scholarly journals Analysis of Methylation Status in Promoter Region of Γ- Globin Gene in Carrier and Affected Β-Thalassemia Patients with High Level of Fetal Hemoglobin in Comparison with Normal Individuals

2021 ◽  
Author(s):  
Mohammad Hamid ◽  
Zohreh Shojaei
Keyword(s):  
Promoter Region ◽  
Globin Gene ◽  
Methylation Status ◽  
Fetal Hemoglobin ◽  
Methylation Pattern ◽  
Beta Thalassemia ◽  
Sequencing Analysis ◽  
Gamma Globin ◽  
Cpg Sites ◽  
Normal Individuals

Introduction: Among the factors that may be associated with the re-expression gamma-globin in adults is the methylation pattern of the promoter region. The study aimed to determine the association between promoter methylation pattern of the gamma-globin gene in the carriers and affected beta-thalassemia individuals and its expression levels. Methods: This study has been done as a case control- study. After taking blood samples from 30 patients and beta-thalassemia carriers and affected patients with fetal hemoglobin elevated as well as 30 normal individuals, genomic DNA was extracted. Six CpG sites of the promoter region and exon1 of the gamma-globin gene were analyzed by the bisulfite sequencing analysis method. Statistical analysis was carried out using a t-test. The values of p ≤ 0.05 were considered significant for comparing two studied groups. Data were analyzed using SPSS version 16 software. Results: In this study, hypomethylation of the gamma-globin promoter region in the patients and carriers compared to showed a significant differences in three CpG sites +6, -53 and -162, respectively (p<0.05). In addition, in three cases, CpG site in position -162 was semi methylated; this situation was markedly different from other samples of the patients and normal cases. Conclusion: Hypomethylation of the gamma-globin gene promoter probably has an auxiliary role in fetal hemoglobin increase.

scholarly journals Molecular analysis of Japanese delta beta-thalassemia

Blood ◽  
1988 ◽  
Vol 72 (5) ◽  
pp. 1771-1776
Author(s):  
S Shiokawa ◽  
H Yamada ◽  
Y Takihara ◽  
E Matsunaga ◽  
Y Ohba ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Beta Thalassemia ◽  
Globin Genes ◽  
Beta Globin ◽  
Beta Globin Gene ◽  
Gamma Globin ◽  
Large Deletions ◽  
Deletion Junction

A DNA fragment containing the deletion junction region from a Japanese individual with homozygous delta beta-thalassemia has been cloned. A clone containing the normal DNA surrounding the 3′ breakpoint of this deletion and a clone carrying the G gamma- and A gamma-globin genes of this patient were also isolated. Sequences of the deletion junction and both gamma-globin genes were determined. A comparison of these sequences with previously determined sequences of the normal counterparts revealed that the 5′ breakpoint is located between 2,134 and 2,137 base pairs (bp) 3′ to the polyA site of the A gamma-globin gene, the 5′ breakpoint is located just downstream of the 3′ border of the fetal gamma-globin duplication unit, and no molecular defects are evident within the gamma-globin gene region. A comparison between the sequences of the normal DNA surrounding the 3′ breakpoint and the normal DNA surrounding the 5′ breakpoint shows that deletion is the result of a nonhomologous recombination event. There are A+T-rich stretches near the 5′ and 3′ breakpoints in the normal DNA, and a portion of an Aly repeat is located in the region 3′ to the 3′ breakpoint. Southern blot analysis using probes 3′ to the beta-globin gene showed that the deletion extends in the 3′ direction further than any other deletions associated with delta beta-thalassemia and hereditary persistence of fetal hemoglobin (HPFH) heretofore reported. These results are discussed in terms of the mechanism generating large deletions in mammalian cells and three models for the regulation of gamma-globin and beta-globin gene expression in humans.

scholarly journals Molecular analysis of Japanese delta beta-thalassemia

Blood ◽  
1988 ◽  
Vol 72 (5) ◽  
pp. 1771-1776 ◽  
Author(s):  
S Shiokawa ◽  
H Yamada ◽  
Y Takihara ◽  
E Matsunaga ◽  
Y Ohba ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Beta Thalassemia ◽  
Globin Genes ◽  
Beta Globin ◽  
Beta Globin Gene ◽  
Gamma Globin ◽  
Large Deletions ◽  
Deletion Junction

