Antiviral Activity of β- l -2′,3′-Dideoxy-2′,3′-Didehydro-5-Fluorocytidine in Woodchucks Chronically Infected with Woodchuck Hepatitis Virus

Abstract

The L-nucleoside analog ␤-L-2,3-dideoxy-2,3-didehydro-5-fluorocytidine (␤-L-Fd4C) was first shown to exhibit potent activity against hepatitis B virus (HBV) in tissue culture and then to significantly inhibit viral spread during acute infection in the duck HBV model (F. Le Guerhier et al., Antimicrob. Agents Chemother. 44:111-122, 2000). We have therefore examined its antiviral activity in a mammalian model of chronic HBV infection, the woodchuck chronically infected with woodchuck hepatitis virus (WHV). Side-by-side comparison of ␤-L-Fd4C and lamivudine administered intraperitoneally during short-term and long-term protocols demonstrated a more profound inhibition of viremia in ␤-L-Fd4C-treated groups. Moreover, ␤-L-Fd4C induced a marked inhibition of intrahepatic viral DNA synthesis compared with that induced by lamivudine. Nevertheless, covalently closed circular (CCC) DNA persistence explained the lack of clearance of infected hepatocytes expressing viral antigens and the relapse of WHV replication after drug withdrawal. Liver histology showed a decrease in the inflammatory activity of chronic hepatitis in woodchucks receiving ␤-L-Fd4C. An electron microscopy study showed the absence of ultrastructural changes of hepatic mitochondria, biliary canaliculi, and bile ducts. However, a loss of weight was observed in all animals, whatever the treatment, as was a transient skin pigmentation in all woodchucks during ␤-L-Fd4C treatment. There was no evidence that lamivudine or ␤-L-Fd4C could prevent the development of hepatocellular carcinoma with the protocols used. These results indicate that ␤-L-Fd4C exhibits a more potent antiviral effect than lamivudine in the WHV model but was not able to eradicate CCC DNA and infected cells from the liver at the dosage and with the protocol used.Chronic hepatitis B virus (HBV) infection remains a major public health problem worldwide due to its natural history, which includes the progression to chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma (25). The recent approval of lamivudine [␤-L(Ϫ)-2Ј,3Ј-dideoxy-3Ј-thiacytidine, also called 3TC or L(Ϫ)SddC] has opened new perspectives for the therapy of chronic hepatitis B. In all clinical trials, 3TC was shown to be well tolerated and to be a very potent inhibitor of HBV replication (7, 24). However, due to the long half-life of infected hepatocytes and the persistence of viral covalently closed circular (CCC) DNA in infected hepatocytes, long-term treatment is required to control or eradicate viral infection (29, 34). The spontaneous HBV genome variability gives rise to a progressive selection of drug-resistant variants in 16 to 43% of the patients after 1 year of therapy (7, 24). These mutations located in the conserved B and C domains of the HBV reverse transcriptase confer resistance to 3TC (2, 49).Therefore, the design and evaluation of new molecules with anti-HBV activity remain major goals. With this aim, new antiviral compounds are usually assessed in experimental models of hepadnavirus replication (30), includ...

A newly developed in vitro model of the human epithelial airway barrier to study the toxic potential of nanoparticles

“…The level of inhibition of [ 32 P]-labelled DNA synthesis was assessed by quantifying the incorporation of [α-32 P]dCtP into the nascent viral DNA. Although no exogenous dNtPs were added in the reactions and since there is a pool of endogenous dNtP-tPs in the HuH-7 extracts (as was shown elsewhere with the use of the DHBV RNA in the reticulocyte lysate; le Guerhier et al, 2001), the results showed that the incorporation into the nascent viral DNA was significantly inhibited with the presence of ddNtPs triphosphate (Fig. 8).…”

“…Several animal studies have been performed in the past for the screening of new antiviral molecules that inhibit the hepadnavirus replication, either in vivo with studies involving the sacrifice of the experimental animals (Addison et al, 2002;Cao and tavis, 2006;Cavanaugh et al, 1997;Colledge et al, 1997;Cullen et al, 1997;Fourel et al, 1994;Genovesi et al, 2000;Hafkemeyer et al, 1996;Howe et al, 1996;Lin et al, 1998;Lofgren et al, 1996;Luscombe et al, 1996;Nicoll et al, 2000;Nicoll et al, 1998;Offensperger et al, 1993a;Rahn et al, 1997;Rajagopalan et al, 1996;Seigneres et al, 2000;Severini et al, 1995;Tomita et al, 2000;Urban et al, 2001;Witcher et al, 1997) or in vivo with additional in vitro studies using the rabbit reticulocyte lysate system (Aguesse-Germon et al, 1998;Doong et al, 1991;Guo et al, 2007;Jacquard et al, 2006;Le Guerhier et al, 2001;Offensperger et al, 1993b;Robaczewska et al, 2005;Seifer et al, 1998;Seigneres et al, 2001;Seigneres et al, 2003;Shaw et al, 1996;Zoulim et al, 1996) (Tab. 1).…”

Persistence of cccDNA during the natural history of chronic hepatitis B and decline during adefovir dipivoxil therapy1 ☆

“…Studies performed in animal hepadnavirus models suggested that the long half-life of viral cccDNA is responsible for viral persistence and reactivation following drug cessation 6,32,33 . Here we show, for the first time, that antiviral therapy with a potent HBV polymerase inhibitor significantly reduces cccDNA in CH-B patients.…”

Residual integrated viral DNA after hepadnavirus clearance by nucleoside analog therapy

“…focused primarily on a strategy of inhibiting viral DNA synthesis through the use of nucleoside analogs that are highly specific for the viral DNA polymerase (1)(2)(3)(4)(5)(6)(7)(8)(9). Treatment of patients with such inhibitors can be remarkably effective in reducing viremias to very low levels (2,3).…”