scholarly journals Complete Scanning of the Hereditary Hemochromatosis Gene (HFE) by Use of Denaturing HPLC

2001 ◽  
Vol 47 (9) ◽  
pp. 1633-1640 ◽  
Author(s):  
Gerald Le Gac ◽  
Catherine Mura ◽  
Claude Férec
Keyword(s):  
Hereditary Hemochromatosis ◽  
Hfe Gene ◽  
Test Accuracy ◽  
Melting Profile ◽  
Coding Region ◽  
Hemochromatosis Gene ◽  
Hfe Mutations ◽  
Methods Analytical ◽  
Positive Controls ◽  
Experimental Melting

Abstract Background: Between 4% and 35% of hereditary hemochromatosis (HC) probands are C282Y or H63D heterozygotes or lack both of these two common HFE mutations, and 15 novel HFE mutations have been described recently. We evaluated denaturing HPLC (DHPLC) for screening of the whole HFE coding region and further defined whether HC probands with an incomplete HFE genotype carry uncommon mutations. Methods: Analytical conditions for each coding exon were determined by a combination of computer melting profile predictions and experimental melting curves. To test accuracy for scanning the complete HFE coding region and optimize DHPLC running conditions, each melting domain was investigated with at least one mutation or one polymorphism as reference. We tested 100 DNA samples harboring the C282Y, H63D, or S65C mutations and 17 artificially created positive controls that carried either 1 of the 14 other known HFE mutations or 3 selected polymorphisms. Results: Investigations on each of the coding exons 1, 2, 4, 5, and 6 could be performed at one analysis temperature. Coding exon 3 displayed a more complex melting profile and required two analysis temperatures. DHPLC detected all known HFE mutations as well as the three selected polymorphisms. Conclusions: DHPLC can be used to scan the HFE gene in HC probands in whom at least one chromosome lacks an assigned mutation.

scholarly journals Precipitating factors of porphyria cutanea tarda in Brazil with emphasis on hemochromatosis gene (HFE) mutations. Study of 60 patients

2013 ◽  
Vol 88 (4) ◽  
pp. 530-540 ◽  
Author(s):  
Fatima Mendonca Jorge Vieira ◽  
Maria Cristina Nakhle ◽  
Clarice Pires Abrantes-Lemos ◽  
Eduardo Luiz Rachid Cancado ◽  
Vitor Manoel Silva dos Reis
Keyword(s):  
Hepatitis C ◽  
Alcohol Abuse ◽  
Porphyria Cutanea Tarda ◽  
Hereditary Hemochromatosis ◽  
Control Group ◽  
Hfe Gene ◽  
Precipitating Factors ◽  
Serological Tests ◽  
Hfe Mutations ◽  
Precipitating Factor

BACKGROUND: Porphyria cutanea tarda is the most common form of porphyria, characterized by the decreased activity of the uroporphyrinogen decarboxylase enzyme. Several reports associated HFE gene mutations of hereditary hemochromatosis with porphyria cutanea tarda worldwide, although up to date only one study has been conducted in Brazil. OBJECTIVES: Investigation of porphyria cutanea tarda association with C282Y and H63D mutations in the HFE gene. Identification of precipitating factors (hepatitis C, HIV, alcoholism and estrogen) and their link with HFE mutations. METHODS: An ambispective study of 60 patients with PCT was conducted during the period from 2003 to 2012. Serological tests for hepatitis C and HIV were performed and histories of alcohol abuse and estrogen intake were investigated. HFE mutations were identified with real-time PCR. RESULTS: Porphyria cutanea tarda predominated in males and alcohol abuse was the main precipitating factor. Estrogen intake was the sole precipitating factor present in 25% of female patients. Hepatitis C was present in 41.7%. All HIV-positive patients (15.3%) had a history of alcohol abuse. Allele frequency for HFE mutations, i.e., C282Y (p = 0.0001) and H63D (p = 0.0004), were significantly higher in porphyria cutanea tarda patients, compared to control group. HFE mutations had no association with the other precipitating factors. CONCLUSIONS: Alcohol abuse, hepatitis C and estrogen intake are prevalent precipitating factors in our porphyria cutanea tarda population; however, hemochromatosis in itself can also contribute to the outbreak of porphyria cutanea tarda, which makes the research for HFE mutations necessary in these patients

scholarly journals HFE Gene Mutation Status Predicts Response to Gemtuzumab Ozogamicin in AML

