Viral hepatitis: Targeted tests and therapies contribute to improved outcomes

An estimated 4.4 million Americans are living with chronic viral hepatitis—the leading cause of liver cancer—even though most do not know they are infected, according to the Centers for Disease Control and Prevention (CDC). About 80,000 new infections occur each year.

The growing hepatitis epidemic presents a variety of clinical and laboratory challenges, yet relatively recent developments in diagnostic testing and targeted therapies offer promising solutions. The emergence of a personalized healthcare approach to patient management may ultimately lead to more cost-effective treatment protocols and even help for patients who previously had no hope for a cure.

Type prevalence and routes of transmission

In the United States, viral hepatitis is usually caused by one of five distinct viruses; hepatitis A, B, C, D, or E. The majority of viral hepatitis cases in the U.S. are caused by hepatitis A (HAV), hepatitis B (HBV) or hepatitis C (HCV). Hepatitis D and E viruses are more common in Far East Asia.

Routes of transmission differ among the viruses. HAV is transmitted through fecal-oral route exposure, and results in approximately 25,000 new infections in the U.S. annually. Rates have declined by 92% since hepatitis A vaccine became available in 1995. HBV is transmitted by percutaneous or mucosal exposure to blood or body fluids from an infected individual. Approximately one million individuals in the U.S. are chronically infected with HBV, with an estimated 43,000 new infections annually. HCV is transmitted primarily through percutaneous or mucosal exposure. In the U.S., an estimated 3 to 4 million individuals are chronically infected with HCV and about 16,000 new HCV infections occur annually.^1,2

While treatment protocols vary by virus type, an increasing variety of targeted diagnostic tests and therapies are available to help healthcare professionals adopt a more personalized approach to patient care, improve outcomes, and manage the growing epidemic.

Immunoassay diagnostic testing

There are numerous serologic markers available for the diagnosis of suspected viral hepatitis. Non-specific markers of hepatic inflammation include elevation of hepatic transaminases—alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Both of these enzymes will be elevated in the serum of a patient with acute viral hepatitis. Elevation of these enzymes may also be present in chronic viral hepatitis.

Acute hepatitis A. The serologic diagnosis of acute hepatitis A depends on the detection of hepatitis A antibody (HAV IgM) in the patient’s serum. The presence of HAV IgM indicates recent exposure to the hepatitis A virus. HAV IgM becomes detectable in the blood within two weeks of exposure, persisting at high levels for approximately 13 weeks. Levels of HAV IgM will then gradually decline to undetectable levels at six months post-exposure.³ After the disappearance of HAV IgM, HAV IgG becomes detectable. The HAV IgG indicates prior infection and immunity. Hepatitis A is a self-limited illness with symptomatic management. Unlike hepatitis B and C, there is no chronic form of infection of hepatitis A.⁴

Acute hepatitis B. In acute HBV infection, hepatitis B surface antigen (HBsAg), a protein on the surface of the hepatitis B virus, appears before the onset of clinical symptoms and can be detected as soon as one month after exposure to the virus, but detection can range for up to 24 weeks. The antibody to HBsAg (Anti-HBs) becomes detectable at approximately six weeks after exposure and remains detectable indefinitely. The appearance of anti-HBs with the absence of HBsAg indicates the resolution of infection and protective immunity. Hepatitis B core antibody (anti-HBc) appears within one to two weeks after the appearance of HBsAg, and before detectable levels of anti-HBs.

Due to the variability in the time of the appearance of anti-HBs after HBV infection, a gap of several weeks (weeks 16-32) may separate the HBsAg and the appearance of anti-HBs. This gap in time is referred to as the “window period.” During the window period, anti-HBc may provide the only serologic evidence of current or recent HBV infection. Determining the immunoglobulin class of anti-HBc levels can also help clinicians distinguish recent HBV infection from remote infection. IgM anti-HBc predominates during the first six months after acute infection, and IgG anti-HBc predominates beyond six months after the initial exposure to HBV.

Another serologic marker of HBV infection, hepatitis B e antigen (HBeAg), becomes detectable in the patient’s serum around the same time as HBsAg. The appearance of HBeAg is indicative of high levels of viral replication and high infectivity. HBeAg levels decline and become undetectable shortly after the aminotransferase levels (ALT and AST) peak. After the HBeAg levels become undetectable, antibody to hepatitis B e antigen (anti-HBe) becomes detectable at approximately week 16. The appearance of anti-HBe is indicative of lower infectivity. Approximately 90% of immunocompetent adults will initiate a rigorous immune response resulting in self-limited hepatitis (Figure 1).

