Mechanism Of Apoptosis Induction Research Articles

Nebivolol, an antihypertensive agent, has new application in inhibiting melanoma.

Repurposing existing drugs for cancer therapy has become an important strategy because of its advantages, such as cost reduction, effect and safety. The present study was designed to investigate the antimelanoma effect and possible mechanisms of action of nebivolol, which is an approved and widely prescribed antihypertensive agent. In this study, we explored the effect of nebivolol on cell proliferation and cell activity in melanoma in vitro and the potential antimelanoma mechanism of nebivolol through a series of experiments, including the analysis of the effects with regard to cell apoptosis and metastasis. Furthermore, we evaluated the antimelanoma effect on xenograft tumor models and inspected the antimelanoma mechanism of nebivolol in vivo using immunohistochemical and immunofluorescence staining assays. As results in this work, in vitro , nebivolol possessed a strong activity for suppression proliferation and cell cycle arrest on melanoma. Moreover, nebivolol significantly induced cell apoptosis in melanoma through a mitochondrial-mediated endogenous apoptosis pathway. Additionally, nebivolol inhibited melanoma cell metastasis. More importantly, nebivolol exhibited significantly effective melanoma xenograft models in vivo , which related to the mechanism of apoptosis induction, proliferation inhibition, metastasis blocking and angiogenesis arrest. Overall, the data of the present study recommend that nebivolol holds great potential in application as a novel agent for the treatment of melanoma.

Abstract 2102: Understanding the mechanism of action of rigosertib in recessive dystrophic epidermolysis bullosa-associated squamous cell carcinoma through single cellanalysis

Abstract The allosteric inhibitor rigosertib (an experimental therapeutic) was previously identified as an effective inducer of G2M arrest and apoptosis in squamous cell carcinoma (SCC) keratinocytes isolated from patients with the rare genetic disease recessive dystrophic epidermolysis bullosa (RDEB), both in vitro and in vivo. Early clinical trial data treating RDEB-associated SCC with rigosertib has shown remarkable efficacy in a sub-set of patients and understanding the mechanism of action of rigosertib in this lethal cancer would provide rationale for patient stratification. Multiple mechanisms of action were previously attributed to rigosertib, including polo-like kinase-1 (PLK1), PI3-kinase, activated RAS and microtubule polymerization inhibition. Our published data comparing PLK1 siRNA with PI3-Kinase inhibitors, MAPK inhibitors and microtubule disruption favored PLK1 as a mechanism of action without definitive conclusion. We therefore sought to determine the mechanism of action using transcriptomic profiling. Flow cytometry experiments identified three main categories of time to increase G2M in nine separate RDEB SCC keratinocyte populations in culture ranging from 1 to 6 hours after exposure to 1uM rigosertib. Bulk-RNA sequencing of representative populations from time to G2M groupings failed to identify consistent transcriptional change after exposure to rigosertib. However, single cell sequencing was able to identify increases in sub-populations of cells associated with G2M as well as transient emergence of unique populations that were specific to individual patient populations after exposure to rigosertib in culture. Of note a transient population of RDEB SCC keratinocytes only present after 1 hour of exposure to rigosertib was characterized by increased expression of the potassium regulator KCNRG which has previously been associated with apoptosis in chronic lymphocytic leukemia. Ongoing work is determining the role this gene may play in the mechanism of apoptosis induction in RDEB SCC keratinocytes after exposure to rigosertib. Citation Format: Andrew P. South, Suhao Han, Paolo Fortina. Understanding the mechanism of action of rigosertib in recessive dystrophic epidermolysis bullosa-associated squamous cell carcinoma through single cellanalysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2102.

