Asian Pacific Association for the Study of the Liver consensus statements on the diagnosis, management and treatment of hepatitis C virus infection

Abstract

Past exposure to hepatitis C virus (HCV) is mostly determined by testing for specific antibodies using an approved enzyme immunoassay (EIA). The presence of antibody shows that the patient has been infected with the virus but does not indicate whether the infection is acute, chronic or resolved. The absence of antibody usually shows that the patient has not been infected. However, antibody might not be detectable in the first few weeks after initial infection (known as the window period) or in patients who are immunosuppressed. Furthermore, there is some evidence that in patients who resolve their infection, antibody levels might decrease and become undetectable many years later. Several countries in the Asia–Pacific region have developed their own individual testing algorithms for anti-HCV testing. For confirmatory testing, some of these approaches include: Repeat testing of reactive samples in the same EIA Retesting reactive samples in a second, independent EIA Testing by immunoblot Presumption that a high signal-to-cut-off ratio for a sample in a specific EIA is highly predictive of an authentic anti-HCV positive result Use of a nucleic acid test (NAT) for detection of HCV RNA. Note that although a NAT for HCV RNA can be helpful in diagnosis, it cannot be considered a true confirmatory test. Ideally, all samples shown to be anti-HCV reactive should be retested using an assay with high specificity to confirm reactivity. However, for some laboratories financial constraints can preclude this approach. Anti-HCV testing is important for determining exposure to the virus but does not identify whether the patient has current infection. However, this information can be provided by an appropriately performed NAT for HCV RNA. Qualitative testing for HCV RNA can offer some important advantages, including: Determination of chronicity Monitoring response to antiviral therapy Assessment of anti-HCV indeterminate samples. Testing for the presence of HCV RNA should be strongly considered in patients at high risk of infection but who might be anti-HCV negative or indeterminate because of immunosuppression (by therapy or disease, such as patients on hemodialysis or with HIV infection). HCV RNA isolation is also necessary for determination of HCV genotypes. A number of HCV genotype classification schemes have been used. In the most recent, HCV has been classified into six major genotypes, which can be further divided into subtypes. Some genotypes such as HCV 1, 2 and 3 are widely distributed, although others are more geographically restricted. Interferon (IFN)-based therapy has become the mainstay of chronic HCV treatment and improved outcomes have been achieved as knowledge is gained about the predictors of response to therapy. Virus genotype and viral load have been shown to be key viral characteristics to guide treatment and clinical management of patients with chronic infection. Several methods are available to determine HCV genotype. The method used will vary from country-to-country and might depend on approval by relevant health authorities and/or available funding. Methods include: Direct sequencing of PCR product (region amplified could be 5′ untranslated region (UTR), core, NS5A and NS5B) Reverse-phase hybridization (e.g. line probe assay) Type-specific PCR Restriction fragment length polymorphism after PCR amplification Melting curve analysis after real-time PCR amplification Typing using genotype-specific antibodies Restriction fragment mass polymorphism analysis by mass spectrometry. The 5′ UTR is well conserved but has sufficient nucleotide sequence divergence to discriminate between most genotypes. It is the target region for most diagnostic HCV RNA PCR assays and genotyping based on the 5′ UTR has a high concordance with genotype determined by sequencing of NS5B. A possible disadvantage of relying on the 5′ UTR for genotype determination is its inability to discriminate the HCV genotype 6c-l, highly prevalent in the Asia–Pacific region, which can be mistyped as HCV genotype 1/1b because of sequence homology. Sequence analysis of the core region is sufficient to identify the HCV genotype 6c-l and a new generation line probe assay designed with core-specific oligonucleotides is also being evaluated. At present, the inability to distinguish HCV genotype 6c-l might impact on predicted sustained virological response (SVR) in patients with apparent genotype 1/1b who are to receive IFN-based therapies. Currently trials are underway to evaluate optimal treatment duration for genotype 6. The final use of HCV RNA isolation is that of viral load estimation. There does not appear to be any association between disease activity, progression to chronicity and HCV viral load. However, viral load has been shown to be a prognostic indicator of therapy outcome. Monitoring of viral load