XmnI Polymorphism Research Articles

Molecular characterization of \u03b2-thalassemia intermedia in the West Bank, Palestine

BackgroundWe aimed to investigate the molecular basis of β-Thalassemia intermedia (TI) in the West Bank region and its management practices.MethodsThis was a case series multi-center study and included 51 cases of TI. DNA sequencing was used to analyze β-globin gene mutations. Common α-globin gene mutations were screened by Gap-PCR (−α3.7, −α4.2, −-MED, αααanti3.7) or DNA sequencing (α2-IVS II 5 nt del). XmnI -158 C > T polymorphisms of Gγ-globin gene was determined by RFLP-PCR.ResultsSeven β-globin gene mutations were observed, namely IVS-I -6 C > T, IVS-I-110 G > A, IVS-II-1 G > A, IVS-I-1 G > A, Codon 37 Trp > Stop, beta − 101 and IVS-II-848 C > A. Ten genotypes were observed. Homozygosity for IVS-I-6 accounted for the majority of TI cases with a frequency of 74.5%. The second common β-globin gene genotype was homozygote IVS-I-110 G > A (5.8%) and homozygote IVS-II-1 G > A (5.8%). The remaining seven genotypes were each detected in about 2% of patients. α-Thalassemia mutations were seen in five patients (9.8%), and included (−α3.7, αααanti3.7 and α2-IVSII-5 nt del). XmnI polymorphism was observed in four patients (7.8%), three homozygotes and one heterozygote.ConclusionsHomozygosity for the mild β-globin gene IVS-I-6 allele was the major contributing factor for the TI phenotype among the study subjects. The role of XmnI SNP and α-thalassemia mutations in ameliorating the TI phenotype was observed in few patients for each factor. The beta − 101 C > T mutation was diagnosed in one patient in homozygote state for the first time in Palestine.

Consanguinity, the driving force behind inheritance of HbS-β thalassemia in Southern Districts of KP.

To determine the clinical, haematological and genetic factors responsible for variable phenotypes of sickle haemoglobin, sickle haemoglobin-beta, and beta-thalassemia patients. The study was conducted in Bannu, Lakki, Tank and Dera Ismail Khan districts of Khyber Pakhtunkhwa province of Pakistan from September 2016 to November 2017, and comprised sickle haemoglobin, sickle haemoglobin-beta, and beta-thalassemia patients. Clinical, haematological and genetic determinants were evaluated using haemoglobin electrophoresis and allele-specific primers through polymerase chain reaction to determine alpha and beta thalassemia, and CgT substitution at position -158 (referred to as Xmn-I polymorphism) in gamma-globin gene. Data was analysed using SPSS 20. Eight b-thalassemia mutations were identified that included IVS I-5(G C), codon 8/9 (+G), codon 30 (G C), -88 (C T), Cap+1(A G), codon 41/42 (-TCTT), IVS I-1(G T) and codon 16(-C). Codon 30 (G C) and -88 (C T) were found only in Pashtoon subjects, Cap+1(A G) and IVS I-1(G T) in Balochi subjects, while 75% of IVS I-5(G C) mutation cases were found in Punjabi ethnic group. In the Pashtoon group, 13 sickle haemoglobin homozygous patients were identified for the first time. Both alpha thalassemia and Xmn-I polymorphism in homozygous condition were common among those with mild phenotype. Phenotypic expression of sickle haemoglobin beta thalassemia was found to be extremely variable and alpha thalassemia and Xmn-I polymorphism in homozygous condition were found to be additional genetic modifiers of the disease.

Genetic determinants related to pharmacological induction of foetal haemoglobin in transfusion-dependent HbE-β thalassaemia.

