H63D Mutations Research Articles

S4368 Atypical Presentation of Beta-Thalassemia Trait with Heterozygous H63D Mutation: Hemolysis, Iron Overload, and Liver Cirrhosis

S4368 Atypical Presentation of Beta-Thalassemia Trait with Heterozygous H63D Mutation: Hemolysis, Iron Overload, and Liver Cirrhosis

Prevalence of C282Y, H63D and S65C gene mutations in patients with hyperferritinemia undergoing therapeutic phlebotomy suspected of having hereditary hemochromatosis

Background: Hemochromatosis is an autosomal recessive disorder caused by excessive iron overload in parenchymal organs. In Brazil, the prevalence of hereditary hemochromatosis (HH) may vary compared to other countries due to different ethnical backgrounds according to the geographic region studied. The prevalence of C282Y mutation of the HFE gene ranges from three to eight times lower in Brazilians than in Caucasians of Northern European origin. Methods: A retrospective cross-sectional study was carried out in 222 patients submitted to therapeutic phlebotomy between March 2011 and July 2014 in a tertiary hospital blood center. The prevalence of HH was determined in 89 (47.3%) out of 188 (84.6%) patients presenting hyperferritinemia, and divided in two genotype groups: C282Y mutation and other mutations. The HH patients’ geographical and clinical characteristics, serum ferritin rate (SF) and transferrin saturation (TS) were analyzed. Results: The HH group consisted of 71 males (79.7%) and 18 (20.3%) females, with a median age of 51.5 ± 10.6 years. There were 36 (40.0%) symptomatic patients and 65 (73.0%) tested positive for the HFE mutation. The initial baseline TS values were significantly higher in primary HH patients (48.6%, IC 95% 43.2% - 55.7%) compared to those with secondary HH (35.0%, IC 95% 27.5% 41.8%, p=0.001). Conclusion: The prevalence of C282Y mutation was higher in this population from the South region compared to previous reports in the Brazilian population, yet inferior to that of Northern Europeans.

The Exploring of Growth Differentiation Factor-15 and H63D Gene Polymorphisms in β-thalassemia Major: Implications for Cardiovascular Risk and Iron Overload

BACKGROUND: β-thalassemia major (βTM) is a genetic disorder characterized by a deficiency in hemoglobin production, ineffective erythropoiesis, chronic hemolysis, lifelong blood transfusions, iron overload, and increased risk of cardiac complications. OBJECTIVE: The study aimed to evaluate the growth differentiation factor-15 (GDF-15) concentration in βTM patients and its correlation with cardiac complications. H63D refers to a specific mutation in the HFE gene, which is associated with hereditary hemochromatosis (HH), a genetic disorder characterized by excessive accumulation of iron in the body. This mutation involves a change of histidine (H) to aspartic acid (D) at position 63 in the HFE protein. This mutation is often only written abbreviated as (H63D). MATERIALS AND METHODS: This case–control study was done on 120 subjects. A total of 60 patient samples were randomly collected from the Genetic Hematology Center at the Babylon Hospital, with an age range of 10–26 years. In addition, 60 samples were collected from healthy children in the same age range as the control group; patients and controls were subdivided into (10–18) and (18–26) year groups. GDF-15 was measured by enzyme-linked immunosorbent assay, and the genotyping of mutation was done by amplification refractory mutation system-polymerase chain reaction technique. RESULTS: The study revealed a significant increase in ferritin (FER) and GDF-15 levels in the patients compared to controls (P < 0.001). GDF-15 showed a direct correlation with age (r = 0.244, P = 0.02) and FER (r = 0.215, P = 0.04). There was a significant difference in H63D mutations between controls and patients (P = 0.044), with a higher proportion of the C-G (heterozygous for the mutant allele) genotype observed in βTM patients (31.67%). Additionally, a notable effect of the H63D mutation on serum ferritin (higher) levels within the βTM group was observed. CONCLUSION: Elevations of the GDF-15 in βTM patients indicate a high risk of cardiovascular complications in patients with βTM. The H63D mutation of the hemostatic iron regulator (HFE) gene is frequently found in βTM. Although a significant effect of the mutation was obtained on serum FER levels, it did not act as a risk factor in βTM patients. However, the frequent presence of the H63D mutation in patients indicated a possible association between single-nucleotide polymorphism and the iron regulation pathway.

