In vitro and animal model studies conducted by scientists at the University College London (UCL) have demonstrated the feasibility of a gene therapy approach to correct the deficiency of a regulatory protein, CTLA4, in immune T lympthocytes, that results in a rare inborn error of immunity. The study, published in Science Translational Medicine, could lead to a new treatment for CTLA4 insufficiency that affects regulatory and effector T cells, resulting in autoimmunity and frequently severe clinical phenotype.

CTLA4 insufficiency occurs in individuals with only one functional copy of CTLA4 who produce too little of the protein to adequately regulate the immune system. The condition also decreases immune memory, such that patients suffer from repeated infections by the same pathogens and may lead to a type of blood cancer (lymphomas).

In this study, the researchers targeted the faulty gene in primary T cells collected from patients with CTLA-4 insufficiency. Using homology-directed repair (HDR) CRISPR/Cas9 gene editing, they inserted the normal CTLA-4 complementary DNA sequence into the first intron (noncoding region) of the CTLA-4 genomic locus. This restored the levels of CTLA-4 in the patient-derived cells to those seen in healthy T cells and rescued the normal function of the cells in vitro.

“Genes that play critical roles in controlling immune responses are not switched on all the time and are very tightly regulated,” said Emma Morris, PhD, a professor of clinical cell and gene therapy, director of UCL’s Division of Infection and Immunity, and co-senior author of this study. “The technique we have used allows us to leave the natural (endogenous) mechanisms controlling gene expression intact, at the same time as correcting the mistake in the gene itself.”

The team also conducted experiments on mouse models of the disease that do not express CTLA-4 in any cell. The researchers grafted T cells with normal CTL4 expression into these mice and demonstrated that it prevented abnormal expansion of T cell populations (lymphoproliferation) in vivo.

“It’s really exciting to think about taking this treatment forward to patients. If we can improve their symptoms and reduce their risk of getting lymphoproliferative disease this will be a major step forward,” said Claire Booth, PhD, a professor of gene therapy at UCL and co-senior author of the study. “This paper is important because we are using the newest gene editing techniques to precisely correct these T cells, which is a new approach in inborn errors of immunity.”

At present, CTLA-4 insufficiency is treated using bone marrow transplantation that replaces stem cells responsible for producing T cells. Transplantation, however, is accompanied by risks of rejection, extended immunosuppression therapy, infections, and long hospital visits.

“Correcting the patient’s T cells can improve many of the symptoms of the disease, at the same time as being much less toxic than a bone marrow transplant,” said Booth. “Collecting and correcting T cells is easier. With this approach, the amount of time in hospital the patients would need would be far less.”

The study was led by Thomas Fox, PhD, a Wellcome Trust Clinical PhD Fellow at UCL, and built on work by co-author Pietro Genovese, PhD, researcher at the Dana-Farber Cancer Institute.

The researchers claim the gene editing therapy they have developed in this proof-of-principle study could be adapted to tackle other conditions. “It allows us to correct genes that are dysregulated or overactive, but also allows us to understand much more about gene expression and gene regulation,” said Morris.

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