Hemochromatosis

Corwin Q. Edwards

James C. Barton

IRON OVERLOAD

Iron overload can result from any process that causes iron accumulation in excess of iron loss. Examples of such processes include (a) erythrocyte transfusions in individuals who do not experience blood loss (e.g., patients with hemolytic anemia, aplastic anemia, and sideroblastic anemia); (b) absorption of excessive amounts of normal dietary iron (hemochromatosis types 1 through 4, disorders of ineffective erythropoiesis); and (c) absorption of increased amounts of excessive dietary iron (e.g., high-dose medicinal iron and African iron overload).

HEMOCHROMATOSIS

Hemochromatosis Type 1, HFE Hemochromatosis

The definition of “classical,” autosomal recessive hemochromatosis (type 1) typical of Western European whites once required demonstration of the rare triad of cutaneous hyperpigmentation (“bronzing”), diabetes mellitus, and cirrhosis that was presumed to represent complications of severe iron overload. The eventual ability to measure iron in liver and other tissues, serum, and by phlebotomy to achieve iron depletion revealed that hemochromatosis was more common than previously believed and that the diagnosis could be defined by severity of iron overload. In 1996, it was discovered that most patients with this type of hemochromatosis were homozygous for a common missense mutation (C282Y) of the HFE gene on chromosome 6p21.3. Thus, most current definitions of hemochromatosis in Western European whites include specification of the iron phenotype and HFE genotype. Three important relationships of phenotype and genotype definitions are: (a) HFE C282Y homozygosity is a genetic configuration that markedly increases the risk that iron overload will eventually develop without other readily demonstrable cause; (b) iron phenotypes and organ-specific complications of HFE C282Y homozygotes vary greatly due to genetic and environmental factors; and (c) iron overload, not HFE C282Y homozygosity or abnormal HFE protein, causes tissue injury in target organs.

Hemochromatosis Type 2, Juvenile Hemochromatosis

Hemochromatosis type 2 is characterized by nontransfusion iron overload, often severe, that typically occurs in children or young adults.

Type 2a, Mutations of Hemojuvelin Gene (HJV)

Hemochromatosis type 2a is an autosomal recessive disorder due to pathogenic mutations in the hemojuvelin gene (HJV; chromosome 1q21).¹, ² The iron overload phenotype of patients with HJV hemochromatosis is much more severe at the same age than the phenotype of typical patients with hemochromatosis due to HFE C282Y homozygosity and the penetrance of hemochromatosis type 2a genotypes (risk to cause iron overload) is also greater. Hemochromatosis type 2a often causes abdominal pain and hepatomegaly in children, hypogonadotrophic hypogonadism in teenagers, arthropathy, and severe heart failure, cardiac arrhythmias, and cirrhosis before the age of 30 years.³ Based on the volume of blood removed to achieve iron depletion in some patients, it can be deduced that they absorbed 3.2 to 3.9 mg of dietary iron per day.⁴ In severe HFE hemochromatosis, 2 mg of iron or less is absorbed daily.⁵, ⁶

Two pathogenic HJV mutations (either homozygous or compound heterozygous configuration) occur in each patient. The most common HJV mutation causes a change in nucleotide 959G→T, which codes for substitution of the normal glycine by valine in position 320 of the polypeptide sequence of the HJV protein (HJV G320V). This mutation has been detected in patients with juvenile-onset hemochromatosis of diverse European ethnicities.⁷, ⁸ About 30 pathogenic HJV mutations have been reported. Many are novel (not detected in unrelated patients with hemochromatosis type 2a.)⁷, ⁸, ⁹, ¹⁰, ¹¹ Some HJV hemochromatosis patients also have an abnormal HFE genotype but the latter does not account for iron overload in most cases.¹²

Hemojuvelin is involved in the up-regulation of hepcidin synthesis, not in the regulation of HFE function. Decreased hemojuvelin activity decreases hepcidin synthesis, thus decreasing hepcidin binding and inactivation of ferroportin. Consequently, regulation of iron transport out of absorptive enterocytes into the plasma via ferroportin is decreased, even in the presence of increased storage iron.

Type 2b, Mutations of Hepcidin Gene (HAMP)

Hemochromatosis type 2b is associated with iron overload of variable severity and age of onset and is also transmitted as an autosomal recessive trait.¹², ¹³ This disorder is rare. Hemochromatosis type 2b is due to mutations in the hepcidin gene (HAMP, chromosome 19q13). HAMP is an acronym for hepcidin antimicrobial peptide. The word hepcidin is derived from hepatic bactericidal protein.

