Atractylenolide II reverses the influence of lncRNA XIST/miR‐30a‐5p/ROR1 axis on chemo‐resistance of colorectal cancer cells

This investigation was conducted to elucidate whether atractylenolide II could reverse the role of lncRNA XIST/miR‐30a‐5p/ROR1 axis in modulating chemosensitivity of colorectal cancer cells. We totally collected 294 pairs of colorectal cancer tissues and adjacent normal tissues and also purchased colorectal cancer cell lines and human embryonic kidney cell line. 5‐fluorouracil, cisplatin, mitomycin and adriamycin were designated as the chemotherapies for colorectal cell lines, and atractylenolides were arranged as the Chinese drug. The expressions of XIST, miR‐30a‐5p and ROR1 were quantified with aid of qRT‐PCR or Western blot, and luciferase reporter gene assay was implemented to determine the relationships among XIST, miR‐30a‐5p and ROR1. Our results demonstrated that XIST and ROR1 expressions were dramatically up‐regulated, yet miR‐30a‐5p expression was down‐regulated within colorectal cancer tissues (P < 0.05). The overexpressed XIST and ROR1, as well as under‐expressed miR‐30a‐5p, were inclined to promote viability and proliferation of colorectal cells under the influence of chemo drugs (P < 0.05). In addition, XIST could directly target miR‐30a‐5p, and ROR1 acted as the targeted molecule of miR‐30a‐5p. Interestingly, atractylenolides not only switched the expressions of XIST, miR‐30a‐5p and ROR1 within colorectal cancer cells but also significantly intensified the chemosensitivity of colorectal cancer cells (P < 0.05). Finally, atractylenolide II was discovered to slow down the viability and proliferation of colorectal cancer cells (P < 0.05). In conclusion, the XIST/miR‐30a‐5p/ROR1 axis could be deemed as pivotal markers underlying colorectal cancer, and administration of atractylenolide II might improve the chemotherapeutic efficacy for colorectal cancer.     

MicroRNA‐425‐5p regulates chemoresistance in colorectal cancer cells via regulation of Programmed Cell Death 10

Acquired chemoresistance represents a major obstacle in cancer treatment, the underlying mechanism of which is complex and not well understood. MiR‐425‐5p has been reported to be implicated tumorigenesis in a few cancer types. However, its role in regulating chemoresistance has not been investigated in colorectal cancer (CRC) cells. Microarray analysis was performed in isogenic chemosensitive and chemoresistant HCT116 cell lines to identify differentially expressed miRNAs. miRNA quantitative real‐time PCR was used to detect miR‐425‐5p expression levels between drug resistant and parental cancer cells. MiR‐425‐5p mimic and inhibitor were transfected, followed by CellTiter‐Glo^® assay to examine drug sensitivity in these two cell lines. Western Blot and luciferase assay were performed to investigate the direct target of miR‐425‐5p. Xenograft mouse models were used to examine in vivo function of miR‐425‐5p. Our data showed that expression of miR‐425‐5p was significantly up‐regulated in HCT116‐R compared with parental HCT116 cells. Inhibition of miR‐425‐5p reversed chemoresistance in HCT116‐R cells. Programmed cell death 10 (PDCD10) is the direct target of miR‐425‐5p which is required for the regulatory role of miR‐425‐5p in chemoresistance. MiR‐425‐5p inhibitor sensitized HCT116‐R xenografts to chemo drugs in vivo. Our study demonstrated that miR‐425‐5p regulates chemoresistance of CRC cells by modulating PDCD10 expression level both in vitro and in vivo. MiR‐425‐5p may represent a new therapeutic target for the intervention of CRC.     

miR‐148a inhibits early relapsed colorectal cancers and the secretion of VEGF by indirectly targeting HIF‐1α under non‐hypoxia/hypoxia conditions

