Researchers have used a gene editing technique to treat a rare immune disorder that could help overcome concerns over the use of gene therapy in these patients. The treatment targets deficiencies in recombination activating genes (RAGs), which can affect the production of lymphocytes and cause severe immune disorders. Specifically, it deploys homology-directed repair (HDR) processes that are commonly used for mending double-stranded breaks in their DNA.This offers the unique advantage of editing the affected gene while preserving its normal regulation.
The finding could help overcome obstacles in gene therapy for patients with mutations in RAG1, which has shown promise but has safety concerns due to the resulting constitutive expression in this gene.
“Our results support the implementation of the exonic gene editing strategy toward clinical application for the treatment of RAG1 deficiency,” report Maria Castiello, PhD, from San Raffaele-Telethon Institute for Gene Therapy in Milan, and co-workers in the journal Science Translational Medicine.
RAG gene defects lead to reduced survival and poor quality of life unless treated by hematopoietic stem cell transplantation (HSCT). However, this treatment of choice is limited by donor availability and outcomes can be compromised by several factors including age, clinical status at the time of transplantation and toxicity.
By contrast, HDR–mediated knockin of a corrective sequence provides an alternative approach for in situ correction of most disease-causing mutations while preserving the physiological regulation of the targeted gene.
The researchers used a combined knockout and knockin strategy to correct RAG1 genes in HSPCs from patients, which rescued protein expression.Transplanting the edited HSPCs into humanized mice showing a version of severe combined immunodeficiency improved B cell production.
Gene editing strategies that targeted exon 2 outperformed that targeting intron 1.
“Overall, we showed that HDR editing of hRAG1 exon 2 allows functional rescue of RAG1 defects by restoring its expression and activity, overcoming lymphoid differentiation block, and leading to the generation of mature T and B cells,” the team writes.
“This study represents a proof-of-concept demonstration of therapeutically relevant HDR correction of the hRAG1 locus for the treatment of RAG1 deficiency, supporting further study of this exonic GE platform toward clinical testing.”