Growth Of A549 Cells Research Articles

Erythrocyte membrane-camouflaged gefitinib/albumin nanoparticles for tumor imaging and targeted therapy against lung cancer

Conventional chemotherapeutic drugs may cause serious side effects such as hepatotoxicity and renal toxicity due to lack of targeting, which affects therapy outcome and the prognosis of patients. Therefore, biomimetic nanoparticles with long blood circulation and active targeting have attracted increasing attention. In this work, we fabricated a biomimetic R-RBC@GEF-NPs nano-system by encapsulating gefitinib-loaded albumin nanoparticles (GEF-NPs) inside cRGD-modified red blood cell (RBC) membranes. The complete RBC membrane structure and membrane proteins enabled the NPs to escape phagocytosis by macrophages. In addition, the cRGD moiety significantly improved tumor cell targeting and uptake. R-RBC@GEF-NPs inhibited the growth of A549 cells in vitro in a dose- and time-dependent manner by inducing apoptosis and cell cycle arrest at the G1 phase. Likewise, the R-RBC@GEF-NPs also decreased tumor weight and volume in the mice injected with A549 cells and prolonged survival time. In addition, the 99Tc-labeled R-RBC@GEF-NPs selectively accumulated in the tumor tissues in vivo, and enabled real time tumor imaging. Finally, blood and histological analyses showed that R-RBC@GEF-NPs did not cause any obvious systemic toxicity. Taken together, the biomimetic R-RBC@GEF-NPs is a promising therapeutic formulation for the treatment of lung cancer.

Investigation of 3-sulfamoyl coumarins against cancer-related IX and XII isoforms of human carbonic anhydrase as well as cancer cells leads to the discovery of 2-oxo-2H-benzo[h]chromene-3-sulfonamide - A new caspase-activating proapoptotic agent.

Herein we report the synthesis of a set of seventeen 3-sulfonamide substituted coumarin derivatives. Prepared compounds were tested invitro for inhibition of four physiologically relevant isoforms of the metalloenzyme human carbonic anhydrase (hCA, EC 4.2.1.1). Several coumarin sulfonamides displayed low nanomolar KI values against therapeutically relevant hCA II, IX, and XII, whereas they did not potently inhibit hCA I. Some of these compounds exerted a concentration-dependent antiproliferative action toward RT4 human bladder cancer and especially A431 human epidermoid carcinoma cell lines. In the meantime, the viability of non-tumorigenic hTERT immortalized human foreskin fibroblast cell line Bj-5ta was not significantly affected by the obtained derivatives. Interestingly, compound 10q (2-oxo-2H-benzo [h]chromene-3-sulfonamide) showed a profound and selective dose-dependent inhibition of A431cell growth with low nanomolar IC50 values. We demonstrated that 10q possessed a concentration-dependent apoptosis induction activity associated with caspase 3/7 activation in cancer cells. As carbonic anhydrase isoforms in question were not potently inhibited by this compound, its antiproliferative effects likely involve other mechanisms, such as DNA intercalation. Compound 10q clearly represents a viable lead for further development of new-generation anticancer agents.

Identifying PIF1 as a Potential Target of Wenxia Changfu Formula in Promoting Lung Cancer Cell Apoptosis: Bioinformatics Analysis and Biological Evidence.

Lung cancer remains the leading cause of cancer-related deaths worldwide. Traditional Chinese medicine (TCM) is a valuable resource of active natural products and plays an important role in cancer treatment with the advantages of high efficiency and safety. Wenxia Changfu formula (WCF) is modified from Dahuang Fuzi decoction from Han Dynasty and has been used for treating lung cancer in China. Our previous research showed that WCF had an antitumor effect in vivo and in vitro, while the mechanism has not been well illustrated. In this study, the effect of WCF on the proliferative ability in three lung cancer cells and one noncancerous human cell line was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. WCF suppressed A549, H460, and PC-9 cell viability in a dose-dependent manner, with no inhibition of noncancerous MRC-5 cells after 48 h treatment with WCF (0–50 mg/mL). Furthermore, we screened for genes in A549 cells using four WCF-treated samples and four control samples on a gene expression profile microarray. 21 genes were significantly downregulated by WCF, which may potentially play an important role in the proliferation of A549 cells. High-content screening evaluated whether silencing the 21 genes affected A549 cell growth. The results showed that PIF1 knockdown exhibited the most potent inhibition of cell proliferation compared with the other genes. Downregulation of PIF1 increased A549 cell apoptosis and the activity of caspase 3/7. Besides, RT-PCR showed that the expression levels of PIF1 mRNA decreased significantly in A549, H460, and PC-9 cells after WCF treatment. In conclusion, the present observations indicate that WCF may inhibit lung cancer cell proliferation by promoting apoptosis via regulating the expression of PIF1.

