Intracellular Metabolism and In Vitro Activity of Tenofovir against Hepatitis B Virus

Abstract

Tenofovir is an acyclic nucleotide analog with activity against human immunodeficiency virus (HIV) and hepatitis B virus (HBV). Tenofovir disoproxil fumarate (tenofovir DF), a bis-alkoxyester prodrug of tenofovir, is approved for the treatment of HIV and is currently being developed to treat chronic hepatitis B. In this report, we further characterize the in vitro activity of tenofovir against HBV as well as its metabolism in hepatic cells. We show that tenofovir is efficiently phosphorylated to tenofovir diphosphate (TFV-DP) in both HepG2 cells and primary human hepatocytes. TFV-DP has a long intracellular half-life (95 h) and is a potent and competitive inhibitor of HBV polymerase (K i ‫؍‬ 0.18 M). Tenofovir has a 50% effective concentration of 1.1 M against HBV in cell-based assays, and potency is improved >50-fold by the addition of bis-isoproxil progroups. Tenofovir has previously demonstrated full activity against lamivudine-resistant HBV in vitro and clinically. Here we show that the major adefovir resistance mutation, rtN236T, confers three-to fourfoldreduced susceptibility to tenofovir in cell culture; the clinical significance of this susceptibility shift has not yet been determined. The rtA194T HBV polymerase mutation recently identified in tenofovir DF-treated HIV/ HBV-coinfected patients did not confer in vitro resistance to tenofovir as a single mutation or in a lamivudineresistant viral background. Overall, the antiviral and metabolic profile of tenofovir supports its development for the treatment of chronic hepatitis B.

Hepatitis B virus mutations potentially conferring adefovir/tenofovir resistance in treatment-naive patients

“…rtL180M and rtM204V, has been observed in individuals who are co-infected with HBV and HIV-1 [14,15] . However, the analysis reported by other authors has not provided a clear association between rtA194T and viral load rebound [4] . Thus, the potential impact of this mutation on tenofovir susceptibility deserves further study.…”

“…While each nucleotide or nucleoside produces efficient viral suppression, they induce only modest rates of HBV surface antigen (HBsAg) seroconversion and require long-term administration to control disease in patients. The need for long-term therapy necessitates drug safety and the ability to delay or manage the emergence of resistant HBV strains [3][4][5] . The clinical benefit of these therapies has been compromised by the emergence of resistant viral strains that carry specific mutations in the polymerase gene [6,7] .…”

Antiviral drug-resistant HBV

“…Aminotransferase levels remained normal in both patients and results of phenotypic studies are conflicting. 59 Data on TDF resistance in patients who received tenofovir monotherapy are not available because tenofovir has been used predominantly in patients with HIV/HBV coinfection, in combination with lamivudine or emtricitabine.…”

“…Furthermore, in the latter patients, a rapid reemergence of LAM-resistant mutations has been observed on reintroduction of lamivudine. 59,60 Case studies have reported that patients with primary nonresponse to adefovir experience further viral suppression when treatment is switched to tenofovir 69 presumably due to the higher dose of tenofovir used in clinical practice. For the same reason, tenofovir may result in some degree of viral suppression in patients with ADV-resistant HBV but the efficacy is likely limited due to in vitro evidence of cross resistance.…”

Low-dose tenofovir is more potent than adefovir and is effective in controlling HBV viremia in chronic HBeAg-negative hepatitis B

“…TDF is also active against hepatitis B virus (HBV) and is equipotent to adefovir in vitro [1,2] , but because of its lower nephrotoxicity, it can be used at higher dosage (e.g. 300 mg/d) and is more active than its parent compound against HBV in vivo [3][4][5] .…”