Lung Cancer Cell Line H1299 Research Articles

Phytochemical evaluation and Chemical characterization of Bauhinia variegata L. bark extract by TGA/ DSC, FT-IR and GC-MS analytical techniques: Pharmaceutical Aspects

Bauhinia variegata (Linn.), Caesalpiniaceae, known as Kachnar in Ayurvedic texts has been shown to have promising anticancer potential. The presence of bioactive molecules in plants which can be commercialized has led to research in knowing their chemical properties. Previous studies from our laboratory have validated the claim of anticancer property of Bauhinia variegata by providing biochemical evidence for the effect of petroleum ether bark extract (PEBE) and chloroform bark extract (CBE) on A549 and H460 lung cancer cell lines respectively. This work aimed at the application of various analytical techniques (thermal analysis, Fourier infrared transform spectroscopy and GC-MS) in the characterization of dried extracts of Bauhinia variegata with medicinal properties. TG-DSC-FTIR-GC-MS was applied to monitor the thermal stability and chemical properties of Bauhinia variegata bark extracts. The TG analysis of PEBE and CBE showed mass loss within three steps in different temperature ranges. The DSC curves for the dried extracts of Bauhinia variegata showed that thermal processes occur between 50 – 100°C for PEBE and 50-70°C for CBE. The FTIR spectroscopic studies of petroleum ether and chloroform bark extracts of Bauhinia variegata revealed different characteristic peak values with various functional groups in the extracts. The GC-MS study of PEBE revealed 18 compounds and CBE revealed 8 compounds. Thus, we have been successful in narrowing down to very few probable anticancer compounds from Bauhinia variegata bark extracts and also through this study, it was possible to substantiate the possibility of applying these techniques for use in the herbal medicines production from medicinal plants.

Abstract 422: Novel approach to destabilization of oncogenic mutant p53 and therapeutic implications

Abstract Introduction: Mutant p53 drives several hallmarks of cancer through a gain-of-function oncogenic program. The oncogenicity of mutant p53 includes sustained proliferation, increased survival and resistance to therapy, increased invasion and metastasis, and genomic instability. The oncogenicity of mutant p53 is related to its stability and accumulation in cells by evading degradation by the proteasome. Therefore, destabilization of mutant p53 has been sought as a therapeutic strategy, but so far without clinical success. We and others observe that proteasome inhibitors cause paradoxical depletion of mutant p53 in lung cancer cell lines. Objectives: In this project, we sought to determine the mechanism(s) of mutant p53 depletion by proteasome inhibition and its therapeutic implications in lung cancer. Methods: Human lung cancer cell lines bearing mutant p53 and wildtype p53 (WTp53) were used in this study. An isogenic panel of cell lines with mutant p53, WTp53 and a vector control was also used to determine mutant p53 dependency of the observed effects. Western blotting, immunoprecipitation, siRNA technology, DNA transfection were used for protein detection, protein-protein interactions, gene knock-down and overexpression respectively. Cycloheximide chase assay was used to determine protein degradation. The cancer genome atlas (TCGA) analysis was performed using the cBioPortal platform. Results: Proteasome inhibitors resulted in depletion of mutant p53 in the lung cancer cell line H1975. This effect was also confirmed using the isogenic model described above. Depletion of mutant p53 occurred through a degradation mechanism and was time dependent. While WTp53 accumulated at initial timepoints of proteasome inhibition, mutant p53 did not accumulate but instead depleted with time. This suggested that the proteasome is not the primary mechanism for mutant p53, in contrast to WTp53. Since autophagy is an alternative mechanism of protein degradation in eukaryotes, we evaluated for, and found evidence of autophagy upon proteasome inhibition. We further showed that autophagic degradation of mutant p53 could involve the heat shock protein 70. Our TCGA data analysis showed proteasome genes amplification in mutant p53 lung cancers, suggesting a possible functional dependency. Furthermore, RNA sequencing analysis showed upregulation of most proteasome genes upon stable expression of the R273H mutant in p53-null lung cancer cell line. Finally, in vitro cell viability assays using two different proteasome inhibitors showed increased sensitivity of the lung cancer cell line with mutant p53 compared to WTp53. Conclusion: The cumulative evidence of proteasome hyperactivity, degradation of mutant p53 and in vitro evidence of increased susceptibility of mutant p53 lung cancer cell lines suggests that proteasome inhibitors warrant a closer attention for therapeutic targeting of mutant p53 in lung cancer. Citation Format: Eziafa I. Oduah, Larisa Litovchick, Steven R. Grossman. Novel approach to destabilization of oncogenic mutant p53 and therapeutic implications [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 422.

