Apoptosis In Lung Adenocarcinoma Cells Research Articles

MLLT3 knockdown suppresses proliferation and cell mobility in human lung adenocarcinoma.

Lung cancer is emerging as one of the most frequently encountered malignancies around the world that carries high morbidity and mortality. Lung adenocarcinoma (LUAD) has become the most common subtype of lung cancer. MLLT3 or named AF9 was first characterized in acute myeloid leukemia and can downregulate the expression of several critical genes. The aim of this study was to explore the function of MLLT3 in the progression of LUAD and related molecular mechanisms. Immunohistochemistry was employed to assess MLLT3 expression in LUAD tissues, while quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot were utilized to detect MLLT3 expression levels in lung adenocarcinoma cell lines. These results revealed a significant overexpression of MLLT3 in LUAD cell lines and tissues. We further uncovered an association between MLLT3 expression profiles in LUAD tissues and metastasis, as well as TNM stage. Survival analysis showed that elevated MLLT3 correlated with a poorer survival rate. We also found that MLLT3 knockdown repressed LUAD cells invasion, migration, and proliferation in vitro, while inducing cell cycle arrest and apoptosis of LUAD cells. Moreover, knocking down MLLT3 inhibited tumor growth in vivo. Specific markers of apoptosis, cell cycle, epithelial-mesenchymal transition (EMT), and MLLT3-induced signaling were examined by Western blot. We demonstrated that MLLT3 knockdown inhibited the activity of the EGFR-MAPK/ERK signaling pathway, and MLLT3 might be a novel diagnostic biomarker and therapeutic target in lung adenocarcinomas.

Foxd3/SLC5A6 axis regulates apoptosis in LUAD cells by controlling mitochondrial biotin uptake.

Lung cancer remains one of the leading causes of cancer-related mortality worldwide, with non-small cell lung cancer (NSCLC) accounting for over 85 % of cases. Lung adenocarcinoma (LUAD) is the most common subtype of NSCLC, and while targeted therapies and immune checkpoint inhibitors have improved outcomes, many patients exhibit resistance, necessitating the development of novel treatments. This study explores the role of the SLC5A6 gene, which encodes a sodium-dependent multivitamin transporter critical for mitochondrial function, in LUAD progression. We found that SLC5A6 is significantly upregulated in LUAD tissues and is associated with poor prognosis. Overexpression of SLC5A6 enhanced cell proliferation and migration, while knockout of SLC5A6 impaired these processes and induced apoptosis by disrupting mitochondrial function. Additionally, we identified Foxd3 as a key transcription factor regulating SLC5A6 expression. In vivo experiments demonstrated that SLC5A6 knockout effectively inhibited tumor growth. These findings suggest that SLC5A6 is a potential therapeutic target for LUAD, offering a new avenue for treatment strategies.

Paeoniae radix overcomes resistance to EGFR-TKIs via aurora B pathway suppression in lung adenocarcinoma

Targeted therapies using epidermal growth factor receptor (EGFR) inhibitors have markedly improved survival rates and quality of life for patients with EGFR-mutant lung adenocarcinoma (LUAD). Despite these advancements, resistance to EGFR inhibitors remains a significant challenge, limiting the overall effectiveness of the treatment. This study explored the synergistic effects of combining Paeoniae Radix (PR) with first-generation EGFR-tyrosine kinase inhibitors (TKIs), erlotinib and gefitinib, to overcome this resistance. Transcriptomic analysis of EGFR-mutant LUAD cell lines revealed that PR treatment could potentially reverse the gene signatures associated with resistance to EGFR-TKIs, primarily through the suppression of the Aurora B pathway. Experimental validation demonstrated that combining PR with erlotinib and gefitinib enhanced drug responsiveness by inhibiting Aurora kinase activity and inducing apoptosis in LUAD cells. Additionally, gene expression changes confirmed these combined effects, with the suppression of the Aurora B pathway and upregulation of the apoptotic pathway, which was accompanied by increased expression of multiple pro-apoptotic genes. Our findings contribute to the development of natural product-based therapeutic strategies to mitigate drug resistance in LUAD.

Identification of potential biomarkers for lung adenocarcinoma: a study based on bioinformatics analysis combined with validation experiments.