Abstract A DNA fragment containing the deletion junction region from a Japanese individual with homozygous delta beta-thalassemia has been cloned. A clone containing the normal DNA surrounding the 3′ breakpoint of this deletion and a clone carrying the G gamma- and A gamma-globin genes of this patient were also isolated. Sequences of the deletion junction and both gamma-globin genes were determined. A comparison of these sequences with previously determined sequences of the normal counterparts revealed that the 5′ breakpoint is located between 2,134 and 2,137 base pairs (bp) 3′ to the polyA site of the A gamma-globin gene, the 5′ breakpoint is located just downstream of the 3′ border of the fetal gamma-globin duplication unit, and no molecular defects are evident within the gamma-globin gene region. A comparison between the sequences of the normal DNA surrounding the 3′ breakpoint and the normal DNA surrounding the 5′ breakpoint shows that deletion is the result of a nonhomologous recombination event. There are A+T-rich stretches near the 5′ and 3′ breakpoints in the normal DNA, and a portion of an Aly repeat is located in the region 3′ to the 3′ breakpoint. Southern blot analysis using probes 3′ to the beta-globin gene showed that the deletion extends in the 3′ direction further than any other deletions associated with delta beta-thalassemia and hereditary persistence of fetal hemoglobin (HPFH) heretofore reported. These results are discussed in terms of the mechanism generating large deletions in mammalian cells and three models for the regulation of gamma-globin and beta-globin gene expression in humans.

The -195C→G Substitution In Brazilian Hereditary Persistence of Fetal Hemoglobin Decreases NF-E1/YY1 Binding and Increases PAX1 Binding to the A Gamma Globin Promoter Region

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2066-2066 ◽  
Author(s):  
Fernanda Marconi Roversi ◽  
Anderson Ferreira Cunha ◽  
Carolina Lanaro ◽  
Ana Flavia Brugnerotto ◽  
Maria Emília Favero ◽  
...  
Keyword(s):  
Dna Binding ◽  
Promoter Region ◽  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Globin Genes ◽  
Dna Array ◽  
Gamma Globin ◽  
Nuclear Extracts ◽  
Hereditary Persistence ◽  
Globin Promoter

Abstract Abstract 2066 Hereditary persistence of fetal hemoglobin (HPFH) is a condition that prevents hemoglobin switching and the consequent silencing of the gamma globin genes, resulting in continued hemoglobin (Hb) F synthesis in adults. Two types of HPFH are responsible for this phenotype: deletional HPFH – deletions in the end of the beta globin locus – and non-deletional HPFH (ndHPFH) – single point mutations in the proximal promoter of both gamma globin genes. Sickle cell anemia patients or beta-thalassemia patients that present HPFH show high levels of HbF that are associated with less severe clinical course in these diseases. The development of new therapies based on the reactivation of gamma globin expression may be important for the treatment of these patients. The Brazilian ndHPFH type is characterized as a C→G substitution in the A gamma globin promoter at position –195 and the molecular mechanism responsible for the reactivation of this gene in the Brazilian ndHPFH type remains unclear. In contrast to the British ndHPFH type (-198), where the mechanism responsible for the increase of HbF levels is mediated by the raising in the affinity for the Sp1 transcription factor (TF), the Brazilian ndHPFH mutation does not affect Sp1 binding. Thus, other TF may be involved in the reactivation of the A gamma globin gene in the Brazilian ndHPFH type. The aim of this study was to investigate the mechanism involved in the reactivation or repression of the A gamma globin gene in the Brazilian ndHPFH type and identify possible TF responsible for this phenotype. In vitro primary human erythroblast cultures, derived from human CD34+ hematopoietic cells from 4 Brazilian ndHPFH type subjects and 4 control subjects, were proliferated and differentiated into late stage erythroblasts. The nuclear extracts from predominantly basophilic and polychromatic erythroblasts were used to profile TF activity using Protein-DNA Array method. The analysis of the array densitometry identified a number of TF whose DNA binding activities were either enhanced or repressed in the Brazilian ndHPFH cultures. Among the TF analyzed, the NF-E1/YY1 and the PAX-1 were selected for this study. Since this assay requires a secondary method to confirm these results, nuclear extracts were used to conduct chromatin immunoprecipitation (ChIP) and electrophoretic mobility shift assay (EMSA). ChIP was carried out using antibodies against NF-E1/YY1 and PAX-1 to quantify the binding to these TF to the –195 A gamma globin promoter region. EMSA was performed using probes with the same sequence spotted on the array membrane to analyze the activity of NF-E1/YY1 and PAX-1. Both methods confirmed and validated the previous array results. NF-E1/YY1 is a transcription factor that represses embryonic (epsilon) and fetal (gamma) globin genes. Protein-DNA array and EMSA showed a decreased binding of NF-E1/YY1 in Brazilian ndHPFH nuclear extracts and ChIP analysis revealed diminished NF-E1/YY1 occupancy at the –195 A gamma globin promoter region of Brazilian ndHPFH. The consensus binding site for NF-E1/YY1 is a CCAN motif that is observed between the –195 and –192 position in the A gamma globin promoter region. The C→G substitution at –195 position may disrupt this DNA binding site, cause decreased NF-E1/YY1 interaction and probably allows the binding of PAX-1, a transcriptional activator with a paired box DNA-binding domain that has as a DNA binding core motif, the sequence TTCCGC. This sequence, located between the –199 and –194 position in the A gamma globin promoter, is only presente in the Brazilian type of ndHPFH. Our protein-DNA array and EMSA results showed an increased binding of PAX-1 in the Brazilian ndHPFH nuclear extracts and quantitative ChIP analysis with anti-PAX-1 antibody showed that PAX-1 binds to the –195 A gamma globin promoter region only in the presence of this C→G substitution. These results suggest that the –195 site (C→G) in the A gamma globin promoter region may decrease NF-E1/YY1 binding and increase PAX-1 binding in this DNA region, probably resulting in the reactivation of the A gamma globin gene. The increase in the HbF levels in the Brazilian ndHPFH occurs differently from the British ndHPFH type and represents a novel mechanism of A gamma globin reactivation. Such findings may lead to the development of future therapeutic strategies for HbF induction in the treatment of other hemoglobinopathies. Support by FAPESP and CNPq. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Fetal hemoglobin silencing in humans

