Wiskott-Aldrich Syndrome (WAS) is a rare X-linked primary immunodeficiency affecting approximately 1 in 100,000 live XY births in North America and is caused by a mutation to the WAS gene which is expressed across hematopoietic lineages. The WAS protein (WASp) plays a role in regulating actin polymerization. On a cellular level, there are a variety of effects of a lack of WASp or expression of a dysfunctional WASp protein for patients including issues with migration, adhesion, chemotactic response, phagocytosis, activation, and proliferation across different cell types in addition to reduced platelet size and output. This can lead to several systematic effects for the patients however because mutations to the WAS gene are not limited to one location or type there is a great amount of variability between patient symptoms making it challenging to diagnose. Major symptoms include frequent and recurrent infections, uncontrolled bleeding episodes, issues associated with autoimmunity, and malignancy, the most common form being lymphoma. Without treatment, the life expectancy of an individual diagnosed with WAS is 14 years of age, and the only curative treatment strategy available is hematopoietic stem and progenitor cell transfer (HSPCT). If not performed with an HLA-identical donor, which is available to less than 10% of patients, and within the first two years of life, the risk of graft versus host disease (GvHD) increases drastically for the patient. A gene therapy using autologous and genetically corrected CD34+ cells would be advantageous to the patients due to a reduction in preparative conditioning, immunosuppressive aftercare, and the risk of GvHD. CSL Behring is currently in the development of a lentiviral gene therapy to fulfill this gap in care, however, to develop the assays required to assess and characterize the drug substance usually an uncorrected patient sample is compared with a gene-edited sample. The limitation here is that due to the risk of infection and bleeding patient sample is very limited and therefore the development of a mock patient sample is necessary for early development. The goal of the project is to develop a WAS-KO protocol utilizing CRISPR/Cas9 and its characterization.
| Identifer | oai:union.ndltd.org:CALPOLY/oai:digitalcommons.calpoly.edu:theses-4389 |
| Date | 01 June 2023 |
| Creators | Hanna, Julia C |
| Publisher | DigitalCommons@CalPoly |
| Source Sets | California Polytechnic State University |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Master's Theses |
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