on therapy has also proven useful, especially for patients infected with HCV genotype 1, with the lack of an early virological response predictive of long-term treatment failure. HCV load testing for patients infected with HCV genotypes 2 or 3 is not recommended as nearly all have an early virological response and a qualitative NAT for HCV RNA is preferred. Commercial signal amplification and target amplification assays are available for quantification of HCV RNA. In addition, several laboratories have developed their own in-house load assays. Traditional end-point PCR for viral load has several disadvantages, in particular a limited dynamic range, so laboratories persisting with in-house testing should adopt the real-time PCR format. Such assays can be calibrated to the World Health Organization (WHO) International Standard. Most recently, commercial HCV core antigen assays have become available. In some circumstances these might be an alternative to HCV RNA assays. The HCV core antigen assays show a good correlation with HCV RNA assays but due to limited sensitivity, they are probably not suited to the monitoring of patients on therapy. The major role these assays might play is in the identification of blood donors in the seroconversion window. Consensus statements: HCV infection and laboratory testing Anti-HCV antibody testing should be by approved anti-HCV third or fourth generation EIA. (II-2)* Samples negative in an approved EIA can be reported as anti-HCV negative. However, it should be noted that individuals on hemodialysis or coinfected with HIV might be HCV RNA positive but anti-HCV negative. (II-2) Samples reactive in an approved single EIA can be reported as anti-HCV positive provided the signal-to-cut-off ratio is sufficiently high to be predictive of a true positive.# (III) For samples that do not reach this threshold or have reactivity close to the cut-off a qualitative NAT for HCV RNA should be considered and/or a further follow-up sample be obtained for both anti-HCV and HCV RNA NAT. (III) HCV RNA qualitative and quantitative testing requires appropriate contamination controls. (II-2) A dedicated sample/aliquot not derived from other test samples is preferred for HCV RNA qualitative and quantitative (viral load) testing. (II-2) HCV RNA quantitation should be reported in IU/mL (optional to include copies/mL).** (III) HCV genotype testing is important for assessing treatment duration and efficacy of antiviral therapy. However, it should be recognized that genotype discrimination based on primers from the 5′ UTR do not distinguish some of the genotype 6 subtypes prevalent in South-East Asia and instead classifies them as genotype 1 or 1b. (II-2) Participation in an external quality assurance program for all testing is ideal. (II-2) Internal quality assurance testing is required for all testing. (II-2) *Numbers in parentheses refer to levels of evidence.9 # An option, which can be used as an alternative to a secondary confirmatory assay, is to use the initial screening assay signal-to-cut-off ratio (S/CO) to estimate the probability of a patient's true antibody status. The ratio is calculated by dividing the optical density (OD) value of the test sample by the OD of the assay cut-off.An example of the calculation of the ratio:Sample OD = 2.991, cut-off OD = 0.377, therefore S/CO ratio = 7.934.For most standard EIAs (i.e. those that do not use chemilumescence), an S/CO ratio greater than 3–4 should be indicative of the presence of true anti-HCV antibodies. Such tests include:• Murex Anti-HCV (version 4.0)• Bio-Rad Monolisa Anti-HCV Plus Version 2• Ortho HCV Version 3.0 ELISA Test System.Choosing the appropriate S/CO ratio for a laboratory will depend on several factors, including the type of EIA, the prevalence of anti-HCV in the population being tested and degree of confidence required in the positive predictive value. For example, using the Ortho HCV Version 3.0 ELISA Test System, the Centers for Disease Control and Prevention (CDC) found that an S/CO ratio of ≥3.8 was predictive of a true anti-HCV result ≥95% of the time, regardless of the population being tested.8 **The high sensitivity of amplification technologies must be counterbalanced by the potential for contamination leading to false positive results. The possibility of contamination cannot be eliminated but good laboratory design and work practices can minimize the chances of it occurring.The greatest potential source of contamination is amplified material generated from the previous PCR; however, the possibility of contamination from the nucleic acid extraction step must also be considered.Ideally, separate rooms should be used for each step of the PCR process: PCR reagent preparation (“clean” room), nucleic acid extraction, amplification and post-PCR analysis. In general, the use of commercial HCV RNA assays and the use of real-time PCR make the requirement for a post-PCR area redundant, provided the amplified material is disposed of correctly. Each