Genipin Analogue (G300) Inhibits Adipogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells through the Suppression of Adipogenic Promoting Factors.

While obesity is a well-known health threatening condition worldwide, effective pharmacological interventions for obesity suppression have been limited due to adverse effects. Therefore, it is important to explore alternative medical treatments for combating obesity. Inhibition of the adipogenesis process and lipid accumulation are critical targets for controlling and treating obesity. Gardenia jasminoides Ellis is a traditional herbal remedy for various ailments. A natural product from its fruit, genipin, has major pharmacological properties; it is anti-inflammatory and antidiabetic. We investigated the effects of a genipin analogue, G300, on adipogenic differentiation in human bone marrow mesenchymal stem cells (hBM-MSCs). G300 suppressed the expression of adipogenic marker genes and adipokines secreted by adipocytes at concentrations of 10 and 20 µM, which effectively reduced the adipogenic differentiation of hBM-MSCs and lipid accumulation in adipocytes. It also improved adipocyte function by lowering inflammatory cytokine secretion and increasing glucose uptake. For the first time, we show that G300 has the potential to be a novel therapeutic agent for the treatment of obesity and its related disorders.

Study on the absolute configuration and biological activity of rotenoids from the leaves and twigs of Millettia pyrrhocarpa Mattapha, Forest & Hawkins, sp. Nov.

BACKGROUND: M. pyrrhocarpa is a new plant in the Fabaceae: Faboideae family that is found in Thailand. A literature search revealed that the Milletia genus is rich in bioactive compounds possessing a wide range of biological activities. In this study, we aimed to isolate novel bioactive compounds and to study their bioactivities. METHODS: The hexane, ethyl acetate, and methanol extracts from the leaves and twigs of M. pyrrhocarpa were isolated and purified using chromatography techniques. These extracts and pure compounds were tested in vitro for their inhibitory activities against nine strains of bacteria, as well as their anti-HIV-1 virus activity and cytotoxicity against eight cancer cell lines. RESULTS: Three rotenoids, named 6aS, 12aS, 12S-elliptinol (1), 6aS, 12aS, 12S-munduserol (2), dehydromunduserone (3), and crude extracts were evaluated for antibacterial, anti-HIV, and cytotoxic activities. It was found that compounds 1-3 inhibited the growth of nine strains of bacteria, and the best MIC/MBC values were obtained at 3/ > 3 mg/mL. The hexane extract showed anti-HIV-1 RT with the highest %inhibition at 81.27 at 200 mg/mL, while 6aS, 12aS, 12S-elliptinol (1) reduced syncytium formation in 1A2 cells with a maximum EC50 value of 4.48 µM. Furthermore, 6aS, 12aS, 12S-elliptinol (1) showed cytotoxicity against A549 and Hep G2 cells with maximum ED50 values of 2.27 and 3.94 µg/mL. CONCLUSION: This study led to the isolation of constituents with potential for medicinal application, providing compounds (1-3) as lead compounds against nine strains of bacteria. The hexane extract showed the highest %inhibition of HIV-1 virus, Compound 1 showed the best EC50 in reducing syncytium formation in 1A2 cells, and it also showed the best ED50 against human lung adenocarcinoma (A549) and human hepatocellular carcinoma (Hep G2). The isolated compounds from M. pyrrhocarpa offered significant potential for future medicinal application studies.

Pyranonaphthoquinones and Naphthoquinones from the Stem Bark of Ventilago harmandiana and Their Anti-HIV-1 Activity.

Seven previously undescribed compounds, including five pyranonaphthoquinones (ventilanones L-P) and two naphthoquinones (ventilanones Q and R), along with 15 known compounds were isolated from the stem bark of Ventilago harmandiana (Rhamnaceae). The structures were established by extensive analysis of their spectroscopic data. The absolute configuration of ventilanone L was established from single crystal X-ray crystallographic analysis using Cu Kα radiation and from its electronic circular dichroism data. Anti-HIV-1 activity using a syncytium inhibition assay and the cytotoxic activities of some isolated compounds were evaluated. Compounds 12, 13, 15, and 16 showed activity against syncytium formation with half maximal effective concentration (EC50) values ranging from 9.9 to 47 µM (selectivity index (SI) 2.4-4.5).

