Liver Hepatitis B Virus DNA Levels Research Articles

The cyclophilin inhibitor CRV431 inhibits liver HBV DNA and HBsAg in transgenic mice.

Hepatitis B virus (HBV) infection is a major health burden worldwide with 240 million chronically infected individuals. Nucleos(t)ide analogs and interferons are the current standards of care due to their suppression of HBV replication, but the treatments rarely eradicate HBV from individuals. Similar to current treatments for human immunodeficiency virus type-1 (HIV-1) and hepatitis C virus (HCV) patients, improved HBV therapies will require the combination of multiple drugs which target distinct steps of the HBV life cycle. In this study, we tested the potential of a cyclophilin inhibitor, CRV431, to affect HBV replication in transgenic mice. We found that oral treatment with CRV431 (50 mg/kg/day) for a period of 16 days significantly reduced liver HBV DNA levels and moderately decreased serum HBsAg levels. We observed an additive inhibitory effect on liver HBV DNA levels in mice treated with a combination of low doses of CRV431 (10 mg/kg/day) and the nucleotide prodrug, tenofovir exalidex (TXL), (5 mg/kg/day). No toxicity was observed in CRV431-treated mice. Although it is well known that CRV431 neutralizes the peptidyl-prolyl isomerase activity of cyclophilins, its anti-HBV mechanism(s) of action remains unknown. Nevertheless, this study provides the first demonstration of a beneficial effect of a cyclophilin inhibitor in vivo in an HBV transgenic mouse model. Altogether our data reveal the potential of CRV431 to be part of improved new therapies for HBV patients.

Serum miR-181b Is Correlated with Hepatitis B Virus Replication and Disease Progression in Chronic Hepatitis B Patients.

Previously, we reported that microRNA-181b (miR-181b) activates hepatic stellate cells partly through the phosphatase and tensin homolog deleted on chromosome 10 (PTEN)/Akt pathway. The main objective of this study was to ascertain whether serum miR-181b expression is correlated with that of liver hepatitis B virus (HBV) DNA and disease progression in chronic hepatitis B (CHB) patients. Serum miR-181b and liver HBV DNA levels were quantified in 64 CHB patients with real-time PCR. Liver fibrosis and necroinflammation were graded according to the Ishak scoring system. Serum miR-181b levels were evaluated in the CHB group, compared with healthy controls. Expression in patients with HBsAg (+) was higher than that in patients with HBsAg (-). Notably, serum miR-181b and liver HBV DNA levels were significantly correlated (P<0.05). Serum miR-181 levels were higher in patients with serum HBV DNA>10(3) IU/ml (P=0.017), histologic activity index (HAI) >8 (P=0.001) and fibrosis score >4 (P<0.0001). Liver HBV DNA levels were higher in patients with abnormal alanine aminotransferase (ALT) values (P=0.004), serum HBV DNA levels>10(3) IU/ml (P=0.034) and fibrosis score >4 (P=0.006). Using multivariate logistic regression analysis, serum miR-181b was identified as an independent predictor of disease progression (OR 4.172, 95% CI 1.838-9.473, P=0.009 for HAI >8; OR 5.387, 95% CI 2.067-14.036, P=0.001 for fibrosis score >4). Serum miR-181b is correlated with liver and serum HBV DNA levels as well as disease progression in CHB.

