Guideline:The liver in thalassemia

Under normal circumstances, about one-third of storage iron (ferritin and haemosiderin) in the body is found in the liver. Approximately 98% of hepatic iron is found in hepatocytes, which make up 80% of total liver mass; the remaining 1.5-2% of total liver iron is found in reticuloendothelial cells, endothelial cells, bile ductular cells and fibroblasts. Iron that enters the cell in excess of that required accumulates in the major storage forms of iron, ferritin, and haemosiderin. Progressive accumulation of storage iron is associated with cellular toxicity, although the specific pathophysiologic mechanisms for hepatocytes injury and liver fibrosis are not entirely understood. These include lipid peroxidation of organelle membranes, increased lysosomal fragility and decreased mitochondrial oxidative metabolism. Iron also has a direct effect on collagen synthesis and/or degradation, and alterations in microsomal enzymes.

The liver plays a central role in iron homeostasis. In addition to iron released from transfused red cells, an enhanced rate of gastrointestinal iron absorption has been suggested. This excess iron is initially confined to the Kupffer cells but when transfusion requirements produce massive iron overload, spillover to hepatic parenchyma cells quickly occurs, with the risk of late development of fibrosis and cirrhosis. In patients with ‚-thalassaemia, in absence of co-factors, the threshold hepatic iron concentration for the development of fibrosis is about 16 mg/g dry weight liver (Angelucci, 2002). Clinical studies suggest a relationship between hepatic iron concentration and the development of ironinduced hepatotoxicity.

Hepatic iron concentration (HIC) is the gold standard for the measurement of body iron overload (HIC in mg/g dry weight x 10.6 = whole body iron store in mg/kg) (Angelucci, 2000). Non-invasive techniques used to assess hepatic iron include computed tomography, biomagnetic liver susceptometry (SQUID) and magnetic resonance imaging (MRI). Of these, relaxation rates R2 (1/T2) and R2* (1/T2*) measured by MRI appear to be the most promising and accurate (Wood, 2005).

''Hepatic iron stores are closely correlated with cumulative transfusional iron load and have been used as a marker for the effectiveness of chelation therapy and prognosis. An increase in hepatic iron is associated with an increased risk of impaired glucose tolerance, diabetes mellitus, cardiac disease and death.''

Hepatitis C Virus (HCV)
This RNA virus was first characterised in 1989,having previously been termed non-A non-B hepatitis. The majority of HCV isolates studied so far can be divided into six major groups, designated genotypes 1-6, with subdivisions in each (subtype a, b, c, etc.). Antibodies that develop after infection are not protective but rather are indicative of current or past infection. Active infection is diagnosed by the presence of circulating HCV RNA in blood Sharara, 1996)

Preventative measures to minimise the risk of posttransfusional hepatitis C include careful selection of voluntary donors and appropriate blood donor screening.

Natural history and complications of infection
Acute infection: generally benign, with >80% asymptomatic. Anicteric Fulminant Hepatitis is very rare.

Chronic infection: develops in 70-80% of cases, leading to chronic liver disease. However, the clinical outcome is highly variable, for reasons that are not completely understood. Determinants of disease severity or chronicity as well as response to therapy include age at acquisition, as well as hostspecific(e.g. immunity) and virus-specific (e.g. genotype) factors and, most important,co-morbidities.

Cirrhosis: develops in a variable percentage of HCV-infected patients, ranging from < 5% in young, healthy people, to approximately 25-35% of cases of patients with relevant comorbidities. Age and co-morbidities appear to be the most important factors affecting the risk of developing cirrhosis. Cirrhosis usually takes as long as two to three decades to develop from the time of acquisition. Fiveyear survival in patients with compensated cirrhosis is 91%, with a 79% rate of 10-year survival. When cirrhosis is decompensated however, 5-year survival falls to just 50%.

Reversibility: The reversibility of advanced fibrosis and even early cirrhosis (Child A – compensated or well-compensated*) has been documented in thalassaemia once causes of liver injury (iron overload and HCV infection are removed) (Muretto, 2002).

End-stage liver disease: should lead to consideration of liver transplantation. Hepatitis C is currently the commonest reason for liver transplantation worldwide. Recurrent hepatitis C infection occurs in > 90% of cases after transplant but is usually mild. Long-term survival after liver transplantation for hepatitis C is similar to that for other diagnoses, averaging 65% after 5 years (Gane, 1996).

