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BACKGROUND/AIM Cancer metastasis is the main cause of mortality in cancer patients. As lung cancer patients are mostly detected at metastatic stages, strategies that inhibit cancer metastasis may offer effective therapies. Activation of FAK and Akt/mTOR pathways promotes the highly metastatic phenotypes of epithelial to mesenchymal transition (EMT). We unraveled EMT inhibitory action of pongamol and the mechanism controlling cell dissemination in lung cancer cells. MATERIALS AND METHODS Cytotoxic and antiproliferative effects of pongamol were determined by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. Apoptosis and necrosis induction in response to pongamol treatment was observed and visualized by nuclei staining assay. Wound healing migration, invasion, and anchorage-dependent growth assay were conducted to evaluate metastatic behaviors. EMT protein expression and FAK pathway were detected by western blot analysis. RESULTS Pongamol at 0-100 μM exhibited significant inhibition on migration, and invasion of cancer cells. Regarding anoikis resistance potential, the compound significantly inhibited survival and growth of cancer cells in an anchorage-independent manner, as indicated by the depletion of growing colonies in pongamol-pretreated cells. Protein level analysis further showed that pongamol exerted its anti-metastasis effect by inhibiting EMT, as indicated by a decrease of several mesenchymal proteins (N-cadherin, vimentin, Snail, and Slug). Regarding the up-stream mechanisms, we found that pongamol inhibited activation of FAK and Akt/mTOR signaling pathways. CONCLUSION Pongamol exhibits potent anti-metastatic activity through suppressing key potentiating factors of cancer metastasis EMT and FAK.

The most prevalent lung cancer is non-small cell lung cancer (NSCLC). This lung cancer type often develops other organ-specific metastases that are critical burdens in the treatment process. Orchid species in the genus

Bioactive natural compounds derived from plants are the source for the development of new drugs. Numerous in vitro studies have explored the anti-inflammatory effect of eurycomanone and fargesin, derived from Eurycoma longifolia and Flosmagnoliae, respectively. However, before anti-inflammatory investigation is conducted, it is important to obtain the safe doses of these compounds to ensure the validity of the anti-inflammatory results. Therefore, the present study was aimed to investigate the cytotoxicity of eurycomanone and fargesin towards macrophage RAW 264.7. cells to determine the safe doses of these compounds. Different concentrations of eurycomanone and fargesin were subjected to RAW 264.7 cells. The cytotoxicity of the compounds was evaluated by MTT assay and 50% inhibitory concentration (IC50) of these compounds was determined. Morphological changes of RAW 264.7 cells upon exposure to these compounds were also observed. Eurycomanone exhibited its cytotoxic effect by reducing RAW 264.7 cell viability dose-dependently with the IC50 of 94.17 µM. Meanwhile, fargesin had slight cytotoxicity towards RAW 264.7 cells with the IC50 of 173.5µM. Eurycomanone was more cytotoxic towards RAW 264.7 cells compared to fargesin. In conclusion, eurycomanone and fargesin at concentration up to 25 µM, was not toxic to the RAW 264.7 murine macrophages cells and the findings can be applied in the future anti-inflammatory study.

From the whole plant of Dendrobium senile, a new phenanthrene namely 2,5,7-trihydroxy-4-methoxyphenanthrene (1) was isolated, together with seven known compounds including moscatin (2), 2,5-dihydroxy-4,9-dimethoxyphenanthrene (3), moscatilin (4), aloifol I (5), 4,4',8,8'-tetramethoxy[1,1'-biphenanthrene]-2,2',7,7'-tetrol (6), 2,2',7,7'-tetrahydroxy-4,4'-dimethoxy-1,1'-biphenanthrene (7) and bleformin G (8). The structure of the new compound was elucidated by analysis of its spectroscopic data. Moscatin (2) and 2,5-dihydroxy-4,9-dimethoxyphenanthrene (3) showed appreciable pancreatic lipase inhibitory effects when compared with the positive control orlistat.

Cancer stem-like cells (CSCs) are key mediators driving tumor initiation, metastasis, therapeutic failure, and subsequent cancer relapse. Thus, targeting CSCs has recently emerged as a potential strategy to improve chemotherapy. In this study, the anticancer activity and stemness-regulating capacity of 4,5,4′-trihydroxy-3,3′-dimethoxybibenzyl (TDB), a bibenzyl extracted from Dendrobium ellipsophyllum, are revealed in CSCs of various human lung cancer cells. Culture with TDB (5–10 μM) strongly abolished tumor-initiating cells in lung cancer H460, H23, and A549 cells in both anchorage-dependent and anchorage-independent colony formation assays. Through the 3D single-spheroid formation model, attenuation of self-renewal capacity was observed in CSC-enriched populations treated with 1–10 μM TDB for 7 days. Flow cytometry analysis confirmed the attenuation of %cell overexpressing CD133, a CSC biomarker, in TDB-treated lung cancer spheroids. TDB at 5–10 μM remarkably suppressed regulatory signals of p-Akt/Akt, p-GSK3β/GSK3β, and β-catenin corresponding to the downregulated mRNA level of stemness transcription factors including Nanog, Oct4, and Sox2. Moreover, the antiapoptosis Bcl-2 and Mcl-1 proteins, which are downstream molecules of Akt signaling, were evidently decreased in CSC-enriched spheroids after culture with TDB (1–10 μM) for 24 h. Interestingly, the diminution of Akt expression by specific siAkt effectively reversed suppressive activity of TDB targeting on the CSC phenotype in human lung cancer cells. These findings provide promising evidence of the inhibitory effect of TDB against lung CSCs via suppression of Akt/GSK3β/β-catenin cascade and related proteins, which would facilitate the development of this bibenzyl natural compound as a novel CSC-targeted therapeutic approach for lung cancer treatment.

Background/aim Integrin-targeting compounds have shown clinically significant benefits in many patients. Here, we examined the activity of millettocalyxin B, extracted from the stem bark of Millettia erythrocalyx, in lung cancer cells. Materials and methods The viability of human lung cancer cells was investigated by the 3-(4,5-dimethylthiazol-2-yl)-2,5diphenyl tetrazoliumbromide (MTT) assay. Migration and invasion assays were performed. Phalloidin-rhodamine staining was used to determine the formation of filopodia. Western blot analysis and immunofluorescence staining were used to identify the signaling proteins involved in migration regulation. Results Non-toxic concentrations (0-25 μM) of millettocalyxin B reduced migration and invasion of lung cancer A549 cells. Filopodia were significantly reduced in millettocalyxin B-treated cells. The migration regulatory proteins including integrin α5, active FAK, active Akt, and Cdc42 were significantly decreased in Millettocalyxin B-treated cells. Conclusion Our findings revealed a novel anti-migration and anti-invasion effects and the underlying mechanism of millettocalyxin B, which may be exploited for cancer treatment.