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1307-1307 ◽  
Author(s):  
Etienne Paubelle ◽  
Alice Marceau ◽  
Florence Zylbersztejn ◽  
Mickael Dussiot ◽  
Ivan Cruz Moura ◽  
...  
Keyword(s):  
Total Dose ◽  
Conflicts Of Interest ◽  
Hereditary Hemochromatosis ◽  
Gemtuzumab Ozogamicin ◽  
Hfe Gene ◽  
Study Cohort ◽  
Dependent Manner ◽  
Wild Type ◽  
Significant Difference ◽  
Hfe Mutations

Abstract BACKROUND: The protein defective in type 1 hereditary hemochromatosis, called HFE, is similar to MHC class I-type proteins, associates with beta2-microglobulin and is implicated in membranous protein recycling. Gemtuzumab ozogamicin (GO), a monoclonal antibody directed against CD33 linked to a cytotoxic agent, has been used with controversial effects in acute myeloid leukemia (AML). Internalization of GO is required for its anti-leukemic effect. Therefore, we hypothesized that H63D or C282Y HFE gene mutations may impair GO activity by preventing its internalization. METHODS: Wild type, C282Y and H63D HFE leukemic cells and primary cell were used to assess effects of GO alone or in combination with cytarabine on cell proliferation and apoptosis. Flow cytometry, confocal and AMNIS stream analysis were used to evaluate GO internalization. HFE mutations were analyzed by PCR analysis on DNA from patients included in clinical studies. Post-hoc subgroup analysis was performed to assess in vivo the role of HFE status on GO efficacy and toxicity among the 280 patients of the ALFA-0701 study as a study cohort (patients aged 50-70 years; GO 3mg/m² on days 1, 4, and 7 of chemotherapy and on day 1 of the first and second induction ; total dose 15 mg/m² ) and then on the GOELAMS-LAM 2006 IR study (patients aged 18-60 years; GO 6 mg/m² on day 4 of chemotherapy during the induction and the first consolidation; total dose 12 mg/m²) and UK NCRI AML17 study (patients aged 18-81 years; GO 3 vs 6 mg/m² on day 1 of chemotherapy but not during consolidation) as validating cohorts. RESULTS: GO induced cell death by apoptosis in AML cell lines and primary cells in a dose-dependent manner and synergistically in combination with cytarabine. However, the IC5O of GO was significantly higher in HFE mutated cells (125 vs 10 ng/mL p<0.001). To further understand this phenomenon, the CD33 internalization was analyzed upon GO or anti-CD33 antibody exposure. In line with the cytotoxic effect, CD33 was significantly less internalized in HFE mutant cells (17.4% vs 65.1% after 1 hour, p<0.01). HFE mutations were screened in 242 of the 280 ALFA-0701 patients with DNA available. There were 155 non-mutated patients (64%), 68 (28%) heterozygous for H63D, 11 (5%) heterozygous for C282Y, and 8 (3%) homozygous for H63D, which is consistent with the prevalence of the various mutations in the French population. Median age was 62 years (50-71) and the M/F ratio was 0.5, equally distributed among the different groups. No significant difference was observed with respect to the diagnosis of various hematological parameters, including white blood cell count, blasts number, cytogenetic subgroups, molecular mutation incidences (FLT3, NPM1, CEBPA, IDH, DNMT3A). In the ALFA-0701 study cohort, HFE wild-type (WT) patients had a higher overall survival (OS) when treated in the GO arm (median, not reached vs 19.5 months, p=0.0193). In contrast, OS was similar among patients with heterozygotes HFE mutations treated in the GO and the control arm (median, 19.9 vs 21.9, p=0.9675). In confirmatory cohorts, GO treatment led to a trend to increased OS in the GOELAMS-LAM 2006 IR cohort only in HFE WT patients (4-year OS with GO 62% vs 48%, p=0.08) but not in mutated patients (4-year OS with GO 73% vs 64%, p=0.45). Furthermore, in this cohort in the FLT3 WT patients subgroup, GO further improve OS in WT patients (4-year OS with GO 72% vs 50%, p=0.017), but not in patients with heterozygous HFE mutation (4-year OS with GO 80% vs 65%, p=0.23) In the UK NCRI AML17 cohort, which used GO only during induction, 245 patients were randomized between GO at 3mg/m2 and 6mg/m2 and evaluated for HFE status. Overall there was no effect of GO dose on outcomes, and no evidence of either any heterogeneity by HFE, nor any subgroup, which showed a differential effect of GO dose. CONCLUSIONS: Future studies should focus on optimising the fractionated schedule for GO at the 3mg/m2. Fractionated high doses (3x3mg/m2) during induction and single dose during consolidation seems to be the best schedule. Most importantly, the effect of GO treatment differed between HFE WT and heterozygote mutated AML patients. GO only increased OS among HFE WT patients. This is likely related to impaired internalization of the CD33 target. Our data suggest that HFE status should be used as a companion test to predict outcome of AML treated with GO. Disclosures No relevant conflicts of interest to declare.