Molecular mechanism analysis of apoptosis induced by silk fibroin peptides

Silk fibroin derived from silkworm cocoons exhibits excellent mechanical properties, good biocompatibility, and low immunogenicity. Previous studies showed that silk fibroin had an inhibitory effect on cells, suppressing proliferation and inducing apoptosis. However, the source of the toxicity and the mechanism of apoptosis induction are still unclear. In this study, we hypothesized that the toxicity of silk fibroin might originate from the crystalline region of the heavy chain of silk fibroin. We then verified the hypothesis and the specific induction mechanism. A target peptide segment was obtained from α-chymotrypsin. The potentially toxic mixture of silk fibroin peptides (SFPs) was separated by ion exchange, and the toxicity was tested by an MTT assay. The results showed that SFPs obtained after 4 h of enzymatic hydrolysis had significant cytotoxicity, and SFPs with isoelectric points of 4.0–6.8 (SFPα II) had a significant inhibitory effect on cell growth. LC-MS/MS analysis showed that SFPα II contained a large number of glycine-rich and alanine-rich repetitive sequence polypeptides from the heavy-chain crystallization region. A series of experiments showed that SFPα II mediated cell death through the apoptotic pathway by decreasing the expression of Bcl-2 protein and increasing the expression of Bax protein. SFPα II mainly affected the p53 pathway and the AMPK signaling pathway in HepG2 cells. SFPα II may indirectly increase the expression of Cers2 by inhibiting the phosphorylation of EGFR, which activated apoptotic signaling in the cellular mitochondrial pathway and inhibited the Akt/NF-κB pathway by increasing the expression of PPP2R2A.

Uncaria-derived alkaloids: A review on the mechanism in inducing apoptosis in oral squamous cell carcinoma

Alkaloids, the major secondary metabolite compounds identified in the genus Uncaria, exhibit the ability to induce apoptosis in oral squa-mous cell carcinoma (OSCC). Various potential alkaloids in herbal medicine have been explored and provided promising results, one of which is the potential to induce apoptosis. This study aims to provide a more comprehensive review of the mechanism of alkaloids derived from Uncaria in inducing apoptosis in OSCC. In the systematic search conducted, the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines were adhered to conduct this review through the databases, including PubMed, ScienceDirect, Wiley and a manual search using Google Scholar. According to the findings, this review states that several types of alkaloids derived from Uncaria that have been reviewed, such as hirsutine, rhynchophylline, isorhynchophylline, and hirsuteine, have different mechanisms in in-ducing apoptosis in OSCC, both through intrinsic and extrinsic pathways. The evidence presented in the present study provides an oppor-tunity for future research to determine the appropriate therapeutic dose based on the specific alkaloid, duration of therapeutic administration, and in vivo and/or in human testing.

The safety evaluation of Shenze Shugan capsule and mechanism of apoptosis induced by five potentially nephrotoxic components

Ethnopharmacological relevanceShenze Shugan capsule is a prescription of traditional Chinese medicine for nonalcoholic steatohepatitis treatment. It includes Rhei Radix et Rhizoma (RR), Cassiae Semen (CS) and Alismatis Rhizoma(AR), which widely contains rhein, emodin, aurantio-obtusin, alisol A and alisol B 23-monoacetate. Aim of the studyIn this study, we aimed to explore the safety of the medicine, and further elucidate the mechanism of apoptosis induction in HK-2 cells by five components, including rhein, emodin, aurantio-obtusin, alisol A and alisol B 23-monoacetate. Materials and methodsWe investigated the nephrotoxicity of Shenze Shugan capsule, including RR, CS, AR and mixed herbs given for two months in rats. Superoxide dismutase (SOD) in kidney tissues, urea nitrogen (BUN) and creatinine (CRE) in serum were detected, and renal pathology analysis was performed. In cell experiments, the apoptotic rate and cell cycle distribution of HK-2 cells were tested by flow cytometry. The levels of mitochondrial membrane potential (ΔΨm) and related protein expression in mitochondrial pathway were measured as well. ResultsWe confirmed that two months of administering high doses(60 times the dose for clinical use in adults) of RR, CS or mixed herbs upregulated the levels of CRE and RUN, inhibited SOD activity, and increased the degree of tubular degeneration and glomerular dilatation, but Shenze Shugan capsule has no significant differences in renal structure or renal function. In addition, we found that five components all concentration-dependently inhibited HK-2 cells proliferation and induced apoptosis, especially aurantio-obtusin as the novel nephrotoxic component. Rhein and emodin significantly induced S/M accumulation, but aurantio-obtusin, alisol A and alisol B 23-monoacetate significantly induced G1/M accumulation in HK-2 cells. Similarly, they could induce Caspase3 activation, loss of mitochondrial membrane potential (ΔΨm), and down-regulation of Bcl-2 and up-regulation of Bax. ConclusionsThrough a two-month subchronic toxicity study in rats, our preliminary determination is that this formulation is safe and reliable for long-term use. Interestingly, the potentially toxic herbs such as RR, CS, AR can reduce toxicity by drug compatibility. When further exploring the mechanism of action of toxic herbs, we found that mitochondrial pathway is involved in the apoptosis of HK -2 cells induced by rhein, emodin, aurantio-obtusin, alisol A and alisol B 23-monoacetate. Our findings provide new ideas for safety studies of Shenze Shugan capsule.

