H1299 Cells Research Articles

Bone marrow mesenchymal stem cells-derived exosomal miR-145-5p reduced non-small cell lung cancer cell progression by targeting SOX9

BackgroundThe role of miR-145-5p in non-small cell lung cancer (NSCLC) has been studied, however, the regulation of hBMSCs-derived exosomes (Exo) transmitted miR-145-5p in NSCLC was still unknown. This study aimed to investigate the role of hBMSCs-derived exosomes (Exo) in the progression of NSCLC.MethodsThe Exo was extracted from hBMSCs and added to A549 and H1299 cell culture, followed by the detection of cell proliferation, migration, and invasion. The correlation between the expression of miR-145-5p and SOX9, as well as their binding relationship was determined by correlation analysis, luciferase gene reporter assay and RNA pull-down assays. The in vivo animal model was established to further verify the impact of hBMSCs-Exo.ResultsIt showed that miR-145-5p was downregulated and SOX9 was upregulated in NSCLC tissues. HBMSCs-derived Exo, and hBMSCs-Exo with overexpression of miR-145-5p could inhibit cell proliferation, migration, and invasion of both A549 and H1299 cells, and prevent against tumor progression in vivo. MiR-145-5p and SOX9 were found to be able to bind to each other, and a negative correlation were observed between the expression of them in NSCLC tissues. Furthermore, inhibition of SOX9 could reversed the suppressed role of miR-145-5p in vitro and in vivo.ConclusionTherefore, HBMSCs-Exo effectively transmitted miR-145-5p, leading to the suppression of malignant development in NSCLC through the regulation of SOX9.

3,4,5-tri-O-caffeoylquinic acid attenuates influenza A virus induced inflammation through Toll-like receptor 3/7 activated signaling pathway

Background3,4,5-tri-O-caffeoylquinic acid (3,4,5-TCQA), a natural polyphenolic acid, has been shown to be effective against influenza A virus (IAV) infection. Although it was found to inhibit the neuraminidase of IAV, it may also perturb other cellular functions, as polyphenolic acids have shown antioxidant, anti-inflammatory and other activities. PurposeThis study aimed to investigate the effect of 3,4,5-TCQA at a cell level, which is critical for protecting host cell from IAV infection. Study design and methodsWe explored the effect of 3,4,5-TCQA on H292 cells infected or un-infected with Pr8 IAV. The major genes and related pathway were identified through RNA sequencing. The pathway was confirmed by qRT-PCR and western blot analysis. The anti-inflammatory activity was evaluated using nitric oxide measurement assay. ResultsWe showed that 3,4,5-TCQA downregulated the immune response in H292 cells, and reduced the cytokine production in Pr8-infected cells, through Toll-like receptor (TLR) signaling pathway. In addition, 3,4,5-TCQA showed anti-inflammatory activity in LPS-activated RAW264.7 cells. ConclusionCollectively, our results indicated that 3,4,5-TCQA suppressed inflammation caused by IAV infection through TLR3/7 signaling pathway. This provides a new insight into the antiviral mechanism of 3,4,5-TCQA.

Combination of Vismodegib and Paclitaxel Enhances Cytotoxicity via Bak-mediated Mitochondrial Damage in EGFR-Mutant Non-Small Cell Lung Cancer Cells.

Half of NSCLC patients harbor epidermal growth factor receptor (EGFR) mutations, and their therapeutic responses are remarkably different from patients with wild-type EGFR (EGFR-WT) NSCLC. We previously demonstrated that the hedgehog inhibitor vismodegib (Vis) potentiates paclitaxel (PTX)-induced cytotoxicity via suppression of Bax phosphorylation, which promotes accumulation of mitochondrial damage and apoptosis in EGFR-WT NSCLC cells. In this study, we further delineated the anticancer activity and underlying mechanisms of this combination treatment in EGFR-mutant NSCLC cells. MTS/PMS activity and trypan blue exclusion assays were used to assess cell viability. Apoptosis was monitored by chromosome condensation, annexin V staining, and cleavage of PARP and caspase-3. Western blots were conducted to track proteins of interest after treatment. Reactive oxygen species (ROS) level was monitored by 2',7'-dichlorodihydrofluorescein diacetate. Mitochondrial status was analyzed by tetramethylrhodamine, ethyl ester. Hedgehog signaling was induced by PTX, which rendered H1975 and PC9 cells insensitive to PTX-induced mitochondrial apoptosis via suppression of Bak. However, Vis enhanced PTX-induced Bak activation, leading to mitochondrial damage, ROS accumulation, and subsequent apoptosis. Our findings suggest that the combination of Vis and PTX could be a potential therapeutic strategy to increase PTX sensitivity of EGFR-mutant NSCLC.

