Tunisian Newborn's Cord Blood: Reference Values of Complete Blood Count and Hemoglobin Fractions.

OBJECTIVE: This study was aimed to establish local reference values for hematological indices and hemoglobin (Hb) fractions in umbilical cord blood (UCB) for the northern population of Tunisia. STUDY DESIGN: Our study included full-term newborns by vaginal deliveries. Hematological parameters were collected using an automated blood cell counter. The amounts of Hb fractions were measured by capillary electrophoresis of Hb. Statistical analysis was performed using R software. RESULTS: A total of 328 cord blood samples were analyzed. Among them, 154 (male: 44.8%, female: 55.2%) were used to establish reference values. The normal reference values of complete blood count (CBC) and Hb fractions were calculated. Mean neonatal Hb was 14.75 ± 2.26 g/dL. Gestational age affects the expression of CBC values as red blood cell (RBC), Hb, hematocrit (Hct), mean corpuscular volume (MCV), white blood cell (WBC), and the Hb profile. Umbilical blood hemogram parameters and Hb profile are affected by the environment; higher in newborns from urban regions but not affected by gender ratio. CONCLUSION: Reference ranges of normal CBC indices and Hb fractions have been successfully established in Tunisian neonates' UCB. Our data suggest reference values that could be useful for neonatal patients' laboratory results and clinical interpretation. KEY POINTS: · Reference values for CBC and hemoglobin fractions have been established.. · Hematological reference for UCB is useful to identify hemolytic anemia cases early.. · UCB hematological values are influenced by gestational age and probably by environmental factors..

Enhanced Eryptosis in Glucose-6-Phosphate Dehydrogenase Deficiency.

BACKGROUND/AIMS: Defects in the Glucose-6-Phosphate Dehydrogenase (G6PD) enzyme enhance cellular oxidative damage, thus impairing erythrocytes and radically shortening their lifespan. We aimed to study programmed erythrocyte cell death in G6PD-deficient patients, describe the molecular genetics basis of G6PD and investigate phenotype-genotype correlations. METHODS: We explored eryptosis using the annexin V-binding assay, taken as an indicator of PS exposure at the erythrocyte surface. We assessed reactive oxygen species (ROS) production, intracellular calcium concentrations and ceramide formation at the cell surface. Prior to and following treatments, cells were analyzed by flow cytometry. Finally, we explored G6PD gene mutations through PCR-Sanger sequencing. RESULTS: Before stimulation, PS-exposing erythrocytes were significantly higher in G6PD-deficient patients than in healthy volunteers. This was paralleled by a significant increase in reactive oxygen species production, suggesting that oxidative stress is the main trigger of PS exposure in G6PD-deficient erythrocytes. Five previously described mutations were detected in our patients. Two genotypes correlated with a significantly higher percentage of PS-exposing cells. CONCLUSION: Our study uncovers a novel effect detected in G6PD-deficient erythrocytes which is cell membrane scrambling with PS translocation to the erythrocyte surface. Our findings shed a light on the mechanisms of premature erythrocyte clearance in G6PD deficiency.

Biochemical, Cellular, and Proteomic Characterization of Hereditary Spherocytosis Among Tunisians.

BACKGROUND/AIMS: Hereditary Spherocytosis (HS) is the most common erythrocyte membrane disorder causing hemolytic anemia. The wide heterogeneity of both clinical and laboratory manifestations of HS contributes to difficulties associated with the diagnosis of this disorder. Although massive data previously reported worldwide, there is yet no data on HS among the Tunisian population. Here we aim to characterize HS in Tunisian patients at biochemical and cellular levels, identify the membrane protein deficiency, and compare the accuracy of the diagnostic tests to identify the most appropriate assay for HS diagnosis. METHODS: We investigated 81 patients with hemolytic anemia and 167 normal controls. The exploration of HS based on clinical and family history, physical examination, and the results of laboratory tests: blood smear, osmotic fragility test (OFT), cryohemolysis test (CT), pink test (PT), eosine-5'-maleimide (EMA) test, and erythrocyte membrane protein electrophoresis. RESULTS: We identified 21 patients with HS, classified as severe (6/21;28.5%), moderate (10/21;47.6%), and mild (5/21;23.8%). The most prevalent protein deficiency was the band 3 protein detected in ten Tunisian HS patients. The EMA test showed a high specificity (97.5%) and sensitivity (94.7%) for HS diagnosis compared to the other screening tests. Interestingly, fourteen among sixteen patients presenting with homozygous sickle cells HbSS showed an increase of EMA fluorescence intensity compared to other anemic patients. CONCLUSION: Our study highlights the efficiency of the EMA dye for the detection of HS whatever the nature of the involved protein deficiency. We report for the first time, the most prevalent protein deficiency among Tunisians with HS. Moreover, we found that the combination of the EMA-binding test with PT or incubated OFT improves the diagnosis sensitivity while maintaining a good specificity.