Abstract LB214: Discovery of EPIM-001, a tumor targeted engineered IL2 with biased IL2Rβ agonist activities

Abstract Background: The cytokine Interleukin 2 (IL2) has been approved for cancer treatment. However, its application has been limited due to severe toxicities associated with its high concentration administration. These side effects are attributed to the strong binding of IL2 to its receptor IL2Rα which is enriched on regulatory T Cells and Lung epithelial cells. We have engineered IL2 to significantly reduce IL2Rα binding and enhance IL2Rβ binding. We further developed EPIM-001, a bi-functional IL2Rβ agonist and Anti-PDL1 biologic, aiming to treat a broad set of solid tumors with improved therapeutic index. Methods: We used mSCAFold࣪, a unique protein engineering platform to redefine and improve pharmacologic properties of the cytokine through structure-based rationally designed libraries and mRNA display selection. The optimal IL2 candidate with enhanced IL2Rβ and reduced IL2Rα binding was engineered with an internally discovered anti-PD-L1 antibody to generate EPIM-001, a bispecific IL2/PD-L1 biologic drug candidate. EPIM-001 was characterized in vitro for receptor binding activity, immune cell activation, proliferation, and tumor cell killing assays. In vivo efficacy was evaluated in xenograft humanized mouse models, and PK/PD and toxicity has been studied in Non-Human Primates (NHPs). Results: We have shown that EPIM-001 is a strong agonist of IL2Rβ, with significantly reduced binding to IL2Rα. It demonstrated much stronger CD8+ T cells and NK cell activation and proliferation compared WT-IL2, whilst only weaker activation and minimal proliferation with Tregs. In vitro cell killing assays showed that EPIM-001 promoted better immune cell killing of PD-L1 expressing target cells than WT IL2. In xenograft in vivo studies, EPIM-001 has shown significant tumor growth inhibition against the human lung cancer cell line H1975, which whilst expressing high levels of PD-L1, does not respond to PD-L1/PD-1 blockade. Furthermore, in the human breast cancer cell line MDA-MB-231, EPIM-001 showed stronger tumor growth inhibition compared to Keytruda. PK/PD and toxicity studies in NHPs showed that EPIM-001 is well tolerated. EPIM-001 greatly expanded CD8+ T cells, whilst having minimal effect on TRegs and Eosinophils associated with toxicity. Conclusions: EPIM-001 is a promising immune modulating drug candidate that selectively activates CD8+ T cells and NK Cells with strong anti-tumor activity and safe profile for both immune checkpoint sensitive and insensitive PD-L1 positive tumors. Citation Format: Kehao Zhao, Sam Hassan, Yan Chen, Jenna Nguyen, Ning Jiang, Gening Zhang, Irene Li, Ryan Feng. Discovery of EPIM-001, a tumor targeted engineered IL2 with biased IL2Rβ agonist activities [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB214.

Bone morphogenetic protein signaling regulation of AMPK and PI3K in lung cancer cells and C. elegans