USP32 facilitates non-small cell lung cancer progression via deubiquitinating BAG3 and activating RAF-MEK-ERK signaling pathway

The regulatory significance of ubiquitin-specific peptidase 32 (USP32) in tumor is significant, nevertheless, the biological roles and regulatory mechanisms of USP32 in non-small cell lung cancer (NSCLC) remain unclear. According to our research, USP32 was strongly expressed in NSCLC cell lines and tissues and was linked to a bad prognosis for NSCLC patients. Interference with USP32 resulted in a significant inhibition of NSCLC cell proliferation, migration potential, and EMT development; on the other hand, USP32 overexpression had the opposite effect. To further elucidate the mechanism of action of USP32 in NSCLC, we screened H1299 cells for interacting proteins and found that USP32 interacts with BAG3 (Bcl2-associated athanogene 3) and deubiquitinates and stabilizes BAG3 in a deubiquitinating activity-dependent manner. Functionally, restoration of BAG3 expression abrogated the antitumor effects of USP32 silencing. Furthermore, USP32 increased the phosphorylation level of the RAF/MEK/ERK signaling pathway in NSCLC cells by stabilizing BAG3. In summary, these findings imply that USP32 is critical to the development of NSCLC and could offer a theoretical framework for the clinical diagnosis and management of NSCLC patients in the future.

Combining [177Lu]Lu-DOTA-TOC PRRT with PARP inhibitors to enhance treatment efficacy in small cell lung cancer.

Small cell lung cancer (SCLC) is a highly aggressive tumor with neuroendocrine origin. Although SCLC frequently express somatostatin receptor type 2 (SSTR2), a significant clinical benefit of SSTR2-targeted radionuclide therapies of SCLC was not observed so far. We hypothesize that combination treatment with a PARP inhibitor (PARPi) could lead to radiosensitization and increase the effectiveness of SSTR2-targeted therapy in SCLC. SSTR2-ligand uptake of the SCLC cell lines H69 and H446 was evaluated in vitro using flow cytometry, and in vivo using SPECT imaging and cut-and-count biodistribution. Single-agent (Olaparib, Rucaparib, [177Lu]Lu-DOTA-TOC) and combination treatment responses were determined in vitro via cell viability, clonogenic survival and γH2AX DNA damage assays. In vivo, we treated athymic nude mice bearing H69 or H446 xenografts with Olaparib, Rucaparib, or [177Lu]Lu-DOTA-TOC alone or with combination treatment regimens to assess the impact on tumor growth and survival of the treated mice. H446 and H69 cells exhibited low SSTR2 expression, i.e. 60 to 90% lower uptake of SSTR2-ligands compared to AR42J cells. In vitro, combination treatment of [177Lu]Lu-DOTA-TOC with PARPi resulted in 2.9- to 67-fold increased potency relative to [177Lu]Lu-DOTA-TOC alone. We observed decreased clonogenic survival and higher amounts of persistent DNA damage compared to single-agent treatment for both Olaparib and Rucaparib. In vivo, tumor doubling times increased to 1.6-fold (H446) and 2.2-fold (H69) under combination treatment, and 1.0 to 1.1-fold (H446) and 1.1 to 1.7-fold (H69) in monotherapies compared to untreated animals. Concurrently, median survival was higher in the combination treatment groups in both models compared to monotherapy and untreated mice. Fractionating the PRRT dose did not lead to further improvement of therapeutic outcome. The addition of PARPi can markedly improve the potency of SSTR2-targeted PRRT in SCLC models in SSTR2 low-expressing tumors. Further evaluation in humans seems justified based on the results as novel treatment options for SCLC are urgently needed.

