The advent of hepatitis C testing.
Since the early 1970s it has been apparent that the majority of cases of transfusion-transmitted hepatitis are caused by a virus other than hepatitis A or B. Epidemiologic studies established that this relatively common transfusion complication was infectious. In the absence of a test for the responsible agent or agents, the illness became known by default as non-A, non-B hepatitis.
Although investigators were unable to isolate and identify the causative virus(es), they discovered a great deal about non-A, non-B hepatitis. It was found to be readily transmissible by blood products and to occur in 7 to 10 per cent of all transfusion recipients, usually as an inapparent illness. Now we have a test, and universal screening of all blood units for hepatitis C has been implemented in the last few months.
Individual risk for hepatitis C is a function of the number of blood products received. As occurs with hepatitis B but not with hepatitis A, a chronic carrier state can develop after hepatitis C infection. Only 5 to 10 per cent of individuals infected with hepatitis B, but 50 per cent of those with hepatitis C, become chronic carriers. Persons who harbor the non-A, non-B virus can remain infectious for years. Many eventually succumb to chronic hepatitis or cirrhosis.
Although transfusion-transmitted cases of non-A, non-B hepatitis were relatively easy to document, sporadic (community-acquired) cases predominated. CDC estimated 150,000 new cases per year in the United States. According to data available before the blood supply was routinely tested for anti-HCV, only 7,500 to 15,000 of these cases are transfusion related. It is clear that individuals who acquire non-A, non-B virus in a manner unrelated to transfusion can transmit it to others if they donate blood.
In the absence of a specific test, blood banks turned to surrogate markets to eliminate potential donors likely to be infected with the non-A, non-B virus. Although a handful of blood banks had begun such screening a few years earlier, ALT and anti-HBc testing was empirically instituted by all U.S. blood banks in 1987 with the expectation of decreasing the incidence of non-A, non-B, hepatitis by 50 per cent. Eliminating donors with positive surrogate markers would cause the donor pool to contract only minimally.
The identification of the hepatitis C virus by researchers at Chiron Corp. in Emeryville, Calif., announced in 1988, was a milestone event for at least three reasons:
1. The recombinant DNA technology used by Chiron's molecular biologists to identify and clone hepatitis C viral proteins was unique in that it enabled the scientists to obtain cDNA clones without prior knowledge of the virus's structure. The same generic procedure can be used in the future to identify other unknown infectious agents.
2. Identifying the hepatitis C virus and developing tests to detect antibody to its proteins will effect yet another significant decrease in the incidence of post-transfusion hepatitis.
3. Having tests available to identify antibodies to hepatitis C finally enables investigators to study the complete spectrum of this elusive disease, including its relationship to chronic hepatitis, cirrhosis, and liver cancer; the co-factors necessary for progression of disease; and the degree to which surrogate markers for the disease identify previously and currently infected individuals. The test allows investigators to test the theory that non-A, non-B hepatitis is caused by a single agent, not by multiple ones. * Research path. Simply stated, the elegant technology that led to the identification of HCV involved the following procedure. Large volumes of plasma from an experimentally infected chimpanzee with a high infectious titer of non-A, non-B virus were ultracentrifuged. The resultant pellet of nucleic acid was recovered and denatured. Reverse transcriptase was used to synthesize complementary DNA (cDNA) from any RNA and/or DNA present in the denatured specimen. Random oligomer primers were used because the nature of the virus was still unknown.
Next, a bacteriophage vector was employed to express cDNA-encoded polypeptides in microbes. This "library" of bacteriophage-expressed polypeptides was subsequently screened against the serum of a patient with documented chronic non-A, non-B hepatitis. Investigators hoped to detect in the index patient's serum the presence of antibody to possible viral polypeptides. After screening about one million bacteriophage-modified microbial clones, they noted a single polypeptide product, now known as 5-1-1, which reacted with antibody present in the patient's serum.
Subsequent investigation revealed that 5-1-1 was part of a larger protein called 81, and of another called C-100-3. (As a point of historical interest, when Novel laureate Paul Ehrlich was searching for chemicals to destroy syphilis-causing spirochetes, he rejected the first 605 he had synthesized before settling on the arsenical "Salvarsan 606." That was in 1910.)