BackgroundBone morphogenetic protein (BMP) is a phylogenetically conserved signaling pathway required for development that is aberrantly expressed in several age-related diseases including cancer, Alzheimer’s disease, obesity, and cardiovascular disease. Aberrant BMP signaling in mice leads to obesity, suggesting it may alter normal metabolism. The role of BMP signaling regulating cancer metabolism is not known.MethodsTo examine BMP regulation of metabolism, C. elegans harboring BMP gain-of-function (gof) and loss-of-function (lof) mutations were examined for changes in activity of catabolic and anabolic metabolism utilizing Western blot analysis and fluorescent reporters. AMP activated kinase (AMPK) gof and lof mutants were used to examine AMPK regulation of BMP signaling. H1299 (LKB1 wild-type), A549 (LKB1 lof), and A549-LKB1 (LKB1 restored) lung cancer cell lines were used to study BMP regulation of catabolic and anabolic metabolism. Studies were done using recombinant BMP ligands to activate BMP signaling, and BMP receptor specific inhibitors and siRNA to inhibit signaling.ResultsBMP signaling in both C. elegans and cancer cells is responsive to nutrient conditions. In both C. elegans and lung cancer cell lines BMP suppressed AMPK, the master regulator of catabolism, while activating PI3K, a regulator of anabolism. In lung cancer cells, inhibition of BMP signaling by siRNA or small molecules increased AMPK activity, and this increase was mediated by activation of LKB1. BMP2 ligand suppressed AMPK activation during starvation. BMP2 ligand decreased expression of TCA cycle intermediates and non-essential amino acids in H1299 cells. Furthermore, we show that BMP activation of PI3K is mediated through BMP type II receptor. We also observed feedback signaling, as AMPK suppressed BMP signaling, whereas PI3K increased BMP signaling.ConclusionThese studies show that BMP signaling suppresses catabolic metabolism and stimulates anabolic metabolism. We identified feedback mechanisms where catabolic induced signaling mediated by AMPK negatively regulates BMP signaling, whereas anabolic signaling produces a positive feedback regulation of BMP signing through Akt. These mechanisms were conserved in both lung cancer cells and C. elegans. These studies suggest that aberrant BMP signaling causes dysregulation of metabolism that is a potential mechanism by which BMP promotes survival of cancer cells.

Exosomal lncRNA LINC01356 Derived From Brain Metastatic Nonsmall-Cell Lung Cancer Cells Remodels the Blood-Brain Barrier.

Brain metastasis is a severe complication that affects the survival of lung cancer patients. However, the mechanism of brain metastasis in lung cancer remains unclear. In this study, we constructed an in vitro BBB model and found that cells from the high-metastatic nonsmall cell lung cancer (NSCLC) cell line H1299 showed a higher capacity to pass through the blood–brain barrier (BBB), as verified by Transwell assays, than cells from the low-metastatic NSCLC cell line A549. Brain microvascular endothelial cells (BMECs) could internalize H1299-derived exosomes, which remarkably promoted A549 cells across the BBB. The BBB-associated exosomal long noncoding RNA (lncRNA) was selected from the RNA-Seq dataset (GSE126548) and verified by real-time PCR and Transwell assays. LncRNA LINC01356 was significantly upregulated in H1299 cells and exosomes derived from these cells compared to that of A549 cells. Moreover, LINC01356 was also upregulated in serum exosomes of patients with NSCLC with brain metastasis compared with those without metastasis. In addition, BMECs treated with LINC01356-deprived exosomes expressed higher junction proteins than those treated with the control exosomes, and silencing LINC01356 in exosomes derived from H1299 cells could inhibit A549 cells from crossing the BBB. These data might indicate that the exosomal lncRNA LINC01356 derived from brain metastatic NSCLC cells plays a key role in remodeling the BBB system, thereby participating in brain metastasis in lung cancer.

<i>Bauhinia variegata</i> Bark Extract: Assessment of its Anti-proliferative and Apoptotic Activities on A549 and H460 Lung Cancer Cell Lines

The hunt for novel anticancer drugs with minimal side effects continues. This study strengthens the claim by providing biochemical evidences of anticancer activities of<em> Bauhinia variegata</em> bark extracts on lung carcinoma cells (A549 and H460). Bark extracts of <em>Bauhinia variegata</em> were prepared by different solvents using Soxhlet apparatus and tested for their antioxidant potential by DPPH assay. The lung cancer cell lines were treated with <em>Bauhinia variegata</em> bark extracts and viability of cells was measured by MTT assay; metastatic ability was determined through Scratch assay and effect on DNA integrity was shown by gel electrophoresis. The Petroleum Ether Bark Extract (PEBE) inhibits proliferation (A549, IC50 = 1.5 mg/ml) at 48 h treatment. DNA damage was observed in A549 cells by agarose gel electrophoresis. The Chloroform Bark Extract (CBE) inhibited proliferation of H460 (IC50 = 1 mg/ml) with DNA damage after 24 h treatment. Soft agar assay indicated decreased ability to form colonies and scratch test showed impaired migration of A549 and H460 to PEBE and CBE treatment respectively. Apoptosis was detected using fluorescent dye staining in A549 and H460 cells. Caspase 3 activity was increased significantly in A549 and H460 cells. PEBE and CBE decrease the mitochondrial membrane potential gradient (??m) of A549 and H460 cells respectively. This study categorically proves the cytotoxic activity of <em>Bauhinia variegata</em> bark extracts on A549 and H460 cells.