PRRX1-OLR1 axis supports CAFs-mediated lung cancer progression and immune suppression

ObjectiveTo investigate the mechanism by which cancer-associated fibroblasts (CAFs) affect the growth and immune evasion of lung cancer cells.MethodsInitially, datasets comparing CAFs with normal fibroblasts were downloaded from the GEO dataset GSE48397. Genes with the most significant differential expression were selected and validated using clinical data. Subsequently, CAFs were isolated, and the selected genes were knocked down in CAFs. Co-culture experiments were conducted with H1299 or A549 cells to analyze changes in lung cancer cell growth, migration, and immune evasion in vitro and in vivo. To further elucidate the upstream regulatory mechanism, relevant ChIP-seq data were downloaded from the GEO database, and the regulatory relationships were validated through ChIP-qPCR and luciferase reporter assays.ResultsOLR1 was significantly overexpressed in CAFs and strongly correlated with adverse prognosis in lung cancer patients. Knockdown of OLR1 markedly inhibited CAFs’ support for the growth and immune evasion of lung cancer cells in vitro and in vivo. ChIP-seq results demonstrated that PRRX1 can promote OLR1 expression by recruiting H3K27ac and H3K4me3, thereby activating CAFs. Knockdown of PRRX1 significantly inhibited CAFs’ function, while further overexpression of OLR1 restored CAFs’ support for lung cancer cell growth, migration, and immune evasion.ConclusionPRRX1 promotes OLR1 expression by recruiting H3K27ac and H3K4me3, activating CAFs, and thereby promoting the growth, migration, and immune evasion of lung cancer cells.

Potential Application of the Myocardial Scintigraphy Agent [123I]BMIPP in Colon Cancer Cell Imaging.

[123I]β-methyl-p-iodophenyl-pentadecanoic acid ([123I]BMIPP), which is used for nuclear medicine imaging of myocardial fatty acid metabolism, accumulates in cancer cells. However, the mechanism of accumulation remains unknown. Therefore, this study aimed to elucidate the accumulation and accumulation mechanism of [123I]BMIPP in cancer cells. We compared the accumulation of [123I]BMIPP in cancer cells with that of [18F]FDG and found that [123I]BMIPP was a much higher accumulation than [18F]FDG. The accumulation of [123I]BMIPP was evaluated in the presence of sulfosuccinimidyl oleate (SSO), a CD36 inhibitor, and lipofermata, a fatty acid transport protein (FATP) inhibitor, under low-temperature conditions and in the presence of etomoxir, a carnitine palmitoyl transferase I (CPT1) inhibitor. The results showed that [123I]BMIPP accumulation was decreased in the presence of SSO and lipofermata in H441, LS180, and DLD-1 cells, suggesting that FATPs and CD36 are involved in [123I]BMIPP uptake in cancer cells. [123I]BMIPP accumulation in all cancer cell lines was significantly decreased at 4 °C compared to that at 37 °C and increased in the presence of etomoxir in all cancer cell lines, suggesting that the accumulation of [123I]BMIPP in cancer cells is metabolically dependent. In a biological distribution study conducted using tumor-bearing mice transplanted with LS180 cells, [123I]BMIPP highly accumulated in not only LS180 cells but also normal tissues and organs (including blood and muscle). The tumor-to-intestine or large intestine ratios of [123I]BMIPP were similar to those of [18F]FDG, and the tumor-to-large-intestine ratios exceeded 1.0 during 30 min after [123I]BMIPP administration in the in vivo study. [123I]BMIPP is taken up by cancer cells via CD36 and FATP and incorporated into mitochondria via CPT1. Therefore, [123I]BMIPP may be useful for imaging cancers with activated fatty acid metabolism, such as colon cancer. However, the development of novel imaging radiotracers based on the chemical structure analog of [123I]BMIPP is needed.

Impact of the redox-active MnTnHex-2-PyP5+ and cisplatin on the metabolome of non-small cell lung cancer cells