The prognosis for lung adenocarcinoma (LUAD) remains dismal, with a 5-year survival rate of <20%. Therefore, the purpose of this study was to identify potentially reliable biomarkers in LUAD by machine learning combination with Mendelian randomization (MR). TCGA-LUAD, GSE40791, and GSE31210 were employed this study. Key module differential genes were identified through differentially expressed analysis and weighted gene co-expression network analysis (WGCNA). Furthermore, candidate biomarkers were derived from protein-protein interaction network (PPI) and machine learning. Ultimately, biomarkers were confirmed using MR analysis. In addition, immunohistochemistry was used to detect the expression levels of genes that have a causal relationship to LUAD in the LUAD group and the control group. Cell experiments were conducted to validate the effect of screening genes on proliferation, migration, and apoptosis of LUAD cells. The correlation between the screened genes and immune infiltration was determined by CIBERSORT algorithm. In the end, the gene-related drugs were predicted through the Drug-Gene Interaction database. In total, 401 key module differential genes were obtained by intersecting of 5,702 differentially expressed genes (DEGs) and 406 key module genes. Thereafter, GIMAP6, CAV1, PECAM1, and TGFBR2 were identified. Among them, only TGFBR2 had a significant causal relationship with LUAD (p=0.04, b=-0.06), and it is a protective factor for LUAD. Subsequently, sensitivity analyses showed that there were no heterogeneity and horizontal pleiotropy in the univariate MR results, and the results were not overly sensitive to individual SNP loci, further validating the reliability of univariate Mendelian randomization (UVMR) results. However, no causal relationship was found between them by reverse MR analysis. Meanwhile, TGFBR2 expression was decreased in LUAD group through immunohistochemistry. TGFBR2 can inhibit proliferation and migration of lung adenocarcinoma cell line A549 and promote apoptosis of A549 cells. Immune infiltration analysis suggested a potential link between TGFBR2 expression and immune infiltration. Finally, Irinotecan and Hesperetin were predicted through DGIDB database. In this study, TGFBR2 was identified as a biomarker of LUAD, which provided a new idea for the treatment strategy of LUAD and may aid in the development of personalized immunotherapy strategies.

Fractionated Leaf Extracts of Ocimum gratissimum Inhibit the Proliferation and Induce Apoptosis of A549 Lung Adenocarcinoma Cells.

Previous in vitro studies in our laboratory demonstrated that ethyl acetate (P2) and water- soluble (PS/PT1) fractionated leaf extracts of Ocimum gratissimum inhibit the proliferation of prostate cancer cells. It has been reported that the crude aqueous extract induces apoptosis in lung adenocarcinoma cells; however, the efficacy of the fractionated extracts against these cells remains unclear. In the present study, we hypothesized that the ability of the fractionated extracts to inhibit proliferation and induce apoptosis is associated with the activation of pro-apoptotic proteins and induction of DNA condensation in A549 cells. Ocimum gratissimum was cultivated and its leaves were harvested, extracted, and fractionated to produce fractions P2 and PS/PT1. Anti-proliferative activity was assessed by direct cell count. For morphological characterization of apoptosis, 4',6-diamidino-2-phenylindole staining was employed. Western blot analysis was performed to evaluate the apoptotic activity of the fractionated extracts. In data generated from anti-proliferation studies, P2 significantly inhibited cell proliferation in a concentration-dependent manner; PS/PT1 elicited a decrease in the viability of cells, occurring at 500 µg/mL. 4',6-diamidino-2-phenylindole staining revealed the induction of apoptosis, as evidenced by the formation of apoptotic bodies. Increased levels of pro-apoptotic proteins were observed as the concentrations of the fractionated extracts increased. These results suggest that fractionated leaf extracts of Ocimum gratissimum inhibit the proliferation and induce apoptosis of A549 cells.

The Nucleolar Protein C1orf131 Is a Novel Gene Involved in the Progression of Lung Adenocarcinoma Cells through the AKT Signalling Pathway.

