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Sickle cell disease (SCD) is a group of inherited blood disorders characterized by abnormal hemoglobin production, affecting individuals worldwide with varying prevalence across different populations. Manifestations vary, ranging from severe to mild. SCD is characterized by the presence of hemoglobin S (HbS), which distorts erythrocytes upon deoxygenation, leading to sickling. This results in hemolytic anemia, painful vaso-occlusive crises (VOC), and multiple organ damage, including bones, due to microinfarcts. Sickle cell trait (SCT), or carrier status, is not considered an SCD and often runs a benign course. We report a 44-year-old man of African descent presenting with a one-month history of pain in his ankles and feet. He had a prior diagnosis of sickle cell "trait" without previous VOC. Hematological indices were normal. Hemoglobin electrophoresis showed absent HbA, elevated HbS, elevated HbF, and normal HbA2. X-rays and MRI revealed bilateral bone infarction in diaphyses of right proximal and bilateral distal tibias. Molecular analysis of [Formula: see text]-globin revealed compound heterozygous hemoglobin S and type 2 deletion of persistence of fetal hemoglobin (HPFH). Pulmonary function tests revealed restrictive lung disease. A literature review from 1946 to May 2024 via PubMed, EMBASE, and Medline was performed, revealing two cases of HbS-HPFH with avascular necrosis affecting the femoral neck were briefly reported more than 60 years ago. Although pulmonary function tests in SCD typically show a mild restrictive pattern with decreased diffusion capacity and rarely an obstructive pattern, no cases of HbS-HPFH were identified. In conclusion, multiple bone infarctions are extremely rare in HbS-HPFH. Lung and bone diseases might be unrecognized in this unique disorder.

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δß-thalassemia is a rare type of thalassemia characterized by increased Hb F levels, including mainly Chinese Gγ(Aγδß)0-thalassemia, Yunnanese Gγ(Aγδß)0-thalassemia, Cantonese Gγ(Aγδß)0-thalassemia in China. Due to the low rate of δß-thalassemia carriers, there are few reports of δß-thalassemia combined with ß-thalassemia causing ß-thalassemia major. Herein, we described the combination of Chinese Gγ(Aγδß)0-thalassemia and ß-thalassemia leading to ß-thalassemia major in a Chinese patient. Hemoglobin analysis was performed by capillary electrophoresis (CE). Routine genetic analysis was carried out by gap-polymerase chain reaction (Gap-PCR) and PCR and reverse dot blot (PCR-RDB). Multiple ligation-dependent probe amplification (MLPA) was used to detect the large deletion, and Gap-PCR confirmed the deletion. A CE result showed an elevated Hb F level of 98.7% and 11.7% in the proband and her mother, but the proband was diagnosed with ßCD17M/ßCD17M using routine genetic analysis. However, her father was heterozygous for CD17 in ß-globin, and her mother was detected as SEA heterozygous. The further analysis presented that the proband had actually missed the diagnosis of Chinese Gγ(Aγδß)0-thalassemia by MLPA and PCR-RDB. Finally, the genotype of the proband was corrected from ßCD17M/ßCD17M to ßCD17M/ßGγ(Aγδß)0. This is the first report of Chinese Gγ(Aγδß)0-thalassemia combined with ß-thalassemia resulting in ß-thalassemia major in China. Screening for δß-thalassemia by Hb analysis could be an effective method.

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Beta-hemoglobinopathies are the most common genetic disorders worldwide, caused by a wide spectrum of mutations in the ß-globin locus, and associated with morbidity and early mortality in case of patient non-adherence to supportive treatment. Allogeneic transplantation of hematopoietic stem cells (allo-HSCT) used to be the only curative option, although the indispensable need for an HLA-matched donor markedly restricted its universal application. The evolution of gene therapy approaches made possible the ex vivo delivery of a therapeutic ß- or γ- globin gene into patient-derived hematopoietic stem cells followed by the transplantation of corrected cells into myeloablated patients, having led to high rates of transfusion independence (thalassemia) or complete resolution of painful crises (sickle cell disease-SCD). Hereditary persistence of fetal hemoglobin (HPFH), a syndrome characterized by increased γ-globin levels, when co-inherited with ß-thalassemia or SCD, converts hemoglobinopathies to a benign condition with mild clinical phenotype. The rapid development of precise genome editing tools (ZFN, TALENs, CRISPR/Cas9) over the last decade has allowed the targeted introduction of mutations, resulting in disease-modifying outcomes. In this context, genome editing tools have successfully been used for the introduction of HPFH-like mutations both in HBG1/HBG2 promoters or/and in the erythroid enhancer of BCL11A to increase HbF expression as an alternative curative approach for ß-hemoglobinopathies. The current investigation of new HbF modulators, such as ZBTB7A, KLF-1, SOX6, and ZNF410, further expands the range of possible genome editing targets. Importantly, genome editing approaches have recently reached clinical translation in trials investigating HbF reactivation in both SCD and thalassemic patients. Showing promising outcomes, these approaches are yet to be confirmed in long-term follow-up studies.