Redox-based cancer therapeutic strategies aim to raise reactive oxygen species (ROS) levels in cancer cells, thus modifying their redox status, and eventually inducing cell death. Promising compounds, known as superoxide dismutase mimics (SODm), e.g. MnTnHex-2-Py5+ (MnTnHex), could increase intracellular H2O2 in cancer cells with deficient ROS removal systems and therefore enhance radio- and chemotherapy efficacy. We have previously shown that MnTnHex was cytotoxic either alone or combined with cisplatin to non-small cell lung cancer (NSCLC) cells. To gain a deeper understanding of the effects and safety of this compound, it is crucial to analyze the metabolic alterations that take place within the cell. Our goal was thus to study the intracellular metabolome (intracellular metabolites) of NSCLC cells (A549 and H1975) using nuclear magnetic resonance (NMR) spectroscopy-based metabolomics to evaluate the changes in cellular metabolism upon exposure to MnTnHex per se or in combination with cisplatin. 1H NMR metabolomics revealed a higher number of significantly altered metabolites in A549 cells exposed to MnTnHex alone or combined with cisplatin in comparison with non-treated cells (nine dysregulated metabolites), suggesting an impact on aminoacyl-tRNA biosynthesis, glycolysis/gluconeogenesis, taurine, hypotaurine, glycerophospholipid, pyruvate, arginine and proline metabolisms. Regarding H1975 cells, significant alterations in the levels of six metabolites were observed upon co-treatment with MnTnHex and cisplatin, suggesting dysregulations in aminoacyl-tRNA biosynthesis, arginine and proline metabolism, pyruvate metabolism, and glycolysis/gluconeogenesis. These findings help us to understand the impact of MnTnHex on NSCLC cells. Importantly, specific altered metabolites, such as taurine, may contribute to the chemosensitizing effects of MnTnHex.

Acovenosigenin A β-glucoside mediates JAK2-STAT3 signaling pathway by targeting GP130 in A549 and H460 cells based on integrative analysis of transcriptome and proteome and biological verification

Acovenosigenin A β-glucoside (AAG) is a cardiac glycoside derived from Streptocaulon juventas (Lour.) Merr, which exhibited the potential in treating lung cancer in our previous research. However, the action mechanism remains unclear. In this research, JAK2-STAT3 signaling pathway was predicted to be the critical regulation pathway based on the integrative analysis of transcriptome and proteome. Western blotting and qPCR assays were performed to identify that AAG can regulate JAK2-STAT3 signaling pathway and its downstream genes, such as c-Myc, Survivin, Cyclin B1, CDK1, Bcl-2. And this action of AAG depended on the suppression of STAT3 phosphorylation and its nuclear translocation through the experiments of Immunofluorescence, transient transfection and cryptotanshinone treatment. Additionally, AAG was discovered to mediate the JAK2-STAT3 pathway in IL-6-driven A549 and H460 cells, which in turn inhibited cell proliferation, promoted mitochondria-related apoptosis, and arrested the cell cycle progression. By molecular docking analysis, CETSA and SIP experiments, the protein of GP130 was identified as the specific target of AAG in A549 and H460 cells. Further studies suggested that AAG inhibited JAK2-STAT3 pathway and its downstream genes by targeting GP130 in nude mice xenograft model in vivo. This research presented that AAG exhibits the promising potential in the treatment of NSCLC.

Intriguing steroid glycosides for cancer therapy by suppressing the DNA damage response and mTOR/S6K signaling pathways

Two rare 8-hydroxysteroid glycosides (6–7), and their downstream metabolites (1–5) with an unprecedented 6/6/5/5/5-pentacyclic scaffold, together with seven known analogues (8–14) were isolated from the twigs and leaves of Strophanthus divaricatus. Their structures were fully assigned by analysis of the spectroscopic and ECD data, NMR calculations, X-ray crystallographic study, and chemical methods. In addition, the inhibitory effects of 1–14 on liver and lung cancer cell lines were evaluated, and preliminary structure–activity relationship was discussed. Data-independent acquisition (DIA)-based quantitative proteomic analysis and biological verification of H1299 cells suggested that this family of compounds may play an anticancer role by suppressing both DNA damage response (DDR) and mTOR/S6K signaling pathways.

CALU promotes lung adenocarcinoma progression by enhancing cell proliferation, migration and invasion

BackgroundLung cancer is the second most common cancer with the highest mortality in the world. Calumenin as a molecular chaperone that not only binds various proteins within the endoplasmic reticulum but also plays crucial roles in diverse processes associated with tumor development. However, the regulatory mechanism of calumenin in lung adenocarcinoma remains elusive. Here, we studied the impact of calumenin on lung adenocarcinoma and explored possible mechanisms.Methods5-ethynyl-2’-deoxyuridine assay, colony formation, transwell and wound healing assays were performed to explore the effects of calumenin on the proliferation and migration of lung adenocarcinoma cells. To gain insights into the underlying mechanisms through which calumenin knockdown inhibits the migration and proliferation of lung adenocarcinoma, we performed Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, Gene Set Enrichment Analysis and Ingenuity Pathway Analysis based on transcriptomics by comparing calumenin knockdown with normal A549 cells.ResultsThe mRNA and protein levels of calumenin in lung adenocarcinoma are highly expressed and they are related to an unfavorable prognosis in this disease. Calumenin enhances the proliferation and migration of A549 and H1299 cells. Gene Set Enrichment Analysis revealed that knockdown of calumenin in A549 cells significantly inhibited MYC and V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog signaling pathways while activating interferon signals, inflammatory signals, and p53 pathways. Ingenuity pathway analysis provided additional insights, indicating that the interferon and inflammatory pathways were prominently activated upon calumenin knockdown in A549 cells.ConclusionsThe anti-cancer mechanism of calumenin knockdown might be related to the inhibition of MYC and KRAS signals but the activation of interferon signals, inflammatory signals and p53 pathways.

