Apoptosis In Non-small Cell Lung Cancer Cells Research Articles

Inhibiting ALK-TOPK signaling pathway promotes cell apoptosis of ALK-positive NSCLC

T-LAK cell-oriented protein kinase (TOPK) is a potential therapeutic target in tumors. However, its role in anaplastic lymphoma kinase (ALK)-positive non-small cell lung cancer (NSCLC) has not been reported. Here, we found that TOPK was highly expressed in ALK-positive NSCLC. Additionally, ALK was identified as another upstream kinase of TOPK by in vitro kinase assay screening. Then, it was proven that ALK phosphorylated TOPK at Y74 in vitro and ex vivo, and the pathways downstream of ALK-TOPK were explored by phosphoproteomic analysis. Subsequently, we demonstrated that inhibiting TOPK enhanced tumor sensitivity to alectinib (an ALK inhibitor). The combination of alectinib and HI-032 (a TOPK inhibitor) suppressed the growth and promoted the apoptosis of ALK-positive NSCLC cells ex vivo and in vivo. Our findings reveal a novel ALK-TOPK signaling pathway in ALK-positive NSCLC. The combination of alectinib and HI-032 might be a promising therapeutic strategy for improving the sensitivity of ALK-positive NSCLC to targeted therapy.

Fasudil Increased the Sensitivity to Gefitinib in NSCLC by Decreasing Intracellular Lipid Accumulation

Simple SummaryThe anti-tumor role of Fasudil in EGFR-mutation NSCLC as well as its mechanism are largely unknown. Here, we show that Fasudil could effectively inhibit EGFR-mutation cell growth and enhance the sensitivity of gefitinib-resistant NSCLC cells to gefitinib by suppressing intracellular lipid accumulation. Mechanistic investigations showed that Fasudil could reverse gefitinib-induced SCD1 expression by suppressing AMPK activity, and combination therapy had a greater inactivation effect on the EGFR/PI3K/AKT pathway than either treatment alone. These findings highlight the clinical significance of Fasudil in EGFR mutation NSCLC therapy.Tyrosine kinase inhibitors (TKIs) resistance is a challenge in patients with epidermal growth factor receptor (EGFR)-mutant non-small-cell lung cancer (NSCLC). Here, we examined the effect of Fasudil in reversing TKIs resistance. The results of CCK8 assay, clone formation assay, cell cycle arrest analysis, and apoptosis analysis show that Fasudil treatment effectively suppressed the growth and induced apoptosis of the EGFR-mutant NSCLC cells. Furthermore, Fasudil in combination with gefitinib showed a synergistic anti-tumor effect in gefitinib-resistant NSCLC cells. RNA-seq analysis and immunoblotting indicated that Fasudil treatment significantly inhibited intracellular lipid accumulation and EGFR/PI3K/AKT pathway activation. Mechanistic investigations showed that Fasudil regulated lipogenic gene expressions via AMPK signal pathway. In vivo, Fasudil and gefitinib co-administration significantly attenuated the growth of H1975 nude mouse xenograft models, suggesting that Fasudil treatment combined with gefitinib can be applied as a therapy for gefitinib-resistant NSCLC cells.

Biomimetic Red Blood Cell Membrane-Mediated Nanodrugs Loading Ursolic Acid for Targeting NSCLC Therapy.

Simple SummaryLung cancer is the second most common cancer after breast cancer. Non-small-cell lung cancer, which represents more than 85% of all lung cancer subtypes, is known for its tumor progression and metastasis, resulting in poor clinical outcomes. Conventional therapies for NSCLC, such as surgery, chemotherapy, and radiotherapy, always fail due to therapeutic resistance. In recent years, ursolic acid (UA), a natural pentacyclic triterpenoid compound, has been shown to be a promising antitumor drug by regulating multiple signaling pathways in cancers. Unfortunately, the poor water solubility, low bioavailability, and systemic toxicity of UA limit its clinical application. In this study, a biomimetic red blood cell membrane nanocarrier was developed to deliver UA to targeted tumor sites efficiently, and it inhibited tumor growth by inducing the apoptosis and autophagy of cancer cells both in vitro and in vivo.As one of the most common cancers worldwide, non-small-cell lung cancer (NSCLC) treatment always fails owing to the tumor microenvironment and resistance. UA, a traditional Chinese medicine, was reported to have antitumor potential in tumor models in vitro and in vivo, but showed impressive results in its potential application for poor water solubility. In this study, a novel biomimetic drug-delivery system based on UA-loaded nanoparticles (UaNPs) with a red blood cell membrane (RBCM) coating was developed. The RBCM-coated UANPs (UMNPs) exhibited improved water solubility, high stability, good biosafety, and efficient tumor accumulation. Importantly, the excellent antitumor efficiency of the UMNPs was confirmed both in vitro and in vivo in cancer models. In addition, we further investigated the antitumor mechanism of UMNPs. The results of Western blotting showed that UMNPs exerted an anticancer effect by inducing the apoptosis and autophagy of NSCLC cells, which makes it superior to free UA. In addition, body weight monitoring, hematoxylin and eosin (HE) analysis, and immunohistochemical (IHC) analysis showed no significant difference between UMNPs and the control group, indicating the safety of UMNPs. Altogether, the preparation of biomimetic UMNPs provides a promising strategy to improve outcomes in NSCLC.