Serum Erythroferrone Levels in a Large General Population: Towards a Better Understanding of Connections between Erythropoiesis and Iron Homeostasis

Introduction: Iron is essential for various biological functions [ Ganz T., Physiol Rev 2013], especially for erythropoiesis which is the most iron-consumer process daily. Hepcidin is the key hormone of iron homeostasis, preventing both iron deficiency and excess, by modulating iron absorption, recycling, and mobilization [ Camaschella C. et al, Haematologica 2020]. Erythroferrone (ERFE) constitutes the principal erythroid regulator of hepcidin: in the presence of enhanced erythropoiesis, expanding erythroblasts produce ERFE which, by impeding bone morphogenetic proteins (BMPs) signaling in hepatocytes, suppress hepcidin, in turn increasing iron availability for the bone marrow [ Ganz T., Free Rad Biol Med 2019]. Increased ERFE levels have been demonstrated in expanded/ineffective erythropoiesis (IE) (e.g., in thalassemia intermedia), where chronic ERFE-mediated hepcidin suppression may cause progressive iron accumulation even in subjects not regularly transfused. In view of this, ERFE has been proposed as a potential biomarker of IE and an appealing target for treating iron overload deriving from IE [ Srole D.N. and Ganz T., J Cell Physiol 2020]. However, studies that fully elucidate the role of ERFE in the general population under physiological conditions are still lacking. Our project evaluated age- and sex-related variations in ERFE levels, and their correlations with hematological and iron status parameters, in a large subpopulation from the Cooperative Health Research In South Tyrol (CHRIS) study. Subjects and methods: The CHRIS study is a population-based study carried out in South Tyrol (Northern Italy) aimed at investigating acquired and genetic basis of common chronic conditions associated with aging [ Pattaro C. et al, J Transl Med 2015]. Blood samples were tested for several biochemical and molecular parameters, including those related to iron status, such as ferritin, transferrin saturation (TSat), soluble transferrin receptor (sTfR), hepcidin, and C282Y and H63D mutations. Serum ERFE was measured in stored aliquots by the Human Erythroferrone IETM ELISA kit (Intrinsic Lifesciences - The BioIron Company TM, USA), according to manufacturer instructions. Results: We measured ERFE in 3688 subjects aged >18 years (55% female). A total of 777 subjects (21% of the whole population; 57% females) had ERFE concentrations below the lower limit of detection for our method (we assigned them an arbitrary value of 0.16 for statistical analyses). Figure 1 illustrates age- and sex-dependent variations of ferritin, sTfR, hepcidin, and ERFE in the whole population. After stratification by hemoglobin (Hb) levels, we observed that ERFE was significantly higher in anemic (i.e., Hb<12 g/dl in females and <13 g/dl in males; n=100) than in non-anemic subjects (respectively 3.74 and 0.65 ng/ml; p<0.0001). In anemic subjects, ERFE showed a strong negative correlation with Hb (r = -0.5; p < 0.001), TSat (r = -0.58; p < 0.001), ferritin (r = -0.48; p < 0.001) and hepcidin (r = -0.32; p = 0.001), and a positive correlation with sTfR (r = 0.58; p < 0.001). Moreover, ERFE was higher in iron-deficient (ID) subjects (i.e., ferritin <30 mg/l; n=773) than in non-ID subjects (0.92 vs. 0.63 ng/ml; p<0.0001). Although ID subjects had lower Hb levels than non-ID subjects (13.7 vs 15 g/dl; p<0.0001), Hb levels were broadly within the normal range.To better explore the relationship between ERFE and iron status, we divided the population into 4 subgroups, according to sTfR and ERFE levels. Considering the skewed distribution of the variables, we used sTfR above or below the 3° tertile (1316 ng/ml), in order to identify subjects having higher or lower iron needs, respectively. Analogously, we further divided the population in subjects with ERFE above or below the 3° tertile (1.21 ng/ml). As illustrated in Figure 2, after stratification, we observed that hepcidin reduced, in subjects with high ERFE levels, when iron requirement was high (0.94 vs 2.10 nM; p < 0.0001), but it didn't when iron requirement was low (3.35 vs. 3.05 nM; p = 0.72), even in presence of normal Hb levels (Hb 14.3 g/dl). Discussion: our data confirmed an association among ERFE, Hb and iron status parameters under conditions of anemia (when erythropoiesis is stimulated); moreover, if confirmed in further studies, it could suggest an effect of ERFE on hepcidin regulation could be linked to depletion of iron stores, even before anemia develops.

Common Mutation in the HFE Gene Modifies Recovery After Intracerebral Hemorrhage.

