Cancer-secreted exosomal miRNAs are emerging mediators of cancer-stromal cross-talk in the tumor environment. Our previous miRNAs array of cervical squamous cell carcinoma (CSCC) clinical specimens identified upregulation of miR-221-3p. Here, we show that miR-221-3p is closely correlated with peritumoral lymphangiogenesis and lymph node (LN) metastasis in CSCC. More importantly, miR-221-3p is characteristically enriched in and transferred by CSCC-secreted exosomes into human lymphatic endothelial cells (HLECs) to promote HLECs migration and tube formation in vitro, and facilitate lymphangiogenesis and LN metastasis in vivo according to both gain-of-function and loss-of-function experiments. Furthermore, we identify vasohibin-1 (VASH1) as a novel direct target of miR-221-3p through bioinformatic target prediction and luciferase reporter assay. Re-expression and knockdown of VASH1 could respectively rescue and simulate the effects induced by exosomal miR-221-3p. Importantly, the miR-221-3p-VASH1 axis activates the ERK/AKT pathway in HLECs independent of VEGF-C. Finally, circulating exosomal miR-221-3p levels also have biological function in promoting HLECs sprouting in vitro and are closely associated with tumor miR-221-3p expression, lymphatic VASH1 expression, lymphangiogenesis, and LN metastasis in CSCC patients. In conclusion, CSCC-secreted exosomal miR-221-3p transfers into HLECs to promote lymphangiogenesis and lymphatic metastasis via downregulation of VASH1 and may represent a novel diagnostic biomarker and therapeutic target for metastatic CSCC patients in early stages.

The accumulation of tumour-associated macrophages (TAMs) in the hypoxic tumour microenvironment (TME) is associated with malignant progression in cancer. However, the mechanisms by which the hypoxic TME facilitates TAM infiltration are not fully understood. This study showed that high ZEB1 expression in hypoxic cervical cancer cell islets was positively correlated with CD163⁺ TAM accumulation. ZEB1 in hypoxic cancer cells promoted the migration of TAMs in vitro and altered the expression of multiple chemokines, especially CCL8. Mechanistically, hypoxia-induced ZEB1 activated the transcription of CCL8, which attracted macrophages via the CCR2–NF-κB pathway. Furthermore, ZEB1 and CCL8 were independent prognostic factors in cervical cancer patients based on The Cancer Genome Atlas (TCGA) data analysis. In conclusion, hypoxia-induced ZEB1 exerts unexpected functions in cancer progression by fostering a prometastatic environment through increased CCL8 secretion and TAM recruitment; thus, ZEB1 may serve as a candidate biomarker of tumour progression and provide a potential target for disrupting hypoxia-mediated TME remodelling.

Cancer-associated lymphatic endothelial cells (LECs) are an active barrier to the effector arm of the anti-tumor immune response; however, it remains unclear how LECs become immunosuppressive in the tumor microenvironment (TME). Exosomal miRNAs have recently been implicated in intercellular crosstalk within the TME. Here we report a mechanistic model via which cervical cancer-secreted, exosome-encapsulated miR-1468-5p promotes lymphatic PD-L1 upregulation and lymphangiogenesis to impair T cell immunity. Subsequently, exosomal miR-1468-5p epigenetically activates the JAK2/STAT3 pathway in LECs by directly targeting HMBOX1 in the SOCS1 promoter, activating an immunosuppressive program that allows cancer cells to escape anti-cancer immunity. Furthermore, clinical data reveal that high serum exosomal miR-1468-5p levels correlate with TME immunosuppressive status and poor prognosis in cervical cancer (CCa) patients. Taken together, our results suggest that cancer-secreted exosomal miR-1468-5p instructs LECs to form an integrated immunosuppressive TME component and may be a prognostic biomarker and therapeutic target for CCa.

Clinical response to immunotherapy is closely associated with the immunosuppressive tumour microenvironment (TME), and influenced by the dynamic interaction between tumour cells and lymphatic endothelial cells (LECs). Here, we show that high levels of miR-142-5p positively correlate with indoleamine 2,3-dioxygenase (IDO) expression in tumour-associated lymphatic vessels in advanced cervical squamous cell carcinoma (CSCC). The miR-142-5p is transferred by CSCC-secreted exosomes into LECs to exhaust CD8⁺ T cells via the up-regulation of lymphatic IDO expression, which was abrogated by an IDO inhibitor. Mechanistically, miR-142-5p directly down-regulates lymphatic AT-rich interactive domain-containing protein 2 (ARID2) expression, inhibits DNA methyltransferase 1 (DNMT1) recruitment to interferon (IFN)-γ promoter, and enhances IFN-γ transcription by suppressing promoter methylation, thereby leading to elevated IDO activity. Furthermore, increased serum exosomal miR-142-5p levels and the consequent IDO activity positively correlate with CSCC progression. In conclusion, exosomes secreted by CSCC cells deliver miR-142-5p to LECs and induce IDO expression via ARID2–DNMT1–IFN-γ signalling to suppress and exhaust CD8⁺ T cells. Our study suggests that LECs act as an integral component of the immune checkpoint(s) in the TME and may serve as a potential new target for CSCC diagnosis and treatment.