Poor Prognosis In Non-small Cell Lung Cancer Patients Research Articles

SLC12A8 promotes the migration and invasion of non-small cell lung cancer (NSCLC) cells.

This study aims to investigate the impacts of SLC12A8 on the invasion, migration, and epithelial-mesenchymal transition (EMT) of non-small cell lung cancer (NSCLC) cells. GEPIA database was employed to examine SLC12A8 expression pattern in lung cancer cells. Subsequently, qRT-PCR and Western blot analyses were conducted to assess SLC12A8 expression in NSCLC tissues and cell lines. The overall prognosis of NSCLC patients was evaluated using Kaplan-Meier plot and univariate and multivariate COX regression curves. The knockdown of SLC12A8 was established using lentivirus-mediated shRNA in A549 and H1299 cells. Cell proliferation, invasion, migration, and apoptosis were evaluated using CCK-8 assay, transwell, and flow cytometry techniques, respectively. Western blot analysis was performed to measure the expression levels of EMT-related proteins (E-cadherin and vimentin). The expression level of SLC12A8 was found to be significantly higher in both NSCLC cell lines and tissues. High SLC12A8 expression was correlated with a poor prognosis in NSCLC patients. Knocking down SLC12A8 led to a significant decrease in proliferation, migration, and invasion abilities, while promoting apoptosis in NSCLC cells. Additionally, SLC12A8 knockdown resulted in decreased levels of N-cadherin and vimentin, along with increased E-cadherin expression. The results indicate that reducing SLC12A8 expression may suppress the malignant phenotype of NSCLC cells, as well as the EMT. SLC12A8 may serve as a target for the clinical management of NSCLC progression.

The COP9 signalosome stabilized MALT1 promotes Non-Small Cell Lung Cancer progression through activation of NF-κB pathway

MALT1 has been implicated as an upstream regulator of NF-κB signaling in immune cells and tumors. This study determined the regulatory mechanisms and biological functions of MALT1 in non-small cell lung cancer (NSCLC). In cell culture and orthotopic xenograft models, MALT1 suppression via gene expression interference or protein activity inhibition significantly impaired malignant phenotypes and enhanced radiation sensitivity of NSCLC cells. CSN5, the core subunit of COP9 signalosome, was firstly verified to stabilize MALT1 via disturbing the interaction with E3 ligase FBXO3. Loss of FBXO3 in NSCLC cells reduced MALT1 ubiquitination and promoted its accumulation, which was reversed by CSN5 interference. An association between CSN5/FBXO3/MALT1 regulatory axis and poor prognosis in NSCLC patients was identified. Our findings revealed the detail mechanism of continuous MALT1 activation in NF-κB signaling, highlighting its significance as predictor and potential therapeutic target in NSCLC.Graphical

DRD1 suppresses cell proliferation and reduces EGFR activation and PD-L1 expression in NSCLC.

Dopamine (DA) acts in various key neurological and physiological processes as both a neurotransmitter and circulating hormone. Over the past several decades, the DA signaling network has been shown to regulate the progression of several types of solid tumors, and considerable evidence has shown it is a druggable pathway in the cancer cell context. However, the specific activity and effect of these pathway components appears to be tissue-type and cell-context-dependent. In the present study, expression and methylation of dopamine receptor D1 (DRD1) were measured using RNA sequencing (RNAseq) and reverse transcription polymerase chain reaction (RT-PCR) in non-small cell lung cancer (NSCLC) samples, and validated using publicly available datasets, including The Cancer Genome Atlas (TCGA). In vitro and in vivo functional experiments were performed for cell proliferation and tumor growth, respectively. Mechanistic analyses of the transcriptome and kinome in DRD1-modulated cells informed further experiments, which characterized the effects on the epidermal growth factor receptor (EGFR) pathway and programmed cell death 1 ligand 1 (PD-L1) proteins. Through these experiments, we identified the DRD1 gene as a negative regulator of disease progression in NSCLC. We show that DRD1, as well as other DA pathway components, are expressed in normal human lung tissue, and that loss of DRD1 expression through promoter hypermethylation is a common feature in NSCLC patients and is associated with worse survival. At the cellular level, DRD1 affects proliferation by inhibiting the activation of EGFR and mitogen-activated protein kinase 1/2 (ERK1/2). Interestingly, we also found that DRD1 regulates the expression of PD-L1 in lung cancer cells. Taken together, these results suggest that DRD1 methylation may constitute a biomarker of poor prognosis in NSCLC patients while other components of this pathway could be targeted to improve response to EGFR- and PD-L1-targeted therapies.

