Circular RNAs are a class of regulatory RNA transcripts, which are ubiquitously expressed in eukaryotes. In the current study, we evaluate the function of a novel circRNA derived from the β-catenin gene locus, circβ-catenin. Circβ-catenin is predominantly localized in the cytoplasm and displays resistance to RNase-R treatment. We find that circβ-catenin is highly expressed in liver cancer tissues when compared to adjacent normal tissues. Silencing of circβ-catenin significantly suppresses malignant phenotypes in vitro and in vivo, and knockdown of this circRNA reduces the protein level of β-catenin without affecting its mRNA level. We show that circβ-catenin affects a wide spectrum of Wnt pathway-related genes, and furthermore, circβ-catenin produces a novel 370-amino acid β-catenin isoform that uses the start codon as the linear β-catenin mRNA transcript and translation is terminated at a new stop codon created by circularization. We find that this novel isoform can stabilize full-length β-catenin by antagonizing GSK3β-induced β-catenin phosphorylation and degradation, leading to activation of the Wnt pathway. Our findings illustrate a non-canonical function of circRNA in modulating liver cancer cell growth through the Wnt pathway, which can provide novel mechanistic insights into the underlying mechanisms of hepatocellular carcinoma.

Emerging evidence indicate that cancer-associated fibroblasts (CAFs) affect tumor progression by reshaping the tumor microenvironment. Neutrophils are prominent components of solid tumors and important in cancer progression. Whether the phenotype and function of neutrophils in hepatocellular carcinoma (HCC) are influenced by CAFs is not well understood. Herein, we investigated the effect of HCC-derived CAFs (HCC-CAFs) on the neutrophils and explored the biological role of this effect. We found that HCC-CAFs induced chemotaxis of neutrophils and protected them from spontaneous apoptosis. Neutrophils were activated by the conditioned medium from HCC-CAFs with increased expression of CD66b, PDL1, IL8, TNFa, and CCL2, and with decreased expression of CD62L. HCC-CAF-primed neutrophils impaired T-cell function through the PD1/PDL1 signaling pathway. We revealed that HCC-CAFs induced the activation of STAT3 pathways in neutrophils, which are essential for the survival and function of activated neutrophils. In addition, we demonstrated that HCC-CAF-derived IL6 was responsible for the STAT3 activation of neutrophils. Collectively, our results suggest that HCC-CAFs regulate the survival, activation, and function of neutrophils within HCC through an IL6–STAT3–PDL1 signaling cascade, which presents a novel mechanism for the role of CAFs in remodeling the cancer niche and provides a potential target for HCC therapy.

Although carcinoma-associated fibroblasts (CAFs) in tumor microenvironments have a critical role in immune cell modulation, their effects on the generation of regulatory dendritic cells (DCs) are still unclear. In this study, we initially show that CAFs derived from hepatocellular carcinoma (HCC) tumors facilitate the generation of regulatory DCs, which are characterized by low expression of costimulatory molecules, high suppressive cytokines production and enhanced regulation of immune responses, including T-cell proliferation impairment and promotion of regulatory T-cell (Treg) expansion via indoleamine 2,3-dioxygenase (IDO) upregulation. Our findings also indicate that STAT3 activation in DCs, as mediated by CAF-derived interleukin (IL)-6, is essential to IDO production. Moreover, IDO inhibitor, STAT3 and IL-6 blocking antibodies can reverse this hepatic CAF-DC regulatory function. Therefore, our results provide new insights into the mechanisms by which CAFs induce tumor immune escape as well as a novel cancer immunotherapeutic approach (for example, targeting CAFs, IDO or IL-6).

The role of exosomal circular RNAs (circRNAs) in Hepatocellular carcinoma (HCC) cells with high metastatic potential has been little studied. Exosomal circRNA from cells with non-metastatic (HepG2), low metastatic (97L), and high metastatic (LM3) potential were sequencing. Metastatic-related circRNAs in serum from HCC patients were measured and their association with clinical prognosis was evaluated. Furthermore, candidate functional circRNAs in LM3-derived exosomes was assessed. LM3 exosomes enhanced the cell migration and invasion potential of HepG2 and 97 L cells. CircPTGR1, a circRNA with three isoforms, was specifically expressed in exosomes from 97 L and LM3 cells, upregulated in serum exosomes from HCC patients and was associated with the clinical stage and prognosis. Knockdown of circPTGR1 expression suppressed the migration and invasion of HepG2 and 97L cells induced by co-culturing with LM3 exosomes. Bioinformatics, co-expression analysis, and a luciferase assay indicated that circPTGR1 competed with MET to target miR449a. Higher metastatic HCC cells can confer this potential on those with lower or no metastatic potential via exosomes with circPTGR1, resulting in increased migratory and invasive abilities in those cells. FUND: National Natural Science Foundation of China (No. 81470870, 81670601, 81570593), Guangdong Natural Science Foundation (No. 2015A030312013, 2015A030313038), Sci-tech Research Development Program of Guangdong Province (2014B020228003), Sci-tech Research Development Program of Guangzhou City (No. 201508020262, 201400000001-3, 201604020001, 201607010024), Innovative Funds for Small and Medium-Sized Enterprises of Guangdong Province (2016A010119103), Pearl River S&T Nova Program of Guangzhou (201710010178), and National 13th Five-Year Science and Technology Plan Major Projects of China (No. 2017ZX10203205-006-001).

Purpose: Elevated levels of neutrophils have been associated with poor survival in various cancers, but direct evidence supporting a role for neutrophils in the immunopathogenesis of human cancers is lacking. Experimental Design: A total of 573 patients with gastric cancer were enrolled in this study. Immunohistochemistry and real-time PCR were performed to analyze the distribution and clinical relevance of neutrophils in different microanatomic regions. The regulation and function of neutrophils were assessed both in vitro and in vivo. Results: Increased neutrophil counts in the peripheral blood were associated with poor prognosis in gastric cancer patients. In gastric cancer tissues, neutrophils were enriched predominantly in the invasive margin, and neutrophil levels were a powerful predictor of poor survival in patients with gastric cancer. IL17+ neutrophils constitute a large portion of IL17-producing cells in human gastric cancer. Proinflammatory IL17 is a critical mediator of the recruitment of neutrophils into the invasive margin by CXC chemokines. Moreover, neutrophils at the invasive margin were a major source of matrix metalloproteinase-9, a secreted protein that stimulates proangiogenic activity in gastric cancer cells. Accordingly, high levels of infiltrated neutrophils at the invasive margin were positively correlated with angiogenesis progression in patients with gastric cancer. Conclusions: These data provide direct evidence supporting the pivotal role of neutrophils in gastric cancer progression and reveal a novel immune escape mechanism involving fine-tuned collaborative action between cancer cells and immune cells in the distinct tumor microenvironment. Clin Cancer Res; 23(6); 1575–85. ©2016 AACR.