Lappaol F regulates the cell cycle by activating CDKN1C/p57 in human colorectal cancer cells.

CONTEXT: Lappaol F (LAF), a natural lignan from Arctium lappa Linné (Asteraceae), inhibits tumor cell growth in vitro and in vivo. The underlying mechanism involves the suppression of the Yes-associated protein. However, the specific role of LAF in cell cycle regulation remains unknown. OBJECTIVE: This study determined the molecular mechanism by which LAF regulates cell cycle progression. MATERIALS AND METHODS: Various colon cancer cell lines (SW480, HCT15, and HCT116) were treated with LAF (25, 50, and 75 µmol/L) for 48 h. The effects of LAF on cell proliferation and cell cycle were determined using sulforhodamine B and flow cytometry assays. Differentially expressed proteins (DEPs) were identified using quantitative proteomics. Bioinformatic analysis of DEPs was conducted via Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. Expression levels of DEPs in the cell cycle pathway were analyzed using RT-qPCR and western blotting. RESULTS: LAF suppressed the proliferation of SW480, HCT15, and HCT116 cells (IC50 47.1, 51.4, and 32.8 µmol/L, respectively) and induced cell cycle arrest at the S phase. A total of 6331 proteins were identified and quantified, of which 127 were differentially expressed between the LAF-treated and untreated groups. GO and KEGG enrichment analyses revealed that DEPs mainly participated in the cell cycle. CDKN1C/p57 showed the most significant differential expression, with the highest fold-change (3.155-fold). Knockdown of CDKN1C/p57 attenuated the S phase cell cycle arrest and proliferation inhibition induced by LAF. CONCLUSION: LAF exerts antitumor effects via S phase arrest by activating CDKN1C/p57 in colorectal cancer cells.

Lappaol F, an anticancer agent, inhibits YAP via transcriptional and post-translational regulation.

CONTEXT: Lappaol F (LAF), a natural lignan from Arctium lappa Linné (Asteraceae), inhibits tumour cell growth by inducing cell cycle arrest. However, its underlying anticancer mechanism remains unclear. OBJECTIVE: The effects of LAF on the Hippo-Yes-associated protein (YAP) signalling pathway, which plays an important role in cancer progression, were explored in this study. MATERIALS AND METHODS: Cervical (HeLa), colorectal (SW480), breast (MDA-MB-231) and prostate (PC3) cancer cell lines were treated with LAF at different concentrations and different durations. BALB/c nude mice bearing colon xenografts were intravenously injected with vehicle, LAF (10 or 20 mg/kg) or paclitaxel (10 mg/kg) for 15 days. The expression and nuclear localisation of YAP were analysed using transcriptome sequencing, quantitative PCR, western blotting and immunofluorescence. RESULTS: LAF suppressed the proliferation of HeLa, MDA-MB-231, SW480 and PC3 cells (IC50 values of 41.5, 26.0, 45.3 and 42.9 µmol/L, respectively, at 72 h), and this was accompanied by significant downregulation in the expression of YAP and its downstream target genes at both the mRNA and protein levels. The expression of 14-3-3σ, a protein that causes YAP cytoplasmic retention and degradation, was remarkably increased, resulting in a decrease in YAP nuclear localisation. Knockdown of 14-3-3σ with small interfering RNA partially blocked LAF-induced YAP inhibition and anti-proliferation effects. In colon xenografts, treatment with LAF led to reduced YAP expression, increased tumour cell apoptosis and tumour growth inhibition. CONCLUSION: LAF was shown to be an inhibitor of YAP. It exerts anticancer activity by inhibiting YAP at the transcriptional and post-translational levels.

Cajanolactone A from Cajanus cajan Promoted Osteoblast Differentiation in Human Bone Marrow Mesenchymal Stem Cells via Stimulating Wnt/LRP5/ß-Catenin Signaling.

Cajanolactone A (CLA) is a stilbenoid discovered by us from Cajanus cajan (L.) Millsp. In our study, CLA was found to promote osteoblast differentiation in human bone marrow mesenchymal stem cells (hBMSCs), as judged by increased cellular alkaline phosphatase activity and extracellular calcium deposits, and elevated protein expression of Runx2, collagen-1, bone morphogenetic protein-2, and osteopontin. Mechanistic studies revealed that hBMSCs undergoing osteoblast differentiation expressed upregulated mRNA levels of Wnt3a, Wnt10b, LRP5/6, Frizzled 4, ß-catenin, Runx2, and Osterix from the early stage of differentiation, indicating the role of activated Wnt/ß-catenin signaling pathway in osteoblast differentiation. Addition of CLA to the differentiation medium further increased the mRNA level of Wnt3a, Wnt10b, Frizzled 4, LRP5, and ß-catenin, inferring that CLA worked by stimulating Wnt/LRP5/ß-catenin signaling. Wnt inhibitor dickkopf-1 antagonized CLA-promoted osteoblastogenesis, indicating that CLA did not target the downstream of canonical Wnt signaling pathway. Treatment with CLA caused no changes in mRNA expression level, as well as protein secretion of osteoprotegerin (OPG) and receptor activator of nuclear factor kappa-B ligand (RANKL), indicating that CLA did not affect the OPG/RANKL axis. Our results showed that CLA, which promoted osteoblast differentiation in hBMSCs, through activating Wnt/LRP5/ß-catenin signaling transduction, is a promising anti-osteoporotic drug candidate.

