Preclinical evaluation for engraftment of CD34+ cells gene-edited at the sickle cell disease (SCD) locus in xenograft mouse and non-human primate models

April 20, 2021

Associate Professor Naoya Uchida of The Institute of Medical Science, The University of Tokyo in Japan and co-researchers at National Heart Lung and Blood Institutes (NHLBI), National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), and National Institute of Allergy and Infectious Diseases (NIAID), National Institutes of Health (NIH) as well as MaxCyte, Inc., in the U.S. developed a viral vector-free non-footprint gene correction method for the SCD mutation at therapeutic levels in DNA (~30%) and proteins (~80%) and demonstrated engraftment of gene-edited CD34+ cells post-transplant in xenograft mice and non-human primates. It was published in "Cell Reports Medicine" on April 20, 2021.

 

Abstract

Sickle cell disease (SCD) is caused by a 20A>T mutation in the β-globin gene. Genome editing technologies have the potential to correct the SCD mutation in hematopoietic stem cells (HSCs), producing adult hemoglobin while simultaneously eliminating sickle hemoglobin. Here, we developed high-efficiency viral vector-free non-footprint gene correction in SCD CD34+ cells with electroporation to deliver SCD mutation-targeting guide RNA, Cas9 endonuclease, and 100-mer single strand donor DNA encoding an intact β-globin sequence, achieving therapeutic-level gene correction at the DNA (~30%) and protein (~80%) levels. Gene-edited SCD CD34+ cells contributed corrected cells 6 months post-xenograft mouse transplant without off-target δ-globin editing. We then developed a rhesus β-to-βs-globin gene conversion strategy to model HSC-targeted genome editing for SCD, and demonstrate engraftment of gene-edited CD34+ cells 10-12 months post-transplant in rhesus macaques. In summary, gene-corrected CD34+ HSCs are engraftable in xenograft mice and non-human primates. These findings will aid the design of HSC-targeted gene correction trials.



Development of hematopoietic stem cell (HSC)-targeted gene-correction therapy for sickle cell disease (SCD). We developed a high-efficiency gene correction method for the SCD mutation and demonstrated engraftment of gene-edited CD34+ hematopoietic stem cells post-transplant in xenograft mice and non-human primates.