Lung adenocarcinoma (LUAD) is the most widespread cancer in the world, and its development is associated with complex biological mechanisms that are poorly understood. Here, we revealed a marked upregulation in the mRNA level of C1orf131 in LUAD samples compared to non-tumor tissue samples in The Cancer Genome Atlas (TCGA). Depletion of C1orf131 suppressed cell proliferation and growth, whereas it stimulated apoptosis in LUAD cells. Mechanistic investigations revealed that C1orf131 knockdown induced cell cycle dysregulation via the AKT and p53/p21 signalling pathways. Additionally, C1orf131 knockdown blocked cell migration through the modulation of epithelial-mesenchymal transition (EMT) in lung adenocarcinoma. Notably, we identified the C1orf131 protein nucleolar localization sequence, which included amino acid residues 137-142 (KKRKLT) and 240-245 (KKKRKG). Collectively, C1orf131 has potential as a novel therapeutic marker for patients in the future, as it plays a vital role in the progression of lung adenocarcinoma.

GLUT1 promotes cell proliferation via binds and stabilizes phosphorylated EGFR in lung adenocarcinoma

BackgroundWhile previous studies have primarily focused on Glucose transporter type 1 (GLUT1) related glucose metabolism signaling, we aim to discover if GLUT1 promotes tumor progression through a non-metabolic pathway.MethodsThe RNA-seq and microarray data were comprehensively analyzed to evaluate the significance of GLUT1 expression in lung adenocarcinoma (LUAD). The cell proliferation, colony formation, invasion, and migration were used to test GLUT1 ‘s oncogenic function. Co-immunoprecipitation and mass spectrum (MS) were used to uncover potential GLUT1 interacting proteins. RNA-seq, DIA-MS, western blot, and qRT-PCR to probe the change of gene and cell signaling pathways.ResultsWe found that GLUT1 is highly expressed in LUAD, and higher expression is related to poor patient survival. GLUT1 knockdown caused a decrease in cell proliferation, colony formation, migration, invasion, and induced apoptosis in LUAD cells. Mechanistically, GLUT1 directly interacted with phosphor-epidermal growth factor receptor (p-EGFR) and prevented EGFR protein degradation via ubiquitin-mediated proteolysis. The GLUT1 inhibitor WZB117 can increase the sensitivity of LUAD cells to EGFR-tyrosine kinase inhibitors (TKIs) Gefitinib.ConclusionsGLUT1 expression is higher in LUAD and plays an oncogenic role in lung cancer progression. Combining GLUT1 inhibitors and EGFR-TKIs could be a potential therapeutic option for LUAD treatment.

Arsenic-induced downregulation of BRWD3 suppresses proliferation and induces apoptosis in lung adenocarcinoma cells through the p53 and p65 pathways.

Bromodomain and WD-repeat domain-containing protein 3 (BRWD3) exhibits high expression in lung adenocarcinoma (LUAD) tissues and cells; however, its function in arsenic-induced toxicological responses remains unclear. This study aimed to investigate BRWD3 expression in response to arsenic-induced conditions and its impact on the proliferation and apoptosis of LUAD cell line SPC-A1 upon BRWD3 knockdown. The results revealed a decrease in BRWD3 expression in SPC-A1 cells treated with sodium arsenite (NaAsO2), but not sodium arsenite's metabolites. BRWD3 knockdown suppressed cell proliferation and induced apoptosis in SPC-A1 cells. Western blot analysis revealed that BRWD3 knockdown resulted in the upregulation of p53, phospho-p53-Ser392, and its downstream factors including MDM2, Bak, and Bax. Additionally, we observed the downregulation of p65, phospho-p65-Ser276, phospho-p65-Ser536, and its downstream factors, including IκBα, BIRC3, XIAP and CIAP1. Moreover, polymerase chain reaction analysis showed that BRWD3 knockdown also resulted in the downregulation of proliferation-related genes and upregulation of apoptosis-related genes. In conclusion, BRWD3 mediated proliferation and apoptosis via the p53 and p65 pathways in response to arsenic exposure, suggesting potential implications for LUAD treatment through BRWD3 downregulation by arsenic.

DHEA down-regulates mitochondrial dynamics and promotes apoptosis of lung adenocarcinoma cells through FASTKD2.

