Background: Long non-coding RNAs (lncRNAs) possess significant regulatory functions in multiple biological and pathological processes, especially in cancer. Dysregulated lncRNAs in hepatocellular carcinoma (HCC) and their therapeutic applications remain unclear.Methods: Differentially expressed lncRNA profile in HCC was constructed using TCGA data. LINC00958 expression level was examined in HCC cell lines and tissues. Univariate and multivariate analyses were performed to demonstrate the prognostic value of LINC00958. Loss-of-function and gain-of-function experiments were used to assess the effects of LINC00958 on cell proliferation, motility, and lipogenesis. Patient-derived xenograft model was established for in vivo experiments. RNA immunoprecipitation, dual luciferase reporter, biotin-labeled miRNA pull-down, fluorescence in situ hybridization, and RNA sequencing assays were performed to elucidate the underlying molecular mechanisms. We developed a PLGA-based nanoplatform encapsulating LINC00958 siRNA and evaluated its superiority for systemic administration.Results: We identified a lipogenesis-related lncRNA, LINC00958, whose expression was upregulated in HCC cell lines and tissues. High LINC00958 level independently predicted poor overall survival. Functional assays showed that LINC00958 aggravated HCC malignant phenotypes in vitro and in vivo. Mechanistically, LINC00958 sponged miR-3619-5p to upregulate hepatoma-derived growth factor (HDGF) expression, thereby facilitating HCC lipogenesis and progression. METTL3-mediated N⁶-methyladenosine modification led to LINC00958 upregulation through stabilizing its RNA transcript. A PLGA-based nanoplatform loaded with si-LINC00958 was developed for HCC systemic administration. This novel drug delivery system was controlled release, tumor targeting, safe, and presented satisfactory antitumor efficacy.Conclusions: Our results delineate the clinical significance of LINC00958 in HCC and the regulatory mechanisms involved in HCC lipogenesis and progression, providing a novel prognostic indicator and promising nanotherapeutic target.

Background: Metastasis is a key feature of malignant tumors and significantly contributes to their high mortality, particularly in hepatocellular carcinoma (HCC). Therefore, it is imperative to explore the mechanism of tumor metastasis. Recently, tumor-associated macrophages (TAMs) have been demonstrated to promote tumor progression, while TAM-derived molecules involved in HCC metastasis warrant further investigation. Methods: THP-1 was treated with IL-4 (Interleukin-4) and IL-13 (Interleukin-13) for M2 polarized macrophages. Exosomes derived from M2 macrophages were characterized. Then, HCC cells or human umbilical vein endothelial cells (HUVECs) were co-cultured with M2 macrophages or treated with M2 macrophage-secreted exosomes. Next, Transwell®, Scratch assay, tube formation, and endothelial permeability assays were performed. Moreover, RT-PCR, western blotting, immunofluorescence, and ELISA were used to assess mRNA and protein expression levels. Finally, the miRNA expression profiles of exosomes derived from M2 and M0 macrophages were analyzed. Results: M2 macrophage infiltration was correlated with metastasis and a poor prognosis in HCC patients. M2-derived exosomes were absorbed by HCC and HUVEC cells and promoted the epithelial-mesenchymal transition (EMT), vascular permeability, and angiogenesis. Notably, MiR-23a-3p levels were significantly higher in M2-derived exosomes and hnRNPA1 mediated miR-23a-3p packaging into exosomes. Phosphatase and tensin homolog (PTEN) and tight junction protein 1 (TJP1) were the targets of miR-23a-3p, as confirmed by luciferase reporter assays. Lastly, HCC cells co-cultured with M2-derived exosomes secreted more GM-CSF, VEGF, G-CSF, MCP-1, and IL-4, which in turn further recruited M2 macrophages. Conclusions: Our findings suggest that M2 macrophage-derived miR-23a-3p enhances HCC metastasis by promoting EMT and angiogenesis, as well as increasing vascular permeability.

Background and Aims: The development and progression of hepatocellular carcinoma (HCC) is dependent on its local microenvironment. Tumor‐associated macrophages (TAMs) are deemed a key factor for the tumor microenvironment and attribute to contribute to tumor aggressiveness. However, the detailed mechanism underlying the pro‐metastatic effect of TAMs on HCC remains undefined. Approach and Results: The present study proved that TAMs were enriched in HCC. TAMs were characterized by an M2‐polarized phenotype and accelerated the migratory potential of HCC cells in vitro and in vivo. Furthermore, we found that M2‐derived exosomes induced TAM‐mediated pro‐migratory activity. With the use of mass spectrometry, we identified that integrin, α_Mβ₂ (CD11b/CD18), was notably specific and efficient in M2 macrophage–derived exosomes (M2 exos). Blocking either CD11b and/or CD18 elicited a significant decrease in M2 exos–mediated HCC cell metastasis. Mechanistically, M2 exos mediated an intercellular transfer of the CD11b/CD18, activating the matrix metalloproteinase‐9 signaling pathway in recipient HCC cells to support tumor migration. Conclusions: Collectively, the exosome‐mediated transfer of functional CD11b/CD18 protein from TAMs to tumor cells may have the potency to boost the migratory potential of HCC cells, thus providing insights into the mechanism of tumor metastasis.