area should have dedicated gowns, gloves and equipment (e.g. pipettes and centrifuges). Aerosol barrier tips (or equivalent) should be used for all steps, including the reagent preparation. A workflow of reagent preparation to extraction to amplification should be followed; if returning against the workflow, gowns and gloves need to be changed.The PCR reagent preparation needs to take place in a clean room but this could be a shared facility (e.g. media preparation) so long as an extraneous source of contamination is unlikely to be present. The extraction area should have a biological safety cabinet to carry out extractions not only as a contamination prevention measure but also as the samples might themselves represent an exposure hazard to the worker. Aliquotting of reagents both in the PCR reagent preparation and extraction area will minimize problems should contamination be found to occur. The extracted material can be added to the PCR reagents in this room and then transferred to the thermal cycler in the amplification area.It has been recommended that for recycling of equipment (e.g. test-tube racks) and for the cleaning of workbenches that a 1/10 dilution of standard bleach be used. It is likely that cleaning with a germicidal detergent is equally effective. World Health Organization estimates that as many as 170 million persons worldwide might be infected with HCV. In Asia, the estimated figures range from 0.3% of the population in New Zealand to 4% in Cambodia. Data for the Pacific region are difficult to obtain, but estimates of an HCV antibody rate of up to 4.9% have been recorded in some parts of the Pacific. In the Middle East, levels of 12% have been reported in some centers. Data available on the incidence of new cases of hepatitis C are scanty because of the difficulty in differentiating between new cases and the initial diagnosis of chronically infected subjects. As the relative importance of the various modes of transmission of HCV varies from country to country, the incidence of new cases will also vary. Better data are required to confirm any changes in incidence. However, there is little doubt that the epidemiology of HCV infection in the Asia–Pacific region is changing. In the new millennium, with the introduction of universal screening of blood products and the abolition of paid blood donation, injecting drug use (IDU) has become the most common route of HCV transmission. In Australia, IDU is the admitted risk factor for chronic HCV infection in 65% of patients seen in hospital clinics. Furthermore, IDU is considered responsible for nearly all new infections. Some data indicate that the risk of transmission of HCV can be reduced by harm reduction methods (e.g. needle exchange programs and public education). However, the high prevalence of IDU in the community has meant a continuing high incidence of new HCV infections. Despite the illegal status of illicit drug injection, consideration should be given to more widespread application of harm reduction strategies. The data concerning risk factors for transmission of hepatitis C in other regions in Asia are relatively sparse. In Japan, the lower prevalence of hepatitis C in younger compared to older people suggests that the incidence of HCV infection is decreasing. The use of traditional therapies (including acupuncture, folk remedies and Suidama) has been a major source of past transmission for HCV in some regions of Japan and Taiwan. In some countries, tattooing, including eyebrow tattooing in Vietnam and Cambodia, might be an important risk factor for transmission of the infection. There is evidence that medical practices such as using non-disposable glass syringes and needles have been an important mode of HCV transmission. Transfusion of blood and blood products prior to HCV screening have also been a major mode of HCV transmission. However, the introduction of universal blood donor screening can virtually eliminate post-transfusion hepatitis C. The lack of universal screening of blood donors in some areas of Asia is still responsible for new cases of hepatitis C in these areas. Some contemporary medical practices still carry a risk of HCV transmission from blood contamination, particularly hemodialysis. Such transmission can be prevented with the use of universal infection control precautions. Mother-to-baby spread has been demonstrated in approximately 7% of HCV RNA-positive mothers. Although there have been a few studies on the prevalence and risk of mother-to-infant transmission of HCV from the Asia–Pacific region, the different definition of mother-to-infant transmission used in these studies makes comparison of the data difficult. Possible criteria for more rigorous definition of mother-to-infant transmission of HCV infection include: (i) detectable anti-HCV in an infant aged older than 18 months; (ii) detection of HCV RNA in an infant aged 3–6 months of age; (iii) detection of HCV RNA in an infant on at least two occasions; (iv) elevated