Intracerebral hemorrhage (ICH) is characterized by bleeding into the brain parenchyma. During an ICH, iron released from the breakdown of hemoglobin creates a cytotoxic environment in the brain through increased oxidative stress. Interestingly, the loss of iron homeostasis is associated with the pathological process of other neurological diseases. However, we have previously shown that the H63D mutation in the homeostatic iron regulatory (HFE) gene, prevalent in 28% of the White population in the United States, acts as a disease modifier by limiting oxidative stress. The following study aims to examine the effects of the murine homolog, H67D HFE, on ICH. An autologous blood infusion model was utilized to create an ICH in the right striatum of H67D and wild-type mice. The motor recovery of each animal was assessed by rotarod. Neurodegeneration was measured using fluorojade-B and mitochondrial damage was assessed by immunofluorescent numbers of CytC+ (cytochrome C) neurons and CytC+ astrocytes. Finally, the molecular antioxidant response to ICH was quantified by measuring Nrf2 (nuclear factor-erythroid 2 related factor), GPX4 (glutathione peroxidase 4), and FTH1 (H-ferritin) levels in the ICH-affected and nonaffected hemispheres via immunoblotting. At 3 days post-ICH, H67D mice demonstrated enhanced performance on rotarod compared with wild-type animals despite no differences in lesion size. Additionally, H67D mice displayed higher levels of Nrf2, GPX4, and FTH1 in the ICH-affected hemisphere; however, these levels were not different in the contralateral, non-ICH-affected hemisphere. Furthermore, H67D mice showed decreased degenerated neurons, CytC+ Neurons, and CytC+ astrocytes in the perihematomal area. Our data suggest that the H67D mutation induces a robust antioxidant response 3 days following ICH through Nrf2, GPX4, and FTH1 activation. This activation could explain the decrease in degenerated neurons, CytC+ neurons, and CytC+ astrocytes in the perihematomal region, leading to the improved motor recovery. Based on this study, further investigation into the mechanisms of this neuroprotective response and the effects of the H63D HFE mutation in a population of patients with ICH is warranted.

Iron overload leading to cirrhosis in a patient with hereditary spherocytosis and heterozygosity for H63D mutation in the HFE gene

Hereditary spherocytosis (HS) refers to a group of autosomal dominantly inherited heterogeneous hereditary haemolytic anaemias (HHA). Significant iron overload in HS is uncommon. Iron overload has been described as a complication of HS when there is co-inheritance of hereditary haemochromatosis (HH). H63D mutation accounts for a minority of hereditary hemochromatosis cases and does not cause iron overload in an otherwise healthy heterozygous carrier state. We present a 64-year-old man , diagnosed with HS presenting with cirrhosis without significant on-going haemolysis. He had a markedly high serum ferritin and transferrin saturation. Magnetic Resonance Imaging for liver iron concentration revealed haemochromatosis of the liver. HFE genotyping showed heterozygosity for the H63D mutant allele. Haemolysis in HS does not usually result in clinically significant iron excess leading to haemochromatosis and the presence of H63D heterozygous mutation will increase the risk of clinically significant iron overload which can lead to hepatic iron deposition and fibrosis. Therefore, if a patient with H63D mutation demonstrates clinically significant iron overload, clinicians should search for other factors that increase iron excess such as on-going haemolysis, alcohol abuse and presence of metabolic syndrome.

HFE hemochromatosis gene polymorphism and porphyria cutanea tarda

Purpose. To study the frequency of genotypes and alleles of C282Y and H63D mutations in the HFE gene in patients with porphyria cutanea tarde of the West Siberian region and their influence on the clinical and biochemical characteristics of the disease. Materials and methods. We observed 14 patients with porphyria cutanea tarde (PCT), who underwent a comprehensive general clinical and instrumental examination. The indicators of the excretory profile of porphyrin metabolism indicators were purposefully determined, a molecular genetic examination was carried out to determine the genotypes and alleles of the C282Y and H63D mutations of the hemochromatosis gene HFE. Results and discussion. Molecular genetic research found that the HFE gene mutation was found in 7 patients (50.0%). Polymorphism for the C282Y allele was found in 2 (14.3%) patients, and for the H63D allele - in 5 patients (35.7%). Risk factors for the manifestation of PCT included frequent alcohol consumption and chronic HCV infection. Genotype 1b was recorded more often (9 people), less often - genotype 3a (4 people). The excretory profile of porphyrin metabolism indices in patients of both groups exceeded the control values. Porphyrin metabolism parameters in patients without HFE gene mutations were significantly higher than those in patients without HFE gene mutations. Conclusions. Polymorphism of the hemochromatosis gene HFE in PCT was detected in 50% of patients. The most common mutation was the H63D allele. The level of porphyrin metabolism disorders in patients with HFE hemochromatosis gene mutations is significantly lower. The clinical picture of PCT in all observed patients did not differ. Chronic viral hepatitis C is assessed as a risk factor for the manifestation of PCT.