Novel coumarin-piperazine-2(5H)-furanone hybrids as potential anti-lung cancer agents: Synthesis, biological evaluation and molecular docking studies

Novel coumarin-piperazine-2(5H)-furanone hybrids 5a–l were efficiently synthesized by introducing a furanone scaffold into coumarin using piperazine as a linker. The cytotoxicity of all hybrids 5a–l were evaluated by MTT assay on human lung cancer A549 cells and normal human lung fibroblast WI-38 cells with cytarabine (CAR) as a positive control. Hybrid 5l (IC50 = 11.28 μM) was the most toxic to A549 cells, 18-fold more toxic than the reference CAR (IC50 = 202.57 μM). Moreover, hybrid 5l (IC50 = 411.93 μM) was less toxic to WI-38 cells, with a much higher selectivity (5l, SI ≈ 37, WI-38/A549) than CAR (SI ≈ 2). Structure–activity relationship analysis showed that both the cytotoxicity against A549 cells and selectivity (WI-38/A549) were greatly improved when the bornyl group was incorporated in the hybrids (5c, 5f, 5i and 5l). Further, hybrid 5l was more toxic and selective against four types of human lung cancer cells (A549, Calu-1, PC-9 and H460; IC50 = 5.72–45.46 μM; SI ≈ 9–72) than three other types of human cancer cells (SK-BR-3, 786-O and SK-OV-3, IC50 = 39.07–130.82 μM; SI ≈ 0–2), showing remarkable specificity. In particular, hybrid 5l (IC50 = 5.72 μM) showed the highest cytotoxicity against H460 cells with the highest selectivity of up to 72 (WI-38/H460). Flow cytometric analysis showed that hybrid 5l induced apoptosis in H460 cells in a concentration-dependent manner. Molecular docking studies revealed a high binding affinity of hybrid 5l with CDK2 protein. Hybrid 5l is expected to be a leading candidate for anti-lung cancer agents.

Inducing Effect of Corylus avellana on Cytotoxic Activity in Lung and Breast Cancer Cells via Apoptosis

Turkish hazelnut (Corylus avellana L. cv Tombul) is a widely used nut in the chocolate industry and is also rich in polyphenol content, which promises anticancer effects. The anti-cancer and apoptotic effects of hazelnut leaves extracts examined on lung and breast cancer cells. Sulforhodamine B (SRB) and Adenosine 5’- triphosphate (ATP) assays were carried out for cell viability measurement. The mode of cell death was shown morphologically by the double fluorescence staining. Apoptosis was determined by performing caspase-mediated cytokeratin 18 (M30 ELISA) and western blot analysis. PARP, caspase 3, caspase 8, DR4, and GAPHD (Glyceraldehyde-3-phosphate Dehydrogenase) protein bands were visualized as markers of apoptosis. A wound healing test was employed to measure cell migration. Methanol extract of hazelnut leaf exhibited inhibition of cell growth activities in a dose-dependent manner. IC50 values were determined as 32.17 µg/ml in MCF-7, 32.16 µg/ml in MDA-MB-231, 20.40 µg/ml in A549 and 12.04 µg/ml in H1299 cells for ethanol extract while it was determined as 21.08 µg/ml in MCF-7, 40.16 µg/ml in MDA-MB-231, 22.04 µg/ml in A549 and 5.91 µg/ml in H1299 cells in methanol extract. In comparison, methanol leaf extracts were more effective in H1299 cells (IC50 value was 5.91 µg/ml).In comparison, ethanol leaf extracts were more effective in H1299 cells (IC50 value was 9.722 µg/ml). Western blot analysis demonstrated that hazelnut leaf extract treatment of cancer cells led to cell death via apoptosis and inhibited cell migration in lung and breast cancer cell lines. The cytotoxic effects of hazelnut extract on breast and lung cancer cells might be valuable and promising in elucidating cell death mechanisms for the development of new methods in cancer treatment.

