The International Journal of Biochemistry & Cell Biology

Long non-coding RNA CCAT1 is overexpressed in endometrial cancer and regulates growth and transcriptome of endometrial adenocarcinoma cells

Long non-coding RNAs (lncRNAs) play important roles in regulation of gene expression and are involved in pathogenesis of different diseases including cancer. Recent studies suggested the lncRNA Colon cancer associated transcript-1 (CCAT1) to act as putative oncogene. In this study, to elucidate the role of this lncRNA in endometrial cancer, we examined its expression in normal endometrium and type 1 endometrial cancer and knocked down its expression in endometrial cancer cell lines followed by transcriptome and pathway analyses. CCAT1 expression was examined in 100 tissue samples of normal endometrium and type 1 endometrial cancer tissues by means of RT-qPCR. Knockdown of CCAT1 expression in HEC-1B and RL95/2 endometrial cancer cells was performed by siRNA transfection. Affymetrix GeneChip arrays were used to elucidate the effect of both lncRNAs on the transcriptome of these cell lines. Median CCAT1 expression was found to be 9.3-fold higher in endometrial cancer when compared to normal endometrium (p < 0.05). In contrast to premenopausal endometrium and G1, G2 and G3 graded endometrial cancer, CCAT1 expression was nearly absent in postmenopausal tissue. Knockdown of CCAT1 by transient siRNA transfection significantly reduced proliferation of HEC-1B cancer cells in vitro by 35.5 % 6 days after transfection and notably reduced their colony formation ability. Affymetrix microarray and Ingenuity pathway analyses revealed a set of up- or down-regulated genes in transfected ERα-negative HEC-1B cells forming a network controlled by the key regulators TNF and TP53, including genes known to be involved in growth control, providing putative molecular mechanisms underlying the observed growth inhibition of HEC-1B cells. In contrast, CCAT1 knockdown in ERα-positive RL95/2 cells did not significantly affect proliferation, but resulted in down-regulation of a network of ERα target genes. Given that the lncRNA CCAT1 was found to be overexpressed in endometrial cancer, affected the growth of HEC-1B cells and the expression of growth regulatory genes, our data suggest CCAT1 to exert oncogenic functions in endometrial cancer and encourage further studies to examine to what extent this lncRNA might be a potential therapy target in this cancer entity.

PRMT6 promotes endometrial cancer via AKT/mTOR signaling and indicates poor prognosis

Arginine methylation plays essential roles in post-transcriptional modification and signal transduction. Dysregulation of protein arginine methyltransferases (PRMTs) has been reported in human cancers, yet the expression and biological function of PRMT6 in endometrial cancer (EMC) remains unclear. Here, we show that PRMT6 is upregulated in EMC and exhibits oncogenic activities via activation of AKT/mTOR pathway. The expression of PRMT6 in EMC is much higher than that in the adjacent nontumorous tissues. Elevated PRMT6 expression is significantly associated with higher histological tumor grade and unfavorable prognosis in two independent cohorts consisting of a total of 564 patients with EMC. In vitro data demonstrate that PRMT6 expression was identified as a downstream target of miR-372-3p. Ectopic expression of miR-372-3p downregulates PRMT6. Overexpression of PRMT6 promotes EMC cell proliferation and migration, whereas knockdown of PRMT6 leads to opposite phenotypes. Mechanistically, PRMT6 induces the phosphorylation of AKT and mTOR in EMC cells. Inhibition of AKT/mTOR signaling by MK2206 or rapamycin attenuates the PRMT6-mediated EMC progression. In clinical samples, high expression of PRMT6 was correlated to low expression of miR-372-3p and high expression of phosphorylated AKT. Collectively, our findings suggest PRMT6 may function as an oncogene to promote tumor progression, and be of prognostic value to predict disease-free survival of patients with EMC. The newly identified miR-372-3p/PRMT6/AKT/mTOR axis represents a new promising target for EMC management.

Xiaoyou decoction suppresses cervical precancerous lesions through the activation of cellular autophagy and the upregulation of p53 expression

Persistent high-risk human papilloma virus (HR-HPV) infection is a key factor in the progression of cervical lesions to cervical cancer. This study explores the molecular mechanisms through which the traditional Chinese medicine Xiaoyou Decoction (XYD) inhibits cervical intraepithelial neoplasia (CIN) lesions, offering new insights into its potential therapeutic application. Network pharmacology analysis was employed to identify the potential active ingredients and key target genes of XYD in treating CIN. Functional enrichment analysis was utilized to pinpoint the critical biological pathways affected by XYD. Clinical randomized trials were performed to evaluate the clinical efficacy of XYD. In vitro experiments were conducted to explore the functional effects and underlying molecular mechanisms of XYD. A total of 209 potential target genes of XYD associated with CIN lesions were identified. In addition, the active ingredients of XYD exhibited a strong association with autophagy-related proteins. Clinical randomized trials demonstrated that XYD treatment effectively alleviated HR-HPV infection, and after a 6-month follow-up, 90.3 % of patients exhibited negative conversion, successfully reversing the progression of CIN lesions. In vitro experiments confirmed that XYD inhibited CIN cell proliferation by activating the autophagy pathway and upregulating p53 protein expression. In conclusion, our study reveals that XYD effectively prevents the persistence of HR-HPV infection and reverses the progression of CIN lesions by activating the autophagy pathway and upregulating p53 expression. These findings provide preliminary insights into the biological effects and specific mechanisms of XYD in CIN, offering a novel perspective for treating persistent HR-HPV infections.

