Base editing of Artemis mutations ex vivo sheds light on gene therapy for Artemis-deficient SCID

Abstract

Artemis is an endonuclease that cleaves DNA hairpins during V(D)J recombination, a critical step enabling coding-joint formation and essential for lymphocyte development. Artemis-deficient severe combined immunodeficiency (ART-SCID) is a monogenic disorder marked by impaired lymphocyte development and poor responsiveness to allogeneic hematopoietic stem cell transplantation. In this study, we explored the use of base editors to correct mutations in the Artemis gene. Specifically, we firstly documented the pathogenic mutations of Artemis from previous studies and newly identified from ClinVar database. Then, we performed ex vivo assays using cytidine base editors (CBEs) to repair the c.181T > C mutation, and adenosine base editors (ABEs) to target the pathogenic c.49G > A and c.404G > A variants associated with ART-SCID. Targeted deep sequencing revealed that rAPOBEC1-SpRY-HF1-BE4max achieved efficient editing (~ 50%) at the c.181T > C site, while ABE8e reached ~ 35% and ~ 20% editing efficiency at the c.49G > A and c.404G > A sites, respectively. Importantly, base editors restored Artemis endonuclease activity in a 293T^{Artemis−/−} system, with no detectable off-target effects at the predicted sites as assessed by targeted deep sequencing. These findings provide proof-of-concept that base editing can correct ART-SCID-associated mutations, highlighting its potential for future therapeutic development.

Keywords

Artemis / ART-SCID / Gene editing / CBE / ABE

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Funding

Guangdong S&T Program(2024B1111130003)

National Natural Science Foundation of China(32170888)

RIGHTS & PERMISSIONS

The Author(s)