Editing outside the body: Ex vivo gene-modification for β-hemoglobinopathy cellular therapy

Efficacy of CRISPR-Based Gene Editing in a Sickle Cell Disease Patient as Measured through the Eye

“…This case confirms that CRISPR-based gene editing in SCD, while substantially effective, may not completely eliminate vaso-occlusive activity. This is consistent with prior reports that show that the success of gene editing varies between individuals and gene editing results in a phenotypic spectrum [ 3 ]. What was unclear in our case, since % HbS after gene editing was 50%, is whether the continued vaso-occlusive events in our patient were due to residual circulating sickled RBCs, persistent dysfunction of the endothelium and coagulation cascade, or a combination of both.…”

“…Evidence suggests that SCD exists on a phenotypic spectrum with variable genetic expressivity, making it difficult to stratify risk of complications for a given individual [ 2 ]. Even with current and emerging SCD treatments, including CRISPR-based gene editing of bone-marrow-derived hematopoietic stem cells, studies have shown that a variable degree of phenotypic mosaicism persists, with a variable proportion of red blood cells (RBCs) still able to sickle and cause disease [ 3 ].…”

Recent advances in “sickle and niche” research - Tribute to Dr. Paul S Frenette -

“…HSCT for SCD using genetically modified autologous HSC donor cells is a promising experimental approach that circumvents some toxicities of allogeneic HSCT ( Doerfler et al., 2021 ; Drysdale et al., 2021 ; Rosanwo and Bauer, 2021 ). One approach for HSC modification is transduction with a lentiviral vector encoding a modified β-globin gene with anti-sickling properties (LentiGlobin), which results in the production of HbA ( Figure 2 ).…”

Fluorinated Silane-Modified Filtroporation Devices Enable Gene Knockout in Human Hematopoietic Stem and Progenitor Cells

“…Autologous gene therapies are elegant and promising alternatives, whereby the patient’s own cells are modified at the genomic level to correct genotypes and alleviate disease phenotypes. To date, most clinical progress has been made in the field of viral vector-mediated gene modification, , which harnesses viruses’ natural ability to enter cells and to modify DNA. The manufacturing of these viral-based therapies has been burdened with extremely high costs, while populations that are frequently affected by prevalent hematological disorders are often located in medically underserved and/or low-resource regions of the world, underscoring the need for intracellular delivery technologies that are accessible and easy to use and require little training to operate. , Additionally, issues with potential insertional mutagenesis due to semirandom gene insertion mediated by viral carriers have driven the gene-editing field away from utilizing viral vectors and toward more targeted strategies such as those employing zinc-finger nucleases, transcription activator-like effector nucleases (TALENs), clustered regularly interspaced palindromic repeats (CRISPR)-Cas, and, more recently, prime and base editors. ,, However, these important gene-modifying biomolecules are often large proteins that need to be delivered to cells using non-cytotoxic and effective intracellular delivery strategies, because the latest favored viral vectors suffer from size limitations and are thus unable to carry the large DNA constructs encoding these proteins. ,− Additionally, gene manipulation through targeted knockouts is an important research tool to elucidate functional gene roles and pathways that may inform clinical targets and outcomes …”