Hereditary hemochromatosis: An update vision of the laboratory diagnosis

Haemochromatosis (HC) is an inherited disorder of iron metabolism. The 85–90% of Hereditary hemochromatosis cases are caused by mutations in HFE gene (HC type 1). The remaining 10–15% of HC cases are caused by mutations in other non-HFE genes (HJV, HAMP, TRF2, SLC40A1, BMP6). The study of patients for the diagnosis of HC has an important laboratory approached: analysis of biochemical parameters and genetic studies. To confirm a case, it is necessary to carry out a genetic study of the C282Y and H63D mutations. The presence of C282Y mutation in homozygosis is compatible with the diagnosis of HC type 1. Due to the incomplete penetrance of this mutation and the variable phenotypic expression, the severe forms of the disease are relatively rare. The study of variants in non-HFE genes allows more detailed study of both non-classic HC cases and those with more severe clinical expression. The genotype characterization of a patient not always justified the phenotype expression of the symptoms in this disease. All laboratory clinicians must consider recommendation provide by the experts in the Materia.

5613463 PREVALENCE OF HEMOCHROMATOSIS HFE GENE POLYMORPHISM AMONG EGYPTIAN βTHALASSEMIA PATIENTS AND ITS RELATION TO DEVELOPMENT OF DIABETES MELLITUS

Background: β-thalassemia constitutes a major health problem in Mediterranean countries. Impaired glucose tolerance and diabetes mellitus are observed complications in patients with thalassemia. However, the mechanisms leading to diabetes are still poorly understood.H63D gene polymorphism, is the second variant of hemochromatosis (HFE) gene, that causes hemochromatosis. Aims: We aimed to assess the frequency of diabetes mellitus in β -thalassemia patients and the frequency of H63D gene polymorphism as well as its relation to development prediabetes and diabetes mellitus in β -thalassemia patients. Methods: Case control study was conducted on 50 patients with β-thalassemia and 50 healthy control with matched age and sex. The age ranged from 6-18y. Study subjects were investigated with oral glucose tolerance test, fasting C-Peptide, fasting insulin, anti -gad and screened for frequency of H63D mutation of HFE gene using restriction fragment length polymorphism. Results: 32% of patients had normal glycemic state, 52% had impaired glucose tolerance and 16% were diabetic P<0.001. H63D mutation was more frequent in thalassemia patient. GG genotype was significantly higher in β- thalassemia patients versus healthy controls (68%vs24%) P<0.001. GG genotype was prevalent in 77%and 75% of prediabetes and diabetic thalassemia patients respectively, versus 50% in patients with normal glycemic state P<0.001. Conclusions: GG genotype of H63D mutation of hemochromatosis HFE gene was frequent in our β- thalassemia patients which may be considered as a genetic risk factor for development of impaired glucose tolerance and diabetes in patients with. β-thalassemia. Keywords: thalassemia; diabetes; HFE gene mutation

Effect of HFE Gene Mutations on Iron Metabolism of Beta-Thalassemia Carriers

The human hemochromatosis protein HFE is encoded by the HFE gene and participates in iron regulation. The aim of this study was to detect the most frequent HFE gene mutations in a control population and in β-thalassemia trait (BTT) carriers, and to study their relationship with iron metabolism. Total blood count, hemoglobin electrophoresis at alkaline pH, HbA2 quantification, iron (Fe), total Fe binding capacity and ferritin were assayed. HFE gene mutations were analyzed by real-time PCR. A total of 119 individuals (69 normal and 50 BTT) were examined. In the control group, 9% (6/69) presented a codon 282 heterozygous mutation (C282Y), and 19% a codon 63 mutation (H63D) (13/69, 11 heterozygotes and 2 homozygotes). In the BTT group, 3 carriers (6%) were heterozygous for C282Y, 14 (28%) for H63D, 1 (2%) for a codon 65 mutation and 1 (2%) was H63D and C282Y double heterozygous. Control group Fe metabolism did not show significant differences (p &gt; 0.05) according to whether or not they carried an HFE gene mutation; while the BTT group with and without HFE mutation showed higher Fe and ferritin than the control group (p &lt; 0.05). However, no increases in iron parameters were detected in BTT carriers that simultaneously exhibited an H63D mutation compared to BTT subjects without a mutation. Therefore, the iron metabolism alterations observed in BTT carriers could not be attributed to the presence of HFE gene mutations. It is likely that BTT individuals have other genetic modifiers that affect their iron balance.