The effect of Thymbra spicata extract and its bioactive component Thymol on non-small-cells lung cancer cell line A549

IntroductionAlternative medicine is important in cancer treatment. The apoptotic effect of Thymol and extracted Thymol from Thymbra spicata on non-small-cells lung cancer was studied.Material and methodsThymol was evaluated in Thymbra spicata extract by HPCL. Cell viability was assessed by MTT method. DCF and flu3-AM probe was used for ROS and cai2+ analysis, respectively. Western blotting was performed to measure NOX2 and Bax/Bcl-2 ratio.ResultsObtained data showed that Thymol was 1.51 mg/g in Thymbra spicata extract. Treatment with Thymol and extracted Thymol from Thymbra spicata resulted in cell death at high concentrations [LC50= 111±4.5 and 119±5.2 μM, respectively]. Subsequently, Thymbra spicata extract and its bioactive component increased ROS and Cai2+ production, NOX2, and Bax/Bcl-2 ratio.ConclusionsThis study revealed the anticancer effects of Thymol and Thymbra spicata extract on non-small-cells lung cancer and at least part of that effect was related to the increase in the NOX2 and Bax/Bcl-2 ratio. Our results demonstrated that TSE and Thymol at high concentrations (180, 120, and 80 μM) decreased the growth of A549 cells. It appeared that cytotoxic activity was exerted through activation of NOX2, ROS generation, increase in Cai2+, and Bax/Bcl-2 ratio. Present results demonstrated that TSE and thymol may be potential therapeutic agents for human lung cancer.

Depletion of ST6GALNACIII retards A549 non-small cell lung cancer cell proliferation by downregulating transferrin receptor protein 1 expression

Alterations in sialylation of terminal residues of glycoproteins have been implicated in forming tumor-associated glycans. ST6GALNAC transfers sialyl moiety to N-acetylgalactosamine residue via α2,6 linkage. Although the oncogenic characteristics of ST6GALNACI or II have been demonstrated in various cancer cells, the impact of ST6GALNACIII on tumor progression remains undefined. In this study, we evaluated the effect of ST6GALNACIII knockdown on the growth of A549 non-small cell lung cancer cells. ST6GALNACIII depletion resulted in significant retardation in growth of A549 cells under various culture conditions, including collagen-supported 3D culture and anchorage-independent soft agar culture conditions. Liquid chromatography with tandem mass spectrometry revealed that two glycopeptides of transferrin receptor protein 1 (TFR1) containing N-acetylhexosamine-sialic acid were not detected in ST6GALNACIII-depleted A549 cells compared with control cells. Subsequent lectin binding assay, western blotting, and real-time RT-PCR indicated that TFR1 sialylation was not significantly changed, but TFR1 protein and mRNA expressions were decreased after ST6GALNACIII knockdown. However, cell growth retardation by ST6GALNACIII knockdown was partially rescued by TFR1 overexpression. Additionally, TFR1 mRNA degradation was accelerated following ST6GALNACIII knockdown with concomitant reduction in mRNA levels of iron regulatory protein 1 and 2, the upstream regulators of TFR1 mRNA stability. Therefore, our results indicated an important role of ST6GALNACIII in promoting A549 cell growth through quantitative regulation of TFR1 expression and provided therapeutic implications for ST6GALNACIII targeting in tumor growth suppression in vivo.