Tumor Cell-Derived Exosomal Hybrid Nanosystems Loaded with Rhubarbic Acid and Tanshinone IIA for Sepsis Treatment.

Sepsis continues to exert a significant impact on morbidity and mortality in clinical settings, with immunosuppression, multi-organ failure, and disruptions in gut microbiota being key features. Although rheinic acid and tanshinone IIA show promise in mitigating macrophage apoptosis in sepsis treatment, their precise targeting of macrophages remains limited. Additionally, the evaluation of intestinal flora changes following treatment, which plays a significant role in subsequent cytokine storms, has been overlooked. Leveraging the innate inflammation chemotaxis of tumor cell-derived exosomes allows for their rapid recognition and uptake by activated macrophages, facilitating phenotypic changes and harnessing anti-inflammatory effects. We extracted exosomes from H1299 cells using a precipitation method. Then we developed a tumor cell-derived exosomal hybrid nanosystem loaded with rhubarbic acid and tanshinone IIA (R+T/Lipo/EXO) for sepsis treatment. In vitro studies, we verify the anti-inflammatory effect and the mechanism of inhibiting cell apoptosis of nano drug delivery system. The anti-inflammatory effects, safety, and modulation of intestinal microbiota by the nanoformulations were further validated in the in vivo study. Nanoformulation demonstrated enhanced macrophage internalization, reduced TNF-α expression, inhibited apoptosis, modulated intestinal flora, and alleviated immunosuppression. R+T/Lipo/EXO presents a promising approach using exosomal hybrid nanosystems for treating sepsis.

Knockdown of GNL3 inhibits LUAD cell growth by regulating Wnt-β-catenin pathway.

Lung adenocarcinoma (LUAD) is a leading cause of tumor-associated mortality, and it is needed to find new target to combat this disease. Guanine nucleotide-binding -protein-like 3 (GNL3) mediates cell proliferation and apoptosis in several cancers, but its role in LUAD remains unclear. To explore the expression and function of Guanine nucleotide-binding protein-like 3 (GNL3) in lung adenocarcinoma (LUAD) and its potential mechanism in inhibiting the growth of LUAD cells. We evaluated the expression of GNL3 in LUAD tissues and its association with patient prognosis using databases and immunohistochemistry. Cell proliferation was assessed by CCK-8 assay as well as colony formation, while apoptosis was evaluated by FCM. The effect of GNL3 knockdown on the Wnt/β-catenin axis was investigated by Immunoblot analysis. GNL3 is overexpressed in LUAD tissues and is correlated with poor prognosis. Knockdown of GNL3 significantly inhibited the growth as well as induced apoptosis in A549 as well as H1299 cells. Furthermore, we found that the inhibitory effect of GNL3 knockdown on LUAD cell growth is associated with the downregulation of the Wnt/β-catenin axis. GNL3 is key in the progression of LUAD by metiating Wnt/β-catenin axis. Targeting GNL3 may represent a novel therapeutic method for LUAD treatment.

Anticancer activity of Araguspongine C via inducing apoptosis, and inhibition of oxidative stress, inflammation, and EGFR-TK in human lung cancer cells: An in vitro and in vivo study.

The advanced non-small cell lung cancer (NSCLC) that harbors epidermal growth factor receptor(EGFR) mutations has put a selective pressure on the discovery and development of newer EGFR inhibitors. Therefore, the presentstudy intends to explore the pharmacological effect of Araguspongine C (Aragus-C) as anticancer agent against lung cancer. The effect of Aragus-C was evaluated on the viability of the A549 and H1975 cells. Further biochemical assays were performed to elaborate the effect of Aragus-C, on the apoptosis, cell-cycle analysis, and mitochondrial membrane potential in A549 cells. Western blot analysis was also conducted to determine the expression of EGFR in A549 cells. Tumor xenograft mice model from A549 cells was established to further elaborate the pharmacological activity of Aragus-C. Results suggest that Aragus C showed significant inhibitory activity against A549 cells as compared to H1975 cells. It has been found that Aragus-C causes the induction of apoptosis and promotes cell-cycle arrest at the G2/M phase of A549 cells. It also showed a reduction in the overexpression of EGFR in A549 cells.In tumor xenograft mice model, it showed a significant reduction of tumor volume in a dose-dependent manner, with maximum inhibitory activity was reported by the 8 mg/kg treated group. It also showed significant anti-inflammatory and antioxidant activity by reducing the level of TNF-α, IL-1β, IL-6, and MDA, with a simultaneous increase of superoxide dismutase and glutathione peroxidase. We have demonstrated the potent anti-lung cancer activity of Aragus-C, and it may be considered as a potential therapeutic choice for NSCLC treatment.