Type 1 Hemochromatosis: A Review of the History, Inheritance and Treatment of Iron Overload Disorder

Hereditary hemochromatosis (also known as type 1 hemochromatosis, iron overload disorder, or the Celtic Curse) is a genetic disorder characterized by an autosomal recessive inheritance pattern. This review project focused on gathering comprehensive data about the history, causes, inheritance, diagnosis, and treatment of iron overload disorder. Through such research, it was discovered that type 1 hemochromatosis may occur due to one of two nucleotide sequence changes (either the C282Y mutation or the H63D mutation) in the HFE gene, which codes for the construction of the homeostatic iron regulator protein and regulates production of the hormone hepcidin. As a result of these mutations, individuals affected by hemochromatosis have elevated iron levels and excess deposits of iron in tissues and organs. Such deposits may cause a variety of symptoms and complications, including fatigue, skin discoloration, heart failure, cirrhosis, liver cancer, and diabetes. Hemochromatosis is one of the most common genetic disorders in the world, affecting one in every three hundred individuals. The majority of hemochromatosis patients are of northern European descent. Currently, there is no cure for type 1 hemochromatosis; however, the primary treatment method for this disorder is phlebotomy. Due to the widespread prevalence of the disorder, researchers continue to explore experimental treatment methods, including gene therapy and iron chelation therapy.

DIFFERENCES IN THE FREQUENCY OF POLYMORPHISMS IN THE HFE GENE (H63D AND C282Y) IN A POPULATION GROUP FROM NORTHWEST SÃO PAULO, BRAZIL

Hereditary hemochromatosis (HH) is an autosomal recessive disease characterized by excess iron. People with polymorphisms in the HFE gene ( H63D and C282Y ) may develop inflammatory processes resulting from changes in iron metabolism. The ability of the HFE protein to make connections with transferrin receptors, located on cell membranes, allows it to help control the absorption of iron in the body. This work sought to investigate the frequency of HFE gene variants in a sample from the city of São José do Rio Preto, SP, Brazil. DNA samples from 105 people were subjected to molecular analysis to confirm mutations in the HFE gene ( H63D and C282Y ), using PCR-RFLP. The group was separated by male and female gender, age group of 10 in 10 years, and ethnic groups. Statistical analyzes were performed using the IBM SPSS Statistics 20 software and data were presented as percentages, with a significance level of 0.05. The associations between the wild-type homozygote, heterozygote, and mutant homozygote genotypes for each mutation were tested by the Chi-Square test. Then, the allele frequencies in the genotypes were calculated. Fisher's exact test was performed to verify the association between gender, age group, and ethnicity with mutations in the HFE gene. The sample included: 55 men (52.38%) and 50 women (47.62%), with ages ranging from 18 to 62 years, with an average of 30 years. A total of 210 alleles were evaluated and for the H63D mutation, 78 wild-type homozygotes, 26 heterozygotes, and 1 mutant homozygote were found. For the C282Y mutation, 99 wild-type homozygotes, 6 heterozygotes and no mutant homozygote were found. Allelic frequency analysis indicated the occurrence of significant differences between wild-type homozygous, heterozygous, and mutant homozygous genotypes for the mutations (p < 0.05). Fisher's exact test indicated no association between gender, age, and ethnicity for the H63D and C282Y polymorphisms (p > 0.05). The heterozygous genotype for H63D was more frequent in men (15-14%) than women 11 (10%). For C282Y equal frequencies (3-3%) were found for heterozygosity between men and women. H63D heterozygosity was more frequent in Caucasians (22-21%). In African-derived, only one individual was heterozygous for H63D . The frequency of HH genotypes varies among different populations. The low incidence or absence of homozygous alleles is due to the different degrees of miscegenation in the formation of the Brazilian population. The H63D mutation is an important marker for the Brazilian population, being frequent in Africa, although it has low phenotypic expression. The C282Y is predominantly associated with homozygosity for clinical cases, however, it is rare in African populations and frequent in Caucasians. Our results corroborate the data in the literature, with a predominance of the H63D mutation in Caucasians, African-derived, and Afro-descendants, and C282Y in Caucasians. The frequency of H63D and C282Y polymorphisms in the HFE gene showed a higher occurrence of heterozygotes for H63D in this population group. Studies to characterize HH are needed to devise strategies for effective support to patients and, therefore, improve quality of life.

S3055 Late Manifestation of Heterozygous H63D Mutation as Hereditary Hemochromatosis in 89-Year-Old Man