Photodynamic Therapy and Immunological View in Gastrointestinal Tumors.

Gastrointestinal cancers are a specific group of oncological diseases in which the location and nature of growth are of key importance for clinical symptoms and prognosis. At the same time, as research shows, they pose a serious threat to a patient's life, especially at an advanced stage of development. The type of therapy used depends on the anatomical location of the cancer, its type, and the degree of progression. One of the modern forms of therapy used to treat gastrointestinal cancers is PDT, which has been approved for the treatment of esophageal cancer in the United States. Despite the increasingly rapid clinical use of this treatment method, the exact immunological mechanisms it induces in cancer cells has not yet been fully elucidated. This article presents a review of the current understanding of the mode of action of photodynamic therapy on cells of various gastrointestinal cancers with an emphasis on colorectal cancer. The types of cell death induced by PDT include apoptosis, necrosis, and pyroptosis. Anticancer effects are also a result of the destruction of tumor vasculature and activation of the immune system. Many reports exist that concern the mechanism of apoptosis induction, of which the mitochondrial pathway is most often emphasized. Photodynamic therapy may also have a beneficial effect on such aspects of cancer as the ability to develop metastases or contribute to reducing resistance to known pharmacological agents.

In silico and in vitro study on apoptosis in MCF-7 human breast cancer cells lines via caspase-3 activation of Markhacanasin A

Markhacanasin A (MA), a cycloartane-type triterpenoid was isolated from the leaves of Markhamia stipulata var. canaense V.S. Dang, collected in Ninh Thuan province, Vietnam. In our previous study, it was found that MA possessed potential cytotoxicity against MCF-7 human breast cancer cell lines, with an IC50=16.51±0.22 μM. However, the mechanism by which MA triggers apoptosis in the MCF-7 cells has not been explored yet. Therefore, this study has been conducted to investigate the mechanism of apoptosis induction in the MCF-7 cell lines by MA. The study utilised the flow cytometry method and molecular docking patterns analysis of caspase-3, caspase-8, caspase-9, Apaf-1, and survivin. Interestingly, the results have demonstrated that MA induces apoptosis in the MCF-7 cell line by activating the caspase-3 enzyme, both in vitro and in silico, which suggests that MA may have therapeutic potential as a breast cancer treatment agent. This study provides new insights into the mechanism of action of MA and highlights its potential as a promising therapeutic option for breast cancer.

Isolation and Anticancer Progression Evaluation of the Chemical Constituents from Bridelia balansae Tutcher.

The dichloromethane extract of the roots of Bridelia balansae Tutcher (Phyllanthaceae) was found to show potential anticancer activity against HCT116 colorectal cancer cell. Our bioassay-guided phytochemical investigation of the roots of B. balansae led to the identification of 14 compounds including seven lignans (1-7), three phenylbenzene derivatives (8-10), two flavanone (11-12), and two triterpenoids (13-14). Among them, 4'-demethyl-4-deoxypodophyllotoxin (1) is the first aryltetralin lignan compound identified from this plant species. In addition, the stereochemistry of 1 was validated by X-ray crystallography for the first time, and its distinguished cytotoxic effect on HCT116 cells with an IC50 value at 20 nM was induced via an apoptosis induction mechanism. Compound 1 could also significantly decrease the migration rate of HCT116 cells, indicating its potential application against cancer metastasis. The western blot analysis showed that 1 has the potential to inhibit cell proliferation and metastasis. Treatment of 1 resulted in the downregulation of matrix metalloproteinases 2 (MMP2) and p-Akt, while p21 was upregulated. Collectively, the present study on the phytochemical and biological profile of B. balansae has determined the plant as a useful source to produce promising anticancer lead compounds.

