Inhibitory Effects of Acyclic Nucleoside Phosphonate Analogues on Hepatitis B Virus DNA Synthesis in HB611 Cells

Abstract

SummaryBy using an assay system based on a human hepatoblastoma cell line (HB611) that continuously synthesizes hepatitis B virus (HBV) DNA, 56 acyclic nucleoside phosphonate analogues were examined for their inhibitory effects on HBV DNA synthesis. The following compounds were found to inhibit HBV DNA synthesis at concentrations that were significantly lower than their minimum cytotoxic concentrations; 9-(2-phosphonylmethoxyethyl)adenine (PMEA), 9-(2-phosphonylmethoxyethyl) guanine(PMEG), 9-(2-phosphonylmethoxyethyl) guanine ethyl ester (PMEGEE), 9 -(2 -phosphonylmethoxyethyQ -1 -deazaadenine (PMEC 1A), 9-(2-phosphonylmethoxyethyl)-2,6-diaminopurine (PMEDAP), (S)-9-(3-hydroxy-2-phosphonylmethoxypropyl)adenine (HPMPA), 9-(3-isopropoxy-2-phosphonylmethoxypropyl)adenine (IPPMPA), 9-(RS)-(2-phosphonylmethoxypropyl)adenine (PMPA) and 9-(3-hydroxy-2-phosphonylmethoxypropyl)-2, 6-diaminopurine (HPMPDAP). The most selective compounds (with indexes greater than 100) were PMEDAP, PMEA, IPPMPA, and PMPA. Acyclic pyrimidine nucleoside phosphonate analogues did not prove markedly selective as anti-HBV agents. Diphosphoryl derivatives of some acyclic purine nucleoside phosphonates (i.e. PMEA, PMEDAP, HPMPA) were prepared. They proved inhibitory to HBV DNA polymerase but not cellular DNA polymerase ct.

Colorimetric Assay System for Screening Antiviral Compounds against Hepatitis B Virus

“…Yokota et al (17,18) was also similar to previously reported data (18). However, the EC50 values that we obtained for PMEG and PMEGEE were much lower than those obtained by others, and the EC50 value of PMPA was higher than that of the Southern blot analysis (18).…”

“…However, the EC50 values that we obtained for PMEG and PMEGEE were much lower than those obtained by others, and the EC50 value of PMPA was higher than that of the Southern blot analysis (18). The selective indexes of (S)-HPMPC and araA estimated by this assay system were higher than previous results (17,18). However, these differences should pose no problem in the screening of anti-HBV compounds.…”

“…This colorimetric method allows for the quantitative detection of HBV DNA and can be partially automated. The use of HB611 cells for analysis of the anti-HBV compounds has been reported previously (4,8,14,17,18). However, we have developed a rapid and accurate colorimetric assay system for anti-HBV agents using HB611 cells, immunoaffinity purification, PCR, and hybrid-capture detection.…”

“…However, human hepatoma cell lines transfected with HBV DNA have also been developed (HB611 and HepG2 2.2.15 cells (11,13)) and used for the screening of antiviral drugs (3,4,8,14,17,18). In these studies, drug efficacy was determined by measuring HBV DNA levels using Southern blot analyses with 9)-labeled HBV DNA as the probe.…”

Comparative Anti-Retrovirus and Anti-Hepadnavirus Activity of Three Different Classes of Nucleoside Phosphonate Derivatives

“…Moreover, PMEA showed potent and selective anti-retrovirus activity in several animal models including MSV infection in newborn NMRI mice (Balzarini et al, 1989(Balzarini et al, , 1991, FIV infection in cats (Hartmann et al, 1993) and SIV infection in rhesus monkeys (Egberink et al, 1990;Tsai et al, 1994). Interestingly, PMEA has also shown activity against human and duck hepatitis B virus in human HB611 and Hep G2 2.2.15 cells and in primary duck hepatocyte cultures at 0.2 and 1.2 J..lM, respectively (Heijtink et al, 1993;Yokota et al, 1994). PMEA has recently entered clinical Phase 1/11 trials in AIDS patients.…”