Expression of EPHA5 in lung adenocarcinoma is associated with lymph node metastasis and EGFR mutation

EPHA5 is a member of the Eph family of tyrosine kinase receptors, which affect carcinogenesis. The expression level of the EPHA5 receptor in a set of lung adenocarcinoma tissue samples was checked using immunohistochemistry. The relationship between EPHA5 expression and clinicopathological parameters, and epidermal growth factor receptor (EGFR) and Braf mutations were analyzed. We also checked the expression level of the EPHA5 receptor in four lung cancer cell lines. High expression of EPHA5 was found in NCI-H460 and H1299 cells, while low expression was observed in A549 and SPC-A1 cells. EPHA5 was knocked down in NCI-H460 and H1299 lung cancer cell lines using siRNAs. The proliferation, clone formation, and invasive ability were analyzed in NCI-H460 and H1299 cells with EPHA5 knockdown. The results show that the EPHA5 receptor is differently expressed in lung adenocarcinoma tissues, in which positive and negative expression of EPHA5 was found in 58.1% and 41.9% of tissues, respectively. Positive expression of EPHA5 was associated with lymph node metastasis (p = 0.002), differentiation (p = 0.020), TNM stage (p = 0.002), and EGFR mutation (p = 0.001). The proliferation, clone formation, and invasive ability were significantly decreased after EPHA5 knockdown in NCI-H460 and H1299 cells. Our data suggest that the EPHA5 receptor plays a role in tumor promotion in lung adenocarcinoma and is a potential target for lung adenocarcinoma treatment.

Casticin and chrysosplenol D from Artemisia annua L. induce apoptosis by inhibiting topoisomerase IIα in human non-small-cell lung cancer cells

BackgroundArtemisia annua L. (A. annua) and its active components exhibit antitumour effects in many cancer cells. However, the biological processes and mechanisms involved are not well understood, especially for the treatment of non-small-cell lung cancer (NSCLC). PurposeThis study aimed to comprehensively explore the biological processes of A. annua and its active components in NSCLC cells and to identify the mechanism by which these compounds induce apoptosis. Study designs/methodsCell viability and flow cytometry assays were used to evaluate the cytotoxicity of A. annua active components casticin (CAS) and chrysosplenol D (CHD) in A. annua in NSCLC cells. After treatment with CAS and CHD, A549 cells were subjected to RNA sequencing (RNA-seq) analysis, differentially expressed genes (DEGs) were screened and subjected to functional enrichment analysis (KEGG and GO analysis) as well as protein interaction network analysis. The key targets associated with apoptosis induction in A549 cells were screened by Cytoscape, and the screened DEGs were validated by qRT-PCR. Immunoblotting, immunofluorescence, and molecular docking assays were used to determine whether CAS and/or CHD could induce apoptosis in NSCLC cells by inducing DNA damage through down-regulation of topoisomerase IIα (topo IIα) expression. The same experiments were verified again in the H1299 lung cancer cell line. ResultsCAS and CHD inhibited NSCLC cells proliferation in a time- and dose-dependent manner, and significantly induced apoptosis. A total of 115 co-upregulated DEGs and 277 co-downregulated DEGs were identified in A549 cells following treatment with CAS and CHD. Comprehensive and systematic data about biological processes and mechanisms were obtained. DNA damage pathways and topo IIα targets were screened to study the apoptosis effects of CAS and CHD on NSCLC cells. CAS and CHD may be able to induce DNA damage by binding to topo IIα-DNA and reducing topo IIα activity. ConclusionThis study suggested that CAS and CHD may reduce topo IIα activity by binding to topo IIα-DNA, affecting the replication of DNA, triggering DNA damage, and inducing apoptosis. It described a novel mechanism associated with topo IIα inhibition to reveal a novel role for CAS and CHD in A. annua as potential anticancer agents and/or adjuvants in NSCLC cells.