First Observation of HbM-Saskatoon at the Origin of Neonatal Cyanosis in a Tunisian Baby.

Several causes are known to be at the origin of neonatal cyanosis among them methemoglobinemia is by inheritance of an hemoglobin (Hb) M variant. This is a rare condition never been reported in Tunisia so far. Here, we report a Tunisian newborn with refractory cyanosis since birth. As cardiac and respiratory diseases were ruled out, methemoglobinemia was suspected. Hematological parameters, concentration of methemoglobin, capillary electrophoresis, and amplification sequencing of the HBB gene were performed. Computational analysis was achieved by different in silico tools to investigate the mutation effect. The diagnosis was established by a raised MetHb, confirmed by the presence HbM-Saskatoon [Beta63 (E7) His>Tyr] by capillary electrophoresis and molecular analysis. The identified mutation occurred as a de novo mutation. In silico analysis confirmed the pathogenicity of the mutation. To our knowledge, this is the first time that this mutation has been reported in the Tunisian population. In view of its low incidence rate, clinicians might misdiagnose cyanosis caused by HbM, which can lead to inappropriate treatment and clinical complications. An up-to-date literature review of HbM disease is presented in this study.

Novel mutations in Uridyl-diphosphate-glucuronosyl-transferase 1A1 (UGT1A1) gene in Tunisian patients with unconjugated hyperbilirubinemia.

INTRODUCTION: Unconjugated hyperbilirubinemia (UCB) is a feature of Gilbert's syndrome (GS) and Crigler-Najjar's syndrome (CNS), which are two hereditary defects in bilirubin metabolism. Both syndromes are linked to mutations in the UGT1A1 gene, which cause either the decrease or the absence of the UGT1A1 enzymatic activity. Here, we investigated the molecular basis of the UGT1A1 gene in Tunisian patients presenting with unconjugated hyperbilirubinemia. METHODS: Twenty-four patients with UCB were investigated. The screening protocol for hemoglobinopathies, enzymopathies, and membrane defects was executed in all patients. Afterward, the molecular analysis of the entire UGT1A1 gene was performed by DNA Sanger sequencing. Several bioinformatic tools were used to explore the effects of novel mutations. RESULTS: Fifteen different UGT1A1 variations were identified, among which four are described here for the first time. In exon 5, the c.1412C > G; p.(Ala471Gly) and c.1589C > T; p.(Ser530Phe) mutations were detected in patients presenting with CNS type I and GS, respectively. In the 3'UTR region of UGT1A1, the c.*90C > T mutation was detected in 3 patients with CNS type I. In the same region, the c.*388C > T defect was found in a GS patient. A deleterious and damaging effect on the UGT1A1 protein were predicted for both exonic mutations. Furthermore, novel microRNAs were identified as targetting the mutated sequences for the 3'UTR mutations. CONCLUSION: Our study provides novel data on UCB among Tunisians. Furthermore, we report four novel mutations associated with both GS and CNS. The identification of these mutations increases the spectrum of the UGT1A1 mutations and contributes to an understanding of the molecular abnormalities associated with unconjugated hyperbilirubinemia.

New Deletion at Promoter of HBG1 Gene in Sickle Cell Disease Patients With High HbF Level.

