Abstract Abstract 3350 Introduction: In the absence of serum hepatitis B (HB) surface antigen (HBsAg) and clinical evidence of HB, an occult HB virus (HBV) infection (OBI) is defined as the existence of HBV DNA in the serum, in the peripheral blood mononuclear cells (PBMC), and/or in the liver. Most frequently, OBI follows the resolution of acute hepatitis and continues indefinitely after HBsAg clearance and liver function improvement. Usually, this form of infection, termed secondary OBI (sOBI), is associated with the presence of serum anti-HB core antibodies (anti-HBc). Up to 20% of individuals with OBI do not react for either anti-HBc or any other serological indicator of HBV exposure. Consistent with the woodchuck model, this form of infection, termed primary OBI (pOBI), is characterized by the presence of DNA in both serum and PBMC, but not in the liver. Although reactivation of sOBI has been reported in a high proportion of hemopoietic stem cell transplant (HSCT) recipients, pOBI reactivation risk has not yet been fully investigated in the transplant setting. We followed up 9 long term surviving HSCT recipients with pOBI, in order to define its clinical significance in immune suppressed patients. Patients and Methods: A total of 75 consecutive patients with hematological disease, receiving autologous or allogeneic HSCT between April 2006 and March 2007, were included in a database and retrospectively examined. As a prerequisite for inclusion in the study, each of the following was needed: (i) negative serological indicators of HBV exposure and absence of clinical signs/symptoms of hepatitis at the time of HSCT, (ii) positive HBV DNA, as detected by polymerase chain reaction (PCR), in both serum and PBMC samples drawn within 6 months before transplant, and (iii) availability of stocked frozen serum/PBMC samples collected after HSCT from each patient. Following these criteria, 9 patients (7 males, 2 females) were selected for the analysis. Five patients (1 Hodgkin disease, 3 non-Hodgkin lymphomas, and 1 B-cell chronic lymphocytic leukemia) received autologous HSCT, and 4 patients (1 acute myeloid leukemia, 1 multiple myeloma, 1 myelodysplastic syndrome, and 1 acute lymphoblastic leukemia) received allogeneic HSCT. Median age at transplant was 49 (range: 33–63) years. No patient received lamivudine prophylaxis. Serum and PBMC samples collected after HSCT were analyzed for HBV serology and DNA detection. The latter was performed by in-house PCR amplification by nested primers (detection limit 50 copies/ml) from highly conserved S region encompassing a-determinant of HBV. Direct sequencing of all HBV DNA amplified products was used to confirm the specificity of the reaction. Results: In 8 cases, one serum/PBMC sample per patient was available, at a median of 27 (range: 21–34) months after HSCT; in 1 case, three serum/PBMC samples were available, at 17, 22, and 28 months after HSCT, respectively. During follow-up, positive HBV DNA was found, both in the serum and in the PBMC. All patients were confirmed to be infected with HBV subgenotype D1, with mutations known to impair HBsAg antigenicity. Three patients were confirmed to be further infected with HBV subgenotype A2. So far, 41 to 52 months after transplant (median: 46 months), neither patients showed seroconversion nor developed clinical hepatitis, suggesting pOBI persistence. Conclusions: Although the virus persisting in pOBI remains biologically competent, it is likely neither to reach liver-pathogenic doses nor to induce “classical” HBV reactivation during PBMC-restricted replication. However, prospective studies with large sample size and long-term follow-up are warranted, to define the actual clinical relevance of HBV occult infection. Indeed, OBI with low viremia could induce DNA damage, cellular demise, immune hypo-responsiveness, and oxidative stress in peripheral blood lymphocytes. Disclosures: No relevant conflicts of interest to declare.