The first reported HAMP mutation (nucleotide 208T→C) causes substitution of the normal cysteine by arginine at amino acid 70 (C70R).¹⁴ Approximately 12 other HAMP mutations have been reported.¹⁵, ¹⁶, ¹⁷, ¹⁸ Mutant hepcidin has decreased ability to bind to ferroportin, its only receptor. Consequently, ferroportin is not degraded and thus continues to transport iron across the basolateral membrane of absorptive enterocytes in the presence of excess iron stores.¹⁹ Some hemochromatosis type 2b patients, usually those heterozygous for pathogenic HAMP promoter or coding region mutations, also have HFE mutations that contribute to the pathogenesis of iron overload (“digenic” hemochromatosis).

Hemochromatosis Type 3, Mutations of Transferrin Receptor-2 Gene (TFR2)

Hemochromatosis type 3 is a rare autosomal recessive disorder reported in persons of European or Asian ancestry caused by mutations of the transferrin receptor-2 gene (TFR2) located on chromosome 7q22.²⁰ The clinical phenotype is moderately variable and resembles either that of HFE hemochromatosis or that of juvenile hemochromatosis types 2a and 2b. Fewer than 20 pathogenic TFR2
mutations have been reported.²⁰, ²¹, ²², ²³, ²⁴ The penetrance of hemochromatosis type 3 genotypes is relatively great. Consanguinity has been identified in some affected families. TFR2 Y250X and R455Q have been detected in individuals or kindreds who were not closely related.²⁰ Pathogenic TFR2 mutations either result in gain-of-function or in decreased quantities of functional transferrin receptor-2. TFR2 protein is thought to modulate the signaling pathway that controls hepcidin expression²⁵ with consequent low hepcidin²⁶ and increased iron absorption and deposition in hepatocytes.

Hemochromatosis Type 4, Mutations of Ferroportin Gene (SLC40A1)

Hemochromatosis type 4 is an uncommon heterogeneous disorder transmitted as an autosomal dominant trait. It is caused by mutations of a gene of the solute carrier family 40 (member #1) (SLC40A1, chromosome 2q32), which encodes the synthesis of ferroportin, the sole receptor of hepcidin.²⁷, ²⁸, ²⁹, ³⁰ There are two distinct ferroportin hemochromatosis phenotypes. Loss-of-function SLC40A1 mutations encode ferroportin that either is not presented normally to the cell surface or has defective iron transport activity. Such mutations decrease iron absorption from the intestine and inhibit iron egress from macrophages. Phenotypes of affected patients include normal or low transferrin saturation, mild anemia, and predominance of iron retention in macrophages. Gain-of-function mutations encode ferroportin that cannot bind hepcidin normally or be internalized after hepcidin binding (hepcidin resistance). This increases iron absorption and stimulates iron export by macrophages. Affected patients have elevated transferrin saturation and iron deposition in hepatocytes.

Ferroportin mutations are cosmopolitan. Most reported mutations are restricted to single families. SLC40A1 V162del has been reported in persons with iron overload from Australia, Europe, and Asia. SLC40A1 A77D has been reported in persons with iron overload in Italy, Australia, and India. SLC40A1 Q248H occurs as a polymorphism in natives who reside in diverse areas of sub-Saharan Africa and in African Americans, but probably does not cause iron overload.³¹

HISTORY

Hemochromatosis Type 1, Classical, Hereditary

The first report of a person with hemochromatosis was included in a discussion of diabetes mellitus in 1865.³² The patient was a 28-year-old Parisian man who sold newspapers and who died of a febrile illness that complicated his diabetes. His autopsy revealed hepatomegaly and hypertrophic cirrhosis. The second case of hemochromatosis was reported in 1871 and described a 51-year-old man who lived in Paris.³³ He had skin bronzing of his face and hands. His autopsy revealed a chocolate-colored, enlarged liver. Microscopic examination of his liver showed pigment granules inside and outside of hepatocytes. In 1886, the term pigmentation cirrhosis was used to describe the triad of skin bronzing, diabetes mellitus, and hepatic cirrhosis.³⁴ The name hämochromatose was first used by von Recklinghausen in his 1889 description of 12 individuals who had pigmentation of multiple organs.³⁵ He believed that the iron that accumulated in organs came from red blood cells.