Harvey Alter, M.D., and colleagues at the NIH Clinical Center ultimately provided the link between the bacteriophage-expressed polypeptide, the antibody that reacted to it, and non-A, non-B hepatitis. Dr. Alter maintains a coded pedigreed panel of frozen sera, some of which are known to have transmitted non-A, non-B hepatitis. When the Chiron test for HCV antibody was performed on Alter's panel, 90 per cent of the transfusion-associated hepatitis cases - or their implicated donors - tested positive. This development established hepatitis C as the long-sought causative agent of most cases of non-A, non-B post-transfusion hepatitis.
From these developments and the studies that followed, hepatitis C was determined to be a 10-kilobase, enveloped, single-stranded, positive-sense RNA virus related to togaviruses and flaviviruses. The flaviviruses are associated with human diseases such as yellow fever and dengue.
Ortho Diagnostic Systems in Raritan, N.J., was the first manufacturer to develop an enzyme immunoassay antibody detection test based on the Chiron recombinant protein C-100-3. The firm submitted clinical trial data to the FDA in August 1989 and received FDA licensure on May 2, 1990. In mid-July 1990, Abbott Laboratories, Abbott Park, Ill., received the second FDA license for a test based on the Chiron technology.
Almost immediately after the FDA had authorized the Ortho test kit, testing for HCV antibody began on all blood products transfused in the United States. Those testing positive were removed from inventory and destroyed. Positive donors were advised not to donate again pending the availability of confirmatory tests to distinguish true-positive from false-positive results. The two surrogate tests, ALT and anti-HBc, will be used to screen donors for the foreseeable future. * New information. Beyond protecting the nation's blood supply, the availability of tests for antibody to HCV has begun to provide insights into the spectrum of the disease while elucidating its epidemiology. The professional journals are publishing new knowledge about hepatitis C at such a rate that it has become difficult to remain current. Among the most noteworthy findings:
[Paragraph]Prevalence. Approximately 1 per cent of adult Americans test positive for anti-HCV. How many of these are true positives cannot be known until a confirmatory test is available. Although Ortho has developed a recombinant immunoblot assay (RIBA) for use as a supplemental test, it has not yet been licensed by the FDA and is not really a confirmatory test.
How many of the true positives are infectious and how many represent resolved infections must still be determined. Several studies indicate that anti-HCV - positive individuals who are probably infectious are those with high optical density values on enzyme immunoassay (EIA), elevated ALTs, and positive tests for anti-HBc. Less likely to be infectious are individuals who lose anti-HCV over a period of years and whose ALTs reverted to normal.
(Paragraph) Test comprehensiveness. Using a pedigreed library of frozen serum specimens and the radioimmune assay for anti-HCV (not yet available commercially), Dr. Harvey Alter and colleagues assessed the value of ALT and anti-HBc as surrogate markers for non-A, non-B hepatitis (now hepatitis C). Their data reveal that these two tests would have detected one-half of the anti-HCV - positive donors involved in the transmission of biopsy-confirmed post-transfusion hepatitis. Only 33 per cent of anti-HCV - positive donors had elevated ALTs; only 54 per cent tested positive for anti-HBc.
[Paragraph]Value of surrogate markers. The New York (City) Blood Center examined frozen serum specimens drawn from 456 donors in 1985 and 1986. The investigators found that surrogate markers would have failed to detect one-third to one-half of the units that tested positive for anti-HCV.
[Paragraph]Mode of transmission. Perhaps the single most important question about hepatitis C remains to be resolved: How is it transmitted? Miriam Alter, Ph.D. (no relation to Dr. Harvey Alter of NIH), and associates at the CDC studied 140 patients who had non-A, non-B hepatitis. No commonly recognized source for the infection - that is, transfusion or IV drug use - was identified in 53 per cent. There is some evidence for sexual transmission, but this is tenuous at best.
Statistical analysis subsequently revealed that the factors associated with increased risk of non-A, non-B hepatitis included a lower level of education, household or sexual activity with a person who had had hepatitis within the previous year, and heterosexual activity with more than two partners.