Vascular endothelial growth factor (VEGF) is correlated with angiogenesis and early relapse of colorectal cancer (CRC). This study investigated the role of miR‐148a in the regulation of VEGF/angiogenesis and early relapse of CRC. We established a stable clone with miR‐148a expression in HCT116 and HT29 cell lines and created a hypoxic condition by using CoCl₂ to determine the underlying mechanism of miR‐148a. The effects of miR‐148a on the phosphoryl‐ERK (pERK)/hypoxia‐inducible factor‐1α (HIF‐1α)/VEGF pathway were evaluated through Western blotting and the inhibitory effect of miR‐148a on angiogenesis was demonstrated through a tube formation assay. Sixty‐three CRC tissues (28 early relapse and 35 non‐early relapse) were analysed to assess the relationship between miR‐148a and HIF‐1α/VEGF. The protein expression of pERK/HIF‐1α/VEGF in HCT116 and HT29 cells was significantly decreased by miR‐148a (all P < 0.05). The protein expression of VEGF/HIF‐1α was strongly inversely associated with the expression of miR‐148a in the 63 CRC tissue samples (all P < 0.05). Tube formation assay demonstrated that miR‐148a significantly obliterated angiogenesis. miR‐148a suppresses VEGF through down‐regulation of the pERK/HIF‐1α/VEGF pathway and might lead to the inhibition of angiogenesis; miR‐148a down‐regulation increased the early relapse rate of CRC. This demonstrates that miR‐148a is a potential diagnostic and therapeutic target.     

miR‐1‐3p and miR‐206 sensitizes HGF‐induced gefitinib‐resistant human lung cancer cells through inhibition of c‐Met signalling and EMT

Hepatocyte growth factor (HGF) overexpression is an important mechanism in acquired epidermal growth factor receptor (EGFR) kinase inhibitor gefitinib resistance in lung cancers with EGFR activating mutations. MiR‐1‐3p and miR‐206 act as suppressors in lung cancer proliferation and metastasis. However, whether miR‐1‐3p and miR‐206 can overcome HGF‐induced gefitinib resistance in EGFR mutant lung cancer is not clear. In this study, we showed that miR‐1‐3p and miR‐206 restored the sensitivities of lung cancer cells PC‐9 and HCC‐827 to gefitinib in present of HGF. For the mechanisms, we demonstrated that both miR‐1‐3p and miR‐206 directly target HGF receptor c‐Met in lung cancer. Knockdown of c‐Met mimicked the effects of miR‐1‐3p and miR‐206 transfections Meanwhile, c‐Met overexpression attenuated the effects of miR‐1‐3p and miR‐206 in HGF‐induced gefitinib resistance of lung cancers. Furthermore, we showed that miR‐1‐3p and miR‐206 inhibited c‐Met downstream Akt and Erk pathway and blocked HGF‐induced epithelial‐mesenchymal transition (EMT). Finally, we demonstrated that miR‐1‐3p and miR‐206 can increase gefitinib sensitivity in xenograft mouse models in vivo. Our study for the first time indicated the new function of miR‐1‐3p and miR‐206 in overcoming HGF‐induced gefitinib resistance in EGFR mutant lung cancer cell.     

LncRNA MEG3 inhibits the progression of prostate cancer by modulating miR‐9‐5p/QKI‐5 axis

This study was designed to detecting the influences of lncRNA MEG3 in prostate cancer. Aberrant lncRNAs expression profiles of prostate cancer were screened by microarray analysis. The qRT‐PCR and Western blot were employed to investigating the expression levels of lncRNA MEG3, miR‐9‐5p and QKI‐5. The luciferase reporter assay was utilized to testifying the interactions relationship among these molecules. Applying CCK‐8 assay, wound healing assay, transwell assay and flow cytometry in turn, the cell proliferation, migration and invasion abilities as well as apoptosis were measured respectively. LncRNA MEG3 was a down‐regulated lncRNA in prostate cancer tissues and cells and could inhibit the expression of miR‐9‐5p, whereas miR‐9‐5p down‐regulated QKI‐5 expression. Overexpressed MEG3 and QKI‐5 could decrease the abilities of proliferation, migration and invasion in prostate cancer cells effectively and increased the apoptosis rate. On the contrary, miR‐9‐5p mimics presented an opposite tendency in prostate cancer cells. Furthermore, MEG3 inhibited tumour growth and up‐regulated expression of QKI‐5 in vivo. LncRNA MEG3 was a down‐regulated lncRNA in prostate cancer and impacted the abilities of cell proliferation, migration and invasion, and cell apoptosis rate, this regulation relied on regulating miR‐9‐5p and its targeting gene QKI‐5.     