Gambogic Acid Inhibits Wnt/ß-catenin Signaling and Induces ER Stress-Mediated Apoptosis in Human Cholangiocarcinoma.

OBJECTIVE: Gambogic acid (GA) has been reported to induce apoptosis in cholangiocarcinoma (CCA) cell lines. However, the molecular mechanisms underlying its anti-cancer activity remain poorly understood. This study was aimed to investigate GA's effect on human CCA cell lines, KKU-M213 and HuCCA-1, and its associated mechanisms on Wnt/ß-catenin signaling pathway. METHODS: Cell viability, apoptosis, and cell cycle analysis were conducted by MTT and flow cytometry. The effect of GA mediated Wnt/ß-catenin and ER stress were determined by luciferase-reporter assay, qRT-PCR, and western blot analysis. RESULTS: GA exhibited potent cytotoxicity in CCA cells which was associated with significantly inhibited cell proliferation, promoted G1 arrest, and activated caspase 3 mediated-apoptosis. GA attenuated ß-catenin transcriptional levels, decreased ß-catenin protein, and suppressed the expression of c-Myc, a downstream target gene of Wnt/ß-catenin signaling. GA activated genes involved in ER stress mechanism in KKU-M213 and enhanced CCA's sensitivity to gemcitabine. CONCLUSION: Our findings reveal that the molecular mechanism underpinning anti-cancer effect of GA is partially mediated through the inhibition of Wnt/ß-catenin signaling pathway and induction of ER stress induced-apoptosis. GA may serve as a promising therapeutic modality for amelioration of gemcitabine-induced toxicity in CCA.

Synthesis of propargylamine mycophenolate analogues and their selective cytotoxic activity towards neuroblastoma SH-SY5Y cell line.

Twenty six propargylamine mycophenolate analogues were designed and synthesized from mycophenolic acid 1 employing a key step A3-coupling reaction. Their cytotoxic activity was examined against six cancer cell lines. Compounds 6a, 6j, 6t, 6u, and 6z exhibited selective cytotoxicity towards neuroblastoma (SH-SY5Y) cancer cells and were less toxic to normal cells in comparison to the lead compound, MPA 1 and a standard drug, ellipticine. Molecular docking results suggested that compound 6a is fit well in the key amino acid of three proteins (CDK9, EGFR, and VEGFR-2) as targets in cancer therapy. The propargylamine mycophenolate scaffold might be a valuable starting point for development of new neuroblastoma anticancer drugs.

Andrographolide promotes proliferative and osteogenic potentials of human placenta-derived mesenchymal stem cells through the activation of Wnt/ß-catenin signaling.

INTRODUCTION: The in vitro expansion and differentiation of mesenchymal stem cells derived from bone marrow (BM-hMSCs) are considered as potential therapeutic tools for clinical applications in bone tissue engineering and regenerative medicine. However, invasive sampling and reduction in number and proliferative capacity with age are the major limitations of BM-hMSCs. Recently, human placenta-derived MSCs (PL-hMSCs) obtained by a non-invasive procedure have attracted much interest. Attempts to increase the potential of PL-hMSCs would be an important paradigm in regenerative medicine. Herein, we examined the proliferative and osteogenic effect of andrographolide (AP) on PL-hMSCs. METHODS: Mesenchymal stem cells were isolated from full-term normal human placentas and were characterized before using. Cell cytotoxicity and proliferative effect of AP were examined by MTT and BrdU assay, respectively. The non-toxicity concentrations of AP were further assessed for osteogenic effect determined by alkaline phosphatase (ALP) expression and activity, alizarin red staining, and osteoblast-specific gene expressions. Screening of genes involved in osteogenic differentiation-related pathways modulated by AP was explored by a NanoString nCounter analysis. RESULTS: PL-hMSCs generated in this study met the MSC criteria set by the International Society of Cellular Therapy. The non-cytotoxic concentrations of AP on PL-hMSCs are up to 10 µM. The compound increased PL-hMSC proliferation concomitant with increases in Wnt/ß-catenin level and activity. It also enhanced osteogenic differentiation in association with osteoblast-specific mRNA expression. Further, AP promoted bone formation and increased bone structural protein level, osteocalcin, in osteoblastic cells. Gene screening analysis showed the upregulation of genes related to Wnt/ß-catenin, TGFß/BMP, SMAD, and FGF signaling pathways. CONCLUSION: We demonstrated, for the first time, the potential role of AP in promoting proliferation, osteogenic differentiation, and osteoblast bone formation of PL-hMSCs. This study suggests that AP may be an effective novel agent for the improvement of PL-hMSCs and stem cell-based therapy for bone regeneration.

