Occult hepatitis B virus (HBV) infection is defined as the presence of HBV DNA in the liver, with or without detectable HBV DNA in the serum of individuals tested negative for hepatitis B surface antigen (HBsAg). As both HBV and hepatitis C virus (HCV) share similar modes of transmission, HBV and HCV coinfection is a well-known clinical entity. Likewise, a high prevalence of occult HBV infection, ranging from 11% to 95%, has been reported in patients with chronic hepatitis C (CHC).1–6
These findings raised a series of clinical questions, including its pathogenic and carcinogenic role and possible impact on HCV treatment response. Cacciola and colleagues conducted a retrospective study in Italy and found that among 66 of the 200 patients with CHC and negative serum HBsAg, but detectable HBV DNA in the liver biopsy tissue, 46 were anti-HBc positive and 20 were negative for all HBV markers (P<0.001). The frequency of cirrhosis was significantly higher in CHC patients with occult HBV infection compared with those without occult HBV infection (22/66, 33% vs. 26/134, 19%; P=0.04). The authors concluded that occult HBV infection occurs frequently in patients with CHC and may have clinical significance.4 Another recent study conducted in France reported that only 4.4% (5/113) of patients with CHC and negative serum HBsAg had detectable HBV DNA in the liver biopsy tissue.7 Kao and colleagues conducted a similar study in Taiwan and found that, in 210 patients with CHC and negative serum HBsAg, 32 (14.8%) had HBV DNA in their sera, which was comparable to 100 healthy controls (15/100, 15%). HBV DNA was not detected in the sera of patients negative for serological markers of HBV infection. They concluded that, in patients with CHC, the prevalence of occult HBV infection did not parallel the severity of liver disease (14.5% in patients with chronic hepatitis, 8% in patients with liver cirrhosis, and 22% in patients with hepatocellular carcinoma).8 Clearly, these results are not consistent, and it should be noted that in these studies different methods were used to determine occult HBV infection and all these reports are cross-sectional studies. Therefore, additional prospective studies are required to confirm whether occult HBV infection accelerates the progression of liver injury in patients with CHC.
Another important clinical question is whether occult HBV infection could impact the treatment response in patients with CHC. Unfortunately, the present data related to this issue have also been inconsistent. Although some studies reported that occult HBV infection may provoke a reduction in the response rate to interferon (IFN) therapy in patients with CHC,4–6,8,9 the opposite results have also been reported. In the study by Cacciola et al4 in Italy, CHC patients with negative HBsAg but detectable intrahepatic HBV DNA were found to have a higher nonresponse rate to conventional IFN monotherapy compared with those without occult HBV infection (26/55; 47% vs. 7/28; 25%), but this was not statistically significant (P=0.06). Another study in France reported that the sustained virological response (SVR) rate to conventional IFN combined with ribavirin was significantly lower in CHC patients with negative HBsAg, but positive serum HBV DNA, compared with those without occult HBV infection (11/40; 28% vs. 65/144; 45%; P<0.05). This was also true in those with HCV genotype 1 infection (42% vs. 58%, P=0.004). The same results were also reported in patients who were HBsAg negative and anti-HBc positive, especially in those with HCV genotype 1 and 4 infection, although the sample size was very small.7 However, another study in Italy reported that the SVR rate was about the same in HCV genotype 1–infected patients with or without occult HBV infection (5/13; 38.5% vs. 7/22; 31.8%).10
How about Asian patients? A study by Kao et al8 in Taiwan reported comparable SVR rate to conventional IFN and ribavirin treatment in CHC patients with or without occult HBV infection (38% vs. 39%); the majority of the studied patients had HCV genotype 1 infection. Similar results were also reported by another group in Taiwan in CHC patients who received pegylated (PEG)-IFN and ribavirin combination treatment.11
Clearly, the above-mentioned studies were retrospectively conducted with a small sample size, enrolled heterogenous patients with different HCV genotypes, and involved different regimens of HCV therapy. The data were analyzed with various emphases without a standard way to define occult HBV infection. Therefore, it is impossible to utilize such studies to draw a firm conclusion as to whether occult HBV infection is associated with HCV treatment response.