Tilianin enhances the antitumor effect of sufentanil on non-small cell lung cancer.

Non-small cell cancer (NSCLC) is the most common cancer in the world, but its effective therapeutic methods are limited. Tilianin and sufentanil alleviate various human tumors. This research aimed to clarify the functions and mechanisms of Tilianin and sufentanil in NSCLC. The functions of Tilianin and sufentanil on NSCLC cell viability, apoptosis, mitochondrial dysfunction, and immunity in vitro were examined using Cell Counting Kit-8 assay, flow cytometry, reactive oxygen species level analysis, CD8+ T cell percentage analysis, Western blot, and enzyme-linked immunosorbent assay, respectively. The molecular mechanism regulated by Tilianin and sufentanil in NSCLC was assessed using Western blot, and immunofluorescence assays. Meanwhile, the roles of Tilianin and sufentanil in NSCLC tumor growth, apoptosis, and immunity in vivo were determined by establishing a tumor xenograft mouse model, immunohistochemistry, and Western blot assays. When sufentanil concentration was proximity 2 nM, the inhibition rate of NSCLC cell viability was 50%. The IC50 for A549 cells was 2.36 nM, and the IC50 for H1299 cells was 2.18 nM. The IC50 of Tilianin for A549 cells was 38.7 μM, and the IC50 of Tilianin for H1299 cells was 44.6 μM. Functionally, 0.5 nM sufentanil and 10 μM Tilianin reduced NSCLC cell (A549 and H1299) viability in a dose-dependent manner. Also, 0.5 nM sufentanil and 10 μM Tilianin enhanced NSCLC cell apoptosis, yet this impact was strengthened after a combination of Tilianin and Sufentanil. Furthermore, 0.5 nM sufentanil and 10 μM Tilianin repressed NSCLC cell mitochondrial dysfunction and immunity, and these impacts were enhanced after a combination of Tilianin and Sufentanil. Mechanistically, 0.5 nM sufentanil and 10 μM Tilianin repressed the NF-κB pathway in NSCLC cells, while this repression was strengthened after a combination of Tilianin and Sufentanil. In vivo experimental data further clarified that 1 µg/kg sufentanil and 10 mg/kg Tilianin reduced NSCLC growth, immunity, and NF-κB pathway-related protein levels, yet these trends were enhanced after a combination of Tilianin and Sufentanil. Tilianin strengthened the antitumor effect of sufentanil in NSCLC.

LINC00665 promotes the progression and immune evasion of lung cancer by facilitating the translation of TCF7 protein through dependence on IRES

ObjectiveTo investigate the influence of LINC00665 on the development and immune evasion of lung cancer.MethodsTumor tissues and corresponding adjacent tissues were collected from 84 lung cancer patients, categorized into non-metastatic (n = 58) and metastatic (n = 26) groups. LINC00665 expression in lung cancer and metastatic lung cancer tissues was assessed via qRT-PCR. Pearson correlation analysis was conducted to examine the correlation between LINC00665 and immune-modulating cytokines (TGF-β, IL-10, IL-1β, IFN-γ, IL-2, TNF-α). A549 and H1299 cells, with relatively high LINC00665 expression, were used for in vitro studies. Cells were transfected with LINC00665-targeting shRNA, and changes in proliferation, apoptosis, migration, invasion, and NK cell cytotoxicity were assessed. Downstream molecular mechanisms of LINC00665 were investigated using GEO database analysis, highlighting the association with HHLA2. LINC00665’s role in promoting HHLA2 expression via binding with TCF7 was explored. In low LINC00665-expressing A549/H1299 cells, overexpression of HHLA2 was performed to evaluate effects on malignant behavior and NK cell sensitivity. A xenograft model was established for in vivo validation through tumor volume and weight measurements, Ki-67 immunoreactivity analysis, and flow cytometry analysis of CD107a + NK cells.ResultsLINC00665, TCF7 mRNA, and HHLA2 mRNA expression levels were significantly higher in lung cancer tissues than adjacent tissues, with non-metastatic lung cancer showing higher expression than metastatic lung cancer. In metastatic lung cancer, LINC00665 positively correlated with immune-suppressive cytokines (TGF-β, IL-10, IL-1β) and negatively correlated with anti-tumor cytokines (IFN-γ, IL-2, TNF-α). LINC00665 knockdown significantly inhibited lung cancer cell growth and metastasis, promoting sensitivity to NK cells. Further analysis revealed that LINC00665 recruits transcription factor TCF7 to upregulate HHLA2 expression in lung cancer cells, thereby facilitating lung cancer development and immune escape.ConclusionLINC00665, through recruitment of TCF7 and upregulation of HHLA2, inhibits NK cell cytotoxicity, promoting the development and immune evasion of lung cancer.

