Hemochromatosis and the HFE Gene - Phenotypic Correlation and Clinical Application
Course AuthorsEric W.H. Leong, M.D., and Emmet B. Keeffe, M.D. Eric W. H. Leong, M.D., is Senior Fellow in Gastroenterology, and Emmet B. Keeffe, M.D., is Professor of Medicine (Gastroenterology) and Medical Director, Liver Transplant Unit, Stanford University School of Medicine. Drs. Leong and Keeffe report no commercial conflict of interest. Estimated course time: 1 hour(s). Albert Einstein College of Medicine – Montefiore Medical Center designates this enduring material activity for a maximum of 1.0 AMA PRA Category 1 Credit(s)™. Physicians should claim only the credit commensurate with the extent of their participation in the activity. In support of improving patient care, this activity has been planned and implemented by Albert Einstein College of Medicine-Montefiore Medical Center and InterMDnet. Albert Einstein College of Medicine – Montefiore Medical Center is jointly accredited by the Accreditation Council for Continuing Medical Education (ACCME), the Accreditation Council for Pharmacy Education (ACPE), and the American Nurses Credentialing Center (ANCC), to provide continuing education for the healthcare team.  
Learning Objectives
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In 1865, Trousseau provided the first description of a syndrome involving cirrhosis, diabetes mellitus and increased skin pigmentation.(1) The familial nature of hemochromatosis was not recognized until 1935 by Sheldon.(2) Following linkage of hereditary hemochromatosis to HLA-A3 in 1977 by Simon et al.,(3) Wolff et al.(4) identified the candidate gene in 1996, through positional cloning, in a cohort of patients with hemochromatosis and named it HLA-H. This gene was renamed HFE on the recommendation of the World Health Organization Nomenclature Committee in 1997.(5) Clinical FeaturesHereditary hemochromatosis is an inherited disorder of iron metabolism that is characterized by inappropriate intestinal absorption of iron, progressive iron deposition in organ parenchymal cells and eventual multiorgan failure in untreated individuals with iron overload. The major target organs include the liver, heart, pancreas and pituitary gland, putting untreated individuals at risk for cirrhosis, hepatocellular carcinoma, cardiomyopathy, cardiac arrhythmias, diabetes mellitus and hypogonadotrophic hypogonadism. Fatigue, arthritis and abnormal skin pigmentation are other common features in this disorder. DiagnosisThe widely accepted criteria for the diagnosis of iron overload due to hemochromatosis include 4 g or more of iron removed by phlebotomy (16 units of blood) before the onset of iron-limited erythropoiesis or at least one of the following results derived from liver biopsy: grade 3 or 4 (out of four grades) stainable iron, hepatic iron concentration greater than 80 µmol per gram (dry weight) of liver tissue and hepatic iron index (hepatic iron concentration divided by age) greater than 1.9.(6),(7) The identification of specific mutations in the HFE gene of patients with hereditary hemochromatosis has permitted the introduction of genetic testing in the clinical setting. EtiologyThe HFE gene encodes for a 343-residue, major histocompatibility complex class I-related protein that interacts with class I2-microglobulin and transferrin receptor on the basolateral surface of intestinal crypt cells.(4),(8) This complex may permit sampling of serum iron by modulating the uptake of transferrin-transferrin receptor heterodimeric complexes into the crypt cell cytoplasm, followed by the release of transferrin-bound iron from the endosome.(9),(10) Crypt cell iron levels influence the activity of iron regulatory proteins, which, in turn, act on iron regulatory elements to regulate expression of the divalent metal ion transporter, DMT-1, on the apical surface of absorptive enterocytes. Thus, a mutation in HFE may impair transferrin receptor-mediated iron uptake, falsely signalling low crypt cell iron stores and promote continued upregulation of DMT-1 expression and iron uptake by the villus enterocyte, leading to iron overload.(11) Two Major MutationsTwo major missense HFE mutations have been identified by Feder et al.(4) Substitution of tyrosine for cysteine at amino acid position 282 in the HFE protein, referred to as the C282Y mutation, appears to prevent binding with b2-microglobulin, which is required for intracellular processing and transport of the protein to its functional locale on the cell surface.(12) In contrast, the H63D mutant protein, which substitutes aspartate for histidine at amino acid position 63, correctly localizes to the cell surface and forms a complex with the transferrin receptor but, apparently, fails to reduce its affinity for transferrin,(13) which may produce low cytoplasmic iron levels in the crypt cell. Other mutations in the HFE gene that have been identified in patients with clinical iron overload include the serine-to-cysteine substitution (S65C) and the IVS3+1G → T splice mutation.(14),(15) Inherited and apparently non-HFE-associated disorders of iron overload have also been reported. Besides thalassemia major, these include autosomal dominant hemochromatosis, described in an extended Melanesian family from the Solomon Islands,(16) juvenile hemochromatosis(17) and non-C282Y-associated hemochromatosis in one Italian family.(18) Pedigree analysis and genotyping have shown that hereditary hemochromatosis is inherited predominantly in an autosomal recessive pattern. Furthermore, several studies have established that approximately 85% (range: 64-100%) of individuals with clinically expressed hemochromatosis are C282Y homozygotes and 3.3% (range: 0-7%) are C282Y/H63D compound heterozygotes.(19),(20),(21),(22),(23),(24) Individuals who are homozygous for C282Y appear most likely to develop clinically significant iron overload, followed by C282Y/H63D compound heterozygotes. Iron overloading may also be seen in C282Y heterozygotes, H63D homozygotes, and H63D heterozygotes, but the iron burden is usually mild to moderate and markedly lower than in C282Y homozygotes. In particular, the H63D mutation in isolation appears to contribute only marginally to the iron-overloaded state, based on a study of 1110 French patients by Moirand et al.(25) (See Table 1). Table 1. Details of Genotypes Producing Expression of Hemochromatosis.
