Search

X-linked severe combined immunodeficiency (X-SCID) is a primary immunodeficiency that is caused by mutations in the interleukin-2 receptor gamma

Mutations in the human β-globin gene are the cause of β-hemoglobinopathies, one of the most common inherited single-gene blood disorders in the world. Novel therapeutic approaches are based on lentiviral vectors (LVs) or CRISPR-Cas9-mediated gene disruption to express adult hemoglobin (HbA), or to reactivate the completely functional fetal hemoglobin, respectively. Nonetheless, LVs present a risk of insertional mutagenesis, while gene-disrupting transcription factors (BCL11A, KLF1) involved in the fetal-to-adult hemoglobin switch might generate dysregulation of other cellular processes. Therefore, universal gene addition/correction approaches combining CRISPR-Cas9 and homology directed repair (HDR) by delivering a DNA repair template through adeno-associated virus could mitigate the limitations of both lentiviral gene transfer and gene disruption strategies, ensuring targeted integration and controlled transgene expression. In this study, we attained high rates of gene addition (up to 12%) and gene correction (up to 38%) in hematopoietic stem and progenitor cells from healthy donors without any cell sorting/enrichment or the application of HDR enhancers. Furthermore, these approaches were tested in heterozygous (β0/β+) and homozygous (β0/β0, β+/β+) β-thalassemia patients, achieving a significant increase in HbA and demonstrating the universal therapeutic potential of this study for the treatment of β-hemoglobinopathies.

Metachromatic leukodystrophy (MLD) is a rare genetic disorder caused by mutations in the

Due to pioneering in vitro investigations on gene modification, gene engineering platforms have incredibly improved to a safer and more powerful tool for the treatment of multiple blood and immune disorders. Likewise, several clinical trials have been initiated combining autologous hematopoietic stem cell transplantation (auto-HSCT) with gene therapy (GT) tools. As several GT modalities such as lentivirus and gene editing tools have a long developmental path ahead to diminish its negative side effects, it is hard to decide which modality is optimal for treating a specific disease. Gene transfer by lentiviruses is the platform of choice for loss-of-mutation diseases, whereas gene correction/addition or gene disruption by gene editing tools, mainly CRISPR/Cas9, is likely to be more efficient in diseases where tight regulation is needed. Therefore, in this review, we compiled pertinent information about lentiviral gene transfer and CRISPR/Cas9 gene editing, their evolution to a safer platform for HSCT, and their applications on other types of gene disorders based on the etiology of the disease and cell fitness.

β-Hemoglobinopathies are among the most common single-gene disorders and are caused by different mutations in the β-globin gene. Recent curative therapeutic approaches for these disorders utilize lentiviral vectors (LVs) to introduce a functional copy of the β-globin gene into the patient's hematopoietic stem cells. Alternatively, fetal hemoglobin (HbF) can reduce or even prevent the symptoms of disease when expressed in adults. Thus, induction of HbF by means of LVs and other molecular approaches has become an alternative treatment of β-hemoglobinopathies. Here, we performed a head-to-head comparative analysis of HbF-inducing LVs encoding for: 1) IGF2BP1, 2) miRNA-embedded shRNA (shmiR) sequences specific for the γ-globin repressor protein BCL11A, and 3) γ-globin gene. Furthermore, two novel baboon envelope proteins (BaEV)-LVs were compared to the commonly used vesicular-stomatitis-virus glycoprotein (VSV-G)-LVs. Therapeutic levels of HbF were achieved for all VSV-G-LV approaches, from a therapeutic level of 20% using γ-globin LVs to 50% for both IGF2BP1 and BCL11A-shmiR LVs. Contrarily, BaEV-LVs conferred lower HbF expression with a peak level of 13%, however, this could still ameliorate symptoms of disease. From this thorough comparative analysis of independent HbF-inducing LV strategies, we conclude that HbF-inducing VSV-G-LVs represent a promising alternative to β-globin gene addition for patients with β-hemoglobinopathies.