Hepatocellular carcinoma (HCC): develops in 1-5% of infected individuals after 20 years, particularly after the development of cirrhosis, increasing by 1-4% each year thereafter (Colombo, 1991). Prevention and early detection of HCC are more effective than attempted cure. Cirrhosis patients should undergo a regular six-monthly screening programme, including liver ultrasound examination and alpha-fetoprotein check, for the early detection of hepatocellular carcinoma.

Extra-hepatic manifestations of HCV infection include porphyria cutanea tarda, essential mixed cryoglobulinemia, glomerulonephritis, autoimmune thyroidits and vasculitis (Sharara, 1996).

'*' Liver cirrhosis is divided to 3 stages following the Child-Pugh score, Score 5-6 (Score A) is characterised by: No ascites, Bilirubin < 2mg/dl, Albumin>3.5g/dl. INR <1.7, no encephalopathy. Therefore, Child-Pugh stage A can be defined as “well compensated disease”.

Special features of hepatitis C in thalassaemia major
The severity of chronic hepatitis C in patients with thalassaemia may be greater because of concomitant iron overload, other concurrent viral infections (HBV, HIV) and possible infection with mixed hepatitis C genotypes. It has been demonstrated that iron and HCV infection are independent but mutually reinforcing risk factors for the development of liver fibrosis and cirrhosis, with a reciprocal multiplicative effect (Angelucci, 2002). It appears therefore that patients with thalassaemia, particularly those with poor control of iron overload, face an increased risk of developing cirrhosis.

Diagnosis and monitoring
Antibody testing This is most valuable for screening blood and blood products and as initial testing in patients with chronic unexplained elevation in serum transaminases or those suspected of having chronic liver disease. Confirmatory testing is done using HCV RNA detection by polymerase chain reaction (PCR), the current standard for the confirmation of viremia. Ascertaining the genotype and quantity of HCV RNA in serum is useful only in determining the type and duration of treatment (see below).

Liver biopsy in thalassaemia major Liver biopsy prior to treatment is helpful in determining the extent of liver damage and to guide decisions on therapy and anticipate complications (Angelucci, 1995).

Treatment This is a rapidly changing field and the treatment of hepatitis in patients with thalassaemia should therefore be undertaken in close collaboration with a specialist in liver disease.

Similar to non-thalassaemia patients, treatment of HCV in patients with thalassaemia is aimed at eradication of the virus, improvement in liver histology,reduction of the risk of liver cirrhosis and hepatocellular carcinoma.

Selection of patients for therapy Patients diagnosed with acute HCV infection and persistently positive serum HCV RNA after 12 weeks of exposure or diagnosis should receive treatment (Sharara, 2006).

Initiation of treatment in chronic hepatitis C has traditionally been based on one or more of the following:
 * confirmed presence of HCV-RNA
 * moderate to high serum ALT levels
 * abnormal liver histology

Encouraging results for the treatment of HCV in thalassaemia, combined with the abovementioned risks of greater severity of chronic hepatitis C in such patients, means that the presence of serum HCV-RNA alone is sufficient to consider treatment in patients with thalassaemia, where the patient has no other contraindications to treatment or other significant co-morbidities. Response to treatment Depending on HCV genotype and viral load,40-80% of patients with chronic hepatitis C will respond to the current standard treatment of pegylated interferon and ribavirin. Response is defined on the basis of a negative highly sensitive qualitative HCV RNA PCR assay, carried out 24 weeks after completion of therapy.

Patient responses are classified as follows:
 * Early viral response (EVR): defined as an undetectable HCV RNA or a > 2-log reduction in viral load after 12 weeks of treatment
 * Response at end of treatment(ETR): defined as absence of HCV-RNA at the end of treatment
 * Sustained viral response (SVR):absence of HCV-RNA > 6 months after concluding treatment. This is in practice equivalent to viral eradication of HCV
 * Non-responders: lack of significant decline (defined as > 2-log reduction from baseline) in HCV RNA after 12 weeks of therapy
 * Relapsers: re-emergence of HCV-RNA after a satisfactory end of treatment response

Monitoring response Depending on HCV viral genotype, the current recommendation is to measure the biochemical (serum ALT) and virological (HCVRNA)response after 4 to 12 weeks of therapy, and to continue therapy for an additional 12 to 24 weeks in patients with undetectable HCV-RNA. Because serum ALT may be raised for other reasons in patients with thalassaemia (iron overload, concomitant infections), monitoring response is based on viral HCV RNA.