Blood ◽  
2006 ◽  
Vol 108 (6) ◽  
pp. 2081-2086 ◽  
Author(s):  
Patricia A. Oneal ◽  
Nicole M. Gantt ◽  
Joseph D. Schwartz ◽  
Natarajan V. Bhanu ◽  
Y. Terry Lee ◽  
...  
Keyword(s):  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Beta Thalassemia ◽  
Postnatal Life ◽  
Globin Genes ◽  
Beta Globin ◽  
Globin Mrna ◽  
Beta Globin Gene ◽  
Gamma Globin ◽  
Age Related

Abstract Interruption of the normal fetal-to-adult transition of hemoglobin expression should largely ameliorate sickle cell and beta-thalassemia syndromes. Achievement of this clinical goal requires a robust understanding of gamma-globin gene and protein silencing during human development. For this purpose, age-related changes in globin phenotypes of circulating human erythroid cells were examined from 5 umbilical cords, 99 infants, and 5 adult donors. Unexpectedly, an average of 95% of the cord blood erythrocytes and reticulocytes expressed HbA and the adult beta-globin gene, as well as HbF and the gamma-globin genes. The distribution of hemoglobin and globin gene expression then changed abruptly due to the expansion of cells lacking HbF or gamma-globin mRNA (silenced cells). In adult reticulocytes, less than 5% expressed gamma-globin mRNA. These data are consistent with a “switching” model in humans that initially results largely from gamma- and beta-globin gene coexpression and competition during fetal development. In contrast, early postnatal life is marked by the rapid accumulation of cells that possess undetectable gamma-globin mRNA and HbF. The silencing phenomenon is mediated by a mechanism of cellular replacement. This novel silencing pattern may be important for the development of HbF-enhancing therapies.

scholarly journals The beta-globin gene on the Chinese delta beta-thalassemia chromosome carries a promoter mutation

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1470-1474 ◽  
Author(s):  
GF Atweh ◽  
XX Zhu ◽  
HE Brickner ◽  
CH Dowling ◽  
HH Jr Kazazian ◽  
...  
Keyword(s):  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Beta Thalassemia ◽  
Beta Globin ◽  
Promoter Mutation ◽  
Beta Globin Gene ◽  
Gamma Globin ◽  
New Type

A new type of delta beta-thalassemia characterized by decreased expression of the beta-globin gene and increased expression of both G gamma and A gamma globin gene in the absence of a detectable deletion has recently been described in the Chinese population. In this study we characterize the mutant beta-globin gene from this delta beta- thalassemia chromosome. An A to G transversion is identified in the “ATA” sequence of the promoter region that leads to decreased expression of the beta-globin gene in vivo and in vitro. We also demonstrate the presence of this mutation in every individual with a high fetal hemoglobin phenotype in this family and its absence in every individual with a normal hemoglobin phenotype. This same promoter mutation has recently been detected in Chinese beta-thalassemia genes where it is present on chromosomes of the same haplotype as that of the delta beta-thalassemia chromosome we are studying. These data support the hypothesis that an as yet unidentified mutation occurred on the ancestral chromosome carrying the promoter mutation and subsequently gave rise to the delta beta-thalassemia phenotype.