Induced miR‐99a expression represses Mtor cooperatively with miR‐150 to promote regulatory T‐cell differentiation

Peripheral induction of regulatory T (Treg) cells provides essential protection from inappropriate immune responses. CD4⁺ T cells that lack endogenous miRNAs are impaired to differentiate into Treg cells, but the relevant miRNAs are unknown. We performed an overexpression screen with T‐cell‐expressed miRNAs in naive mouse CD4⁺ T cells undergoing Treg differentiation. Among 130 candidates, the screen identified 29 miRNAs with a negative and 10 miRNAs with a positive effect. Testing reciprocal Th17 differentiation revealed specific functions for miR‐100, miR‐99a and miR‐10b, since all of these promoted the Treg and inhibited the Th17 program without impacting on viability, proliferation and activation. miR‐99a cooperated with miR‐150 to repress the expression of the Th17‐promoting factor mTOR. The comparably low expression of miR‐99a was strongly increased by the Treg cell inducer “retinoic acid”, and the abundantly expressed miR‐150 could only repress Mtor in the presence of miR‐99a. Our data suggest that induction of Treg cell differentiation is regulated by a miRNA network, which involves cooperation of constitutively expressed as well as inducible miRNAs.     

KITLG is a novel target of miR‐34c that is associated with the inhibition of growth and invasion in colorectal cancer cells

MiR‐34c is considered a potent tumour suppressor because of its negative regulation of multiple target mRNAs that are critically associated with tumorigenesis and metastasis. In the present study, we demonstrated a novel target of miR‐34c, KITLG, which has been implicated in colorectal cancer (CRC). First, we found a significant negative relationship between miR‐34c and KITLG mRNA expression levels in CRC cell lines, including HT‐29, HCT‐116, SW480 and SW620 CRC cell lines. In silico analysis predicted putative binding sites for miR‐34c in the 3′ untranslated region (3′UTR) of KITLG mRNA. A dual‐luciferase reporter assay further confirmed that KITLG is a direct target of miR‐34c. Then, the cell lines were infected with lentiviruses expressing miR‐34c or a miR‐34c specific inhibitor. Restoration of miR‐34c dramatically reduced the expression of KITLG mRNA and protein, while silencing of endogenous miR‐34c increased the expression of KITLG protein. The miR‐34c‐mediated down‐regulation of KITLG was associated with the suppression on proliferation, cellular transformation, migration and invasion of CRC cells, as well as the promotion on apoptosis. Knockdown of KITLG by its specific siRNA confirmed a critical role of KITLG down‐regulation for the tumour‐suppressive effects of miR‐34c in CRC cells. In conclusion, our results demonstrated that miR‐34c might interfere with KITLG‐related CRC and could be a novel molecular target for CRC patients.     

MiR‐129‐5p promotes docetaxel resistance in prostate cancer by down‐regulating CAMK2N1 expression

This study focuses on the effect of miR‐129‐5p on docetaxel‐resistant (DR) prostate cancer (PCa) cells invasion, migration and apoptosis. In our study, the expression of CAMK2N1 was assessed by qRT‐PCR in PCa patient tissues and cell lines including PC‐3 and PC‐3‐DR. Cells transfected with miR‐129‐5p mimics, inhibitor, CAMK2N1 or negative controls (NC) were used to interrogate their effects on DR cell invasions, migrations and apoptosis during docetaxel (DTX) treatments. The apoptosis rate of the PCa cells was validated by flow cytometry. Relationships between miR‐129‐5p and CAMK2N1 levels were identified by qRT‐PCR and dual‐luciferase reporter assay. CAMK2N1 was found to be down‐expressed in DR PCa tissue sample, and low levels of CAMK2N1 were correlated with high docetaxel resistance and clinical prediction of poor survival. CAMK2N1 levels were decreased in DR PCa cells treated with DXT. We further explored that up‐regulation of miR‐129‐5p could promote DR PCa cells viability, invasion and migration but demote apoptosis. Involved molecular mechanism studies revealed that miR‐129‐5p reduced downstream CAMK2N1 expression to further impact on chemoresistance to docetaxel of PCa cells, indicating its vital role in PCa docetaxel resistance. Our findings revealed that miR‐129‐5p contributed to the resistance of PC‐3‐DR cells to docetaxel through suppressing CAMK2N1 expression, and thus targeting miR‐129‐5p may provide a novel therapeutic approach in sensitizing PCa to future docetaxel treatment.     

