CD45RO^-CD8⁺ T cell-derived exosomes restrict estrogen-driven endometrial cancer development via the ERβ/miR-765/PLP2/Notch axis

Abstract

<b>Rationale:</b> Estrogen-dependent cancers (e.g., breast, endometrial, and ovarian cancers) are among the leading causes of morbidity and mortality in women worldwide. Recently, exosomes released by tumor-infiltrating CD8⁺ T cells have been under the spotlight in the field of cancer immunotherapy. Our study aims at elucidating the underlying mechanisms of the crosstalk between estrogen signaling and CD8⁺ T cells, and possible intervention values in uterine corpus endometrial cancer (UCEC). <b>Methods:</b> Micro RNA-seq was conducted to screen differentially expressed micro RNA in UCEC. Bioinformatic analysis was processed to predict the target of miR-765. RNA silencing or overexpressing and pharmacologic inhibitors were used to assess the functions of ERβ/miR-765/PLP2/Notch axis in UCEC cell proliferation and invasion <i>in vivo</i> and <i>in vitro</i>. <i>In vivo</i> imaging was performed to evaluate the metastasis of tumor in mice. Combined fluorescent <i>in situ</i> hybridization for miR-765 and immunofluorescent labeling for CD8 was carried out to prove the co-localization between miR-765 and CD8⁺ T cells. Exosomes derived from CD45RO^-CD8⁺ T cells were isolated to detect the regulatory effects on UCEC. <b>Results:</b> miR-765 is characterized as the most downregulated miRNA in UCEC, and there is a negative correlation between miR-765 and Proteolipid protein 2 (PLP2) in UCEC lesion. Estrogen significantly down-regulates miR-765 level, and facilitates the development of UCEC by estrogen receptor (ER) β. Mechanistically, this process is mediated through the miRNAs (e.g., miR-3584-5p, miR-7-5p, miR-150-5p, and miR-124-3p) cluster-controlled regulation of the PLP2, which further regulates Ki-67 and multiple epithelial-mesenchymal transition (EMT)-related molecules (e.g, E-cadherin and Vimentin) in a Notch signaling pathway-dependent manner. Interestingly, the selective ER degrader Fulvestrant alleviates estrogen-mediated miR-765/PLP2 expression regulation and UCEC development in ERβ-dependent and -independent manners. Additionally, CD45RO^-CD8⁺ T cell-derived exosomes release more miR-765 than that from CD45RO⁺CD8⁺ T cells. In therapeutic studies, these exosomes limit estrogen-driven disease development via regulation of the miR-765/PLP2 axis. <b>Conclusions:</b> This observation reveals novel molecular mechanisms underlying estrogen signaling and CD8⁺ T cell-released exosomes in UCEC development, and provides a potential therapeutic strategy for UCEC patients with aberrant ERβ/miR-765/PLP2/Notch signaling axis.