Sufentanil inhibits the proliferation and epithelial mesenchymal transition of lung cancer cells through Wnt/beta-catenin signaling pathway

ABSTRACT Lung cancer is the most common malignancy and leading cause of cancer-related death. Sufentanil is a commonly used opioid anesthetic in clinics. This study aimed to explore the effects of sufentanil on the malignant behavior of lung cancer cells. H460 and H1299 lung cancer cell lines were selected for in vitro experiments. The MTT assay was conducted to detect cell viability. Proliferation ability was determined by colony formation and EdU assays. Transwell assays were performed to measure migration and invasion abilities. Western blotting was used to detect the expression of related proteins. LiCl was used to activate the Wnt/β-catenin signaling pathway. Sufentanil decreased the proliferation, migration, and invasion of H460 and H1299 cells. The protein expression levels of vimentin, N-cadherin, β-catenin, c-Myc, and MMP2 were downregulated, while those of E-cadherin and ZO-1 were upregulated after sufentanil treatment. LiCl treatment reversed the effects of sufentanil on H460 and H1299 cells. Sufentanil inhibited the proliferation, migration, invasion, and epithelial–mesenchymal transition of lung cancer cells by regulating the Wnt/β-catenin signaling pathway.

In vitro anticancer, antioxidant and DNA-binding study of the bioactive ingredient of clove and its isolation

Cancer cannot be ignored since it is the most dangerous disease because it is a major cause of death globally with 15% mortality. Researchers have been attracted to the plant-based solution of this havoc. Among all plants, Syzygium aromaticum has shown tremendous results in many aspects such as anticancer, antioxidant, and others. All the studies that took place, were done on the plant extract only. No one goes further than this. Hence, an advanced computational chemistry-based method for the characterization and identification of the bioactive ingredients isolated from cloves was developed for the first time. First, different extracts of Syzygium aromaticum plant buds were obtained using different solvents (Water, methanol, chloroform, ethyl acetate, 50% ethanol, and hexane), then each extract was evaluated for its anticancer activity against A549 and H1299 lung cancer cell lines. The fractionation of the most active extract was done using flash chromatography. After that, anticancer evaluation of every fraction was done again. One of the obtained fractions showed the highest anticancer activity. For the identification of the most active fraction the experimental IR and NMR data of it was taken and compared with the computational IR and NMR data of 19 compounds found in cloves. Furthermore, DNA binding affinity and antioxidant activity of the fraction showing the highest anticancer activity were also studied. The presented method of the isolation of the most bioactive ingredient will be the most helpful for all the scientists working in the field of separation science and phytomedicine.

The Effect of Beta Adrenoreceptor Blockers on Viability and Cell Colony Formation of Non-Small Cell Lung Cancer Cell Lines A549 and H1299

Beta adrenoblockers are a large class of drugs used to treat cardiovascular diseases, migraines, glaucoma and hyperthyroidism. Over the last couple of decades, the anticancer effects of these compounds have been extensively studied. However, the exact mechanism is still not known, and more detailed studies are required. The aim of our study was to evaluate the anticancer activity of beta adrenoblockers in non-small cell lung cancer cell lines A549 and H1299. In order to find the relationship with their selectivity to beta adrenoreceptors, selective (atenolol, betaxolol, esmolol, metoprolol) and non-selective (pindolol, propranolol and timolol) beta blockers were tested. The effect on cell viability was evaluated by MTT assay, and the activity on cell ability to form colonies was tested by clonogenic assay. The type of cell death was evaluated by cell double staining with Hoechst 33342 and Propidium iodide. The most active adrenoblockers against both tested cancer cell lines were propranolol and betaxolol. They completely inhibited lung cancer cell colony formation at 90% of the EC50 (half-maximal effective concentration) value. Most tested compounds induced cell death through apoptosis and necrosis. There was no correlation established between beta adrenoblocker anticancer activity and their selectivity to beta adrenoreceptors.

Variation in Compositions and Biological Activities of Essential Oils from Four Citrus Species: Citrus limon, Citrus sinensis, Citrus paradisi, and Citrus reticulata.

Species of the genus Citrus are cultivated in many regions of China and are widely used for medicinal purposes. In the present study, essential oils (EOs) were extracted from four different Citrus species using steam distillation. The chemical components of these four essential oils were separated using gas chromatography-mass spectrometry, and 52 compounds were confirmed. D-limonene was found to be the most abundant compound. All four essential oils demonstrated varied but remarkable radical scavenging capacity (IC50 ; 0.77-13.9 %). Citrus paradisi essential oil exhibited excellent antioxidant activity. Compared to ibuprofen, topical application of the four Citrus spp. essential oils significantly inhibited ear edema formation in mice. Furthermore, essential oils from the four Citrus species reduced the expression levels of interleukin-6 (IL-6), cyclooxygenase-2 (COX-2) and nuclear transcription factor kappa B p65 (NF-κB) to different degrees. The cytotoxicity of the four essential oils on BV2 microglial cells was determined using the MTT assay (IC50 ; 321.37-1558.87 μg/mL), wherein Citrus limon essential oil showed the lowest cytotoxicity. The essential oils of Citrus limon, Citrus reticulata, and Citrus paradisi had an inhibitory effect on the lung cancer cell lines H1299 by inducing a G0/G1 cell cycle arrest. Cluster and principal component analyses were used to determine the relationship among the Citrus species. These results suggest that the four Citrus essential oils have potential for use as active ingredients in functional foods or cosmeceutical products.