ETIOLOGY

Cloning of the HFE Gene

In 1996, the hemochromatosis gene was isolated by positional cloning and two common mutations that account for most cases of hemochromatosis in persons of Western European descent were discovered.³⁶ The gene was named HFE (H = hemochromatosis; Fe = iron).³⁷, ³⁸ The cloning of HFE was reported more than 130 years after the first case description of hemochromatosis at autopsy and confirmed reports during an interval of more than two decades that hemochromatosis was transmitted as an autosomal recessive condition linked to the HLA locus on the short arm of chromosome 6.³⁹, ⁴⁰, ⁴¹, ⁴², ⁴³, ⁴⁴, ⁴⁵, ⁴⁶, ⁴⁷

Function of the HFE Gene

The normal HFE gene encodes normal HFE protein which is expressed in duodenal crypt cells and reticuloendothelial cells.⁴⁸ In a normal individual, HFE protein binds to β₂-microglobulin, which decreases the affinity of cell membrane transferrin receptors for transferrin.⁴⁹, ⁵⁰, ⁵¹, ⁵² This decreases iron absorption. In iron deficiency, HFE protein synthesis is decreased, binding of HFE protein to β₂-microglobulin is decreased, the affinity of the transferrin receptor for transferrin is increased, and thus duodenal crypt cells absorb more iron.

Effects of HFE Gene Mutations

The HFE protein binding site for the β₂-microglobulin binding site has two cysteine molecules that form a disulfide bridge. In HFE C282Y homozygotes, tyrosine replaces a cysteine molecule at amino acid position 282 in the product of both HFE gene copies (maternal and paternal) and thus the disulfide bridge is absent. This prevents binding of the abnormal HFE protein to β₂-microglobulin on the cell surface. Excessive iron is then absorbed through the crypt cells and passed into the circulation.⁵³, ⁵⁴, ⁵⁵, ⁵⁶, ⁵⁷, ⁵⁸ Despite these observations, the full function and participation of the HFE protein in iron absorption has not been elucidated. Studies of penetrance of hemochromatosis demonstrate that HFE C282Y homozygosity is necessary but probably not sufficient for iron overload to develop.

PATHOPHYSIOLOGY

Normal adults absorb an amount of iron each day that precisely balances daily losses (men ˜1 mg, women 1 to 3 mg). In healthy children and infants, daily iron absorption corresponds to the iron requirements of growth and development. In acute blood loss or iron deficiency, daily iron absorption increases until the total body deficit has been replaced. In persons with hemochromatosis, the absorption of dietary iron is inappropriately great in comparison with quantities of total body iron. In some persons with HFE hemochromatosis and iron overload, daily iron absorption may exceed 2 mg at a time when iron absorption should have decreased to nearly zero.⁵⁹, ⁶⁰ In some children with juvenile hemochromatosis, average daily iron absorption may exceed 3 mg/day. Phlebotomy therapy to achieve iron depletion in hemochromatosis may increase iron absorption to 10 mg/day and iron absorption may remain inappropriately elevated long after iron deficiency resolves.⁶¹, ⁶², ⁶³ Iron loading in hemochromatosis types 1 to 4 is caused either by an abnormality in hepcidin synthesis or a decrease of its activity via decreased function of ferroportin. HFE, HJV, and TFR2 probably all have a contributory role in regulating hepcidin synthesis.

Effects of Hepcidin

Hepcidin, the central regulator of iron absorption, is a low-molecular-weight thionin-like, defensin-like peptide with many cysteine pairs encoded by the HAMP gene. Hepcidin has antimicrobial activity against bacteria and fungi, is present in plasma, and is excreted in urine. Hepcidin is synthesized in hepatocytes in response to anemia, hypoxia, and inflammatory stimuli mediated by interleukin 6,⁶⁴, ⁶⁵, ⁶⁶, ⁶⁷, ⁶⁸, ⁶⁹, ⁷⁰, ⁷¹ and in the presence of adequate or increased amounts of iron.⁷², ⁷³, ⁷⁴, ⁷⁵ It is also involved in the abnormalities of iron metabolism that occur in the anemia of chronic disease.⁷⁶, ⁷⁷, ⁷⁸, ⁷⁹ Hepcidin levels are inversely related to iron absorption.

Ferroportin, the sole receptor for hepcidin, occurs as a multimer on the surfaces of cells responsible for gathering and recycling iron: enterocytes (basolateral surfaces), macrophages, hepatocytes, and placental syncytiotrophoblasts. The binding of cell surface ferroportin by hepcidin results in ferroportin tyrosine phosphorylation at the plasma membrane, dephosphorylation, and degradation by ubiquitination.¹⁹, ⁶⁹, ⁸⁰ Thus, hepcidin participates in regulation of plasma iron levels and tissue distribution of iron by post-translational regulation of ferroportin.

Hepcidin Effect in Iron Deficiency

In iron deficiency, hepcidin production is down-regulated so the amount of hepcidin available to bind ferroportin is reduced. More iron is exported from absorptive enterocytes by ferroportin than usual, resulting in increased absorption of iron through duodenal enterocytes into the plasma.⁸¹

Hepcidin Effect in Iron Repletion

In normally iron-replete humans, the production of hepcidin is up-regulated. The hepcidin bonds with ferroportin inducing ferroportin internalization and degradation.¹⁹, ⁶⁹, ⁸⁰ Consequently, the amount of ferroportin is reduced and iron absorption from duodenal enterocytes into plasma decreases.