Background: DHEA is a steroid hormone produced by the gonads, adrenal cortex, brain, and gastrointestinal tract. While the anti-obesity, anti-atherosclerosis, anti-cancer, and memory-enhancing effects of DHEA have been substantiated through cell experiments, animal studies, and human trials, the precise mechanisms underlying these effects remain unclear. Altered mitochondrial dynamics can lead to mitochondrial dysfunction, which is closely related to many human diseases, especially cancer and aging. This study was to investigate whether DHEA inhibits lung adenocarcinoma through the mitochondrial pathway and its molecular mechanism. Methods: Through animal experiments and cell experiments, the effect of DHEA on tumor inhibition was determined. The correlation between FASTKD2 expression and DHEA was analyzed by Western blot, Reverse transcription-quantitative PCR, Immunohistochemistry, and TCGA database. Results: In this study, DHEA supplementation in the diet can inhibit the tumor size of mice, and the effect of adding DHEA one week before the experiment is the best. DHEA limits the glycolysis process by inhibiting G6PDH activity, increases the accumulation of reactive oxygen species, and initiates apoptosis in the mitochondrial pathway of cancer cells. Conclusion: DHEA suppresses mitochondrial fission and promotes mitochondrial fusion by downregulating the expression of FASTKD2, thereby inhibiting tumor growth and prolonging the overall survival of lung adenocarcinoma patients, which also provides a new target for the prevention and treatment of lung adenocarcinoma.

Integrating multi-omics and machine learning survival frameworks to build a prognostic model based on immune function and cell death patterns in a lung adenocarcinoma cohort.

The programmed cell death (PCD) plays a key role in the development and progression of lung adenocarcinoma. In addition, immune-related genes also play a crucial role in cancer progression and patient prognosis. However, further studies are needed to investigate the prognostic significance of the interaction between immune-related genes and cell death in LUAD. In this study, 10 clustering algorithms were applied to perform molecular typing based on cell death-related genes, immune-related genes, methylation data and somatic mutation data. And a powerful computational framework was used to investigate the relationship between immune genes and cell death patterns in LUAD patients. A total of 10 commonly used machine learning algorithms were collected and subsequently combined into 101 unique combinations, and we constructed an immune-associated programmed cell death model (PIGRS) using the machine learning model that exhibited the best performance. Finally, based on a series of in vitro experiments used to explore the role of PSME3 in LUAD. We used 10 clustering algorithms and multi-omics data to categorize TCGA-LUAD patients into three subtypes. patients with the CS3 subtype had the best prognosis, whereas patients with the CS1 and CS2 subtypes had a poorer prognosis. PIGRS, a combination of 15 high-impact genes, showed strong prognostic performance for LUAD patients. PIGRS has a very strong prognostic efficacy compared to our collection. In conclusion, we found that PSME3 has been little studied in lung adenocarcinoma and may be a novel prognostic factor in lung adenocarcinoma. Three LUAD subtypes with different molecular features and clinical significance were successfully identified by bioinformatic analysis, and PIGRS was constructed using a powerful machine learning framework. and investigated PSME3, which may affect apoptosis in lung adenocarcinoma cells through the PI3K/AKT/Bcl-2 signaling pathway.

Expression and role of FKBPL in lung adenocarcinoma.

Dysregulated expression of FK506-binding protein like (FKBPL) has been demonstrated to play crucial roles in tumour development. However, the role of FKBPL in lung adenocarcinoma (ADC) remains unclear. Using immunohistochemical staining, we showed that FKBPL expression was significantly lower in lung ADC than the normal tissues (P < 0.0001). Patients with well or moderately differentiated tumours have higher FKBPL expression compared with patients with poor differentiated tumours (P = 0.037). However, no significant associations were found between FKBPL expression and other clinicopathological variables (P > 0.05 for all). Cox univariate analysis showed that high FKBPL expression was correlated with prolonged overall survival (OS) (P = 0.010). Kaplan-Meier analysis further confirmed that the FKBPL-low group showed a significantly shorter OS than the FKBPL-high group (P = 0.0081). FKBPL expression was not shown as an independent prognostic factor for OS in the multivariate analysis (P = 0.063). Moreover, our study demonstrated that FKBPL could suppress the proliferation of lung ADC cells by delaying cell cycle G1/S phase transition. In addition, FKBPL resulted in increased apoptosis in lung ADC cells. Using the Human Apoptosis Array Kit, we observed that overexpression of FKBPL in lung ADC A549 cells significantly decreased the anti-apoptotic proteins, including heat shock protein 32 (HSP32), heat shock protein 27 (HSP27), and paraoxonase-2 (PON2). FKBPL depletion significantly attenuated the pro-apoptotic protein, phospho-p53 (S46), in lung ADC H1975 cells. These new findings provide an experimental basis for further theoretical investigation of lung ADC.