Thalassaemia are the most common inherited autosomal recessive single gene disorders characterized by chronic hereditary haemolytic anaemia due to the absence or reduced synthesis of one or more of the globin chains. Haemoglobin E-β thalassaemia is the genotype responsible for approximately one half of all severe beta-thalassaemia worldwide. This study proposes to evaluate the effect of various molecular parameters on the response of hydroxyurea. Hydroxyurea was started at an initial dose of 10mg/kg of body weight/day on 110 transfusion-dependent HbE-β thalassaemia patients. HbF level was measured by HPLC analysis. β-Thalassaemia mutations, XmnI and five other SNPs, and α-globin gene deletions and triplications were detected by ARMS-PCR, RFLP-PCR and Gap-PCR, respectively. Based on the factors for evaluating hydroxyurea-response, 42 patients were responders as they showed an increment of Hb from a mean baseline value of 6.45g/dl (± 0.70) to 7.78g/dl (± 0.72) post-therapy. Based on increase in HbF above the median value (14.72%) post-therapy, 78 patients were found to be responders. All the 78 responders showed mean decrease in transfusion of 74.26% (± 8.32) with a maximum decrease of 98.43%. There was a significant correlation between decrease in transfusion and increase in HbF level for all 78 responders. XmnI polymorphism showed the strongest association (p < 0.0001) with increase in HbF levels and Hb levels. Patients with α-globin gene deletions were better responders. It was concluded that hydroxyurea treatment is effective in transfusion-dependent HbE-β thalassaemia patients and the response is best in patients having both XmnI polymorphism and α-deletion.

Mutation Screening of the Krüppel-like Factor 1 Gene in Individuals With Increased Fetal Hemoglobin Referred for Hemoglobinopathy Investigation in South of Iran.

Any mutation in the Krüppel-like factor 1 (KLF1) gene may interfere with its proper related function in the erythropoiesis process and lead to alterations in proper activation of its downstream protein through globin switching, which results in an increase in fetal hemoglobin (HbF). This study aimed to investigate whether KLF1 mutation can associate with high level of HbF in individuals with increased fetal hemoglobin referred for screening of hemoglobinopathies in south of Iran. The human KLF1 gene was amplified via the polymerase chain reaction procedure, and sequencing was used to determine any mutation in these patients. Moreover, XmnI polymorphisms in the position of -158 of γ-globin gene promoter were analyzed in all patients by polymerase chain reaction restriction fragment length polymorphism. Analysis of sequencing revealed a missense mutation in the KLF1 gene, p.Ser102Pro (c.304T>C), which was detectable in 10 of 23 cases with elevated HbF level. This mutation was only detected in individuals who had a HbF level between 3.1% and 25.6%. Statistical analysis showed that the frequency of C allele is significantly correlated with a high level of HbF (P<0.05). The allele frequency of positive result of XmnI polymorphism in individuals with increased HbF level was also significant, which showed an association with increased HbF level (P<0.05). To the best of our knowledge, this is the first report of p.Ser102Pro (c.304T>C) in the KLF1 gene in β-thalassemia patients with increased level of fetal hemoglobin. According to statistical results of p.Ser102Pro mutation and XmnI polymorphism, it has been strongly suggested that both polymorphisms have an association with increased HbF samples. These nucleotide changes alone may not be the only elements raising the level of HbF, and other regulatory and modifying factors also play a role in HbF production.

XmnI polymorphism in Egyptian patients with β-thalassemia major and its correlation with the HbF level

Clinical severity of β-thalassaemia depends on the types of β-gene mutations involved. It can also be influenced by genetic factors like concomittant α-thalassaemia and increased γ-chain production. Several loci are implicated in higher production of HbF. The XmnI restriction site at −158 position of the Gγ-gene is associated with increased expression of the Gγ-globin gene and higher production of HbF. This study aims to determine the frequency of the Gγ XmnI polymorphism in β-thalassaemia patients in Egypt and its correlation to the HBF level and clinical severity of the disease. We investigated the XmnI polymorphism in 100 children with β-thalassaemia major using polymerase chain reaction (PCR-RFLP)-restriction fragment length polymorphism. We found that ninety-four children had XmnI (−/−) genotype (94%) and six children had XmnI (+/−) genotype. On the other hand, the study found that the presence of this polymorphism influences HbF concentration and ameliorate the clinical severity of the disease.