Hepcidin Effect in Hemochromatosis

Dysregulation of hepcidin production underlies the pathophysiology of most types of hemochromatosis (types 1, 2a, 2b, and 3). In iron-loaded individuals with HFE hemochromatosis, for example, hepcidin expression by the liver is inappropriately low⁸², ⁸³ and thus iron absorption continues despite increased iron stores. Hepcidin levels are decreased in all types of hemochromatosis except in patients with “classical” ferroportin hemochromatosis (type 4). In those patients, iron overload is caused by a mutation in the ferroportin gene (SLC40A1) that results in diminished or absent functional quantities of ferroportin.⁸¹ In such cases, hepcidin production and regulation are normal.

Molecular Basis of Increased Iron Absorption in Hemochromatosis

Several factors in the duodenum contribute to inappropriately high iron absorption in HFE hemochromatosis. In enterocytes, DMT1 (divalent metal transporter-1) transports iron across the apical microvillous membrane into the cell in the presence of hydrogen ions derived from gastric acid and ferroportin transports iron across the basolateral membrane into the plasma. Untreated C282Y homozygotes have inappropriate up-regulation of DMT1 and ferroportin mRNA expression in proportion to their serum ferritin level.⁸⁴ Absorption of heme iron is also increased in proportion to serum ferritin levels.⁵, ⁶⁰ The interaction of transferrin, transferrin receptor, and HFE protein at the basolateral membranes of duodenal absorption cells may play a minor role in regulating iron absorption in persons with HFE hemochromatosis.⁸⁵ SLC40A1 mutations that encode ferroportin that either is not presented normally to the cell surface or has defective iron export activity are associated with loss-of-function, normal or low transferrin saturation, and predominance of iron retention in macrophages. Such mutations decrease iron absorption from the intestine and inhibit iron export from macrophages.

The complicated interactions of HFE, DMT1, transferrin, transferrin receptors, hemojuvelin, hepcidin, and ferroportin in regulating iron absorption in normal individuals and in persons with hemochromatosis are a topic of intense and productive research.⁷⁵, ⁸⁰, ⁸⁶, ⁸⁷, ⁸⁸, ⁸⁹, ⁹⁰

ORGAN AND CELLULAR INJURY DUE TO IRON OVERLOAD

After many years of increased iron absorption, hepatocyte and Kupffer cell iron storage sites become overloaded and iron accumulates progressively in the myocardium, pancreas, anterior pituitary, spleen, and other organs. Cardiac myocytes, beta-cells of the pancreatic islets, and gonadotroph cells of the anterior pituitary have especially great affinity for otherwise unbound iron. Some individuals with severe iron overload due to hemochromatosis have body iron burdens that are more than 10 times normal. Usually, such individuals do not develop symptoms or signs of illness until after they have had iron overload for three to five decades. Exceptions include the organ damage and death that can occur before the age of 30 years in persons with severe iron overload due to juvenile hemochromatosis.

Iron that is bound to transferrin or stored in modest amounts in ferritin is not toxic. In the presence of excessive iron stores, non-transferrin-bound or non-ferritin iron occurs and generates reactive oxygen species (oxyradicals), especially the hydroxyl radical.⁹¹, ⁹², ⁹³, ⁹⁴, ⁹⁵, ⁹⁶, ⁹⁷ It is likely that hydroxyl, alkoxyl, and peroxyl radicals are involved in lipid peroxidation which damages microsomes, mitochondria, and lysosomes. Hydroxyl radicals are also thought to be involved in iron-related damage of enzymes, proteins, nucleic acids, and polysaccharides. Lipid peroxidation then results in disruption of membrane-dependent functions of lysosomes, mitochondria, endoplasmic reticulum, cell membranes, and DNA.⁹¹, ⁹⁴ Excess iron and iron-induced oxyradicals may also activate stellate cell transformation to fibroblasts in the liver and act as carcinogens in hepatocytes or extrahepatic parenchymal cells.

