Membrane-proximal external region is a superior target for mediating effector activity of HIV-1 specific chimeric antigen receptor modified T cells

Abstract

17The use of chimeric antigen receptor modified-T (CAR-T) cells in adoptive 18 immunotherapy has been popularized through recent success in the field of cancer 19 treatment research. CD4ζ CAR, which targets HIV-1-infected cells, has been developed 20 and evaluated in patients. Though well-tolerated for over a decade, efficacy was 21 disappointingly limited. This result encourages us to develop a novel CAR more effective 22 than CD4ζ CAR. To quantitatively compare anti-HIV-1 activity of different CAR constructs 23 in a highly sensitive and reproducible manner, we developed a multicolor flow cytometry 24 method for assessing anti-HIV-1 effector T-cell activity. "Target" Jurkat cells inducibly 25 expressing an HIV-1 HXBC2 envelope protein and "Non-target" control cells were genetically 26 labeled with red and blue fluorescent protein, respectively, and co-incubated with human 27 primary T cells transduced with anti-HIV-1 "Effector" CARs at various Effector vs Target 28 cell ratios. Absolute cell numbers of each population were collected by MACSQuant 29 Analyzer and used for calculation of relative cytotoxicity. We successfully ranked the