METTL5-mediated m6A modification of SLC7A11 promotes cervical cancer by inhibiting ferroptosis

Ferroptosis could suppress the viability of cervical cancer cells and trigger their death, thereby offering a unique perspective for exploring novel therapeutic approach for cervical cancer. Here, this study tried to explore the role of N

A platinum(II) complex HY1-Pt overcomes cisplatin-induced resistance and attenuates metastasis of epithelial ovarian cancer by cancer cell stemness inhibition

Tumor recurrence, acquired resistance and metastasis have severely limited the effect of clinical treatments for epithelial ovarian cancer. Recent researches reveal that cancer stem cells play important roles in the process of cisplatin-induced resistance and cancer cell metastasis. A platinum(II) complex (HY1-Pt) owning casein kinase 2 specificity reported in our recent research was herein applied to treat cisplatin-sensitive and cisplatin-resistant epithelial ovarian cancers, respectively, anticipating to achieve high anti-tumor efficacy. HY1-Pt showed highly efficient anti-tumor effect with low toxicity for either cisplatin-sensitive or cisplatin-resistant epithelial ovarian cancer both in vitro and in vivo. Biological studies indicated that HY1-Pt as a casein kinase 2 inhibitor could effectively overcome cisplatin resistance through the signaling pathway of Wnt/β-catenin by inhibiting expression of the signature genes of cancer stemness cells in A2780/CDDP cells. Moreover, HY1-Pt could suppress tumor migration and invasion in vitro and in vivo, further proving that HY1-Pt can be a potent novel platinum(II) agent for cisplatin-resistant epithelial ovarian cancer treatment.

Targeting Ku86 enhances X-ray-induced radiotherapy sensitivity in serous ovarian cancer cells

Drug resistance and recurrence are significant contributors to the poor prognosis of serous ovarian cancer (SOC). Radiotherapy is primarily used for the treatment of cancer recurrence; however, it is rarely applied in cases of SOC. Ku86, also known as XRCC5(X-ray repair cross complementing 5), has rarely been reported in the radiotherapy of SOC. Therefore, this study aimed to investigate the role of Ku86 in the development of SOC and in radiotherapy sensitivity induced by X-ray. In vitro experiments and database analysis showed significantly elevated expression of Ku86 in SOC. Further, after down-regulating Ku86 using RNAi technology, cell proliferation was inhibited. Further, the cell cycle was blocked in the G2 phase, and G2/G1 was increased since X-ray irradiation led to an increase in γ-H2AX. Down-regulation of Ku86 in the case of X-ray irradiation will promote the above biological effects, with more γ-H2AX and higher G2/G1. Here, we further deduce that Ku86 promotes the X-ray induced effect is most likely to activate the ATR pathway to block the cell cycle while inhibiting the NHEJ pathway to inhibit DNA damage response(DDR). Together these findings suggest that the down-regulation of Ku86 can improve X-ray-induced radiotherapy by affecting ATR and NHEJ pathways, and provide a new strategy for the treatment of ovarian cancer.

p38 MAPK–mediated upregulation of claudin-3 and claudin-4 by gemcitabine contributes to chemoresistance in ovarian cancer

Chemotherapy is a primary therapeutic option in cancer treatment, but often associated with unwanted side effects and drug resistance. Claudin-3 (CLDN3) and claudin-4 (CLDN4) are essential components of tight junctions, frequently overexpressed in ovarian cancer, serve as potential therapeutic targets. In this study, we utilized flow cytometry, qPCR, Western blot, and animal experiments to investigate the regulation of CLDN3 and CLDN4 by chemotherapy drug, gemcitabine, in the ovarian cancer cell line A2780. We reported that gemcitabine can induce expression of CLDN3 and CLDN4 in ovarian cancer cells. Mechanistically, we showed that gemcitabine induces expression of CLDN3 and CLDN4 through p38 MAP kinase mediated transcriptional regulation. Overexpression of CLDN3 or CLDN4 functionally protected A2780 ovarian cancer from gemcitabine induced cell killing. It appears that gemcitabine induced expression of CLDN3/4 is a chemoresistance mechanism for cancer cells. Gemcitabine-induced upregulation of CLDN3/4 suggests that ovarian cancer cells may be more effectively targeted using claudin-3/4-specific antibodies or antibody-drug conjugates (ADCs) in combination with chemotherapy, which could have clinical implications for ovarian cancer treatment in the future.

