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Query: UMLS:C0019045 (hemoglobinopathies)
 2,704 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Reactivation of fetal hemoglobin genes has been proposed as a potential therapeutic procedure in patients with beta-thalassemia, sickle cell disease, or other beta-hemoglobinopathies. In vitro model systems based on small plasmid globin gene constructs have previously been used in human and mouse erythroleukemic cell lines to study the molecular mechanisms regulating the expression of the fetal human globin genes and their reactivation by a variety of pharmacologic agents. These studies have led to great insights in globin gene regulation and the identification of a number of potential inducers of fetal hemoglobin. In this study we describe the development of enhanced green fluorescence protein (EGFP) reporter systems based on bacterial artificial chromosomes (EBACs) to monitor the activity of the epsilon-, (G)gamma-, (A)gamma-, delta-, and beta-globin genes in the beta-globin locus. Additionally, we demonstrate that transfection of erythroleukemia cells with our EBACs is greatly enhanced by expression of EBNA1, which also facilitates episomal maintenance of our constructs in human cells. Our studies in human cells have shown physiologically relevant differences in the expression of each of the globin genes and also demonstrate that hemin is a potent inducer of EGFP expression from EGFP-modified epsilon-, (G)gamma-, and (A)gamma-globin constructs. In contrast, the EGFP-modified delta- and beta-globin constructs consistently produced much lower levels of EGFP expression on hemin induction, mirroring the in vivo ontogeny. The EGFP-modified beta-globin eukaryotic BAC (EBAC) vector system can thus be used in erythroleukemia cells to evaluate induction of the epsilon- and gamma-globin genes from the intact human beta-globin locus.
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PMID:Development of sensitive fluorescent assays for embryonic and fetal hemoglobin inducers using the human beta -globin locus in erythropoietic cells. 1239 13

The severe hemoglobinopathies, including beta-thalassemia major and sickle cell anemia, are candidate diseases for a genetic treatment based on the transfer of a regulated globin gene in autologous hematopoietic stem cells. Two years ago, May et al reported that an optimized beta-globin transcription unit containing multiple proximal and distal regulatory elements harbored by a recombinant lentiviral vector could efficiently integrate into murine hematopoietic stem cells and express therapeutic levels of the human beta-globin gene. Here, we review the advantages afforded by lentivirus-mediated globin gene transfer and recent studies based on this strategy.
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PMID:Therapeutic globin gene delivery using lentiviral vectors. 1243 53

Patients affected by beta-thalassemia major require lifelong transfusions because of insufficient or absent production of the beta chain of hemoglobin (Hb). A minority of patients are cured by allogeneic bone marrow transplantation. In the most severe of the hitherto available mouse models of beta-thalassemia, a model for human beta-thalassemia intermedia, we previously demonstrated that globin gene transfer in bone marrow cells is curative, stably raising Hb levels from 8.0-8.5 to 11.0-12.0 g/dL in long-term chimeras. To fully assess the therapeutic potential of gene therapy in the context of a lethal anemia, we now have created an adult model of beta(0)-thalassemia major. In this novel model, mice engrafted with beta-globin-null (Hbb(th3/th3)) fetal liver cells succumb to ineffective erythropoiesis within 60 days. These mice rapidly develop severe anemia (2-4 g/dL), massive splenomegaly, extramedullary hematopoiesis (EMH), and hepatic iron overload. Remarkably, most mice (11 of 13) treated by lentivirus-mediated globin gene transfer were rescued. Long-term chimeras with an average 1.0-2.4 copies of the TNS9 vector in their hematopoietic and blood cells stably produced up to 12 g/dL chimeric Hb consisting of mu alpha(2):hu beta(2) tetramers. Pathologic analyses indicated that erythroid maturation was restored, while EMH and iron overload dramatically decreased. Thus, we have established an adult animal model for the most severe of the hemoglobinopathies, Cooley anemia, which should prove useful to investigate both genetic and pharmacologic treatments. Our findings demonstrate the remarkable efficacy of lentivirus-mediated globin gene transfer in treating a fulminant blood disorder and strongly support the efficacy of gene therapy in the severe hemoglobinopathies.
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PMID:A novel murine model of Cooley anemia and its rescue by lentiviral-mediated human beta-globin gene transfer. 1248 Jun 89