Acute and sub-chronic toxicity of Cajanus cajan leaf extracts.

CONTEXT: The leaves of Cajanus cajan (L.) Millsp. (Fabaceae) have diverse bioactivities, but little safety data are reported. OBJECTIVE: This study examines the toxicological profiles of C. cajan leaf extracts. MATERIALS AND METHODS: The leaves were extracted by water or 90% ethanol to obtain water or ethanol extract (WEC or EEC). EEC was suspended in water and successively fractionated into dichloroform and n-butanol extracts (DEC and BEC). Marker compounds of the extracts were monitored by high-performance liquid chromatography (HPLC). Kunming mice were administered with a single maximum acceptable oral dose (15.0 g/kg for WEC, EEC and BEC and 11.3 g/kg for DEC) to determine death rate or maximal tolerated doses (MTDs). In sub-chronic toxicity investigation, Sprague-Dawley rats were orally given WEC or EEC at 1.5, 3.0 or 6.0 g/kg doses for four weeks and observed for two weeks after dosing to determine toxicological symptoms, histopathology, biochemistry and haematology. RESULTS: Flavonoids and stilbenes in the extracts were assayed. In acute toxicity test, no mortality and noted alterations in weight and behavioural abnormality were observed, and the maximum oral doses were estimated as MTDs. In sub-chronic toxicity study, no mortality and significant variances in haematological and biochemical parameters or organ histopathology were observed, but increased kidney weight in 3.0 g/kg WEC- or 3.0 and 6.0 g/kg EEC-treated female rats, and reduced testes and epididymis weight in EEC-treated male rats were recorded. These changes returned to the level of control after recovery period. CONCLUSION: Acute and sub-chronic toxicity of Cajanus cajan leaf extracts was not observed.

[Chemical Constituents from Periploca forrestii].

Objective: To study the chemical constituents from Periploca forrestii. Methods: The constituents were separated by column chromatography and their structures were elucidated by spectroscopic methods. Results: Seven compounds were isolated from Periploca forrestii and identified as wogonin( 1),negletein( 2),vanilline( 3),isovanilline( 4),periplocoside L( 5),ß-sitosterol( 6) and ß-daucosterol( 7). Conclusion: Compounds 1 and 2 are obtained from this genus for the first time,and compounds 3 ~ 5 are isolated from this plant for the first time.

A Halogen-Containing Stilbene Derivative from the Leaves of Cajanus cajan that Induces Osteogenic Differentiation of Human Mesenchymal Stem Cells.

A new natural halogen-containing stilbene derivative was isolated from the leaves of Cajanus cajan (L.) Millsp. and identified as 3-O-(3-chloro-2-hydroxyl-propanyl)-longistylin A by comprehensive spectroscopic and chemical analysis, and named cajanstilbene H (1). It is the first halogen-containing stilbene derivative found from plants. In human mesenchymal stem cells (hMSC) from bone marrow, 1 did not promote cell proliferation, but distinctly enhanced osteogenic differentiation of hMSC in time- and dose-dependent manners. In six human cancer cell lines, 1 showed a moderate inhibitory effect on cell proliferation, with IC50 values of 21.42-25.85 µmol·L(-1).

Total aglycones from Marsdenia tenacissima increases antitumor efficacy of paclitaxel in nude mice.

Marsdeniae tenacissimae Caulis (MTC) is a Chinese herbal medicine used mainly for treatment of cancer, whose pharmacologically active constituents responsible for its in vivo activity and clinical efficacy have not been clearly elucidated. In this study, total aglycones of MTC (ETA) showed the ability to sensitize KB-3-1, HeLa, HepG2 and K562 cells to paclitaxel treatment. More inspiringly, ETA markedly enhanced the antitumor activity of paclitaxel in nude mice bearing HeLa or KB-3-1 xenografts. Compared to treatment with paclitaxel alone, treatment with combination of paclitaxel and ETA achieved significant reduction in volume and weight of HeLa tumors (p<0.05), and remarkable inhibition to the growth of KB-3-1 tumors (p<10⁻6). ETA was characterized by the presence of a group of tenacigenin B ester derivatives, among which four reference compounds, 11α-O-tigloyl-12ß-O-acetyltenacigenin B, 11α,12ß-di-O-tigloyltenacigenin B, 11α-O-2-methylbutanoyl-12ß-O-tigloyltenacigenin B, and 11α-O-(2-methylbutanoyl)-12ß-O-benzoyltenacigenin B, accounted for 42.14% of the total peak area of 19 detectable components assayed by HPLC. Our study has identified ETA as a promising sensitizer for cancer chemotherapy.