OBJECTIVES: The 5' upstream region of the HBG1 gene plays a very important role in the expression of fetal hemoglobin (HbF). In contrast, increased HbF levels can inhibit the deoxygenation-induced polymerization of sickle hemoglobin (α2ßS2), which leads to moderation at the clinical level among sickle cell disease (SCD) patients. Thus, we focused on this article on the study of the 5' upstream region of HBG1 among SCD pediatric patients with high levels of HbF. MATERIALS AND METHODS: Fifteen SCD pediatric patients were chosen during the first time of diagnosis, and the HbF values were determined before hydoxyurea treatment. The ages at entry ranged from 1 to 8 years. The mutational screening of the 5' upstream region of the HBG1, which extends to -587 bp, was performed by polymerase chain reaction/sequencing. RESULTS: HbF values range from 6.9% to 26%. Sequencing results showed the presence of 6 known polymorphisms, which are as follows: RS35993903, RS34844625, RS3020750, RS2860456, RS2860470, and RS12290216. Interestingly, we also found a new deletion of GCAG in the HBG1 promoter at position -273. CONCLUSIONS: We described a new mutation, which is a deletion of GCAG in the HBG1 promoter at position -273. This deletion could affect a binding site of a transcription factor unknown so far and thus modulate the expression of the HBG1 gene.

EZH2, new diagnosis and prognosis marker in acute myeloid leukemia patients.

PURPOSE: Acute myeloid leukemia (AML) is a heterogeneous disease. The discovery of novel discriminative biomarkers remains of utmost value for improving outcome predictions. Enhancer of zeste homolog 2 (EZH2) is a histone methyltransferase of H3K27me3. It is frequently up-regulated in human cancers and associated with silencing of differentiation genes. We aimed herein to investigate the prevalence and prognosis impact of somatic EZH2 mutations and their potential associations with other prognostic markers FLT3, NPM1, DNMT3A and IDH2. MATERIALS AND METHODS: Our study population was composed of 211 Tunisian patients with de novo AML and 14 healthy donors. The 11 last exons coding the set domain of EZH2 were investigated by PCR and Sanger sequencing. RESULTS: EZH2 mutations were identified in 66/211 (31%) patients with a sex ratio of 1.06. The presence of EZH2 mutations was statistically significantly associated with failure consolidation therapy (p = 0.004). There were no differences in the incidence of EZH2 mutations and FLT3-ITD, NPM1, DNMT3A and IDH2 mutations. When EZH2 mutations were associated with those of FLT3 or IDH2, a short duration of progression free survival was observed (p < 0.05). Moreover, CD7 aberrant markers conferred a poor prognosis in EZH2 mutated patients (p < 0.05). CONCLUSIONS: Given these data we conclude that EZH2 mutations are frequent in our patients, and can be used as a prognosis marker in combination with FLT3, IDH2 mutations and CD7 marker, to stratify AML patients and to guide therapeutic decisions.

[Psychosocial and academic consequences of beta-thalassemia major in Tunisia]. / Retentissement psychosocial et scolaire de la bêta-thalassémie majeure en Tunisie.

In Tunisia, beta-thalassemia major is a real public health problem. A study carried out of patients affected shows that for them, this chronic haemoglobinopathy is a disability hampering their physical activities, their social integration, their academic results and their emotional life.

Hb Moscva [ß24(B6)Gly→Asp (GGT>GAT), HBB: c.74G>A]: An Unstable Hemoglobin Newly Detected as a De Novo Mutation in a Mauritanian Patient.

Unstable hemoglobins (Hbs) are a group of Hb disorders that could be the origin of chronic hemolytic anemia. Most of these disorders are caused by point mutations taking place in the globin genes and affecting the stability of the Hb molecule. They are inherited as autosomal dominant diseases and described worldwide. Herein we report a new observation of an unstable variant in the Mauritanian population. The patient was a young girl of Mauritanian origin. She presented with chronic hemolytic anemia with an unknown etiology after being referred to several medical centers. Laboratory investigations based on routine analyses, capillary electrophoresis (CE), cation exchange high performance liquid chromatography (HPLC) and DNA sequencing revealed an abnormal unstable Hb known as Hb Moscva [ß24(B6)Gly→Asp (GGT>GAT), HBB: c.74G>A] that occurred as a de novo mutation newly detected in an African girl of Mauritanian origin.

Molecular study of ABCB1 gene and its correlation with imatinib response in chronic myeloid leukemia.