serum amino transaminases in an infant; or (v) identical genotype between mother and child. Co-infection with HIV and high blood levels of HCV-RNA correlate with the risk of perinatal transmission. Long-term follow-up studies have shown a low prevalence of HCV-related clinical signs and symptoms among vertically infected children in the first 10–15 years of life. Approximately 20% of children appear to clear the infection, 50% have evidence of chronic asymptomatic infection, and about 30% have evidence of chronic infection with elevated transaminase levels. Sexual transmission does occur, but is rare. There are few data regarding prevention in this situation, but concurrent sexually transmitted diseases and sexual practices that could involve blood contamination might increase the risk of HCV transmission. Common sense recommendations to reduce spread of HCV in the household setting (such as not sharing razors, avoiding blood contaminations) and by sexual transmission have been referred to in previous authoritative documents. Consensus Statements: Prevention of HCV infection All countries must introduce universal screening of blood donors for hepatitis C antibody (anti-HCV) with third or fourth generation EIAs. More data on the cost-effectiveness of nucleic acid testing for universal screening of blood products is required. (II-2) In healthcare settings, adherence to universal precautions for infection control is essential. This should include use of disposable or adequately sterilized materials for invasive procedures, and adequate cleansing and sterilization of instruments. (II-2) As transmission of HCV via IDU is an increasing trend in the Asia–Pacific region, it is important to implement an education campaign about the harm of drug use, especially among school-age children. Harm reduction programs such as needle syringe programs should also be implemented. It is important to educate tattooists and practitioners of traditional or alternative therapies about ways to minimize blood contamination. This involves sterilization techniques for procedures that involve skin penetration or breaks to mucosal surfaces. (II-2) The natural history of hepatitis C is quite variable. There are some inherent drawbacks in studying natural history. Firstly, it is difficult to ascertain the exact time of acquirement of infection; second, primary infection is commonly asymptomatic and thirdly disease progression is slow. Natural history data reported in the literature vary according to the type of study (retrospective vs prospective). Different study populations also result in different predictions about natural history (patients attending liver clinic vs blood donors vs community based studies vs post-transfusion cohorts). In spite of these variations there are some generalizations that can be made. In acute HCV infection: 20–30% of patients are symptomatic Fulminant hepatic failure is very uncommon Elevation in serum alanine aminotransferase (ALT) levels occurs approximately 2–8 weeks after exposure HCV RNA can be detected in serum within 1–2 weeks after exposure HCV RNA levels increase progressively and peak before ALT rise and development of symptoms. 20–50% of patients might clear the virus spontaneously. Symptomatic patients and women are more likely to clear the virus Most patients who clear infection do so within the first 12 weeks 50–80% of patients will develop chronic infection. In chronic HCV infection: Up to 20–30% of patients will develop a progressive liver disease leading to cirrhosis and hepatocellular carcinoma (HCC) Cirrhosis rates begin to become significant after 20 years of infection HCC rates begin to become significant after 30 years of infection Factors associated with disease progression include duration of infection, age at the time of acquirement of infection, sex, alcohol consumption, immunosuppression (e.g. HIV coinfection or organ transplant recipients), obesity and insulin resistance, coinfection with other viruses, elevated aminotransferases and genetic factors Although elevated ALT suggests active liver damage, normal ALT does not exclude significant liver disease Progression to cirrhosis can be best predicted on baseline histological parameters such as the activity of necroinflammation and stage of fibrosis Once patients develop cirrhosis, HCC develops at approximately 1–4% per year and is increased in patients with raised α-fetoprotein levels at baseline. Consensus Statements: Natural history of HCV infection Acute hepatitis C is a well-recognized entity. In the stage of acute hepatitis patients should be monitored for spontaneous viral clearance. Patients with symptomatic acute hepatitis and female sex are more likely to clear the virus. In chronic HCV infection, elevated serum ALT suggests progressive liver damage. However, normal ALT does not exclude significant liver disease. A fibrosis score (Metavir score >2 or Ishak score >3) suggests progressive liver disease. In chronic HCV infection, it is well