PRKCDBP Methylation is a Potential and Promising Candidate Biomarker for Non-small Cell Lung Cancer.

Background and objectivesThe occurrence and development of lung cancer are closely linked to epigenetic modification. Abnormal DNA methylation in the CpG island region of genes has been found in many cancers. Protein kinase C delta binding protein (PRKCDBP) is a potential tumor suppressor and its epigenetic changes are found in many human malignancies. This study investigated the possibility of PRKCDBP methylation as a potential biomarker for non-small cell lung cancer (NSCLC).MethodsWe measured the methylation levels of PRKCDBP in the three groups of NSCLC tissues. Promoter activity was measured by the dual luciferase assay, with 5'-aza-deoxycytidine to examine the effect of demethylation on the expression level of PRKCDBP.ResultsThe methylation levels of PRKCDBP in tumor tissues and 3 cm para-tumor were higher than those of distant (> 10 cm) non-tumor tissues. Receiver operating characteristic (ROC) curve analysis between tumor tissues and distant non-tumor tissues showed that the area under the line (AUC) was 0.717. Dual luciferase experiment confirmed that the promoter region was able to promote gene expression. Meanwhile, in vitro methylation of the fragment (PRKCDBP_Me) could significantly reduce the promoter activity of the fragment. Demethylation of 5'-aza-deoxycytidine in lung cancer cell lines A549 and H1299 showed a significant up-regulation of PRKCDBP mRNA levels.ConclusionPRKCDBP methylation is a potential and promising candidate biomarker for non-small cell lung cancer.

Endocrine secretory granule production is caused by a lack of REST and intragranular secretory content and accelerated by PROX1

Endocrine secretory granules (ESGs) are morphological characteristics of endocrine/neuroendocrine cells and store peptide hormones/neurotransmitters. ESGs contain prohormones and ESG-related molecules, mainly chromogranin/secretogranin family proteins. However, the precise mechanism of ESG formation has not been elucidated. In this study, we experimentally induced ESGs in the non-neuroendocrine lung cancer cell line H1299. Since repressive element 1 silencing transcription factor (REST) and prospero homeobox 1 (PROX1) are closely associated with the expression of ESG-related molecules, we edited the REST gene and/or transfected PROX1 and then performed molecular biology, immunocytochemistry, and electron and immunoelectron microscopy assays to determine whether ESG-related molecules and ESGs were induced in H1299 cells. Although chromogranin/secretogranin family proteins were induced in H1299 cells by knockout of REST and the induction was accelerated by the PROX1 transgene, the ESGs could not be defined by electron microscopy. However, a small number of ESGs were detected in the H1299 cells lacking REST and expressing pro-opiomelanocortin (POMC) by electron microscopy. Furthermore, many ESGs were produced in the REST-lacking and PROX1- and POMC-expressing H1299 cells. These findings suggest that a lack of REST and the expression of genes related to ESG content are indispensable for ESG production and that PROX1 accelerates ESG production.Trial registration: Not applicable.