Serum miR‐92a‐1 is a novel diagnostic biomarker for colorectal cancer

Colorectal cancer (CRC) is one of the most common cancers worldwide, with high mortality. Abnormally expressed microRNAs (miRNAs) are considered novel biomarkers in cancer diagnosis. The aim of this study was to investigate the diagnostic value of miR‐92a‐1 in patients with CRC. Serum samples were collected from 148 patients pathologically diagnosed with CRC and 68 gender‐ and age‐matched healthy volunteers. Quantitative real‐time polymerase chain reaction (qRT‐PCR) was used to measure serum miR‐92a‐1 level. Relationship between miR‐92a‐1 and clinicopathological features of CRC cases was analysed via chi‐square test. Receiver operating characteristic (ROC) curve was plotted to estimate the diagnostic value of miR‐92a‐1 in CRC. Serum miR‐92a‐1 was significantly up‐regulated in CRC patients compared with healthy individuals (P < .001). Moreover, miR‐92a‐1 expression was correlated with TNM stage (P = .02), histological stage (P = .003), lymph node metastasis (P = .003) and distant metastasis (P < .001). ROC analysis showed that the area under the ROC curve (AUC) was 0.914, suggesting high diagnostic accuracy of miR‐92a‐1 in ROC. The optimal cut‐off value was 1.485, with a sensitivity of 81.8% and a specificity of 95.6%. MiR‐92a‐1 is increased in CRC patients and correlated with aggressive clinical characteristics. Serum miR‐92a‐1 may be a potential diagnostic biomarker for CRC.     

The contrary functions of lncRNA HOTAIR/miR‐17‐5p/PTEN axis and Shenqifuzheng injection on chemosensitivity of gastric cancer cells

This study was implemented to figure out whether lncRNA HOTAIR/miR‐17‐5p/PTEN axis played a role that was opposite to Shenqifuzheng (SQFZ) injection in regulating the chemosensitivity of gastric cancer cells. The gastric cancer tissues were gathered and four gastric cancer cell lines were prepared, including BGC‐823, MGC‐803, SGC‐7901, and MKN28. Moreover, cisplatin, adriamycin, mitomycin, and 5‐fluoroura were managed as the chemo‐therapeutics, and SQFZ was prepared as a Chinese medicine. Striking distinctions of HOTAIR, miR‐17‐5p, and PTEN expressions were observed between gastric cancer tissues and para‐carcinoma normal tissues (P < 0.05). MKN28 was associated with the highest resistance to cisplatin, adriamycin, mitomycin, and 5‐fluoroura among all the cell types, and SQFZ significantly improved the MKN28 cells’ sensitivity to the drugs (P < 0.05). The over‐expressed HOTAIR and miR‐17‐5p, as well as under‐expressed PTEN tended to significantly facilitate the viability, EMT process and proliferation of MKN28 cells that were subject to treatment of chemo‐therapies (P < 0.05). SQFZ could amplify the effects of si‐HOTAIR, miR‐17‐5p inhibitor, and pcDNA‐PTEN on boosting the chemosensitivity of gastric cancer cells (P < 0.05). In addition, HOTAIR was also found to directly target miR‐17‐5p, and PTEN appeared to be subject to the modification of HOTAIR and miR‐17‐5p in its acting on the viability, proliferation, EMT process, and apoptosis of gastric cancer cells. The HOTAIR/miR‐17‐5p/PTEN axis could be regarded as the potential treatment targets for gastric cancer, and adjuvant therapy of SQFZ injection could assist in further improving the treatment efficacy of chemo‐therapies for gastric cancer.     

Human embryonic stem cells and metastatic colorectal cancer cells shared the common endogenous human microRNA‐26b

The increase in proliferation and the lack of differentiation of cancer cells resemble what occur in the embryonic stem cells during physiological process of embryogenesis. There are also striking similarities in the behaviour between the invasive placental cells and invasive cancer cells. In the present study, microarrays were used to analyse the global expression of microRNAs in a human embryonic stem cell line (i.e. HUES‐17) and four colorectal cancer (CRC) cell lines (i.e. LoVo, SW480, HT29 and Caco‐2) with different metastatic potentialities. Only the expression of miR‐26b was significant decreased in HUES‐17s and LoVo cells, compared with other three cell lines (P < 0.01). The quantitative real‐time PCR analysis confirmed the results of the microarray analysis. Overexpression of miR‐26b expression by miR‐26 mimics transfection and led to the significant suppression of the cell growth and the induction of apoptosis in LoVo cells in vitro, and the inhibition of tumour growth in vivo. Moreover, the potential targets of miR‐26b was predicted by using bioinformatics, and then the predicted target genes were further validated by comparing gene expression profiles between LoVo and NCM460 cell lines. Four genes (TAF12, PTP4A1, CHFR and ALS2CR2) with intersection were found to be the targets of miR‐26b. MetaCore network analysis further showed that the regulatory pathways of miR‐26b were significantly associated with the invasiveness and metastasis of CRC cells. These data suggest that miR‐26b might serve as a novel prognostic factor and a potential therapeutic target for CRC.     