recognized that excessive alcohol and insulin resistance are associated with disease progression. It is recommended that patients consume less than the WHO guidelines for alcohol intake. It is recommended that obesity and insulin resistance be controlled through exercise and dietary intervention to achieve ideal BMI. (II-2) In patients with HCV related liver cirrhosis, risk of hepatic decompensation is approximately 3–4% per year; 1.4–6.9% per year for HCC. In patients with well-compensated HCV cirrhosis the 10-year survival rate is 80%. However, if there are features of decompensation, the survival rate is significantly reduced, i.e. to approximately 25%. HCC is a frequent and life-threatening complication of chronic HCV infection. In cirrhotic patients, a surveillance program for the early detection of HCC should be offered. (II-2) IFN therapy impacts positively on the natural history of HCV-related liver cirrhosis. Among sustained virological responders, the rate of decompensation at 5 years is 1%. In the biochemical responders, the 5-year rate of decompensation is 9.1%. (II-1) Before a discussion of specific therapies, some general points need to be made. The desired end-point of treatment of HCV infection is viral clearance, as indicated by non-detectability of HCV RNA in serum by the most sensitive test available. Rapid virological response (RVR) is defined as non-detectability of serum HCV RNA (<50 IU/mL) after 4 weeks of therapy. Early virological response (EVR) is defined as undetectable HCV RNA (<50 IU/mL) or at least a 2 log decrease in serum HCV RNA from baseline level after 12 weeks of therapy. Studies using pegylated (peg)-IFN showed that 65–72% of subjects with EVR went on to develop a SVR. End-of-treatment virological response (ETVR) is indicated by non-detectability of HCV RNA at the end of therapy. Sustained virological response is defined as undetectable serum HCV RNA (<50 IU/mL) 24 weeks after the end of therapy. Low viral load (LVL) is defined as serum HCV RNA < 400 000 IU/mL High viral load (HVL) is defined as serum HCV RNA > 400 000 IU/mL. An SVR is the best correlate of beneficial changes in hepatic fibrosis, prevention of HCC and improvement in other clinical outcomes. SVR has been shown to have the following beneficial effects: (i) fibrotic regression; (ii) substantially reduced rate of HCC; (iii) decreased rate of other complications, including liver failure and liver-related death; and (iv) improved quality of life. Alcohol intake should be discouraged during treatment. Hepatitis A and B immunization should be advised in patients not immune to hepatitis A virus and hepatitis B virus (HBV). With improved treatment results, the value of liver biopsy is being questioned because of the potential risks of the procedure and concerns regarding sampling error. Clinical, biochemical and imaging findings can identify many patients with advanced cirrhosis, but not those with lesser degrees of fibrosis. A liver biopsy would be useful in this latter group of patients. Although liver fibrosis markers are commercially available, they are currently insufficiently accurate to support their routine use. Specific issues regarding therapy in acute and chronic HCV infection will now be addressed. In acute HCV infection, serum HCV RNA is usually detected before the appearance of anti-HCV and is often the only diagnostic indicator of this condition. Acute hepatitis C infection often becomes chronic, especially in asymptomatic individuals. However, up to 50% of patients who presented with symptoms can spontaneously resolve their infection. Female sex and infection by HCV genotype non-1 increase the chance of spontaneous resolution. Spontaneous resolution is less likely after 12 weeks of infection. Treatment of hepatitis C in the acute stage has resulted in better SVR rates than treatment in the chronic stage. The objective of antiviral treatment in acute hepatitis C is to prevent the development of chronic hepatitis C. Studies using daily induction doses of conventional IFN-α followed by three times weekly IFN-α, as well as those using peg-IFN-α for 24 weeks, have achieved high rates of SVR in acute hepatitis C. Peg-IFN-α has been found to be superior to conventional IFN-α plus ribavirin. Addition of ribavirin to IFN-α or peg-IFN-α has not resulted in significant improvement in SVR rates. HCV genotypes 2, 3 and 4 respond better than HCV genotype 1 and treatment time can be reduced to 12 weeks with peg-IFN-α in subjects infected with these HCV genotypes. Prophylactic IFN is not recommended in needle stick injuries because of low overall infectivity rate. The objective of antiviral treatment for chronic hepatitis C infection is to prevent liver-related complications, including HCC. The following have been shown to influence treatment outcome: (i) age; (ii) sex; (iii) virus genotype; (iv) virus load; and (v) stage of fibrosis, especially F3, F4. The response to IFN-α plus ribavirin of patients with normal serum ALT levels is