DIFFERENTIAL DIAGNOSIS OF IRON OVERLOAD

Many heritable or acquired disorders are associated with increased body iron stores. Some of these disorders appear in Table 25.1,¹, ¹², ¹³, ²⁰, ²¹, ²², ²³, ²⁴, ²⁶, ²⁷, ³⁶, ⁷², ⁷³, ⁷⁴, ⁹⁸, ⁹⁹, ¹⁰⁰, ¹⁰¹, ¹⁰², ¹⁰³, ¹⁰⁴, ¹⁰⁵, ¹⁰⁶, ¹⁰⁷, ¹⁰⁸, ¹⁰⁹, ¹¹⁰, ¹¹¹, ¹¹², ¹¹³, ¹¹⁴, ¹¹⁵, ¹¹⁶, ¹¹⁷, ¹¹⁸, ¹¹⁹, ¹²⁰, ¹²¹, ¹²², ¹²³, ¹²⁴, ¹²⁵, ¹²⁶, ¹²⁷, ¹²⁸, ¹²⁹, ¹³⁰, ¹³¹, ¹³², ¹³³, ¹³⁴, ¹³⁵, ¹³⁶, ¹³⁷, ¹³⁸, ¹³⁹, ¹⁴⁰, ¹⁴¹, ¹⁴², ¹⁴³, ¹⁴⁴, ¹⁴⁵, ¹⁴⁶, ¹⁴⁷, ¹⁴⁸, ¹⁴⁹, ¹⁵⁰, ¹⁵¹, ¹⁵², ¹⁵³, ¹⁵⁴, ¹⁵⁵, ¹⁵⁶, ¹⁵⁷, ¹⁵⁸, ¹⁵⁹, ¹⁶⁰, ¹⁶¹, ¹⁶², ¹⁶³, ¹⁶⁴, ¹⁶⁵, ¹⁶⁶, ¹⁶⁷, ¹⁶⁸, ¹⁶⁹, ¹⁷⁰, ¹⁷¹, ¹⁷², ¹⁷³, ¹⁷⁴, ¹⁷⁵, ¹⁷⁶, ¹⁷⁷, ¹⁷⁸, ¹⁷⁹, ¹⁸⁰, ¹⁸¹, ¹⁸², ¹⁸³, ¹⁸⁴, ¹⁸⁵, ¹⁸⁶, ¹⁸⁷, ¹⁸⁸, ¹⁸⁹, ¹⁹⁰, ¹⁹¹, ¹⁹², ¹⁹³, ¹⁹⁴, ¹⁹⁵, ¹⁹⁶, ¹⁹⁷, ¹⁹⁸, ¹⁹⁹, ²⁰⁰ including gene location, mode of heritability, and gene mutation responsible for the disorder. Table 25.1 provides guidance for physicians who are evaluating and managing patients with iron overload.

HFE HEMOCHROMATOSIS GENE

HFE Location

The HFE gene is located approximately 4 megabases telomeric to the HLA region on the short arm of chromosome 6(6p). HFE is structurally similar to other HLA class I-like genes. HFE is composed of seven exons, of which the first six encode for the six domains of the HFE protein. The seven exons of HFE result in formation of a messenger RNA transcript of 4.2 kilobases. This in turn results in the synthesis of the HFE product, a protein of 343 amino acids. The most common mutation of HFE (C282Y) associated with iron overload is caused by a mutation of one nucleotide base 845G→A in exon 4 of the HFE gene.³⁶

HFE Mutations

More than 60 mutations of the HFE gene have been reported.¹⁵, ³⁶, ²⁰¹, ²⁰², ²⁰³, ²⁰⁴, ²⁰⁵, ²⁰⁶, ²⁰⁷, ²⁰⁸, ²⁰⁹, ²¹⁰, ²¹¹, ²¹², ²¹³, ²¹⁴, ²¹⁵, ²¹⁶, ²¹⁷, ²¹⁸, ²¹⁹, ²²⁰, ²²¹, ²²², ²²³, ²²⁴, ²²⁵, ²²⁶, ²²⁷, ²²⁸, ²²⁹, ²³⁰, ²³¹, ²³², ²³³, ²³⁴, ²³⁵, ²³⁶, ²³⁷, ²³⁸, ²³⁹ The most common mutation that occurs in people who have hemochromatosis is a change in nucleotide 845 from the normal guanidine to adenine in exon 4 of the HFE gene (referred to as 845G→A). This nucleotide change results in the substitution of tyrosine in place of the normal cysteine in amino acid position 282, referred to as Cysteine282Tyrosine, Cys282Tyr, or C282Y. This mutation causes the majority of iron accumulation in subjects who have hemochromatosis.³⁶