scholarly journals Complementary Base Editing Approaches for the Treatment of Sickle Cell Disease and Beta Thalassemia

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3352-3352 ◽  
Author(s):  
Ling Lin ◽  
Adrian P. Rybak ◽  
Conrad Rinaldi ◽  
Jonathan Yen ◽  
Yanfang Fu ◽  
...  
Keyword(s):  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Beta Thalassemia ◽  
Beta Globin ◽  
Cell Disease ◽  
Hematopoietic Stem ◽  
Base Editing ◽  
Gamma Globin

Sickle cell disease (SCD) and Beta thalassemia are disorders of beta globin production and function that lead to severe anemia and significant disease complications across a multitude of organ systems. Autologous transplantation of hematopoietic stem cells engineered through the upregulation of fetal hemoglobin (HbF) or correction of the beta globin gene have the potential to reduce disease burden in patients with beta hemoglobinopathies. Base editing is a recently developed technology that enables precise modification of the genome without the introduction of double strand DNA breaks. Gamma globin gene promoters were comprehensively screened with cytosine and adenine base editors (ABE) for the identification of alterations that would derepress HbF. Three regions were identified that significantly upregulated HbF, and the most effective nucleotide residue conversions are supported by natural variation seen in patients with hereditary persistence of fetal hemoglobin (HPFH). ABEs have been developed that significantly increase the level of HbF following nucleotide conversion at key regulatory motifs within the HBG1 and HBG2 promoters. CD34+ hematopoietic stem and progenitor cells (HSPC) were purified at clinical scale and edited using a process designed to preserve self-renewal capacity. Editing at two independent sites with different ABEs reached 94 percent and resulted in up to 63 percent gamma globin by UPLC. The levels of HbF observed should afford protection to the majority of SCD and Beta thalassemia patients based on clinical observations of HPFH and non-interventional therapy that links higher HbF dosage with milder disease (Ngo et al, 2011 Brit J Hem; Musallam et al, 2012 Blood). Directly correcting the Glu6Val mutation of SCD has been a recent goal of genetic therapies designed for the SCD population. Current base editing technology cannot yet convert mutations like those that result from the A-T transversion in sickle beta globin; however, ABE variants have been designed to recognize and edit the opposite stranded adenine residue of valine. This results in the conversion of valine to alanine and the production of a naturally occurring variant known as Hb G-Makassar. Beta globin with alanine at this position does not contribute to polymer formation, and patients with Hb G-Makassar present with normal hematological parameters and red blood cell morphology. SCD patient fibroblasts edited with these ABE variants achieve up to 70 percent conversion of the target adenine. CD34 cells from healthy donors were then edited with a lead ABE variant, targeting a synonymous mutation in an adjacent proline that resides within the editing window and serves as a proxy for editing the SCD mutation. The average editing frequency was 40 percent. Donor myeloid chimerism documented at these levels in the allogeneic transplant setting exceeds the 20 percent that is required for reversing the sickle phenotype (Fitzhugh et al, 2017 Blood). These next generation editing approaches provide a promising new modality for treating patients with Beta thalassemia and SCD. Disclosures No relevant conflicts of interest to declare.

scholarly journals Benserazide As a Novel Fetal Hemoglobin Inducer: An Observational Study in Non-Carriers of Hemoglobin Disorders

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 2345-2345 ◽  
Author(s):  
Marina Erê A. H. P. Santos ◽  
Felipe Vendrame ◽  
Alvaro Henrique Junqueira Tavares ◽  
Leticia Marani ◽  
Paula C. de Azevedo ◽  
...  
Keyword(s):  
Globin Gene ◽  
Dose Range ◽  
Fetal Hemoglobin ◽  
Beta Thalassemia ◽  
Clinical Use ◽  
Gamma Globin ◽  
Significant Difference ◽  
F Cells ◽  
Chronic Use