TAOK1 Promotes Proliferation and Invasion of Non-Small-Cell Lung Cancer Cells by Inhibition of WWC1.

Background For patients with advanced non-small-cell lung cancer (NSCLC), targeted therapy significantly improves the therapeutic effect of NSCLC patients. With the development of molecular targeted therapy, more and more NSCLC-related genes have been found. Thousand and one amino-acid kinase 1 (TAOK1) has been identified as a potential target for drug research in various cancers. The main objective of this study was to explore the expression and function of TAOK1 in NSCLC. Methods Western blotting was employed to assess TAOK1 expression in NSCLC cell lines. The effects of TAOK1 on biological behaviors, including proliferation, invasion, and apoptosis of NSCLC cells, were assessed. The relationship between TAOK1 and WW and C2 domain containing 1 (WWC1) was assessed by Co-IP assay. The subcutaneous injection of tumor cells in nude mice was used to verify it in vivo. Results As expected, TAOK1 was increased in NSCLC cell lines. Following TAOK1 knockdown, NSCLC cells exhibited a significant decrease in the invasion and increased apoptosis in vitro. Instead, the TAOK1 elevation showed the opposite results. The Co-IP assay identified that TAOK1 specifically interacted with WWC1. Knockdown of WWC1 overturned TAOK1 silencing-mediated malignant phenotype of NSCLC cells. Additionally, subcutaneous tumorigenesis assays in nude mice confirmed that TAOK1 knockdown markedly restrained the proliferation capacity of NSCLC cells in vivo. Conclusion Surprisingly, TAOK1 overexpression in NSCLC promotes tumor cell growth and invasion, which is associated with downregulation of its downstream protein WWC1, and this result might provide a robust research basis to inquire about the precise therapeutic targets for NSCLC.

CircLIFR Inhibits Non-small Cell Lung Cancer Progression by Acting as a miR-429 Sponge to Enhance CELF2 Expression.

Lung cancer is the most commonly diagnosed cancer and the leading reason for tumor-related mortality, while non-small cell lung cancer (NSCLC) is the most usual type of lung cancer. Circular RNAs (circRNAs) have emerged as vital regulators in the development of human cancers, including NSCLC. We aimed to explore the functions of circRNA leukemia inhibitory factor receptor (circLIFR) in NSCLC progression. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to quantify the expression of circLIFR, microRNA-429 (miR-429), and Elav-like family member 2 (CELF2) in NSCLC tissues and cells. Cell proliferation capability of NSCLC cells was determined by Cell Counting Kit-8 (CCK-8) and colony formation assays. The flow cytometry assay was performed to evaluate cell-cycle distribution and apoptosis of NSCLC cells. The abilities of migration and invasion were measured by transwell assay. In addition, the activities of caspase 3 and caspase 9 were measured by the assay kits. The interaction relationship between miR-429 and circLIFR or CELF2 was analyzed by dual-luciferase reporter, RNA immunoprecipitation (RIP), and RNA pull-down assays. The expression levels of related proteins were examined by Western Blot assay. The xenograft experiment was established to explore the role of circLIFR in vivo. CircLIFR, circular, and stable transcript in NSCLC cells, was decreased more than 2 folds in NSCLC tissues and cells than controls (P < 0.0001). Importantly, overexpression of circLIFR impeded cell proliferation, migration, invasion, and inactivated protein kinase B (AKT)/phosphatase and tensin homolog (PTEN)-signaling pathways while enhanced apoptosis and cell-cycle arrest in NSCLC cells, which was overturned by upregulation of miR-429 or silencing of CELF2. Furthermore, the upregulation of circLIFR inhibited NSCLC tumor growth in vivo. Overexpression of circLIFR could suppress NSCLC progress by acting as a sponge of miR-429 to regulate the expression of CELF2 and PTEN/AKT-signaling pathways in NSCLC.