TABLE 25.1 HERITABLE AND ACQUIRED DISORDERS ASSOCIATED WITH IRON OVERLOAD: DIFFERENTIAL DIAGNOSIS

Heritable Disorder		Chromosomal Assignment	Heritability	Cause of Iron Loading	References
HFE Hemochromatosis		6p21.3	Autosomal recessive	Mutations of HFE	36
Juvenile Hemochromatosis
	HJV Hemochromatosis	1q21	Autosomal recessive	Mutations of hemojuvelin	1, 12, 98, 99, 100, 101
	HAMP Hemochromatosis	19q13	Autosomal recessive	Hepcidin antimicrobial peptide gene mutations	13, 72, 73, 74
TFR2 Hemochromatosis		7q22	Autosomal recessive	Inactivation of transferrin receptor 2	20, 21, 22, 23, 24, 26, 102
SLC40A1 Hemochromatosis		2q32	Autosomal dominant	Ferroportin gene mutations	27, 101, 103, 104, 105, 106, 107
H-Ferritin Hemochromatosis		11q 12-q13	Autosomal dominant	H-ferritin gene mutations	108
Porphyria cutanea tarda		1p34	Autosomal dominant or sporadic	Heterogeneous	109, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 128, 129
African iron overload		Unknown	Autosomal dominant	Unknown	130, 131, 132, 133, 134, 135, 136, 137
Neonatal iron overload		Unknown	Heterogeneous	In utero iron transfer	138, 139, 140, 141, 142, 143, 144
Atransferrinemia		3q21	Autosomal recessive	Transferrin gene mutations and red cell transfusions	145, 146
Aceruloplasminemia		3q23-q24	Autosomal recessive	Ceruloplasmin gene mutations	147, 148, 149, 150, 151, 152, 153, 154
Hereditary hyperferritinemia and cataract syndrome		19q13.1-q13.3.3	Autosomal dominant	L-Ferritin gene mutations	155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165
Friedreich ataxia		9p23-p11,9q13	Autosomal recessive	Frataxin gene mutations	166, 167, 168, 169, 170, 171, 172, 173
Panthothenase kinase-associated neurodegeneration		20p13-p12.3	Autosomal recessive	Pantothenase kinase gene mutations	174, 175
β-Thalassemia major		11p15.5	Autosomal recessive	β-Globin gene mutations, chronic hemolysis, red cell transfusions	—
Other chronic hemolytic anemias
Hereditary X-linked sideroblastic anemia		Xp11.21	X-Linked	δ-Aminolevulinic acid synthase gene mutations	176, 177, 178
X-Linked sideroblastic anemia with ataxia		Xq13.1-q13.3	X-Linked	ABCB7 mutations^a	179, 180, 181, 182
MLASA syndrome^b		12q24.33	Autosomal recessive	Pseudouridine synthase-1 mutations	183, 184
GLRX5 sideroblastic anemia		14q32.13	Autosomal recessive	Glutaredoxin 5 mutations	185, 186
DMT1 iron overload^c		12q13	Autosomal recessive	SLC11A2 mutations^d	187, 188
Pyruvate kinase deficiency		1q21	Autosomal recessive	Pyruvate kinase gene mutations	189, 190, 191
G6PD deficiency^e		Xq28	X-Linked	G6PD gene mutations	192, 193
Congenital dyserythropoietic anemias		Type I 15q15.1-q15.3	Autosomal recessive	Ineffective erythropoiesis	194, 195
	Type II 20q11.2	Autosomal recessive	Ineffective erythropoiesis	196, 197, 198
	Type III 15q21	Autosomal dominant	Ineffective erythropoiesis	199, 200
Acquired Disorder		Cause of Iron Loading
Transfusions		Red cell iron infusion
Medicinal iron		Excessive iron ingestion
Iron injections		Parenteral injection
Myelodysplasia with ring sid-eroblasts		Excessive iron absorption; transfusion
Portacaval shunt		Excessive iron absorption
Hemodialysis		Iron infusion
Nonalcoholic fatty liver disease		Excessive iron absorption
^aABCB7, ATP-binding cassette, subfamily B, member 7. ^bMLASA, myopathy with lactic acidosis and sideroblastic anemia. ^cDMT1, divalent metal transporter-1. ^dSLC11A2, solute carrier family 11, member 2. ^eG6PD, glucose-6-phosphate dehydrogenase.

The second most common HFE gene mutation in hemochromatosis patients is a change in nucleotide 187 from the normal cytidine to guanidine 187C→G in exon 2. This results in substitution of aspartate for the normal histidine at amino acid position 63. Both the C282Y and H63D mutations were identified in the initial report of the isolation and cloning of HFE.³⁶

Of the reported mutations of the HFE gene, 31 are missense mutations that result in substitution of the normal amino acid by another. There are 27 reported nonsense mutations of HFE, 12 of which are splice errors that are associated with intron sequence variants. Eight mutations involve a nucleotide change that does not result in an amino acid substitution (synonymous mutations). Some important HFE gene mutations are given in Table 25.2. Other mutations occur in noncoding regions of HFE.²⁴⁰ It is expected that other rare mutations of the HFE gene will be identified in adults with hemochromatosis phenotypes.

