Phenotypic models of CAR-T cell activation elucidate the pivotal regulatory role of CAR down-modulation

Abstract

Adoptive cell immunotherapy with chimeric antigen receptor (CAR) showed limited potency in solid tumors, despite durable remissions for hematopoietic malignancies. Therefore, an investigation of ways to enhance the efficacy of CARs anti-tumor response have been engaged upon. We previously examined the interplay between the biophysical parameters of CAR binding, i.e. affinity, avidity and antigen density, as regulators of CAR-T cell activity, and detected nonmonotonic behaviors of affinity and antigen density, and an interrelation between avidity and antigen density. Here we built an evolving phenotypic model of CAR-T cell regulation, which suggested that receptor down-modulation is a key determinant of CAR-T cell function. We verified this assumption by measuring and manipulating receptor down-modulation and intracellular signaling processes. Indeed, CAR down-modulation inhibition, via actin polymerization inhibition, but not inhibition of regulatory inhibitory phosphatases, was able to increase CAR-T cell responses. Additionally, we documented trogocytosis in CAR-T cells that depends on actin polymerization. In summary, our study modeled the parameters that govern CAR-T cell engagement and revealed an under-appreciated mechanism of T cell regulation. These results have a potential to predict and therefore advance the rational design of CAR-T cell for adoptive treatments.