ITGAL expression in non-small-cell lung cancer tissue and its association with immune infiltrates.

Integrin subunit alpha L (ITGAL) encodes an integrin component of LFA-1 and is a membrane receptor molecule widely expressed on leukocytes. It plays a key role in the interaction between white blood cells and other cells. There was a significant correlation between the expression of ITGAL and the tumor microenvironment in a number of cancers. However, experimental studies targeting ITGAL and immune cell infiltration in non-small-cell lung cancer (NSCLC) and the response to immune checkpoint inhibitor therapy are lacking. Data were obtained from The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), and Clinical Proteomic Tumor Analysis Consortium (CPTAC) databases to explore the relationship between ITGAL expression and prognosis, as well as the immune cell infiltration in patients with NSCLC. In addition, immunohistochemical staining for ITGAL and multiplex immunofluorescence (mIF) staining for ITGAL, CD20, CD68, CD4, and CD8 from tissue microarrays containing 118 tumor tissues and paired paracancerous tissues from patients with NSCLC were performed. The correlation between ITGAL expression and clinical factors, as well as the immunophenotypes of tumor-infiltrating immune cells, were also analyzed. In NSCLC tumor tissues, ITGAL was downregulated compared with matched paracancerous tissues, and low ITGAL expression was associated with a poor prognosis of NSCLC patients. Subsequently, immunohistochemistry results for tissue microarray showed that ITGAL expression was mainly elevated in tumor stroma and areas with highly infiltrated immune cells. ITGAL expression was higher in paracancerous tissues than tumor tissues. Furthermore, mIF results indicated that the patients with ITGAL-high expression tend had significantly higher CD8+ T cells, CD68+ macrophages, CD4+ T cells, and CD20+ B cells infiltration in their tumor tissues. Immunophenotypes were classified into three categories, that is deserted, excluded, and inflamed types, according to each kind of immune cell distribution in or around the cancer cell nest. MIF results showed that ITGAL expression level was correlated with the immunophenotypes. Furthermore, ITGAL expression was associated with the prognosis of NSCLC in patients with immune checkpoint inhibitor therapy and the patients with high ITGAL expression tends have better outcomes. ITGAL may be used as a biomarker for assessing the immune microenvironment in patients with NSCLC.

Elevation of LEM Domain Containing 1 Predicts Poor Prognosis of NSCLC Patients and Triggers Malignant Stemness and Invasion of NSCLC Cells by Stimulating PI3K/AKT Pathway.

Non-small cell lung cancer (NSCLC) is a leading cause of cancer-related death globally. LEM domain containing 1 (LEMD1) function has been identified in several cancers but not in NSCLC. This study aimed to investigate the LEMD1 function in NSCLC. NSCLC tissues were obtained from 66 patients, and LEMD1 expressions were measured using quantitative real-time PCR, immunohistochemical assay, and Western blot. Overall survival of NSCLC patients was estimated by the Kaplan-Meier method. Meanwhile, LEMD1 function and mechanism were assessed using Cell Counting Kit-8, 5-Ethynyl-2'-deoxyuridine analysis, Transwell, Sphere formation assay, and flow cytometry. Furthermore, LEMD1 function in vivo was evaluated by establishing a xenograft tumor model, hematoxylin-eosin staining, and immunohistochemical assay. LEMD1 was highly expressed in NSCLC tissues and was interrelated to tumor differentiation, TNM stage, and lymph node metastasis of patients. Overall survival of NSCLC patients with high LEMD1 was found to be lower than that of patients with low LEMD1. Functionally, interference with LEMD1 restrained NSCLC cell proliferation, invasion, and stemness characteristics. Mechanistically, LEMD1 facilitated the malignant phenotype of NSCLC, and 740 Y-P reversed this impact, prompting that LEMD1 aggravated NSCLC by activating PI3K/AKT pathway. Furthermore, LEMD1 knockdown hindered NSCLC proliferation in vivo. Conclusion: LEMD1 accelerated NSCLC cell proliferation, invasion, and stemness characteristics via activating PI3K/AKT pathway.