In this issue of the journal, Toyoda et al12 reported in their study whether HBV integration to hepatocytes has an impact on the efficacy of PEG-IFN and ribavirin therapy in HBV-negative CHC patients. They analyzed 157 CHC patients who underwent PEG-IFN and ribavirin combination therapy. Using the Alu-PCR assay, HBV integration was detectable in the liver biopsy tissue in 54 of the 157 (34.4%) patients with CHC. Although there were no significant differences between patients with and without HBV integration in terms of baseline characteristics, including liver histology, pretreatment HCV RNA levels, and genetic polymorphisms of IL28B genotype, patients with HCV genotype 1b (n=91) and HBV DNA integration showed a significantly higher SVR rate compared with those without HBV integration (19/30, 63% vs. 19/59, 32%; P=0.0098). The multivariate analysis confirmed that HBV integration and IL28B polymorphisms are the only 2 independent factors associated with SVR. The authors concluded that HBV integration was associated with a higher SVR rate to combination therapy with PEG-IFN and ribavirin in patients with HCV genotype 1b infection.
This study is different from most previous studies in several ways: it (1) was a relatively large and well-conducted retrospective study on this issue in Asian patients with CHC and genotype 1 infection; (2) enrolled patients who received uniform standard PEG-IFN and ribavirin combination therapy as the HCV treatment regimen, and (3) used HBV DNA integration, not routine HBV serology and serum HBV DNA, as an indication of HBV infection. Apart from these strengths, this study also held limitations, which will be discussed below.
First, in the absence of full medical history and HBV serological tests, it is hard to confirm whether these patients truly had a history of HBV infection. This concern is obvious by the fact that the anti-HBc positivity (a test performed in limited patients) was not well associated with HBV DNA integration.
Second, Alu-PCR is technically a relatively old method for detecting joints between cellular chromosome and viral sequences, and the primers were anchored on sites with Alu repeat sequences that may limit detection of different integration sites. Moreover, as authors indicated, using Alu repeats may limit the identification of HBV X sequence integration sites that are far away from the priming site. It should be noted that more sensitive methods for detecting joints of HBV integration have been well established.13–16 Perhaps these newer methods are preferred for future research on the related issue.
In addition, the authors did not present the data on the frequency of detectable HBV DNA integration in each individual liver specimen. As HBV integration could vary from 1/100 to 1/1000 hepatocytes,13,16 the clinical value of the reported HBV integration by Toyoda and colleagues remains to be determined, ideally in association with the frequency of HBV integration in these patients.
Third, in contrast to the traditional definition of occult HBV infection, most of the cases reported by Toyoda and colleagues had isolated HBV DNA integration in the absence of all other HBV biomarkers. HBV DNA integration has been thought to be associated with the development of hepatocellular carcinoma in 4 Japanese CHC patients without cirrhosis who achieved SVR. Two of these 4 patients had detectable HBV and covalently closed circular DNA, although serum HBV DNA was undetectable.17 Clearly, such isolated HBV DNA integration is by no means to meet the present definition of occult HBV infection. Therefore, further studies are required to determine the association of isolated HBV DNA integration with occult HBV infection. Unless this is well elucidated, it is difficult to determine the clinical relevance of the association of isolated HBV DNA integration with a higher SVR rate to PEG-IFN and ribavirin treatment in HCV genotype 1–infected patients.
Fourth, it remains unknown how the isolated HBV DNA integration influences the SVR rate to PEG-IFN and ribavirin treatment in HCV genotype 1–infected patients. Ideally, this could be addressed in the following ways: first, it is important to determine the frequency of HBV integration in each individual patient and further quantitatively correlate it with the SVR rate in these patients. Second, it may also be very valuable to determine which HBV fragments integrate into the host’s hepatocytic genome and what their possible transacting function is. For instance, the same research group has identified HBV X gene integration in liver tissue in noncirrhotic patients with hepatocellular carcinoma and serologically HBV-negative CHC patients, indicating the potential hepatocarcinogenic role of such HBV integration.18 Consistent with their previous report, they were able to confirm HBV X gene integration in some of their studied patients.12 However, as the authors indicated, they did not determine whether the integrated HBV fragment had potential transcriptional and translational function or truncated nonfunction fragment in these patients.
Fifth, significant research advances are being made in developing more effective, convenient, and safer HCV treatment regimens.19 This may raise another question whether occult HBV infection or isolated HBV integration could impact the SVR rates to these newly developing HCV treatment regimens.
In summary, the clinical significance of occult HBV infection in HCV pathogenesis and treatment response remains to be determined. The study by Toyoda and colleagues published in this issue of the journal reported that HBV integration to hepatocytes is associated with increased SVR rate to PEG-IFN and ribavirin therapy in HBV-negative CHC patients with genotype 1 infection. As discussed above, many questions have to be well addressed to reliably answer these important clinical questions.
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