S-(N,N-diethyldithiocarbamoyl)-N-acetyl-l-cysteine for the treatment of non-small cell lung cancer through regulating NF-κB signalling pathway without neurotoxicity

The discovery of novel targeted agents for non-small cell lung cancer (NSCLC) remains an important research landscape due to the limited efficacy, side effects and drug resistance of current treatment options. Among many repurposed drugs, disulphiram (DSF) has shown the potential to target tumours. However, its unpleasant neurotoxicity greatly limits its use. A DSF derivative, S-(N,N-diethyldithiocarbamoyl)-N-acetyl-l-cysteine (DS-NAC), was synthesised against NSCLC. The therapeutic effects, mechanism and toxicities of DS-NAC were evaluated in A549 and H460 cells and the mouse model of in situ lung cancer. The in vitro results exhibited that DS-NAC had potent anti-proliferation, apoptotic, anti-metastasis and epithelial–mesenchymal transition (EMT) inhibition effects. In the orthotopic lung cancer mouse model, therapeutic effects of DS-NAC were better than those of DSF and were similar to docetaxel (DTX). Also, results from western blot and immunohistochemistry showed that DS-NAC in combination with copper exerted therapeutic effects via regulating NF-κB signalling pathway and ROS-related proteins such as HIF-1α, Nrf2 and PKC-δ rather than regulating ROS level directly. Moreover, the safety evaluation study showed that DS-NAC had low haematologic and hepatic toxicities in comparison with DTX as well as low neurological toxicity compared with DSF. DS-NAC could be a promising anti-lung cancer agent with a favourable safety profile.

Cucurbitacin B targets STAT3 to induce ferroptosis in non-small cell lung cancer

Cucurbitacin B (CuB) is a compound found in plants like Cucurbitaceae that has shown promise in fighting cancer, particularly in lung cancer. However, the specific impact of CuB on ferroptosis and how it works in lung cancer cells has not been fully understood. Our research has discovered that CuB can effectively slow down the growth of non-small cell lung cancer (NSCLC) cells. Even in small amounts, it was able to inhibit the growth of various NSCLC cell lines. This inhibitory effect was reversed when ferroptosis inhibitors DFO, Lip-1 and Fer-1 were introduced. CuB was found to increase the levels of reactive oxygen species (ROS), lipid ROS, MDA, and ferrous ions within H358 lung cancer cells, leading to a decrease in GSH, mitochondrial membrane potential (MMP) and changes in ferroptosis-related proteins in a dose-dependent manner. These findings were also confirmed in A549 lung cancer cells. In A549 cells, different concentrations of CuB induced the accumulation of intracellular lipid ROS, ferrous ions and changes in ferroptosis-related indicators in a concentration-dependent manner. Meanwhile, the cytotoxic effect induced by CuB in A549 cells was counteracted by ferroptosis inhibitors DFO and Fer-1. Through network pharmacology, we identified potential targets related to ferroptosis in NSCLC cells treated with CuB, with STAT3 targets showing high scores. Further experiments using molecular docking and cell thermal shift assay (CETSA) revealed that CuB interacts with the STAT3 protein. Western blot and immunofluorescence staining demonstrated that CuB inhibits the phosphorylation of STAT3 (P-STAT3) in H358 cells. Silencing STAT3 enhanced CuB-induced accumulation of lipid ROS and iron ions, as well as the expression of ferroptosis-related proteins. On the other hand, overexpression of STAT3 reversed the effects of CuB-induced ferroptosis. The results indicate that CuB has the capability to suppress STAT3 activation, resulting in ferroptosis, and could be a promising treatment choice for NSCLC.