Abstract Induction of fetal hemoglobin (HbF) production is an effective therapeutic strategy in the management of patients with beta hemoglobinopathies. Hydroxyurea is the only drug with this mechanism approved for clinical use, and 20% or more of patients do not respond or tolerate it, which has led to the search for new HbF inducers. Benserazide (BEN) is a DOPA decarboxylase inhibitor used in combination with levodopa in the treatment of parkinsonism, but it was also noticed to induce increased gamma globin production in preclinical models. The mechanisms by which BEN acts include downregulation of BCL11A, LSD1 and HDAC3 on the promoter region of the gamma globin gene, making it an interesting candidate for clinical studies in hemoglobinopathies. We hypothesized that patients undergoing treatment for parkinsonism with chronic use of BEN-containing medication may develop increase in HbF production and in circulating F-cells. Material and Methods: Peripheral blood samples were collected from patients with parkinsonism during their follow-up at the Neurology Clinic, who had been using BEN for at least 30 days (BEN group), from healthy controls (group AA), and from patients known to have increased of HbF due to sickle cell anemia (group SS), for comparison purposes. Exclusion criteria for BEN and AA groups were: any hemoglobinopathy, transfusion in the last 90 days, and use of HU or any chemotherapeutic agent. Automated complete blood counts with reticulocyte count were performed on a XN-3000 equipment (Sysmex, Japan), HbF levels were determined by HPLC (BioRad, USA), and F-cell percentage was determined by flow cytometry (BD FACSCalibur, USA). Results: Thirty-five patients on BEN, 10 negative controls (AA group) and five positive controls (SS group) were included. One patient taking BEN was excluded due to HPLC compatible with beta thalassemia trait. Patients taking BEN had blood counts within the normal range. There was no statistically significant difference between BEN and AA groups, and the SS group was significantly anemic as expected. We found a strongly positive correlation between HbF and circulating F-cells (p <0.0001, r = 0.946) when analyzing the entire population, but within the subgroup of patients using BEN, this correlation was much weaker (p = 0.0032, r = 0.492). Dose range of BEN used by the patients was 100-700mg daily, at 1.21 to 11.1mg/kg/day, but we found no correlation between dose and HbF levels. Discussion: We found no differences in blood counts, and extensive use of this regimen support that chronic use of BEN is safe in doses up to 11.1mg/kg/day. In vitro studies suggest that BEN is 30 times more potent than HU. Therefore, considering the minimum dose of HU used in clinical practice of 15mg/kg/day, we had expected that some increase in HbF could be observed at 0.5mg/kg/day of BEN. However, our data show that, even with chronic use of doses 20 times higher, there was no increase in HbF or F-cell levels. The lack of effectiveness of BEN in this population may be explained by its hydrolysis in the intestinal mucosa, and by its reported short half-life (48min), which could account for reduced bioavailability. In addition, patients not bearing beta globin mutations may be less likely to respond to gamma globin induction. Conclusion: Although BEN is already in clinical use and is a strong candidate as a new HbF inducer, we did not detect this effect in humans without hemoglobinopathies. Our data impact the experimental design of future clinical trials with BEN, and suggests the need for more studies on its pharmacokinetics and pharmacodynamics to improve the chances of achieving the desired clinical effect. Disclosures Fertrin: Alexion Pharmaceutical Brasil: Speakers Bureau; Apopharma Inc.: Honoraria.

A Zinc-Finger Transcriptional Activator Designed to Interact with the Gamma-Globin Gene Promoters Enhances Fetal Hemoglobin Production in Erythroid Cells Derived From Normal and Beta-Thalassemic CD34+ Cells.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3567-3567
Author(s):  
Andrew Wilber ◽  
Uli Tschulena ◽  
Phillip W. Hargrove ◽  
Yoon-Sang Kim ◽  
Carlos F. Barbas ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Lentiviral Vector ◽  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Erythroid Differentiation ◽  
Beta Thalassemia ◽  
Normal Donor ◽  
Gene Promoters ◽  
Gamma Globin ◽  
Cd34 Cells