Quantitative Trait Loci Influencing Hb F Levels in Southern Thai Hb E (HBB: c.79G>A) Heterozygotes

Variation of fetal hemoglobin (Hb F) expression in heterozygous Hb E (HBB: c.79G>A) individuals is associated with several genetic modifiers and not well understood. This study was undertaken in order to determine the effect of single nucleotide polymorphisms (SNPs), including XmnI Gγ (rs7482144), rs766432 on the BCL11A gene and rs9376074 on the HBS1L gene, on Hb F levels in Southern Thai heterozygous Hb E individuals. A total of 97 Southern Thai subjects carrying heterozygous Hb E were selected for the hematological study. After excluding the samples with α-thalassemia (α-thal) interaction or moderate anemia, because both conditions can affect the hematological parameters, the remaining 74 samples were submitted to SNP analysis. Hematological parameters were measured using an automated hematology analyzer and high performance liquid chromatography (HPLC). The results show that rs766432 was strongly associated with increased Hb F levels and rs7482144 was associated with Hb F levels in each subgroup (genotype) of rs766432. This study suggested that the BCL11A locus has a major effect on Hb F levels compared with the XmnI polymorphism in Hb E heterozygotes. This association of Hb F levels with SNPs is useful for the interpretation of hemoglobin (Hb) typing in heterozygous Hb E samples with high Hb F levels. Future research will need to address the better understanding of the mechanisms of the SNPs that regulate Hb F production without stress erythropoiesis in Hb E heterozygotes.

Genetic Background of the Sickle Cell Disease Pediatric Population of Dakar, Senegal, and Characterization of a Novel Frameshift β-Thalassemia Mutation [HBB: c.265_266del; p.Leu89Glufs*2

Sickle cell disease is a genetic disorder with a large variability in the pattern and severity of clinical manifestations. Different genetic modulators have been identified but very few epidemiologic data are available on these modifier genes in Senegal. This study aimed to determine their prevalence in a Senegalese sickle cell disease pediatric population. The following genetic parameters were genotyped in 295 sickle cell disease children of the Dakar pediatric hospital: sickle cell disease genotype [βS/βS (HBB: c.20A>T), βS/βC (HBB: c.19G>A), βS/β0-thalassemia (β0-thal)], XmnI polymorphism, the five most common α-thalassemia (α-thal) deletions and the A(–) and Betica glucose-6-phosphate-dehydrogenase (G6PD) deficient variants. Despite very few βS/βC and βS/β0-thal children (1.0% each), a novel frameshift β0-thal mutation was characterized: HBB: c.265_266del; p.Leu89Glufs*2. The –α3.7 (rightward) deletion was the only α-thal deletion identified in this cohort (12.0% allelic frequency). Most of βS/βS patients (61.9%) were homozygous for the XmnI polymorphism and assumed to carry a Senegal/Senegal βS haplotype. The remaining haplotypes were predominantly of the Benin type. While the Betica G6PD variant was quite frequent (13.0%), a low frequency of the A(–) variant was detected (1.0–2.0%). The systematic genotyping of the –α3.7 deletion and of the G6PD Betica variant in sickle cell disease patients from Senegal could be useful to identify patients at risk for several complications, such as cerebral vasculopathy, where it has been demonstrated that a normal α-globin genotype and G6PD deficiency are predisposing factors. These patients should be eligible for a transcranial Doppler examination that is not routinely offered in Senegal.