Although many natural products have proven their potential to regulate obesity through the modulation of adipocyte biology, none of them has yet been approved for clinical use in obesity therapy. This work aims to isolate valuable secondary metabolites from an orchid species (Dendrobium heterocarpum) and evaluate their possible roles in the growth and differentiation of 3T3-L1 pre-adipocytes. Six compounds were isolated from the orchid’s methanolic extracts and identified as amoenylin (1), methyl 3-(4-hydroxyphenyl) propionate (2), 3,4-dihydroxy-5,4’-dimethoxybibenzyl (3), dendrocandin B (4), dendrofalconerol A (5), and syringaresinol (6). Among these phytochemicals, compounds 2, 3, and 6 exhibited lower effects on the viability of 3T3-L1 cells, offering non-cytotoxic concentrations of ≲10 µM. Compared to others tested, compound 3 was responsible for the maximum reduction of lipid storage in 3T3-L1 adipocytes (IC50 = 6.30 ± 0.10 µM). A set of protein expression studies unveiled that compound 3 at non-cytotoxic doses could suppress the expression of some key transcription factors in adipocyte differentiation (i.e., PPARγ and C/EBPα). Furthermore, this compound could deactivate some proteins involved in the MAPK pathways (i.e., JNK, ERK, and p38). Our findings prove that D. heterocarpum is a promising source to explore bioactive molecules capable of modulating adipocytic growth and development, which can potentially be assessed and innovated further as pharmaceutical products to defeat obesity.

Background/aim Epithelial to mesenchymal transition (EMT), and focal adhesion kinase (FAK) facilitate lung cancer cell motility and survival. We, therefore, investigated the antimigratory effect of 3,4-dihydroxy-5,4'-dimethoxybibenzyl (DS-1) on human lung cancer cells. Materials and methods Cell viability and proliferation were examined by the 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide assay. Filopodia formation, migration, and anchorage-independent growth assays were performed to assess metastatic behaviors while EMT-related proteins, integrins, and FAK-RhoA pathway were evaluated by western blot analysis. Results We found that DS-1 significantly inhibited the proliferation of lung cancer cells compared to the control. The aggressive behavior of cancer cells, including migration and invasion, was significantly reduced by DS-1. Anchorage-independent growth analysis provided evidence that DS-1 suppressed the growth and survival of cancer cells in detached conditions as indicated by the significant reduction in size and number of colonies. With regard to the mechanisms involved, we found that DS-1-suppressed EMT, as indicated by the reduction of EMT markers, namely N-cadherin, SNAIL and SLUG, and increased levels of the epithelial marker, E-cadherin. In addition, DS-1 was shown to reduce the level of integrin β1 protein and FAK activation. Conclusion DS-1 suppressed lung cancer metastasis via suppressing EMT, integrin β1 expression and FAK-related signaling.

Bacterial meningitis remains one of the most prevalent infectious diseases worldwide. Although advances in medical care have improved mortality and morbidity, neurological complications remain high. Therefore, aside from antibiotics, therapeutic adjuvants targeting neuroinflammation are essential to combat the long-term neuronal sequelae of bacterial meningitis. In the present study, we propose (−)-dendroparishiol as a potential add-on therapy to improve neuroinflammation associated with bacterial meningitis. The biological activity of (−)-dendroparishiol was first predicted by computational analysis and further confirmed in vitro using a cell-based assay with LPS-induced BV-2 microglial cells. Biological pathways involved with (−)-dendroparishiol were identified by applying network pharmacology. Computational predictions of biological activity indicated possible attenuation of several inflammatory processes by (−)-dendroparishiol. In LPS-induced BV-2 microglial cells, (−)-dendroparishiol significantly reduced the expression of inflammatory mediators: iNOS, NO, COX-2, IL-6, and TNF-α. Molecular docking results demonstrated the potential iNOS and COX-2 inhibitory activity of (−)-dendroparishiol. Network pharmacological analysis indicated the plausible role of (−)-dendroparishiol in biological processes involved in oxidative stress and neuroinflammation with enrichment in neuroinflammatory pathways. Overall, this study provides scientific evidence for the potential application of (−)-dendroparishiol in the management of bacterial meningitis-associated neuroinflammation.

Background/aim: Dermal papilla cells (DPCs) regulate hair follicle development. We aimed to investigate the effect of scoparone from Dendrobium densiflorum on DPCs in the induction of stem cell properties and pluripotency-related proteins. Materials and methods: DPC viability was evaluated by the MTT assay. Apoptosis or necrosis of DPCs was determined by Hoecsht33342/PI nuclear staining analysis. Expression of OCT4, NANOG and SOX2 genes was determined using Real-Time Polymerase Chain Reaction (PCR). Immunocytochemistry and western blot analysis were performed to determine pluripotency related proteins. Results: Scoparone increased the expression of pluripotency related transcription factors SOX2 and NANOG, while it had minimal effects on OCT4 levels. Scoparone exerted its stemness-enhancing activity through the up-regulation of Akt-dependent inhibition of GSK3β, resulting in increased cellular levels of β-catenin. Conclusion: Our results show a potential novel activity and mechanism of action of scoparone on human DPCs that could facilitate the development of hair enrichment approaches.

Batatasin III is a stilbenoid compound present in a wide variety of

Large amounts of Morus alba L. (MA) roots are needed as the source of active stilbenes in the industrial production of traditional medicines and cosmeceuticals. A recent investigation demonstrated resveratrol and its derivatives to be promising anti-COVID-19 agents. However, conventional cultivation of MA does not meet the demand for its stilbenes, and root quality usually varies between crops. This study established the in vitro non-GMO root culture of MA and optimized the root density, precursor feeding, and elicitors for stilbene productivity. A root culture with optimal inoculum density (3 g/flask of 30 mL medium) accumulated mulberroside A, oxyresveratrol, and resveratrol at 18.7 ± 1.00 mg/g, 136 ± 5.05 µg/g, and 41.6 ± 5.84 µg/g dry weight (DW), respectively. The feeding of L-tyrosine shortened the time required to reach the stilbene productive stage. Root cultures co-treated with 200 µM methyl jasmonate and 2 mg/mL yeast extract accumulated the highest contents of mulberroside A (30.3 ± 2.68 mg/g DW), oxyresveratrol (68.6 ± 3.53 µg/g DW), and resveratrol (10.2 ± 0.53 µg/g DW). In summary, root culture is a promising and sustainable source of stilbenes for the development of health products and agents for further investigation as potential anti-COVID-19 agents. Electronic supplementary material The online version of this article (10.1007/s00449-020-02474-7) contains supplementary material, which is available to authorized users.