HFE Mutations in Hemochromatosis Patients and in Different Populations

In the first report of the HFE gene and its mutations, 83% of subjects with hemochromatosis were C282Y homozygotes.³⁶ The prevalence of homozygosity for C282Y varies from 33% to 100% among groups of hemochromatosis patients. In the United States, 59% to 89% of hemochromatosis patients are C282Y homozygotes. The percentage of hemochromatosis patients from 14 countries who have the C282Y and H63D genotypes is presented in Table 25.3.³⁶, ⁹⁸, ²²⁸, ²⁴¹, ²⁴², ²⁴³, ²⁴⁴, ²⁴⁵, ²⁴⁶, ²⁴⁷, ²⁴⁸, ²⁴⁹, ²⁵⁰, ²⁵¹, ²⁵², ²⁵³, ²⁵⁴, ²⁵⁵, ²⁵⁶, ²⁵⁷, ²⁵⁸, ²⁵⁹, ²⁶⁰, ²⁶¹, ²⁶², ²⁶³, ²⁶⁴, ²⁶⁵, ²⁶⁶, ²⁶⁷, ²⁶⁸, ²⁶⁹, ²⁷⁰, ²⁷¹, ²⁷², ²⁷³, ²⁷⁴, ²⁷⁵, ²⁷⁶

TABLE 25.2 SIXTY-SEVEN MUTATIONS OF THE HEMOCHROMATOSIS GENE (HFE) IN HEMOCHROMATOSIS TYPE 1: NUCLEOTIDE AND AMINO-ACID CHANGES, TYPE OF MUTATION, EXON OR INTRON INVOLVED, AND EFFECT OF THE MUTATION ON IRON LOADING