Establishment of drug-resistant HBV small-animal models by hydrodynamic injection

In antiviral therapy of hepatitis B virus (HBV) infection, drug resistance remains a huge obstacle to the long-term effectiveness of nucleoside/tide analogs (NAs). Primary resistance mutation (rtM204V) contributes to lamivudine (LAM)-resistance, and compensatory mutations (rtL180M and rtV173L) restore viral fitness and increase replication efficiency. The evaluation of new anti-viral agents against drug-resistant HBV is limited by the lack of available small-animal models. We established LAM-resistance HBV replication mice models based on clinical LAM-resistant HBV mutants. Double (rtM204V+rtL180M) or triple (rtM204V+rtL180M+rtV173L) lamivudine-resistant mutations were introduced into HBV expression vector, followed by hydrodynamic injection into tail vein of NOD/SCID mice. Viremia was detected on days 5, 9, 13 and 17 and liver HBV DNA was detected on day 17 after injection. The serum and liver HBV DNA levels in LAM-resistant model carrying triple mutations are the highest among the models. Two NAs, LAM and entecavir (ETV), were used to test the availability of the models. LAM and ETV inhibited viral replication on wild-type model. LAM was no longer effective on LAM-resistant models, but ETV retains a strong activity. Therefore, these models can be used to evaluate anti-viral agents against lamivudine-resistance, affording new opportunities to establish other drug-resistant HBV small-animal models.

Interleukin‐15 suppresses hepatitis B virus replication via IFN‐β production in a C57BL/6 mouse model

Interleukin-15 (IL-15) is a pleiotropic cytokine known to modulate both innate and adaptive immunity. It is suggested that IL-15 may play an important role in the regulation of immune response to hepatitis B virus (HBV). We investigated whether IL-15 could modulate the immune response to HBV. A mouse model for HBV tolerance was established by hydrodynamical injection of pAAV/HBV1.2 plasmid into C57BL/6 mice. This HBV-carrier mouse was simultaneously hydrodynamically injected with either an IL-15-expression plasmid pLIVE-IL-15 or a mock control vector pLIVE-EGFP. The serum levels of HBsAg and HBeAg were measured by radioimmunoassay. Hydrodynamic injection of the plasmid pLIVE-IL-15 resulted in sustained high level of IL-15 in mouse serum, along with the markedly decreased serum HBsAg and HBeAg titres and liver HBV DNA levels. IL-15 also induced anti-HBV activity in T cell- and B cell-deficient Rag1(-/-) mice. Interestingly, despite an increase in NK cell numbers in both spleen and liver of IL-15 treated mice, the anti-HBV effect of IL-15 was neither dependent on presence of NK cells nor on production of IFN-γ. Furthermore, IL-15 could exert anti-HBV function independent of the common IL-2γ(c) R. Lastly, we found that IFN-β expression in the liver and serum was significantly up-regulated by liver expression of IL-15, and blockade of IFN-β function abrogated the anti-HBV activity of IL-15. Liver over-expression of IL-15 may suppress HBV replication in an IFN-β-dependent manner.

Transmissibility of Hepatitis B Virus (HBV) Infection through Blood Transfusion from Blood Donors with Occult HBV Infection

Studies of the transmissibility of hepatitis B virus (HBV) in occult hepatitis B (OHB) through blood transfusion are scarce. We aimed to determine the transmissibility of HBV in blood donors with OHB through transfusion in animal and human studies. Among 217,595 blood donors, 67 donors with OHB were identified. Four chimeric mice populated with human hepatocytes were inoculated with 2 donor serum samples. Serial serum and liver HBV DNA levels were measured. Forty-nine recipients of blood transfusions traced from 10 donors with OHB (9 of whom were positive for antibody to hepatitis B surface antigen [anti-HBs]) were tested for HBV infection. Homology and phylogenetic analyses between the HBV genomic sequences of donors and recipients were performed. Serum HBV DNA was detectable (10(4) copies/mL) in 1 mouse at weeks 5 and 7 after inoculation. Total HBV DNA and HBV replication template (covalently closed circular DNA) and hepatitis B core antigen were detected in the mouse liver. After transfusion, 45 recipients (91.8%) had no HBV infection (ie, they tested negative for hepatitis B surface antigen and HBV DNA). Four tested positive for HBV DNA. In 3 recipients, 83%-86% homology and distant phylogenetic relatedness with their donor HBV excluded transmission through transfusion. The remaining recipient HBV had 95% sequence homology with her donor HBV, compatible with acquisition of HBV infection from the transfusion. High anti-HBs levels in 7 other recipients suggested recent transfusion-related HBV immune response. OHB donor blood infectivity was shown in our animal and human studies. However, the risk of HBV transmission in humans was low, especially from blood products obtained from donors with OHB who were anti-HBs positive.

