Shortly after birth, fetal hemoglobin is replaced by adult hemoglobin in red cells, a process that reflects a developmental switch in the β-globin locus that favors the expression of β-globin and the ...suppression of γ-globin. Therapies that may abrogate this switch have long been pursued on the basis of observations that the persistence of fetal-hemoglobin production after birth mitigates the phenotypes of sickle cell disease and β-thalassemia major, as well as the absence of signs of either disease when fetal hemoglobin levels are high in utero or at birth.
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Pharmacologic inhibition of the globin developmental switch would require regular and . . .
Genetic diseases of blood cells are prime candidates for treatment through ex vivo gene editing of CD34
hematopoietic stem/progenitor cells (HSPCs), and a variety of technologies have been proposed ...to treat these disorders. Sickle cell disease (SCD) is a recessive genetic disorder caused by a single-nucleotide polymorphism in the β-globin gene (HBB). Sickle hemoglobin damages erythrocytes, causing vasoocclusion, severe pain, progressive organ damage, and premature death. We optimize design and delivery parameters of a ribonucleoprotein (RNP) complex comprising Cas9 protein and unmodified single guide RNA, together with a single-stranded DNA oligonucleotide donor (ssODN), to enable efficient replacement of the SCD mutation in human HSPCs. Corrected HSPCs from SCD patients produced less sickle hemoglobin RNA and protein and correspondingly increased wild-type hemoglobin when differentiated into erythroblasts. When engrafted into immunocompromised mice, ex vivo treated human HSPCs maintain SCD gene edits throughout 16 weeks at a level likely to have clinical benefit. These results demonstrate that an accessible approach combining Cas9 RNP with an ssODN can mediate efficient HSPC genome editing, enables investigator-led exploration of gene editing reagents in primary hematopoietic stem cells, and suggests a path toward the development of new gene editing treatments for SCD and other hematopoietic diseases.
An unprespecified interim analysis of results from a phase 1–2 study of gene therapy for sickle cell disease shows resolution of severe vaso-occlusive events in 25 patients who could be evaluated. In ...the 24 months before enrollment, these patients had a median of 3.5 severe vaso-occlusive events per year.
Gene therapy with CD34+ cells transduced with a lentivirus vector carrying a β-globin gene was performed in 22 patients. At a median of 26 months, all the patients were either transfusion-independent ...or had a major reduction in transfusion requirements.
PURPOSE OF REVIEWHematopoietic cell transplantation (HCT) is a curative therapy for sickle cell disease (SCD) that is utilized very rarely because of limited allogeneic donor availability, limited ...healthcare resources needed to expand the treatment to regions in the world where most affected individuals reside, and by a view among SCD experts that HCT lacks the evidential rigor with short and long-term toxicity profiles that together might support its broader application.
RECENT FINDINGSIn this update, recent advances focused on donor selection, reduced toxicity preparation for HCT, and treatment of young adults will be presented. The current status of conventional bone marrow transplantation with a human leukocyte antigen-identical sibling donor is summarized.
SUMMARYHCT for SCD is curative in almost all children who have a human leukocyte antigen-matched sibling donor. The future of this therapy will hinge on expanding the number of individuals who might be treated.
A growing number of gene therapy‐ and gene editing‐based treatments for patients with sickle cell disease (SCD) are entering clinical trials. These treatments, designed to target the underlying cause ...of SCD, have the potential to provide functional cures, which until now were possible only through allogeneic hematopoietic stem cell transplant. However, as these novel approaches advance from early‐ to late‐stage clinical trials, it is essential to identify physiologically and clinically relevant endpoints that can demonstrate the achievement of a functional cure for SCD. Here, we present an overview of the pathophysiology of SCD and current treatment options, review ongoing SCD clinical trials using gene therapy or gene editing approaches, and identify the most relevant endpoints for demonstrating the attainment of a functional cure for SCD.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Introduction: Sickle cell anemia (SCA) is a hereditary blood disease caused by a single-gene mutation that affects millions of individuals world-wide. In this review, we focus on techniques to treat ...SCA by ex vivo genetic manipulation of hematopoietic stem/progenitor cells (HSPC), emphasizing replacement gene therapy and gene editing.
