Development of Chimeric Antigen Receptor T Cells Against T Cell Lymphoma and Leukemia Using CRISPR/Cas9 Gene-editing

T cell neoplasms include T-cell acute lymphoblastic leukemia (T-ALL) and T-cell non-Hodgkin lymphomas (T-NHL) and are characterized by an overall very poor prognosis. With the exception of allogeneic stem cell transplantation, once relapsed there are no effective treatments for these diseases and developing novel definitive immunotherapies will represent a vertical advancement for the field.

Goals:

• To develop chimeric antigen receptor (CAR) T cells for the treatment of T cell derived neoplasms, specifically including T-cell acute lymphoblastic leukemia (T-ALL) and T-cell non-Hodgkin lymphomas (T-NHL)
• Reduce the toxicity of targeting normal T cells by engineering the normal immune system to be resistant to CART killing.
• Develop a method to reconstituting the host immune system with CART-resistant normal T cells generated by CRISPR/Cas9 KO of normal T cells (temporary T cell reconstitution) or hemopoietic CD34+ stem cells (long-term T cell reconstitution).

Accomplishments:

• Five different designed CART2 constructs and six different CART5 have been constructed.
• CRISPR-Cas9 gRNA for CD2 and CD5 have been designed and validated.
• Several T cell expansion protocols have been optimized for manufacturing of CD2 KO CART2 and CD5 KO CART5.
• CART2 and CART5 have been tested in vitro in a killing assay using the T-ALL cell line Jurkat.

Project Participants:  Carl June, Marco Ruella