Antiproliferative activity and apoptosis-inducing mechanism of Amaryllidaceae alkaloid montanine on A549 and MOLT-4 human cancer cells

The isoquinoline alkaloids found in Amaryllidaceae are attracting attention due to attributes that can be harnessed for the development of new drugs. The possible molecular mechanisms by which montanine exerts its inhibitory effects against cancer cells have not been documented. In the present study, montanine, manthine and a series of 15 semisynthetic montanine analogues originating from the parent alkaloid montanine were screened at a single test dose of 10 μM to explore their cytotoxic activities against a panel of eight cancer cell lines and one non-cancer cell line. Among montanine and its analogues, montanine and its derivatives 12 and 14 showed the highest cytostatic activity in the initial single-dose screening. However, the native montanine exhibited the greatest antiproliferative activity against cancer cells, with a lower mean IC50 value of 1.39 µM, compared to the displayed mean IC50 values of 2.08 µM for 12 and 3.57 µM for 14. Montanine exhibited the most potent antiproliferative activity with IC50 values of 1.04 µM and 1.09 µM against Jurkat and A549 cell lines, respectively. We also evaluated montanine’s cytotoxicity and cell death mechanisms. Our results revealed that montanine triggered apoptosis of MOLT-4 cells via caspase activation, mitochondrial depolarisation and Annexin V/PI double staining. The Western blot results of MOLT-4 cells showed that the protein levels of phosphorylated Chk1 Ser345 were upregulated with increased montanine concentrations. Our findings provide new insights into the mechanisms underlying the cytostatic, cytotoxic and pro-apoptotic activities of montanine alkaloids in lung adenocarcinoma A549 and leukemic MOLT-4 cancer cell types.

P53 status, and G2/M cell cycle arrest, are determining factors in cell-death induction mediated by ELF-EMF in glioblastoma

The average survival of patients with glioblastoma is 12–15 months. Therefore, finding a new treatment method is important, especially in cases that show resistance to treatment. Extremely low-frequency electromagnetic fields (ELF-EMF) have characteristics and capabilities that can be proposed as a new cancer treatment method with low side effects. This research examines the antitumor effect of ELF-EMF on U87 and U251 glioblastoma cell lines. Flowcytometry determined the viability/apoptosis and distribution of cells in different phases of the cell cycle. The size of cells was assessed by TEM. Important cell cycle regulation genes mRNA expression levels were investigated by real-time PCR. ELF-EMF induced apoptosis in U87cells much more than U251 (15% against 2.43%) and increased G2/M cell population in U87 (2.56%, p value < 0.05), and S phase in U251 (2.4%) (data are normalized to their sham exposure). The size of U87 cells increased significantly after ELF-EMF exposure (overexpressing P53 in U251 cells increased the apoptosis induction by ELF-EMF). The expression level of P53, P21, and MDM2 increased and CCNB1 decreased in U87. Among the studied genes, MCM6 expression decreased in U251. Increasing expression of P53, P21 and decreasing CCNB1, induction of cell G2/M cycle arrest, and consequently increase in the cell size can be suggested as one of the main mechanisms of apoptosis induction by ELF-EMF; furthermore, our results demonstrate the possible footprint of P53 in the apoptosis induction by ELF-EMF, as U87 carry the wild type of P53 and U251 has the mutated form of this gene.