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BACKGROUND: Deletions in the ß-globin cluster are uncommon and cause thalassemia (thal) with hereditary persistence of fetal hemoglobin. They constitute a heterogenous group of disorders characterized by absent or reduced synthesis of adult hemoglobin (Hb A) and increased synthesis of fetal hemoglobin (Hb F). Although the clinical severity of these disorders are asymptomatic owing to the increased Hb F levels, the molecular basis is very heterogenous due to the large deletions in the ß-globin cluster spanning both HBD and HBB genes. Here, we describe a Tunisian family carrying a novel deletion mutation causing (δß)°-thalassemia. METHODS: The amounts of hemoglobin fractions were measured by capillary electrophoresis of hemoglobin. Amplification and sequencing of different regions on the ß-gene cluster were performed by Sanger method. RESULTS: Family study and genetic analysis revealed a large deletion mutation in the ß-globin cluster of 14.5 kb (NG_000,007.3:g. 58,253 to g.72837del14584) at the homozygous state in the patient and at heterozygous state at the other members of the family. This deletion removes the HBD and HBB genes. CONCLUSIONS: In our knowledge, this new large deletion is described for the first time in the Tunisian population and in the world, designed Tunisian(δß)0 in Ithanet database (IthaID: 3971). Therefore, it is important to identify the deletion leading to δß-thalassemia carriers at the molecular level, to highlight the importance of recognizing the clinical features and implementing appropriate testing to clarify the diagnosis and manage the condition.

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Reactivation of γ-globin expression is a promising therapeutic approach for ß-hemoglobinopathies. Here, we propose a novel Cas9/AAV6-mediated genome editing strategy for the treatment of ß-thalassemia: Natural HPFH mutations -113A > G, -114C > T, -117G>A, -175T > C, -195C > G, and -198T > C were introduced by homologous recombination following disruption of BCL11A binding sites in HBG1/HBG2 promoters. Precise on-target editing and significantly increased γ-globin expression during erythroid differentiation were observed in both HUDEP-2 cells and primary HSPCs from ß-thalassemia major patients. Moreover, edited HSPCs maintained the capacity for long-term hematopoietic reconstitution in B-NDG hTHPO mice. This study provides evidence of the effectiveness of introducing naturally occurring HPFH mutations as a genetic therapy for ß-thalassemia.

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In the already identified quantitative trait loci (QTL), modulating Hb F levels are cis-acting haplotypes of the ß-globin gene cluster itself, although the single nucleotide polymorphisms (SNPs) accounting more for the association, remain uncertain. In this study, the role in Hb F production of previously reported candidate SNPs within the ß-globin gene cluster was reexamined, along with a yet poorly studied variation in the BGLT3 gene. In a sample of ß-thalassemia (ß-thal) carriers, we succeeded in replicating the significant association between increased Hb F levels and rs7482144 (C>T) (HBG2 XmnI), which is the most well-established variation in the cluster influencing the trait. This SNP was found to be in strong linkage disequilibrium (LD) with a variation in the HBBP1 gene [rs10128556 (G>A)], which consistently revealed a similar association signal. Remarkably, much stronger than the latter associations were those involving both rs968857 (T allele) (3' HBBP1) and rs7924684 (G allele) (BGLT3), two SNPs that were also in strong LD. As the pattern of LD detected in the ß-globin gene cluster does not correlate with a tight linkage between markers, complex interactions between SNPs at the cluster seem to modulate Hb F. Seeing that no such associations were detected in normal subjects, the question can be raised on whether, under erythropoiesis stress, epigenetic mechanisms contribute to change the regulation of the entire ß-globin gene cluster. In conclusion, we provide statistical evidence for a new player within the ß-globin gene cluster, BGLT3, that in cooperation with other regions influences Hb F levels in ß-thal carriers.