New Insights on β-Thalassemia in the Palestinian Population of Gaza: High Frequency and Milder Phenotype Among Homozygous IVS-I-1 (HBB: c.92+1G>A) Patients with High Levels of Hb F

β-Thalassemia (β-thal) is a very common disease in the Palestinian population of the Gaza Strip. We studied their mutation frequency and clinical features. Thirteen different mutations were identified. The most common mutation was IVS-I-1 (G>A) (HBB: c.92+1G>A), which was prevalent in 31.5% of the thalassemia alleles studied. The IVS-I-110 (G>A) (HBB: c.93−21G>A) mutation was found in 25.0% of the alleles. Homozygotes for the IVS-I-1 mutation had higher mean hemoglobin (Hb) levels, required less blood transfusions, and lower transferrin saturation than the homozygotes for the IVS-I-110 mutation. This milder phenotype was, most likely, the result of the persistent production of Hb F; it was 9-fold higher in absolute terms (g/dL) and 7.7-fold higher in relative terms (percentage of total Hb). About half of our IVS-I-1 patients carried the XmnI polymorphism, which is known to be associated with elevated Hb F levels.

The molecular characterization of Beta globin gene in thalassemia patients reveals rare and a novel mutations in Pakistani population

IntroductionA multicentre study (including four cities in Pakistan) aimed to investigate the frequency and spectrum of alpha and beta thalassemia genetic mutations and XmnI polymorphism of the Gamma Globin gene. MethodsOne hundred and sixty one beta thalassemia patients, identified on the ground of haematological parameters, were screened for mutations of the Alpha (HBA2 and HBA1) and Beta (HBB) Globin genes as well as Gamma (HBG2) Globin gene, -158 Gγ XmnI polymorphism, using a combination of multiplex GAP polymerase chain reaction (PCR), Sanger sequencing and restriction fragment length polymerase (RFLP) based PCR. ResultsMutations of at least one HBB gene was identified in 157 of 161 patients screened. Among 16 identified mutations in the beta gene, HBB:c.27_28insG (p. Ser10Valfs*14) was the most prevalent. α−3.7 and α−4.2 deletions were co-inherited with beta thalassemia mutations. Rare mutations such as HBB:c.–138C > T and HBB:c.315 + 1G > A were also identified. One novel variant (HBB:c.-148T > A), two rare mutations [HBB:c.332T > C (p.Leu111Pro); HBB:c.92G > C (p.Arg31Thr] and a novel association, HBB:c.[92G > C (p.Arg31Thr)] and [-92C > G], were reported for the first time in our study. HBG2:c.-211C > T base-pair substitution (historically described as -158 GγXmnI polymorphism) was present in 36% of the patients. ConclusionHeterogeneity in clinical and haematological parameters in TM, show that monogenic disorders can present with a wide spectrum of disease severity. Our studies identified rare and novel mutations that will be useful in the prevention of highly prevalent disease of thalassemia in Pakistan following nationwide awareness campaign.

Structural and Functional Insights on an Uncharacterized Aγ-Globin-Gene Polymorphism Present in Four β0-Thalassemia Families with High Fetal Hemoglobin Levels

Several DNA polymorphisms have been associated with high production of fetal hemoglobin (HbF), although the molecular basis is not completely understood. In order to identify and characterize novel HbF-associated elements, we focused on five probands and their four families (from Egypt, Iraq and Iran) with thalassemia major (either β(0)-IVSII-1 or β(0)-IVSI-1) and unusual HbF elevation (>98 %), congenital or acquired after rejection of bone marrow transplantation, suggesting an anticipated favorable genetic background to high HbF expression. Patient recruitment, genomic DNA sequencing, western blotting, electrophoretic mobility shift assays, surface plasmon resonance (SPR) biospecific interaction analysis, bioinformatics analyses based on docking experiments. A polymorphism of the Aγ-globin gene is here studied in four families with β(0)-thalassemia (β(0)-IVSII-1 and β(0)-IVSI-1) and expressing unusual high HbF levels, congenital or acquired after rejection of bone marrow transplantation. This (G→A) polymorphism is present at position +25 of the Aγ-globin genes, corresponding to a 5'-UTR region of the Aγ-globin mRNA and, when present, is physically linked in chromosomes 11 of all the familiar members studied to the XmnI polymorphism and to the β(0)-thalassemia mutations. The region corresponding to the +25(G→A) polymorphism of the Aγ-globin gene belongs to a sequence recognized by DNA-binding protein complexes, including LYAR (Ly-1 antibody reactive clone), a zinc-finger transcription factor previously proposed to be involved in down-regulation of the expression of γ-globin genes in erythroid cells. We found a novel polymorphism of the Aγ-globin gene in four families with β(0)-thalassemia and high levels of HbF expression. Additionally, we report evidence suggesting that the Aγ-globin gene +25(G→A) polymorphism decreases the efficiency of the interaction between this sequence and specific DNA binding protein complexes.