Alkoxyalkyl Esters of 9-( S )-(3-Hydroxy-2-Phosphonomethoxypropyl) Adenine Are Potent and Selective Inhibitors of Hepatitis B Virus (HBV) Replication In Vitro and in HBV Transgenic Mice In Vivo

Alkoxyalkyl esters of acyclic nucleoside phosphonates have previously been shown to have increased antiviral activity when they are administered orally in animal models of viral diseases, including lethal infections with vaccinia virus, cowpox virus, ectromelia virus, murine cytomegalovirus, and adenovirus. 9-(S)-(3-Hydroxy-2-phosphonomethoxypropyl)adenine [(S)-HPMPA] was previously shown to have activity against hepatitis B virus (HBV) in vitro. To assess the effect of alkoxyalkyl esterification of (S)-HPMPA, we prepared the hexadecyloxypropyl (HDP), 15-methyl-hexadecyloxypropyl (15M-HDP), and octadecyloxyethyl (ODE) esters and compared their activities with the activity of adefovir dipivoxil in vitro and in vivo. Alkoxyalkyl esters of (S)-HPMPA were 6 to 20 times more active than unmodified (S)-HPMPA on the basis of their 50% effective concentrations in 2.2.15 cells. The increased antiviral activity appeared to be due in part to the increased uptake and conversion of HDP-(S)-HPMPA to HPMPA diphosphate observed in HepG2 cells in vitro. HDP-(S)-HPMPA retained full activity against HBV mutants resistant to lamivudine (L180M, M204V), but cross-resistance to a mutant resistant to adefovir (N236T) was detected. HDP-(S)-HPMPA is orally bioavailable and provides excellent liver exposure to the drug. Oral treatment of HBV transgenic mice with HDP-(S)-HPMPA, 15M-HDP-(S)-HPMPA, and ODE-(S)-HPMPA for 14 days reduced liver HBV DNA levels by roughly 1.5 log units, a response equivalent to that of adefovir dipivoxil.

Comparative Study of Anti-hepatitis B Virus RNA Interference by Double-stranded Adeno-associated Virus Serotypes 7, 8, and 9

Using a hepatitis B virus (HBV) transgenic mouse model, we previously showed that a single dose of double-stranded adeno-associated virus (dsAAV) vector serotype 8 carrying a small hairpin RNA (shRNA) effectively reduces HBV replication and gene expression, but the effect gradually decreases with time. In this report, we compared the anti-HBV RNA interference (RNAi) effect of dsAAV8 with those of dsAAV7 and dsAAV9, two other hepatotropic AAV vectors, and examined whether the sequential use of these heterologous AAV vectors could prolong the anti-HBV effect. Our results showed that shRNA delivered by each of the three dsAAV vectors profoundly reduced the serum HBV titer and liver HBV mRNA and DNA levels in the transgenic mice for up to 22 weeks, with dsAAV8 having the greatest inhibitory effect, followed by dsAAV9 and dsAAV7. The potency of dsAAV8 correlated with the presence of higher levels of vector DNA and anti-HBV shRNA in the liver. An in vivo cross-administration experiment showed that preexisting anti-AAV8 antibody completely blocked the anti-HBV RNAi effect of dsAAV8, but had no effect on the potency of dsAAV7 and dsAAV9. Moreover, we demonstrated that a longer anti-HBV effect could be achieved by the sequential use of dsAAV8 and dsAAV9. These results indicate that effective and persistent HBV suppression might be achieved by a combination of the power of RNAi silencing effect and multiple treatments with different AAV serotypes.Molecular Therapy (2009) 17 2, 352-359 doi:10.1038/mt.2008.245.