Introduction: Hereditary hemochromatosis (HH) is an autosomal recessive that usually affects middle-aged people [1]. In most patients, homozygous C282Y mutations in the HFE gene express the disease. But in some occurrences, compound heterozygous, including C282Y and H63D mutations, were also witnessed. We present an atypical case where heterozygous H63D manifested as HH in an 89-year-old man. Case Description/Methods: An 89-year-old Caucasian man of Italian descent presented with confusion, left-sided facial droop, and slurring of speech. Past medical history included end-stage renal disease on hemodialysis and atrial fibrillation. For the past few months before presentation, the patient has been complaining of increasingly tired with intermittent episodes of slurring speech, mainly after he receives dialysis. During the initial physical examination, the patient had confusion but was alert & oriented to self and time. There was mild abdominal distension and positive for asterixis, and the skin was bronze in color. Initial labs showed in table 1. The patient had persistent hypoglycemia and required intravenous glucose administration. Brain imaging was normal. CT scan of the abdomen, pelvis, and chest showed severe cardiomegaly and a moderate amount of ascites. Iron studies were abnormal. Ascites fluid analysis was consistent with cirrhosis. Although, an ultrasound right upper quadrant was negative for any cirrhotic morphology. MRI showed hemosiderosis. 2D Echo showed a severely dilated right ventricle. The patient empirically underwent two sessions of phlebotomy. Genetic testing confirmed heterozygous H63D mutation. The patient followed up for the phlebotomy sessions and demonstrated significant improvement. Discussion: This case highlights the consideration of HH in differential regardless of age if a patient presents with symptoms suspected of HH. Standard genetic testing includes testing C282Y and H63D mutations in the HFE gene on chromosome 6. Less frequently tested S65C, HFE4, TFR2-HHC, and FTH1 gene mutations also have the expression of HH in sporadic cases [2-3]. Previously, heterozygous C282Y carriers with iron overload are assumed to have other genetic changes or influence by environmental factors, such as alcohol or liver disease, that increase disease expression [2]. However, no such literature is available for heterozygous H63D carriers. Given the unusual age for phenotype expression and atypical disease, the presentation makes this unique case for learning the pathophysiology and genetics of HH.Figure 1.: A: MRI of liver showing decreased signal of the liver and spleen on in phase imaging is likely due to hemosiderosis. B: Two-dimensional transthoracic echocardiography showing severely dilated right ventricle and both atria. Moderate pulmonary hypertension was also noticed. Table 1. - Day 1 Day 2 Day 3 Day 4 Day 15 Day 42 Hemoglobin (g/dL) 15.6 14.8 15.1 13.8 13.6 12.2 AST (U/L) 85 102 135 125 109 71 ALT (U/L) 56 57 70 74 72 46 Bilirubin (mg/dL) 3.0 2.9 2.6 2.9 2.6 2.1 INR 1.3 1.4 1.4 1.4 1.2 1.2 Ammonia (µmol/L) 99 45 46 42 31 Platelets (10ˆ3/µL) 103 86 82 63 59 91 Ferritin (ng/ml) 1283 725 Iron (µg/dL) 187 130 Transferrin Saturation (%) 94 74 Total Iron-binding Capacity (µg/dL) 200 276

Far From the Biliary Tree: A Case of Overlapping Autoimmune Liver Disease in a Patient Presenting With Sicca Symptoms

Primary biliary cholangitis (PBC) is a chronic autoimmune condition with many extrahepatic manifestations that are commonly encountered as a patient's primary presenting complaints. Rarely, PBC co-exists as an “overlapping syndrome” with other liver-related autoimmune conditions such as autoimmune hepatitis (AIH). Presented is a rare case of PBC with features of AIH diagnosed in a patient who initially presented with hemoptysis and worsened sicca symptoms due to advanced Sjögren’s syndrome. The patient had a three-year evolution of abnormal liver biochemistry and was found to be a heterozygous carrier for hereditary hemochromatosis (H63D mutation). Given that patients with PBC-AIH are at an increased risk of complications compared to isolated disease from either disorder, early diagnosis and prompt management can help spare patients from cirrhosis, liver failure and transplantation, or even death.

Diagnosis of Liver Cirrhosis on the Background of Mutations H63D of the HFE Gene and H1069Q of the ATP7B Gene in associated with Hemochromatosis and Wilson's Disease (Clinical Case)

The purpose of the study was to ilustrate the analysis of etiological factors of liver cirrhosis using clinical and anamnestic data and the results of instrumental, laboratory and genetic researches. Materials and methods. The data of anamnesis and objective examination, results of instrumental, laboratory and genetic research methods are evaluated and analyzed. Modern protocols and medical literature were used. Results and discussion. Clinical case of the patient, 52 years old. Complaints of weakness, pain in the left hypochondrium, taste of iron, convulsions of the upper and lower extremities. Laboratory and instrumental methods of research allowed to establish the following indicators in the patient: erythrocytopenia, thrombocytopenia, neutropenia, persistent lymphocytosis, lecopenia, decreased platelet count, increased average erythrocyte volume and average hemoglobin content in one erythromycin distribution, albuminemia, increased beta globulin, decreased albumin to globulin ratio, increased liver enzymes (ALT, AST, bilirubin direct) and GGT, blood iron metabolism (COPD and iron levels), iron saturation and iron ferritin saturation, negative immunological analysis for antinuclear antibodies (ANA), HbS Ag and anti-HCV were not detected. The patient was consulted by a hematologist, lymphoproliferative diseases were excluded. On the basis of data on hepatosplenomegaly, portal hypertension, varicose veins of the esophagus, lymphadenopathy, excluding nonalcoholic fatty liver disease, alcoholic fatty liver disease, viral hepatitis, autoimmune hepatitis, biliary cirrhosis, diagnosed with a diagnosis on the detection of mutations that cause hemochromatosis and Wilson's disease. Molecular genetic studies have shown the following results: the H63D mutation of the HFE gene in the heterozygous state and the H1069Q mutation of the ATP7B gene in the heterozygous state were detected. Mutation testing and phenotype prediction based on genotype opens up prospects not only for personalized therapy, but also for the development of new treatment strategies. The literature provides data about new therapies with different mechanisms of action and discusses studies on Bis-choline tetrathiomolybdate in patients, pre-clinical studies of a novel chelator methanobactin and animal studies exploring cures for WD with gene therapy using adeno-associated vectors that introduce ATP7B into liver cells. Conclusion. The clinical case showed the need to involve specialists in various specialties and a set of research methods to establish the etiology of liver cirrhosis and further etiopathogenetic treatment and the formation of risk groups for primary prevention among relatives