Bioactive tetrahydrofuran lignans from roots, stems, leaves and twigs of Anogeissus rivularis.

Four previously undescribed tetrahydrofuran lignans, named anorisols A-D (1-4) and fourteen known compounds (5-18) were isolated from the roots, stems, leaves and twigs of Anogeissus rivularis. The chemical structures were elucidated on the basis of their spectroscopic data and by comparison with the literature data. The absolute configurations of 1-4 were established by comparison of the experimental ECD spectra with the calculated ECD spectra. Some isolated compounds were evaluated for their cytotoxic activity as well as anti-HIV-1 activity employing reverse transcriptase (RT) and syncytium reduction assays using the ΔTat/RevMC99 virus in 1A2 cell line systems. Compound 6 displayed the most potent activity in syncytium inhibition assay with effective concentration at 50% (EC50) value of 13.3 µM (SI >3.0). In the reverse transcriptase assay, compound 1 exhibited moderate activity with IC50 value of 213.9 µM.

Synthesis and cytotoxic activity of new 7-acetoxy-12-amino-14-deoxy andrographolide analogues.

Two new series of 19-silylether- and 19-formyl-7-acetyl-12-amino-14-deoxyandrographolide analogues were designed and synthesized from natural andrographolide via key step reactions including allylic hydroxylation, tandem CAE reaction and one pot formylation. Evaluation of their cytotoxicity against eight cancer cells line found 6e exhibited the highest activity on MCF-7 cancer cell (IC50 2.93) and comparable to the drug elipticin. Replacement of silylether at C-19 with formyl group exhibited selective activity on P-388 cell line. Computational studies revealed the amino group at C-12 and O-acetoxy at C-7 position play significant roles in cytotoxicity against MCF-7 cancer cells. Cytotoxicity of these two series highlights the importance of 12-substituted-14-deoxyandrographolide scaffold and these types of compounds could be employed in future developments against breast cancer.

Boesenmaxane Diterpenoids from Boesenbergia maxwellii.

Three new diterpenoids, boesenmaxanes A-C (1-3), with an unprecedented core skeleton consisting of an unusual C-C bond between C-12 and an exo-cyclic methylene C-13, were isolated from the rhizome extracts of Boesenbergia maxwellii. The structures were elucidated by analysis of spectroscopic and X-ray diffraction data. Electronic circular dichroism spectra were used to determine the absolute configuration. All the isolates were evaluated for their cytotoxic effects, anti-HIV activity, and antimicrobial activity. Boesenmaxanes A and C (1 and 3) showed significant inhibitory activity in the syncytium reduction assay, with EC50 values of 55.2 and 27.5 µM, respectively.

Andrographolide modulates OPG/RANKL axis to promote osteoblastic differentiation in MC3T3-E1 cells and protects bone loss during estrogen deficiency in rats.

A decline of estrogen in menopause women is accompanied with increases in many pro-inflammatory cytokines and osteoporosis. Andrographolide (AP), from Andrographis paniculata, which has an anti-inflammatory activity, may have potential to alleviate osteoporosis during estrogen deficiency. Here we report the promoting effects of AP on the differentiation of mouse pre-osteoblastic (MC3T3-E1) cells by increasing the expression and activity of alkaline phosphatase (ALP), an osteoblastic gene-specific marker. AP also accelerated bone formation and increased bone structural gene production including collagen and osteocalcin. We demonstrate for the first time the promoting effect of AP on the differentiation of osteoblast involved with the OPG/RANKL signaling pathway. AP also protected bone loss in the estrogen-deficient (ovariectomized, OVX) rats after 12 weeks of treatment. It protected the loss of bone mineral density, bone microarchitecture, and improved bone turnover rate in OVX rats. This study provides an essential evidence for clinical applications and development of AP towards treating osteoporosis in post-menopausal women.