Polyphyllin II induced apoptosis of NSCLC cells by inhibiting autophagy through the mTOR pathway

Context Polyphyllin II (PPII) is a steroidal saponin isolated from Rhizoma Paridis. It exhibits significant antitumor activity such as anti-proliferation and pro-apoptosis in lung cancer. Objective To explore whether PPII induce autophagy and the relationship between autophagy and apoptosis in non-small cell lung cancer (NSCLC) cells. Materials and methods The effects of PPII (0, 1, 5, and 10 μM) were elucidated by CCK8 assay, colony formation test, TUNEL staining, MDC method, and mRFP-GFP-LC3 lentivirus transfection in A549 and H1299 cells for 24 h. DMSO-treated cells were selected as control. The protein expression of autophagy (LC3-II, p62), apoptosis (Bcl-2, Bax, caspase-3) and p-mTOR was detected by Western blotting. We explored the relationship between autophagy and apoptosis by autophagy inhibitor CQ (10 μM) and 3-MA (5 mM). Results PPII (0, 1, 5, and 10 μM) inhibited the proliferation and induced apoptosis. The IC50 values of A549 and H1299 cells were 8.26 ± 0.03 and 2.86 ± 0.83 μM. We found that PPII could induce autophagy. PPII promoted the formation of autophagosome, increased the expression of LC3-II/LC3-I (p < 0.05), while decreased p62 and p-mTOR (p < 0.05). Additionally, the co-treatment with autophagy inhibitors promoted the protein expression of c-caspase-3 and rate of Bax/Bcl-2 (p < 0.05), compared with PPII-only treatment group. Therefore, our results indicated that PPII-induced autophagy may be a mechanism to promote cell survival, although it can also induce apoptosis. Conclusions PPII-induced apoptosis exerts its anticancer activity by inhibiting autophagy, which will hopefully provide a prospective compound for NSCLC treatment.

LncRNA NEAT1 induces autophagy through the miR-128-3p/ADAM28 axis to suppress apoptosis of nonsmall-cell lung cancer.

This study aimed to identify the molecular mechanism underlying NEAT1 regulation of non-small-cell lung cancer (NSCLC) autophagy and apoptosis. The expression levels of NEAT1, miR-128-3p, and ADAM28 in NSCLC tissues and cells were examined using qRT-PCR. The relationships between NEAT1, miR-128-3p, and ADAM28 expression levels and prognosis of NSCLC patients were investigated using the Kaplan-Meier analysis method. The interactions between NEAT1, miR-128-3p, and ADAM28 were confirmed by dual-luciferase reporter assay or FISH assay. Cell proliferation and apoptosis were detected by CCK-8 and flow cytometry assays, respectively. Apoptosis and autophagy-related proteins Bax, cleaved caspase-3, cleaved caspase-9, Bcl-2, LC3, Beclin-1, and p62 were analyzed by western blotting. Finally, an in vivo NSCLC mouse xenograft model was established to confirm the in vitro data. We showed NEAT1 and ADAM28 expression levels were upregulated, while the miR-128-3p level was downregulated in NSCLC tissues and cells. NSCLC patients with high NEAT1 expression levels, low miR-128-3p levels, or high ADAM28 levels had significantly reduced overall survival times. Silencing of NEAT1 inhibited NSCLC cell autophagy and promoted apoptosis by sponging miR-128-3p. MiR-128-3p directly targeted ADAM28 and suppressed ADAM28 expression, which led to deactivation of the JAK2/STAT3 signaling pathway. Furthermore, ADAM28 overexpression attenuated miR-128-3p overexpression-induced apoptosis and autophagy inhibition in NSCLC by increasing the phosphorylation of JAK2 and STAT3. NEAT1 depletion or miR-128-3p overexpression inhibited autophagy and promoted apoptosis in vivo by suppressing ADAM28. In other word, silencing NEAT1 inhibited autophagy and then promoted NSCLC cell apoptosis by deactivating the JAK2/STAT3 signaling pathway through regulation of miR-128-3p/ADAM28 axis.

A peptide translated from circPPP1R12A promotes the malignancy of non-small cell lung cancer cells through AKT signaling pathway.