Identification of β-globin haplotypes linked to sickle hemoglobin (Hb S) alleles in Mazandaran province, Iran.

Carrier frequency of the βS allele has been reported to be 0.19% in Mazandaran province, northern Iran. Haplotype analysis of the βS allele helps trace the origin of its encoded hemoglobin (Hb) variant, Hb S, in a region. The aim of this study was to investigate the haplotypes associated with βS alleles in Mazandaran province. Capillary electrophoresis was carried out to detect individuals suspected to have a βS allele(s). DNA analysis (PCR-RFLP) was used for final confirmation. To identify 5' to 3' β-globin gene cluster haplotypes associated with βS alleles, family linkage analysis was applied. Six polymorphic sites (HincII 5' to ε, XmnI 5' to Gγ, HindIII in Gγ, HindIII in Aγ, HincII 3' to ψβ and AvaII in β) were investigated using the PCR-RFLP method. Five different haplotypes were linked to βS alleles, while βA alleles were associated with nine haplotypes. Among the βS alleles, 53.9% were associated with the Benin (----++) haplotype, and the Arab-Indian (+++-++) haplotype had the second-highest frequency (23%). Unlike southern provinces, where the Arab-Indian haplotype is prominent, the Benin haplotype is the most frequent haplotype in northern Iran, and this may represent a founder effect. Since the Benin haplotype does not carry the XmnI polymorphism 5' to the Gγ gene, which is responsible for high expression of Hb F, a severe form of sickle cell disease can be anticipated in patients that are homozygous for the βS allele in the northern region.

Inheritance of IVSI-I Mutation Assures 158 (C-T) XmnI Polymorphism in Thalassemia Intermedia.

Thalassemia intermedia lies between asymptomatic β-thalassemia carrier state to severe thalassemia Major. XmnI polymorphism is one of the modifying factors of intermedia phenotype. This study intendeds to determine the frequency of Xmn-I polymorphism and its association with different beta chain mutations that would contribute towards the phenotype. 100 whole blood samples of thalassemia intermedia patients were tested for the common beta chain mutations found in Pakistan and then tested for Xmn-I polymorphism. The most common mutation identified in these patients was IVSI-5. Xmn-I polymorphism was found in 79% of the patients, 36% being homozygous and 43% being heterozygous. Xmn-I polymorphism was found in 84.7% of the samples with IVSI-5 mutation, 52.1% of Fr 8-9 , 45% of Cap+1, 85.7% of Cd30, 91.66% of HbE, and 33.3% of the samples with del619 mutation. Whereas δβ, HbS, IVSI-I showed 100% positivity for Xmn-I polymorphism. The age of start of transfusion was 3 -9 years and had high hemoglobin F levels. All the samples with IVSI-I mutation were homozygous (+/+) for Xmn-I polymorphism. Thus a firm linkage between the XmnI polymorphism and IVSI-I mutation is suggested and that IVSI-I mutation always accompany homozygosity of Xmn-I polymorphism and the coinheritance of the two mutations will lead to a milder phenotype.