Development and characterization of inhalable transferrin functionalized amodiaquine nanoparticles – Efficacy in Non-Small Cell Lung Cancer (NSCLC) treatment

New drug discovery and development processes encounter significant challenges including requirement of huge investments and lengthy time frames especially in cancer research field. Repurposing of old drugs against cancer provides a possible alternative while associated scale-up complexities with production of nanoparticles at industrial scale could be overcome by using a scalable nanoparticle technique. We previously described use of polymeric nanoparticles for inhaled delivery of amodiaquine (AQ) for non-small cell lung cancer (NSCLC) treatment. In this study, targeting potential of transferrin ligand conjugated inhalable AQ-loaded nanoparticles (Tf-AMQ NPs) was investigated against NSCLC. Tf-AMQ NP (liquid formulation) demonstrated an aerodynamic diameter of 4.4 ± 0.1 µm and fine particle fraction of 83.2 ± 3.0%, representing AQ deposition in the respirable region of airways. Cytotoxicity studies in NSCLC cell line with overexpressed transferrin receptors shown significant reduction in IC50 values with Tf-decorated AQ-loaded nanoparticles compared to AQ or non-targeted NPs, along with significant apoptosis induction (caspase assay) and reduced % colony growth in A549 and H1299 cells with Tf-AMQ NP. Furthermore, 3D spheroid studies (~7-fold reduction in spheroid volume compared to AMQ NPs) explained efficiency of conjugated nanoparticles in penetrating tumor core, and growth inhibition. AQ’s autophagy inhibition ability significantly increased with nanoparticle encapsulation and transferrin conjugation. In conclusion, amodiaquine can be an assuring candidate for repurposing to consider for NSCLC treatment while delivering inhalable transferrin conjugated nanoparticles developed using a scalable HPH process to the target site, thus reducing the dose, side effects.

PolyI:C suppresses TGF-β1-induced Akt phosphorylation and reduces the motility of A549 lung carcinoma cells.

Epithelial mesenchymal transition (EMT) is a critical process involved in the invasion and metastasis of cancer, including lung cancer (LC). Transforming growth factor (TGF)-β is one of factors capable of inducing EMT. Polyinosinic-polycytidylic acid (polyI:C), a synthetic agonist for toll-like receptor (TLR) 3, can enhance immune responses and has been used as an adjuvant for cancer vaccines; however, it remains unclear whether it influences other process, such as EMT. In the present study, we examined the effects of polyI:C on TGF-β-treated A549 human LC cells. By in vitro cell proliferation assay, polyI:C showed no effect on the growth of A549 cells treated with TGF-β1 at the concentration range up to 10μg/ml; however, it markedly suppressed the motility in a cell scratch and a cell invasion assay. By Western blotting, polyI:C dramatically decreased TGF-β1-induced Ak strain transforming (Akt) phosphorylation and increased phosphatase and tensin homologue (PTEN) expression without affecting the Son of mothers against decapentaplegic (Smad) 3 phosphorylation or the expression level of E-cadherin, N-cadherin or Snail, indicating that polyI:C suppressed cell motility independently of the 'cadherin switching'. The Akt inhibitor perifosine inhibited TGF-β1-induced cell invasion, and the PTEN-specific inhibitor VO-OHpic appeared to reverse the inhibitory effect of polyI:C. PolyI:C has a novel function to suppress the motility of LC cells undergoing EMT by targeting the phosphatidylinositol 3-kinase/Akt pathway partly via PTEN and may prevent or reduce the metastasis of LC cells.

Hirsuteine Inhibits the Proliferation of Human Non-Small Cell Lung Cancer (NSCLC) Cells by Inducing Apoptosis and Cell Cycle Arrest

Background: Hirsuteine is an alkaloid compound that can inhibit the proliferative activity of several cancer cell types in vitro, yet no prior reports have explored its ability to inhibit non-small cell lung cancer (NSCLC) growth. As such, herein, we sought to explore the antiproliferative activity of hirsuteine when used to treat human NSCLC A549 and NC I-H1299 cells across a range of tested concentrations (0–25 μM) and to explore the mechanisms underlying its therapeutic efficacy. Methods: The effects of hirsuteine on cell viability was examined via CCK-8 and colony formation assays, while apoptosis was assessed through Hoechst 33,258 staining and flow cytometry. Cell cycle progression was additionally evaluated via propidium iodide staining, while Western blotting and real-time quantitative polymerase chain reaction (qPCR) method were conducted to assess the levels of proteins and genes associated with apoptosis and cell cycle progression, respectively. Results: Hirsuteine markedly suppressed the proliferation of A549 and NCI-H1299 cells in a dose and time-dependent fashion and induced clear changes in cell morphology, resulting in G0-G1 phase cell cycle arrest that was related to the downregulation of Cyclin E and CDK2. Hirsuteine additionally induced robust apoptosis of A549 and NCI-H1299 cells and Bcl-2 downregulation together with the upregulation of Bax, Apaf1, cytoplasmic cytochrome C, cleaved caspase-3 and cleaved caspase-9, together driving this apoptotic cell death. Conclusion: Hirsuteine can effectively suppress the growth of human NSCLC A549 and NCI-H1299 cells and induce apoptotic death in a dose-dependent fashion in vitro , emphasizing the promising of this alkaloid compound as a potent anticancer treatment that warrants study as a treatment for human NSCLC patients.