In contrast, Spanish studies of the female sex partners of infected male IV drug users reported minimal risk of sexual transmission of hepatitis C. While there is no current evidence to suggest a role for arthopod vectors in hepatitis C, it is worth noting that both dengue and yellow fever - diseases caused by related viruses - are transmitted by this route.
[Paragraph]Severity. The seriousness of hepatitis C infection was underscored by information reported in 1989 by Dr. Raymond Gambino based on information presented at a meeting at the New York Blood Center. Dr. Gambino related that 50 per cent of acute cases of non-A, non-B hepatitis become chronic. Twenty per cent of the chronic carriers develop cirrhosis in a mean time of 17 years. There is also an increased risk of developing hepatocellular carcinoma in a mean time of 20 years.[9, 10]
[Paragraph]Additivity. The synergy of hepatitis C and alcohol abuse in the development of cirrhosis and hepatocellular carcinoma (primary liver cancer) was recently addressed by researchers in Barcelona. In this study, 75 per cent of patients with hepatocellular carcinoma were anti-HCV positive, as were 56 per cent of patients with cirrhosis and 7 per cent of controls. The prevalence of anti-HCV was 76 per cent among patients who had both alcoholic cirrhosis and hepatocellular carcinoma.
Most of the knowledge needed to understand the epidemiology, clinical course, laboratory diagnosis, and treatment of hepatitis C has yet to be obtained. Second-generation antibody tests, third-generation tests for viral RNA, and the results of ongoing therapeutic trials with interferon alpha are eagerly anticipated. The results of all this hard work will contribute to a growing body of information about the way viruses work - and how to stop them.
[1.] Tabor, E.; Seef, L.B.; and Gerety, R.J. Chronic non-A, non-B carrier state: Transmissible agent documented in one patient over a six-year period. N. Engl. J. Med. 303: 140-143, 1980.
[2.] Stevens, C.E.; Aach, R.D.; Hollinger, F.B.; et al. Hepatitis B virus antibody in blood donors and the occurence of non-A, non-B hepatitis in transfusion recipients: An analysis of the transfusion-transmitted viruses study. Ann. Intern. Med. 101: 733-738, 1984.
[3.] Choo, Q.L.; Kuo, G.; Weiner, A.J.; et al. Isolation of a cDNA clone derived from a blood-borne non-A, non-B viral hepatitis genome. Science 244: 359-362, 1989.
[4.] Alter, H.J.; Purcell, R.H.; Shih, J.W.; et al. Detection of antibody to hepatitis C virus in prospectively followed transfusion recipients with acute and chronic non-A, non-B hepatitis, N. Engl. J. Med. 321: 1494-1500, 1989.
[5.] Van der Poel, C.L.; Reesink, H.W.; Schaasberg, W.; et al. Infectivity of blood seropositive for hepatitis C virus antibodies. Lancet 335: 558-560, March 10, 1990.
[6.] Stevens, C.E.; Taylor, P.E.; Pindyck, J.; et al. Epidemiology of hepatitis C virus. JAMA 263: 49-53, 1990.
[7.] Alter, M.J.; Coleman, P.L.J.; Alexander, J.; et al. Importance of heterosexual activity in the transmission of hepatitis B and non-A, non-B hepatitis. JAMA 262: 1201-1205, 1989.
[8.] Estaban, J.I.; Vilamoniu, L.; Gonzalez, A.; et al. Hepatitis C virus antibodies among at-risk groups in Spain. Lancet 2: 294-296, 1989.
[9.] Gambino, R. NANB hepatitis: A new antibody test for the hepatitis C virus. Lab Report for Physicians 10(12): 89-93, December 1988.
[10.] Kew, M.C.; Houghton, M.; Choo, Q.L.; et al. Hepatitis C virus antibodies in southern African blacks with hepatocellular carcinoma. Lancet 335: 873-874, April 14, 1990.
[11.] Bruix, J.; Barrera, J.M.; Calvet, X.; et al. Prevalence of antobodies to hepatitis C virus in Spanish patients with hepatocellular carcinoma and hepatic cirrhosis. Lancet 2: 1004-1006, 1989.
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|Author:||Soloway, Henry B.|
|Publication:||Medical Laboratory Observer|
|Date:||Nov 1, 1990|
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