Cleistanthin A induces apoptosis and suppresses motility of colorectal cancer cells.

Colorectal cancer (CRC) is a leading cause of cancer-related deaths worldwide. Here, we investigated the molecular mechanisms that underpin the anticancer effects of cleistanthin A (CA) in two CRC cell lines, HCT 116, and SW480. At 48 h, CA exhibited apoptotic cytotoxic effects in both CRC cell lines, concomitant with reduction of an anti-apoptotic protein, survivin. Mechanistically, CA treatment significantly reduced the expression levels of ß-catenin and active-ß-catenin in a dose-dependent manner in both CRC cell lines. Moreover, CA suppressed the Wnt/ß-catenin signaling pathway by decreasing ß-catenin-mediated transcriptional activity and expression of ß-catenin target genes, AXIN2, CCND1, and survivin. Furthermore, CA also inhibited transcriptional activity in cells overexpressing a constitutively active ß-catenin S33Y, indicating a GSK-3ß-independent mechanism underlying the observed CA effects on CRC cells. Although cytotoxic activity was not observed with CA treatment at 24 h, cell migration and invasion were significantly reduced. In addition, CA suppressed V-type ATPase activity and focal adhesion kinase (FAK) phosphorylation. Collectively, our study reveals that CA has time-dependent effects on CRC cell phenotypes. First, short-term CA treatment inhibited CRC cell migration and invasion partly through the suppression of V-type ATPase activity. This suppression resulted in reduced FAK activation. Second, longer-term CA treatment decreased cell viability which correlated with the suppression of Wnt/ß-catenin signaling induced transcriptional activity. Altogether, our data suggest that CA has the potential to develop as an effective and novel therapeutic drug for CRC patients.

Inhibition of Adipogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells by a Phytoestrogen Diarylheptanoid from Curcuma comosa.

We investigated the effect of a phytoestrogen, (3R)-1,7-diphenyl-(4E,6E)-4,6-heptadien-3-ol (DPHD), from Curcuma comosa Roxb. (Zingiberaceae family) on the adipogenic differentiation of mesenchymal progenitors, human bone marrow-derived mesenchymal stem cells (hBMSCs). DPHD inhibited adipocyte differentiation of hBMSCs by suppressing the expression of genes involved in adipogenesis. DPHD at concentrations of 0.1, 1, and 10 µM significantly decreased triglyceride accumulation in hBMSCs to 7.1 ± 0.2, 6.3 ± 0.4, and 4.9 ± 0.2 mg/dL, respectively, compared to the nontreated control (10.1 ± 0.9 mg/dL) (p < 0.01). Based on gene expression profiling, DPHD increased the expression of several genes involved in the Wnt/ß-catenin signaling pathway, a negative regulator of adipocyte differentiation in hBMSCs. DPHD also increased the levels of essential signaling proteins which are extracellular signal-regulated kinases 1 and 2 (ERK1/2) and glycogen synthase kinase 3 beta (GSK-3ß) that link estrogen receptor (ER) signaling to Wnt/ß-catenin signaling. In conclusion, DPHD exhibited the anti-adipogenic effect in hBMSCs by suppression of adipogenic markers in hBMSCs through the activation of ER and Wnt/ß catenin signaling pathways. This finding suggests the potential role of DPHD in preventing bone marrow adiposity which is one of the major factors that exacerbates osteoporosis in postmenopause.

Design and synthesis of C-12 dithiocarbamate andrographolide analogues as an anticancer agent.

A series of 21 new analogues of C-12 dithiocarbamate andrographolide was designed and synthesized from natural andrographolide isolated from a common Thai plant, Andrographis paniculata. The reaction used to manipulate the andrographolide scaffold was conducted in one pot under mild reaction conditions. This avoided toxic catalysts and gave nearly quantitative yields of new analogues, generally without by-products and can be easily scaled -up for industrial processing. All new analogues were evaluated against nine cancer cell lines, some analogues exhibited greater selective cytotoxic activity to MCF-7 cancer cell than that of the parent andrographolide and cancer drugs.