Phenolic compounds are active compounds in Dendrobium formosum Roxb. ex Lindl. that have antidiabetic and anti-obesity activities. To obtain a high extraction yield of phenolic compounds from D. formosum, optimization of microwave-assisted extraction (MAE) was developed in this study using response surface methodology. The optimal conditions for phenolic compound yield were using absolute ethanol as the solvent, 540 W of microwave power, 49.37 s of extraction time, a 22.67 mL/g solvent-to-feed ratio, and three extraction cycles. The yields of coelonin, erianthridin, moscatilin, lusianthridin, gigantol and batatasin III when extracted under optimal conditions were 0.38 ± 0.04, 0.40 ± 0.05, 1.45 ± 0.18, 0.14 ± 0.01, 0.47 ± 0.05 and 0.58 ± 0.06 mg/g dry weight, respectively, with errors of less than 4% deviation from the predicted values. Our results indicate that MAE can obtain a higher yield of phenolic compounds and have a greater effect on glucose uptake than those of other extraction methods. Therefore, MAE has high efficiency for extracting phenolic compounds from D. Fomosum, yielding extracts with a high amount of compounds that exhibit antidiabetic activity.

Purpose: To examine the glucose uptake stimulatory activity of the root extract of Cissus javana DC. (Vitaceae) in Lδ myotubes of rat, and also to identify the extract’s active principles.Methods: The methanol extract was prepared from Cissus javana tuberous roots and evaluated for glucose uptake stimulatory effects on Lδ rat muscle cells and inhibitory activity against α-glucosidase. The chemical components were isolated using several chromatographic techniques, and their structures characterized by spectroscopic methods. Each isolate was then assayed for glucose uptake stimulatory and α-glucosidase inhibitory activities.Results: The extract (100 μg/ml) exhibited glucose uptake stimulatory effect (70.9 % enhancement) and α-glucosidase enzyme inhibitory activity (100 % inhibition). Through chromatographic separation, bergenin, stigmast-4-en-3-one and β-sitosterol were isolated and identified. Bergenin, at 100 μg/ml (0.3046 mM), increased glucose uptake by Lδ myotubes by 50.5 % without toxicity. At the same concentration, bergenin showed no inhibition on α-glucosidase enzyme, while stigmast-4-en-3-one and β-sitosterol exhibited 98.6 and 40.6 %, inhibition, respectively.Conclusion: This study is the first report on the chemical constituents, and the glucose uptake stimulatory and α-glucosidase inhibitory activities of Cissus javana DC. roots. The findings reveal the antidiabetic potential of the plant and the glucose-uptake enhancing activity of bergenin. Keywords: Cissus javana, α-Glucosidase, Antidiabetes, Rat skeletal muscle cells, Bergenin

(+)-7-O-Methylisomiroestrol (MeI), a novel chromene, was discovered as a phytoestrogen in the Pueraria candollei var. mirifica (Airy Shaw & Suvat.) Niyomdham (PM) root having been used as an active agent against oestrogen depletion disorders. The identification of PM phytochemicals is crucial for the development of standardised botanical drugs of PM. MeI was purified from the root cortex of PM, and its structure was elucidated using NMR and mass spectrometry. The content of MeI in the root bark of the PM root was 2.1–6.5 × 10−3% (w/w). The oestrogenic potency of MeI was stronger than that of isomiroestrol but less than that of deoxymiroestrol and miroestrol. Therefore, MeI is a new oestrogenic biomarker for the effective chemical standardisation of the PM extract for health product development.

In searching for novel targeted therapeutic agents for lung cancer treatment, norcycloartocarpin from Artocarpus gomezianus was reported in this study to promisingly interacted with Akt and exerted the apoptosis induction and epithelial-to-mesenchymal transition suppression. Selective cytotoxic profile of norcycloartocarpin was evidenced with approximately 2-fold higher IC50 in normal dermal papilla cells (DPCs) compared with human lung cancer A549, H460, H23, and H292 cells. We found that norcycloartocarpin suppressed anchorage-independent growth, cell migration, invasion, filopodia formation, and decreased EMT in a dose-dependent manner at 24 h, which were correlated with reduced protein levels of N-cadherin, Vimentin, Slug, p-FAK, p-Akt, as well as Cdc42. In addition, norcycloartocarpin activated apoptosis caspase cascade associating with restoration of p53, down-regulated Bcl-2 and augmented Bax in A549 and H460 cells. Interestingly, norcycloartocarpin showed potential inhibitory role on protein kinase B (Akt) the up-stream dominant molecule controlling EMT and apoptosis. Computational molecular docking analysis further confirmed that norcycloartocarpin has the best binding affinity of -12.52 kcal/mol with Akt protein at its critical active site. As Akt has recently recognized as an attractive molecular target for therapeutic approaches, these findings support its use as a plant-derived anticancer agent in cancer therapy.

From the whole plant of

Three novel actinomycete strains, designated as DR6-1T, DR6-2 and DR6-4, isolated from the roots of Dendrobium heterocarpum Lindl in Thailand were studied using a polyphasic taxonomic approach. The strains grew at 20–37 °C, at pH 5–10 and with 5 % (w/v) NaCl. They contained meso-diaminopimelic acid in the cell-wall peptidoglycan and MK-9(H4) was a major menaquinone. Arabinose and galactose were the major sugars in the cell wall. The predominant cellular fatty acids were iso-C16 : 0 and iso-C15 : 0. The detected polar lipids were diphosphatidylglycerol, hydroxyphosphatidylethanolamine, phosphatidylethanolamine, phosphatidylinositol and phosphatidylglycerol. Strains DR6-1T, DR6-2 and DR6-4 shared 99.9–100 % 16S rRNA gene sequence similarity and were closely related to Amycolatopsis echigonensis JCM 21831T (98.7-98.8%). The approximate genome size of strain DR6-1T was 9.6 Mb with a G+C content of 69.6 mol%. The ANIb and dDDH values between genomic sequences of strain DR6-1T and Amycolatopsis echigonensis JCM21831T, Amycolatopsis rubida JCM 10871T and Amycolatopsis nivea KCTC 39515T were 90.55, 92.25, 92.60%, and 47.20, 52.10 and 52.50%, respectively. Based on the phenotypic, chemotaxonomic and genotypic characteristics, it has been concluded that strains DR6-1T, DR6-2 and DR6-4 represent a novel species of the genus Amycolatopsis for which the name Amycolatopsis dendrobii sp. nov. is proposed. The type strain is DR6-1T (=JCM 33742T=KCTC 49546T=TISTR 2840T).

