3,7-Dihydroxytropolones Inhibit Initiation of Hepatitis B Virus Minus-Strand DNA Synthesis

Abstract

Initiation of protein-primed (-) strand DNA synthesis in hepatitis B virus (HBV) requires interaction of the viral reverse transcriptase with epsilon (ε), a cis-acting regulatory signal located at the 5’ terminus of pre-genomic RNA (pgRNA), and several host-encoded chaperone proteins. Binding of the viral polymerase (P protein) to ε is necessary for pgRNA encapsidation and synthesis of a short primer covalently attached to its terminal domain. Although we identified small molecules that recognize HBV ε RNA, these failed to inhibit protein-primed DNA synthesis. However, since initiation of HBV (-) strand DNA synthesis occurs within a complex of viral and host components (e.g., Hsp90, DDX3 and APOBEC3G), we considered an alternative therapeutic strategy of allosteric inhibition by disrupting the initiation complex or modifying its topology. To this end, we show here that 3,7-dihydroxytropolones (3,7-dHTs) can inhibit HBV protein-primed DNA synthesis. Since DNA polymerase activity of a ribonuclease (RNase H)-deficient HBV reverse transcriptase that otherwise retains DNA polymerase function is also abrogated, this eliminates direct involvement of RNase (ribonuclease) H activity of HBV reverse transcriptase and supports the notion that the HBV initiation complex might be therapeutically targeted. Modeling studies also provide a rationale for preferential activity of 3,7-dHTs over structurally related α-hydroxytropolones (α-HTs).

Recent Advances in Hepatitis B Treatment

“…Bulky substitution in positions R 1 , R 2 and R 3 leads to a decreased inhibitory activity, indicating that sulfonyl- or lactone substituents can increase the efficacy [ 190 , 191 , 193 ]. These findings have been validated by recent studies, which also highlighted the increased efficacy of amide-substituted α-HTs [ 194 , 195 , 196 ].…”

Synthesis of Polyoxygenated Tropolones and their Antiviral Activity against Hepatitis B Virus and Herpes Simplex Virus‐1

“…In collaboration with Le Grice and co-workers, it was recently found that 3,7-dihydroxytropolones are capable of inhibiting initiation of HBV minus strand DNA synthesis. [23] Unfortunately, cytotoxicity of the compounds precluded observing any antiviral activity of the 3,7-dihydroxytropolones previously tested (e. g., 4 a). Consistent with these prior results, none of the 3,7-dihydroxtropolones tested had selective anti-HBV activity (4 b-g and 15).…”

Biosynthesis of the bacterial antibiotic 3,7-dihydroxytropolone through enzymatic salvaging of catabolic shunt products

“…[14][15][16] Typically, (di) hydroxylated tropolones have strong cytotoxic, antimicrobial and antiviral activities when three adjacent oxygens are present, which allows chelation of up to two metal ions (e.g., Fe 2+ , Zn 2+ , Cu 2+ , Mg 2+ ) and thus enables the (un)competitive inhibition of (bi) metallic enzymes such as the medically relevant inositol monophosphatase, matrix metalloproteases and virulence factors (e.g., anthrax lethal factor or HIV reverse transcriptase). 9 Compound 5, e.g., was shown to inhibit the initiation of DNA synthesis of the hepatitis B virus 17 and is furthermore highly cytotoxic against cultured B16 melanoma cells, thereby substantially extending the life span of mice with B16 melanoma. 15 Recent studies suggest that the cytotoxic effects of tropolones (in multiple myeloma cells) may be due to the alteration of cellular iron availability.…”

“… 9 Compound 5, e.g. , was shown to inhibit the initiation of DNA synthesis of the hepatitis B virus 17 and is furthermore highly cytotoxic against cultured B16 melanoma cells, thereby substantially extending the life span of mice with B16 melanoma. 15 Recent studies suggest that the cytotoxic effects of tropolones (in multiple myeloma cells) may be due to the alteration of cellular iron availability.…”