PB2251: TWO POLYCYTHEMIA VERA PATIENTS WITH A DIAGNOSIS OF CONCOMITANT HEREDITARY HEMOCHROMATOSIS

Background: Hereditary Hemochromatosis (HH) is an inherited iron metabolism disorder characterized by excessive iron accumulation in the liver, heart, pancreas and other endocrine organs. Although it is often associated with mutations in the HFE gene, it can also be caused by mutations associated with HJV, TFR-2 and ferroportin. JAK2 V617F (+) Polycythemia Vera (PV) is the most common cause of acquired primary erythrocytosis and has been reported very rarely in patients with HH. Aims: We presented two polycythemia vera patients with a diagnosis of concomitant hereditary hemochromatosis. Methods: Presence of HFE gene mutation and jak-2 V617F mutation was determined by PCR test Results: Case 1: N.G, a 67-year-old male patient with a history of hypertension, insulin-dependent diabetes mellitus and cerebrovascular accident was diagnosed with PV in another center in 2007. The patient was JAK2 V617F (+) and the tests performed in our laboratory after phlebotomies were EPO: 7.8 mIU/ml, ferritin: 34 ng/dl, transferrin saturation: 49%. Bone marrow aspiration biopsy revealed hypercellularity with erythroid and megakaryocytic hyperplasia. Although Hb/Hct levels were within the targeted range with hydroxyurea 500 mg 2x1 treatment, he was followed up with frequent phlebotomies due to persistent pruritus. After phlebotomy, there was a significant improvement in his pruritus. He was using clopidogrel as antiaggregant treatment. During follow-up, there was deterioration in blood glucose regulation and an increase in insulin requirement. In June 2018, he developed widespread hyperpigmentation on his skin. Due to unregulated diabetes, hyperpigmentation, and high ferritin levels despite frequent phlebotomy, he was examined for HH. He was found to be heterozygous positive for the HFE gene H63D mutation. The patient is still being followed up with HU 2X1 and phlebotomy without any problem. Case 2: E.H, a 57-year-old male patient with a history of hypertension and type 2 diabetes mellitus was diagnosed with PV in 2015. He was JAK2 V617F(+) and at diagnosis EPO:3.7 mIU/ml, ferritin:1403 ng/ml, transferrin saturation:63%. Bone marrow aspiration biopsy revealed hypercellularity and erythroid hyperplasia. Since he was a low-risk patient under the age of 60 and had no history of thrombosis, he was followed up with ASA 100 mg/day and intermittent phlebotomies. Similar to the first case, persistent pruritus was present despite the normal Hb/Hct level. Since hyperferritinemia was not expected in polycythemia vera and the patient was diabetic, he was examined for HH. In the HFE gene analysis, the H63D mutation was found to be heterozygous. Due to the frequent need for phlebotomy, the patient is considered to start treatment with hydroxyurea, and the follow-up still continues. Summary/Conclusion: The association of HH and PV has been rarely reported in the literature, and its clinical significance is not clearly known. In the presence of unexplained high ferritin levels in PV, even if the clinical findings are unclear, screening for HH may be recommended as an appropriate approach. The role of HFE genotypes in the development of myeloproliferative diseases is unknown. Clinical data accumulation is needed to elucidate iron metabolism and prognostic features in PV patients with HFE mutations. Keywords: polycythemia vera, hereditary hemochromatosis, JAK2 V617F, HFE mutation