Some individuals have "occult" infection with hepatitis B virus (HBV), defined as presence of HBV genome in the serum or liver tissue without HBV surface antigen (HBsAg) in the serum. The aim of this study was to investigate whether serum antibodies against HBV core antigen in isolation ("anti-HBc alone") are a useful marker of "occult" HBV in patients with or without hepatitis C virus (HCV) infection. "Anti-HBc alone" was detected in the sera of 119/6,544 (1.8%) asymptomatic outpatients referred to the diagnostic laboratory for routine testing for viral hepatitis, 62/607 (10.2%) drug users, and 42/195 (21.5%) patients with hepatocellular carcinoma. Using three in-house nested-PCR amplification assays to detect HBV preS-S (S), precore-core (C), and Pol viral regions, respectively, "occult" HBV sequences were found in 9 of the 223 sera (4.0%) with "anti-HBc alone." The highest prevalence of "occult" HBV sequences (5.9%) was detected in "anti-HBV alone" sera of individuals referred to the diagnostic laboratory without HCV antibodies. Direct sequencing of all PCR products confirmed the specificity of the PCR reactions and revealed the predominance of HBV genotype D. The data presented in this study suggest that detection of "anti-HBc alone" could reflect unrecognized "occult" HBV infection and that physicians should consider investigating such patients with HBV molecular tests.

Tumor recurrence after resection of hepatocellular carcinoma (HCC) can occur early (<2 years) or late (>2 years) as metastases or de novo tumors. Interferon (IFN) has the potential for chemoprevention against hepatitis C virus (HCV)-related cirrhosis. A predetermined group of 150 HCV RNA-positive patients undergoing resection of early- to intermediate-stage HCC was stratified into 80 HCV-pure (hepatitis B anticore antibody [anti-HBc]-negative) and 70 mixed HCV+hepatitis B virus (HBV) (anti-HBc-positive) groups, then randomized to IFN-alpha (3 million units 3 times every week for 48 weeks [n = 76]) versus control (n = 74). The primary end point was recurrence-free survival (RFS); secondary end points were disease-specific and overall survival. Intention-to-treat and subgroup analysis on adherent patients were conducted. Treatment effects on early/late recurrences were assessed using multiple Cox regression analysis. No patient experienced life-threatening adverse events. There were 28 adherent patients (37%). After 45 months of median follow-up, overall survival was 58.5%, and no significant difference in RFS was detectable between the two study arms (24.3% vs. 5.8%; P = .49). HCC recurred in 100 patients (48 IFN-treated, 52 controls), with a 50% reduction in late recurrence rate in the treatment arm. HCC multiplicity and vascular invasion were significantly related to recurrence (P = .01 and .0003). After viral status stratification, while no treatment effect was apparent in the mixed HCV+HBV population and on early recurrences (72 events), there was a significant benefit on late recurrences (28 events) in HCV-pure patients adherent to treatment (HR: 0.3; 95% CI: 0.09-0.9; P = .04). In conclusion, IFN does not affect overall prevention of HCC recurrence after resection, but it may reduce late recurrence in HCV-pure patients receiving effective treatment.

OBJECTIVE:The aim of this study was to investigate the role of alcohol dehydrogenase type 3 (ADH3), glutathione S-transferase M1 (GSTM1) and T1 (GSTT1) polymorphisms in modifying hepatocellular carcinoma (HCC) risk according to alcohol intake.METHODS:A hospital-based case-control study was conducted in two areas of North Italy. Two-hundred cases hospitalized for HCC and 400 controls were recruited. Genotypes were determined using PCR and the PCR/restriction fragment length polymorphism-based method.RESULTS:There was no association of the putative risk genotypes ADH3(1-1), GSTM1 null and GSTT1 null with HCC (odds ratio [OR], 0.8; 95% confidence interval [CI], 0.5-1.3; OR, 1.0; 95% CI, 0.6-1.5; OR, 0.8; 95% CI, 0.4-1.4, respectively). A steady increase in HCC risk with increasing alcohol intake, which did not vary according to ADH3 and GSTT1 genotypes, was observed. Nevertheless, the OR for HCC due to an alcohol intake of >100 g of ethanol per day increased in subjects with GSTM1 null genotype (OR, 8.5; 95% CI, 3.9-18.6) compared to GSTM1 non-null genotype (OR, 4.5; 95% CI, 2.0-10.0).CONCLUSIONS:ADH3(1-1) and GSTT1 null genotypes did not modify the risk of HCC due to alcohol intake whereas an influence of GSTM1 null genotype for high ethanol consumption was suggested.

