Chimeric antigen receptor (CAR) T cells are modified T cells that have been genetically engineered to recognize and kill any cancer cell expressing a specific cell surface tumor antigen. Despite the remarkable success of CAR T cell technologies to date, adoption of the technologies has been limited by severe side-effects such as a potentially fatal cytokine release syndromes (CRS), neurologic toxicities, and on-target off-tumor toxicities, etc. Moreover, chronic exposure of CAR T cells to tumor antigen can lead to CAR T cell exhaustion, which results in inactivation of the CAR T cell therapy. Therefore, an ability to sensitively control CAR T cell fate in vivo, either to suppress its activity when confronted with a cytokine storm, or to rejuvenate its activity when encountering CAR T cell exhaustion, would be highly desirable.
We have developed a CAR T cell regulatory platform that enables delivery of any desired modulator selectively into an engineered CAR T cell without simultaneously permitting any modulator uptake by other cells. For this purpose, a fusion receptor that contains two subdomains, a ligand binding module and a membrane-anchoring/internalizing module, is co-expressed with the classical CAR construct in the desired T cell. In the embodiment described here, the ligand binding domain is comprised of human FK506 binding protein (FKBP), while the membrane-anchoring/internalizing domain is comprised of sequences from human folate receptor alpha (FRα). Because the fusion construct is human, it is not expected to be highly immunogenic. Moreover, because FRα is a GPI-anchored cell surface protein that constitutively internalizes, the fusion construct is predicted to constantly enter cells by endocytosis throughout the lifespan of the CAR T cell.
We show here that this fusion construct can be co-expressed with an anti-CD19 CAR in the same human T cell with a T2A linker in between in one lentiviral vector. We then demonstrate that any CAR T cell modulator can be selectively targeted to CAR T cells in vivo by linking the desired modulator to FK506; i.e. the ligand that binds FKBP. Upon binding FKBP, the FK506-modulator conjugate is shown to rapidly internalize into the targeted CAR T cell via standard FRα endocytosis. Furthermore, when the linker connecting FK506 to the desired modulator is designed to be cleaved within early endosomes, the modulator is demonstrated to be released following entry into the CAR T cell, where it becomes immediately available for modulation of CAR T cell properties. In the panels shown in this poster, examples of the use of this universal CAR T cell regulatory strategy to control the activities of CAR T cells following their infusion into animal models are provided.
Citation Format: Boning Zhang, John V Napoleon, Philip Low. Systemic Control of CAR T Cell Activity Using a Secret Passageway Fusion Receptor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2316.