Effects of Aspergillus fumigatus Conidia on Apoptosis and Proliferation in an In Vitro Model of the Lung Microenvironment.

Background/Aim: Aspergillus is often detected in respiratory samples from patients with chronic respiratory diseases, including pulmonary fibrosis, suggesting that it can easily colonize the airways. To determine the role of Aspergillus colonization in pulmonary fibrosis, we cultured human lung epithelial A549 cells or murine embryo fibroblast NIH/3T3 cells with Aspergillus conidia in 3D floating culture representing the microenvironment. Materials and Methods: Cells were cultured in two-dimensional (2D) and three-dimensional floating (3DF) culture with heat-inactivated Aspergillus fumigatus (AF) 293 conidia at an effector-to-target cell ratio of 1:10 (early-phase model) and 1:100 (colonization model), and RNA-sequencing and Western blots (WB) were performed. Results: AF293 conidia reduced A549 cell growth in 2D and 3DF cultures and induced apoptosis in A549 spheroids in 3DF culture. RNA-sequencing revealed the increased expression of genes associated with interferon-mediated antiviral responses including MX dymamin-like GTPase 1 (MX1). Interestingly, the decreased expression of genes associated with the cell cycle was observed with a high concentration of AF293 conidia. WB revealed that epithelial-mesenchymal transition was not involved. Notably, AF293 conidia increased NIH/3T3 growth only in 3DF culture without inducing an apoptotic reaction. RNA-sequencing revealed the increased expression of genes associated with interferon signalling, including MX2; however, the decreased expression of genes associated with the cell cycle was not observed. Conclusions: AF affects both apoptosis of epithelial cells and the growth of fibroblasts. A deeper understanding of the detailed mechanisms underlying Aspergillus-mediated signaling pathway in epithelial cells and fibroblasts will help us to understand the lung microenvironment.

Abstract LB099: The inhibitory effect of pemetrexed disodium heptahydrate on the growth of A549 cells

Abstract Background: Pemetrexed disodium heptahydrate (pemetrexed) is currently used for the treatment of patients with advanced non-small cell lung cancer (NSCLC). More than 30% of lung cancer cases involve KRAS mutations. In this study, we examined the anticancer ability of pemetrexed in KRAS-dependent A549 lung cancer cells. Methods: A549 cells were treated with pemetrexed for 72 h and then evaluated by a TUNEL assay, ROS assay and Western blotting. Results: The apoptotic effect of pemetrexed was observed through increased levels of cleaved caspase-9 (Asp 315), cleaved caspase-3 (Asp 175) and cleaved PARP. Furthermore, pemetrexed increased ROS-mediated DNA damage and enhanced the proapoptotic Bax protein expression, leading to the loss of mitochondrial membrane potential (MMP). The increased expression of the β-galactosidase protein was correlated with the induction of cellular senescence. Significantly, pemetrexed induced cytotoxicity without inhibiting the RAS/RAF/MEK/ERK signaling pathway, while pemetrexed induced autophagy, which activated the AMPK/mTOR signaling pathway. Conclusion: Taken together, these findings indicate that, in addition to apoptosis, pemetrexed-induced autophagy and cellular senescence, providing a promising direction for the development of KRAS-targeted therapy. Citation Format: Md Mohiuddin, Hideharu Kimura, Takashi Sone, Satoshi Watanabe, Hiroki Matsuoka, Keigo Saeki, Nanao Terada, Miki Abo, Kazuo Kasahara. The inhibitory effect of pemetrexed disodium heptahydrate on the growth of A549 cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB099.

Swertia chirayita suppresses the growth of non-small cell lung cancer A549 cells and concomitantly induces apoptosis via downregulation of JAK1/STAT3 pathway.