PURPOSE: The introduction and success of imatinib mesylate have become a paradigm shift in chronic myeloid leukemia (CML) treatment. However, despite its high efficiency, resistance to imatinib has emerged as a significant problem, which may in part be caused by pharmacogenetic variability. Three single-nucleotide polymorphisms (C1236T, G2677T/A, C3435T) and/or mRNA expression changes of ABCB1 gene were demonstrated to be associated with inter-individual variability of imatinib response in CML patients. In this study, we aimed to examine whether genetic variations and/or altered expression of ABCB1 gene may influence response to imatinib. METHODS: Sixty nine CML Tunisian patients, undergoing imatinib therapy, were enrolled in this study. These were divided into two groups: responders and non-responders to imatinib. The relative transcript expression levels of ABCB1 gene and the distribution of allele and genotype frequency of ABCB1 SNPs were compared between these two categories of patients. Linkage disequilibrium tests and haplotypes analysis were also studied. RESULTS: Our results showed that the mRNA expression level of ABCB1 gene did not differ significantly between the two categories of patients. In addition, results obtained from ABCB1 polymorphisms study and their correlation with imatinib response showed that the optimal response rate to imatinib did not differ significantly between C1236T, G2677T/A or C3435T genotypes. However, haplotype analysis showed that the 1236C-2677A-3435C haplotype was observed only in imatinib non-responders' patients suggesting that CAC haplotype was linked to higher risk of imatinib resistance. CONCLUSION: Furthermore, analyses of ABCB1 haplotypes should be taken into account to study the relationship between ABCB1 genotypes and imatinib efficacy.

Homozygous Mutation on the ß-Globin Polyadenylation Signal in a Tunisian Patient with ß-Thalassemia Intermedia and Coinheritance of Gilbert's Syndrome.

We report here the clinical, hematological and molecular data in a 50-year-old patient with ß-thalassemia intermedia (ß-TI) caused by a homozygous ß+ mutation on the ß-globin gene polyadenylation (polyA) signal (AATAAA>AAAAAA). ß Haplotype analysis was accomplished by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Haplotype and framework analysis showed that this mutation is associated with the [- - - - + + +] ß haplotype and framework 1 (CCGCT) (FW1). This mutation was previously reported in the heterozygous state in association with the codon 9 (+TA) mutation in a ß-TI patient originating from Tunisia. To the best of our knowledge, this is the first report describing this mutation in the homozygous state. The case reported here, coinherited Gilbert's syndrome, which is characterized by hyperbilirubinemia. This conclusion was reached by the investigation of the promoter region [A(TA)nTAA] motif of the UGT1A1 gene, showing the (TA)6/(TA)7 genotype.

hOCT1 gene expression predict for optimal response to Imatinib in Tunisian patients with chronic myeloid leukemia.

PURPOSE: Imatinib mesylate (IM) is considered as a highly effective therapy for chronic myeloid leukemia (CML) patients. However, a minority of patients fail to achieve optimal response due to impaired bioavailability of IM. The human organic cation transporter 1 (OCT1; SLC22A1) has been reported to be the main influx transporter involved in IM uptake into CML cells. Genetic variants and/or hOCT1 expression changes may influence IM response. In this study, we aimed to investigate the impact of both hOCT1 polymorphisms located in exon 7 and hOCT1 mRNA levels on the clinical outcome in CML patients. METHODS: hOCT1 expression profile was determined using the quantitative real-time polymerase chain reaction in 69 CML patients treated with IM (35 responders to IM patients and 34 IM-resistant patients), while genotyping of 69 cases and 51 controls for hOCT1 polymorphisms was performed by direct sequencing after amplification of exon7. RESULTS: Our results showed that the hOCT1 gene was significantly downregulated in the samples of the IM-resistant group when compared with the IM-responder group (p = 0.0211). Moreover, sequencing data show an association in all cases between the SNP 408V>M (g.1222G>A) and an intronic 8 bp (base pairs) insertion of GTAAGTTG (rs36056065) at the 3' end of exon 7. The genotype and allele distribution of the different SNPs did not differ significantly between the two groups of patients. CONCLUSIONS: hOCT1 mRNA expression may serve as a clinical biomarker of response to imatinib and could be useful to predict IM therapy outcome of CML patients.

