Researchers at the UCLA Department of Microbiology, Immunology & Molecular Genetics have developed novel methods to achieve efficient, precise gene integration and effective expression of cDNA cassettes to express normal versions of genes in hematopoietic stem cells.
BACKGROUND:
Hematopoietic stem cells (HSCs)—stem cells that give rise to other blood cells—have great therapeutic potential because of their ability to both self-renew and differentiate into other blood cell types. Many severe primary immune deficiencies (PIDs) are due to defects in lymphoid or hematopoietic cells that can be reconstituted through hematopoietic stem cell transplantation (HSCT). Unlike allogeneic HSCT wherein cells are taken from a donor, gene therapy using a patient’s own HSCs has potential advantages for the absence of graft-versus-host disease and requires less intense conditioning and immune suppression. HSC-targeted gene editing technologies using engineered nucleases such as CRISPR/Cas9 allow for the safe, site-specific correction of disease-causing mutations and have shown promising clinical benefits in multiple diseases. Correction of specific disease-causing mutations extends the potential applications of gene therapy to dominant disorders in addition to those resulting from simple loss-of-function mutations. Despite advancements, development efficient, selective and precise gene-editing therapies is absent for many disease-causing mutations.
INNOVATION:
UCLA researchers have optimized the gene editing process for X-linked hyper-IgM syndrome (XHIM), a genetic condition caused by mutations in the CD40LG gene that affects the immune system. Optimal gene editing with CRISPR/Cas9 requires fine-tuning design features, including the homologous donors (cDNA expression cassettes and homology arms) and target sites for more efficient, precise gene integration. This optimization decreases off-target cuts in undesired locations and allows expression of the transgene to the highest level possible.
POTENTIAL APPLICATIONS:
•Gene therapy of immune deficiencies, including X-linked Hyper IgM Syndrome
•Gene therapy of hemoglobinopathies
•Gene therapy of storage and metabolic diseases
ADVANTAGES:
•More efficient, safe, and effective gene editing
•Better expression of inserted transgenes
DEVELOPMENT-TO-DATE:
The method has been tested in vitro and in vivo in immunodeficient mouse models.
Related Papers (from the inventors only)
Kohn, D.B. and Kuo, C.Y., New frontiers in the therapy of primary immunodeficiency: From gene addition to gene editing. Journal of Allergy and Clinical Immunology, 2017, 139, 726-732.