GIMAP7 inhibits epithelial-mesenchymal transition and glycolysis in lung adenocarcinoma cells via regulating the Smo/AMPK signaling pathway.

GTPase immunity-associated protein 7 (GIMAP7) has been previously recognized as a prognostic marker in pan-cancer. Our objective was to explore the function of GIMAP7 in the progression of lung adenocarcinoma (LUAD). GIMAP7 was overexpressed by transfection with GIMAP7 plasmid, and knocked down using siRNAs. The biological functions of GIMAP7 were examined by employing CCK-8, EdU, colony formation, flow cytometry, wound healing, and transwell assays. The effects of GIMAP7 on the extracellular acidification rate (ECAR), oxygen consumption rate (OCR), lactate production, and glucose uptake were evaluated. In addition, the related mRNA and protein expression was detected employing immunohistochemical, western blot, and qRT-PCR. A xenograft tumor model was established in nude mice to evaluate the effects of GIMAP7 on tumor growth. GIMAP7 was lowly expressed in LUAD tissues and cells. GIMAP7 inhibited the proliferation, mobility, EMT, glycolysis, but promoted apoptosis in LUAD cells. Moreover, we also confirmed that GIMAP7 suppressed Smo/AMPK signaling in LUAD cells. By adding the Smo agonist SAG and AMPK agonist GSK621, the results of rescue experiments further verified that GIMAP7 played a role in LUAD inhibition through inhibition of the Smo/AMPK signaling pathway. Furthermore, the role of GIMAP7 in inhibiting tumor growth was verified in vivo. These results demonstrate that GIMAP7 could inhibit cell proliferation, mobility and glycolysis, but accelerate apoptosis via repressing the Smo/AMPK signaling pathway in LUAD.

CircSFMBT2 Plays an Oncogenic Role in Lung Adenocarcinoma Depending on the miR-1305/SALL4 Axis.

Circular RNAs (circRNAs) exhibit significant functions in diverse malignant tumors, including lung adenocarcinoma (LUAD). In this study, we aimed to elucidate the role of circRNA scm like with four mbt domains 2 (circSFMBT2) in LUAD. Quantitative real-time polymerase chain reaction (qRT-PCR), western blot assay or immunohistochemistry (IHC) assay was performed for quantification of circSFMBT2, microRNA-1305 (miR-1305), spalt like transcription factor 4 (SALL4), proliferating Cell Nuclear Antigen (PCNA) or Ki-67. 5-ethynyl-2'-deoxyuridine (EdU) assay, transwell assay and flow cytometry analysis were applied to analyze cell proliferation, metastasis and apoptosis, respectively. Mouse xenograft model was established to explore the function of circSFMBT2. Dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay were used to estimate the relationship between miR-1305 and circSFMBT2 or SALL4. CircSFMBT2 was upregulated in LUAD and related to advanced TNM stage and poor prognosis. CircSFMBT2 knockdown suppressed cell proliferation, metastasis, glycolysis and induced apoptosis in LUAD cells in vitro as well as tumor formation in vivo. CircSFMBT2 directly targeted miR-1305, and miR-1305 inhibition reversed circSFMBT2 knockdown-mediated inhibitory effects on LUAD malignant behaviors. SALL4 was the target gene of miR-1305. MiR-1305 overexpression repressed the malignant phenotypes of LUAD cells, while SALL4 enhancement abated the effects. CircSFMBT2 aggravated the progression of LUAD by the miR-1305/SALL4 axis, which might provide a diagnostic and prognostic marker for LUAD.

Circ_0004140 promotes lung adenocarcinoma progression by upregulating NOVA2 via sponging miR-330-5p.