Change			Type of Mutation		Effect on Iron Loading		Reference
Nucleotide Complementary DNA Position	AminoAcid	Symbols	Missense	Nonsense	Exon or Intron Affected	Probands or Homozygotes
c.-20G→A		fs	—	Yes	5′UTR	↑	201
88C→T	Leu30Leu	L30L	No	No	Exon 2	None	202
128G→A;^a	Gly43Asp;	G43D;	Yes	—	Exon 2	↑	203
187C→G	His63Asp	H63D
138T→G	Leu46Trp	L46W	Yes	—	Exon 2	↑	204
c.del149-170^b	Leu50fs	L50fs	—	Yes	Exon 2	None	205, 206
128G→A	Gly43Asp	G43D	Yes	—	Exon 2	↑	15
157G→A	Val53Met	V53M	Yes	—	Exon 2	None	207
175G→A	Val59Met	V59M	Yes	—	Exon 2	None	207
187C→G	His63Asp	H63D	Yes	—	Exon 2	↑	36
187C→G^a	His63Asp;	H63D;	Yes	—	Exon 2,4	↑	208
845G→C	Cys282Tyr	C282Y
189T→C	His63His	H63H	No	No	Exon 2	None	207
193A→T	Ser65Cys	S65C	Yes	—	Exon 2	↑	209
196C→T	Arg66Cys	R66C	Yes	__	Exon 2	↑	202
199C→T	Arg67Cys	R67C	Yes	__	Exon 2	↑	206
c.del	203	Val68fs	V68fs	__	Yes	Exon 2	↑↑	210
211C→T	Arg74Stop	R71X^c	—	Yes	Exon 2	↑↑	15, 206, 239
277G→C	Gly93Arg	G93R	Yes	—	Exon 2	↑↑	211
c.del	277	Gly93fs	G93fs^d	__	Yes	Exon 2	↑↑	212
314T→C	Ile105Thr	I105T	Yes	—	Exon 2	↑	211
340G→A	Glu114Lys	E114K	Yes	—	Exon 2	↑	206
IVS2 (+4)T→C	Splice error	—	—	—	Intron 2	None	213
381A→C	Gln127His	Q127H	Yes	—	Exon 3	↑	207
385G→A	Asp129Asn	D129N	Yes	—	Exon 3	None	204
414C→G	Try138Stop	Y138X	—	Yes	Exon 3	↑↑	204
471del	Ala158fs	A158fs	—	Yes	Exon 3	↑↑214
c.del478	Pro160fs	P160fs	—	Yes	Exon 3	↑↑	215
502G→C	Glu168Gln	E168Q	Yes	—	—	↑	216
502G→C;^a	Glu168Gln;	E168Q;	Yes	—	Exon3	↑	217
187C→G	His63Asp	H63D
502G→T	Glu168Stop	E168X	—	Yes	Exon 3	↑↑	218
506G→A	Trp169Stop	W169X	—	Yes	Exon 3	↑↑	218
527	C→T	Ala176Val	A176V	Yes	—	Exon 3	↑	219
548T→C	Leu183Pro	L183P	Yes	—	Exon 3	↑↑	220
IVS3 (+1)G→T	Splice error	Null allele	—	Yes	Exon 3	↑↑	221
IVS3 (+21)T→C	Gly43Asp	G43D	Yes	—	Intron 3	↑	202
IVS3(+21)T→C	Splice error	__	__	Yes	Intron 3	None	202
IVS3(-48)C→G	Splice	error	__	__	Yes	Intron 3	None	222
c.del616-48C→T	fs	X	__	Yes	Exon 4	None	223
636G→C	Val212Val	V212V	No	No	Exon 4	None	224
671G→A	Arg224Gly	R224G	Yes	__	Exon 4	↑	202
689A→T	Tyr230Phe	Y230F	Yes	__	Exon 4	↑↑	204
c.del691-693	Tyr231fs	Y231X	__	Yes	Exon 4	↑↑	225
696C→T	Pro232Pro	P232P	No	No	Exon 4	None	202
697C→T	Gln233Stop	Q233X	__	Yes	Exon 4	↑↑	226
c.dup794	Trp267fs	W267fs	__	Yes	Exon 4	↑↑	227
724G→A	Asp242Asp	D242D	No	No	Exon 4	None	223
747G→A	Lys249Lys	K249K	No	No	Exon 4	None	223
814G→T	Val272Leu	V272L	Yes	—	Exon 4	?	228
829G→A	Glu277Lys	E277K	Yes	—	Exon 4	None	229
845G→A	Cys282Tyr	C282Y	Yes	—	Exon 4	↑↑	36
845G→C	Cys282Ser	C282S	Yes	—	Exon 4	↑↑	230
845G→A^a	Cys282Tyr;	C282Y;	Yes	—	Exon 4	↑	231
842C→A	Thr281Lys	T281K
848A→C	Gln283Pro	Q283P	Yes	__	Exon 4	↑↑	232
884T→A	Val295Glu	V295E	Yes	__	Exon 4	None	223
884T→C	Val295Ala	V295A	Yes	—	Exon 4	?	210
867G→C	Leu289Leu	L289L	No	No	Exon 4	None	202
IVS4 (+37)A→G	Splice error	—	—	Yes	Intron 4	None	207
IVS4 (+48) G→A	Spice error	—	—	Yes	Intron 4	None	233
IVS4 (+109)A→G	Splice error	—	—	Yes	Intron 4	None	207
IVS4 (-44)T→C	Splice error	—	—	Yes	Intron 4	?	234
IVS4(-50) A→G	Splice error	—	—	Yes	Intron 4	?	235
IVS4 (+115)T→C	Splice error	—	—	Yes	Intron 4	None	207
942T→C	Synonymous	—	No	No	—	None	219
989G→T	Arg330Met	R330M	Yes	—	Exon 5	↑↑	207
IVS5 (+1)G→A	Splice error	—	—	Yes	Intron 5	↑	236
IVS5 (-47)G→A	Splice error	—	—	Yes	Intron 5	?	234
c.1022-1034del13	His341fs	H341X	—	Yes	Exon 6	↑↑	234
HFEdel	HFEdel	—	—	Yes		↑ or ↑↑	237, 238
↑, increased; ↑↑, markedly increased; ?, effect unknown; A, adenine; C, cytosine; G, guanine; T, thymine; fs, frameshift; IVS, intron sequence variant; 5′UTR, untranslated region.
^aDouble missense complex allele in cis (on same allele). ^bOriginally published as 370del22.²⁰⁵, ²⁰⁶ ^cOriginally described as R74X.¹⁵, ²⁰⁶ ^dW94fs may be correct designation.²⁰⁶

The prevalence of HFE mutations is high in some populations, such as those of Central, Western, and Northern Europe, and in countries that were populated predominantly by people who originated in these areas of Europe, including the United States, Canada, and Australia.²⁴¹, ²⁷⁷ It is predicted that the prevalence of HFE mutations is very low in parts of the world with little or no ancestry from Central and Western Europe, including sub-Saharan Africa (Native Africans), the Middle East, the Orient, and Native Americans in North, Central, and South America. The frequency of homozygosity for the HFE C282Y mutation in the United States varies from 0% to 1.00% in whites, 0% to 0.22% in Mexican Americans, and 0% to 0.06% in blacks (Table 25.4). The prevalence of C282Y, H63D, and wild-type HFE genotypes from nine studies in the United States is presented in Table 25.4.²¹¹, ²⁴², ²⁴⁴, ²⁷⁸, ²⁷⁹, ²⁸⁰, ²⁸¹, ²⁸², ²⁸³ Extensive data about genotype frequencies in many countries, and from different regions within countries, have been published.²⁴¹, ²⁷⁷, ²⁸⁴

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