Chimeric antigen receptor (CAR) T-cell therapy has shown remarkable success in the treatment of hematologic malignancies. Unfortunately, it has limited efficacy against solid tumors, even when the targeted antigens are well expressed. A better understanding of the underlying mechanisms of CAR T-cell therapy resistance in solid tumors is necessary to develop strategies to improve efficacy. Here we report that solid tumors release small extracellular vesicles (sEV) that carry both targeted tumor antigens and the immune checkpoint protein PD-L1. These sEVs acted as cell-free functional units to preferentially interact with cognate CAR T cells and efficiently inhibited their proliferation, migration, and function. In syngeneic mouse tumor models, blocking tumor sEV secretion not only boosted the infiltration and antitumor activity of CAR T cells but also improved endogenous antitumor immunity. These results suggest that solid tumors use sEVs as an active defense mechanism to resist CAR T cells and implicate tumor sEVs as a potential therapeutic target to optimize CAR T-cell therapy against solid tumors. Significance: Small extracellular vesicles secreted by solid tumors inhibit CAR T cells, which provide a molecular explanation for CAR T-cell resistance and suggests that strategies targeting exosome secretion may enhance CAR T-cell efficacy. See related commentary by Ortiz-Espinosa and Srivastava, p. 2637

Small extracellular vesicles (sEVs) are nanosized vesicles. Death receptor 5 (DR5) mediates extrinsic apoptosis. We engineer DR5 agonistic single-chain variable fragment (scFv) expression on the surface of sEVs derived from natural killer cells. PDGFR transmembrane domain delivers DR5-scFvs to the surface of sEVs. DR5-scFv sEVs rapidly induce apoptosis of different types of DR5 ⁺ cancer cells, myeloid-derived suppressor cells (MDSCs), and cancer-associated fibroblasts (CAFs). DR5-scFv sEVs migrate specifically to DR5 ⁺ tumors in vitro and in vivo. Systemic delivery of DR5-scFv sEVs significantly inhibits the growth of DR5 ⁺ melanoma, liver cancer, and breast cancer and prolongs mouse life span without significant toxicity. DR5-scFv sEVs are significantly more efficacious than DR5 antibodies in vivo. In organotypic patient-derived melanoma slice cultures, DR5-scFv sEVs effectively inhibit melanoma cells and MDSCs and activate CD8 ⁺ T cells. Our studies demonstrate that DR5-scFv sEVs can inhibit tumor growth by targeting tumor cells and immunosuppressive stromal cells in the TME.

Background: Ferroptosis is a programmed cell death caused by iron‐dependent accumulation and cellular lipid peroxides, which is different from apoptosis and pyroptosis. This study investigated the possible effect of ferroptotic response in the pathogenesis of infantile hemangioma (IH). Methods and Results: The staining level of 4‐hydroxynonenal (4‐HNE), the marker of ferroptotic cells, was significantly increased in the involutive IH samples compared with the proliferative samples (9 proliferative versus 12 involutive lesions, P =0.0152). By contrast, the expression of glutathione peroxidase 4 (GPX4), a key enzyme regulating ferroptotic resistance, was significantly increased in the involutive IH samples. Meanwhile, the GPX4 was richly expressed in macrophages of IH. The data from in vitro study showed that the mRNA ( P =0.0002) and protein ( P =0.0385) expression levels of GPX4 were significantly upregulated in macrophages cultured with hemangioma‐derived stem cells conditional medium (HemSC‐CM). Mechanistically, HemSC‐CM promoted the expression of GPX4 in macrophages ( P =0.0482) by increasing nuclear factor erythroid 2‐related factor 2 translocation to the nucleus ( P =0.0026). Additionally, inhibition of GPX4 or inducing ferroptosis in macrophages could inhibit progression of lesion in IH nude mice mode. CONCLUSIONS: Hemangioma‐derived stem cells (HemSCs) could promote macrophage ferroptotic resistance through upregulating expression of GPX4, which is required for the progression of IH.