Isolation, identification, and characterization of α- asarone, from hydromethanolic leaf extract of Acorus calamus L. and its apoptosis-inducing mechanism in A549 cells

Due to the presence of several active secondary metabolites, the traditional Indian and Chinese medicinal herb Acorus calamus L. has been utilized for both medical and culinary purposes since ancient times. A recent report has underscored the promising cytotoxic effect of A. calamus leaves extract against non-small cell lung cancer A549 cells. Thus, we want to separate the bioactive substance from the hydromethanolic extract of A. calamus leaves in the current investigation. Thin-layer chromatography was used to separate the compounds and different spectroscopic methods (UV, FTIR, NMR, and LCMS/MS) were used for the structure prediction. α-asarone was found to be the main bioactive compound present and it was isolated from A. calamus leaves extract. It exerted a good cytotoxic effect with an IC50 value of 21.43 ± 1.27 μM against A549 cells and IC50 value of 324.12 ± 1.32 μM against WI-38 cells. The induction of apoptosis in A549 cells by α-asarone was reaffirmed by the diverse differential staining methods including DAPI, Acridine Orange/Ethidium Bromide, and Giemsa staining. Additionally, α-asarone induced mitochondrial membrane potential (ΔΨm) dissipation with a concomitant increase in the production of ROS. Furthermore, it also increased expressions of caspase-3, caspase-9, caspase-8, DR4, and DR5 genes in A549 cells. In conclusion, α-asarone-induced apoptotic cell death in non-small lung cancer cells (A549) as a result of loss of mitochondrial function, increased ROS production, subsequent activation of an internal and extrinsic caspase pathway, and altered expression of genes controlling apoptosis. As a whole, α-asarone is a plausible therapeutic agent for managing lung cancer. HIGHLIGHTS Isolation of bioactive compound from hydromethanolic leaves extract of Acorus calamus L. by thin layer chromatography. Structural elucidation of the bioactive compound was carried out using different methods like UV analysis, FTIR, NMR, and LC-MS/MS analysis. A plausible mode of action revealed that α-asarone can induce apoptosis in lung cancer cells (A549). Communicated by Ramaswamy H. Sarma

Antiproliferative activity and apoptosis-inducing mechanism of Curcuma longa (Turmimax®) on HeLa cell lines.

Curcumin, the primary polyphenol found in turmeric, is derived from the Curcuma longa plant. Since curcumin is nontoxic and has a wide range of medicinal qualities, including anti-oxidant, analgesic, anti-inflammatory, and antibacterial action, it has been widely employed in Ayurveda medicine for ages. Curcumin has recently been discovered to have anti-cancer properties through its impact on numerous biological pathways involved in carcinogenesis, metastasis, tumorigenesis, cell cycle regulation, mutagenesis, and oncogene expression. In this study, we determined the Antiproliferative activity and apoptosis-inducing mechanism of C. longa (Turmimax®) on human cancer cells. The cytotoxic effect was evaluated against HeLa cell lines using the MTT (3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) assay. Flow cytometric analysis was performed to detect apoptotic cell death. Turmimax® exhibits promising properties as a potential anti-cancer therapeutic agent in human cervical adenocarcinomas and possibly other cancer types, with an IC50 value of 87.89 µg/mL. In HeLa cells treated with Turmimax®, cell cycle arrest was seen in the G0/G1 and S phases. By inducing apoptosis and increasing the number of apoptotic cells in a dose-dependent manner, the experimental data suggest that Turmimax® has considerable promise in cancer prevention and treatment.

Gold nanoparticles loaded TNF-α and CALNN peptide as a drug delivery system and promising therapeutic agent for breast cancer cells

ABSTRACT We investigated the anti-cancer properties of gold nanoparticles loaded TNF- and CALNN peptides, which we proposed as a potential drug delivery system using in vitro and in vivo models. The binding of GNPs-TNF- and GNPs-TNF-CALNN was characterized using a UV, ELISA and SEM analysis. The outcomes demonstrated that a novel drug delivery system had an anti-proliferative activity against breast cancer cell lines through a mechanism of apoptosis induction. In vivo model involved studying the cytotoxic influence of a drug delivery system GNPs, GNPs-TNF-α and GNPs-TNF-α-CALNN when applied to the transplanted AN-3 cell line. tumor sections were examined using microarray. In-vivo studies demonstrated that GNPs alone had less of a growth inhibitory effect on tumors implanted in mice when compared to GNPs-TNF – CALNN combined therapy. The cytotoxic assay showed that GNPs, GNPs-TNF-α and GNPs-TNF-α-CALNN exhibit selective toxicity towards cancer cells, inducing cell apoptosis through activation of caspase-3 and 7, p53 protein.