similar to that of patients with raised ALT levels and these patients should not be denied therapy. Patients with no (F0) or minimal (F1) hepatic fibrosis do not necessarily need antiviral therapy. However, treatment should be considered for those who have disabling symptoms or a higher grade of activity on liver biopsy, and for those persons who wish to be treated regardless. As for all patients, they should receive advice concerning: The natural history of their disease, especially the likelihood and projected timing of any possible liver related complications. The efficacy of available treatments. The cost of available treatments. The adverse effects of available treatments, and need for ongoing contraception after administration of ribavirin. The success rate of obtaining an SVR following HCV treatment has improved since the last APASL consensus statement in 2000. In randomized controlled trials, the highest overall SVR rates have been achieved with the combination of once a week subcutaneous injection of long acting peg-IFN-α combined with daily oral ribavirin for 1 year. This is the current standard treatment, especially for patients infected with HCV genotype 1. Peg-IFN is produced by binding polyethylene glycol to IFN molecules resulting in slower absorption from subcutaneous sites and decreased renal clearance leading to increased half-life. There are two licensed peg-IFN-α products available for use in the Asia–Pacific region. Two large phase 3 studies using weekly doses of the two different peg-IFN-α, both combined with daily ribavirin, given for 48 weeks have demonstrated improved efficacy with higher SVR rates when compared to three times weekly standard IFN plus daily ribavirin combination. Patients infected with HCV genotypes 2 and 3 had SVR rates over 80%. A study showed that 6 months' treatment is sufficient for persons infected with HCV genotypes 2 and 3. It is recommended that patients infected with HCV genotype 1 be treated for 1 year and that those infected with genotypes 2 or 3 be treated for 6 months. Because of small numbers of patients with HCV genotypes 4, 5 and 6 in the various studies, the most appropriate duration of therapy for persons infected with these HCV genotypes is unknown. Patients with LVL have an increased SVR. Recent data showed that patients with low levels of HCV genotype 1 who lose their serum HCV RNA after one month of therapy (rapid virological response; RVR) would need only 6 months of treatment with the standard combination therapy. However, data are conflicting on whether patients with HCV genotype 2 or 3 with LVL who achieved loss of serum HCV RNA after 1 month of treatment should be considered for 12–16 weeks of therapy. Any positive data regarding this applies more to patients with HCV genotype 2 than genotype 3. The recommended dose of peg-IFN-α2a is 180 μg weekly and the recommended dose of peg-IFN-α2b is 1.5 μg/kg bodyweight. Ribavirin doses of 1000 mg daily are recommended for persons up to 75 kg in weight and 1200 mg for persons more than 75 kg in weight. There are few absolute contraindications for use of peg-IFN-α and ribavirin (RBV). They include: Present or past psychosis or severe depression Uncontrolled seizures Hepatic decompensation Pregnancy (RBV) Renal failure (RBV) Severe heart disease (RBV). The relative contraindications for IFN and ribavirin are: History of depression Uncontrolled diabetes mellitus Uncontrolled hypertension Retinopathy Psoriasis Autoimmune thyroiditis or other active autoimmune disorders including autoimmune hepatitis Symptomatic heart disease or severe vascular disease (RBV) Anemia/ischemic vascular disease (RBV). In addition to these contraindications, special caution is required if IFN is administered in the following circumstances: Neutropenia (neutrophil count <1500 cells/µL3) Thrombocytopenia (platelet count <85 000/µL3) Organ transplantation History of autoimmune disease Presence of thyroid autoantibodies Age older than 70 years. Testing prior to starting IFN with ribavirin treatment is indicated to identify those who might be at special risk of adverse effects, and monitoring during therapy is recommended mainly to prevent serious adverse events. The incidence and types of side-effects of peg-IFN-α plus ribavirin are similar to those caused by conventional IFN plus ribavirin. Side-effects related to IFN include: cytopenia, abnormalities of thyroid function, depression, irritability, concentration and memory disturbances, visual disturbances, fatigue, muscle aches, headaches, nausea and vomiting, loss of appetite and weight, low grade fever and skin irritation, insomnia, hearing loss, tinnitus, interstitial fibrosis and hair thinning. Side-effects associated with ribavirin include hemolytic anemia, fatigue, itching, rash, cough, gastrointestinal upset, pharyngitis, gout and birth defects. It is essential that persons who take ribavirin practice strict contraception during treatment and for 6 months after the termination of treatmen