Paphiopedilum callosum (Rchb.f.) Stein belongs to the Orchidaceae family. The phytochemistry and bioactivities of this plant have not been reported. In this study, a bioassay-guided isolation with a yeast cell-based assay was performed to isolate topoisomerase I poison compounds from the roots of P. callosum. One new and five known compounds were isolated. According to their spectroscopic data (ESI-MS, NMR, IR and UV), the new compound (1) is 3′-hydroxy-2,6,5′-trimethoxystilbene. The known compounds (2-6) were identified by comparing their spectroscopic data with those in the literature. According to the yeast cell-based assay, all the compounds were topoisomerase Ι poisons based on their ability to inhibit the growth of yeast cells. As further confirmation, the cytotoxic activities of the isolated compounds were evaluated using human cancer cell lines (MCF-7 and NCI-H187). The compounds exhibited varying degrees of cytotoxicity on the representative cell lines. These isolated compounds could serve as new lead compounds in the development of cancer chemotherapeutics.

Background Oxyresveratrol is a major bioactive component derived from the heartwood of Artocarpus lacucha. This compound exerts several biological activities, including neuroprotective effects in vitro and in vivo. However, there is limited pharmacokinetic information on this compound, especially its distribution in neuronal tissue and its route of excretion. The aim of this study was to investigate the pharmacokinetic profiles of oxyresveratrol alone and in combination with piperine as a bioenhancer in rats. Methods Male Wistar rats were administered with oxyresveratrol 10 mg/kg, oxyresveratrol 10 mg/kg plus piperine 1 mg/kg via intravenous or oxyresveratrol 100 mg/kg, oxyresveratrol 100 mg/kg plus piperine 10 mg/kg via oral gavage. Plasma, internal organs, urine, and feces were collected. Determination of the oxyresveratrol concentration in biological samples was performed by liquid chromatography tandem mass spectrometry. Results The combination with piperine had shown a significantly higher maximum concentration in plasma approximately 1500 μg/L within 1–2 h after oral dosing, and could increase oral bioavailability of oxyresveratrol approximately 2–fold. Oxyresveratrol could widely distributed most of the internal organs with a tissue to plasma ratio of 10–100 fold within 5 min after dosing. Urinary excretion of oxyresveratrol glucuronide was the major route of excretion after administration of oxyresveratrol alone and in combination with piperine. Conclusion The addition of piperine could enhance some of the pharmacokinetic properties of oxyresveratrol via both intravenous and oral administration. This pharmacokinetic information will be useful for appropriate strategies to develop oxyresveratrol as a phytopharmaceutical product. Electronic supplementary material The online version of this article (10.1186/s12906-019-2653-y) contains supplementary material, which is available to authorized users.

Derris scandens (Roxb.) Benth. is a medicinal plant used for treatment of musculoskeletal pain in Thai traditional medicines. Its stem contains active compound genistein-7-O-[α-rhamnopyranosyl-(1 to 6)-β-glucopyranoside] (GTG) which is used as a biomarker for standardization of D. scandens extracts. As an alternative for rapid quantitation of GTG, a monoclonal antibody against GTG was prepared and applied for an indirect competitive enzyme-linked immunosorbent assay (ELISA) to determine GTG in plants and herbal products. The established method provided a quantification range of 0.31-10 µg/mL with a limit of detection of 0.29 µg/mL. The assay was validated for precision and accuracy by intra- and interassay variation analyses, recovery test, and comparison analysis between the amounts of GTG determined by ELISA and HPLC. The results exhibited that the developed ELISA is sensitive and effective for determination of GTG in D. scandens plant materials and herbal products.

Phytostilbenes are responsible for several biological activities of mulberry (Morus sp.), which has been widely used as a raw material in health products. This study aimed to investigate the capability of Morus alba L. cell in bioreactors to produce the major bioactive stilbenes. The cell obtained from air-driven bioreactors such as round bottom, flat bottom, and air-lift vessel shape bioreactors was collected and analyzed for the levels of mulberroside A and oxyresveratrol. The results showed that the cell culture in round bottom and air-lift vessel bioreactors had higher growth rate, as compared with the cell culture in shake flasks (1.38- and 1.41-fold, respectively). The optimized culture condition to produce mulberroside A was obtained from round bottom bioreactor culture (55.56 ± 11.41 μmol/L). Additionally, endogenous stilbenoid hydrolysis of cell from the bioreactor culture was examined. Under optimized hydrolytic conditions, mulberroside A in the cell was readily deglycosylated to give oxyresveratrol within 1 h. These results indicated that the glycoside mulberroside A in the cell is sensitive to the endogenous enzymatic hydrolysis. Interaction of the stilbenoid components with the endogenous hydrolytic enzyme triggered by cell disruption in M. alba samples was suggested to be the major cause of the alteration of the stilbenoid levels. These findings have provided a new approach to producing glycosidic compounds and corresponding aglycones in cell culture.

Integrin-targeting compounds have shown clinically significant benefits in many patients. Here, we examined the activity of millettocalyxin B, extracted from the stem bark of Millettia erythrocalyx, in lung cancer cells.The viability of human lung cancer cells was investigated by the 3-(4,5-dimethylthiazol-2-yl)-2,5diphenyl tetrazoliumbromide (MTT) assay. Migration and invasion assays were performed. Phalloidin-rhodamine staining was used to determine the formation of filopodia. Western blot analysis and immunofluorescence staining were used to identify the signaling proteins involved in migration regulation.Non-toxic concentrations (0-25 μM) of millettocalyxin B reduced migration and invasion of lung cancer A549 cells. Filopodia were significantly reduced in millettocalyxin B-treated cells. The migration regulatory proteins including integrin α5, active FAK, active Akt, and Cdc42 were significantly decreased in Millettocalyxin B-treated cells.Our findings revealed a novel anti-migration and anti-invasion effects and the underlying mechanism of millettocalyxin B, which may be exploited for cancer treatment.

Lusianthridin is a phenanthrene derivative isolated from Dendrobium venustum. Some phenanthrene compounds have antiplatelet aggregation activities via undefined pathways. This study aims to determine the inhibitory effects and potential mechanisms of lusianthridin on platelet aggregation. The results indicated that lusianthridin inhibited arachidonic acid, collagen, and adenosine diphosphate (ADP)-stimulated platelet aggregation (IC50 of 0.02 ± 0.001 mM, 0.14 ± 0.018 mM, and 0.22 ± 0.046 mM, respectively). Lusianthridin also increased the delaying time of arachidonic acid-stimulated and the lag time of collagen-stimulated and showed a more selective effect on the secondary wave of ADP-stimulated aggregations. Molecular docking studies revealed that lusianthridin bound to the entrance site of the cyclooxygenase-1 (COX-1) enzyme and probably the active region of the cyclooxygenase-2 (COX-2) enzyme. In addition, lusianthridin showed inhibitory effects on both COX-1 and COX-2 enzymatic activities (IC50 value of 10.81 ± 1.12 µM and 0.17 ± 1.62 µM, respectively). Furthermore, lusianthridin significantly inhibited ADP-induced suppression of cAMP formation in platelets at 0.4 mM concentration (p &lt, 0.05). These findings suggested that possible mechanisms of lusianthridin on the antiplatelet effects might act via arachidonic acid-thromboxane and adenylate cyclase pathways.