Abstract Abstract 3567 Poster Board III-504 Fetal hemoglobin (α2γ2; HbF) is a potent genetic modifier of the severity of beta-thalassemia and sickle cell anemia. Clinical studies indicate that moderate elevation in production of HbF achieved through heritable persistence of HbF or administration of hydroxyurea, effectively reduce the severity of beta-chain defects. Accordingly, we are exploring strategies to maintain expression of the endogenous gamma-globin genes following lentiviral vector-mediated gene transfer. The artificial zinc-finger transcription factor (GG1-VP64) was designed to interact with sequences in the proximal gamma-globin gene promoters and has been shown to enhance gamma-globin expression in human erythroleukemia cells and mouse marrow cells which are transgenic for the human beta-globin locus. Here, we describe studies designed to evaluate the impact of expression of GG1-VP64 on gamma-globin expression by maturing adult erythroblasts derived from CD34+ cells of normal and thalassemic donors. We utilized an in vitro culture model of human erythropoiesis in which late stage erythroblasts are derived from human CD34+ hematopoietic cells. In this system, cytokine-mobilized peripheral blood or steady state bone marrow CD34+ cells from adults yielded erythroblasts containing 2% or less HbF. The lentiviral vector encodes for bicistronic expression of the GG1-VP64 transactivator and GFP under transcriptional control of the beta-spectrin or ankyrin-1 promoter which give low but progressive increase in expression during erythroid development. Three normal donor CD34+ cells were transduced 48 hours after initiation of culture by overnight exposure to the GG1-VP64 vector or GFP control vector. Approximately 50-60% of the cells were successfully transduced with the control and GG1-VP64 vectors as monitored by flow cytometry analysis for GFP expression. Control vector transduction had no effect on cell proliferation or differentiation monitored by consistent increases in cell numbers and the appearance of CD71 (transferrin receptor) and CD235 (glycophorin A) on most cells (>98% and >80%, respectively) whereas GG1-VP64 gene transfer reduced cell proliferation slightly without affecting erythroid differentiation. Erythroblasts derived from GFP transduced cells expressed low levels of HbF (1.7+/−0.6%) whereas those derived from cells transduced with GG1-VP64 demonstrated induction of HbF ranging from 12-21% with an average vector copy number of 0.8 to 1.0. When cells from a normal donor were sorted into GFP- and GFP+ populations, significant levels of HbF were present only in the GFP+ fraction. We next tested the GG1-VP64 transactivator in three independent studies using bone marrow CD34+ cells from two patients with beta-thalassemia major. Gene transfer was effective as reflected by 74+/−6% (control) and 47+/− 2% (GG1-VP64) GFP marking in bulk cultures. Again, GG1-VP64 gene transfer in beta-thalassemia CD34+ cells reduced cell growth somewhat but did not perturb erythroid differentiation as monitored by the appearance of transferrin receptor (>98%) and Glycophorin A (>80%) as well as cell morphology. Erythroblasts derived from GFP transduced cells expressed levels of HbF in the range of 26+/−5% whereas those derived from cells transduced with GG1-VP64 demonstrated a 2-fold induction of HbF to 52+/−9% with an average vector copy number of 0.5-0.9. Our data show that lentiviral-mediated, enforced expression of GG1-VP64 under the control of erythroid-specific promoters induced significant amounts of HbF in normal and thalassemic erythroblasts derived from adult CD34+ cells without altering their capacity for erythroid maturation following transduction. These observations demonstrate the potential for sequence specific enhancement of HbF in patients with beta-thalassemia or sickle cell anemia. Disclosures: No relevant conflicts of interest to declare.

scholarly journals The beta-globin gene on the Chinese delta beta-thalassemia chromosome carries a promoter mutation

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1470-1474 ◽  
Author(s):  
GF Atweh ◽  
XX Zhu ◽  
HE Brickner ◽  
CH Dowling ◽  
HH Jr Kazazian ◽  
...  
Keyword(s):  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Beta Thalassemia ◽  
Beta Globin ◽  
Promoter Mutation ◽  
Beta Globin Gene ◽  
Gamma Globin ◽  
New Type

Abstract A new type of delta beta-thalassemia characterized by decreased expression of the beta-globin gene and increased expression of both G gamma and A gamma globin gene in the absence of a detectable deletion has recently been described in the Chinese population. In this study we characterize the mutant beta-globin gene from this delta beta- thalassemia chromosome. An A to G transversion is identified in the “ATA” sequence of the promoter region that leads to decreased expression of the beta-globin gene in vivo and in vitro. We also demonstrate the presence of this mutation in every individual with a high fetal hemoglobin phenotype in this family and its absence in every individual with a normal hemoglobin phenotype. This same promoter mutation has recently been detected in Chinese beta-thalassemia genes where it is present on chromosomes of the same haplotype as that of the delta beta-thalassemia chromosome we are studying. These data support the hypothesis that an as yet unidentified mutation occurred on the ancestral chromosome carrying the promoter mutation and subsequently gave rise to the delta beta-thalassemia phenotype.

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