The lack of small-animal models that are suitable for evaluation of agents used to treat infection with hepatitis C virus (HCV) severely hinders the assessment of potential new therapies for the disease. This study created such a model, termed the "HCV-Trimera" model. The HCV-Trimera model was developed by using lethally irradiated mice, reconstituted with SCID mouse bone marrow cells, in which human liver fragments infected ex vivo with HCV had been transplanted. Viremia (positive-strand HCV RNA levels) in HCV-Trimera mice peaked at approximately day 18 after liver transplantation, and an infection rate of 85% was reached. Viral replication in liver grafts was evidenced by the presence of specific negative-strand HCV RNA. The usefulness of this model for evaluation of anti-HCV agents was demonstrated by the ability of a small molecule (an HCV internal ribosomal entry site inhibitor) and an anti-HCV human monoclonal antibody (HCV AB(XTL)68) to reduce virus loads in HCV-Trimera mice in a dose-dependent manner.

Hepatitis B virus (HBV) infection presents a major public health problem worldwide. Persistent HBV infection occurs in 5% to 10% of patients after acute hepatitis and may progress to chronic liver disease including chronic active hepatitis, cirrhosis, and hepatocellular carcinoma. 1 Newborns to HBV carrier mothers and immunocompromised individuals such as liver transplant recipients are at high risk for contracting HBV infection. 2,3 Recombinant HBV vaccines provide a safe and effective means for prevention of HBV conferring long-term immunity through active immunization. 4 In contrast to the gradual onset of protection following active immunization against HBV, passive immunization with antibodies to hepatitis B immunoglobulin (HBIG) provides immediate but short-term protection against viral transmission and infection. Administration of HBIG is commonly used in recipients of liver transplants who were HBV carriers to prevent HBV reinfection of the graft. 5-9 HBIG is a plasma derived, polyclonal antibody preparation obtained from blood donors who were anti-hepatitis B surface antigen (HBsAg) antibody positive. The limited availability of anti-HBsAg high-titer donors as well as safety concerns as to the use of a blood derived product, especially in immunosuppressed patients, urge for an alternative source for such immunotherapy. Human monoclonal antibodies (mAbs) to HBV may offer such an alternative. For clinical use, mAbs would be advantageous by presenting a stable and reproducible source for prolonged immunotherapy. Human mAbs to HBV generated by fusion of B cells derived from peripheral blood lymphocytes of vaccine immunized patients were described. [10][11][12][13][14][15] It was shown that mAbs might protect chimpanzees against challenge with HBV. 14,16,17 Recently we have described the development of high affinity, fully human mAbs against HBsAg generated in the Trimera system. These mAbs were previously characterized in terms of their affinity, specificity, and ability to bind HBV-infected human liver tissue. 18 In the present report, we have characterized the anti-HBV effect of 2 human mAbs, 17.1.41 and 19.79.5 by evaluating their specificity and reactivity to HBsAg derived from different viral subtypes and genotypes. The anti-HBV effect was evaluated in vivo in 2 different animal model systems, in HBVTrimera mice, [19][20][21] and in HBV chronic-carrier chimpanzees. The antibodies were able to reduce viral load in both animal models and to inhibit HBV infection in the HBV-Trimera model. The 2 mAbs were more potent than Hepatect (Biotest Pharma, Dreieich, Germany) in reducing viral load and inhibiting HBV infection in this model, suggesting a potential clinical use.Abbreviations: HBV, hepatitis B virus; HBIG, hepatitis B immunoglobulin; HBsAg, hepatitis B surface antigen; mAb, monoclonal antibody; ␤-L-5FddC, ␤-L-5-fluoro-2Ј, 3Ј-dideoxycytidine; ELISA, enzyme-linked immunosorbent assay; PBS, phosphate-buffered saline; BSA, bovine serum albumin.From

An approach to develop fully human monoclonal antibodies in a human/mouse radiation chimera, the Trimera system, is described. In this system, functional human lymphocytes are engrafted in normal strains of mice which are rendered immuno-incompetent by lethal total body irradiation followed by radioprotection with severe combined immunodeficient (SCID) mouse bone marrow. Following transplantation, human lymphocytes colonize murine lymphatic organs and secrete human immunoglobulins. We have established this system as a tool to develop fully human monoclonal antibodies, and applied it for the generation of monoclonal antibodies specific for hepatitis B virus surface antigen. A strong memory response to hepatitis B surface antigen was elicited in Trimera engrafted with lymphocytes from human donors positive for antibodies to hepatitis B surface antigen. The human specific antibody fraction in the Trimera was 10(2)-10(3)-fold higher as compared with that found in the donors. Spleens were harvested from Trimera mice showing high specific-antibody titres and cells were fused to a human-mouse heteromyeloma fusion partner. Several stable hybridoma clones were isolated and characterized. These hybridomas produce high-affinity, IgG, anti-hepatitis B surface antigen antibodies demonstrating the potential of the Trimera system for generating fully human monoclonal antibodies. The biological function and the neutralizing activity of these antibodies are currently being tested.

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In chronic hepatitis B virus (HBV) infectionInfection with the hepatitis B virus (HBV) is one of the most common infectious diseases with an estimated 300 million chronic HBV carriers worldwide. 1 The high risk of patients with chronic hepatitis B (CHB) developing liver cirrhosis and hepatocellular carcinoma is associated with a high morbidity and mortality. Because HBV is a noncytopathic virus, liver injury is mainly mediated by the host immune response against virus-infected liver cells. 2 Strong HBV core/precore (HBc/e) specific T helper (Th) cell activities are detectable in acute self-limited hepatitis B or during exacerbations of the chronic disease in concert with strong HBV envelope, core, and polymerase-specific cytotoxic T lymphocyte reactivities, whereas such T-cell responses are weak in chronic disease. [3][4][5][6] This HBV-specific T-cell failure of chronic HBV carriers is believed to account for viral persistence.Furthermore, the B-and Th-cell response to the HBV surface antigen (HBsAg) seems to be insufficient in chronic hepatitis B. Whereas in patients with acute hepatitis B, the disappearance of HBsAg and the occurrence of anti-hepatitis B surface antigen (HBs) antibodies in serum mark viral elimination with subsequent development of protective immunity, in patients with CHB such antibodies are usually undetectable. Accordingly, low frequencies of anti-HBssecreting B cells are detected in the latter patients in contrast to very high B-cell numbers in patients with acute self-limited hepatitis B. 7 However, from current in vitro studies it remains unclear whether chronic HBV carriers exhibit an HBsAgspecific Th cell defect because proliferative T-cell responses to HBsAg are very weak or undetectable not only in chronic infection but also during acute hepatitis B or during acute exacerbations of the chronic disease. 4,5 Moreover, in a recent study, antienvelope antibodies were detected in immune complexes with HBsAg in sera of patients with chronic HBV infection. 8 However, the epitope specificity of such antibodies was not further specified (i.e., anti-preS1/2 or anti-HBs) and the antibody levels were not compared with those of HBVimmunized or HBs-vaccinated individuals. Thus, our current knowledge about the cellular and molecular mechanisms of the presumed HBs-specific B-and T-cell defect of chronic HBV carriers is controversial and limited.Anti-HBs antibodies might mediate important antiviral effector functions because anti-HBs antibodies are virus neutralizing, 9,10 HBsAg is expressed on the membrane of infected liver cells 11 and anti-HBs-mediated cytotoxicity has