Focal adhesion kinase (FAK) controls several cancer aggressive potentials of cell movement and dissemination. As epithelial–mesenchymal transition (EMT) and the migratory-associated integrins, known influencers of metastasis, have been found to be linked with FAK activity, this study unraveled the potential pharmacological effect of artocarpin in targeting FAK resulting in the suppression of EMT and migratory behaviors of lung cancer cells. Treatment with artocarpin was applied at concentrations of 0–10 μM, and the results showed non-cytotoxicity in lung cancer cell lines (A549 and H460), normal lung (BEAS-2B) cells and primary metastatic lung cancer cells (ELC12, ELC16, and ELC20). We also found that artocarpin (0–10 µM) had no effect on cell viability, proliferation, and migration in BEAS-2B cells. For metastasis-related approaches, artocarpin significantly inhibited cell migration, invasion, and filopodia formation. Artocarpin also dramatically suppressed anchorage-independent growth, cancer stem cell (CSC) spheroid formation, and viability of CSC-rich spheroids. For molecular targets of artocarpin action, computational molecular docking revealed that artocarpin had the best binding affinity of −8.0 kcal/mol with FAK protein. Consistently, FAK-downstream proteins, namely active Akt (phosphorylated Akt), active mTOR (phosphorylated mTOR), and Cdc42, and EMT marker and transcription factor (N-cadherin, Vimentin, and Slug), were found to be significantly depleted in response to artocarpin treatment. Furthermore, we found the decrease of Caveolin-1 (Cav-1) accompanied by the reduction of integrin-αν and integrin-β3. Taken together, these findings support the anti-metastasis potentials of the compound to be further developed for cancer therapy.

Cancer metastasis is a major cause of the high mortality rate in lung cancer patients. The cytoskeletal rearrangement and degradation of extracellular matrix are required to facilitate cell migration and invasion and the suppression of these behaviors is an intriguing approach to minimize cancer metastasis. Even though Erianthridin (ETD), a phenolic compound isolated from the Thai orchid Dendrobium formosum exhibits various biological activities, the molecular mechanism of ETD for anti-cancer activity is unclear. In this study, we found that noncytotoxic concentrations of ETD (≤ 50 μM) were able to significantly inhibit cell migration and invasion via disruption of actin stress fibers and lamellipodia formation. The expression of matrix metalloproteinase-2 (MMP-2) and MMP-9 was markedly downregulated in a dose-dependent manner after ETD treatment. Mechanistic studies revealed that protein kinase B (Akt) and its downstream effectors mammalian target of rapamycin (mTOR) and p70 S6 kinase (p70S6K) were strongly attenuated. An in silico study further demonstrated that ETD binds to the protein kinase domain of Akt with both hydrogen bonding and van der Waals interactions. In addition, an in vivo tail vein injection metastasis study demonstrated a significant effect of ETD on the suppression of lung cancer cell metastasis. This study provides preclinical information regarding ETD, which exhibits promising antimetastatic activity against non-small-cell lung cancer through Akt/mTOR/p70S6K-induced actin reorganization and MMPs expression.

A phytochemical investigation on the whole plants of Aerides multiflora revealed the presence of three new biphenanthrene derivatives named aerimultins A–C (1–3) and a new natural phenylpropanoid ester dihydrosinapyl dihydroferulate (4), together with six known compounds (5–10). The structures of the new compounds were elucidated by analysis of their spectroscopic data. All of the isolates were evaluated for their a-glucosidase inhibitory activity. Aerimultin C (3) showed the most potent activity. The other compounds, except for compound 4, also exhibited stronger activity than the positive control acarbose. Compound 3 showed non-competitive inhibition of the enzyme as determined from a Lineweaver–Burk plot. This study is the first phytochemical and biological investigation of A. multiflora.

Cancer stem-like cells (CSCs) are key mediators driving tumor initiation, metastasis, therapeutic failure, and subsequent cancer relapse. Thus, targeting CSCs has recently emerged as a potential strategy to improve chemotherapy. In this study, the anticancer activity and stemness-regulating capacity of 4,5,4′-trihydroxy-3,3′-dimethoxybibenzyl (TDB), a bibenzyl extracted from Dendrobium ellipsophyllum, are revealed in CSCs of various human lung cancer cells. Culture with TDB (5–10 μM) strongly abolished tumor-initiating cells in lung cancer H460, H23, and A549 cells in both anchorage-dependent and anchorage-independent colony formation assays. Through the 3D single-spheroid formation model, attenuation of self-renewal capacity was observed in CSC-enriched populations treated with 1–10 μM TDB for 7 days. Flow cytometry analysis confirmed the attenuation of %cell overexpressing CD133, a CSC biomarker, in TDB-treated lung cancer spheroids. TDB at 5–10 μM remarkably suppressed regulatory signals of p-Akt/Akt, p-GSK3β/GSK3β, and β-catenin corresponding to the downregulated mRNA level of stemness transcription factors including Nanog, Oct4, and Sox2. Moreover, the antiapoptosis Bcl-2 and Mcl-1 proteins, which are downstream molecules of Akt signaling, were evidently decreased in CSC-enriched spheroids after culture with TDB (1–10 μM) for 24 h. Interestingly, the diminution of Akt expression by specific siAkt effectively reversed suppressive activity of TDB targeting on the CSC phenotype in human lung cancer cells. These findings provide promising evidence of the inhibitory effect of TDB against lung CSCs via suppression of Akt/GSK3β/β-catenin cascade and related proteins, which would facilitate the development of this bibenzyl natural compound as a novel CSC-targeted therapeutic approach for lung cancer treatment.