Chronic hepatitis B. In chronic HBV infection, HBsAg remains detectable beyond six months after the initial infection, IgG anti-HBc continues to be detectable, and anti-HBs is either undetectable or detectable at low levels. In the early phase of chronic HBV infection, HBV DNA can be detected in the patient’s serum. This is typically referred to as the replicative phase of chronic HBV infection. Replicative chronic HBV infection is characterized as a period of maximal infectivity and liver injury. In chronic HBV infection, HBeAg is a qualitative marker and HBV DNA is a quantitative marker. Eventually patients convert from a replicative phase to a non-replicative phase of chronic HBV infection. This occurs at a rate of approximately 10% per year, and is accompanied by seroconversion from HBeAg positive to anti-HBe positive. This seroconversion is often accompanied with a transient elevation of aminotransferase activity that is thought to be due to cell-mediated immune clearance of virus-infected hepatocytes. In the non-replicative phase of chronic HBV infection, liver injury tends to diminish, and these patients are referred to as inactive HBV carriers. HBV infection can trigger a sequence of events which may result in hepatocellular carcinoma⁵ (Figure 2).

Acute Hepatitis C. Patients with acute HCV infection are typically asymptomatic and infection is commonly not recognized. Only 20% to 30% of patients with acute hepatitis C infection experience symptoms of fatigue, abdominal pain, anorexia, and jaundice. A hepatitis C antibody (anti-HCV) test detects antibodies to HCV. Anti-HCV becomes detectable approximately four to 10 weeks after initial HCV infection. The presence of hepatitis C antibody does not distinguish between acute or chronic infection. Anti-HCV is typically used as a screening test for hepatitis C infection. Because most patients infected with hepatitis C are asymptomatic, the diagnosis is typically made through screening of blood donations or when elevations in aminotransferase levels (ALT or AST) are discovered. In the chronic state, HCV RNA stays present in the blood and ALT does not drop to normal levels. The natural course of chronic hepatitis C varies from person to person. The HCV recombinant immunoblast assay (RIBA) can be used as a confirmatory test; however it is no longer regularly used in clinical practice. The RIBA test has primarily been replaced by molecular tests for HCV RNA using polymerase chain reaction (PCR) technology.⁶

Molecular tests for screening, diagnosis and monitoring

A number of molecular diagnostic tests are available for donor blood screening and for diagnosis and monitoring of patients with viral hepatitis B or C.

Hepatitis B. A hepatitis B DNA quantitative test uses PCR technology to measure HBV DNA in the serum or plasma of infected patients. The presence of HBV DNA is a reliable marker of active HBV replication. HBV DNA levels are detectable within 30 days following infection, generally reach peak levels at the time of acute symptomatic hepatitis, and gradually decline and disappear as the infection spontaneously resolves. In cases of acute viral hepatitis, testing with HBV DNA in serum may be a helpful adjunct in the diagnosis of acute HBV infection, since HBV DNA can be detected approximately 21 days before HBsAg appears in the serum.

In chronic HBV infection, HBV DNA levels in serum are useful in determining the status of the HBV infection. Patients with chronic HBV infection fail to clear the virus and remain HBsAg positive. Patients can be further distinguished as having chronic active (replicative) HBV infection by the presence of high HBV DNA levels and a positive HBeAg; or having chronic inactive (non-replicative) HBV infection by the presence of low or undetectable HBV DNA levels and a negative HBeAg. Patients with chronic active HBV infection are at greater risk for severe liver damage and are more infectious than patients with inactive HBV infection. Additionally, HBV DNA is the only reliable marker of active HBV replication with reactivation of inactive chronic HBV infection when HBeAg remains negative. The ultimate goal in treating patients with chronic HBV infection who are HBeAg positive is seroconversion to HBeAg negative and long-term viral suppression demonstrated by undetectable HBV DNA. In patients who are HBeAg negative, the therapeutic goal is long-term viral suppression.

Real-time PCR methods have a broad dynamic range and can quantify HBV titers from single IUs to tens of millions of IUs. Results may be reported as “detected” with the comment “HBV DNA level is below the limit of quantification” indicating the assay cannot accurately quantify HBV below a predetermined level. A quantitative result provided in IU/mL and log (10) IU/mL is indicative of the degree of HBV viral replication. Monitoring HBV DNA levels enables a healthcare provider to assess the patient’s disease progression and response to therapy.⁷

Hepatitis C. Molecular testing for hepatitis C RNA includes both qualitative and quantitative Nucleic Acid Tests (NATs). Qualitative tests for HCV RNA can be used to confirm viremia (especially low-level viremia) and to screen blood donations. The use of nucleic acid testing for HCV among blood donations has dramatically reduced the incidence of post-transfusion Hepatitis C. Quantitative HCV RNA tests are used clinically to guide therapy in the treatment of patients with chronic HCV infection. The rate of virologic response is an important parameter to monitor during treatment. Monitoring patients for changes in HCV RNA levels (viral load) after four weeks and 12 weeks of therapy predicts the likelihood for sustained virologic response (SVR), which is considered the gold standard.⁸ It is defined as the absence of HCV RNA from serum by a sensitive PCR assay 24 weeks following discontinuation of therapy. This is generally regarded as a “virologic cure” (Figure 3).

Genotyping and the evolution of personalized medicine

Hepatitis genotyping, which plays a fundamental role in the development of targeted therapies and personalized medicine, is performed by either reverse-hybridization line probe assay, direct sequencing test, or restriction fragment length polymorphism assay. These assays have been found to have a high degree of concordance and reliability. Genotypes can be further categorized into subtypes and quasi-species for more detailed characterization, although this is largely limited to research settings for epidemiological purposes.