Kinetochore scaffold 1 downregulation suppressed the development of non-small cell lung cancer by inactivating the phosphatidylinositol 3 kinase/protein kinase B (AKT)/nuclear factor-kappa B pathway.

Kinetochore scaffold 1 (KNL1) is indispensable for generating motile micro-tubule attachments and isolating chromosomes. KNL1 is highly expressed in multiple middle-route tissues and promotes tumor development. However, how it functions in non-small cell lung cancer (NSCLC) is unclear. Real-time quantitative PCR (RT-qPCR) and Western blotting (WB) were used to determine KNL1 expression in NSCLC tissues and cells. The sh-KNL1 or oe-KNL1 was transfected into NSCLC cells. The colony formation assay, cell counting kit-8 (CCK-8) assay, and flow cytometry were used to evaluate cell proliferation and apoptosis. A transwell assay was used to monitor invasion and migration. The CCK-8 assay was used to measure NSCLC cell sensitivity to chemotherapy drugs. WB confirmed the protein levels of apoptosis-related proteins, cell cycle-associated proteins, and the phosphatidylinositol 3 kinase (PI3K)/protein kinase B (AKT)/nuclear factor kappaB (NF-κB) pathway. A PI3K/AKT/NF-κB pathway inhibitor was used to intervene in NSCLC cell transfection along with oe-KNL1, thus revealing the function of the pathway in carcinogenicity mediated by KNL1. In result KNL1 expression was substantially increased in NSCLC tissues and cells. High-level KNL1 expression is related to the poor prognosis of NSCLC patients. KNL1 silencing bolstered promoted NSCLC cell apoptosis and inhibited proliferation, cell cycle progression, invasion, and EMT, whereas KNL1 silencing had the opposite effect. KNL1 knockdown increased NSCLC cell sensitivity to chemical drugs. KNL1 promoted PI3K/AKT/NF-κB pathway activation, while PI3K/AKT/NF-κB pathway inhibition weakened the procancer effect mediated by KNL1 overexpression but had little influence on KNL1 levels. We conclude that KNL1 activates the PI3K/AKT/NF-κB pathway to increase NSCLC progression and attenuate NSCLC sensitivity to chemotherapy drugs.

ALKBH5 regulates paclitaxel resistance in NSCLC via inhibiting CEMIP-mediated EMT

N6-methyladenosine (m6A) is the most prevalent mRNA modification, and it is verified to be closely correlated with cancer occurrence and progression. The m6A demethylase ALKBH5 (alkB homolog 5) is dysregulated in various cancers. However, the role and underlying mechanism of ALKBH5 in the pathogenesis and especially the chemo-resistance of non-small cell lung cancer (NSCLC) is poorly elucidated. The current study shows that ALKBH5 expression is reduced in paclitaxel (PTX) resistant NSCLC cells and down-regulation of ALKBH5 usually implies poor prognosis of NSCLC patients. Over-expression of ALKBH5 in PTX-resistant cells can suppress cell proliferation and enhance chemo-sensitivity, while knockdown of ALKBH5 exerts the opposite effect, which further supports the tumor suppressive role of ALKBH5. Over-expression of ALKBH5 can also reverse the epithelial-mesenchymal transition (EMT) process in PTX-resistant cancer cells. Mechanistically, data from RNA-seq, real-time PCR and western blotting indicate that CEMIP (cell migration inducing hyaluronidase 1), also known as KIAA1199, may be the downstream target of ALKBH5. Furthermore, ALKBH5 negatively regulates the CEMIP level by reducing the stability of CEMIP mRNA. Collectively, the current data demonstrate that the ALKBH5/CEMIP axis modulates the EMT process in NSCLC, which in turn regulates the chemo-sensitivity of cancer cells to PTX.

Hsa_circ_0022383 promote non-small cell lung cancer tumorigenesis through regulating the miR-495-3p/KPNA2 axis

Hsa_circ_0022383 (circ_0022383) is a newly discovered circRNA. Its functions and relevant molecular mechanisms in tumorigenesis have not been reported. Here we aimed to explore how circ_0022383 regulates the tumorigenesis of non-small-cell lung cancer (NSCLC). We found thatcirc_0022383 expression was dramatically elevated in NSCLC tissues and cell lines. Upregulation of circ_0022383 was associated with poor prognosis in NSCLC patients. Silencing of circ_0022383 repressed cell proliferation and migration in vitro and inhibited oncogenesis and tumor metastasis in vivo. Moreover, our results discovered that circ_0022383 was mainly located in the cytoplasm of NSCLC cells. Mechanistically, circ_0022383 sponged miR-495-3p to modulate KPNA2 expression, thereby regulating NSCLC tumorigenesis and progression. In conclusion, our study demonstrates that circ_0022383 facilitates NSCLC tumorigenesis by regulating the miR-495-3p/KPNA2 axis, providing new insights into NSCLC development.