BackgroundRecent literature have indicated that the malignancy of cancer cells is modulated by hsa_circ_0000423 (named circPPP1R12A) through the way of translating protein. Herein, we investigated the role and latent mechanisms of circPPP1R12A in Non‐Small Cell Lung Cancer (NSCLC).MethodsCircPPP1R12A expression was measured by qRT‐PCR. The malignancy of NSCLC was determined by CCK‐8, TUNEL assay, Wound healing, Transwell and Western blotting assays. The underlying mechanisms of circPPP1R12A were confirmed by Western blotting and qRT‐PCR assays.ResultsCircPPP1R12A expression in NSCLC tissues was higher than that of neighboring normal tissues. CircPPP1R12A showed an upregulated expression in NSCLC cells. Upregulation of circPPP1R12A could promote the cell viability of NSCLC cells and reduce the apoptosis of NSCLC cells, while it could not promote cell invasion and migration. The reduction of cell viability and apoptosis was discovered in NSCLC cells with the silencing of circPPP1R12A, but circPPP1R12A knockdown does not inhibit cell invasion and migration. There was something interesting that circPPP1R12A encoding protein circPPP1R12A‐73aa was found in NSCLC cells. Mutations in circPPP1R12a‐73AA might disrupt the function of circPPP1ra‐73AA in A549 and H1299 cells. Next, we found that circPPP1R12A caused the increased growth of NSCLC cells by activating AKT signaling pathway.ConclusionIn summary, our study proved that NSCLC cell proliferation was promoted by circPPP1R12A‐73aa translated from circPPP1R12A through the AKT pathway, which could throw some light on the understanding of the mechanism of NSCLC.

Neferine, a novel ROCK1-targeting inhibitor, blocks EMT process and induces apoptosis in non-small cell lung cancer.

The compounds derived from Traditional Chinese Medicines have shown various pharmacological activities with unique advantages, especially in the aspect of antitumor. Neferine (Nef), a natural compound, extracted from green seed embryos of Lotus (Nelumbo nucifera Gaertn.) also exerts antitumor effects on cancers. In this study, the effects and mechanisms of Nef on epithelial-to-mesenchymal transition (EMT) process in non-small cell lung cancer (NSCLC) were evaluated. The results showed that Nef had the antitumor effects in vivo and in vitro. Nef significantly suppressed cell viability and induced apoptosis in NSCLC cells, with elevated reactive oxygen species and reduced BCL2/BAX ratio. Nef was also demonstrated to inhibit the invasion, metastasis and EMT process of NSCLC cells, and attenuate EMT-related changes of E-cadherin, N-cadherin and Vimentin at both transcriptional and translational levels. Moreover, we concluded that the inhibitory effects of Nef on EMT was achieved by targeting Rho-associated protein kinase 1, a protein mediating the process of EMT in various cancers. These results showed that Nef had a significant antitumor effect on NSCLC cells by inducing apoptosis and blocking EMT, providing the therapeutical prospect on NSCLC treatment.

The Ethanolic Extract of Trichosanthes Kirilowii Root Exerts anti-Cancer Effects in Human Non-Small Cell Lung Cancer Cells Resistant to EGFR TKI

The aim of this study was to investigate whether the ethanol extract of the Trichosanthes kirilowii root (ETK), traditionally used to treat lung diseases, exhibits anticancer activity in epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI)-resistant non-small cell lung cancer (NSCLC) cells. ETK treatment suppressed the growth of EGFR TKI-resistant NSCLC cells, including H1299, H1975, PC9/ER (erlotinib-resistant PC9) and PC9/GR (gefitinib-resistant PC9) cells, in a concentration- and time-dependent manner. Dose-dependent decline in anchorage-dependent and -independent colony formation was also detected following ETK treatment. We demonstrate that the growth-inhibitory effect of ETK was related to apoptosis induction, based on flow cytometry results showing ETK-induced increase in the percentage of cells with sub-G1 DNA and the population of annexin V-positive cells. Consistently, ETK induced chromatin condensation and cleavage of poly(ADP-ribose) polymerase (PARP). As a molecular mechanism, the phosphorylation level of signal transducer and activator of transcription 3 (STAT3) and Src was decreased by ETK. ETK-induced apoptosis was partially reversed by transfection of constitutively activated STAT3, indicating that STAT3 inactivation mediated ETK-induced apoptosis in EGFR TKI-resistant NSCLC cells. Our results provide basic evidence supporting the role of ETK as a novel therapeutic in EGFR TKI-resistant NSCLC.