Lung carcinoma is the leading cause of cancer-related mortalities worldwide, and present therapeutical interventions are not successful enough to treat this disease in many cases. Recent years have witnessed a surge in exploring natural compounds for their antiproliferative efficacy to expedite the characterization of novel anticancer chemotherapeutics. Swertia chirayita is a valued medicinal herb and possess intrinsic pharmaceutical potential. However, elucidation of its anticancer effects at molecular levels remains unclear and needs to be investigated. We assessed the anticancer and apoptotic efficacy of S. chirayita ethanolic extract (Sw-EtOH) on non-small cell lung cancer (NSCLC) A549 cells during this exploratory study. The results elucidated that S. chirayita extract induced toxic effects within lung cancer cells by ~1 fold during cytotoxicity and LDH release assay at a 400 μg/ml concentration. Sw-EtOH extract elevates the level of ROS, resulting in the disruption of Δψm and release of cytosolic cytochrome c by 3.15 fold. Activation of caspases-3, -8 & -9 also escalated by ~1 fold, which further catalyze the augmentation of PARP cleavage (~3 folds), resulting in a four-fold increase in Sw-EtOH induced apoptosis. The gene expression analysis further demonstrated that Sw-EtOH extracts inhibited JAK1/STAT3 signaling pathway by down-regulating the levels of JAK1 and STAT3 to nearly half a fold. Treatment of Sw-EtOH modulates the expression level of various STAT3 associated proteins, including Bcl-XL, Bcl-2, Mcl-1, Bax, p53, Fas, Fas-L, cyclinD1, c-myc, IL-6, p21 and p27 in NSCLC cells. Thus, our study provided a strong impetus that Sw-EtOH holds the translational potential of being further evaluated as efficient cancer therapeutics and a preventive agent for the management of NSCLC.

Knockdown of Myeloid Cell Leukemia-1 by MicroRNA-101 Increases Sensitivity of A549 Lung Cancer Cells to Etoposide.

Background: Studies have shown that myeloid cell leukemia-1 (Mcl-1) is the target gene for microRNA -101 (miRNA-101), and decreased levels of miRNA-101 are associated with elevated levels of Mcl-1 and lung cancer survival. The objective of the present study was to investigate the effect of miRNA-101 on the sensitivity of A549 lung cancer cells to etoposide. Methods: The study was conducted during 2018 and 2019 at Arak University of Medical Sciences, Arak, Iran. The effect of miRNA-101 on Mcl-1 expression was assessed using reverse transcription-quantitative polymerase chain reaction 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT), and trypan blue exclusion assays were performed to determine the effect of treatments on cell survival and proliferation, respectively. The interaction between miRNA-101 and etoposide was evaluated using the combination index analysis of Chou-Talalay. Apoptosis was quantified using ELISA cell death assay. ANOVA and Bonferroni’s tests were used to determine statistical differences between the groups (P<0.05). GraphPad Prism software (version 6.01) was used for data analysis.Results: The results showed that miRNA-101 clearly inhibited the expression of Mcl-1 and reduced the growth of A549 cells, relative to blank control and negative control miRNA (P<0.05). Transfection of miRNA-101 synergistically enhanced the sensitivity of the A549 cells to etoposide. Apoptosis assay data also showed that miRNA-101 triggered apoptosis and augmented the etoposide-mediated apoptosis. Conclusion: Up-regulation of miRNA-101 inhibited cell survival and proliferation, and sensitized A549 cells to etoposide by suppressing Mcl-1 expression. miRNA-101 replacement therapy can be considered as an effective therapeutic strategy in non-small cell lung cancer.

Decreased TMEM40 expression is associated with malignant behavior of cutaneous squamous cell carcinoma and inhibits tumor progression.

Cutaneous squamous cell carcinoma (CSCC) is one of the most common types of skin cancer in humans worldwide. The identification and characterization of cancer-associated transmembrane proteins are important for understanding the molecular biology of CSCC. The aim of the present study was to evaluate the expression pattern of transmembrane protein 40 (TMEM40) in CSCC and its clinical significance. The underlying mechanisms were also examined. Reverse transcription-quantitative PCR, western blot and immunohistochemistry analysis were used to determine the relative expression of TMEM40 in CSCC cell lines and clinical tissue samples. The effect of TMEM40 gene silencing on cell proliferation was also evaluated using Cell Counting Kit-8 assays. Wound healing assays, flow cytometry and Transwell assays were used to explore the migration, cell cycle distribution/apoptosis and invasion of CSCC cells following TMEM40 silencing, respectively. In the present study, increased TMEM40 expression was observed in CSCC tissue samples, compared with normal skin, and TMEM40 expression was associated with large tumor size in patients with CSCC. In vitro functional assays indicated that TMEM40 was involved in the regulation of A431 and SCL1 cell growth through its effects on the cell cycle and apoptosis. Silencing TMEM40 in A431 and SCL1 cells resulted in cell cycle arrest at the G0/G1 phase and promoted apoptosis. In addition, migration and invasion were significantly inhibited following silencing of TMEM40 expression in CSCC cells. Taken together, the results of the present study indicated that reduced TMEM40 expression could inhibit CSCC development and that TMEM40 may represent a therapeutic target in CSCC.