Prediction of poor response Negative predictors in all patients with hepatitis C are:
 * High baseline HCV-RNA level and the absence of its early decay (4-12 weeks) upon initiation of treatment
 * HCV genotypes 1 or 4
 * Presence of bridging fibrosis or cirrhosis
 * Co-existence of other viruses (HBV, HIV)

Controversial in this specific setting is the role of iron overload.

Since no baseline factor is specifically predictive of treatment success or failure, withholding therapy on the basis of factors suggesting a poor response is unwarranted. Because of the possible role of iron overload in reducing the likelihood of successful treatment of hepatitis C and for general, well-known clinical reasons, effective chelation therapy should be strongly considered before initiation of antiviral therapy in patients with bad control of transfusional iron.

Treatment regimens
The gold standard is combination therapy with pegylated interferon and ribavirin. An example of an algorithm used for Hepatitis C managament is presented in Figure 1 see page 96.

Type of interferon: Pegylated interferon α-2a or α-2β‚ given subcutaneously once weekly

Duration: 24 to 48 weeks, depending on genotype

Side effects: Typical side effects in most patients include flu-like symptoms, insomnia, and cognitive and mood changes, especially in the first two weeks after starting interferon. Dose-dependent neutropenia and thrombocytopenia commonly occur during interferon therapy. Particular attention should be paid to this complication in patients with thalassaemia and hypersplenism. Since both deferiprone and interferon may cause neutropenia, there are theoretical risks associated with their combined use, and this combination should be initiated with caution and under careful monitoring. Hypothyroidism is an important complication of interferon treatment.

Some patients have experienced exacerbation of local reactions at the site of desferrioxamine infusion during interferon treatment. Heart failure has been seen in a few patients with thalassaemia receiving interferon, and special care should be given if prescribing interferon for patients with pre-existing heart disease.

Monitoring for side effects: Close monitoring for hypothyroidism is mandatory in patients receiving interferon, and testing for thyroid function and the presence of anti-thyroid antibodies should precede initiation of therapy. Regular monitoring of blood counts is also necessary, to identify neutropenia or thrombocytopenia. Cessation of therapy should be considered if the absolute neutrophil count falls below 1,000. Ribavirin is a nucleoside (guanosine)analogue, well absorbed orally, and typically given in doses of 800-1200mg/d. Alone, it has limited antiviral activity in hepatitis C but in combination therapy with interferon has been shown to significantly increase sustained response rates compared to interferon alone.

Side effects: Haemolysis occurs in most patients without thalassaemia, with a decrease in haemoglobin of 10-20% from baseline levels.

In thalassaemia major this may be associated with a more marked haemolysis and a 30% increase in transfusion requirement, which requires careful adjustment of the transfusion interval and intensification of iron chelation therapy (Li, 2002; Inati, 2005).

It is important to note that dose-reduction of ribavirin is associated with an inferior sustained viral response and it is hence recommended that transfusion-chelation requirements be adjusted to compensate for ribavirin-associated haemolysis rather than altering the recommended ribavirin dose (Inati, 2005).

Treatment duration and viral load monitoring: Depends primarily on the HCV genotype. For genotypes 1 or 4, treatment is administered for 48 weeks provided there is a positive early viral response (EVR) at 12 weeks. In the absence of EVR, treatment is usually discontinued and further treatment options considered.

This approach has been validated in patients with thalassaemia where an SVR of 64% is seen in patients infected with genotype 1 and 4 and who have exhibited an undetectable HCV RNA at 12 weeks of treatment (Inati, 2005). For genotypes 2 or 3, treatment is limited to 24 weeks. Given the high rate of SVR for genotypes 2 and 3, approaching 80%, a 12-week determination of viral load is not usually necessary.

Treatment options for nonresponders These have not been firmly established and are currently considered experimental. An expedited second treatment option may need to be considered in patients with advanced fibrosis on liver biopsy.