Circular RNAs (circRNAs) play a significant role in the tumorigenesis and progression of diverse human cancers, including lung adenocarcinoma. A previous study suggested that circ_0004140 expression was increased in lung adenocarcinoma cells. However, the molecular mechanism of circRNA circ_0004140 involved in lung adenocarcinoma is poorly defined. Circ_0004140, microRNA-330-5p (miR-330-5p), and NOVA alternative splicing regulator 2 (NOVA2) expression were determined by real-time quantitative polymerase chain reaction (RT-qPCR). Cell proliferation, apoptosis, migration, invasion, and angiogenesis ability were assessed using 5-ethynyl-2'-deoxyuridine (EdU), flow cytometry, wound healing, transwell, and capillary-like network formation assays. Proliferating cell nuclear antigen (PCNA), cyclin D1, and NOVA2 protein levels were detected using Western blot assay. The interaction between miR-330-5p and circ_0004140 or NOVA2 was verified by dual-luciferase reporter assay. Xenograft tumor model was utilized to assess the role of circ_0004140 in tumor growth in vivo. Circ_0004140 was upregulated in lung adenocarcinoma tissues and cell lines. Circ_0004140 silencing suppressed cell proliferation, migration, invasion and tube formation ability, and triggered the apoptosis of lung adenocarcinoma cells. Circ_0004140 acted as a molecular sponge for miR-330-5p, and miR-330-5p silencing largely reversed circ_0004140 knockdown-induced effects in lung adenocarcinoma cells. NOVA2 was a target of miR-330-5p, and NOVA2 overexpression might largely overturn miR-330-5p overexpression-induced influences in lung adenocarcinoma cells. Circ_0004140 upregulated NOVA2 expression via sponging miR-330-5p in lung adenocarcinoma cells. Circ_0004140 silencing restrained xenograft tumor growth in vivo. Circ_0004140 knockdown might suppress the malignant biological behaviors of lung adenocarcinoma cells via miR-330-5p-dependent regulation of NOVA2.

A risk score model based on lipid metabolism-related genes could predict response to immunotherapy and prognosis of lung adenocarcinoma: a multi-dataset study and cytological validation

BackgroundLipid metabolism is a key factor in tumorigenesis and drug resistance, and models related to lipid metabolism have shown potential to predict survival and curative effects of adjuvant therapy in various cancers. However, the relationship between lipid metabolism and prognosis and treatment response of lung adenocarcinoma (LUAD) are still unclear.MethodsWe enrolled seven bulk RNA-sequence datasets (GSE37745, GSE19188, GSE30219, GSE31547, GSE41271, GSE42127, and GSE72094) from the GEO database and one single-cell RNA-sequencing dataset (GSE117570) from the TISCH2 database. Non-negative matrix factorization (NMF) was utilized to construct the risk score model based on lipid score calculated by GSVA algorithm. Phs000452.v3, PMID: 26359337, PMID: 32472114, PRJEB23709 datasets were used to test the response to immunotherapy. Drug sensitivity analysis was assessed according to the GDSC database, and immunotherapy response was evaluated using the Wilcoxon test. Cellular function assays including clone formation, EDU assays and flow cytometry were implemented to explore the phenotype alteration caused by the knockdown of PTDSS1, which is one of key gene in risk score model.ResultsWe analyzed both bulk and single-cell RNA sequencing data to establish and validate a risk score model based on 18 lipid metabolism-related genes with significant impact on prognosis. After divided the patients into two groups according to risk score, we identified differences in lipid-related metabolic processes and a detailed portrait of the immune landscapes of high- and low-risk groups. Moreover, we investigated the potentials of our risk score in predicting response to immunotherapy and drug sensitivity. In addition, we silenced PTDSS1 in LUAD cell lines, and found that the proliferation of the cells was weakened, and the apoptosis of the cells was increased.ConclusionOur study highlights the crucial roles of lipid metabolism in LUAD and provides a reliable risk score model, which can aid in predicting prognosis and response to immunotherapy. Furthermore, we investigated the roles of PTDSS1 in LUAD carcinogenesis, which showed that PTDSS1 regulated proliferation and apoptosis of LUAD cells.

MCM4 acts as a biomarker for LUAD prognosis.