HDAC6 suppresses microRNA-199a transcription and augments HPV-positive cervical cancer progression through Wnt5a upregulation

High-risk human papillomavirus (HR-HPV) infection is a major risk factor for the initiation and progression of cervical cancer (CC). This study aimed to explore the role of histone deacetylase 6 (HDAC6) in HPV-positive CC and the molecules implicated. Differentially expressed genes between HPV-positive and HPV-negative tissues, and differentially expressed microRNAs (miRNAs) in cells after HDAC6 downregulation were identified using microarray analyses. The expression profiles of HDAC6 and miR-199a and their cellular functions were investigated via loss-of-function studies. Xenograft tumors were induced in mice for in vivo studies. HDAC6 and Wnt5a were highly expressed, whereas miR-199a was poorly expressed in HPV-positive CC tissues. Downregulation of HDAC6 reduced proliferation, migration, invasion, and resistance to apoptosis of HPV-positive CC cells. HDAC6 suppressed the transcription of miR-199a, and miR-199a targeted Wnt5a to inactivate the Wnt signaling pathway. Further downregulation of miR-199a blocked the inhibitory effect of HDAC6 silencing on CC cell growth both in vivo and in vitro, whereas further artificial inhibition of Wnt5a inactivated Wnt signaling and blocked the malignant behaviors of CC cells. This study showed that HDAC6 suppresses the transcription of miR-199a and enhances the progression of HPV-positive cervical cancer through upregulation of Wnt5a.

IFNγ induces JAK1/STAT1/p65 NFκB-dependent interleukin-8 expression in ovarian cancer cells, resulting in their increased migration

Interferon-γ (IFNγ) is a pleiotropic cytokine that has a crucial role in immune response and tumor immunity. Because of its anti-tumor effects, IFNγ has been used in cancer treatment. However, IFNγ also has tumor-promoting functions that are less well understood. Here, we show that IFNγ induces expression of the pro-inflammatory and pro-angiogenic chemokine interleukin-8 (IL-8, CXCL8) in ovarian cancer (OC) cells. The IFNγ-induced IL-8 expression is dependent on JAK1, STAT1, and p65 NFκB, and is associated with an increased occupancy of K314/315 acetylated p65 NFκB and Ser-727 phosphorylated STAT1 at the IL-8 promoter. Neutralization of IL-8 using anti-IL-8 antibody reduces IFNγ-induced migration of OC cells, and their invasion ability in 3D spheroids. Together, these findings identify IL-8 as a novel target induced by IFNγ/JAK1/STAT1/p65 NFκB signaling, and indicate that the IFNγ-induced IL-8 contributes to IFNγ pro-tumorigenic effects in ovarian cancer cells.

Wnt antagonist as therapeutic targets in ovarian cancer

Ovarian cancer is a fatal malignancy in women with a low survival rate that demands new therapeutic paradigms. Cancer cells acquire various exclusive alterations to proliferate, invade, metastasize, and escape cell death, acting independently of growth-inducing or growth-inhibiting signals. The nature of cellular signaling in tumorigenesis is interwoven. Wnt signaling is an evolutionarily conserved signaling cascade that has been shown to regulate ovarian cancer pathogenesis. The molecular mechanism of Wnt signaling underlying the development of ovarian cancer, drug resistance, and relapse is not completely understood. Extracellularly secreted Wnt signaling inhibitors are crucial regulators of ovarian cancer tumorigenesis and malignant properties of cancer stem cells. Wnt inhibitors arbitrated modifications affecting Wnt pathway proteins on the cell membranes, in the cytoplasm, and in the nucleus have been shown to span essential contributions in the initiation, progression, and chemoresistance of ovarian cancer. Although many extrinsic inhibitors developed targeting the downstream components of the Wnt signaling pathway, investigating the molecular mechanisms of endogenous secreted inhibitors might substantiate prognostic or therapeutic biomarkers development. Given the importance of Wnt signaling in ovarian cancer, more systematic studies combined with clinical studies are requisite to probe the precise mechanistic interactions of Wnt antagonists in ovarian cancer. This review outlines the latest progress on the Wnt antagonists and ovarian cancer-specific regulators such as micro-RNAs, small molecules, and drugs regulating these Wnt antagonists in ovarian tumourigenesis.