Hyperbaric oxygen suppressed tumor progression through the improvement of tumor hypoxia and induction of tumor apoptosis in A549-cell-transferred lung cancer

Tumor cells have long been recognized as a relative contraindication to hyperbaric oxygen treatment (HBOT) since HBOT might enhance progressive cancer growth. However, in an oxygen deficit condition, tumor cells are more progressive and can be metastatic. HBOT increasing in oxygen partial pressure may benefit tumor suppression. In this study, we investigated the effects of HBOT on solid tumors, such as lung cancer. Non-small cell human lung carcinoma A549-cell-transferred severe combined immunodeficiency mice (SCID) mice were selected as an in vivo model to detect the potential mechanism of HBOT in lung tumors. HBOT not only improved tumor hypoxia but also suppressed tumor growth in murine xenograft tumor models. Platelet endothelial cell adhesion molecule (PECAM-1/CD31) was significantly increased after HBOT. Immunostaining of cleaved caspase-3 was demonstrated and apoptotic tumor cells with nuclear debris were aggregated starting on the 14th-day after HBOT. In vitro, HBOT suppressed the growth of A549 cells in a time-dependent manner and immediately downregulated the expression of p53 protein after HBOT in A549 cells. Furthermore, HBOT-reduced p53 protein could be rescued by a proteasome degradation inhibitor, but not an autophagy inhibitor in A549 cells. Our results demonstrated that HBOT improved tissue angiogenesis, tumor hypoxia and increased tumor apoptosis to lung cancer cells in murine xenograft tumor models, through modifying the tumor hypoxic microenvironment. HBOT will merit further cancer therapy as an adjuvant treatment for solid tumors, such as lung cancer.

Dual Enzyme-Like Performances of PLGA Grafted Maghemite Nanocrystals and Their Synergistic Chemo/Chemodynamic Treatment for Human Lung Adenocarcinoma A549 Cells.

In recent years, the emergence of non-toxic but catalytically active inorganic nanoparticles has attracted great attention for cancer treatment, but the therapeutic effect has been affected by the limited reactive oxygen species in tumors. Therefore, the combination of chemotherapy and chemodynamic therapy is regarded as a promising therapeutic strategy. In this paper, we reported the preparation and bioactivity evaluation of poly(lactic acid-co-glycolic acid) (PLGA) grafted-γ-Fe₂O₃ nanoparticles with dual response of endogenous peroxidase and catalase like activities. Our hypothesis is that PLGAgrafted γ-Fe₂O₃ nanoparticles could be used as a drug delivery system for the anti-tumor drug doxorubicin to inhibit the growth of lung adenocarcinoma A549 cells; meanwhile, based on its mimic enzyme properties, this kind of nanoparticles could be combined with doxorubicin in the treatment of A549 cells. Our experimental results showed that the PLGAgrafted γ-Fe₂O₃ nanoparticles could simulate the activity of catalase and decompose hydrogen peroxide into H₂O and oxygen in neutral tumor microenvironment, thus reducing the oxidative damage caused by hydrogenperoxide to lung adenocarcinoma A549 cells. In acidic microenvironment, PLGA grafted γ-Fe₂O₃ nanoparticles could simulate the activity of peroxidase and effectively catalyze the decomposition of hydrogen peroxide to generate highly toxic hydroxyl radicals, which could cause the death of A549 cells. Furthermore, the synergistic effect of peroxidase-like activity of PLGA-grafted γ-Fe₂O₃ nanoparticles and doxorubicin could accelerate the apoptosisand destruction of A549 cells, thus enhancing the antitumor effect of doxorubicin-loaded PLGA-grafted γ-Fe₂O₃ nanoparticles. Therefore, this study provides an effective nanoplatform based on dual inorganic biomimetic nanozymes for the treatment of lung cancer.