Apoptosis Detection Assays.

Many apoptosis assays are available since there are many proteins regulated at multiple points and involved in apoptosis signaling cascade. To detect apoptosis accurately, two or more assays should be used since there are many overlapped features between apoptosis and necrosis. There are six major groups of available assays to detect apoptosis: membrane alteration, mitochondrial assays, cytomorphological alterations, DNA fragmentation, detection of caspase, cleaved substrate, inhibitors and regulators, and detection of apoptosis in whole mounts. Among those assay, early apoptosis could be detected through annexin V, which is based on the loss of the cellular membrane integrity. Also, there are many assays that can detect midphase of apoptosis using caspase activation and molecular processing including PARP degradation. Late phase of apoptosis could be detected with DNA fragmentation assays. Combinations of these assays allow us to identify the mechanisms of apoptosis induction after specific stimulus. This chapter will introduce three apoptosis detection assays including annexin assay, DNA/chromatin condensation assays, and TUNEL assay.

The role of some lipids and their metabolites in programmed cell death (lipoapoptosis).

In recent years, the understanding of the mechanisms involved in the regulation of lipoapoptosis signaling pathways has expanded considerably. However, many mechanisms of apoptosis induction by lipids as well as molecules mediating intracellular and systemic signals belonging to AOS/enzyme-dependent phospholipid metabolites are not completely clear.This review summarizes the current understanding of the mechanisms of apoptotic cell death induction by some lipid molecules. Literature search was performed in the database “PubMed”, “eLIBRARY” using key words: “apoptosis”, “lipids”, “fatty acids”, “eicosanoids”, “reactive oxygen species”.A brief characterization of the signaling pathways of apoptosis is given. The role of reactive oxygen species and their dependent products of lipid peroxidation in the regulation of the main signaling pathways of apoptosis are shown. Particular attention is paid to the product of phospholipid metabolism – 4-hydroxynonenal.Pro- and anti-apoptotic effects of some prostaglandins are demonstrated. Arguments are presented that prostaglandins of series J and D are pro-apoptotic in most cells, and this effect depends on activation of the prostanoid receptor DP2 and on reduction of AKT kinase activity. In contrast, the E-series prostaglandins and hydroxyecosatetraenoic acid act opposite to the J-series and D-series prostaglandins, reducing apoptosis by activating AKT and increasing Bcl-2 protein expression.The role of individual fatty acids involved in the initiation and transduction of pro-apoptotic and anti-apoptotic signals is assessed. It was shown that saturated fatty acids have the maximum damaging potential than their unsaturated counterparts. An in-depth understanding and deciphering of the mechanisms by which lipids and their metabolites modulate the activation of signaling pathways of programmed cell death can help to develop therapeutic strategies to prevent a number of diseases associated with impaired regulation of apoptosis.

Eltanexor Effectively Reduces Viability of Glioblastoma and Glioblastoma Stem-Like Cells at Nano-Molar Concentrations and Sensitizes to Radiotherapy and Temozolomide.

Current standard adjuvant therapy of glioblastoma multiforme (GBM) using temozolomide (TMZ) frequently fails due to therapy resistance. Thus, novel therapeutic approaches are highly demanded. We tested the therapeutic efficacy of the second-generation XPO1 inhibitor Eltanexor using assays for cell viability and apoptosis in GBM cell lines and GBM stem-like cells. For most GBM-derived cells, IC50 concentrations for Eltanexor were below 100 nM. In correlation with reduced cell viability, apoptosis rates were significantly increased. GBM stem-like cells presented a combinatorial effect of Eltanexor with TMZ on cell viability. Furthermore, pretreatment of GBM cell lines with Eltanexor significantly enhanced radiosensitivity in vitro. To explore the mechanism of apoptosis induction by Eltanexor, TP53-dependent genes were analyzed at the mRNA and protein level. Eltanexor caused induction of TP53-related genes, TP53i3, PUMA, CDKN1A, and PML on both mRNA and protein level. Immunofluorescence of GBM cell lines treated with Eltanexor revealed a strong accumulation of CDKN1A, and, to a lesser extent, of p53 and Tp53i3 in cell nuclei as a plausible mechanism for Eltanexor-induced apoptosis. From these data, we conclude that monotherapy with Eltanexor effectively induces apoptosis in GBM cells and can be combined with current adjuvant therapies to provide a more effective therapy of GBM.