Five compounds including a new bisbibenzyl named dendropachol (1) and four known compounds (2–5) comprising 4,5-dihydroxy-2,3-dimethoxy-9,10-dihydrophenanthrene (2), gigantol (3), moscatilin (4) and 4,5,4′-trihydroxy-3,3′-dimethoxybibenzyl (5) were isolated from a methanolic extract of Dendrobium pachyglossum (Orchidaceae). The chemical structures of the isolated compounds were characterized by spectroscopic methods. Dendropachol (1) was investigated for its protective effects on hydrogen peroxide (H2O2)-induced oxidative stress in HaCaT keratinocytes. Compound 1 showed strong free radical scavenging compared to the positive control. For the cytoprotective effect, compound 1 increased the activities of GPx and CAT and the level of GSH but reduced intracellular reactive oxygen species (ROS) generation and accumulation. In addition, compound 1 significantly diminished the expression of p53, Bax, and cytochrome C proteins, decreased the activities of caspase-3 and caspase-9, and increased Bcl-2 protein. The results suggested that compound 1 exhibited antioxidant activities and protective effects in keratinocytes against oxidative stress induced by H2O2.

Four new phenanthrene derivatives, gastrobellinols A-D (1&ndash, 4), were isolated from the methanolic extract of Gastrochilus bellinus (Rchb.f.) Kuntze, along with eleven known phenolic compounds including agrostophyllin (5), agrostophyllidin (6), coniferyl aldehyde (7), 4-hydroxybenzaldehyde (8), agrostophyllone (9), gigantol (10), 4-(methoxylmethyl)phenol (11), syringaldehyde (12), 1-(4&prime, hydroxybenzyl)-imbricartin (13), 6-methoxycoelonin (14), and imbricatin (15). Their structures were determined by spectroscopic methods. Each isolate was evaluated for &alpha, glucosidase inhibitory activity. Compounds 1, 2, 3, 7, 9, 13, and 15 showed higher activity than the drug acarbose. Gastrobellinol C (3) exhibited the strongest &alpha, glucosidase inhibition with an IC50 value of 45.92 &mu, M. A kinetic study of 3 showed competitive inhibition on the &alpha, glucosidase enzyme. This is the first report on the phytochemical constituents and &alpha, glucosidase inhibitory activity of G. bellinus.

Oxidative stress in dermal fibroblasts is strongly correlated with the aging process of the skin. The application of natural compounds that can increase the ability of dermal fibroblasts to counteract oxidative stress is a promising approach to promote skin health and beauty. Eriodictyol is a flavonoid that exerts several pharmacological actions through its antioxidant properties. However, its protective effects on dermal fibroblasts have not yet been investigated. In this study, we investigated whether eriodictyol protects human dermal fibroblasts (BJ fibroblasts) from the harmful effects of hydrogen peroxide (H2O2). Eriodictyol pretreatment significantly prevented necrotic cell death caused by H2O2 exposure. In addition, the level of 2′,7′-dichloro-dihydro-fluorescein oxidation was decreased, and that of glutathione was maintained, indicating that the beneficial effects of eriodictyol against H2O2 were closely associated with oxidative-stress attenuation. Eriodictyol mediates its antioxidant effects on dermal fibroblasts against H2O2 through (i) the direct neutralization of reactive oxygen species; (ii) the enhancement of the activities of H2O2-detoxifying enzymes, including catalase and glutathione peroxidase; and (iii) the induction of the expressions of catalase and glutathione peroxidase 1 via the activation of the Nrf2 signaling system. These results support the potential application of eriodictyol as an ingredient in skincare products for cosmeceutical and pharmaceutical purposes.

Four new phenanthrene derivatives, gastrobellinols A-D (1–4), were isolated from the methanolic extract of Gastrochilus bellinus (Rchb.f.) Kuntze, along with eleven known phenolic compounds including agrostophyllin (5), agrostophyllidin (6), coniferyl aldehyde (7), 4-hydroxybenzaldehyde (8), agrostophyllone (9), gigantol (10), 4-(methoxylmethyl)phenol (11), syringaldehyde (12), 1-(4′-hydroxybenzyl)-imbricartin (13), 6-methoxycoelonin (14), and imbricatin (15). Their structures were determined by spectroscopic methods. Each isolate was evaluated for α-glucosidase inhibitory activity. Compounds 1, 2, 3, 7, 9, 13, and 15 showed higher activity than the drug acarbose. Gastrobellinol C (3) exhibited the strongest α-glucosidase inhibition with an IC50 value of 45.92 μM. A kinetic study of 3 showed competitive inhibition on the α-glucosidase enzyme. This is the first report on the phytochemical constituents and α-glucosidase inhibitory activity of G. bellinus.

Due to the high mortality of lung cancer, natural derivative compounds have been promoted as versatile sources for anticancer drug discovery. Erianthridin, a phenanthrene compound isolated from Dendrobium formosum, exhibits intriguing apoptosis-inducing effects in non-small cell lung cancer cells. Apoptotic nuclei staining assays showed that apoptotic cells with DNA fragmentation and apoptotic bodies were apparent, and an increase in annexin V-FITC-positive cells were found in cells treated with erianthridin. The apoptosis protein markers for cleaved caspase-3 and cleaved poly-ADP-ribose polymerase were significantly upregulated in response to erianthridin. A mechanistic investigation revealed that erianthridin was able to attenuate extracellular signal-regulated kinase activity and thereby mediate apoptosis through the modulation of Bcl-2 family protein levels. U0126, an extracellular signal-regulated kinase inhibitor, augmented the apoptosis-inducing effect of erianthridin; in contrast, overexpression of exogenous extracellular signal-regulated kinase substantially abrogated erianthridin activity. Furthermore, an in vitro 3D tumorigenesis assay showed that erianthridin was able to potentially suppress lung cancer cell proliferation. This study is the first to report a promising cytotoxic effect of erianthridin, which provides preclinical evidence for further research and development of this compound.

Two new compounds, dihydrodengibsinin (1) and dendrogibsol (2), were isolated from the whole plant of Dendrobium gibsonii, together with seven known compounds (3&ndash, 9). The structures of the new compounds were elucidated by their spectroscopic data. All these isolates were evaluated for their &alpha, glucosidase inhibitory activities. Dendrogibsol (2) and lusianthridin (7) showed strong &alpha, glucosidase inhibitory activity when compared with acarbose. An enzyme kinetic study revealed that dendrogibsol (2) is a noncompetitive inhibitor of &alpha, glucosidase.

Two new compounds, dihydrodengibsinin (1) and dendrogibsol (2), were isolated from the whole plant of Dendrobium gibsonii, together with seven known compounds (3–9). The structures of the new compounds were elucidated by their spectroscopic data. All these isolates were evaluated for their α-glucosidase inhibitory activities. Dendrogibsol (2) and lusianthridin (7) showed strong α-glucosidase inhibitory activity when compared with acarbose. An enzyme kinetic study revealed that dendrogibsol (2) is a noncompetitive inhibitor of α-glucosidase.