CircRNA 0009043 suppresses non-small-cell lung cancer development via targeting the miR-148a-3p/DNAJB4 axis

BackgroundCircular RNAs (circRNAs) are important regulators of the development and progression of non-small-cell lung cancer (NSCLC) and many other malignancies. The functional importance of circ_0009043 in NSCLC, however, has yet to be established.MethodsThe expression of circ_0009043, miR-148a-3p, and DnaJ heat shock protein family (Hsp40) member B4 (DNAJB4) in NSCLC cells was assessed via qPCR. The proliferative activity of these cells was examined through EdU uptake and CCK-8 assays, while flow cytometry approaches were used to examine apoptotic cell death rates. Protein expression was measured through Western immunoblotting. Interactions between miR-148a-3p and circ_0009043 or DNAJB4 were detected through RNA immunoprecipitation (RIP) and dual-luciferase reporter assays. The in vivo importance of circ_0009043 as a regulator of oncogenic activity was assessed using murine xenograft models.ResultsBoth NSCLC cells and tissue samples were found to exhibit circ_0009043 upregulation, and lower circ_0009043 expression levels were found to be related to poorer NSCLC patient overall survival. Knocking down circ_0009043 resulted in the enhancement of NSCLC cell proliferative activity and the suppression of apoptotic tumor cell death in vitro, while also driving more rapid in vivo tumorigenesis. Mechanistically, circ_0009043 was found to function as a molecular sponge that sequestered miR-148a-3p, which was in turn able to directly suppress DNAJB4 expression. When miR-148a-3p was overexpressed, this reversed the impact of knocking down circ_0009043 on the apoptotic death and proliferation of NSCLC cells. Conversely, miR-148a-3p inhibition resulted in the suppression of NSCLC cell apoptosis and the enhancement of tumor cell growth, while the downregulation of DNAJB4 reversed these changes.ConclusionCirc_0009043 acts as a tumor suppressor in NSCLC cells, promoting DNAJB4 upregulation via the sequestration of miR-148a-3p.

The diagnostic value of has_circ_0006423 in non-small cell lung cancer and its role as a tumor suppressor gene that sponges miR-492

The diagnosis and treatment of non-small cell lung cancer (NSCLC) are not ideal. We identified NSCLC-related has_circ_0006423 in database. qRT-PCR was used to measure expression levels of hsa_circ_0006423 and miR-492 in the plasma and tissue samples, and 3 NSCLC cell lines, respectively. We analyzed the relationship between expression levels of hsa_circ_0006423 and clinicopathological factors and miR-492 expression in plasma and tissue samples. Assess the diagnostic value of hsa_circ_0006423 and miR-492 in NSCLC. Cell function vitro experiment to explore the effect of has_circ_0006423 on NSCLC. We found has_circ_0006423 is lower expressed in NSCLC and miR-492 is opposite, has_circ_0006423 and miR-492 has diagnostic value in NSCLC. In A549 and NCI-H1299 cells, hsa_circ_0006423 inhibited the proliferation, migration, and invasion of NSCLC cells by sponging miR-492 and accelerating NSCLC cell apoptosis. This effect may be due to the combination of has_circ_0006423 and miR-492 affecting the progression of NSCLC.

Knockdown of circLRWD1 weakens DDP resistance via reduction of SIRT5 expression through releasing miR-507 in non-small cell lung cancer.

Cisplatin (DDP) is an antineoplastic agent for non-small cell lung cancer (NSCLC). Hsa_circ_0081664 (circLRWD1) is overexpressed in DDP-resistant NSCLC cells, but its function is unclear. Thus, this study is to investigate whether circLRWD1 participates in DDP resistance in NSCLC. Changes in circLRWD1 expression were determined by real-time quantitative PCR. Effects of circLRWD1 inhibition on DDP-resistant NSCLC cell viability, proliferation, migration, invasion, and apoptosis were analyzed. The sponge function of circLRWD1 was predicted by bioinformatics analysis and verified by dual-luciferase reporter, RNA immunoprecipitation, and RNA pull-down assays. The function of circLRWD1 in DDP resistance was verified by xenograft models. CircLRWD1 was unconventionally overexpressed in DDP-resistant NSCLC samples and cells. Moreover, circLRWD1 silencing decreased IC 50 value, restrained cell proliferation, reduced cell migration and invasion, and facilitated cell apoptosis in DDP-resistant NSCLC cells. Also, circLRWD1 knockdown elevated DDP-resistant NSCLC cell sensitivity to DDP in xenograft models. Furthermore, circLRWD1 regulated SIRT5 expression via adsorbing miR-507. SIRT5 overexpression weakened circLRWD1 silencing-mediated suppression of cell resistance to DDP in DDP-resistant NSCLC cells. In conclusion, circLRWD1 elevated SIRT5 expression via adsorbing miR-507, resulting in promoting NSCLC cell resistance to DDP, providing evidence to explain the significant role of circLRWD1 in DDP resistance in NSCLC.

