block
Andrea
A
Cuconati
Haitao
H
Guo
D000081:Acetamides / Q000494:pharmacology*; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D001549:Benzamides / Q000494:pharmacology*; D002460:Cell Line; D004261:DNA Replication / Q000187:drug effects; D004270:DNA, Circular / Q000378:metabolism*; D004279:DNA, Viral / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D004372:Ducks; D056945:Hep G2 Cells; D015611:Hepatitis B Virus, Duck / Q000187:drug effects / Q000502:physiology; D006513:Hepatitis B e Antigens / Q000378:metabolism; D006515:Hepatitis B virus / Q000187:drug effects* / Q000502:physiology; D019694:Hepatitis B, Chronic / Q000188:drug therapy / Q000821:virology; D022781:Hepatocytes / Q000821:virology; D006801:Humans; D008826:Microbial Sensitivity Tests; D011725:Pyridines / Q000494:pharmacology*; D013449:Sulfonamides / Q000494:pharmacology*; D013844:Thiazoles / Q000494:pharmacology*; D014779:Virus Replication / Q000235:genetics
C578163:2-(benzenesulfonyl-(2-chloro-5-trifluoromethyl-phenyl)-amino)-N-pyridin-4-ylmethyl-acetamide; C578164:4-(benzyl-methyl-sulfamoyl)-N-(2-methyl-benzothiazol-5-yl)-benzamide; D000081:Acetamides; D000998:Antiviral Agents; D001549:Benzamides; D004270:DNA, Circular; D004279:DNA, Viral; D006513:Hepatitis B e Antigens; D011725:Pyridines; D013449:Sulfonamides; D013844:Thiazoles; D004259:DNA-Directed DNA Polymerase