The emergence of genotype characterization and targeted therapies has led to an evolution in hepatitis C treatment in recent years. Six hepatitis C genotypes have been described, with genotype 1 being the most prevalent (74%) in the U.S. In the course of patient management, quantitation of HCV genotypes 1-6 can be measured by HCV RNA levels at baseline and during antiviral treatment. The assay can be utilized to predict sustained and nonsustained virological response to HCV therapy.

Unfortunately, the most prevalent HCV genotype (1) has traditionally been the least responsive to antiviral therapy. With the approval of the protease inhibitors boceprevir and telaprevir for the treatment of chronic HCV infection in 2011, the treatment guidelines have been updated. Many patients who previously failed treatment, whether they were discontinuers, relapsers, or non-responders, now have new opportunities for a cure. New combinations of standard peginterferon and ribavirin therapy are frequently given in combination with boceprevir or telaprevir for patients infected with HCV genotype 1. These new regimens require close monitoring of HCV RNA levels to assess response to treatment.⁹

As a result, new treatment guidelines will likely result in more frequent testing of patients to assess responses to therapy and evidence of viral resistance to the protease inhibitors. In addition, improved hepatitis diagnostics may lead to a greater number of people being identified as candidates for targeted antiviral therapies, which may further drive up the overall volume of laboratory testing and potentially shift the assay mix within the lab.

Overcoming laboratory challenges

As this potential increase in demand for hepatitis testing looms on the horizon, labs already face several challenges in managing heavy test volumes.

Despite strong commitment from the laboratory community, a widespread lack of staffing resources sometimes leads to infrastructure-related issues like the absence of standardized laboratory policies, limited strategic planning, inadequate synergies between clinical and research laboratories, and limited staff productivity. All of these factors can potentially compromise the quality of test results and impact patient management.

Careful, strategic planning on the part of labs will enable them to manage growth in demand successfully and continue to provide significant value to ordering physicians and their patients. One critical strategy, for example, is prudent inventory management, including the careful selection of assay menus, efficient management of reagents, and performance of scheduled equipment maintenance. Another strategy is improving workflow efficiency for greater staff productivity. To help compensate for shortages in staffing, labs can adopt automated platforms for both immunoassay and molecular testing. For example, molecular systems are available that can conduct viral load testing for hepatitis B, hepatitis C and HIV in parallel, simultaneously, on a single automated platform.

As the viral hepatitis epidemic continues to present its own challenges in the U.S. and globally, careful strategic planning can help labs take advantage of the opportunity—with the advent of new immunoassay and molecular tests—to contribute to better diagnostics and patient management using targeted hepatitis therapies.

Detection, quantification, and genotyping tests are clinically important and help healthcare providers determine the most effective patient management strategy. Management of hepatitis with a personalized healthcare approach may give some patients new opportunities for a cure, allow others to discontinue medication when therapy has a low probability of efficacy, and increase the likelihood of successful treatment outcomes—in the process helping to reduce the overall costs of healthcare.

References

1. CDC. Surveillance for acute viral hepatitis—United States, 2007. Surveillance Summaries, May 22, 2009. MMWR 2009;58 (No. SS-3).
2. CDC. Viral hepatitis surveillance United States, 2009. http://www.cdc.gov/hepatitis/Statistics/2009Surveillance/PDFs/2009HepSurveillanceRpt.pdf. Accessed February 2012.
3. Mayo Clinic, Mayo Medical Laboratories. http://www.mayomedicallaboratories.com/test-catalog/print.php?unit_code=8342. Accessed February 2012.
4. American College of Physicians. MKSAP 15. Gastroenterology and Hepatology:60.
5. Dienstag JL. Chapter 304. Acute viral hepatitis. In: Longo DL, et al, eds. Harrison’s Principles of Internal Medicine. 18th ed. New York: McGraw-Hill;2012. http://www.accessmedicine.com/content.aspx?aID=9133433. Accessed February 2012.
6. CDC. Hepatitis C information for health professionals. http://www.cdc.gov/hepatitis/hcv/hcvfaq.htm#c5. Accessed February 2012.
7. Mayo Clinic. Mayo Medical Laboratories. http://www.mayomedicallaboratories.com/test-catalog/print.php?unit_code=88634. Accessed February 2012.
8. Scott JD, Gretch DR. JAMA. 2007;297(7);724-732. http://jama.ama-assn.org/content/297/7/724.full. Accessed February 2012.
9. Ghany MG, Nelson DR, Strader DB, Thomas DL, Seeff LB. An update on treatment of genotype 1 chronic hepatitis C virus infection: 2011 practice guideline by the American Association for the Study of Liver Diseases. Hepatology. 2011:54:1433-1444.doi: 10.1002/hep.24641.

Johnnie A. Lee, MD, MPH, FACP, Medical Director, Medical and Scientific Affairs; Michelle Payette, MD, Medical and Scientific Affairs Manager; and John Osiecki, PhD, Medical and Scientific Affairs Manager, at Roche Diagnostics in Indianapolis.