*One Normal or "Wild Type" allele at this position to produce the heterozygote. However, studies of Italian patients with hemochromatosis and sporadic porphyria cutanea tarda, in which iron is a key modulating factor in the function of uroporphyrinogen decarboxylase, provide contrary data, suggesting that the C282Y mutation plays a less important role in clinically expressed iron overload.(26) In fact, among C282Y homozygotes, only 50% fully express the diseased state characterized by symptoms and biochemical indices consistent with iron overload.(27) Partial penetrance of the C282Y mutation, therefore, provides one explanation for the observed disparity between the documented prevalence of the mutation and patients' relatively infrequent presentation with symptomatic iron overload. Population studies show that the allele frequency of the C282Y mutation ranges from 0.4% among Melanesians to 10% among the Irish, with a worldwide carrier frequency of 1.9%.(28),(29) Homozygosity for the C282Y mutation is seen in 0.5% of individuals of northern European ancestry. The allele frequency of the H63D mutation is as high as 30% among the Basque, whereas the worldwide carrier frequency for this mutation is 8.1%.(28) Genetic ScreeningGiven the one in 300-400 incidence of expressed disease among Caucasians, as well as the potentially life-threatening complications of unrecognized iron overload, the availability of genetic testing for the two major HFE mutations provides an attractive tool for early identification of individuals who are at risk for progressive iron accumulation. However, it is necessary to consider the clinical context, feasibility of implementation and cost-effectiveness of any test before its adoption in a widespread screening strategy. Three populations should be considered in the context of screening for hemochromatosis: relatives of hemochromatotic patients, individuals undergoing routine medical examination and the general population.(30) Powell at al. recommend that all first-degree relatives of patients known to have hereditary hemochromatosis undergo screening for the C282Y mutation; those who are homozygous for the mutation should have blood samples drawn for iron studies (transferrin saturation and ferritin) and liver enzymes. In addition, the spouse of the proband should be tested for the C282Y mutation. If the spouse is heterozygous for the C282Y mutation, the children should undergo genetic testing because each is 50% at risk of being homozygous for the mutation. If the spouse does not possess the C282Y mutation, testing of the children is not necessary.(31) This strategy of screening for the C282Y mutation among first-degree relatives of patients with hereditary hemochromatosis has been shown to be cost-effective.(32) Many hepatologists recommend that determination of the fasting serum transferrin saturation should be performed as a component of the routine medical examination at about age 30. A value that exceeds 45% should raise clinical suspicion of hemochromatosis, as this screening threshold identifies 98% of affected individuals with relatively few false-positive results.(33) Subsequent measurement of serum ferritin, accepting the possibility of a false elevation in the setting of infection, inflammation or malignancy, provides information on the iron burden because ferritin increases linearly with total body iron stores.(34) Genotyping and measurement of liver enzymes should also be performed at this point. If the individual is not a C282Y homozygote or C282Y/H63D compound heterozygote, non-HFE-associated causes of iron overload must be considered. If the genotype, on the other hand, indicates a predisposition to HFE-related hemochromatosis, the serum ferritin and liver enzyme profile should be evaluated. Clinical ManagementPatients who have persistently elevated transferrin saturations with normal ferritin and normal aminotransferase levels can be classified as having non-expressed hemochromatosis; clinical observation and repetition of the iron indices with aminotransferase levels in one to two years is reasonable in this setting.(35),(36) Phlebotomy is indicated in patients whose ferritin is 300-1000 µg/L with normal liver enzymes. Liver biopsy is necessary to confirm and identify the degree of parenchymal iron overload when the patient has a ferritin in excess of 1000 µg/L and/or elevated aminotransferase levels,(37) as the risk of developing a hepatoma and dying increases significantly once the hepatic iron concentration is 400 µmol per gram or more.(38) However, young patients under 40 years of age who are genotypically at risk for expressing the hemochromatosis phenotype are unlikely to have significant hepatic fibrosis or cirrhosis and liver biopsy may be unnecessary in this group.(39) Should Everyone Be Screened?Although hereditary hemochromatosis meets the World Health Organization criteria for disease screening in the general population, some issues merit further reflection. First, population studies document the heterogeneity of C282Y allele frequencies around the world,(27),(28),(29) and in areas with low prevalence rates, the concern is that fewer homozygotes will be identified, compounded by the higher cost of finding each homozygote. Second, as the penetrance of the mutation in the screened population decreases, the cost of identifying homozygotes rises, while the incremental cost saving for both men and women falls.(40) Thus, an argument can be made for phenotypic screening with iron studies. In the United States, this approach would target 1.8 to 3.8 million adults for further testing, using the cut-off values for elevated serum transferrin saturation (>60% in men and >55% in women) recommended by the College of American Pathologists.(41) In addition, at a cost of US $12 for the initial transferrin saturation determination, the cost to the health care system would be substantial. Measurement of the unbound iron binding capacity has been shown to be cost-effective,(42) allowing the cost per screening test to drop below US $1. Adams and Valberg have identified two cost-effective strategies for population screening for HFE-related hemochromatosis.(40) In the first approach, phenotypic testing for iron overload is performed, followed by confirmatory genotyping. In the second approach, initial genotyping saves money if the cost of the genetic test is under US $28. Therefore, at a cost of US $173 per test, genotyping cannot be recommended for population screening for hereditary hemochromatosis. ConclusionAt the present time, it may be reasonable to introduce phenotypic screening by including iron studies with other screening blood tests. In the future, it is possible that screening by genotype will be feasible if costs of the genetic test are reduced and any relevant ethical issues associated with genetic screening are resolved. |