Cellular Regulation of Kynurenic Acid-Induced Cell Apoptosis Pathways in AGS Cells.

Kynurenic acid was included in the three compounds (caffeic acid, chlorogenic acid, and kynurenic acid) that showed high antioxidant and anti-inflammatory potential among the phenolic compounds contained in Gynura procumbens. In this study, the mechanism of cancer cell death induced by kynurenic acid (KYNA), which has the highest molecular binding affinity, in the gastric cancer cell line AGS was confirmed in molecular docking analysis. KYNA showed the most cancer cell death effect on AGS cells among several gastric cancer cell lines (MKN, AGS, and SNU). AGS cells were used for later experiments, and KYNA concentrations of 0, 150, 200, and 250 µM were used. KYNA inhibited cell migration and proliferation in AGS cells in a concentration-dependent manner. G2/M phase cell cycle arrest and reduction of related proteins (Cdc25C, CDK1 and CyclinB1) were confirmed in KYNA-treated AGS cells. Apoptosis of KYNA-treated AGS cells was confirmed by Annexin V/propidium iodide (PI) staining flow cytometry analysis. As a result of morphological chromatin condensation through DAPI (4′,6-diamidino-2-phenylindole), intense blue fluorescence was confirmed. The mechanism of apoptosis induction of KYNA-treated AGS cells was confirmed by western blotting. In the extrinsic pathway, apoptosis induction markers FasL, Fas, and Caspase-3 and -8 were increased in a concentration-dependent manner upon KYNA treatment. In the intrinsic pathway, the expression of anti-apoptotic factors PI3K, AKT, and Bcl-xL was down-regulated, and the expression of apoptosis-inducing factors BAD, Bak, Bax, Cytochrom C, and Caspase-9 was up-regulated. Therefore, in the present study, we strongly imply that KYNA induces apoptosis in AGS gastric cancer cells. This suggests that KYNA, a natural compound, could be the basis for drug for the treatment of gastric cancer.

Bioinformatics Analysis Reveals FOXM1/BUB1B Signaling Pathway as a Key Target of Neosetophomone B in Human Leukemic Cells: A Gene Network-Based Microarray Analysis.

Abnormal expression of Forkhead box protein M1 (FOXM1) and serine/threonine kinase Budding uninhibited by benzimidazoles 1 (BUB1B) contributes to the development and progression of several cancers, including chronic myelogenous leukemia (CML). However, the molecular mechanism of the FOXM1/BUB1B regulatory network and the role of Neosetophomone-B (NSP-B) in leukemia remains unclear. NSP-B, a meroterpenoid fungal secondary metabolite, possesses anticancer potential in human leukemic cells lines; however, the underlying mechanism has not been elucidated. The present study aimed to explore the role of NSP-B on FOXM1/BUB1B signaling and the underlying molecular mechanism of apoptosis induction in leukemic cells. We performed gene expression profiling of NSP-B-treated and untreated leukemic cells to search for differentially expressed genes (DEGs). Interestingly BUB1B was found to be significantly downregulated (logFC -2.60, adjusted p = 0.001) in the treated cell line with the highest connectivity score among cancer genes. Analysis of TCGA data revealed overexpression of BUB1B compared to normal in most cancers and overexpression was associated with poor prognosis. BUB1B also showed a highly significant positive correlation with FOXM1 in all the TCGA cancer types. We used human leukemic cell lines (K562 and U937) as an in vitro study model to validate our findings. We found that NSP-B treatment of leukemic cells suppressed the expression of FOXM1 and BUB1B in a dose-dependent manner. In addition, NSP-B also resulted in the downregulation of FOXM1-regulated genes such as Aurora kinase A, Aurora kinase B, CDK4, and CDK6. Suppression of FOXM1 either by siRNA or NSP-B reduced BUB1B expression and enhanced cell survival inhibition and induction of apoptosis. Interestingly combination treatment of thiostrepton and NSP-B suppressed of cell viability and inducted apoptosis in leukemic cells via enhancing the activation of caspase-3 and caspase-8 compared with single-agent treatment. These results demonstrate the important role of the FOXM1/BUB1B pathway in leukemia and thus a potential therapeutic target.

