United States

Chairman

Dr. Robert H. Purcell
(1990-   , Member 1979-1989)
Head, Hepatitis Viruses Section
Laboratory of Infectious Diseases
National Institute of Allergy and Infectious Diseases
National Institutes of Health
Building 7, Room 202
7 Center Drive MSC 0740
Bethesda, Maryland 20892-0740

Japan

Chairman

Dr. Kusuya Nishioka
(1988-   , Member 1979-1988)
Technical Advisor
The Japanese Red Cross Society
4-1-31 Hiroo, Shibuya-ku
Tokyo 150, Japan

Panel Members

Dr. Miriam Alter (1994-1997)
Chief, Epidemiology Section
Hepatitis Branch
Centers for Disease Control and Prevention
Mail Stop G37, Bldg. 6, Rm. 271
1600 Clifton Road
Atlanta, Georgia 30333

Dr. Francis V. Chisari (1992-1995)
Head, Division of Experimental Pathology
Department of Molecular and Experimental Medicine
The Scripps Research Institute
10666 North Torrey Pines Road
Room SR106 SBR-10
La Jolla, California 92037

Dr. Jay Hoofnagle (1991-1995)
Director, Division of Digestive Diseases and Nutrition
National Institute of Diabetes and Digestive and Kidney Diseases
National Institutes of Health
Building 31, Room 9A23
Bethesda, Maryland 20892-2560

Dr. Stanley M. Lemon (1991-1995)
Division of Infectious Diseases
Department of Medicine
University of North Carolina at Chapel Hill
547 Burnett-Womack CB #7030
Chapel Hill, North Carolina 27599-7030

Dr. Makoto Mayumi (1988-   )
Professor
Jichi Medical College
3311-1 Yakushiji, Minami-Kawachi-gun
Tochigi 329-04, Japan

Dr. Hiroshi Suzuki (1988-   )
President
Yamanashi Medical College
1100 Shimo-Gatoh, Tamaho-cho, Nakakoma-gun
Yamanashi 409-38, Japan

Dr. Kyuichi Tanikawa (1994-   )
Professor
Kurume University
School of Medicine
7, Asahimachi, Kurume

830, Japan Dr. Hiroshi Yoshikura (1994-   )
Professor
Faculty of Medicine
University of Tokyo
7-3-1 Hongo, Bunkyo-ku
Tokyo 113, Japan

Guidelines

Hepatitis Panels USJCMSP

The long-term objective of the United States-Japan Hepatitis Program is to define conditions for the control of viral hepatitis by improved environmental conditions, therapy, or prophylaxis. To achieve these objectives, the research areas of high interest to the program are:

1. Biochemical and biophysical characterization of hepatitis agents
2. Seroepidemiologic studies in Japan, the United States, and Asian countries to identify prevalence and incidence of infections caused by the various hepatitis viruses. Implicit in these objectives is appropriate work on in vitro cultivation of hepatitis agents.
3. Development and evaluation of vaccines, immunoglobulins, and antiviral compounds for prevention and treatment of illness with particular emphasis on therapeutic measures for chronic carriers of HBV and HCV
4. Long-term studies to determine the incidence of cirrhosis and hepatocellular carcinoma as sequelae of chronic viral hepatitis.

Five-Year Summary

Broad Goals

The major objectives of the Hepatitis Panels are to identify hepatitis viruses and define their attributes, to develop vaccines to prevent infections caused by hepatitis viruses, to design new antiviral substances to ameliorate/ cure chronic disease, and to safeguard the public by ensuring that its blood supply remains free of these infectious agents. A strong molecular biological component is associated with this program as investigators study key factors that influence organ targeting, the immune responses, protein processing, and the replication and packaging of viruses. Identifying suitable animal models and developing methods for cultivating all hepatitis viruses are additional goals.

Just 10 years ago, it was estimated that there were 150 million chronic carriers of hepatitis B virus (HBV) throughout the world. Today the number has increased to over 350 million. As more information on the recently discovered (1989) hepatitis C virus emerges, its impact on the prevalence of hepatitis and associated chronic liver disease will become more and more apparent. It is incumbent, therefore, to maintain a concerted effort to define and resolve chronic hepatitis. This also should include studies on the coinfectant, hepatitis D (HDV), which can cause more severe disease in HBV carriers.