Inhibition effect of oxyepiberberine isolated from Coptis chinensis franch. On non-small cell lung cancer based on a network pharmacology approach and experimental validation

Ethnopharmacological relevanceAs an important Chinese herb, Coptis chinensis Franch. (Huanglian, HL) has a long history of usage for clearing heat, eliminating dampness, purging fire and detoxification in Traditional Chinese Medicine (TCM). HL, also called goldthread, was frequently used for the treatment of typhoid, tuberculosis, epidemic cerebrospinal meningitis, pertussis, and other lung-related diseases. Modern research has shown that HL and its main compounds also have anti-tumor effects. However, studies have not reported whether its main compounds inhibit Non-small cell lung cancer (NSCLC) development and progression. ObjectiveThis study aimed to find out the potential targets and mechanisms of Oxyepiberberine (OPB) isolated from HL in the treatment of NSCLC, using network pharmacology and biological experimental. MethodsSilica gel chromatography column was used to isolate OPB from HL, and the structure of OPB was elucidated using different spectroscopic analysis methods, including 1H-nuclear magnetic resonance (NMR), 13C-NMR and electrospray ionization mass spectrometry (ESI/MS). MTT assay was performed to determine cell proliferation of OPB on A549, H1975 and BEAS-2B cells. Then, the potential targets, pathways and hub genes of OPB for treating NSCLC were screened out through network pharmacology. Based on the results of network pharmacology, core targets of OPB for treating NSCLC were docking with OPB via molecular docking. Wound healing, plate clone, Hoechst staining, and western blot assay were used to verify the function of OPB in treatment of NSCLC. ResultsOPB was isolated from the HL, its molecular formula was identified as C20H17NO5. Through MTT, OPB significantly inhibited the proliferation of H1975 cells and A549 cells, and A549 was chosen as the test cancer cell. Through network pharmacology, 22 potential targets, 156 related-pathways, and 6 hub genes were screened out. The results of molecular docking showed that SRC, BRAF, and MMP9 were the core targets of OPB against NSCLC. Through biological experimental, it was found that OPB inhibited growth and migration of A549 cells. In addition, OPB induced apoptosis in A549 cells. Through western blot assay, the expressions of Src, ERK1/2 and other four proteins were down-regulated, which suggested that OPB inhibited the proliferation of lung cancer cells by down-regulating SRC-FAK-RAS-RAF-MEK-ERK pathway, so as to achieve the anti-NSCLC effect. ConclusionOur study demonstrated that anti-NSCLC effect of OPB through network and experiments, which provided a theoretical basis for the clinical antitumor of OPB, and provided a foundation for further study of OPB.

Nelfinavir restricts A549 cell growth by inhibiting STAT3 signaling.

ObjectiveTo investigate the anticancer effect of nelfinavir (NFV) on human A549 cells.MethodsThe inhibitory effects of NFV on the proliferation of human A549 cells were assessed using a MTT assay. Apoptotic cells were observed by fluorescence microscopy following Hoechst 33342 staining. Apoptosis of A549 cells was assessed using Annexin-V/propidium iodide staining and flow cytometry. Expression levels of signal transducer and activator of transcription 3 (STAT3) and p-STAT3 were measured by western blotting. STAT3 RNA silencing was used to investigate the pro-apoptotic mechanism of NFV in A549 cells.ResultsNFV dose-dependently suppressed proliferation of human A549 cells and induced significant apoptosis. Western blotting showed that the antitumor function of NFV might be mediated by STAT3 inhibition. A549 cell apoptosis in response to 20 µM NFV was significantly increased following STAT3 silencing. NFV significantly impeded the expression of the anti-apoptotic proteins Bcl-xL and Bcl-2, by increased the expression of the pro-apoptotic protein Cle-PARP.ConclusionsOur findings highlight STAT3 as a promising therapeutic target. NFV is a novel anti-cancer drug for the treatment of non-small-cell lung cancer.

A Combined Phytochemistry and Network Pharmacology Approach to Reveal Potential Anti-NSCLC Effective Substances and Mechanisms in Marsdenia tenacissima (Roxb.) Moon (Stem).