ShRNA‑mediated knockdown of KNTC1 inhibits non-small-cell lung cancer through regulating PSMB8

In view of the important roles played by Kinetochore proteins in mitosis, we believed that they may contribute to the development and progression of human cancers, which has been reported recently elsewhere. Kinetochore-associated 1 (KNTC1) participates in the segregation of sister chromatids during mitosis, the effects of which on non-small-cell lung cancer (NSCLC) remain unclear. Here, we sought to identify the biological significance of KNTC1 in NSCLC. KNTC1 protein expression in NSCLC tissues was investigated by immunohistochemistry. Lentivirus delivered short hairpin RNA (shRNA) was utilized to establish KNTC1 silence NSCLC cell lines. The effects of KNTC1 depletion on NSCLC cell proliferation, migration, apoptosis, and tumor formation were analyzed by MTT assay, wound-healing assay, transwell assay, flow cytometry assay, and in nude mouse models in vivo. After KNTC1 reduction, NSCLC cell viability, proliferation, migration, and invasion were restrained. A xenograft tumor model was also provided to demonstrate the inhibited tumorigenesis in NSCLC. In addition, the downstream mechanism analysis indicated that KNTC1 depletion was positively associated with PSMB8. The findings of the present study suggested that KNTC1 may have a pivotal role in mediating NSCLC progression and may act as a novel therapeutic target for NSCLC.

Circ_0001955 promotes the progression of non-small cell lung cancer via miR-769-5p/EGFR axis

ABSTRACT To elaborate on the role of circular RNA 0001955 (circ_0001955) on the proliferation and apoptosis of non-small cell lung cancer (NSCLC) cells and its underlying mechanism. Circ_0001955 expression in NSCLC was screened out through bioinformatics analysis based on GEO database. Circ_0001955, microRNA-769-5p (miR-769-5p), and epidermal growth factor receptor (EGFR) expression in NSCLC tissues and cell lines was examined using quantitative real-time PCR (qRT-PCR) and Western blot. Cell proliferation and apoptosis were examined using the CCK-8 method, BrdU experiment and flow cytometry analysis, respectively. Bioinformatics prediction, dual-luciferase reporter gene experiment and RNA immunoprecipitation (RIP) experiments were applied to validate the targeting relationship between miR-769-5p and circ_0001955 and the 3’ UTR of EGFR. Pearson’s correlation analysis was employed to validate the correlations among them. Circ_0001955 expression was up-regulated in NSCLC tissues and cell lines, and its overexpression was strongly associated with increased tumor TNM stage and lymph node metastasis. Circ_0001955 overexpression enhanced the proliferation and restrained the apoptosis in NSCLC cells, whereas knocking down circ_0001955 exerted the opposite effects. Circ_0001955 directly targeted miR-769-5p and negatively regulated its expression. EGFR, a target gene of miR-769-5p, could be indirectly and positively regulated by circ_0001955. Correlation analysis indicated that circ_0001955 was negatively correlated with miR-769-5p expression, while circ_0001955 was positively correlated with EGFR expression. Circ_0001955 facilitates the proliferation and represses the apoptosis of NSCLC cells by modulating miR-769-5p/EGFR axis.