Physiology of iron transport and the hemochromatosis gene

2002 ◽  
Vol 282 (3) ◽  
pp. G403-G414 ◽  
Author(s):  
Antonello Pietrangelo
Keyword(s):  
Iron Homeostasis ◽  
Cell Damage ◽  
Hereditary Hemochromatosis ◽  
Hfe Gene ◽  
Cell Functions ◽  
Cellular Iron ◽  
Histocompatibility Complex ◽  
Biological Studies ◽  
Hemochromatosis Gene ◽  
Main Protein

Iron is essential for fundamental cell functions but is also a catalyst for chemical reactions involving free radical formation, potentially leading to oxidative stress and cell damage. Cellular iron levels are therefore carefully regulated to maintain an adequate substrate while also minimizing the pool of potentially toxic “free iron.” The main control of body iron homeostasis in higher organisms is placed in the duodenum, where dietary iron is absorbed, whereas no controlled means of eliminating unwanted iron have evolved in mammals. Hereditary hemochromatosis, the prototype of deregulated iron homeostasis in humans, is due to inappropriately increased iron absorption and is commonly associated to a mutated HFE gene. The HFE protein is homologous to major histocompatibility complex class I proteins but is not an iron carrier, whereas biochemical and cell biological studies have shown that the transferrin receptor, the main protein devoted to cellular uptake of transferrin iron, interacts with HFE. This review focuses on recent advances in iron research and presents a model of HFE function in iron metabolism.

scholarly journals Multiplex Allele-Specific PCR for Simultaneous Detection of H63D and C282Y HFE Mutations in Hereditary Hemochromatosis

2018 ◽  
Vol 3 (1) ◽  
pp. 10-17
Author(s):  
Heleen H Arts ◽  
Barry Eng ◽  
John S Waye
Keyword(s):  
Hereditary Hemochromatosis ◽  
Simultaneous Detection ◽  
Turnaround Time ◽  
Molecular Diagnostic ◽  
Hfe Gene ◽  
Specific Pcr ◽  
Liver And Pancreas ◽  
Allele Specific ◽  
Hfe Mutations ◽  
Allele Specific Pcr

Abstract Background Hereditary hemochromatosis (HH) is characterized by excessive iron absorption in the intestine, which can lead to failure of vital organs such as the heart, liver, and pancreas. Among northern Europeans, HH is most often associated with the C282Y and H63D mutations of the HFE gene. We developed a test that allows screening for both mutations in a single reaction. Methods A multiplex allele-specific PCR was developed for simultaneous genotyping of the H63D and C282Y HFE mutations. PCR fragments were designed such that the resulting PCR product can be analyzed in a single polyacrylamide gel lane. Results Test results from our multiplex assay were concordant with genotypes of 55 Canadian patients with suspected hemochromatosis, which had previously been established by allele-specific PCRs that targeted H63D and C282Y in separate reactions. Conclusions Molecular diagnostic detection of H63D and C282Y mutations can be achieved by a variety of methods, but these are not necessarily time-efficient or economical. Multiplex allele-specific PCR is an excellent tool for molecular diagnostic screening for H63D and C282Y mutations in patients with suspected hemochromatosis. This method is inexpensive, accurate, and highly efficient in terms of labor, throughput, and turnaround time.

scholarly journals Hereditary hemochromatosis gene (HFE) mutations C282Y, H63D and S65C in patients with idiopathic dilated cardiomyopathy