ADAMTS8 inhibited lung cancer progression through suppressing VEGFA

BackgroundADAMTS8 expression has been identified to be low in many cancers including lung cancer. However, the specific functions and regulatory system of ADAMTS8 remain to be unveiled. PurposeTo study the potential modulatory mechanism of ADAMTS8 in lung cancer in cell and xenograft mice models. MethodsDifferential expression of ADAMTS8 in lung cancer was analyzed on online tools. So was the overall survival curve in association with ADAMTS8/VEGFA expression in lung cancer patients. RT-qPCR was applied to validate the ADAMTS8 expression in lung cancer cell lines H460 and A549, with the normal lung epithelial cell Beas-2b as a control. Thereafter, overexpressed and knockdown plasmids were constructed for transfection. Colony and flow cytometry methods were used for cell proliferation and apoptosis. RT-qPCR and Western blot methods validated the changes in VEGFA after ADAMTS8 regulation in cells. Tube formation and Transwell methods were applied to observe the changes in tube formation and migration in HUVECs induced by tumor conditioned medium (TCM). Stable-transfected cells were injected subcutaneously into nude mice. H&E and Immunohistochemistry were applied to analyze the pathological differences and protein changes of ADAMTS8, VEGFA and CD31. ResultsHigh ADAMTS8 was correlated with high overall survival rate in lung cancer patients. ADAMTS8 was also abnormally downregulated in NSCLC cells. Upregulation of ADAMTS8 suppressed cell proliferation and enhanced apoptosis while downregulation of ADAMTS8 promoted cell proliferation and decreased apoptosis. VEGFA was negatively correlated with ADAMTS8 in lung cancer tissues. Upregulation of ADAMTS8 inhibited VEGFA in mRNA and protein levels. Further, knockdown of ADAMTS8 induced tube formation and migration of HUVECs and upregulation of ADAMTS8 inhibited this. In addition, upregulation of ADAMTS8 in nude mice inhibited tumor growth and also suppressed VEGFA and CD31 in tumors. ConclusionADAMTS8 inhibited lung cancer progression through suppressing VEGFA in lung cancer.

Antitumor activity of a lectibody targeting cancer-associated high-mannose glycans

Aberrant protein glycosylation is a hallmark of cancer, but few drugs targeting cancer glycobiomarkers are currently available. Here, we showed that a lectibody consisting of the high-mannose glycan-binding lectin Avaren and human immunoglobulin G1 (IgG1) Fc (AvFc) selectively recognizes a range of cell lines derived from lung, breast, colon, and blood cancers at nanomolar concentrations. Binding of AvFc to the non-small cell lung cancer (NSCLC) cell lines A549 and H460 was characterized in detail. Co-immunoprecipitation proteomics analysis revealed that epidermal growth factor receptor (EGFR) and insulin-like growth factor 1 receptor (IGF1R) are among the lectibody's common targets in these cells. AvFc blocked the activation of EGFR and IGF1R by their respective ligands in A549 cells and inhibited the migration of A549 and H460 cells upon stimulation with EGF and IGF1. Furthermore, AvFc induced potent Fc-mediated cytotoxic effects and significantly restricted A549 and H460 tumor growth in severe combined immunodeficiency (SCID) mice. Immunohistochemistry analysis of primary lung tissues from NSCLC patients demonstrated that AvFc preferentially binds to tumors over adjacent non-tumor tissues. Our findings provide evidence that increased abundance of high-mannose glycans in the glycocalyx of cancer cells can be a druggable target, and AvFc may provide a new tool to probe and target this tumor-associated glycobiomarker.

LSD1 inhibitors from the roots of Pueraria lobata

One new 6a,11a-dehydropterocarpan derivative, 6-O-methyl-anhydrotuberosin (1), one new 6a-hydroxypterocarpan, (6aR,11aR,11bR)-hydroxytuberosone (7), and seven known compounds including two 6a,11a-dehydropterocarpans (2 and 4), two coumestans (3 and 5), one isoflavonoid (6) and two other phenolic compounds (8 and 9) were isolated from the roots of Pueraria lobata. The structures of the isolated compounds were elucidated with spectroscopic and spectrometric methods (1 D and 2DNMR, HRESIMS). Compounds 1, 2, 4–5 showed potent LSD1 inhibitory activities with IC50 values ranging from 1.73 to 4.99 μM. Furthermore, compound 2 showed potent cytotoxicity against gastric cancer cell lines MGC-803 and BGC-823, and lung cancer cell lines H1299 and H460.