Inhibition of PKA/CREB1 pathway confers sensitivity to ferroptosis in non-small cell lung cancer

Ferroptosis is a type of regulated cell death characterized by iron accumulation and lipid peroxidation. The molecular mechanisms underlying ferroptosis regulation in non-small cell lung cancer (NSCLC) are poorly understood. In this study, we found that protein kinase A (PKA) inhibition enhanced ferroptosis susceptibility in NSCLC cells, as evidenced by reduced cell viability and increased lipid peroxidation. We further identified cAMP-responsive element protein 1 (CREB1), a transcription factor and a substrate of PKA, as a key regulator of ferroptosis. Knockdown of CREB1 sensitized NSCLC cells to ferroptosis inducers (FINs) and abolished the effects of PKA inhibitor and agonist, revealing the pivotal role of CREB1 in ferroptosis regulation. Using a high-throughput screening approach and subsequent validation by chromatin immunoprecipitation (ChIP) and dual-luciferase assays, we discovered that CREB1 transcriptionally activated stearoyl-CoA desaturase (SCD), an enzyme that catalyzes the conversion of saturated fatty acids to monounsaturated fatty acids. SCD conferred ferroptosis resistance by decreasing the availability of polyunsaturated fatty acids for lipid peroxidation, and its overexpression rescued the effect of CREB1 knockdown on ferroptosis in vitro. Besides, CREB1 knockdown suppressed xenograft tumor growth in the presence of Imidazole Ketone Erastin (IKE), a potent FIN, and this effect was reversed by SCD. Finally, we showed that high expression of CREB1 was associated with poor prognosis in NSCLC patients from public datasets and our institution. Collectively, this study illustrates the effect of PKA/CREB1/SCD axis in regulating ferroptosis of NSCLC, targeting this pathway may provide new strategies for treating NSCLC patients.

Targeting Immunosuppressive Myeloid Cells and Exhausted CD8+ T Cells Overcomes Radioresistance in NSCLC

Although stereotactic body radiation therapy (SBRT) has achieved great success in the treatment of non-small cell lung cancer (NSCLC), local relapses still occur and abscopal effects are rarely seen even when combined with immune checkpoint blockers (ICBs). Therefore, it is necessary to thoroughly understand the immune responses after SBRT. We characterized the dynamic changes of tumor-infiltrating immune cells at early and late time points after SBRT in a therapy-resistant murine tumor model using single-cell transcriptomes and T-cell receptor sequencing. At the early stage, the innate and adaptive immune systems were activated, including activation of NKs and NKTs, and infiltration of cytotoxic CD8+ T cells. At the late stage, however, the tumor immune microenvironment (TIME) shifted into immunosuppressive properties, containing enrichment of immunosuppressive tumor-associated neutrophils (TANs), M2-like tumor-associated macrophages (TAMs), and terminal exhausted CD8+ T cells. Furthermore, our study revealed that inhibition of CD39 combined with SBRT preferentially reinvigorated exhausted CD8+ T cells and promoted their proliferation, infiltration, and cytotoxicity. Meanwhile, it also promoted M1-like macrophage infiltration and DCs maturation. On the other hand, consequently increased infiltration of immunosuppressive myeloid cells after SBRT could be a potential mechanism mediating CD8+ T cell dysfunction. Moreover, we found that combination treatment with anti-VISTA and SBRT synergistically reduced immunosuppressive myeloid cells, containing TANs, M-MDSCs, and M2-like TAMs, and further activated CD8+ T cells. Clinically, high VISTA expression was associated with poor prognosis in NSCLC patients. Altogether, our data provides deep insight into acquired resistance to SBRT from an immune perspective and presents rational combination strategies.