Management of special patient populations Consultation with a physician experienced in the management of liver disease is especially important in the clinical management of the following patient populations:
 * Children
 * Patients with cirrhosis
 * Immunosuppressed patients
 * Pregnant patients
 * Patients with acute hepatitis C

Prevention There is currently no vaccine or immunoglobulin to prevent hepatitis C. The following recommendations are made to reduce the risk of non-parenteral transmission:

Sexual transmission risk is generally low. However, insufficient data exist to recommend changes in current recommendations: that patients encourage their sexual partners to be tested for hepatitis C, and that safe sexual practices should be encouraged.

General measures, such as avoiding sharing toothbrushes, razors, etc. are advised, to avoid transmission to family members. However, the risk of transmission is low, and special measures such as to segregate towels and eating utensils are probably unnecessary.

Hepatitis B Virus(HBV)
Incidence ''Vaccination strategies, screening of blood donors for HBsAg, and other public health measures, have led to a significant reduction in hepatitis B infections in most countries of Europe and North America, as well as other parts of the world. Hepatitis B nevertheless remains a formidable medical problem, mainly in developing countries.''

Current HBsAg positivity in thalassaemia major ranges from <1% to >20% and Hepatitis B infection remains a significant cause of chronic liver disease and hepatocellular carcinoma in patients with thalassaemia in many regions of the developing world.

Clinical significance of HBV markers Despite the availability of good screening tests for hepatitis B, the interpretation of results may be difficult or misleading.


 * Acute infection. HBsAg is a reliable marker(can be present for 4-5 months). HBeAg is also transiently present (1-3 months). Anti-HBc IgM is the most reliable test for the diagnosis of acute HBV infection.

Chronic infection (overt carrier) is marked by the presence of HBsAg and anti-HBc in the blood (usually accompanied by HBeAg or anti-HBe). In accordance with international definitions, overt carriers can be classified as:


 * active carriers, identified by the presence of HBeAg or anti-HBe antibodies and a viral load ≥5 log10 copies/ml (although others cite a figure of ≥4 log10 copies/ml), corresponding to about 17,200 IU/ml, according to most recent standardisations. The great majority of active carrier cases are associated with the presence of hepatic disease.


 * inactive carriers, characterised by the persistent normality of transaminase in an anti-HBe-positive subject, associated with levels of viremia below the threshold (<5 Log10) and, eventually, with IgM anti-HBc <0.2 IMx Index. In the majority of such subjects, the histological finding, when available, does not reveal significant liver disease (necroinflammatory activity <4 HAI), while in a small minority of cases it is possible to observe effects of a chronic (sometimes even cirrhotic) disease which have became silent spontaneously or thanks to the effect of the antiviral treatment.


 * previous infection: the presence of anti-Hbc antibodies ± anti-Hbs indicates previous infection. In particular circumstances, such as deep immunosuppression (i.e. hemopoietic stem cell transplantation), the possibility of HBV reactivation after a previous infection has been demonstrated. This category of patients can therefore also be defined as potential occult carriers (Marzano, 2007).


 * vaccination: the presence of HBsAg antibodies (if anti-HBc is not present)indicates vaccination.

Patients with thalassaemia should be screened for all serological markers of hepatitis B and classified according to Table 1 (see page 100), which provides a list of possible interpretations of screening results.

Natural history
Acute hepatitis: This is the most common presentation, with an incubation period of 4- 20 weeks. Severity is variable, with an icteric period often preceded by a prodromal illness with arthralgia and urticaria. Progression to fulminant hepatic failure is rare (≤1%). Acute hepatitis B is usually managed by supportive measures alone.

Progression to chronic hepatitis B occurs in 5-10% of otherwise healthy adults and in 90% of neonates. In acute icteric hepatitis B in adults, transition to chronicity appears to be rare, probably occurring in less than 2% of cases. For patients with chronic hepatitis B infection, co-infection with hepatitis C may increase the severity and rate of progression of liver disease.

Cirrhosis occurs at a rate of 1-2.2% per year. Iron loading in thalassaemia may increase the risk, as may concomitant HCV infection.

Hepatocellular carcinoma is a wellrecognised complication of chronic hepatitis B infection.