Hypoxic tumour cell-derived exosomal miR-340-5p promotes radioresistance of oesophageal squamous cell carcinoma via KLF10

Abstract Background: Radiotherapy resistance is a major obstacle in the treatment of oesophageal squamous cell carcinoma (OSCC). Hypoxia is a critical cause of radioresistance. However, the communication between hypoxic cells and aerobic cells via exosomes during the transfer of radiation resistance remains unclear. Methods: Exo-miR-340-5p levels were analysed by RNA-seq and qRT-PCR. We co-cultured OSCC cells with isolated normoxic and hypoxic exosomes to study their impact on radiosensitivity. We used a specific exo-miR-340-5p mimic and knock-down retrovirus to explore the role of this miRNA in the transfer of radioresistance from hypoxic to normoxic cells. Dual-luciferase reporter and RIP assays were used to verify KLF10 as a putative target of miR-340-5p. Several in vitro assays were conducted and xenograft models were established to investigate the effect of exo-miR-340-5p on OSCC radiosensitivity. The plasma exo-miR-340-5p levels in OSCC patients were analysed to study the clinical value of this parameter.Results: Hypoxic exosomes alleviated radiation-induced apoptosis and accelerated DNA damage repair. miR-340-5p was highly expressed in hypoxic exosomes and was transferred into normoxic cells, where it induced radioresistance. Overexpression of miR-340-5p in normoxic OSCC cells mimicked the radioresistance of cells co-cultured with hypoxic exosomes. Knockdown of miR-340-5p in hypoxic exosomes reversed the radioresistance effect, indicating that exo-miR-340-5p is critical for hypoxic EV-transferred radioresistance. KLF10 was identified as the direct target of miR-340-5p. Moreover, metformin was found to increase the expression of KLF10 and enhance the radiosensitivity of OSCC. Higher levels of miR-340-5p in the plasma exosomes from OSCC patients are related to a poorer radiotherapy response and prognosis. Conclusions: Hypoxic tumour cell-derived exosomal miR-340-5p confers radioresistance in OSCC by targeting KLF10/UVRAG, suggesting that miR-340-5p could be a potential biomarker and therapeutic target for the enhancement of radiosensitivity in OSCC. Metformin can increase KLF10 expression, which ameliorates the radioresistance induced by exo-miR-340-5p transfer. Therefore, metformin could be further investigated as a therapeutic option for the treatment of OSCC.

Download Full-text

Hypoxic tumour cell-derived exosomal miR-340-5p promotes radioresistance of oesophageal squamous cell carcinoma via KLF10

Journal of Experimental & Clinical Cancer Research ◽

10.1186/s13046-021-01834-9 ◽

2021 ◽

Vol 40 (1) ◽

Author(s):

Fangyu Chen ◽

Bing Xu ◽

Jie Li ◽

Xi Yang ◽

Junjie Gu ◽

...

Keyword(s):

Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Squamous Cell ◽

Tumour Cell ◽

Therapeutic Option ◽

In Vitro Assays ◽

Luciferase Reporter ◽

Oesophageal Squamous Cell Carcinoma ◽

Potential Biomarker ◽

Hypoxic Tumour Cell

Abstract Background Radiotherapy resistance is a major obstacle in the treatment of oesophageal squamous cell carcinoma (OSCC). Hypoxia is a critical cause of radioresistance. However, the communication between hypoxic cells and aerobic cells via exosomes during the transfer of radiation resistance remains unclear. Methods Exo-miR-340-5p levels were analysed by RNA-seq and qRT-PCR. We co-cultured OSCC cells with isolated normoxic and hypoxic exosomes to study their impact on radiosensitivity. We used a specific exo-miR-340-5p mimic and knock-down retrovirus to explore the role of this miRNA in the transfer of radioresistance from hypoxic to normoxic cells. Dual-luciferase reporter and RIP assays were used to verify KLF10 as a putative target of miR-340-5p. Several in vitro assays were conducted and xenograft models were established to investigate the effect of exo-miR-340-5p on OSCC radiosensitivity. The plasma exo-miR-340-5p levels in OSCC patients were analysed to study the clinical value of this parameter. Results Hypoxic exosomes alleviated radiation-induced apoptosis and accelerated DNA damage repair. miR-340-5p was highly expressed in hypoxic exosomes and was transferred into normoxic cells, where it induced radioresistance. Overexpression of miR-340-5p in normoxic OSCC cells mimicked the radioresistance of cells co-cultured with hypoxic exosomes. Knockdown of miR-340-5p in hypoxic exosomes reversed the radioresistance effect, indicating that exo-miR-340-5p is critical for hypoxic EV-transferred radioresistance. KLF10 was identified as the direct target of miR-340-5p. Moreover, metformin was found to increase the expression of KLF10 and enhance the radiosensitivity of OSCC. Higher levels of miR-340-5p in the plasma exosomes from OSCC patients are related to a poorer radiotherapy response and prognosis. Conclusions Hypoxic tumour cell-derived exosomal miR-340-5p confers radioresistance in OSCC by targeting KLF10/UVRAG, suggesting that miR-340-5p could be a potential biomarker and therapeutic target for the enhancement of radiosensitivity in OSCC. Metformin can increase KLF10 expression, which ameliorates the radioresistance induced by exo-miR-340-5p transfer. Therefore, metformin could be further investigated as a therapeutic option for the treatment of OSCC.