LINC00921 Diminishes Lung Cancer Radiosensitivity by Bestabilizing NUDT21 and Driving Aberrant MED23 Alternative Polyadenylation

Alternative polyadenylation (APA) plays a major role in controlling transcriptome diversity and therapeutic resistance of cancers. However, long-noncoding RNAs (lncRNAs) involved in pathological APA remain poorly defined. Here, we functionally identified a MED13L/P300-induced oncogenic lncRNA, LINC00921, diminished lung cancer radiosensitivity by destabilizing NUDT21 and driving aberrant MED23 alternative polyadenylation. ChIP-seq screening, RNA-seq and real-time PCR were used to identified LINC00921 in NSCLC. We performed RNA pulldown, RIP-qPCR, western blotting and Co-immunoprecipitation to investigate the function of LINC00921, which induced destabilization of NUDT21 and promoted 3' UTR shortening of MED23 via APA. Through H3K27ac ChIP-seq screening, we functionally characterize LINC00921, a MED13L/P300-induced oncogenic lncRNA, required for global regulation of APA in non-small-cell lung cancer (NSCLC). LINC00921 shows significant potential for reducing radiosensitivity of NSCLC and high LINC00921 levels were associated with poor prognosis for NSCLC patients treated with radiotherapy. Mechanistically, LINC00921 directly interacts withNUDT21 via binding to its RNA-binding motif-2. LINC00921 controls NUDT21 stability via facilitating binding of NUDT21 with its newly identified E3 ligase TRIP12. Intriguingly, 3'UTR APA profiles reveal that LINC00921-induced destabilization of NUDT21 decreases the percentage of distal polyadenylation sites (PAS) usage index, resulting in the 3' UTR shortening of MED23 mRNA, which, in turn, leads to elevated MED23 protein levels in cancer cells. MED23 further increases nuclear translocation of β-catenin, and, thereby, activates expression of multiple β-Catenin/TCF/LEF-regulated core oncogenes (c-Myc, CCND1, and BMP4). Taken together, our data revealed a novel model that integrates a lncRNA into regulation of malignant APA, radiotherapy resistance and NSCLC progression. These findings highlight the importance of functionally annotating lncRNAs controlling APA and unlock the clinical potential of novel therapeutics for advanced NSCLC.

METTL14 drives growth and metastasis of non-small cell lung cancer by regulating pri-miR-93-5p maturation and TXNIP expression.

Non-small cell lung cancer (NSCLC) is a prevalent and aggressive malignancy responsible for a significant number of cancer-related deaths worldwide. Unraveling the molecular mechanisms governing NSCLC growth and metastasis is crucial for the identification of novel therapeutic targets and the development of effective anti-cancer strategies. One such mechanism of interest is the involvement of METTL14, an RNA methyltransferase implicated in various cellular processes, in NSCLC progression. The objective of this study was to investigate the role of METTL14 in NSCLC development and metastasis and to elucidate the underlying molecular mechanisms. By understanding the impact of METTL14 on NSCLC pathogenesis, the study aimed to identify potential avenues for targeted therapies in NSCLC treatment. We used bioinformatics and high-throughput transcriptome sequencing analyses to screen regulatory mechanisms affecting NSCLC. The Kaplan-Meier method assessed the correlation between METTL14 expression and the prognosis of NSCLC patients. The effects of manipulated METTL14 on malignant phenotypes of NSCLC cells were examined by colony formation assay, flow cytometry, scratch assay, and Transwell assay. The tumorigenic capacity and metastatic potential of NSCLC cells in vivo were evaluated in nude mice. METTL14 was overexpressed in NSCLC tissues and cell lines. Its high expression indicated a poor prognosis for NSCLC patients. METTL14 silencing promoted apoptosis and repressed proliferation, migration, and invasion of NSCLC cells. miR-93-5p targeted and inhibited TXNIP. METTL14 increased miR-93-5p expression and matured pri-miR-93-5p through m6A alteration to inhibit TXNIP, thereby inhibiting NSCLC cell apoptosis. By controlling the miR-93-5p/TXNIP axis, METTL14 increased the tumorigenic potential and lung metastasis of NSCLC cells in nude mice. This study revealed a role for METTL14 in the contribution to NSCLC development and metastasis and identified METTL14 as a potential target for NSCLC treatment.

USP12 promotes nonsmall cell lung cancer progression through deubiquitinating and stabilizing RRM2.