MCM4 forms the pre-replication complex (MCM2-7) with five other minichromosome maintenance (MCM) proteins. This complex binds to replication origins at G1 stage in cell cycle process, playing a critical role in DNA replication initiation. Recently, MCM4 is reported to have a complex interaction with multiple cancer progression, including gastric, ovarian and cervical cancer. Here, this study mainly focused on the expression of MCM4 and its values in lung adenocarcinoma (LUAD). MCM4 was highly expressed in LUAD tumours and cells, and had an important effect on the overall survival. Overexpression of MCM4 promoted the proliferation, and suppressed the apoptosis in LUAD cells. However, MCM4 silence led to the opposite results. In vivo, knockdown of MCM4 inhibited tumour volume and weight in xenograft mouse model. As a member of DNA helicase, knockdown of MCM4 caused cell cycle arrest at G1 stage through inducing the expression of P21, a CDK inhibitor. These findings indicate that MCM4 may be a possible new therapeutic target for LUAD in the future.

Hydromethanolic leaves extract of Dalbergia sissoo Roxb. ex DC. induces apoptosis in lung adenocarcinoma cells

Plants have a wide range of active secondary metabolites that are frequently utilized to treat cancer. For the research, Dalbergia sissoo Roxb. ex DC. leaves (DS) were hydromethanolically extracted, and their phytochemical content was determined. Total phenolics and flavonoids were quantified along with the measurement of in vitro antioxidant and cytotoxic activities. The bioactive components in the extract were identified with the use of Gas Chromatography with High-Resolution Mass Spectrometry (GC-HRMS). The crude extract contained 177.500 ± 0.019 mg/ml of flavonoids and 296.122 ± 0.002 mg/ml of Gallic Acid Equivalent (GAE) phenolics. Moreover, plant crude extract showed significant 2,2-diphenyl-1-picrylhydrazyl (DPPH) activity (IC50, 14.06 ± 0.18 μg/ml), 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) [ABTS] activity (IC50, 25.97 ± 1.04 μg/ml), superoxide scavenging activity (IC50, 149.91 ± 0.39 μg/ml), and hydrogen peroxide scavenging activity (IC50, 133.37 ± 2.30 μg/ml). The thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) methods were used to confirm the presence of phenolic compounds. DS effectively scavenges nitric oxide. The crude extract of DS exhibited good cytotoxic effects on lung cancer cells (A549) with an IC50 value of 90.56 ± 2.32 (μg/ml), and less toxicity on normal lung cells (WI-38) with an IC50 value of 381.10 ± 1.58 (μg/ml). IC50 value of methotrexate (the standard drug) was 10.20 ± 1.82 μg/ml on A549 cells and 26.21 ± 1.14 μg/ml on WI-38 cells. Various staining techniques [4, 6-diamidino-2-phenylindole (DAPI), Acridine orange/Ethidium Bromide (AO/EB) staining, Giemsa staining] were used to determine the plausible mechanism of DS to induce apoptosis. The inhibiting mechanism of the crude extract was further demonstrated by the clonogenic assay and qualitative and quantitative measurements of Reactive Oxygen Species (ROS). Real-time polymerase chain reaction (RT-PCR) analysis revealed the induction of apoptosis in A549 cells through the activation of caspases 9 and 3 together with TRAIL receptors. In a nutshell, hydromethanolic extract of DS resulted in distinct apoptotic morphological alterations, ROS production, the initiation of apoptosis via activating the TRAIL receptors, caspase 9 and 3, and the suppression of colony formation in A549 cells.

Combined inhibition of pyruvate dehydrogenase kinase 1 and lactate dehydrogenase a induces metabolic and signaling reprogramming and enhances lung adenocarcinoma cell killing

