This study develops ways to treat patients with inborn T cell defects by correcting their malfunctioning T cells using genomic scissors. The scissors need to be guided to the defective gene by a tag called guide RNA (gRNA). The gRNA prioritization for each patient is laborious and time-consuming. We will use different screening and computational techniques to streamline the personalized gRNA selection, in order to develop gene correction options for a larger number of patients.
Primary immunodeficiencies (PIDDs) are genetic diseases of the white blood cells. The patients present with a constellation of infection susceptibility, autoimmune disease, and early cancer. PIDDs are caused by mutations in more than 300 genes. The individual mutations are ultra-rare, and two patients with the same mutation can present with vastly different conditions.
We hypothesize that PIDD patients with defective T cell function can be managed by correcting the mutation in the T cells and infusing the cells back as an autologous transplant. Particularly the patients with life-threatening viral and fungal infections would benefit, but even other T cell related diseases, such as certain autoimmune diseases and viral-induced cancers, could respond. T cell infusion requires only a mild lymphodepleting pre-treatment, which makes the procedure safe and applicable to ill patients.
The T cell mutations can be corrected by CRISPR-Cas9 gene editing. The therapy requires the design of guide RNAs (gRNAs) that lead the CRISPR editing complex to the mutant locus. The gRNAs are designed computationally, but the current softwares do not predict mutation-specific gRNAs with sufficient accuracy. In this study, we propose to develop an improved, clinically applicable gRNA design software for personalized gene therapy. We will evaluate a set of gRNAs for on- and off-target efficiency in patient T cells, as well as in pooled and arrayed screening conditions. We will then use the resulting data to refine the existing gRNA design algorithms.
In parallel to software development, we will scale up the autologous T cell editing protocol for automatized good manufacturing practice (GMP)-level pipeline. We will study the genomic integrity of the corrected T cell products and transplant them to mice to evaluate the safety of the procedure.
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