Modeling Human Severe Combined Immunodeficiency and Correction by CRISPR/Cas9-Enhanced Gene Targeting
Chia-Wei Chang,
Yi-Shin Lai,
Erik Westin,
Alireza Khodadadi-Jamayran,
Kevin M. Pawlik,
Lawrence S. Lamb Jr.,
Frederick D. Goldman,
Tim M. Townes
Affiliations
Chia-Wei Chang
Department of Biochemistry and Molecular Genetics, Schools of Medicine and Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294, USA
Yi-Shin Lai
Department of Biochemistry and Molecular Genetics, Schools of Medicine and Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294, USA
Erik Westin
Department of Biochemistry and Molecular Genetics, Schools of Medicine and Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294, USA
Alireza Khodadadi-Jamayran
Department of Biochemistry and Molecular Genetics, Schools of Medicine and Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294, USA
Kevin M. Pawlik
Department of Biochemistry and Molecular Genetics, Schools of Medicine and Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294, USA
Lawrence S. Lamb Jr.
Department of Medicine, Division of Hematology/Oncology, Schools of Medicine and Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294, USA
Frederick D. Goldman
Department of Pediatrics, Division of Hematology/Oncology, Schools of Medicine and Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294, USA
Tim M. Townes
Department of Biochemistry and Molecular Genetics, Schools of Medicine and Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294, USA
Mutations of the Janus family kinase JAK3 gene cause severe combined immunodeficiency (SCID). JAK3 deficiency in humans is characterized by the absence of circulating T cells and natural killer (NK) cells with normal numbers of poorly functioning B cells (T–B+NK–). Using SCID patient-specific induced pluripotent stem cells (iPSCs) and a T cell in vitro differentiation system, we demonstrate a complete block in early T cell development of JAK3-deficient cells. Correction of the JAK3 mutation by CRISPR/Cas9-enhanced gene targeting restores normal T cell development, including the production of mature T cell populations with a broad T cell receptor (TCR) repertoire. Whole-genome sequencing of corrected cells demonstrates no CRISPR/Cas9 off-target modifications. These studies describe an approach for the study of human lymphopoiesis and provide a foundation for gene correction therapy in humans with immunodeficiencies.