The role of rs1984112_G at CD36 gene in increasing reticulocyte level among sickle cell disease patients.

AIMS AND BACKGROUND: Mediators of adhesion become a potential new target for pharmacological therapy to struggle the complications of sickle cell disease (SCD). Several mechanisms for increased adherence have been postulated and the well-studied are CD36 and VLA4 which encoded by ITGA4. Herein, we sought to determine whether one polymorphism of CD36 namely: rs1984112 and three exons of ITGA4 (4, 5, and 6) are implicated in hemolytic status and clinical events among SCD Tunisian patients. MATERIAL AND METHODS: This study enrolled 99 unrelated Tunisian subjects (63SS and 36Sß). All SCD patients are children (less than 16 years old). The rs1984112 and the ITGA4's exons 4, 5, and 6 were analyzed for all subjects by PCR/sequencing. The association of each genotype found with both clinical complications and hemolytic status was performed using t-test. Clinical events studied included vaso-occlusive crisis (VOC), osteonecrosis, stroke, frequent infection, priapism, and acute syndrome. RESULTS: The results show that rs1984112_G allele at CD36 gene revealed to be associated with higher levels of reticulocyte count (p < 0.01). The statistical result show a near significance of homozygous mutant GG genotype with VOC (p = 0.051). No association between rs1984112_G allele and the clinical severity of SCD were found. Mutational screening of exon 4, 5, and 6 of ITGA4 gene revealed absence of mutated variant. CONCLUSION: Our results are similar to those found in Portuguese population which reported the role of rs1984112_G in increasing reticulocyte level among SCD patients. Consequently, the rs1984112_G of CD36 could be considered as a reliable biomarker for predicting patients at high risk for vascular occlusions and thus, allows earlier and more effective therapeutic management.

Downregulation of miR-451 in Tunisian chronic myeloid leukemia patients: potential implication in imatinib resistance.

OBJECTIVES: Resistance to imatinib has been recognized as a major challenge for the treatment of chronic myeloid leukemia (CML). Aberrant expression of miR-451 has been reported to participate in anticancer drug resistance. However, the role of miR-451 in imatinib resistance has not been investigated. The present study was undertaken to determine the expression of miR-451 in order to find a possible association between the expression of this miRNA and imatinib resistance in Tunisian CML patients. METHODS: First, real-time RT-PCR was performed to identify the expression of miR-451 in peripheral leukocytes of 59 CML patients treated with imatinib. Then, bioinformatics analysis was carried out to understand the regulatory roles of miR-451 in imatinib-resistant process. RESULTS: Downregulated miR-451 was observed in imatinib-resistant CML cases. In silico analysis identified MYC as a potential target of miR-451. We further revealed the existence of an MYC-binding site in MiR-451 promoter region. On the other hand, increased level of MYC was detected in imatinib-resistant CML cases which may explain the causative role of MYC in CML cases and the downregulation of miR-451. CONCLUSION AND DISCUSSION: Taken together, our findings suggest that miR-451 and MYC form together a regulatory loop which may act as a potential therapeutic target, and disruption of suggested regulatory loop could help to improve CML therapy.

Regulatory network analysis of microRNAs and genes in imatinib-resistant chronic myeloid leukemia.

Targeted therapy in the form of selective breakpoint cluster region-abelson (BCR/ABL) tyrosine kinase inhibitor (imatinib mesylate) has successfully been introduced in the treatment of the chronic myeloid leukemia (CML). However, acquired resistance against imatinib mesylate (IM) has been reported in nearly half of patients and has been recognized as major issue in clinical practice. Multiple resistance genes and microRNAs (miRNAs) are thought to be involved in the IM resistance process. These resistance genes and miRNAs tend to interact with each other through a regulatory network. Therefore, it is crucial to study the impact of these interactions in the IM resistance process. The present study focused on miRNA and gene network analysis in order to elucidate the role of interacting elements and to understand their functional contribution in therapeutic failure. Unlike previous studies which were centered only on genes or miRNAs, the prime focus of the present study was on relationships. To this end, three regulatory networks including differentially expressed, related, and global networks were constructed and analyzed in search of similarities and differences. Regulatory associations between miRNAs and their target genes, transcription factors and miRNAs, as well as miRNAs and their host genes were also macroscopically investigated. Certain key pathways in the three networks, especially in the differentially expressed network, were featured. The differentially expressed network emerged as a fault map of IM-resistant CML. Theoretically, the IM resistance process could be prevented by correcting the included errors. The present network-based approach to study resistance miRNAs and genes might help in understanding the molecular mechanisms of IM resistance in CML as well as in the improvement of CML therapy.