Quantification of intrahepatic total hepatitis B virus DNA in chronic hepatitis B patients and its relationship with liver histology

Aims:This study was conducted to evaluate the relationship between total intrahepatic hepatitis B virus (HBV) DNA levels and liver histology in terms of the degree of histological activity index (HAI)...

P.090 Entecavir (ETV) treatment through 96 weeks results in substantial virologic and biochemical improvement and HBeAg seroconversion in HBeAg(+) chronic hepatitis B (CHB) patients (study ETV-022)

In antiviral therapy of hepatitis B virus (HBV) infection, drug resistance remains a huge obstacle to the long-term effectiveness of nucleoside/tide analogs (NAs). Primary resistance mutation (rtM204V) contributes to lamivudine (LAM)-resistance, and compensatory mutations (rtL180M and rtV173L) restore viral fitness and increase replication efficiency. The evaluation of new anti-viral agents against drug-resistant HBV is limited by the lack of available small-animal models. We established LAM-resistance HBV replication mice models based on clinical LAM-resistant HBV mutants. Double (rtM204V+rtL180M) or triple (rtM204V+rtL180M+rtV173L) lamivudine-resistant mutations were introduced into HBV expression vector, followed by hydrodynamic injection into tail vein of NOD/SCID mice. Viremia was detected on days 5, 9, 13 and 17 and liver HBV DNA was detected on day 17 after injection. The serum and liver HBV DNA levels in LAM-resistant model carrying triple mutations are the highest among the models. Two NAs, LAM and entecavir (ETV), were used to test the availability of the models. LAM and ETV inhibited viral replication on wild-type model. LAM was no longer effective on LAM-resistant models, but ETV retains a strong activity. Therefore, these models can be used to evaluate anti-viral agents against lamivudine-resistance, affording new opportunities to establish other drug-resistant HBV small-animal models.

Intrahepatic Hepatitis B Virus Covalently Closed Circular DNA Can Be a Predictor of Sustained Response to Therapy

This study aimed to determine whether intrahepatic hepatitis B virus (HBV) covalently closed circular (ccc) DNA and total HBV DNA levels at the end of therapy would predict sustained response to therapy. Hepatitis B e antigen (HBeAg)-positive chronic hepatitis B patients receiving either lamivudine monotherapy or combination of peginterferon and lamivudine had liver biopsy at the end of 1 year therapy and were followed for 52 more weeks after cessation of therapy. Serum HBV DNA, intrahepatic HBV ccc DNA, and total HBV DNA levels were determined. Forty-seven patients, including 34 males and 13 females, were studied. Twenty-seven patients received combination therapy, and 20 patients received lamivudine monotherapy. Twenty-nine patients had end-of-treatment virologic response, and 15 patients had sustained response 52 weeks after therapy. At the end of treatment, log serum HBV DNA levels correlated well with log intrahepatic HBV cccDNA and log intrahepatic total HBV DNA levels. Log intrahepatic cccDNA and log intrahepatic total DNA levels were significantly lower among patients with sustained virologic response. The adjusted odds ratio for log cccDNA was 5.3 (95% CI: 1.5-18.2, P = .009) and, for log intrahepatic HBV DNA, was 4.4 (95% CI: 1.3-14.7, P = .015) to predict sustained virologic response. Using log cccDNA at -0.80 copies/genome equivalent as cutoff, the sensitivity, specificity, and positive and negative predictive values and accuracy of predicting sustained virologic response were 73%, 78%, 56%, 86%, and 77% respectively. Intrahepatic HBV cccDNA and intrahepatic total HBV DNA levels at the end of therapy are superior to serum HBV DNA as surrogates of sustained virologic response.

Occult HBV infection—both hidden and mysterious

Occult HBV infection—both hidden and mysterious