Xanthotoxol suppresses non-small cell lung cancer progression and might improve patients’ prognosis

BackgroundDeveloping novel and effective drugs with less toxicity is urgent for non-small cell lung cancer (NSCLC) therapy. Xanthotoxol (Xan) is the major natural component of the medical plant Angelica dahurica with potential anti-cancer activities. PurposeIn this study, we aimed to demonstrate the effect and underlying mechanism of Xan in NSCLC and evaluate the effectiveness of Xan in NSCLC patients. MethodsCCK8, colony formation, EdU, flow cytometry, and transwell assays were carried out to investigate the anti-NSCLC activity of Xan in vitro. In addition, the xenograft mouse model was established to evaluate the anti-NSCLC effect of Xan in vivo. Moreover, bioinformatics analysis was performed to establish a prediction model based on RNA sequencing data. Furthermore, Western blot was used to detect the expression of proteins regulated by Xan. ResultsXan inhibited the cell viability, colony formation capacity, DNA replication, cell cycle transition, migration and invasion, as well as inducing apoptosis of NSCLC cells. In addition, Xan suppressed NSCLC xenograft growth in vivo without obvious toxicity. Interestingly, bioinformatics analyses based on the RNA sequencing data indicated that Xan exerted inhibitory effects on NSCLC cells by down-regulating signals contributing to NSCLC progression and demonstrated that Xan was effective in ameliorating the prognosis of NSCLC patients with a new proposed prediction model. Moreover, Xan was shown to regulate cell cycle arrest, apoptosis, and epithelial-mesenchymal transition (EMT)-associated genes through downregulating PI3K-AKT signaling, thus suppressing NSCLC proliferation and metastasis. ConclusionsTaken together, our work proved that Xan induced cell cycle arrest, facilitated apoptosis, and inhibited EMT processes through downregulating the PI3K-AKT pathway to suppress NSCLC progress. Moreover, we also proposed a new model for evaluating Xan as a novel and effective drug in NSCLC treatments.

MiR-630 suppresses non-small cell lung cancer by targeting vimentin.

ObjectiveThis study aimed to clarify the function of miR‐630 on non‐small cell lung cancer (NSCLC) cells.MethodsQuantitative real‐time PCR was utilized to detect the mRNA expression of miR‐630 and vimentin (VIM) in NSCLC tissues and cells. The protein expression of VIM, P53, Caspase‐3, Bcl‐2, Bax and JAK2/STAT3 was evaluated via Western blot. Dual‐luciferase reporter assay was applied to evaluate whether VIM is the target gene of miR‐630. The migration, invasion, proliferation and apoptosis of NSCLC cells were examined by wound‐healing assay, transwell assay, CCK‐8 assay, and flow cytometry, respectively.ResultsMiR‐630 was lowly expressed in NSCLC tissues and cells, while VIM was highly expressed in NSCLC cells. Dual‐luciferase reporter assay data validated that miR‐630 directly targeted VIM. MiR‐630 overexpression inhibited VIM expression, but the inhibition of miR‐630 upregulated VIM expression. Besides, miR‐630 mimics restrained cell migration, invasion, and proliferation, and promoted NSCLC cell apoptosis. Whereas, VIM overexpression partly attenuated the inhibitory effect of miR‐630 on NSCLC cells. Moreover, miR‐630 mimics impeded p‐JAK2 and p‐STAT3 protein expression; and miR‐630 inhibitor upregulated p‐STAT3 and VIM protein expression, which was reversed after the addition of STAT3 inhibitor C188‐9.ConclusionMiR‐630 constrained the progression of NSCLC by inhibiting JAK2/STAT3 pathway and downregulating VIM expression.

Fucoxanthin Inactivates the PI3K/Akt Signaling Pathway to Mediate Malignant Biological Behaviors of Non-Small Cell Lung Cancer

Although lung cancer treatment strategies have improved in recent years, the 5-year overall survival of non-small cell lung cancer (NSCLC) remains less than 15%. Chemotherapy is considered the most promising option in the comprehensive treatment of NSCLC. Fucoxanthin (FX) is a natural product derived from brown algae and has extensive applications in medicine. Previous studies reported that FX effectively inhibits the growth of NSCLC cells in vitro and in vivo. However, the mechanism underlying the anti-NSCLC effect of FX remains unknown. In this study, NSCLC cell lines and a xenograft nude mouse model were used to examine the anti-NSCLC activities of FX in vitro and in vivo. Network pharmacology analysis and inhibitors or activators of the PI3K/Akt signaling pathway were used to explore the anti-NSCLC mechanisms of FX. The results indicated that FX could inhibit proliferation, migration, and invasion, arrest cell cycle at the G0/G1 phase, and induce apoptosis of NSCLC cells in vitro. Additionally, FX suppressed tumor growth in vivo. The PI3K/Akt signaling pathway was found to be involved in the anti-NSCLC activity of FX. In conclusion, FX inhibits malignant biological behaviors of NSCLC by suppressing the phosphorylation of both PI3K and AKT, and subsequently inactivating PI3K/AKT signaling pathway.