The phytochemical investigation of Huberantha jenkinsii resulted in the isolation of two new and five known compounds. The new compounds were characterized as undescribed 8-oxoprotoberberine alkaloids and named huberanthines A and B, whereas the known compounds were identified as allantoin, oxylopinine, N-trans-feruloyl tyramine, N-trans-p-coumaroyl tyramine, and mangiferin. The structure determination was accomplished by spectroscopic methods. To evaluate therapeutic potential in diabetes and Parkinson&rsquo, s disease, the isolates were subjected to assays for their &alpha, glucosidase inhibitory activity, cellular glucose uptake stimulatory activity, and protective activity against neurotoxicity induced by 6-hydroxydopamine (6-OHDA). The results suggested that mangiferin was the most promising lead compound, demonstrating significant activity in all the test systems.

The phytochemical investigation of Huberantha jenkinsii resulted in the isolation of two new and five known compounds. The new compounds were characterized as undescribed 8-oxoprotoberberine alkaloids and named huberanthines A and B, whereas the known compounds were identified as allantoin, oxylopinine, N-trans-feruloyl tyramine, N-trans-p-coumaroyl tyramine, and mangiferin. The structure determination was accomplished by spectroscopic methods. To evaluate therapeutic potential in diabetes and Parkinson’s disease, the isolates were subjected to assays for their α-glucosidase inhibitory activity, cellular glucose uptake stimulatory activity, and protective activity against neurotoxicity induced by 6-hydroxydopamine (6-OHDA). The results suggested that mangiferin was the most promising lead compound, demonstrating significant activity in all the test systems.

Background/aim Epithelial to mesenchymal transition (EMT) is a cellular process that facilitates cancer metastasis. Therefore, therapeutic approaches that target EMT have garnered increasing attention. The present study aimed to examine the in vitro effects of ephemeranthol A on cell death, migration, and EMT of lung cancer cells. Materials and methods Ephemeranthol A was isolated from Dendrobium infundibulum. Non-small cell lung cancer cells H460 were treated with ephemeranthol A and apoptosis was evaluated by Hoechst 33342 staining. Anoikis resistance was determined by soft agar assay. Wound healing assay was performed to test the migration. The regulatory proteins of apoptosis and cell motility were determined by western blot. Results Treatment with ephemeranthol A resulted in a concentration-dependent cell apoptosis. At non-toxic concentrations, the compound could inhibit anchorage-independent growth of the cancer cells, as indicated by the decreased colony size and number. Ephemeranthol A also exhibited an inhibitory effect on migration. We further found that ephemeranthol A exerts its antimetastatic effects via inhibition of EMT, as indicated by the markedly decrease of N-cadherin, vimentin, and Slug. Furthermore, the compound suppressed the activation of focal adhesion kinase (FAK) and protein kinase B (Akt) proteins, which are key regulators of cell migration. As for the anticancer activity, ephemeranthol A induced apoptosis by decreasing Bcl-2 followed by the activation of caspase 3 and caspase 9. Conclusion The pro-apoptotic and anti-migratory effects of ephemeranthol A on human lung cancer cells support its use for the development of novel anticancer therapies.

Supplemental material, Supplementary material, for α-Glucosidase Inhibitory and Glucose Uptake Stimulatory Effects of Phenolic Compounds From Dendrobium christyanum by Htoo Tint San, Panitch Boonsnongcheep, Waraporn Putalun, Wanwimon Mekboonsonglarp, Boonchoo Sritularak and Kittisak Likhitwitayawuid in Natural Product Communications

Background/aim Lung cancer is by far the most common cause of cancer mortality, accounting for nearly 20% of all global cancer deaths. Therefore, potent and effective compounds for treatment of this cancer type are essential. Phoyunnanin E, isolated from Dendrobium venustum (Orchidaceae), has promising pharmacological activities; however, it is unknown if phoyunnanin E affects apoptosis of lung cancer cells. Materials and methods The apoptosis-inducing activity of phoyunnanin E on H460 lung cancer cells was investigated by Hoechst 33342, and annexin V-fluorescein isothiocyanate/propidium iodide staining. The underlying mechanism was determined via monitoring apoptosis-regulatory proteins by western blot analysis. The apoptotic effect of the compound was confirmed in H23 lung cancer cells. Results Phoyunnanin E significantly induced apoptotic cell death of H460 lung cancer cells, as indicated by condensed and fragmented nuclei with the activation of caspase-3 and -9 and poly (ADP-ribose) polymerase cleavage. Phoyunnanin E mediated apoptosis via a p53-dependent pathway by increasing the accumulation of cellular p53 protein. As a consequence, anti-apoptotic proteins including induced myeloid leukemia cell differentiation protein (MCL1) and B-cell lymphoma 2 (BCL2) were found to be significantly depleted, while pro-apoptotic BCL-2-associated X protein (BAX) protein was up-regulated. Furthermore, it was found that expression of an inhibitor of apoptosis, survivin, markedly reduced in response to phoyunnanin E treatment. The apoptosis-inducting effect was also found in phoyunnanin E-treated H23 lung cancer cells. Conclusion These results indicate the promising effect of phoyunnanin E in induction of apoptosis, that may be useful for the development of novel anticancer agents.

Lung cancer appears to have the highest rate of mortality among cancers due to its metastasis capability. To achieve metastasis, cancer cells acquire the ability to undergo a switch from epithelial to mesenchymal behaviour, termed the epithelial-to-mesenchymal transition (EMT), which is associated with poor clinical outcomes. Drug discovery attempts have been made to find potent compounds that will suppress EMT. Cypripedin, a phenanthrenequinone isolated from Thai orchid, Dendrobium densiflorum, exhibits diverse pharmacological activities. In this study, we found that cypripedin attenuated typical mesenchymal phenotypes, including migratory behaviour, of non-small cell lung cancer H460 cells, with a significant reduction of actin stress fibres and focal adhesion and with weakened anchorage-independent growth. Western blot analysis revealed that the negative activity of this compound on EMT was a result of the down-regulation of the EMT markers Slug, N-Cadherin and Vimentin, which was due to ATP-dependent tyrosine kinase (Akt) inactivation. As a consequence, the increase in the Slug degradation rate via a ubiquitin-proteasomal mechanism was encouraged. The observation in another lung cancer H23 cell line also supported this finding, indicating that cypripedin exhibits a promising pharmacological action on lung cancer metastasis that could provide scientific evidence for the further development of this compound.