The Apolipoprotein A1 polymorphisms were associated with decreased risk for metabolic syndrome in Koreans

Apolipoprotein A1 (ApoA1) plays an important role in reverse cholesterol transport. The functional abnormality of ApoA1 is known to be the major risk for lipid disorders or cardiovascular diseases. In this study, we examined the associations of two ApoA1 polymorphisms (XmnI and MspI/rs670) with the Korean patients with metabolic syndrome (MetS). A total of 835 subjects were investigated, which includes 320 patients with MetS and 515 healthy controls. Genotyping of the ApoA1 polymorphisms was analyzed by conventional polymerase chain reaction and digestions with restriction endonucleases. The mutant genotypes of the XmnI and MspI polymorphisms were associated with the decreased risk of MetS. The X1-A and X2-G haplotypes for the XmnI and MspI polymorphisms were also protective to susceptibility for MetS. The XmnI and MspI polymorphisms showed significant associations with DBP (P = 0.047) and body mass index (P = 0.037) among MetS risk factors, respectively. Stratified analysis for MetS risk components was significantly associated between the XmnI polymorphism and blood pressure. Based on these results, the variants and haplotypes of the ApoA1 polymorphisms may be protective against MetS susceptibility in Koreans.

Thalassemia Modular Stratification System for Personalized Therapy of Beta-Thalassemia (THALAMOSS)

Thalassemia Modular Stratification System for Personalized Therapy of Beta-Thalassemia (THALAMOSS)

Molecular Basis of β-Thalassemia Intermedia in Erbil Province of Iraqi Kurdistan

β-Thalassemia intermedia (β-TI) is a clinical term describing a range of clinical phenotypes that are intermediate in severity between the carrier state and β-thalassemia major (β-TM). To characterize the molecular basis of β-TI in Erbil Province, Northern Iraq, 83 unrelated patients were investigated. Detection of β-globin gene mutations was carried out by reverse hybridization assay and direct gene sequencing. All patients were screened for the XmnI polymorphism by direct sequencing of HBG2 (Gγ promoter gene). Detection of α-globin gene deletions and triplication was carried out using the reverse hybridization assay. Four main molecular patterns were identified in association with the β-TI phenotype, namely: β+/β+ (38.5%), β+/β0 (21.6%), β0/β0 (31.3%), and β0/wild type (8.4%). IVS-I-6 (T > C) was the most frequently encountered mutation (55 alleles, 34.6%), followed by IVS-II-1 (G > A) and codon 8 (−AA); furthermore, we report for the first time from Iraq two β+ mutations, −87 (C > G) and 5′ untranslated region (5′UTR) +22 (G > A). The XmnI polymorphism was detected in 47.0% of patients, mainly in association with the β0/β0 genotype. The α-globin gene deletions were encountered in four cases, including one case with (– –FIL) double gene deletion, a report that is the first from our country. The α-globin gene triplication was detected in five of the seven heterozygous β-thalassemia (β-thal) patients. Similar to other Mediterranean countries, inheritance of mild β-globin mutations was the main molecular pattern underlying β-TI in our patients followed by the ameliorating effect of the XmnI polymorphism.

Genotype–phenotype correlation and report of novel mutations in β-globin gene in thalassemia patients

Heterogeneity in thalassemia is due to various modifying factors viz. coinheritance of α-gene defects, abnormal hemoglobin, XmnI polymorphism, variation in repeat sequences present in LCR, and silencer region of the gene. The present work on populations from eastern regions of India was undertaken to study the genetic profile of heterogeneity in thalassemia patients. Mutation analysis in 126 index families revealed the presence of 3 novel mutations: CD2 (-A) in the 1st exon, -42 (C-G), and -223 (T-C) in the promoter region of β-globin gene. The modifying effect of coexisting α-gene defects, and abnormal Hb (HbS) was clearly observed in our study, however ameliorating effect of T allele of XmnI polymorphism was not found. Analysis of the regulatory regions (LCR) exhibited new combinations (CA(15)TA(5) and CA(13)TA(8)) in HS1 region and one (AT)(10)T(3) in (AT)(x)T(y )silencer region. Thus disparate factors, when considered together, were able to explain several of the thalassemic phenotypes, otherwise not explained by the β globin mutations. However, there were still some cases in this group whose molecular origin could not be ascertained. Our findings confirm not only the extensive genotypic and clinical heterogeneity in β thalassemia but also the need to look for more modulators and modifiers to better understand the genotype-phenotype correlation in thalassemia.