Green synthesized zinc oxide nanoparticles induce apoptosis by suppressing PI3K/Akt/mTOR signaling pathway in osteosarcoma MG63 cells

This study aimed to assess the apoptosis-inducing mechanism of zinc oxide nanoparticles (ZnO NPs) stabilized by Solanum xanthocarpum plant extract in human osteosarcoma MG63 cells. In the present study, we synthesized ZnO NPs from S. xanthocarpum extract and evaluated its anticancer mechanism on MG 63 cells. The synthesized ZnO NPs were characterized by ultraviolet spectroscopy, X-ray crystallography, transmission electron microscopy, energy dispersive X-ray, and Fourier-transform infrared spectroscopy analysis. The mean size of the synthesized ZnO NPs was 21.62 ± 7.45 nm and spherical in shape. The cytotoxicity of ZnO NPs on MG63 cells was determined by MTT assay. The Western blot analysis was carried out to examine the expression of apoptotic and autophagy-related proteins in MG63 cells. The findings of the study reveal that ZnO NPs treatment showed concentration-dependent cytotoxicity, increased lipid peroxidation, decreased antioxidant activity, increased reactive oxygen species generation, and increased DNA damage. In addition, ZnO NPs treatment increased the expression of apoptotic members such as p53, Bax, caspase-3, -8, and -9 while downregulating Bcl-2 expression in MG63 cells. Furthermore, ZnO NPs treatment suppressed the P13K/AKT/mTOR signaling pathway and increased the expression of LC3 and beclin-1 in MG63 cells. The present study demonstrated that ZnO NPs induced apoptosis and autophagy in MG63 cells through modifying apoptotic and autophagy-related proteins.

Design, synthesis, evaluation of new 3-acetylisoxazolines and their hybrid analogous as anticancer agents: In vitro and in silico analysis

3-Acetylisoxazolines were synthesized by the reaction of natural (R)-limonene and (R)-carvone with acetone in the presence of iron (III) nitrate. The reaction showed to be highly peri- and regioselective. Next, using a 1,3-dipolar cycloaddition reaction, the mono-3-acylisoxazolines derived from these monoterpenes were evaluated for their reactivity with nitrilimines. Only the enone of carvone-isoxazoline was regioselectively reactive, providing a new fused isoxazoline-carvone-pyrazolines. The structure of all the newly synthesized mono-cycloadducts (3 &5) and bis-cycloadducts (4 &7a–c) were fully identified based on their HRMS and NMR spectral data. They have also been screened for their cytotoxic activity against four human cancer cell lines: fibrosarcoma (HT-1080), lung carcinoma (A-549), and breast (MCF-7 and MDA-MB-231) cell lines. The obtained results showed that compound 4 was a potent cytotoxic agent against all selected cells. The possible mechanism of apoptosis induction by compound 4 was investigated using Annexin-V binding assay, caspase-3/7 activity and analysis cell cycle progression. The compound 4 induced the early apoptosis of both MCF-7 and MDA-MB-231 through caspase-3/7 activation, and the compound 4 have elicited S and G2/M phase arrest in MCF-7and MDA-MB-231 cancer cells, respectively. For further target investigations, a molecular docking study was employed and it showed that compound 4 has an inhibitory activity against Pim-1 protein kinase. Molecular dynamics study showed that compound 4/Pim-1 complex was stable during the simulation run at different time intervals. In-Silico ADMET predicted that compound 4 has good pharmacokinetic properties with high estimated oral bioavailability.