A large number of mutations have been described in the human beta-globin locus causing thalassemia or various hemoglobinopathies. However, only a very limited number of these mutations have been studied in animal model systems in the context of the human beta-globin locus. We report here the use of the GET Recombination system with an EcoRI/Kan(R) counterselection cassette to facilitate the introduction of the HbE (codon 26, GAG-->AAG mutation and the codon 41-42 (-TTCT) deletion, two mutations found in high frequency in South-East Asia, into the human beta-globin locus. The counterselection cassette was first inserted into the target sequence in the beta-globin gene, and then a PCR fragment carrying the required modification was used to replace it. Efficient counterselection depends upon the tight regulation of the highly toxic EcoRI endonuclease gene by expression of lacI(q). Induction by IPTG during counterselection efficiently eliminates non-recombinant bacterial clones. The technique can be performed on any known gene sequence using current BAC technology, allowing identification and comparative functional analysis of key regulatory elements, and the development of accurate animal models for human genetic disorders.
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PMID:Insertion of common mutations into the human beta-globin locus using GET Recombination and an EcoRI endonuclease counterselection cassette. 1252 64

Hemoglobin (Hb) Hekinan (alpha27; Glu-Asp) is a rare alpha-chain variant found mainly in Japanese and Chinese whereas Hb E (beta26; Glu-Lys) is common among Southeast Asians. We report a hitherto undescribed condition in which these two variants co-segregate. The proband was a 25-yr-old Thai woman who was encountered with the presence of mild hypochromic microcytosis with Hb 8.2 g/dL, hematocrit (Hct) 26.0%, Mean Corpuscular Value (MCV) 68.6 fL, Mean Corpuscular Hemoglobin (MCH) 21.6 pg and Mean Corpuscular Hemoglobin Concentration (MCHC) 31.5 g/dL. Although Hb electrophoresis at alkaline pH did not show any abnormal band except Hb E in addition to Hb A, high performance liquid chromatography analysis revealed abnormal peaks at the Hb A and Hb E positions. DNA analysis of the proband revealed a GAG-GAT mutation at codon 27 of the minor alpha1-globin gene for Hb Hekinan in trans to the South-east Asian (SEA) deletional alpha-thalassemia 1 determinant and a GAG-AAG mutation at codon 26 of the beta-globin gene for Hb E. She was therefore a triple heterozygote for these three anomalies. Family study identified that her mother was a double heterozygote for Hb Hekinan and Hb E without alpha-thalassemia whereas her father was a classical Hb H disease patient. The genotype-phenotype relationship observed in this Thai family with complex hemoglobinopathies is presented and a simple DNA assay based on the polymerase chain reaction methodology for rapid diagnosis of Hb Hekinan is described.
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PMID:Complex interaction of Hb Hekinan [alpha27(B8) Glu-Asp] and Hb E [beta26(B8) Glu-Lys] with a deletional alpha-thalassemia 1 in a Thai family. 1269 66

The correction of mutant beta-globin genes has long been a therapeutic goal for patients with beta-thalassemia or hemoglobinopathies. The use of homologous recombination (HR) to achieve this goal is an attractive approach because it eliminates the need to include regulatory sequences in the therapeutic construct, and it eliminates mutagenesis induced by random integration. However, HR is a very inefficient process for gene correction, and its efficiency is probably locus dependent. The length of targeting arms is thought to be a determinant of targeting efficiency, so we compared the ability of standard (8-kb) versus very long (16-, 24-, and 110-kb) regions of homology to correct a mutant murine beta-globin gene in embryonic stem cells. Increasing the length of the targeting sequences did not increase the efficiency of HR in this locus, suggesting that alternative approaches will be required to improve the efficiency of this approach for globin gene correction.
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PMID:Long targeting arms do not increase the efficiency of homologous recombination in the beta-globin locus of murine embryonic stem cells. 1273 Jan 7