Download Full-text

Hypoxic Tumour Cell-derived Exosomal miR-340-5p Promotes Radioresistance of Oesophageal Squamous Cell Carcinoma via KLF10

10.21203/rs.3.rs-75570/v1 ◽

2020 ◽

Author(s):

Fangyu Chen ◽

Bing Xu ◽

Jie Li ◽

Xi Yang ◽

Junjie Gu ◽

...

Keyword(s):

Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Squamous Cell ◽

Tumour Cell ◽

Clinical Samples ◽

Oesophageal Squamous Cell Carcinoma ◽

Potential Biomarker ◽

Underlying Mechanisms ◽

Induced Apoptosis ◽

Hypoxic Tumour Cell

Abstract Background: Radiotherapy resistance is a major obstacle in the treatment of oesophageal squamous cell carcinoma (OSCC). Hypoxia is a critical cause of radioresistance. However, the communications between hypoxic cells and aerobic cells via exosomes during radiation resistance remain unclear. Methods: We previously detected miR-340-5p is highly expressed in hypoxic OSCC exosomes by RNA-seq. The effects and mechanisms of hypoxic EVs and miR-340-5p on radiosensitivity were evaluated by qPCR, western blot IHC, flow cytometry, TUNEL assay, etc. Tumour xenografts and clinical samples were used for evaluating radiosensitivity and dissecting underlying mechanisms. Results: Hypoxic exosomes can alleviate radiation induced apoptosis and accelerate DNA damage repair. miR-340-5p is highly expressed in hypoxic exosomes and causes radioresistance. Knockdown miR-340-5p in hypoxic EVs reversed whose radioresistant effect, indicating miR-340-5p is critical for hypoxic EV-induced radioresistance. KLF10 is the direct target of miR-340-5p. Moreover, metformin was found to increase the expression of KLF10 and enhance radiosensitivity of OSCC. And higher levels of miR-340-5p in the plasma exosomes of OSCC patients are related to poor radiotherapy response and prognosis. Conclusions: Hypoxic tumour cell-derived exosomal miR-340-5p confers radioresistance in OSCC by targeting KLF10/UVRAG, suggesting that miR-340-5p could be a potential biomarker and therapeutic target for the enhancement of radiosensitivity in OSCC. Combined use of metformin and radiotherapy might benefit OSCC patients.

Download Full-text

circPARD3 drives malignant progression and chemoresistance of laryngeal squamous cell carcinoma by inhibiting autophagy through the PRKCI-Akt-mTOR pathway

Molecular Cancer ◽

10.1186/s12943-020-01279-2 ◽

2020 ◽

Vol 19 (1) ◽

Author(s):

Wei Gao ◽

Huina Guo ◽

Min Niu ◽

Xiwang Zheng ◽

Yuliang Zhang ◽

...

Keyword(s):

Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Squamous Cell ◽

Laryngeal Squamous Cell Carcinoma ◽

Mtor Pathway ◽

Luciferase Reporter ◽

Circular Rnas ◽

Autophagic Flux ◽

In Vivo Models ◽

Potential Biomarker

Abstract Background Laryngeal squamous cell carcinoma (LSCC) is the second most common malignant tumor in head and neck. Autophagy and circular RNAs (circRNAs) play critical roles in cancer progression and chemoresistance. However, the function and mechanism of circRNA in autophagy regulation of LSCC remain unclear. Methods The autophagy-suppressive circRNA circPARD3 was identified via RNA sequencing of 107 LSCC tissues and paired adjacent normal mucosal (ANM) tissues and high-content screening. RT-PCR, Sanger sequencing, qPCR and fluorescence in situ hybridization were performed to detect circPARD3 expression and subcellular localization. Biological functions of circPARD3 were assessed by proliferation, migration, invasion, autophagic flux, and chemoresistance assays using in vitro and in vivo models. The mechanism of circPARD3 was investigated by RNA immunoprecipitation, RNA pulldown, luciferase reporter assays, western blotting and immunohistochemical staining. Results Autophagy was inhibited in LSCC, and circPARD3 was upregulated in the LSCC tissues (n = 100, p < 0.001). High circPARD3 level was associated with advanced T stages (p < 0.05), N stages (p = 0.001), clinical stages (p < 0.001), poor differentiation degree (p = 0.025), and poor prognosis (p = 0.002) of LSCC patients (n = 100). Functionally, circPARD3 inhibited autophagy and promoted LSCC cell proliferation, migration, invasion and chemoresistance. We further revealed that activation of the PRKCI-Akt-mTOR pathway through sponging miR-145-5p was the main mechanism of circPARD3 inhibited autophagy, promoting LSCC progression and chemoresistance. Conclusion Our study reveals that the novel autophagy-suppressive circPARD3 promotes LSCC progression and chemoresistance through the PRKCI-Akt-mTOR pathway, providing new insights into circRNA-mediated autophagy regulation and potential biomarker and target for LSCC treatment. Graphical abstract

Download Full-text

miR-375 Suppresses IGF1R Expression and Contributes to Inhibition of Cell Progression in Laryngeal Squamous Cell Carcinoma

BioMed Research International ◽

10.1155/2014/374598 ◽

2014 ◽

Vol 2014 ◽

pp. 1-11 ◽

Cited By ~ 15

Author(s):

Jie Luo ◽

Jianhui Wu ◽

Zenghong Li ◽

Hao Qin ◽

Bin Wang ◽

...

Keyword(s):

Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Squamous Cell ◽

Noncoding Rna ◽

Molecular Mechanisms ◽

Laryngeal Squamous Cell Carcinoma ◽

Suppressive Effect ◽

In Vitro Assays ◽

Luciferase Reporter

MicroRNAs (miRNAs) are small noncoding RNA molecules which are involved in tumorigenesis and development. To investigate their role in primary laryngeal squamous cell carcinoma (LSCC), miRNA GeneChips were used to screen the differentially expressed miRNA, and then validated by real-time quantitative PCR in LSCC samples, we found that miR-375 was frequently downregulated in primary LSCC tissues. The tumor-suppressive effect of miR-375 was determined by in vitro assays; through gain-of-function studies we demonstrated that miR-375 can inhibit LSCC cell (SNU-48 and SNU-899) proliferation, motility, and invasion, and promote their apoptosis. In addition, bioinformatics tools TargetScan, PicTar, and Miranda were used to investigate the potential target of miR-375; bioinformatics analysis and dual-luciferase reporter assay indicated that IGF1R was a novel direct target of miR-375. Ectopic transfection of miR-375 led to a significant reduction in IGF1R and its downstream signaling molecule AKT at both the mRNA and protein levels in LSCC cells. Our results suggested that downregulation of miR-375 is one of the molecular mechanisms for the development and progression of LSCC by directly targeting IGF1R and affecting its downstream AKT signaling pathways. Furthermore, miR-375 and IGF1R may serve as a novel therapeutic target for LSCC.

Download Full-text

LINC00958 promotes proliferation, migration, invasion, and epithelial-mesenchymal transition of oesophageal squamous cell carcinoma cells

PLoS ONE ◽

10.1371/journal.pone.0251797 ◽

2021 ◽

Vol 16 (5) ◽

pp. e0251797

Author(s):

Biqi Wang ◽

Duo Tang ◽

Zijia Liu ◽

Qian Wang ◽

Shan Xue ◽

...

Keyword(s):

Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Oesophageal Cancer ◽

Squamous Cell ◽

Epithelial Mesenchymal Transition ◽

Quantitative Polymerase Chain Reaction ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Oesophageal Squamous Cell Carcinoma ◽

Mesenchymal Transition

Oesophageal cancer is one of the deadliest cancers in the world. Oesophageal squamous cell carcinoma (ESCC) is the most prevalent histological type of oesophageal cancer. Oesophageal cancer has a poor prognosis because of its invasiveness. Thus, it is especially important to seek effective treatment methods. Research indicates that long non-coding RNAs (lncRNAs) play a significant role in the occurrence and development of oesophageal cancer. The aim of this study was to describe the role of LINC00958 in ESCC. Bioinformatics and real-time quantitative polymerase chain reaction (RT-qPCR) methods were utilized to predict and verify the expression of LINC00958 in ESCC. Related functional experiments, including cell proliferation, migration and invasion, were performed. In addition, a western blot and a dual luciferase reporter gene experiment were used to study the detailed carcinogenic mechanism of LINC00958. The results indicated there was a high expression of LINC00958 in ESCC, which promoted proliferation, migration, invasion and Epithelial–Mesenchymal Transition (EMT) of ESCC cells, and this effect may be via regulating miR-510-5p.