VEGFa/VEGFR2 autocrine and paracrine signaling promotes cervical carcinogenesis via β-catenin and snail

VEGF secretion into the tumor microenvironment by cancer cells regulates several oncogenic signaling pathways and cancer-regulated angiogenesis. VEGFR receptors are exclusively present on endothelial cells to maintain their biological homeostasis. The acquisition of unique VEGFR2 receptor and VEGFa in cervical cancer (CC) cells reflects VEGFa/VEGFR2 autocrine machinery. Given the critical role of VEGFR2 in endothelial cell proliferation, migration, and angiogenesis, we explored its function in CC epithelial-mesenchymal transition (EMT) and stemness. Here we report that VEGFR2 regulates cancer-induced angiogenesis and EMT-linked stemness in CC cells via AKT/GSK3β/β-catenin and Snail pathway. Receptor tyrosine kinase inhibitor (RTKi) of VEGFR, Pazopanib (PAZ), shows potential anti-VEGFR2 activity and inhibits VEGFa induced metastatic events such as migration, invasion, and anoikis resistance in CC cells. Similarly, PAZ also attenuates cancer-regulated angiogenesis by inhibiting VE-cadherin internalization in endothelial cells followed by inhibition of endothelial cell migration. Selective depletion of VEGFR2 ligand VEGFa in CC cells also attenuates EMT, metastatic events, and inhibition of cancer-induced angiogenesis. In addition, blocking of VEGFR2 signaling in CC cells via PAZ or shRNA alters the formation of cervical tumorspheres (TS) and their successive generation. Collectively, inhibition of functional VEGFa/VEGFR2 autocrine and paracrine axis ceases the cancer-promoting events in cervical cancer cells. Based on the finding in this study, this oncogenic pathways could be used as a potential therapeutic target in a clinical setting with conventional radio-chemotherapy to achieve synergistic killing of CC cells.

Iron overload increases the sensitivity of endometriosis stromal cells to ferroptosis via a PRC2-independent function of EZH2

Given the high concentration of iron in the micro-environment of ovarian endometriosis, it is plausible to hypothesize that ectopic endometrial cells may be more susceptible to undergoing ferroptosis. Manipulation of ferroptosis has been explored as a potential therapeutic strategy to treat related diseases. In this study, we examined the impact on ectopic endometrial stromal cells (EESCs) of iron overload and an inducer of ferroptosis. We found that the iron concentration in the ovarian endometriosis was much higher than control samples. Treatment of cultured EESCs with ferric ammonium citrate (FAC) increase the sensitivity to undergo ferroptosis. By analyzing the RNA-seq results, it was discovered that zeste 2 polycomb repressive complex 2 subunit (EZH2) was significantly downregulated in ferroptosis induced EESCs. Moreover, overexpression of EZH2 effectively prevented the induction of ferroptosis. In addition, the activity or expression of EZH2 is directly and specifically inhibited by the methyltransferase inhibitor GSK343, which raises the sensitivity of stromal cells to ferroptosis. Taken together, our findings revealed that EZH2 act as a suppressor in the induced cell ferroptosis through a PRC2-independent methyltransferase mechanism. Therefore, blocking EZH2 expression and inducing ferroptosis may be effective treatment approaches for ovarian endometriosis.

Hypomethylated gene RAC3 induces cell proliferation and invasion by increasing FASN expression in endometrial cancer

Endometrial cancer (EC) is one of the most prevalent gynecological cancers with a 5-year survival rate of 20-60%. Feasible prognostic molecular biomarkers of EC are necessary for accurate prediction of EC prognosis. RAC3 is a member of the Rho GTPases. Public databases including Gene Expression Profiling Interactive Analysis (GEPIA2), Tumor Immune Estimation Resource (TIMER), LinkedOmics, Search Tool for the Retrieval of Interacting Genes/Proteins (STRING), TISIDB and cBioPortal were employed to analyze the differential expression, clinicopathologic characteristics, functional networks, immune cell infiltrates and genetic alteration of RAC3 in EC patients. RAC3 expression was elevated in EC patients analyzed by TIMER and GEPIA. Overexpression of RAC3 was obviously correlated with clinical stage, histological type, histological grade and DNA hypomethylation. Patients with high RAC3 expression displayed poor overall survival. Functional enrichment analysis showed that RAC3 was involved in translational initiation, DNA replication and mRNA processing. RAC3 expression was negatively associated with infiltrating levels of B cells, CD8 + T cells, macrophages and dendritic cells in EC. Experiments in vitro showed that RAC3 was upregulated in EC tissues and cell lines, and RAC3 induced cell proliferation and invasion by increasing fatty acid synthase (FASN) expression. High expression of RAC3iscorrelated with poor prognosis and low infiltration of immune cells in EC. RAC3 promotes cell proliferation and invasion via FASN. These results demonstrate thatRAC3 functions as an EC oncogene and reveal its underlying mechanism in EC progression, suggesting that RAC3 may serve as a potential therapeutic target in EC.