RRM2 is the catalytic subunit of ribonucleotide reductase (RNR), which catalyzes de novo synthesis of deoxyribonucleotide triphosphates (dNTPs) and plays critical roles in cancer cell proliferation. RRM2 protein level is controlled by ubiquitination mediated protein degradation system; however, its deubiquitinase has not been identified yet. Here we showed that ubiquitin-specific peptidase 12 (USP12) directly interacts with and deubiquitinates RRM2 in non-small cell lung cancer (NSCLC) cells. Knockdown of USP12 causes DNA replication stress and retards tumor growth in vivo and in vitro. Meanwhile, USP12 protein levels were positively correlated to RRM2 protein levels in human NSCLC tissues. In addition, high expression of USP12 was associated with poor prognosis in NSCLC patients. Therefore, our study reveals that USP12 is a RRM2 regulator and targeting USP12 could be considered as a potential therapeutical strategy for NSCLC treatment.

Overexpression of LINC00607 inhibits cell growth and aggressiveness by regulating the miR-1289/EFNA5 axis in non-small-cell lung cancer.

Long non-coding RNAs (lncRNAs) play a key role in cancer progression, including non-small-cell lung cancer (NSCLC). LncRNA long intergenic non-protein-coding RNA 00607 (LINC00607) was previously discovered to be downregulated in lung adenocarcinoma tissues. Nevertheless, the potential role of LINC00607 in NSCLC is still unclear. The expression of LINC00607, miR-1289, and ephrin A5 (EFNA5) in NSCLC tissues and cells was tested by reverse transcription quantitative polymerase chain reaction. Cell viability, proliferation, migration, and invasion were measured by 3-(4,5-dimethylthiazole-2-y1)-2,5-diphenyl tetrazolium bromide, colony formation, wound healing, and Transwell assays. The relationship among LINC00607, miR-1289, and EFNA5 in NSCLC cells was verified by the luciferase reporter assay, RNA pull-down assay, and RNA immunoprecipitation assay. In this study, LINC00607 was downregulated in NSCLC, and its low level is associated with poor prognosis of NSCLC patients. Furthermore, LINC00607 overexpression repressed NSCLC cell viability, proliferation, migration, and invasion. LINC00607 bound with miR-1289 in NSCLC. EFNA5 was a downstream target of miR-1289. EFNA5 overexpression also inhibited NSCLC cell viability, proliferation, migration, and invasion. EFNA5 knockdown antagonized the influence of LINC00607 overexpression on NSCLC cell phenotypes. Overall, LINC00607 serves as a tumor suppressor gene in NSCLC through binding with miR-1289 and modulating the level of EFNA5.

Genomics-based screening of novel therapeutic targets and biomarkers for lung cancer.

e15093 Background: Since the number of lung cancer patients who show objective response to standard treatments is still limited, development of new anti-cancer agents with minimum risk of side effects and specific cancer biomarkers is urgently required. Methods: We have been screening new therapeutic targets and their companion biomarkers for lung cancer as follows; i) To identify overexpressed genes in lung cancers by the gene expression profile analysis, ii) To verify the candidate genes for their scarce expression in normal tissues, iii) To validate the clinicopathological importance of their protein expression by tissue microarray covering hundreds of lung cancers, and iv) To characterize their function for the growth and/or survival of lung cancer cells by siRNAs. Results: We identified dozens of candidate oncoproteins and selected a lung cancer cell division-associated protein 1 (LCDAP1). Immunohistochemical analysis showed that LCDAP1 was expressed in 171 of 267 (64%) non-small cell lung cancer (NSCLC) tissues but not in normal lung epithelia. LCDAP1 expression was significantly associated with poor prognosis for NSCLC patients. To investigate its function, we screened LCDAP1-binding protein in lung cancer cells and found an interaction of LCDAP1 with a member of 5'-to-3' DNA helicase that was also up-regulated in lung cancer cells. Knockdown of either of LCDAP1 or its binding partner expression with siRNAs significantly inhibited the growth of cancer cells. Conclusions: Our genomics-based screening could be useful to develop diagnostic and prognostic biomarkers as well as therapeutic targets for small molecules, monoclonal antibodies, and nucleic acid drugs against lung cancer.

LncRNA GAS5 regulated by FTO-mediated m6A demethylation promotes autophagic cell death in NSCLC by targeting UPF1/BRD4 axis.

