Cancer is the leading cause of death in people living with HIV (PLWH) on combination antiretroviral therapy (cART) in the United States and worldwide. As we approach the fourth decade of the HIV/AIDS epidemic, the burden of one such cancer, HIV-related lymphoma, remains substantial. Diffuse large B cell lymphoma (DLBCL) is one of the most common HIV-associated malignancies, and represents a significant medical need. Two landmark trials have led to the FDA approval of two different CD19 directed chimeric antigen receptor (CAR) T cell (CAR19) products in the United States for the treatment of relapsed or refractory (RR) large B cell lymphoma after two or more lines of systemic therapy: tisagenlecleucel (Kymriah™) and axicabtagene ciloleucel (Yescarta ®). Unfortunately, in both trials, patients with known HIV infection were excluded. Therefore, PLWH in the U.S. are not able to receive this potentially curative treatment. γδ T cells represent an important arm of the innate immune system known to be associated with a favorable prognosis in cancers including DLBCL and in fighting HIV infection, but are also known to be susceptible to HIV infection and to represent a latent viral reservoir. PLWH uniformly have depletion of the Vδ2 subset that highly expresses the CCR5 receptor, with associated decreased responsiveness to infection and tumor antigens. This project proposes to explore the concept of engineering γδ T cells, particularly the key Vγ9Vδ2 subset, to target DLBCL by knocking a CAR19 construct into the CCR5 gene. By engineering cells to have a new property we are combining the principles of genome editing with synthetic biology. The resulting γδCAR19-CCR5KO cells would have the potential to recognize and kill CD19 expressing lymphoma and normal CD19 cells via the CAR19 receptor, and to kill lymphoma and HIVinfected cells via the endogenous γδ TCR and innate cytotoxic properties of γδ T cells. They should proliferate in response to activation through the CAR19, and since they would be resistant to HIV infection they should not be killed by HIV, nor act as a future harbor for latent HIV. These proposed studies will address the feasibility of engineering and expanding γδCAR19-CCR5KO cells in culture, and then testing their ability to kill lymphoma cells and possibly clear HIV infection in cell culture and mouse tumor models. The results of these translational investigations would be used to support development of this concept to pilot manufacturing process development, serve as IND-enabling studies, and lead to a clinical trial.