Glucose metabolic reprogramming from oxidative to aerobic glycolysis, referred as the Warburg effect, is a hallmark of tumor cells. Accumulating evidence suggests that a subset of microRNAs play pivotal roles in modulating such reprogramming of glucose metabolism in cancer cells. miR-3662 has been implicated previously in both pro-tumorigenic and anti-tumorigenic effects in several types of cancer. The expression level of miR-3662 is downregulated in acute myeloid leukemia, whereas increased miR-3662 expression is observed in lung adenocarcinoma. However, the roles and underlying mechanisms of miR-3662 in hepatocellular carcinoma (HCC) metabolic reprogramming remain unclear. Our present study revealed that miR-3662 was frequently downregulated in HCC tissues and cell lines. The low expression level of miR-3662 was associated with tumor size, tumor multiplicity, Edmondson grade, and tumor-node-metastasis stage. Gain-of-function and loss-of-function assays showed that miR-3662 dampened glycolysis by reducing lactate production, glucose consumption, cellular glucose-6-phosphate level, ATP generation, and extracellular acidification rate, and increasing oxygen consumption rate in HCC cells after treatment with the hypoxia mimetic CoCl₂. Moreover, miR-3662 suppressed cell growth in vitro and in vivo, and induced G1/S cell cycle arrest. miR-3662 inhibited the activation of ERK and JNK signaling pathways in HCC. By combined computational and experimental approaches, hypoxia-inducible factor-1α (HIF-1α) was determined as a direct target of miR-3662. After treatment with the hypoxia mimetic CoCl₂, miR-3662 regulated the Warburg effect and HCC progression via decreasing HIF-1α expression. Our findings uncover a mechanistic role for miR-3662/HIF-1α axis in HCC metabolic reprogramming, providing a potential therapeutic strategy in liver cancer.

Background: Hypoxia is a hallmark of cancer, and is closely intertwined with tumor immune evasion. Circular RNAs (circRNAs) have been implicated in tumor response to immune checkpoint blockades. However, hypoxia-associated circRNAs that orchestrate the association between hypoxia and response to immunotherapy remain poorly understood. Here, we aimed to determine the roles of hypoxia-associated circRNAs in immune escape of hepatocellular carcinoma (HCC) cells. Methods: Differentially expressed hypoxia-associated circRNAs were determined using high-throughput sequencing technology. HCC patients treated with PD-1 blockade were enrolled to assess the clinical significance of circPRDM4. RT-qPCR, western blotting, flow cytometry, T cell-mediated tumor cell killing assay, and enzyme linked immunosorbent assay were used to investigate the roles of circPRDM4 in immune escape of HCC cells in vitro. Patient-derived xenograft mouse models and adoptive human tumor infiltrating lymphocyte-CD8⁺ T cell transfer were adopted to evaluate the effects of circPRDM4 in vivo. RNA pull-down, mass spectrometry, RNA immunoprecipitation, chromatin immunoprecipitation, chromatin isolation by RNA purification, dual-luciferase reporter assays, dot blotting, DNA in situ hybridization, and immunoprecipitation were utilized to examine the interaction between circPRDM4, HIF-1α, and CD274 promoter. Results: We identified circPRDM4 as a hypoxia-associated circRNA in HCC. circPRDM4 was upregulated in responders to PD-1 blockade and associated with therapeutic efficacy. In vitro and in vivo experiments showed that circPRDM4 induced PD-L1 expression and promoted CD8⁺ T cell-mediated immune escape under hypoxic conditions. Mechanistically, circPRDM4 acted as a scaffold to recruit HIF-1α onto CD274 promoter, and cemented their interaction, ultimately promoting the HIF-1α-mediated transactivation of PD-L1. Conclusions: These findings illustrated that circPRDM4 promoted immune escape of HCC cells by facilitating the recruitment of HIF-1α onto the promoter of CD274 under hypoxia, thereby inhibiting CD8⁺ T cell infiltration in the tumor microenvironment. This work may provide a novel prognostic biomarker and therapeutic candidate for HCC immunotherapy.