TET2-mediated 5-hydroxymethylcytosine induces genetic instability and mutagenesis.

The family of Ten-Eleven Translocation (TET) proteins is implicated in the process of active DNA demethylation and thus in epigenetic regulation. TET 1, 2 and 3 proteins are oxygenases that can hydroxylate 5-methylcytosine (5-mC) into 5-hydroxymethylcytosine (5-hmC) and further oxidize 5-hmC into 5-formylcytosine (5-fC) and 5-carboxylcytosine (5-caC). The base excision repair (BER) pathway removes the resulting 5-fC and 5-caC bases paired with a guanine and replaces them with regular cytosine. The question arises whether active modification of 5-mC residues and their subsequent elimination could affect the genomic DNA stability. Here, we generated two inducible cell lines (Ba/F3-EPOR, and UT7) overexpressing wild-type or catalytically inactive human TET2 proteins. Wild-type TET2 induction resulted in an increased level of 5-hmC and a cell cycle defect in S phase associated with higher level of phosphorylated P53, chromosomal and centrosomal abnormalities. Furthermore, in a thymine-DNA glycosylase (Tdg) deficient context, the TET2-mediated increase of 5-hmC induces mutagenesis characterized by GC>AT transitions in CpG context suggesting a mutagenic potential of 5-hmC metabolites. Altogether, these data suggest that TET2 activity and the levels of 5-hmC and its derivatives should be tightly controlled to avoid genetic and chromosomal instabilities. Moreover, TET2-mediated active demethylation might be a very dangerous process if used to entirely demethylate the genome and might rather be used only at specific loci.

Ritonavir-Induced Suicidal Death of Human Erythrocytes.

The antiviral drug ritonavir has been shown to trigger suicidal death or apoptosis of tumour cells and has thus been considered for the treatment of malignancy. In analogy to apoptosis of nucleated cells, erythrocytes may enter eryptosis, the suicidal erythrocyte death characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Triggers of eryptosis include Ca(2+) entry with increase in cytosolic Ca(2+) activity ([Ca(2+) ]i ), oxidative stress and ceramide. The present study explored whether and how ritonavir induces eryptosis. To this end, flow cytometry was employed to estimate cell volume from forward scatter, phosphatidylserine exposure at the cell surface from Annexin-V binding, [Ca(2+) ]i from Fluo3 fluorescence, abundance of reactive oxygen species (ROS) from 2',7'-dichlorodihydrofluorescein diacetate (DCFDA) fluorescence and ceramide abundance utilizing specific antibodies. As a result, a 48-hr exposure of human erythrocytes to ritonavir significantly increased the percentage of Annexin-V-binding cells (≥5 µg/ml), significantly decreased forward scatter (≥5 µg/ml), significantly increased Fluo3 fluorescence (20 µg/ml), slightly, but significantly increased DCFDA fluorescence (20 µg/ml) and slightly, but significantly increased ceramide abundance (20 µg/ml). The effect of ritonavir on Annexin-V binding was significantly blunted, but not fully abolished by the removal of extracellular Ca(2+) . In conclusion, ritonavir triggers erythrocyte shrinkage and phosphatidylserine translocation at the erythrocyte cell membrane, an effect in part due to the stimulation of Ca(2+) entry, oxidative stress and ceramide.

Association of genetic variation in IKZF1, ARID5B, CDKN2A, and CEBPE with the risk of acute lymphoblastic leukemia in Tunisian children and their contribution to racial differences in leukemia incidence.