miR‐515‐5p controls cancer cell migration through MARK4 regulation

Here, we show that miR‐515‐5p inhibits cancer cell migration and metastasis. RNA‐seq analyses of both oestrogen receptor receptor‐positive and receptor‐negative breast cancer cells overexpressing miR‐515‐5p reveal down‐regulation of NRAS, FZD4, CDC42BPA, PIK3C2B and MARK4 mRNAs. We demonstrate that miR‐515‐5p inhibits MARK4 directly 3′ UTR interaction and that MARK4 knock‐down mimics the effect of miR‐515‐5p on breast and lung cancer cell migration. MARK4 overexpression rescues the inhibitory effects of miR‐515‐5p, suggesting miR‐515‐5p mediates this process through MARK4 down‐regulation. Furthermore, miR‐515‐5p expression is reduced in metastases compared to primary tumours derived from both in vivo xenografts and samples from patients with breast cancer. Conversely, miR‐515‐5p overexpression prevents tumour cell dissemination in a mouse metastatic model. Moreover, high miR‐515‐5p and low MARK4 expression correlate with increased breast and lung cancer patients' survival, respectively. Taken together, these data demonstrate the importance of miR‐515‐5p/MARK4 regulation in cell migration and metastasis across two common cancers.     

CiRS‐7 targeting miR‐7 modulates the progression of non‐small cell lung cancer in a manner dependent on NF‐κB signalling

The purpose of this study was to figure out the effect of ciRS‐7/miR‐7/NF‐κB axis on the development of non‐small cell lung cancer (NSCLC). In response, the expressions of ciRS‐7, miR‐7 and NF‐κB subunit (ie RELA) within NSCLC tissues and cell lines were determined with real‐time polymerase chain reaction (RT‐PCR) and Western blot. Moreover, the NSCLC cells were transfected with pcDNA3‐ciRS‐7‐ir, pcDNA3‐ciRS‐7, miR‐NC and miR‐7 mimic. Furthermore, the targeted relationships between ciRS‐7 and miR‐7, as well as between miR‐7 and RELA, were confirmed by luciferase reporter assay. The proliferation, migration and apoptosis of NSCLC cells were, successively, measured using CCK‐8 assay, wound‐healing assay and flow cytometry test. Consequently, ciRS‐7, miR‐7, histopathological grade, lymph node metastasis and histopathological stage could independently predict the prognosis of patients with NSCLC (all P < .05). Moreover, remarkably up‐regulated ciRS‐7 and RELA expressions, as along with down‐regulated miR‐7 expressions, were found within NSCLC tissues and cells in comparison with normal ones (P < .05). Besides, overexpressed ciRS‐7 and underexpressed miR‐7 were correlated with increased proliferation, migration and invasion, yet reduced apoptosis rate of NSCLC cells (P < .05). More than that, ciRS‐7 specifically targeted miR‐7 to reduce its expressions (P < .05). Ultimately, the NSCLC cells within miR‐7 + RELA group were observed with superior proliferative, migratory and invasive capabilities than those within miR‐7 group (P < .05), and RELA expression was also significantly modified by both ciRS‐7 and miR‐7 (P < .05). In conclusion, the ciRS‐7/miR‐7/NF‐kB axis could exert pronounced impacts on the proliferation, migration, invasion and apoptosis of NSCLC cells.     

MicroRNA‐1271 suppresses the proliferation and invasion of colorectal cancer cells by regulating metadherin/Wnt signaling

Recent studies have reported an important role for microRNA‐1271 (miR‐1271) in tumorigenesis. However, the role of miR‐1271 in colorectal cancer remains unknown. Here, we found that miR‐1271 was significantly decreased in colorectal cancer tissues and cell lines. Overexpression of miR‐1271 inhibited cell proliferation, colony formation, cell invasion, and induced cell cycle arrest in colorectal cancer cells. Metadherin (MTDH) was identified as a target gene of miR‐1271. Moreover, miR‐1271 negatively regulated MTDH expression in colorectal cancer cells and reversely correlated with MTDH expression in colorectal cancer specimens. Additionally, miR‐1271 also regulated the activation of Wnt signaling in colorectal cancer cells. The restoration of MTDH expression significantly reversed the antitumor effect of miR‐1271 in colorectal cancer cells. These findings indicate an important role for miR‐1271/MTDH in the tumorigenesis of colorectal cancer, and suggest that miR‐1271 may be a novel therapeutic target for colorectal cancer.