1,721,197 research outputs found
Genetic modification of human hematopoietic stem cells
No full textTransplantation of genetically modified hematopoietic stem cells is a potential therapy for a variety of genetic and acquired blood disorders, such as severe combined immunodeficiencies, thalassemia and AIDS. Genetic modification of stem cells can be carried out ex vivo, by transducing bone marrow or peripheral blood stem cell-rich fractions with viral vectors carrying therapeutic genes. These vectors must be able to transduce long-term repopulating stem cells, and allow appropriate transgene expression. Recent improvements in cell culture and vector technology are providing new tools for obtaining clinically relevant numbers of genetically modified hematopoietic stem cells from a standard bone marrow harvest. The proof of the therapeutic potential of this technology is the successful therapy of two different forms of severe combined immunodeficiencies, rare genetic disorders associated with recurrent infections that may be fatal in the first years of life. These pivotal trials, however, have also uncovered the oncogenic potential of random retroviral insertions into the human genome. For non life-threatening disorders, the risk of insertional oncogenesis may be unacceptable, and new research is needed to develop new, safer and more efficient gene transfer vectors for clinical application
Alternative splicing caused by lentiviral integration in the human genome
No full textGene transfer vectors derived from murine oncoretroviruses or human lentiviruses are widely used in human gene therapy. Integration of these vectors in the human genome may, however, have genotoxic effects, caused by deregulation of gene expression at the transcriptional or posttranscriptional level. In particular, integration of lentiviral vectors within transcribed genes has a significant potential to affect their expression by interfering with splicing and polyadenylation of primary transcripts. Aberrant splicing is caused by the usage of both constitutive and cryptic splice sites located in the retroviral backbone as well as in the gene expression cassettes. We describe a set of simple methods that allow the identification of chimeric transcripts generated by the insertion of a lentiviral vector within genes and the evaluation of their relative abundance. Identification of the splice sites, either constitutive or cryptic, that are frequently used by the cell splicing machinery within a given vector provides a useful resource to attempt recoding of the vector with the objective of reducing its potential genotoxicity in a clinical context. © 2012 Elsevier Inc. All rights reserved
Gene Therapy for Hemoglobinopathies
Full text linkGene therapy for β-Thalassemia and sickle-cell disease is based on transplantation of genetically corrected, autologous hematopoietic stem cells. Preclinical and clinical studies have shown the safety and efficacy of this therapeutic approach, currently based on lentiviral vectors to transfer a β-globin gene under the transcriptional control of regulatory elements of the β-globin locus. Nevertheless, a number of factors are still limiting its efficacy, such as limited stem-cell dose and quality, suboptimal gene transfer efficiency and gene expression levels, and toxicity of myeloablative regimens. In addition, the cost and complexity of the current vector and cell manufacturing clearly limits its application to patients living in less favored countries, where hemoglobinopathies may reach endemic proportions. Gene-editing technology may provide a therapeutic alternative overcoming some of these limitations, though proving its safety and efficacy will most likely require extensive clinical investigation
Specificity of HOX protein function depends on DNA-protein and protein- protein interactions, both mediated by the homeo domain
Full text linkTranscription of human HOX gene promoters in cultured cells is positively and negatively regulated by HOX proteins interacting with specific target sequences. The human HOXD9 protein activates transcription of the HOXD9 promoter by interacting with the HCR sequence and is antagonized by the HOXD8 protein. HOXD8 is not intrinsically a repressor, since it can activate transcription on different targets. Complete or partial HOXD8/HOXD9 homeo domain swapping indicates that the ability to recognize, and activate transcription from, the HCR target in vivo depends on the amino terminus and helix 1 of the homeo domain. The inhibitory activity of HOXD8 is not affected by deletion of the homeo domain helix 2/3 region, whereas it requires the amino terminus/helix 1 region and an additional, effector domain located at the protein amino-terminal end. This activity is therefore DNA-binding independent, and possibly mediated by protein-protein interactions. Affinity chromatography experiments show that the homeo domain amino terminus/helix 1 region is able to mediate direct interactions between HOX proteins in solution. These data indicate that specificity of HOX protein function in vivo depends on both DNA-protein and protein-protein interactions, mediated by the same sub region of the homeo domain
Gene therapy of inherited skin adhesion disorders
No full textGene therapy is a potential treatment for devastatinginherited diseases for which there is little hope offinding a conventional cure. These include lethal diseaseslike immunodeficiencies and metabolic disorders,and non-lethal conditions associated with poorquality of life and life-long symptomatic treatments,like muscular dystrophy, cystic fibrosis or thalassemia.Skin adhesion defects belong to both groups. For thenon-lethal forms, gene therapy, or transplantation ofcultured skin derived from genetically corrected epidermalstem cells, represents a very attractive therapeuticoption, and potentially a definitive treatment.Recent advances in gene transfer and stem cell culturetechnology are making this option closer than ever.This paper critically reviews the progress and prospectsof gene therapy for skin adhesion defects, andthe technical and non-technical factors currently limitingits development.Gene therapy is a potential treatment for devastating inherited diseases for which there is little hope of finding a conventional cure. These include lethal diseases like immunodeficiencies and metabolic disorders, and non-lethal conditions associated with poor quality of life and life-long symptomatic treatments, like muscular dystrophy, cystic fibrosis or thalassemia. Skin adhesion defects belong to both groups. For the non-lethal forms, gene therapy, or transplantation of cultured skin derived from genetically corrected epidermal stem cells, represents a very attractive therapeutic option, and potentially a definitive treatment. Recent advances in gene transfer and stem cell culture technology are making this option closer than ever. This paper critically reviews the progress and prospects of gene therapy for skin adhesion defects, and the technical and non-technical factors currently limiting its development. © 2009 Elsevier Ltd. All rights reserved
Haemoglobin lepore in Italy: Haematological and structural studies
No full textThis paper records the mean haematological and haemoglobin data and the structural characterization of 43 Hb Lepore heterozygotes from 27 apparently unrelated families detected either during screening programs or during haematological investigations of patients in the following parts of Italy: 1. Sicily (12 Hb Lepore heterozygotes from 10 apparently unrelated families); 2. Calabria (12 Hb Lepore heterozygotes from 4 apparently unrelated families); 3. Pyglia (2 related Hb Lepore heterozygotes); 4. Basilicata (3 related Hb Lepore heterozygotes); 5. Campania (7 apparently unrelated Hb Lepore heterozygotes); 6. Abruzzo (4 Hb Lepore heterozygotes from 2 apparently unrelated families); 7. Marche (2 related Hb Lepore heterozygotes); 8. Tuscany (1 Hb Lepore heterozygote)
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