Long non-coding RNA (lncRNA) growth arrest-specific transcript 5 (GAS5) has been shown to be a regulator for many cancers, including non-small cell lung cancer (NSCLC). Therefore, its role and mechanism in the process of NSCLC deserve to be further revealed. The expression levels of GAS5, fat mass and obesity-associated protein (FTO) and bromodomain-containing protein 4 (BRD4) were detected by quantitative real-time PCR. Western blot analysis was used to examine the protein expression of FTO, BRD4, up-frameshift protein 1 (UPF1) and autophagy-related markers. Methylated RNA immunoprecipitation was used to assess the m6A level of GAS5 regulated by FTO. Cell proliferation and apoptosis were determined using MTT assay, EdU assay and flow cytometry. Autophagy ability was assessed by immunofluorescence staining and transmission electron microscope. Xenograft tumor model was constructed to explore the effects of FTO and GAS5 on NSCLC tumor growth in vivo. The interaction between UPF1 and GAS5 or BRD4 was confirmed by pull-down assay, RIP assay, dual-luciferase reporter assay, and chromatin immunoprecipitation. Fluorescent in situ hybridization was used to analyze the co-localization of GAS5 and UPF1. Actinomycin D treatment was employed to evaluate BRD4 mRNA stability. GAS5 was downregulated in NSCLC tissues and was associated with poor prognosis in NSCLC patients. FTO was highly expressed in NSCLC, and it inhibited GAS5 expression by reducing GAS5 m6A methylation level. GAS5 suppressed by FTO could promote the autophagic death of NSCLC cells in vitro and inhibit NSCLC tumor growth in vivo. In addition, GAS5 was able to interact with UPF1 to reduce the mRNA stability of BRD4. Knockdown of BRD4 reversed the inhibition of GAS5 or UPF1 silencing on the autophagic cell death of NSCLC. The findings of the study showed that lncRNA GAS5 mediated by FTO could contribute to the autophagic cell death of NSCLC by interacting with UPF1 to reduce BRD4 mRNA stability, suggesting that GAS5 might be a vital therapy target for NSCLC progression.

Critical Role of the Sulfiredoxin-Peroxiredoxin IV Axis in Urethane-Induced Non-Small Cell Lung Cancer.

Non-small cell lung cancer (NSCLC), the most common type of lung cancer, etiologically associates with tobacco smoking which mechanistically contributes to oxidative stress to facilitate the occurrence of mutations, oncogenic transformation and aberrantly activated signaling pathways. Our previous reports suggested an essential role of Sulfiredoxin (Srx) in promoting the development of lung cancer in humans, and was causally related to Peroxiredoxin IV (Prx4), the major downstream substrate and mediator of Srx-enhanced signaling. To further explore the role of the Srx-Prx4 axis in de novo lung tumorigenesis, we established Prx4-/- and Srx-/-/Prx4-/- mice in pure FVB/N background. Together with wild-type litter mates, these mice were exposed to carcinogenic urethane and the development of lung tumorigenesis was evaluated. We found that disruption of the Srx-Prx4 axis, either through knockout of Srx/Prx4 alone or together, led to a reduced number and size of lung tumors in mice. Immunohistological studies found that loss of Srx/Prx4 led to reduced rate of cell proliferation and less intratumoral macrophage infiltration. Mechanistically, we found that exposure to urethane increased the levels of reactive oxygen species, activated the expression of and Prx4 in normal lung epithelial cells, while knockout of Prx4 inhibited urethane-induced cell transformation. Moreover, bioinformatics analysis found that the Srx-Prx4 axis is activated in many human cancers, and their increased expression is tightly correlated with poor prognosis in NSCLC patients.

WDR74 rs11231247 contributes to the susceptibility and prognosis of non-small cell lung cancer.