Background/Aims: The aim of the present, open-labeled, randomized study was to determine the efficacy and safety of different doses of consensus interferon plus ribavirin in the initial treatment of chronic hepatitis C.Methods: One hundred and one genotype 2/3 patients were randomized to receive 9 mcg (group A, n = 48) or 18 mcg (group B, n = 53) of consensus interferon thrice weekly plus ribavirin (1000/1200 mg/daily) for 24 weeks and 92 genotype 1 patients to receive 9 mcg (group C, n = 47) or 18 mcg (group D, n = 45) of consensus interferon plus ribavirin for 48 weeks.Results: In an intention-to-treat analysis, the sustained virologic response at 24-week follow-up was 69% and 66% for group A and B (P = 0.77) and 40% and 36% for group C and D (P = 0.63). The overall sustained response was 67% and 38% in patients with genotype 2/3 and 1, respectively. Among genotype 1 patients the sustained virologic response was 39% and 41% for high or low baseline viremia levels.Conclusions: Higher consensus interferon dose does not increase sustained virologic response. Naive genotype 1 patients may achieve significant response rate of approximately 40% if treated with 9 mcg of consensus interferon plus ribavirin for 48 weeks. (C) 2003 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Objective: Intrahepatic cholangiocarcinoma (ICC) is a rare type of primary liver cancer (PLC) arising from intrahepatic bile ducts. We carried out a case–control study to assess the association between ICC and hepatitis B and C virus (HBV and HCV) infections, alcohol intake, and hepatolithiasis in Brescia, North Italy. Methods: Among 370 subjects with histology-based diagnosis of PLC who were resident in the area and hospitalized in 1995–2000, 26 (7%) ICC cases were identified. A total of 824 subjects unaffected by hepatic diseases and frequency-matched with PLC cases by age, sex, date, and hospital of admission were recruited as controls. Results: Among ICC cases the mean age was 65 years, 80.8% were males, and 38.5% had cirrhosis. Seropositivity for anti-HCV, HBsAg, alcohol intake > 80 g/day and history of hepatolithiasis were found in 25%, 13%, 23.1%, and 26.9% of ICC cases and in 5.8%, 6.7%, 32.9%, and 10.6% of controls, respectively. The odds ratios adjusted for demographic factors by logistic regression (95% confidence interval; 95% CI) were 9.7 (1.6–58.9) for anti-HCV, 2.7 (0.4–18.4) for HBsAg, and 6.7 (1.3–33.4) for hepatolithiasis, whereas no association was found with alcohol drinking. Conclusions: HCV and hepatolithiasis may be risk factors for ICC in Western countries.

Objectives: We carried out a case–control study to investigate the role of history of liver cancer in a first-degree relative as a risk factor for hepatocellular carcinoma (HCC).

We performed a case-control study to evaluate the association of a new human DNA virus named TT virus (TTV) with hepatocellular carcinoma (HCC). We recruited 174 subjects hospitalized for HCC (84% males; mean age: 64 years) and 118 patients hospitalized for non-liver diseases in Brescia, northern Italy, as controls (94% males; mean age: 66 years). TTV DNA was found in serum by polymerase chain reaction (PCR) in 26 cases (15%) and 11 controls (9.3%) (P >. 1). TTV group 2 infection was identified in 16 cases (61.5%) and 4 controls (36.4%) (P >.1) using a type-specific PCR method. Sequence analysis of 222 nt of TTV DNA demonstrated that the remaining 10 cases and 7 controls were all infected by group 1. The odds ratio (OR) for TTV-DNA positivity, adjusted for demographic variables, hepatitis B surface antigen (HBsAg), hepatitis C virus (HCV) RNA, and heavy alcohol intake was 1.8 (95% CI: 0.7-4.8; P >.1). The OR did not change when the analysis was restricted to 14 HCC cases and 56 controls who were negative for each known risk factor for HCC (OR = 1.7; 95% CI: 0.8-4.0). TTV-DNA positivity was not associated with transfusion history. The prevalence of TTV DNA was higher among HCC cases positive for HBsAg (10 of 38 [26.3%]) than among those positive for HCV RNA (8 of 62 [12.9%]) or negative for hepatitis B virus (HBV), HCV, and hepatitis G virus (HGV) infections (5 of 62 [8. 1%]) (P =.02). This study does not support the hypothesis of an association between TTV infection and HCC.