Progress/Accomplishments

Both Japanese and U.S. investigators have examined ways to improve the growth of enterically-transmitted hepatitis A virus (HAV) in vitro and have found that mutations in the 5' nontranslatable and P-2 regions of the genome increase in vitro replication. Such efforts could result in the development of a live attenuated HAV vaccine. In the United States, two candidate formalin-killed HAV vaccines have been assessed in Phase III clinical trials. One, HAVRIX by SmithKlein Beecham Biologicals, was recently licensed for use in the United States. The other, by Merck, Sharp and Dohme Research, should be licensed fairly soon. The Japanese are examining similar candidate vaccines. Although the incidence of HAV is decreasing, HAV still accounts for significant illness in developed countries. HAV infection is more common in children in developing countries and poses a greater health risk to susceptible adults in the United States and Japan.

The perinatal spread of HBV is being curtailed significantly in Japan and the United States by vaccinating neonates. Today, a high percentage (85 to 95+ percent) of babies born to mothers who are carriers of HBV are vaccinated against HBV and are given hepatitis B gamma globulin (HBIG). The HBV vaccine is recommended by the Children's Vaccine Initiative for use in infants born in the United States. Currently, 40 percent of all infants are vaccinated against HBV.

The duck hepatitis B virus (DHBV) has been used to create the first successful in vitro hepadnaviral DNA synthesis assay, an excellent way to screen for some types of antiviral substances. Here, the polymerase region of the viral genome plays a role not only in reverse transcription as a primer, but also in viral encapsidation. Studies of acute infections by HBV and associated hepadnaviruses in animal models indicate that not all hepatocytes infected with the virus are lysed by the immune system. Rather, many cells are actually cured by immune mechanisms that impede viral replication. Efforts are now underway to use the HBV nucleocapsid as a vaccine vector for other infectious agents such as malaria. Initial success has been noted in animal models. Naked DNA vaccines against HBV represent a new vaccine paradigm. Advanced diagnostics have identified more silent cases of hepatitis B. This is an important advance for screening transplant and transfusion patients. Mutant viruses are of concern as precore mutants of HBV cause a more fulminant disease and "escape" mutants pose a threat to vaccination programs. Only alpha interferon is licensed as an antiviral to treat chronic HBV infections.

Since it was cloned and sequenced in 1989, HCV has generated extensive research in

• Defining the modes of transmission
• Analyzing structural and nonstructural proteins
• Preventing transmission in transfusion, transplantation, and hemodialysis patients
• Identifying HCV as a cofactor in cryoglobulinemia, porphyria cutanea tarda, hemochromatosis, and autoimmune hepatitis
• Identifying, cloning, and sequencing many serotypes and genotypes
• Improving HCV diagnostic tests, that are now in their third generation
• Studying the immune response to HCV infection
• Studying the effects of HCV in patients coinfected with HBV and/or HIV.

As with HBV, infection by HCV can lead to chronicity. However, unlike HBV, a high proportion of individuals infected with HCV become chronic carriers. Because most HCV carriers are asymptomatic, their propensity to spread the disease is high.

Research on HCV is limited as efforts to establish tissue culture systems for growing virus and for screening antivirals are still in preliminary stages of development. In addition, the chimp is the only known animal model, although efforts to produce genetically-defined mouse strains are underway. A rudimentary vaccine made from structural proteins was tested in chimps and found to provide some protection upon challenge. However, the goal of making a vaccine against HCV will not be easy to achieve as the virus mutates quickly after infection, thereby enabling subsequent infections with the same or different strains to occur. Neutralizing antibodies have been detected but are difficult to measure.

Efforts to identify nonA, nonB, and nonC viral agents as contaminants of the blood supply are ongoing. Results of initial efforts already have been published. Clearly, much more work is needed to confirm and expand on the present work.

As a viral subparticle, HDV is dependent on HBV's surface protein for packaging to complete its life cycle. RNA self-editing proceeds from a single amino acid mutation, which causes two lengths of the delta antigen to be copied. One assists in replication; the other is involved in packaging. Because HDV successfully coinfects woodchucks, this animal model enables re-searchers to study the influence of antiviral and vaccine candidates. In tissue culture, the HDV genome has been successfully replicated in the absence of coinfection with HBV. Of the three genotypes identified to date, one from South America appears to be the most virulent, and alpha interferon has been used to treat infected patients.