WD repeat-containing protein 74 (WDR74) has been linked with the development of lung cancer. This study aims to investigate the relationship between WDR74 rs11231247 and non-small cell lung cancer (NSCLC) susceptibility and the prognosis of NSCLC patients. UALCAN, MethPrimer, ensembl and Pancan meQTL databases were used for bioinformatics analysis. The case-control study included 462 NSCLC patients and 462 health controls. WDR74 rs11231247 genotype was determined by PCR-RFLP. Logistic regression model was used to calculate odds ratio (OR) and 95% confidence interval (95% CI) for analyzing the association of WDR74 SNP with the risk of NSCLC. Log-rank test and Cox regression analysis were used to evaluate the effect of WDR74 genetic variation on the prognosis of NSCLC. Compared with normal tissues, WDR74 expression level was higher and methylation level was lower in LUAD tissues. There were two CpG islands presented in the promoter of WDR74. And rs11231247 was in the second CpG island. We then discovered that rs11231247 CC and CT were more likely modified by methylation. LUAD case-control study demonstrated that rs11231247 CC genotype was associated with NSCLC risk with OR (95%CI) of 5.29 (2.59-10.79). Stratified analysis showed that rs11231247 T>C polymorphism could increase NSCLC risk in younger subjects (age≤58) (OR = 1.64, 95%CI = 1.06-2.54, P=0.027). Survival analysis and Cox regression analysis showed rs11231247 CC genotype contributed to a poor prognosis of NSCLC patients (MST=21, HR=2.09, 95%CI=1.17-3.75). WDR74 rs11231247 polymorphism affected the risk and prognosis of NSCLC.

Polo-like kinase (PLK) 5, a new member of the PLK family, serves as a biomarker to indicate anabatic tumor burden and poor prognosis for resectable non-small cell lung cancer.

A review argues that polo-like kinase 5 (PLK5) may be linked to unfavorable prognosis in non-small cell lung cancer (NSCLC) patients, which contradicts the discoveries from The Human Protein Atlas database (derived from TCGA analysis). This study intended to comprehensively confirm the association of PLK5 with clinical characteristics and prognosis in NSCLC patients. This two-center, retrospective, cohort study enrolled 210 NSCLC patients receiving surgical resection. PLK5 protein and mRNA were detected by immunohistochemistry and RT-qPCR in tumor and nontumor tissues. Moreover, RNA FPKM data for 994 lung cancer patients were obtained from The Human Protein Atlas database. PLK5 protein was decreased in tumor tissue compared to nontumor tissue (P < 0.001). Additionally, decreased PLK5 protein was linked with increased pathological grade (P = 0.002), lymph node metastasis presence (P = 0.001), elevated tumor-node-metastasis (TNM) stage (P = 0.003), and abnormal cancer antigen 125 (CA125) (P = 0.002). Meanwhile, low PLK5 protein was correlated with shortened disease-free survival (DFS) (P = 0.007) and overall survival (OS) (P = 0.038); further multivariable Cox regression analysis revealed that low PLK5 protein independently predicted unfavorable DFS (hazard ratio = 0.573, P = 0.022). PLK5 mRNA was reduced in tumor tissue compared with nontumor tissue (P < 0.001); its decline was linked with enhanced pathological grade (P = 0.034), climbed TNM stage (P = 0.032), and abnormal CA125 (P = 0.002). Furthermore, low PLK5 mRNA was correlated with unfavorable DFS (P = 0.046). The Human Protein Atlas database also disclosed the link between low PLK5 mRNA and worse OS (P = 0.046). A PLK5 decrement reflects anabatic tumor burden and poor prognosis in NSCLC patients.

CircSCN8A suppresses malignant progression and induces ferroptosis in non-small cell lung cancer by regulating miR-1290/ACSL4 axis

ABSTRACT Circular RNAs (CircRNAs) are reported to exert vital regulatory roles in the occurrence and development of various human malignancies, including non-small cell lung cancer (NSCLC). Bioinformatics methods identified the down-regulation of circSCN8A (circBase ID: hsa_circ_0026337) in NSCLC tissues. However, its biological functions and molecular mechanisms in NSCLC remain unknown. In this study, we found that circSCN8A expression was down-regulated in NSCLC tissues and cells. Low circSCN8A expression was positively associated with aggressive clinicopathological characteristics and poor prognosis in NSCLC patients. CircSCN8A suppressed cell proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) in vitro and blocked tumor growth in vivo. Moreover, circSCN8A promoted cell ferroptosis in NSCLC. Mechanistically, circSCN8A acted as a competing endogenous RNA (ceRNA) by sponging miR-1290 to enhance the expression of long-chain acyl-CoA synthetase-4 (ACSL4). Furthermore, the knockdown of ACSL4 or overexpression of miR-1290 reversed the effect of circSCN8A on facilitating ferroptosis and inhibiting cell proliferation and metastasis. In summary, circSCN8A represses cell proliferation and metastasis in NSCLC by regulating the miR-1290/ACSL4 axis to induce ferroptosis. Thus, circSCN8A may represent a promising therapeutic target against NSCLC.