Recent genome-wide association studies (GWAS) focusing on pediatric acute lymphoblastic leukemia (ALL), the most common malignancy in children younger than 15 years old, have found evidence that single-nucleotide polymorphisms (SNPs) in IKZF1 (7p12.2), ARID5B (10q21.2), CDKN2A (9p21.3), and CEBPE (14q11.2) are strongly associated to the risk of developing pediatric ALL. These studies have been conducted in European and Thai populations, and it is unclear whether these observations generalize to other populations with a lower incidence of pediatric ALL. In order to explore the impact of these variants on pediatric ALL risk in the Tunisian population, we genotyped 58 cases of pediatric ALL and 150 controls for SNPs rs4132601 (7p12.2), rs7089424 (10q21.2), rs3731217 (9p21.3), and rs2239633 (14q11.2). Our results, which are consistent with findings in European populations, show that 3 SNPs, i.e., rs4132601 (P = .00116, odds ratio [OR] = 2.78, 95% confidence interval [CI] = [1.42, 5.87]), rs7089424 (P = .0022, OR = 0.49, 95% CI = [0.31, 0.79]), and rs2239633 (P = .0010, OR = 0.47, 95% CI = [0.29, 0.75]) are significantly associated with a higher risk of developing pediatric ALL (P < .05). Furthermore, we show differences in allele frequencies in SNPs between Tunisian and Caucasian and/or Thai populations (e.g., CEBPE, rs2239633; population attributable risk [PAR] ∼15-fold the PAR of Thai population). These differences, combined with differences in linkage disequilibrium structure between populations and differences in size between populations, may contribute to racial differences in pediatric ALL incidence.

rs11886868 and rs4671393 of BCL11A associated with HbF level variation and modulate clinical events among sickle cell anemia patients.

AIMS: Fetal hemoglobin (HbF) modulates the phenotype of sickle cell anemia (SCA) by inhibiting deoxy sickle hemoglobin (HbS) polymerization. HbF genes are genetically regulated, and the level of HbF and its distribution among sickle erythrocytes is highly variable. Herein, we aimed to determine whether two functional polymorphisms of BCL11A are implicated in the variation of HbF and clinical events in SCA Tunisian patients. MATERIAL AND METHODS: The studied population consisted of 148 SCA patients with SS phenotype. The group of patients was divided into two subgroups according to the threshold point of %HbF which is 15%. Genotyping of rs11886868 and rs4671393 was performed using PCR/Sequencing. To test for trait association with the candidate SNPs, genotype and allele frequencies between 'group who had %HbF < 15' and 'group who had %HbF >15' (controls) were compared using Pearson's chi-square test (compare 2, version 1.02). The association of each genotype and the combined genotype with complications was performed by logistic regression test. RESULTS: Our findings showed that the majority of patients carried genotype CT of rs11886868 and genotypes AG and GG of rs4671393 present HbF level < 15%. RR = 0.08, RR = 0.176, and RR = 0.189, respectively. The results showed a significant association between the alleles T of rs11886868 and G of rs4671393 and %HbF < 15% with P = 0.016; RR = 0.39 and P = 8.9 × 10(-3): RR = 0.567, respectively. Interestingly, the C allele of the rs11886868 and the A allele of the rs46713939 were associated with an ameliorated phenotype in patient's SCA. The combination of the genotypes GG and CT explains more phenotypic variance than the sum of the two BCL11A SNPs taken individually.

P53 activation inhibits all types of hematopoietic progenitors and all stages of megakaryopoiesis.

TP53 also known as p53 is a tumor suppressor gene mutated in a variety of cancers. P53 is involved in cell cycle, apoptosis and DNA repair mechanisms and is thus tightly controlled by many regulators. Recently, strategies to treat cancer have focused on the development of MDM2 antagonists to induce p53 stabilization and restore cell death in p53 non-mutated cancers. However, some of these molecules display adverse effects in patients including induction of thrombocytopenia. In the present study, we have explored the effect of SAR405838 not only on human megakaryopoiesis but also more generally on hematopoiesis. We compared its effect to MI-219 and Nutlin, which are less potent MDM2 antagonists than SAR405838. We found that all these compounds induce a deleterious effect on all types of hematopoietic progenitors, as well as on erythroid and megakaryocytic differentiation. Moreover, they inhibit both early and late stages of megakaryopoiesis including ploidization and proplatelet formation. In conclusion, MDM2 antagonists induced a major hematopoietic defect in vitro as well as an inhibition of all stages of megakaryopoiesis that may account for in vivo thrombocytopenia observed in treated patients.