Genetic Modifiers of Sickle Cell Disease: A Genotype-Phenotype Relationship Study in a Cohort of 82 Children on Mayotte Island

Sickle cell disease presents a great clinical variability that remains largely misunderstood. New disease protective genetic modifiers acting mainly through an increased Hb F level have recently been described. We studied relations between clinical and hematological phenotypes and known sickle cell disease genetic modifiers in patients from Mayotte Island, a remote French territory located in the Indian Ocean. Eighty-two children with sickle cell disease were enrolled; their median age was 5.9 years (range 1-18). Clinical and hematological features of sickle cell disease were retrospectively collected. Genetic studies included determination of β-globin genotypes [Hb SS, Hb S-β0-thalassemia (Hb S-β0-thal), Hb S-β+-thal], βS-globin locus haplotype, α-thalassemia (α-thal), and single nucleotide polymorphisms (SNPs) located in quantitative trait loci for Hb F expression (XmnI polymorphism, BCL11A rs4671393 and rs11886868, intergenic region of HBS1L-MYB rs28384513, rs4895441 and rs9399137). Univariate and multivariate analyses were conducted. Twenty-eight percent of the patients had Hb S-β-thal (eight different mutations in 21 patients), 55.0% had the −α3.7 (rightward) deletion and 88.0% of the homozygous Hb SS patients were carrying a homozygous Bantu haplotype. In the multivariate model, the prognosis role of the SNP BCL11A rs4671393 was confirmed in the studied population showing a significant association with an elevated Hb F level and with a low hospitalization rate. The −α3.7 deletion, XmnI polymorphism and intergenic region HBS1L-MYB SNPs were not significantly linked to any clinical criteria of severity. This report, the first to describe the main features of children with sickle cell disease on Mayotte Island, highlights the protective effect of the BCL11A polymorphism in this population.

Genetic Modifiers in β-Thalassemia Intermedia: A Study on 102 Iraqi Arab Patients.

To determine the molecular basis of β-thalassemia intermedia (TI) and the contribution of the three hemoglobin F (HbF) quantitative trait loci (QTLs) on chromosomes 11, 2, and 6 to the milder phenotype, a total of 102 Iraqi Arab patients with TI were studied. The β and α genotypes as well as HBG2 g. 158 C>T (rs7482144), BCL11A (rs1427407 and rs10189857), and HBS1L-MYB (rs28384513 and rs9399137) by multiplex polymerase chain reaction and reverse hybridization were studied. A total of 21 different β-thalassemia mutations arranged in 35 different genotypes were identified. The genotypes encompassed β(+)/β(+) mutations in 33 cases, β(+)/β(0) in 17 cases, β(0)/β(0) in 47 cases, β(0)/wild type in 3 and β(0)/Hb E in 2 cases. The most common was IVS-II-1 (G>A)/IVS-II-1 (G>A), followed by IVS-I-6 (T>C)/IVS-I-6 (T>C) and IVS-I-110 (G>A)/IVS-I-110 (G>A), in 31.4%, 17.6%, and 6.9%, respectively. Alpha-thalassemia mutations were found in 15.2% of those homozygous for the β-mutations, while α gene triplication was identified in all three heterozygotes. Of the five QTLs tested, only rs7482144 and rs10189857 were significantly associated with β(0)/β(0) when compared to β(+)/β(+), with odds ratios of 6.4 (95% confidence interval [CI] 2.9-14.0) and 3.2 (95% CI 1.2-8.6), respectively. In conclusion, this study has demonstrated that among Iraqi patients with thal intermedia, the main contributors to the milder phenotype were β(+) alleles, XmnI polymorphism, and BCL11A (rs10189857), while other QTLs on chromosomes 2 and 6, as well as alpha-thalassemia, were not significantly relevant.

