CAR T cells targeting αvβ3 integrin are effective against advanced cancer in preclinical models

Abstract

Integrins are heterodimeric receptors that convey cell-to-cell and cell-to-matrix interactions. Integrin α_vβ₃ 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 α_vβ₃-specific chimeric antigen receptor (CAR) T cells and analyzed their antitumor function in preclinical models in vitro and in vivo. α_vβ₃-CARs comprising a super-humanized hLM609 targeting domain with either high or low affinity (hLM609v7, K_d = 3 nM vs hLM609v11, K_d = 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. α_vβ₃-CAR T cells eliminated α_vβ₃-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 α_vβ₃-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 α_vβ₃-CAR. Notably, a single administration of hLM609v11 α_vβ₃-CAR T cells was able to induce complete elimination of melanoma lesions, leading to long-term tumor-free survival. These data establish α_vβ₃ 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. α_vβ₃-CAR T cells have therapeutic potential in several prevalent solid tumors, including melanoma and triple-negative breast cancer.