Investigation of Antimicrobial, DNA Protective and Cytotoxic Activity of Red Cabbage (Brassica Oleracea L. Var. Capitata F. Rubra) Plant

The need to search for natural products that offer a safe alternative to drugs and industrial products that often cause side effects and harm human health is growing over the years. In this study, the water and methanol extracts of the anthocyanin-rich red cabbage (Brassica oleracea L. var. capitata f. rubra) which is available on markets throughout the year, were examined for their antibacterial activity on Escherichia coli, Staphylococcus aereus and Stenotrophomonas maltophilia, and for their antifungal activity on Yarrowia lipolytica, Candida albicans and Candida maltophilia. The DNA protective activity of extracts on the pBR322 plasmid was evaluated by exposing pBR322 to UV and adding hydrogen peroxide. Furthermore, the MTT assay was performed on the H1299 lung cancer cell line and the HUVEC cell line as a negative control to evaluate the cytotoxicity of the extracts. The red cabbage extracts were found to have no antibacterial or antifungal activity on the tested microorganisms. The results of DNA protective activity have shown that at a concentration of 50 mg/mL there is a DNA protective activity of the extracts. No effect of the red cabbage extracts on the H1299 cell line as shown by MTT assay results. On the other hand, the decrease in the viability of HUVEC cells started at a concentration of 25 μg/mL and above, reaching 70% at 100 μg/ mL for the water extract and 74 % at 100 μg/ mL for the methanol extract. Further studies to investigate the active components of the DNA protective activity in the red cabbage extracts are required. Further studies are also needed to explore the possibility of incorporating these components in cosmetic products such as sunscreens without causing skin damage.

Investigation of Antimicrobial, DNA Protective and Cytotoxic Activity of Red Cabbage (Brassica Oleracea L. Var. Capitata F. Rubra) Plant

The need to search for natural products that offer a safe alternative to drugs and industrial products that often cause side effects and harm human health is growing over the years. In this study, the water and methanol extracts of the anthocyanin-rich red cabbage (Brassica oleracea L. var. capitata f. rubra) which is available on markets throughout the year, were examined for their antibacterial activity on Escherichia coli, Staphylococcus aereus and Stenotrophomonas maltophilia, and for their antifungal activity on Yarrowia lipolytica, Candida albicans and Candida maltophilia. The DNA protective activity of extracts on the pBR322 plasmid was evaluated by exposing pBR322 to UV and adding hydrogen peroxide. Furthermore, the MTT assay was performed on the H1299 lung cancer cell line and the HUVEC cell line as a negative control to evaluate the cytotoxicity of the extracts. The red cabbage extracts were found to have no antibacterial or antifungal activity on the tested microorganisms. The results of DNA protective activity have shown that at a concentration of 50 mg/mL there is a DNA protective activity of the extracts. No effect of the red cabbage extracts on the H1299 cell line as shown by MTT assay results. On the other hand, the decrease in the viability of HUVEC cells started at a concentration of 25 μg/mL and above, reaching 70% at 100 μg/ mL for the water extract and 74 % at 100 μg/ mL for the methanol extract. Further studies to investigate the active components of the DNA protective activity in the red cabbage extracts are required. Further studies are also needed to explore the possibility of incorporating these components in cosmetic products such as sunscreens without causing skin damage.

A small molecule targeting CHI3L1 inhibits lung metastasis by blocking IL-13Rα2-mediated JNK-AP-1 signals.

Our previous big data analyses showed a high level of association between chitinase 3 like1 (CHI3L1) expression and lung tumor development. In the present study, we investigated whether a CHI3L1‐inhibiting chemical, 2‐({3‐[2‐(1‐cyclohexen‐1‐yl)ethyl]‐6,7‐dimethoxy‐4‐oxo‐3,4‐dihydro‐2‐quinazolinyl}sulfanyl)‐N‐(4‐ethylphenyl)butanamide (K284), could inhibit lung metastasis and studied its mechanism of action. We investigated the antitumor effect of K284 both in vitro and in vivo. K284 (0.5 mg·kg−1 body weight) significantly inhibited lung metastasis in in vivo models after injection of murine melanoma cells (B16F10) or adenocarcinomic human alveolar basal epithelial cells (A549). K284 significantly and concentration‐dependently also inhibited cancer cell proliferation and migration in the A549 and H460 lung cancer cell lines. We found that the binding of K284 to the chitin‐binding domain (CBD) of CHI3L1 prevented the binding of CHI3L1 to its receptor, interleukin‐13 receptor subunit alpha‐2 (IL‐13Rα2), thereby suppressing the CHI3L1 signal. This blocking of the CHI3L1‐IL‐13Rα2 signal caused the inhibition of c‐Jun N‐terminal kinase (JNK)‐activator protein 1 (AP‐1) signals, resulting in the prevention of lung metastasis and cancer cell growth. Our data demonstrate that K284 may serve as a potential candidate anticancer compound targeting CHI3L1.