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INTRODUCTION: Sickle cell disease and ß thalassemia are the principal ß hemoglobinopathies. The complex pathophysiology of sickle cell disease is initiated by sickle hemoglobin polymerization. In ß thalassemia, insufficient ß-globin synthesis results in excessive free α globin, ineffective erythropoiesis, and severe anemia. Fetal hemoglobin (HbF) prevents sickle hemoglobin polymerization; in ß thalassemia HbF compensates for the deficit of normal hemoglobin. When HbF constitutes about a third of total cell hemoglobin, the complications of sickle cell disease are nearly totally prevented. Similarly, sufficient HbF in ß thalassemia diminishes or prevents ineffective erythropoiesis and hemolysis. AREAS COVERED: This article examines the pathophysiology of ß hemoglobinopathies, the physiology of HbF, intracellular distribution, and the regulation of HbF expression. Inducing high levels of HbF by targeting its regulatory pathways pharmacologically or with cell-based therapeutics provides major clinical benefit and perhaps a 'cure.' EXPERT OPINION: Erythrocytes must contain about 10 pg of HbF to 'cure' sickle cell disease. If HbF is the only hemoglobin present, much higher levels are needed to 'cure' ß thalassemia. These levels of HbF can be obtained by different iterations of gene therapy. Small molecule drugs that can achieve even modest pancellular HbF concentrations are a major unmet need.

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Red blood cell exchange transfusion may be beneficial and should be considered in the early management of patients with sickle cell disease and COVID-19 to prevent the need for intubation and intensive care unit admission due to respiratory distress.

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The expression of the fetal Gγ- and Aγ-globin genes in normal development is confined to the fetal period, where two γ-globin chains assemble with two α-globin chains to form α2γ2 tetramers (HbF). HbF sustains oxygen delivery to tissues until birth, when ß-globin replaces γ-globin, leading to the formation of α2ß2 tetramers (HbA). However, in different benign and pathological conditions, HbF is expressed in adult cells, as it happens in the hereditary persistence of fetal hemoglobin, in anemias and in some leukemias. The molecular basis of γ-globin differential expression in the fetus and of its inappropriate activation in adult cells is largely unknown, although in recent years, a few transcription factors involved in this process have been identified. The recent discovery that fetal cells can persist to adulthood and contribute to disease raises the possibility that postnatal γ-globin expression could, in some cases, represent the signature of the fetal cellular origin.

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Acute splenic sequestration crisis (ASSC) is a life-threatening complication of sickle cell disease characterized by acute splenic enlargement and a rapid drop in hemoglobin (Hb) levels. Although ASSC is predominantly a disease of childhood, rare cases of ASSC have been reported in adults, frequently in association with heterozygous sickle cell disease. We describe a case of ASSC in a 29-year-old woman with homozygous Hb S (HBB: c.20A>T) disease and suspected hereditary persistence of fetal Hb (HPFH), which was accompanied by marked leukocytosis.

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Fetal hemoglobin (HbF) usually consists of 4 to 10% of total hemoglobin in adults of African descent with sickle cell anemia. Rarely, their HbF levels reach more than 30%. High HbF levels are sometimes a result of ß-globin gene deletions or point mutations in the promoters of the HbF genes. Collectively, the phenotype caused by these mutations is called hereditary persistence of fetal hemoglobin, or HPFH. The pancellularity of HbF associated with these mutations inhibits sickle hemoglobin polymerization in most sickle erythrocytes so that these patients usually have inconsequential hemolysis and few, if any, vasoocclusive complications. Unusually high HbF can also be associated with variants of the major repressors of the HbF genes, BCL11A and MYB. Perhaps most often, we lack an explanation for very high HbF levels in sickle cell anemia.

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The expression of the human ß-like globin genes follows a well-orchestrated developmental pattern, undergoing two essential switches, the first one during the first weeks of gestation (ε to γ), and the second one during the perinatal period (γ to ß). The γ- to ß-globin gene switching mechanism includes suppression of fetal (γ-globin, HbF) and activation of adult (ß-globin, HbA) globin gene transcription. In hereditary persistence of fetal hemoglobin (HPFH), the γ-globin suppression mechanism is impaired leaving these individuals with unusual elevated levels of fetal hemoglobin (HbF) in adulthood. Recently, the transcription factors KLF1 and BCL11A have been established as master regulators of the γ- to ß-globin switch. Previously, a genomic variant in the KLF1 gene, identified by linkage analysis performed on twenty-seven members of a Maltese family, was found to be associated with HPFH. However, variation in the levels of HbF among family members, and those from other reported families carrying genetic variants in KLF1, suggests additional contributors to globin switching. ASF1B was downregulated in the family members with HPFH. Here, we investigate the role of ASF1B in γ- to ß-globin switching and erythropoiesis in vivo. Mouse-human interspecies ASF1B protein identity is 91.6%. By means of knockdown functional assays in human primary erythroid cultures and analysis of the erythroid lineage in Asf1b knockout mice, we provide evidence that ASF1B is a novel contributor to steady-state erythroid differentiation, and while its loss affects the balance of globin expression, it has no major role in hemoglobin switching.