Lung adenocarcinoma (LUAD) is one of the most prevalent and aggressive types of lung cancer. Metabolic reprogramming plays a critical role in the development and progression of LUAD. Pyruvate dehydrogenase kinase 1 (PDK1) and lactate dehydrogenase A (LDHA) are two key enzymes involved in glucose metabolism, whilst their aberrant expressions are often associated with tumorigenesis. Herein, we investigated the anticancer effects of combined inhibition of PDK1 and LDHA in LUAD in vitro and in vivo and its underlying mechanisms of action. The combination of a PDK1 inhibitor, 64, and a LDHA inhibitor, NHI-Glc-2, led to a synergistic growth inhibition in 3 different LUAD cell lines and more than additively suppressed tumor growth in the LUAD xenograft H1975 model. This combination also inhibited cellular migration and colony formation, while it induced a metabolic shift from glycolysis to oxidative phosphorylation (OXPHOS) resulting in mitochondrial depolarization and apoptosis in LUAD cells. These effects were related to modulation of multiple cell signaling pathways, including AMPK, RAS/ERK, and AKT/mTOR. Our findings demonstrate that simultaneous inhibition of multiple glycolytic enzymes (PDK1 and LDHA) is a promising novel therapeutic approach for LUAD.

Ivermectin induces nonprotective autophagy by downregulating PAK1 and apoptosis in lung adenocarcinoma cells.

LUAD (Lung adenocarcinoma), the most common subtype of lung carcinoma and one of the highest incidences and mortality cancers in the world remains still a substantial treatment challenge. Ivermectin, an avermectin derivative, has been traditionally used as an antiparasitic agent in human and veterinary medicine practice during the last few decades. Though ivermectin has been shown to be effective against a variety of cancers, however, there is few available data reporting the antitumor effects of ivermectin in LUAD. The effect of ivermectin on cell viability and proliferative ability of LUAD cells was evaluated using CCK-8 and colony formation assay. Apoptosis rate and autophagy flux were detected using flow cytometry based on PI/Annexin V staining and confocal laser scanning microscope based on LC3-GFP/RFP puncta, respectively. Western blotting experiment was conducted to verify the results of changes in apoptosis and autophagy. LUAD-TCGA and GEO databases were used to analyse the expression and predictive value of PAK1 in LUAD patients. Xenograft model and immumohistochemical staining were used for verification of the inhibitor effect of ivermectin in vivo. Ivermectin treatment strikingly impeded the colony formation, and the viability of the cell, along with cell proliferation, and caused the apoptosis and enhanced autophagy flux in LUAD cells. In addition, ivermectin-induced nonprotective autophagy was confirmed by treating LUAD cells with 3-MA, an autophagy inhibitor. Mechanistically, we found that ivermectin inhibited PAK1 protein expression in LUAD cells and we confirmed that overexpression of PAK1 substantially inhibited ivermectin-induced autophagy in LUAD cells. Based on TCGA and GEO databases, PAK1 was highly expressed in LUAD tissues as compared with normal tissues. Furthermore, LUAD patients with high PAK1 level have poor overall survival. Finally, in vivo experiments revealed that ivermectin efficiently suppressed the cellular growth of LUAD among nude mice. This study not only revealed the mechanism of ivermectin inhibited the growth of LUAD but also supported an important theoretical basis for the development of ivermectin during the therapy for LUAD.

Dual inhibition of KIF11 and BCL2L1 induces apoptosis in lung adenocarcinoma cells

EGFR-mutant lung adenocarcinoma (LUAD) mostly depends on EGFR for survival and consequently responds well to EGFR inhibitors. However, resistance to the drugs develops almost universally during treatment. We previously demonstrated that EGFR-mutant LUAD cell lines, HCC827 and H1975, have subpopulations of cells, which we termed HCC827 GR2 and H1975 WR7 cells, that can thrive independently of EGFR signaling. These EGFR-independent EGFR-mutant cancer cells are difficult to treat because they lack sensitivity to EGFR inhibitors. Therefore, the development of novel strategies to target EGFR–independent EGFR-mutant LUAD is particularly important. We found that high expression of kinesin family member 11 (KIF11) correlated with poor survival in patients with LUAD. We also observed that KIF11 silencing caused cell cycle arrest at G2/M in HCC827 GR2 and H1975 WR7 cells. Furthermore, dual silencing of KIF11 plus BCL2L1, an anti-apoptotic BCL2 family member, in these two EGFR-independent sublines resulted in marked apoptosis levels. Dual inhibition of KIF11 plus BCL2L1 also induced apoptosis in HCC827 and H1975 parental cells and a KRAS-mutant LUAD cell line, H441. These findings collectively suggest that dual inhibition of KIF11 plus BCL2L1 may be a new approach for the treatment of LUAD.