Cytotoxic compounds from the leaves and stems of the endemic Thai plant Mitrephora sirikitiae.

Context: Mitrephora sirikitiae Weeras., Chalermglin & R.M.K. Saunders (Annonaceae) is a plant endemic to Thailand. Its constituents and their biological activities are unknown.Objective: Isolation and identification of the compounds in the leaves and stems of M. sirikitiae and determination of their cytotoxicity.Materials and methods: Methanol extracts of the leaves and stems of M. sirikitiae were separated by chromatography, and spectroscopic methods were used to determine the structures of the components. The cytotoxicity of the extracts and pure compounds was evaluated using the sulforhodamine B assay with several cell lines. The cells were treated with the compounds at concentrations of 0.16-20 µg/mL for 48 or 72 h.Results: The investigation of the extracts of M. sirikitiae leaves and stems resulted in the isolation of a new lignan, mitrephoran, and 15 known compounds. Among these compounds, 2-(3,4-dimethoxyphenyl)-6-(3,5-dimethoxyphenyl)-3,7-dioxabicyclo[3.3.0]octane, ciliaric acid, 6-methoxymarcanine A, and stepharanine were isolated from this genus for the first time. The alkaloids liriodenine and oxoputerine exhibited strong cytotoxicity against all tested cells (IC50 values of 6.59-11.02 µM). In contrast, magnone A, 3',4-O-dimethylcedrusin, and 6-methoxymarcanine A inhibited the growth of some of the tested cells (IC50 values of 2.03-19.73 µM). Magnone A and 6-methoxymarcanine A showed low toxicity for Hek 293 cells (IC50 >20 µM).Discussion and conclusions: M. sirikitiae is a source of cytotoxic lignans and alkaloids. Among the cytotoxic compounds, magnone A and 6-methoxymarcanine A are potentially useful lead compounds for the further development of anticancer agents because of their selective inhibitory effects on cancer cell lines.

Inhibition of topoisomerase IIα and induction of DNA damage in cholangiocarcinoma cells by altholactone and its halogenated benzoate derivatives.

Topoisomerase IIα enzyme (Topo IIα) plays a critical function in DNA replication process and is considered to be a promising target of anti-cancer drugs. In the present study, we reported that the altholactone derivatives modified by adding a halogenated benzoate group showed greater inhibitory activity on Topo IIα enzyme in cell-free system concomitant with cytotoxicity against the CCA cell lines (KKU-M055 and KKU-M213) than those of the parent altholactone. However, the cytotoxic activities of four halogenated benzoate altholactone derivatives including iodo-, fluoro-, chloro-, and bromobenzoate derivatives (compound 1, 2, 3, and 4, respectively) were of equal potency. The fluorobenzoate derivative (compound 2) was chosen for investigating the underlying mechanism in CCA cells. Compound 2 arrested CCA cell cycle at sub G1 phase and induced apoptotic cell death. It markedly inhibited Topo IIα protein expression in both KKU-M055 and KKU-M213 cells, which was accompanied by DNA double-strand breaks demonstrated by an increase in phosphorylated H2A.X protein. Interestingly, KKU-M055 cells, which express higher Topo IIα mRNA compared to KKU-M213 cells, showed greater sensitivity to the compound, indicating the selectivity of the compound to Topo IIα enzyme. By computational docking analysis, the binding affinity of altholactone (-52.5 kcal/mol) and compound 2 (-56.7 kcal/mol) were similar to that of the Topo II poison salvicine (-53.7 kcal/mol). The aromatic moiety of both altholactones embedded in a hydrophobic pocket of Topo II ATPase domain. In addition, compound 2 induced the formation of linear DNA in Topo II-mediated cleavage assay. Collectively, our results demonstrate that the addition of fluorobenzoyl group to altholactone enhances potency and selectivity to inhibit type IIα topoisomerases. Atholactone and fluorobenzoate derivative act as Topo II cleavage complexes stabilizing compounds or Topo II poisons preferentially through binding at ATPase domain of Topo IIα, leading to DNA double-strand breaks and apoptosis induction. Such activity of 3-fluorobenzoate derivative of altholactone should be further explored for the development of an anti-cancer drug for CCA.