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Hereditary persistence of fetal hemoglobin (HPFH) is a benign condition in which significant fetal hemoglobin production continues well into adulthood, disregarding the normal shutoff point after which only adult-type hemoglobin should be produced. The percentage of incorrect expression might be as low as 10%-15% or as high as 100% of the total hemoglobin, usually higher in homozygotes than in heterozygotes. The present case is a typical example of homozygous HPFH.

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Hemoglobin is a tetrameric protein composed of two α and two ß chains, each containing a heme group that reversibly binds oxygen. The composition of hemoglobin changes during development in order to fulfill the need of the growing organism, stably maintaining a balanced production of α-like and ß-like chains in a 1:1 ratio. Adult hemoglobin (HbA) is composed of two α and two ß subunits (α2ß2 tetramer), whereas fetal hemoglobin (HbF) is composed of two γ and two α subunits (α2γ2 tetramer). Qualitative or quantitative defects in ß-globin production cause two of the most common monogenic-inherited disorders: ß-thalassemia and sickle cell disease. The high frequency of these diseases and the relative accessibility of hematopoietic stem cells make them an ideal candidate for therapeutic interventions based on genome editing. These strategies move in two directions: the correction of the disease-causing mutation and the reactivation of the expression of HbF in adult cells, in the attempt to recreate the effect of hereditary persistence of fetal hemoglobin (HPFH) natural mutations, which mitigate the severity of ß-hemoglobinopathies. Both lines of research rely on the knowledge gained so far on the regulatory mechanisms controlling the differential expression of globin genes during development.

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BACKGROUND: Deletional hereditary persistence of fetal hemoglobin (HPFH)/δß-thalassemia and δ-thalassemia are rare inherited disorders which may complicate the diagnosis of ß-thalassemia. The aim of this study was to reveal the frequency of these two disorders in Southwestern China. METHODS: A total of 33,596 subjects were enrolled for deletional HPFH/δß-thalassemia, and positive individuals with high fetal hemoglobin (Hb F) level were diagnosed by multiplex ligation-dependent probe amplification (MLPA). A total of 17,834 subjects were analyzed for mutations in the δ-globin gene. Positive samples with low Hb A2 levels were confirmed by δ-globin gene sequencing. Furthermore, the pathogenicity and construction of a selected δ-globin mutation were analyzed. RESULTS: A total of 92 suspected cases with Hb F ≥5.0% were further characterized by MLPA. Eight different deletional HPFH/δß-thalassemia were observed at a frequency of 0.024%. In addition, 195 cases suspected to have a δ-globin gene mutation (Hb A2 ≤2.0%) were characterized by molecular analysis. δ-Globin gene mutation was found at a frequency of 0.49% in Yunnan. The pathogenicity and construction for a selected δ-globin mutation was predicted. CONCLUSION: Screening of these two disorders was analyzed in Southwestern China, which could define the molecular basis of these conditions in this population.

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Disorders in hemoglobin (hemoglobinopathies) were the first monogenic diseases to be characterized and remain among the most common and best understood genetic conditions. Moreover, the study of the ß-globin locus provides a textbook example of developmental gene regulation. The fetal γ-globin genes (HBG1/HBG2) are ordinarily silenced around birth, whereupon their expression is replaced by the adult ß-globin genes (HBB primarily and HBD). Over 50 years ago it was recognized that mutations that cause lifelong persistence of fetal γ-globin expression ameliorate the debilitating effects of mutations in ß-globin. Since then, research has focused on therapeutically reactivating the fetal γ-globin genes. Here, we summarize recent discoveries, focusing on the influence of genome editing technologies, including CRISPR-Cas9, and emerging gene therapy approaches.