P1556: HYPER -FERRITINEMIA IN PEDIATRIC ONCOLOGY PATIENTS SECONDARY TO TREATMENT: DIAGNOSTIC APPROACH AND OUTCOME

Background: Multiple red blood cell transfusions during cancer therapy are the leading cause of iatrogenic hyper- ferritinemia which of notice is often overlooked. Patients with genetic predisposition for hemochromatosis have a higher chance of developing hyper-ferritinemia, with a potential to promote significant organ dysfunction. Aims: The aim of this study was to identify pediatric oncology patients with high levels of ferritin post cancer treatment, and refer their management and outcome. Methods: Selected patients’ charts of our Department were retrospectively reviewed. Patient and treatment characteristics, ferritin levels and their progression over time, genetic testing for hemochromatosis genes, treatment and outcome were recorded and analyzed. Results: Twelve (12) patients were identified with increased (>600 ng/ml) ferritin levels, diagnosed with Acute Lymphoblastic Leukemia (N=5), Acute Myeloid Leukemia (N=1), Non-Hodgkin Lymphoma (N=1), Neuroblastoma (N =2), Medulloblastoma (N=1), Rhabdomyosarcoma (N=2) from 1995 to 2020. Mean patient age at diagnosis was 7.1 years (range 3.0-15.2 years). Three patients had undergone Hematopoietic Stem Cell Transplantation, 2 of them twice. Mean blood transfusions per patient were 42 (range: 9-62), with 6 patients receiving more than 50 transfusions each (10-15 ml/kg). Of 6 patients tested for hemochromatosis genes, 2 were negative for the common mutations of HFE, TfR2 FPN1 genes, 1 was found double heterozygote for the His63Asp and Cys282Tyr HFE mutations and 1 patient carried the HFE H63D mutation. Eight (8/12) patients were observed only, and ferritin reduction of 5.7% to 69.7% was observed, within 12 months of follow-up, but none normalized ferritin levels, the lowest value being 560 ng/ml (Image). One patient received chelation therapy (deferasinox po) for 13 months, 1 patient (heterozygous for beta-thalassemia trait) received 37 phlebotomy sessions and 2 received combination of chelation (deferasirox only or with deferoxamine) and short-course phlebotomies. Out of the later intervention group of patients, none achieved normal ferritin levels after an observation period of 12 months. With a mean follow up of 3.7 years for all patients (range 1,1-15,7 years), mean ferritin levels after cancer therapy and at +12 months, were 2056 ng/ml (range: 621-4750 ng/ml) and 1227 ng/ml (range: 590-3550 ng/ml), respectively. Only 1 patient achieved ferritin levels under 500 ng/ml following 3-years of phlebotomies, and she has remained so, for another 4 years of follow-up (b-thalassemia heterozygote and Η63D HFE heterozygote for the hemochromatosis genes). Image:Summary/Conclusion: Iatrogenic iron overload post antineoplastic therapy in pediatric patients is a common and often overlooked diagnosis. Prompt evaluation and appropriate treatment is necessary. Patients presenting with hyper-ferritinemia for extended time period could bear hemochromatosis gene mutations. Since the symptoms are absent initially, pediatric patients after cancer treatment should be screened for ferritin levels. If persistently high levels are documented, evaluation of relevant mutations and appropriate treatment is necessary. The long-term effect of high Ferritin levels is not fully documented, but it has the potential to promote significant organ dysfunction. Further research is necessary in this field.

20-Year-Old Man With Elevated Liver Tests

20-Year-Old Man With Elevated Liver Tests

Comparison between H63D and G71D gene mutation effects on iron overload in Iraqi patients with β-thalassemia major: A case–control study

Comparison between H63D and G71D gene mutation effects on iron overload in Iraqi patients with β-thalassemia major: A case–control study

Screening for hereditary haemochromatosis in patients undergoing knee arthroplasty : a retrospective cohort study of 2,035 patients.

AimsHereditary haemochromatosis is a genetic disorder that is caused by several known mutations in the human homeostatic iron regulator protein (HFE) gene. Abnormal accumulation of iron causes a joint disease that resembles osteoarthritis (OA), but appears at a relatively younger age and is accompanied by cirrhosis, diabetes, and injury to other organs. Increased serum transferrin saturation and ferritin levels are known markers of haemochromatosis with high positive predictive values.MethodsWe have retrospectively analyzed the iron studies of a cohort of 2,035 patients undergoing knee joint arthroplasty due to OA.ResultsNo patients had HFE gene C282Y, S65C, or H63D mutations testing. In total, 18 patients (2.96%) of the male cohort and 51 (3.58%) of the female cohort had pathologically increased ferritin levels that may be indicative of haemochromatosis. Seven patients (0.34%) had serum transferrin saturation above 45%.ConclusionThe awareness for the diagnosis of this disorder in Orthopaedics is low and needs improvement. Osteoarthritic patients undergoing knee arthroplasty should be routinely screened for haemochromatosis by iron studies and referred to genetic testing when needed.Level of evidence: Level III - Retrospective cohort study.Cite this article: Bone Jt Open 2021;2(12):1062–1066.