Correction of the sickle cell mutation in embryonic stem cells

Abstract

Sickle cell anemia is one of the most common genetic diseases worldwide. Patients often suffer from anemia, painful crises, infections, strokes, and cardiopulmonary complications. Although current management has improved the quality of life and survival of patients, cure can be achieved only with bone marrow transplantation when histocompatible donors are available. The ES cell technology suggests that a therapeutic cloning approach may be feasible for treatment of this disease. Using a transgenic͞knockout sickle cell anemia mouse model, which harbors 240 kb of human DNA sequences containing the ␤ S -globin gene, we prepared ES cells from blastocysts that had the sickle cells anemia genotype and carried out homologous recombination with DNA constructs that contained the ␤ A -globin gene. We obtained ES cells in which the ␤ S was corrected to the ␤ A sequence. Hematopoietic cells differentiated from these ES cells produced both hemoglobin A and hemoglobin S. This approach can be applied to human ES cells to correct the sickle mutation as well as ␤-thalassemia mutations.correction of mutation ͉ homologous recombination ͉ sickle cell anemia ͉ hematopoietic differentiation ͉ ␤-thalassemia

Beta-thalassemia

“…148,149 Specifically, the vector developed by Sadelain et al containing HS2, 3, and 4 from LCR associated with an extended beta globin gene led to partial correction of anemia in a beta-thalassemia mouse. 147 Regarding the alternative approach, in a mouse model of sickle cell anemia, Chang et al 150 were able to correct the molecular defect by homologous recombination by transfecting the embryonic stem cells from the affected mice with a DNA fragment containing the normal beta globin gene sequences. Hematopoietic stem cells, derived from the corrected embryonic stem cells ex vivo, were able to produce HbA and HbS, thereby leading to a phenotype similar to human sickle cell trait.…”

Customizing the genome as therapy for the β-hemoglobinopathies

“…Genetic correction of the SCD and b-thalassemia mutations Classical gene-targeting approaches have been used to repair the SCD mutation in embryonic stem cells, 75 but this approach cannot be applied HbF: Figure 1. Network of potential targets for genomeediting-based therapy of the b-globin disorders.…”

Transgenes Delivered by Lentiviral Vector are Suppressed in Human Embryonic Stem Cells in A Promoter-Dependent Manner

“…The potential of these cell lines can be further enhanced by genetic modifications, which may promote controlled differentiation of stem cells along a specific developmental pathway (3), facilitate purification of a certain type of cells in a mixed population of ES-derived cells (4), or correct genetic defects (5). In spite of all the potential this approach presents, the standard chemical or mechanical methods for transgene delivery exhibited very low efficiency in human ES cells, with the most effective approaches yielding only approximately 1 stably transfected cell per 10 5 cells (6,7).…”