Polymorphic variations influencing fetal hemoglobin levels: association study in beta-thalassemia carriers and in normal individuals of Portuguese origin.

Three major loci have been associated with HbF levels, including -158C/T (XmnI) at HBG2 promoter region, and several polymorphisms at BCL11A intron-2 and HBS1L-MYB (HMIP) intergenic region. Mutations in the KLF1 gene were recently associated with increased HbF levels. This study aims to evaluate whether genetic variability at these loci influences HbF levels in β-thalassemia carriers and in normal individuals of Portuguese origin. Sixty five β-thalassemia carriers, HbF levels ranging from 0.2% to 9.5%, and 60 individuals with normal hematological parameters, HbF levels ranging from 0.2% to 7.4%, were selected for this study. In β-thal carriers linear regression models revealed a strong statistical significant association for HBG2 (XmnI) rs7482144 (β=0.455; P=5.858×10(-7)), and nominal significance for BCL11A rs766432 (β=0.215; P=0.029) and HMIP rs9399137 (β=0.209; P=0.011). In normal individuals, a case (HbF>2%; n=15) vs. control (HbF<1.7%; n=45) model, showed nominal significant associations for BCL11A SNPs rs11886868 (OR=4; P=0.001), rs766432 (OR=3.7; P=0.002) and rs7606173 (OR=0.36; P=0.032). KLF1 rs3817621 was not found associated with HbF levels. Our results suggest that in Portuguese β-thal carriers the HBG2 XmnI polymorphism is strongly associated with HbF levels. In normal individuals, BCL11A polymorphisms, but not HMIP or HBG2 (XmnI) loci, are nominally associated with HbF expression.

XmnI polymorphism: Relation to β-thalassemia phenotype and genotype in Egyptian Children

Backgroundβ-Globin mutations with Xmn1 site might be associated with elevated HbF expression which may in turn ameliorate the severity of β-thalassemia phenotype. Aim of the studyTo investigate the frequency of −158 (C>T) XmnI polymorphism among Egyptian Children and young adults with β-thalassemia, to examine the relationship between XmnI polymorphism and β-thalassemia genotypes and phenotypes and to assess the possible relation of XmnI polymorphism and response to hydroxyurea (Hu) therapy. Patients and methodsSeventy-two β-thalassemia patients (37 females; M/F ratio 0.95) with a mean age of 7.53±6.99 were included. Laboratory investigations included Complete blood count (CBC), Hb electrophoresis by high performance liquid chromatography (HPLC), β-thalassemia mutation identification by the reverse dot blot hybridization technique (RDB) and detection of XmnlGg polymorphism by RFLP. ResultsThe frequency of positive heterozygote XmnI gene polymorphism was 8.3%. Eighty-three percent of XmnIGγ+/− patients were never transfused (p=0.001) and had higher total hemoglobin compared to XmnIGγ−/− (p=0.01); while mean HbF was higher among XmnIGγ+/− patients compared to the other group but the difference was marginally insignificant (p=0.06). β-Thalassemia mutation IVS II-1 showed relatively higher XmnI polymorphism frequency (50%) and followed by its frequency among 10 undefined β-thalassemia mutations which was 20%. The frequency of positive heterozygote XmnI gene polymorphism was 11.6% among the TI group vs. 3.5% among the TM group (p=0.4). Among 20 cases who received HU; 5/14 responders vs. 1/6 none responder had positive heterozygote XmnI gene polymorphism (p=1.0). Conclusions and recommendationsIn conclusion, molecular determination of genetic markers in childhood will help to identify phenotypes of our patients and to avoid over or under treatment strategies. Further prospective studies concerning the genetic markers that could predict the response to hemoglobin F inducers like hydroxyurea are highly recommended.