Pyranonaphthoquinone and anthraquinone derivatives from Ventilago harmandiana and their potent anti-inflammatory activity.

The chemical study on the heartwoods extract of Ventilago harmandiana (Rhamnaceae) resulted in the isolation of ten previously undescribed pyranonaphthoquinones (ventilanones A-J), an undescribed anthraquinone (ventilanone K), together with eight known anthraquinone derivatives. Their structures were elucidated by extensive analysis of their spectroscopic data. The absolute configuration of ventilanone A was established from single crystal X-ray crystallographic analysis of its p-bromobenzenesulfonate ester derivative using Cu Kα radiation. The absolute configurations of the other related compounds were identified by comparison of their ECD data with those of ventilanone A and related known compounds. Cytotoxic and anti-inflammatory activities of some of the isolated compounds were evaluated. Ventilanone A and ventilanone C exhibited moderate cytotoxicity against P-388 cell line. Ventilanone D exhibited significant anti-inflammatory activity while ventilanone A and ventilanone C showed moderate activity.

Cytotoxic polyoxygenated cyclohexene derivatives from the aerial parts of Uvaria cherrevensis.

Three previously undescribed polyoxygenated cyclohexene derivatives named cherrevenol M (1), cherrevenol N (2), and cherrevenone (3), together with nine related known analogues 4-12 were isolated from the ethyl acetate fraction partitioned from the methanol extract of the aerial parts of Uvaria cherrevensis (Annonaceae). The determination of the structures and their relative configurations of the isolated compounds were established by spectroscopic techniques, electronic circular dichroism (ECD) analysis as well as comparison with the literature data. For cherrevenone (3), the relative and absolute configurations were also confirmed by using X-ray diffraction and ECD techniques, respectively. Compounds isolated except for compounds 8 and 10 were evaluated for their cytotoxic activity and cherrevenone (3) showed moderate cytotoxic activity against all cancerous cell lines except for ASK cell line with ED50 values ranging from 1.04 ±â€¯0.13 to 10.09 ±â€¯4.31 µM.

The anti-cancer activity of an andrographolide analogue functions through a GSK-3ß-independent Wnt/ß-catenin signaling pathway in colorectal cancer cells.

The Wnt/ß-catenin signaling pathway plays a key role in the progression of human colorectal cancers (CRCs) and is one of the leading targets of chemotherapy agents developed for CRC. The present study aimed to investigate the anti-cancer effects and molecular mechanisms of 19-O-triphenylmethyl andrographolide (RS-PP-050), an andrographolide analogue and determine its activity in the Wnt/ß-catenin pathway. RS-PP-050 was found to potently inhibit the proliferation and survival of HT-29 CRC cells. It induces cell cycle arrest and promotes apoptotic cell death which was associated with the activation of PARP-1 and p53. Furthermore, RS-PP-050 exerts inhibitory effects on ß-catenin transcription by suppressing T-cell factor/lymphocyte enhancer factor (TCF/LEF) activity in cells overexpressing ß-catenin and by down-regulating the endogenous expression of Wnt target genes. RS-PP-050 also decreased the protein expression of the active form of ß-catenin but functions independently of GSK-3ß, a negative regulator of Wnt. Interestingly, RS-PP-050 extensively blocks phosphorylation at Ser675 of ß-catenin which links to interference of the nuclear translocation of ß-catenin and might contribute to Wnt inactivation. Collectively, our findings reveal the underlying anti-cancer mechanism of an andrographolide analogue and provide useful insight for exploiting a newly chemotherapeutic agent in Wnt/ß-catenin-overexpressing CRC cells.

Synthetic analogues of durantoside I from Citharexylum spinosum L. and their cytotoxic activity.

New iridoid glycoside derivatives from durantoside I, the latter from the dried flowers and leaves of Citharexylum spinosum, were synthesized by variously modifying a sugar moiety by silylation or acetylation and/or removal of cinnamate group at C-7 position and subsequent screening for comparative cytotoxicity against several cancer cell lines. Addition of alkylsilane to durantoside I and removal of cinnamate group were most effective in improving cytotoxicity.