CAR T cells targeting α_vβ₃integrin are effective against advanced cancer in preclinical models

Abstract

Integrins are heterodimeric receptors that convey cell-to-cell and cell-to-matrix interactions. Integrin αvβ3 is expressed in several tumor entities including melanoma, glioblastoma, breast, pancreatic, and prostate cancer, where it promotes tumor cell survival and metastasis. Here, we generated αvβ3-specific chimeric antigen receptor (CAR) T cells and analyzed their antitumor function in preclinical models in vitro and in vivo. αvβ3-CARs comprising a super-humanized hLM609 targeting domain with either high or low affinity (hLM609v7, Kd = 3 nM vs hLM609v11, Kd = 160 nM) and equipped with either a long or a short IgG4-Fc extracellular spacer (229 vs 12 amino acids) were expressed in CD8+ and CD4+ T cells through lentiviral transduction. αvβ3-CAR T cells eliminated αvβ3-positive tumor cells rapidly and specifically, produced IFN-γ and IL-2 (CD4+ > CD8+) and exhibited productive proliferation. In vitro, we observed the strongest reactivity with the higher affinity hLM609v7 αvβ3-CAR in the short spacer configuration, consistent with the tumor membrane-distal localization of the hLM609 epitope. In a murine xenograft model of metastatic A-375 melanoma, the strongest antitumor effect was mediated by the lower affinity hLM609v11 αvβ3-CAR. Notably, a single administration of hLM609v11 αvβ3-CAR T cells was able to induce complete elimination of melanoma lesions, leading to long-term tumor-free survival. These data establish αvβ3 integrin as a novel target for CAR T-cell immunotherapy and affirm our previous notion that binding domain affinity and spacer length can be calibrated to augment CAR reactivity. αvβ3-CAR T cells have therapeutic potential in several prevalent solid tumors, including melanoma and triple-negative breast cancer.