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OBJECTIVE: To reveal the prevalence and molecular characterization of (δß)0 -thalassemia [(δß)0 -thal] and hereditary persistence of fetal hemoglobin (HPFH) in the Chinese Zhuang population. METHODS: A total of 105 subjects with fetal hemoglobin (Hb F) level ≥5% from 14 204 unrelated ones were selected for the study. Multiplex ligation dependent probe amplification was firstly used to analyze dosage changes of the ß-globin gene cluster for associated with (δß)0 -thal and HPFH mutations. The gap polymerase chain reaction was then performed to identify the deletions using the respective flanking primers. Hematologic data were recorded and correlated with the molecular findings. RESULTS: Twenty-one (0.15%) subjects were diagnosed with Chinese G γ(A γδß)0 -thal. Nine (0.06%) were diagnosed with Southeast Asia HPFH (SEA-HPFH) deletion. Seventy-five (0.53%) cases remained uncharacterized. Three genotypes for Chinese G γ(A γδß)0 -thal and SEA-HPFH deletion were identified, respectively. The genotype-phenotype relationships were discussed. CONCLUSION: Our study for the first time demonstrated that (δß)0 and HPFH were not rare events, and molecular characterized G γ(A γδß)0 -thal and HFPH mutations in the Chinese Zhuang population. The findings in our study will be useful in genetic counseling and prenatal diagnostic service of ß-thalassemia in this populations.

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The gene for hereditary persistence of fetal hemoglobin (HPFH) in the Caribbean is much more common than previously estimated. To avoid labeling persons with the benign syndrome Hb S (HBB: c.20A>T)/HPFH as a disease and wasting scarce resources, parental studies are recommended when newborn screening reveals a pattern consistent with an SS phenotype.

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The implications of lncRNAs regarding fetal hemoglobin (HbF) induction in hemoglobin disorders remain poorly understood. In this study, microarray analysis was performed to profile lncRNAs, miRNAs and mRNAs in individuals with hereditary persistence of fetal hemoglobin (HPFH), ß-thalassemia carriers with high HbF levels and healthy controls. The results show aberrant expression of 862 lncRNAs, 568 mRNAs and 63 miRNAs in the high-HbF group compared with the control group. Altered NR_001589, NR_120526, T315543, miR-486-3p, miR-19b-1-5p and miR-20a-3p expression was confirmed by quantitative reverse transcription-polymerase chain reaction, and Spearman correlation coefficients revealed significant positive correlations with HbF. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses showed the hematopoietic cell lineage and apoptosis to be most significantly dysregulated in HbF induction. We analyzed coding genes near the lncRNAs and constructed a coding-noncoding co-expression network. Based on the results, lncRNAs likely contribute to increased HbF levels by activating expression of HBE1 and hematopoietic cell lineage-inducible molecules and by inhibiting that of apoptosis-inducible molecules. Finally, through construction of a competing endogenous RNA network, we found that 6 lncRNAs could bind competitively with miR-486-3p, resulting in increased HbF levels. Taken together, our findings provide new insights into the mechanisms of HbF induction and potentially provide new targets for the treatment of ß-thalassemia major.

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Hereditary persistence of fetal hemoglobin deletion type-2 (HPFH-2) and Sicilian-δß-thalassemia are conditions described as large deletions of the human ß-like globin cluster, with absent ß-globin chains and a compensatory variable increase in γ-globin. HPFH, in general, may be distinguished from DB-Thalassemia by higher fetal hemoglobin (HbF) levels, absence of anemia and hypochromic and microcytic erythrocytes. MicroRNAs (miRNAs) regulate a range of cellular processes including erythropoiesis and regulation of transcription factors such as the BCL11A and SOX6 genes, which are related to the regulation of γ-globin expression. In this report, a possible association among the overexpression of miRNAs and the expression of the γ-globin gene was analyzed in these two conditions. Forty-nine differentially expressed miRNAs were identified by microarrays in CD34+-derived erythroid cells of two subjects heterozygous for Sicilian-δß-thalassemia, 2 for HPFH-2 and 3 for controls after 13 days of culture. Some of these miRNAs may participate in γ-globin gene regulation and red blood cell function. The BCL11A gene was found to be potentially targeted by 12 miRNAs that were up-regulated in HPFH-2 or in DB-Thal. A down-regulation of BCL11A gene expression in HPFH-2 was verified by quantitative polymerase chain reaction. These data suggest an important action for miRNA that may partially explain the phenotypic differences between HPFH-2 and Sicilian δß-thalassemia and the increased expression of γ-globin in these conditions.

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