Oxidative stress and inflammatory responses are the main causes of various diseases and disorders. To prevent these incidents, antioxidant and anti-inflammatory agents are needed. Dendrobium parishii, known in Thai as “Ueang Khrang Sai San” was studied for chemical constituents by solvent extraction, followed by column chromatography. Seven compounds including two new (1–2) and five known compounds (3–7) were isolated and screened for antioxidant activity using DPPH, ORAC and deoxyribose assays. The new compound (compound 2) showed the highest antioxidant activities and was further evaluated for its antioxidant and anti-inflammatory effects, as well as mechanisms of action in RAW264.7 murine macrophage cells under oxidative stress and inflammation induced by hydrogen peroxide (H2O2) and lipopolysaccharide (LPS), respectively. Compound 2 at 12.5, 25 and 50 μg/mL significantly decreased ROS in H2O2 treated RAW264.7 cells in a dose dependent manner and enhanced antioxidant enzyme (SOD, GPx and CAT) activities. For the anti-inflammatory activity, compound 2 reduced the expression of iNOS and COX-2 in LPS treated RAW264.7 cells. The results indicate that compound 2 has the potential to be developed as an antioxidant and anti-inflammatory agent.

The life-threatening potential of lung cancer has increased over the years due to its acquisition of chemotherapeutic resistance, especially to cisplatin, a first-line therapy. In response to this development, researchers have turned their attention to several compounds derived from natural origins, including cypripedin (CYP), a phenanthrenequinone substance extracted from Dendrobium densiflorum. The aim of the present study was to investigate the ability of CYP to induce apoptosis and enhance cisplatin-mediated death of human lung cancer NCI-H460 cells using cell viability and apoptosis assays. The induction of apoptosis by CYP was observed at a concentration of > 50 μM with the appearance of morphological changes, including DNA condensation and chromatin fragmentation. Together with, CYP was able to activate caspase-3 and downregulate the anti-apoptotic proteins Bcl-2 and Bcl-xL. Also, a non-cytotoxic dose of CYP synergistically potentiated the effect of cisplatin in non-small cell lung cancer line H460 cells, which clearly exhibited the apoptotic phenotype. Western blot analysis revealed that the underlying mechanism involved the downregulation of anti-apoptotic Bcl-xL, whereas the levels of other apoptotic regulatory proteins were not altered. This study provides interesting information on the potent effect of CYP as a chemotherapeutic sensitizer that could be further developed to improve the clinical outcomes of lung cancer patients.

The polyphenol compound, oxyresveratrol (OXY) possesses potent antioxidant and neuroprotective properties of potential utility in the treatment of Alzheimerʼs disease. However, the low oral bioavailability limits its neuroprotective effect and clinical application. The neuroprotective effect of orally administered OXY-loaded self-microemulsifying drug delivery system (OXY-SMEDDS) was compared with free OXY in vivo. Mice were orally administered either free OXY or OXY-SMEDDS once daily at a dose of 90, 180, or 360 mg/kg for 14 d. Mice received a single intracerebroventricular injection of the neurotoxic amyloid β (Aβ)25 – 35 peptide at day 8 during oral treatment. The OXY-SMEDDS formulation resulted in four-times reduction of the free OXY dose required for prevention of neurotoxicity effects due to Aβ 25 – 35 peptide as demonstrated by a significant decline in behavior impairments, lipid oxidation levels, and neuronal cell loss in all hippocampal subfields (p

Cancer stem cells (CSCs) are an important therapeutic target. The therapeutic agents targeting CSCs should lead to improved clinical outcomes. Here we have demonstrated the CSC-suppressing activity of pongol methyl ether (PME), a pure compound from Millettia erythrocalyx. Methods: CSC-suppressing effects were evaluated by spheroid formation assay and detection of CSC markers. The related CSC cell signals were evaluated by Western blot, immunofluorescence and molecular docking analysis. Proteins affected by PME treatment were subjected to bioinformatic analysis. Protein–protein interaction (PPI) networks were constructed by the Search Tool for Interactions of Chemicals (STITCH). The Kyoto Encyclopedia of Genes and Genomes (KEGG) mapper were used to confirm the underlying pathways. Results: PME (5–25 µM) significantly suppressed the ability of lung cancer cells to form colonies, grow in an anchorage-independent manner and generate tumour spheroids. PME at 25 µM significantly decreased the CSC markers (CD133 and ALDH1A1) and pluripotent transcription factors (Oct4 and Nanog). Akt, the key upstream signal of CSC control, was significantly decreased by the PME treatment. The molecular docking indicated that PME was bound to Akt-1 with a binding affinity of −9.2 kcal/mol greater than the Akt-1 inhibitor (reference compound, CQW). The STITCH network identified a total of 15 proteins interacted in PPI networks, and Akt-1 was identified as a central protein. The KEGG mapper indicated that the selected CSC markers were mostly involved in the ‘signalling pathways regulating pluripotency of stem cells’ pathway map and Akt, Oct4 and Nanog were the regulatory proteins in the dominant pathway. In addition, PME (10–25 µM) can suppress spheroid formation and reduce CSC-specific marker expression in patient-derived primary lung cancer cells. Conclusions: Our study revealed a novel pharmacological effect and the underlying mechanism of PME that can attenuate CSC phenotypes in lung cancer cells and may be developed for lung cancer therapy.

Cancer stem cells (CSCs) have been recognized as rare populations driving cancer progression, metastasis, and drug resistance in leading cancers. Attempts have been made toward identifying compounds that specifically target these CSCs. Therefore, investigations of novel therapeutic strategies for CSC targeting are required. The cytotoxic effects of chrysotoxine on human non-small cell lung cancer-derived H460 and H23 cells were evaluated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The effects of chrysotoxine suppression of CSC-like phenotypes were determined in CSC-rich populations and primary CSCs in three-dimensional (3D) culture and in an extreme limiting dilution assay. Expression of CSC markers and associated proteins was determined by Western blot analyse and flow cytometry. We have reported herein the CSC-suppressing activity of chrysotoxine, a bibenzyl compound isolated from Dendrobium pulchellum We have shown, to our knowledge for the first time, that chrysotoxine dramatically suppresses CSC-like phenotypes of H460 and H23 cells. Treatment with chrysotoxine significantly reduced the viability of 3D CSC-rich populations and concomitantly decreased known CSC markers. Chrysotoxine suppressed CSC phenotypes through downregulation of Src/protein kinase B (Akt) signaling. Active (phosphorylated Y416) Src was shown to regulate cancer stemness, since ectopic overexpression of Src strongly activated Akt and subsequently enhanced pluripotency transcription factor SRY (sex-determining region Y)-box 2 (Sox2)- mediating CSC phenotypes, whereas the short hairpin RNA of Src and an Src inhibitor (dasatinib) suppressed Akt, Sox2, and CSC properties. Importantly, chrysotoxine was shown to suppress active Src/Akt signaling and in turn depleted Sox2-mediated CSCs. Our findings indicate a novel CSC-targeted role of chrysotoxine and its regulation by Src/Akt and Sox2, which may be exploited for cancer treatment.