2005 ◽  
Vol 7 (1) ◽  
pp. 103-108 ◽  
Author(s):  
Jokke Hannuksela ◽  
Mari Leppilampi ◽  
Keijo Peuhkurinen ◽  
Satu Kärkkäinen ◽  
Eija Saastamoinen ◽  
...  
Keyword(s):  
Dilated Cardiomyopathy ◽  
Hereditary Hemochromatosis ◽  
Idiopathic Dilated Cardiomyopathy ◽  
Hemochromatosis Gene ◽  
Hfe Mutations

scholarly journals From Environment to Genome and Back: A Lesson from HFE Mutations

2020 ◽  
Vol 21 (10) ◽  
pp. 3505
Author(s):  
Raffaela Rametta ◽  
Marica Meroni ◽  
Paola Dongiovanni
Keyword(s):  
Dietary Habits ◽  
Epigenetic Modification ◽  
Iron Status ◽  
Hereditary Hemochromatosis ◽  
Phenotypic Expression ◽  
Bioactive Molecules ◽  
Hfe Gene ◽  
Human Populations ◽  
Environmental Pressures ◽  
Hfe Mutations

The environment and the human genome are closely entangled and many genetic variations that occur in human populations are the result of adaptive selection to ancestral environmental (mainly dietary) conditions. However, the selected mutations may become maladaptive when environmental conditions change, thus becoming candidates for diseases. Hereditary hemochromatosis (HH) is a potentially lethal disease leading to iron accumulation mostly due to mutations in the HFE gene. Indeed, homozygosity for the C282Y HFE mutation is associated with the primary iron overload phenotype. However, both penetrance of the C282Y variant and the clinical manifestation of the disease are extremely variable, suggesting that other genetic, epigenetic and environmental factors play a role in the development of HH, as well as, and in its progression to end-stage liver diseases. Alcohol consumption and dietary habits may impact on the phenotypic expression of HFE-related hemochromatosis. Indeed, dietary components and bioactive molecules can affect iron status both directly by modulating its absorption during digestion and indirectly by the epigenetic modification of genes involved in its uptake, storage and recycling. Thus, the premise of this review is to discuss how environmental pressures led to the selection of HFE mutations and whether nutritional and lifestyle interventions may exert beneficial effects on HH outcomes and comorbidities.

scholarly journals Analysis of HFE gene mutations and HLA-A alleles in Brazilian patients with iron overload

2006 ◽  
Vol 124 (2) ◽  
pp. 55-60 ◽  
Author(s):  
Rodolfo Delfini Cançado ◽  
Aline Cristiane de Oliveira Guglielmi ◽  
Carmen Silvia Vieitas Vergueiro ◽  
Ernani Geraldo Rolim ◽  
Maria Stella Figueiredo ◽  
...  
Keyword(s):  
Iron Overload ◽  
Binding Capacity ◽  
Hereditary Hemochromatosis ◽  
Gene Mutations ◽  
Hfe Gene ◽  
Outpatient Unit ◽  
Laboratory Variables ◽  
Hfe Gene Mutations ◽  
Hfe Mutations ◽  
Iron Binding Capacity

CONTEXT AND OBJECTIVE: Hemochromatosis is a common inherited disorder of iron metabolism and one of the most important causes of iron overload. The objective was to analyze the presence of C282Y, H63D and S65C mutations in the HFE gene and HLA-A alleles for a group of Brazilian patients with iron overload, and to correlate genotype with clinical and laboratory variables. DESIGN AND SETTING: Prospective study, in Discipline of Hematology and Oncology, Faculdade de Ciências Médicas da Santa Casa de Misericórdia de São Paulo. METHODS: We studied 35 patients with iron overload seen at our outpatient unit between January 2001 and December 2003. Fasting levels of serum iron and ferritin, and total iron-binding capacity, were assayed using standard techniques. Determinations of C282Y, H63D and S65C mutations in the HFE gene and of HLA-A alleles were performed by polymerase chain reaction (PCR). RESULTS: Twenty-six out of 35 patients (74%) presented at least one of the HFE gene mutations analyzed. Among these, five (14%) were C282Y/C282Y, four (11%) C282Y/H63D, one (3%) H63D/H63D, six (17%) C282Y/WT and ten (29%) H63D/WT. No patients had the S65C mutation and nine (25%) did not present any of the three HFE mutations. Four out of five patients with C282Y/C282Y genotype (80%) and three out of four patients with C282Y/H63D genotype (75%) were HLA A*03. CONCLUSION: Analysis of HFE gene mutations constitutes an important procedure in identifying patients with hereditary hemochromatosis, particularly for patients with iron overload.