Serial serum samples from 19 patients with post-transfusion hepatitis C, followed for an average of 8 years, were tested for HCV antibody by individual synthetic peptide-based ELISA (core, NS3/4, El, E2/NS1 and NS5) and four-antigen RIB A, and for HCV-RNA by PCR amplification with nested primers from 5′ noncoding region and type-specific primers from core and NS5 regions. Ten patients progressed from acute to chronic hepatitis and 9 recovered. An expanded antibody pattern seems to correlate with a higher frequency of HCV-RNA-positive patients. The disappearance with time of HCV antibody was observed only in patients who cleared HCV. The persistence of HCV-RNA in 5 of 9 recovered patients suggests the existence of a ‘healthy’ carrier state of HCV. Most patients with chronic hepatitis had type I or II HCV infection, whereas most asymptomatic HCV carriers were infected with type IV HCV.

Hepatitis C virus is the most frequent cause of chronic non-A, non-B hepatitis, and the antibodies to structural and nonstructural proteins encoded by viral genome have been suggested to be markers of ongoing HCV infection. We studied the behavior of these antibodies during interferon therapy in 18 patients with chronic hepatitis C and also during a follow-up period of at least four years. A significant decrease of anti-HCV titer was found only in patients who had shown positive response to therapy and all of them were anti-HCV negative at the end of follow-up. Analysis by recombinant immunoblotting assay showed that only anti-c100 were affected by interferon therapy, whereas anti-c22 and anti-c33 were not modified. Using polymerase chain reaction to detect small amounts of HCV genome in serum, we could confirm that the behavior of HCV-RNA during and after interferon therapy is similar to that of anti-HCV and the loss of anti-c100 seems to be closely related to HCV-RNA disappearance from serum. Our patients with chronic hepatitis C were found to be of type 1b and 2, according to the recent score of Simmonds, and the clearance of serum HCV-RNA during treatment and its sustained negative status are closely related to genotype 2 and to long-term positive response to interferon.

During the first year (1986) of blood donor screening for antibody to HIV, 201,750 subjects were tested in 40 blood banks of Lombardia (Italy). All sera repeatedly positive by ELISA were submitted to our reference center for confirmation by Western blot (WB). Only 40 (0.02%) of 286 repeatedly reactive donors were positive by WB, whereas another 45 (0.022%) gave atypical antibody reactivities on WB, mainly directed against HIV core proteins. Of the 16 donors with inconclusive WB results followed for 4-12 months, 3 developed a full-blown antibody response, 5 maintained the anti-core reactivity throughout the follow-up period, and 8 lost all reactivities. The use of recombinant env and core antigen ELISAs seems to decrease the proportion of sera with inconclusive WB reactions, and to identify as true positive all seroconverting donors in advance of the WB test. The large majority (35 out of 40) of WB-positive donors and all seroconverters for antibody to HIV admitted to belong to a group at risk for AIDS. Among the 19 first-time donors with HIV infection, we found 3 subjects with serological evidence of LAV-2 infection. We describe also the diagnostic and ethical issues when a donor notification policy is based on WB confirmatory procedures.

Abstract. During the first year (1986) of blood donor screening for antibody to HIV, 201, 750 subjects were tested in 40 blood banks of Lombardia (Italy). All sera repeatedly positive by ELISA were submitted to our reference center for confirmation by Western blot (WB). Only 40 (0.02%) of 286 repeatedly reactive donors were positive by WB, whereas another 45 (0.022%) gave atypical antibody reactivities on WB, mainly directed against HIV core proteins. Of the 16 donors with inconclusive WB results followed for 4–12 months, 3 developed a full‐blown antibody response, 5 maintained the anti‐core reactivity throughout the follow‐up period, and 8 lost all reactivities. The use of recombinant env and core antigen ELISAs seems to decrease the proportion of sera with inconclusive WB reactions, and to identify as true positive all seroconverting donors in advance of the WB test. The large majority (35 out of 40) of WB‐positive donors and all seroconverters for antibody to HIV admitted to belong to a group at risk for AIDS. Among the 19 first‐time donors with HIV infection, we found 3 subjects with serological evidence of LAV‐2 infection. We describe also the diagnostic and ethical issues when a donor notification policy is based on WB confirmatory procedures.