Reactivation of fetal hemoglobin (HbF) expression using pharmacological agents represents a potential strategy for the therapy of beta-thalassemia, sickle cell disease, HbE and other beta-hemoglobinopathies. However, the drugs currently available have low efficacy and specificity and are associated with high toxicity. We describe the development of stable cellular genomic reporter assays (GRAs) based on the green fluorescence protein (EGFP) gene under the Ggamma-globin promoter in the intact human beta-globin locus. We show that human erythroleukemic cell lines stably transfected with a Ggamma-EGFP beta-globin locus construct can maintain a uniform basal level of EGFP expression over long periods of continuous culture and that induction of EGFP expression parallels the induction of the endogenous globin genes. We compared the EGFP-induction potency of a number of chemotherapeutic agents, including histone deacetylase inhibitors and DNA-binding agents. We show that hydroxyurea and butyrate result in moderate levels of induction (70-80%) but with an additive inductive effect. Among the DNA-binding agents tested, cisplatin was the most potent inducer of HbF expression, (442+/-32%), a level which is comparable to hemin (764+/-145%). These results indicate that cellular GRAs containing Ggamma-EGFP-modified beta-globin locus constructs can be used to develop novel inducers of HbF synthesis for the therapy of beta-hemoglobinopathies.
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PMID:Cellular genomic reporter assays for screening and evaluation of inducers of fetal hemoglobin. 1464 8

We have studied 918 chromosomes for mutations leading to beta-thalassemia and sickle cell anemia, which are the two most frequently found monogenic disorders in Antalya, Turkey. Three hundred and seventy-seven postnatal and 82 prenatal cases were studied between 2000 and May 2003 in our center using reverse dot blot hybridization (RDBH) with 22 probes specific for Mediterranean populations. In this study, IVSI-110 (G-->A) appeared to be the most common mutation with an occurrence rate of 44.4% among the 16 different mutations found to be associated with beta-thalassemia. Heterozygosity for IVSI-110 was the most prevalent combination, whereas 34 of our 377 postnatal cases showed homozygosity for this mutation, a genotype leading to beta-thalassemia major. The total percentage of postnatal patients clinically diagnosed as beta-thalassemia major was 18.6%, whereas 5% of the cases were diagnosed clinically as beta-thalassemia intermedia. One new Hb variant, Hb Antalya, and one new mutation, Cod 3 (+T) were found. HbS accounted for 10.3% of all mutations; homozygosity was found in 1.9% of all cases. Of the 82 cases analysed prenatally for beta-globin gene mutations and by cytogenetic techniques for possible chromosomal abnormalities, 21 fetuses were found to be affected with beta-globin gene mutations. One of these fetuses was also found to have a 45,X karyotype, and 1 had a 46,XY/47,XY,+22 karyotype. Quite a high rate of consanguineous marriages in Antalya (35.17%) renders mutation screening, genetic counseling, and educational programs held by our Thalassemia Unit essential. This study was the first to be performed specifically in our region where hemoglobinopathies are most frequent as a consequence of migrations of racially and culturally distinct groups to the area in the distant past.
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PMID:Molecular analysis of beta-thalassemia and sickle cell anemia in Antalya. 1515 12

beta-Thalassemia and sickle cell disease constitute the most common single gene defect in humans and are characterized by absent/reduced and abnormal beta-globin, respectively. Patients with these hemoglobinopathies require chronic blood transfusions and rely on bone marrow transplant for a potential cure. Gene therapy provides a viable alternative by permanent correction of the defect in hematopoietic stem cells, but it has suffered from problems of vector instability, low viral titers, and variable expression of globin genes for more than a decade. This paper reviews the field of gene therapy for hemoglobinopathies with emphasis on the complexities of the beta-globin gene and its regulation, obstacles that have impeded progress, and recent advances in vector technology that will take this field forward toward the goal of successful genetic correction of these devastating diseases.
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PMID:Gene therapy for hemoglobinopathies: are we there yet? 1521 60

Hemoglobinopathies are caused by abnormal structure or synthesis of hemoglobin chains and represent serious monogenic disorders. A new study demonstrates that lentiviral vectors can express clinically relevant levels of human transgenic beta-globin in red cells of xenografted mice. While some safety concerns must be addressed, this study is an important step toward potential clinical trials of gene therapy for hemoglobinopathies.
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PMID:Lenti in red: progress in gene therapy for human hemoglobinopathies. 1546 34


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