scholarly journals Frequency of the Hemochromatosis Gene (HFE) Variants in a Jordanian Arab Population and in Diabetics from the Same Region

2009 ◽  
Vol 27 (1) ◽  
pp. 17-22 ◽  
Author(s):  
Asem Alkhateeb ◽  
Amal Uzrail ◽  
Khaldon Bodoor
Keyword(s):  
Blood Donor ◽  
Hereditary Hemochromatosis ◽  
University Hospital ◽  
European Ancestry ◽  
Hfe Gene ◽  
Arab Population ◽  
Population Type ◽  
Significant Difference ◽  
Hemochromatosis Gene

Hereditary HFE-linked hemochromatosis is a frequent recessive disorder among individuals of northern European ancestry. The clinical characteristic of this disease is the gradual accumulation of iron in internal organs, which ultimately may lead to organ damage and death. Three allelic variants of HFE gene have been correlated with hereditary hemochromatosis: C282Y is significantly associated with hereditary hemochromatosis in populations of Celtic origin, H63D and S65C are associated with milder form of iron overload. In this study we performed mutation analysis to identify allele frequency of the three variants of HFE gene in Jordanian Arab population, to assess deviations of these frequencies from those detected elsewhere, and to determine if there is an increased frequency of these variants in a diabetic population (Type 2 diabetes) from the same area. DNA was extracted from blood samples of 440 individuals attending King Abdullah University Hospital for ambulatory services. We used polymerase chain reaction (PCR) to amplify exons 2 and 4 of the HFE gene then restriction fragment length polymorphism (RFLP) method to detect the variants. There were neither homozygous nor heterozygous for C282Y variant. For the H63D variant, 0.68% were homozygous and 21.1% were heterozygous. For the S65C variant, there were no homozygous and 0.23% were heterozygous. Allelic frequencies were, 0%, 11.25%, and 0.11% for C282Y, H63D, and S65C, respectively. Our samples were subdivided into two categories of type 2 diabetic (89 cases) and controls (blood donors, 204 cases) and compared with regard to the H63D variant. Both groups did not have homozygous H63D variant. H63D heterozygous in diabetics were 23.60% and in blood donor controls 22.55%. Allelic frequency of the mutant H63D allele was 11.80% in diabetics and 11.27% for the blood donor controls. This is the first study to show the frequency of the three hemochromatosis gene variants in Jordan with the interesting finding of no C282Y allele detected in 440 samples. Additionally, no significant difference was observed in H63D variant frequency in type 2 diabetics as compared to controls.

scholarly journals From Environment to Genome and Back: A Lesson from HFE Mutations

2020 ◽  
Author(s):  
Raffaella Rametta ◽  
Marica Meroni ◽  
Paola Dongiovanni
Keyword(s):  
Dietary Habits ◽  
Epigenetic Modification ◽  
Iron Status ◽  
Hereditary Hemochromatosis ◽  
Phenotypic Expression ◽  
Bioactive Molecules ◽  
Hfe Gene ◽  
Human Populations ◽  
Environmental Pressures ◽  
Hfe Mutations

The environment and the human genome are closely entangled and many genetic variations that occur in human populations are the result of adaptive selection to ancestral environmental (mainly dietary) conditions. However, the selected mutations may become maladaptive when environmental conditions change, thus becoming candidates for diseases. Hereditary hemochromatosis (HH) is a potentially lethal disease leading to iron accumulation mostly due to mutations in the HFE gene. Indeed, homozygosity for the C282Y HFE mutation is associated with the primary iron overload phenotype. However, both penetrance of the C282Y variant and the clinical manifestation of the disease are extremely variable, suggesting that other genetic, epigenetic and environmental factors play a role in the development of HH, as well as, and in its progression to end-stage liver diseases. Alcohol consumption and dietary habits may impact on the phenotypic expression of HFE-related hemochromatosis. Indeed, dietary components and bioactive molecules can affect iron status both directly by modulating its absorption during digestion and indirectly by the epigenetic modification of genes involved in its uptake, storage and recycling. Thus, the premise of this review is to discuss how environmental pressures led to the selection of HFE mutations and whether nutritional and lifestyle interventions may exert beneficial effects on HH outcomes and comorbidities.

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