Areas covered: Viral transduction of an anti-sickling β-like globin gene has been tested in pre-clinical and early-phase clinical studies, and shows promising preliminary results. Targeted editing of endogenous genes by site-directed nucleases has been developed more recently, and several approaches also are nearing clinical translation.
Expert commentary: The indications and timing of gene therapy for SCA in lieu of supportive care treatment and allogeneic hematopoietic cell transplantation are still undefined. In addition, ensuring access to the treatment where the disease is endemic will present important challenges that must be addressed. Nonetheless, gene therapy and gene editing techniques have transformative potential as a universal curative option in SCA.
Sickle Cell Disease and ß-thalassemia, which are caused by defective or deficient adult ß-globin (HBB) respectively, are the most common serious genetic blood diseases in the world. Persistent ...expression of the fetal ß-like globin, also known as 𝛾-globin, can ameliorate both disorders by serving in place of the adult ß-globin as a part of the fetal hemoglobin tetramer (HbF). Here we use CRISPR-Cas9 gene editing to explore a potential 𝛾-globin silencer region upstream of the δ-globin gene identified by comparison of naturally-occurring deletion mutations associated with up-regulated 𝛾-globin. We find that deletion of a 1.7 kb consensus element or select 350 bp sub-regions from bulk populations of cells increases levels of HbF. Screening of individual sgRNAs in one sub-region revealed three single guides that caused increases in 𝛾-globin expression. Deletion of the 1.7 kb region in HUDEP-2 clonal sublines, and in colonies derived from CD34+ hematopoietic stem/progenitor cells (HSPCs), does not cause significant up-regulation of 𝛾-globin. These data suggest that the 1.7 kb region is not an autonomous 𝛾-globin silencer, and thus by itself is not a suitable therapeutic target for gene editing treatment of ß-hemoglobinopathies.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Despite advances in supportive therapy to prevent complications of sickle cell disease (SCD), access to care is not universal. Hematopoietic cell transplantation is, to date, the only curative ...therapy for SCD, but its application is limited by availability of a suitable HLA-matched donor and lack of awareness of the benefits of transplant. Included in this study are 1000 recipients of HLA-identical sibling transplants performed between 1986 and 2013 and reported to the European Society for Blood and Marrow Transplantation, Eurocord, and the Center for International Blood and Marrow Transplant Research. The primary endpoint was event-free survival, defined as being alive without graft failure; risk factors were studied using a Cox regression models. The median age at transplantation was 9 years, and the median follow-up was longer than 5 years. Most patients received a myeloablative conditioning regimen (n = 873; 87%); the remainder received reduced-intensity conditioning regimens (n = 125; 13%). Bone marrow was the predominant stem cell source (n = 839; 84%); peripheral blood and cord blood progenitors were used in 73 (7%) and 88 (9%) patients, respectively. The 5-year event-free survival and overall survival were 91.4% (95% confidence interval, 89.6%-93.3%) and 92.9% (95% confidence interval, 91.1%-94.6%), respectively. Event-free survival was lower with increasing age at transplantation (hazard ratio HR, 1.09; P < .001) and higher for transplantations performed after 2006 (HR, 0.95; P = .013). Twenty-three patients experienced graft failure, and 70 patients (7%) died, with the most common cause of death being infection. The excellent outcome of a cohort transplanted over the course of 3 decades confirms the role of HLA-identical sibling transplantation for children and adults with SCD.
•HLA-identical sibling transplantation for SCD offers excellent long-term survival.•Mortality risk is higher for older patients; event-free survival has improved in patients transplanted after 2006.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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