Marsdeniae tenacissimae Caulis is a traditional Chinese medicine, named Tongguanteng (TGT), that is often used for the adjuvant treatment of cancer. In our previous study, we reported that an ethyl acetate extract of TGT had inhibitory effects against adenocarcinoma A549 cells growth. To identify the components of TGT with anti-tumor activity and to elucidate their underlying mechanisms of action, we developed a technique for isolating compounds, which was then followed by cytotoxicity screening, network pharmacology analysis, and cellular and molecular experiments. We isolated a total of 19 compounds from a TGT ethyl acetate extract. Two novel steroidal saponins were assessed using an ultra-performance liquid chromatography-photodiode array coupled with quadrupole time-of-flight mass (UPLC-ESI-Q/TOF-MS). Then, we screened these constituents for anti-cancer activity against non-small cell lung cancer (NSCLC) in vitro and obtained six target compounds. Furthermore, a compound-target-pathway network of these six bioactive ingredients was constructed to elucidate the potential pathways that controlled anticancer effects. Approximately 205 putative targets that were associated with TGT, as well as 270 putative targets that were related to NSCLC, were obtained from online databases and target prediction software. Protein–protein interaction networks for drugs as well as disease putative targets were generated, and 18 candidate targets were detected based on topological features. In addition, pathway enrichment analysis was performed to identify related pathways, including PI3K/AKT, VEGF, and EGFR tyrosine kinase inhibitor resistance, which are all related to metabolic processes and intrinsic apoptotic pathways involving reactive oxygen species (ROS). Then, various cellular experiments were conducted to validate drug-target mechanisms that had been predicted using network pharmacology analysis. The experimental results showed the four C21 steroidal saponins could upregulate Bax and downregulate Bcl-2 expression, thereby changing the mitochondrial membrane potential, producing ROS, and releasing cytochrome C, which finally activated caspase-3, caspase-9, and caspase-8, all of which induced apoptosis in A549 cells. In addition, these components also downregulated the expression of MMP-2 and MMP-9 proteins, further weakening their degradation of extracellular matrix components and type IV collagen, and inhibiting the migration and invasion of A549 cells. Our study elucidated the chemical composition and underlying anti-tumor mechanism of TGT, which may be utilized in the treatment of lung cancer.

Alopecuroidines A−C, three matrine-derived alkaloids from the seeds of Sophora alopecuroides

Three matrine-derived alkaloids, alopecuroidine A (1), alopecuroidine B (2a) and alopecuroidine C (2b) were isolated from the seeds of Sophora alopecuroides. Their structures were elucidated by extensive spectroscopic analyses and X-ray diffraction. Three compounds possess an unprecedented rearranged fused 7/6/5/6 tetracyclic skeleton with a diazacycloheptane structure. Their plausible biosynthetic pathway was also proposed. The anti-proliferative activities of compounds 1 and 2a were examined by the MTT assay. Compound 1 inhibited the viability of human lung cancer A549 cells, having a half maximal inhibitory concentration (IC50) of 7.58 ± 2.47 µmol/L at 72 h. The flow cytometric analysis suggested that 1 inhibited A549 cell growth by inducing apoptosis and cell-cycle arrest. Additionally, 1 induced the loss of mitochondrial membrane potential, elevated intracellular reactive oxygen species, increased the Bax/Bcl-2 ratio, stimulated cleaved-caspase-3 and P53 protein levels, and suppressed the pro-caspase-3 level. Thus, 1 appeared to induce A549 cells apoptosis through a mitochondria-mediated apoptotic pathway.

Saturated Oxo Fatty Acids (SOFAs): A Previously Unrecognized Class of Endogenous Bioactive Lipids Exhibiting a Cell Growth Inhibitory Activity

The discovery of novel bioactive lipids that promote human health is of great importance. Combining "suspect" and targeted lipidomic liquid chromatography-high-resolution mass spectrometry (LC-HRMS) approaches, a previously unrecognized class of oxidized fatty acids, the saturated oxo fatty acids (SOFAs), which carry the oxo functionality at various positions of the long chain, was identified in human plasma. A library of SOFAs was constructed, applying a simple green photochemical hydroacylation reaction as the key synthetic step. The synthesized SOFAs were studied for their ability to inhibit in vitro the cell growth of three human cancer cell lines. Four oxostearic acids (OSAs) were identified to inhibit the cell growth of human lung carcinoma A549 cells. 6OSA and 7OSA exhibited the highest cell growth inhibitory potency, suppressing the expression of both STAT3 and c-myc, which are critical regulators of cell growth and proliferation. Thus, naturally occurring SOFAs may play a role in the protection of human health.