HEV, a water-borne calici-like hepatitis virus, is found primarily in Mexico and in developing countries of Asia and Africa. However, serologic tests have improved since the time this genome was first cloned in 1990 and its antigens expressed by recombinant methods. It has been determined that 1 to 2 percent of the world's population have antibody to HEV. Although it is not as infectious as HAV, which may account for periodic epidemics in unprotected adults, HEV can cause a greater risk to adults and exhibits high mortality in pregnant women. A single-dose vaccine candidate currently being developed in the United States shows protection against both disease and infection in monkeys.

Future Goals

Future goals include:

• Continued unravelling of the natural history, pathology, immunology, and biochemical/physical attributes of hepatitis viruses
• Implementation of effective vaccination strategies for HAV vaccines
• Improvement of universal programs to control HBV by vaccination
• Monitoring of HBV vaccine efficacy and the preparation of third generation vaccines
• Identification of effective antiviral therapies for short-term/long-term use in chronic hepatitis patients
• Improvement of diagnostics to make them easier to use in developing countries
• Identification of in vitro and in vivo models for studying the progression of disease and for analyzing vaccine/antiviral interdiction for hepatitis C
• Licensure of future vaccines for HCV, HDV, and HEV
• Identification and study of new hepatitis viruses.

Selected References

United States

1. Guidotti LG, Ando K, Hobbs MV, Ishikawa T, Runkel L, Schreiber RD, Chisari FV. Cytotoxic T lymphocytes inhibit hepatitis B virus gene expression by a noncytolytic mechanism in transgenic mice. Proc Natl Acad Sci USA 1994 Apr 26; 91 (9):3764-8.
2. Pollack JR, Ganem D. Site-specific RNA binding by a hepatitis B virus reverse transcriptase initiates two distinct reactions: RNA packaging and DNA synthesis. J Virol 1994 Sep; 68 (9):5579-87.
3. Fourel I, Cullen JM, Saputelli J, Aldrich CE, Schaffer P, Averett DR, Pugh J, Mason WS. Evidence that hepatocyte turnover is required for rapid clearance of duck hepatitis B virus during antiviral therapy of chronically infected ducks. J Virol 1994 Dec; 68 (12):8321-30.
4. Tsarev SA, Tsareva TS, Emerson SU, Govindarajan S, Shapiro M, Gerin JL, Purcell RH. Successful passive and active immunization of cynomolgus monkeys against hepatitis E. Proc Natl Acad Sci USA 1994 Oct 11; 91 (21):10198-202.
5. Farci P, Alter HJ, Govindarajan S, Wong DC, Engle R, Lesniewski RR, et al. Lack of protective immunity against reinfection with hepatitis C virus. Science 1992 Oct 2; 258 (5079):135-40.

Japan

1. Mishiro S, Hoshi Y, Takeda K, Yoshikawa A, Gotanda T, Takahashi K, Akahane Y, Yoshizawa H, Okamoto H, Tsuda F, Peterson D, Muchmore E. Non-A, non-B hepatitis specific antibodies directed at host-derived epitope: implication for an autoimmune process. Lancet 1990; 336:1400-3.
2. Kosaka Y, Takase K, Kojima M, Shimizu M, Inoue K, Yoshiba M, Tanaka S, Akahane Y, Okamoto H, Tsuda F, Miyakawa Y, Mayumi M. Fulminant hepatitis B: Induction by hepatitis B virus mutants defective in the precore region and incapable of encoding, antigen. Gastroenterology 1991; 100:1087-94.
3. Shimizu YK, Iwamoto A, Hijikata M, Purcell RH, Yoshikura H. Evidence for in vitro replication of hepatitis C virus genome in a human T-cell line. Proc Natl Acad Sci USA 1992; 89:547-81.
4. Suzuki H, Iino S, Shiraki K, Akahane Y, Okamoto H, Domoto K, Mishiro S. Safety and efficacy of a recombinant yeast-derived preS2+S-containing hepatitis B vaccine (TGP-943): Phase 1, 2, and 3 clinical testing. Vaccine 1994; 12:1090-6.
5. Ohto H, Terazawa S, Sasaki N, Sasaki N, Hino K, Ishiwata C, Kako M, Ujiie N, Endo C, Matsui A, Okamoto, H, Mishiro S. The vertical transmission of hepatitis C virus collaborative study group: Transmission of hepatitis C virus from mothers to infants. N Engl J Med 1994; 330:744-50.