Induction of Ferroptosis by Ophiopogonin-B Through Regulating the Gene Signature AURKA in NSCLC.

Ferroptosis is a new type of iron-dependent programmed cell death. In recent years, its role in the diagnosis and treatment of multiple tumors, including non-small cell lung cancer (NSCLC), has been continuously observed. The relationship between the ferroptosis-related genes and the prognosis of patients with NSCLC needs to be clarified. In this study, The Cancer Genome Atlas (TCGA) and the Gene Expression Synthesis database (Gene Expression Omnibus, GEO) were used to build a model of ferroptosis-related differentially expressed genes (DEGs). A total of 101 ferroptosis-related DEGs were screened using R language, and a 12-gene signature was finally established through univariate Cox regression analysis and least absolute shrinkage and selection operator (LASSO)-penalized Cox regression analysis. According to the risk scores, the patients were divided into a high-risk or a low-risk group, with patients in the low-risk group showing better prognosis. AURKA, one of the genes in the 12-gene signature, was found to be highly expressed in tumors. In addition, further study verified AURKA to be a negative regulator of ferroptosis in NSCLC cells. Ophiopogonin B (OP-B) had been reported to induce apoptosis, mitotic catastrophe, and autophagy in NSCLC cells. Herein, proteomic sequencing analysis and OP-B administration revealed the upregulation of AURKA and the downregulation of PHKG2 and SLC7A5 in the 12-gene signature, indicating that OP-B induced ferroptosis in NSCLC. Determination of the concentrations of malondialdehyde (MDA), glutathione (GSH), and intracellular iron and the mitochondrial membrane potential (MMP) confirmed the induction of ferroptosis by OP-B in vitro. Furthermore, transmission electron microscopy (TEM) examination of lung cancer xenotransplantation in nude mice confirmed that OP-B induced ferroptosis in vivo. Further study of the molecular mechanism showed that the ferroptosis effect caused by OP-B can be partially reversed by the overexpression of AURKA. Overall, our study established a new ferroptosis-related risk prediction model for the prognosis of patients with NSCLC, revealed the enrichment pathways of ferroptosis in NSCLC, and discovered the negative regulation of AURKA in ferroptosis. On this basis, we demonstrated that OP-B can induce ferroptosis in NSCLC and clarified the specific molecular mechanism of OP-B inducing ferroptosis by regulating the expression of AURKA.

CircDLG1 promotes malignant development of non-small cell lung cancer through regulation of the miR-630/CENPF axis.

Circular RNAs (circRNAs) have been reported to be crucial modulatory molecules in the etiology of non-small cell lung cancer (NSCLC). This study aimed to probe the precise role and mechanism of circRNA discs large MAGUK scaffold protein1 (circDLG1) in the malignant progression of NSCLC. The abundances of circDLG1, miR-630, and centromere proteinF (CENPF) mRNAs were gauged by quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation was tested in 3‑(4, 5‑dimethylthiazol-2-yl)-2, 5‑diphenyltetrazolium bromide (MTT) assay and 5‑ethynyl-2'-deoxyuridine (EdU)-incorporation assay. Cell apoptosis was analyzed by flow cytometry. Cell migration and invasion were assessed by transwell assay. Western blot was exploited to examine the levels of all proteins. The interaction between miR-630 and circDLG1 or CENPF was verified by dual-luciferase reporter, RNA pull-down, and/or RNA immunoprecipitation assays. Tumor xenograft assay and immunohistochemistry (IHC) were executed for the role of circDLG1 in tumor growth in vivo. CircDLG1 and CENPF were highly expressed in NSCLC, while miR-630 was downregulated. CircDLG1 silencing repressed proliferation, migration, and invasion, and expedited apoptosis of NSCLC cells in vitro. Mechanistically, circDLG1 deficiency modulated NSCLC cell malignant development through interacting with miR-630. Furthermore, CENPF was targeted by miR-630, and circDLG1 could positively control CENPF expression through acting as an miR-630 sponge. Furthermore, CENPF overexpression reversed the repressive impacts of circDLG1 inhibition in the malignant behaviors of NSCLC cells. Besides, circDLG1 interference hindered tumor growth in vivo. CircDLG1 knockdown could impede NSCLC advancement through modulating the miR-630/CENPF axis, manifesting as apromising molecular target for NSCLC treatment.