Download Full-text

The oncogenetic role of microRNA-31 as a potential biomarker in oesophageal squamous cell carcinoma

Clinical Science ◽

10.1042/cs20110207 ◽

2011 ◽

Vol 121 (10) ◽

pp. 437-447 ◽

Cited By ~ 94

Author(s):

Tengfei Zhang ◽

Qiming Wang ◽

Dan Zhao ◽

Yaling Cui ◽

Bangrong Cao ◽

...

Keyword(s):

Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Squamous Cell ◽

Luciferase Reporter ◽

Oesophageal Squamous Cell Carcinoma ◽

Validation Group ◽

Serum Samples ◽

Total N ◽

Normal Controls

miR-31 (microRNA-31) is frequently altered in numerous cancers. The aim of the present study was to investigate the role of miR-31 in ESCC (oesophageal squamous cell carcinoma). We measured miR-31 in 45 paired ESCC tissues and 523 serum samples using real-time RT (reverse transcription)–PCR. The serum samples were divided into a discovery group (120 ESCCs and 121 normal controls), a validation group (81 ESCCs and 81 controls), and a final group comprising six other common tumours (colorectal, liver, cervical, breast, gastric and lung cancers; total n=120). A Mann–Whitney U test and Wilcoxon matched-pairs test were used for the statistics. miR-31 was up-regulated in 77.8% of the ESCC tissues. Serum miR-31 levels in ESCC patients were significantly higher than in normal controls (P<0.001). It yielded an ROC (receiver operating characteristic) AUC (area under the curve) of 0.902 [95% CI (confidence interval), 0.857–0.936] in the discovery group and a similar result in the validation group [ROC AUC, 0.888 (95% CI, 0.819–0.939)]. Patients with high-levels of serum miR-31 also had a poorer prognosis in relapse-free survival (P=0.001) and tumour-specific survival (P=0.005). In vitro studies showed that miR-31 promoted ESCC colony formation, migration and invasion. Luciferase reporter and Western blot assays confirmed that three tumour suppressor genes, namely EMP1 (epithelial membrane protein 1), KSR2 (kinase suppressor of ras 2) and RGS4 (regulator of G-protein signalling 4), were targeted by miR-31. We conclude that miR-31 plays oncogenetic functions and can serve as a potential diagnostic and prognostic biomarker for ESCC.

Download Full-text

TGF-β-induced PLEK2 promotes metastasis and chemoresistance in oesophageal squamous cell carcinoma by regulating LCN2

Cell Death and Disease ◽

10.1038/s41419-021-04155-z ◽

2021 ◽

Vol 12 (10) ◽

Author(s):

Feng Wang ◽

Chaoqi Zhang ◽

Hong Cheng ◽

Chengming Liu ◽

Zhiliang Lu ◽

...

Keyword(s):

Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Squamous Cell ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Oesophageal Squamous Cell Carcinoma ◽

Promoter Sequences ◽

Regulatory Processes

AbstractOesophageal squamous cell carcinoma (ESCC) has a relatively unfavourable prognosis due to metastasis and chemoresistance. Our previous research established a comprehensive ESCC database (GSE53625). After analysing data from TCGA database and GSE53625, we found that PLEK2 predicted poor prognosis in ESCC. Moreover, PLEK2 expression was also related to the overall survival of ESCC patients undergoing chemotherapy. Repression of PLEK2 decreased the proliferation, migration, invasion and chemoresistance of ESCC cells in vitro and decreased tumorigenicity and distant metastasis in vivo. Mechanistically, luciferase reporter assay and chromatin immunoprecipitation assay suggested that TGF-β stimulated the process that Smad2/3 binds to the promoter sequences of PLEK2 and induced its expression. RNA-seq suggested LCN2 might a key molecular regulated by PLEK2. LCN2 overexpression in PLEK2 knockdown ESCC cells reversed the effects of decreased migration and invasion. In addition, TGF-β induced the expression of LCN2, but the effect disappeared when PLEK2 was knockdown. Moreover, AKT was phosphorylated in all regulatory processes. This study detected the major role of PLEK2 in driving metastasis and chemoresistance in ESCC by regulating LCN2, which indicates the potential use of PLEK2 as a biomarker to predict prognosis and as a therapeutic target for ESCC.

Download Full-text