Aikeqing, a kidney- and spleen-tonifying compound Chinese medicine granule, prevented ovariectomy-induced bone loss in rats via the suppression of osteoclastogenesis.

Postmenopausal women are prone to osteoporosis due to increased osteoclast activation and bone resorption caused by oestrogen deficiency. In Traditional Chinese Medicine theory, medicines with spleen- and kidney-nourishing effects are commonly used in postmenopausal osteoporosis (PMOP) treatment. Aikeqing (AKQ) is a compound Chinese medicinal granule with spleen- and kidney-nourishing effects. Herein, we investigate the in vitro and in vivo anti-osteoporotic effects of AKQ, its underlying mechanisms and pharmacodynamic basis. In vitro antiosteoporotic effects of AKQ were assessed by its ability to promote osteoblastogenesis in MC3T3-E1 and/or inhibit RANKL-induced osteoclastogenesis in murine bone marrow monocytes (BMMs). The protective effect of AKQ on bone loss induced by oestrogen deficiency was evaluated in ovariectomized rats. The underlying mechanisms were studied in BMMs by detecting the effects of AKQ on the RANKL-induced expression of genes and proteins involved in the regulation of osteoclastogenesis. The main chemical constituents of AKQ in the granule were analyzed by UPLC-QTOF-MS. Our findings show that AKQ did not affect osteoblastogenesis, but it inhibited RANKL-induced osteoclastogenesis. In the ovariectomized rats, oral administration of AKQ (4 g/kg/d) for 90 d effectively prevented oestrogen deficiency-induced bone loss. Mechanistic studies in BMMs revealed that AKQ inhibited RNAKL-induced activation of NF-κB (p65) and MAPKs (p38 and JNK) via blocking the RANK-TRAF6 interaction, subsequently suppressing the translocation and expression of NFATc1 and c-Fos. UPLC-QTOF-MS analysis quantified the 123 main components of AKQ. Taken together, AKQ was demonstrated for the first time as a novel alternative therapy for osteoclast-associated bone diseases.

Chemical constituents from the Streptomyces morookaensis strain Sm4-1986.

Three new compounds, including 6-methoxy-3,4,5,7-tetramethylisochromane-3,8-diol (1), 3,4,5,7-tetramethylisochromane-3,6,8-triol (2), streptimidone derivative (3), along with ten known compounds (4-13) were isolated from the Streptomyces morookaensis strain Sm4-1986. Their chemical structures were established based on the information from UV, IR, NMR (1H NMR, 13C NMR, 1H-1H COSY, HSQC, HMBC, NOESY), and mass spectroscopic. Moreover, all the isolated new compounds were evaluated for antibacterial activities (S. aureus, B. cereus, S. epidermids and methicillin-resistant S. aureus) and their cytotoxicities against MCF-7, A549, Hela tumor cell lines and Marc-145 normal cell line.

An ester derivative of tenacigenin B from Marsdenia tenacissima (Roxb.) Wight et Arn reversed paclitaxel-induced MDR in vitro and in vivo by inhibiting both P-gp and MRP2.

ETHNOPHARMACOLOGICAL RELEVANCE: Marsdenia tenacissima is a medicinal plant, used as a raw material for cancer treatment in China. In our previous studies, 11α-O-2-methylbutanoyl-12ß-O-tigloyl-tenacigenin B (MT2), the main steroid aglycone isolated from M. tenacissima, was found to significantly enhance the antitumor activity of paclitaxel (PTX) in vivo. However, it is unclear whether MT2 reverses multidrug resistance (MDR) in tumors. AIM OF THE STUDY: To determine the role and mechanism of MT2 in reversing tumor MDR. MATERIALS AND METHODS: MDR cell line HeLa/Tax was established from the human cervical carcinoma cell line HeLa by long-term exposure to subtoxic concentrations of PTX and was used to evaluate the ability of MT2 to restore chemosensitivity of cells both in vitro and in a nude mouse model. The expression of P-glycoprotein (P-gp) and multidrug resistance-associated protein 2 (MRP2) was determined using western blotting and immunohistochemistry. The substrate transport function was assessed using an MDR function assay kit. The binding modes of MT2 and P-gp were determined using the conformation-sensitive anti-P-gp antibodies. The permeability and transport properties of MT2 were analyzed in Caco-2 cell monolayers. RESULTS: Compared to parental cells, HeLa/Tax cells overexpress P-gp and MRP2 and are approximately 100-360 fold more resistant to the anticancer drugs PTX, docetaxel, and vinblastine. MT2 at 5 or 10 µmol/L significantly increased the sensitivity of HeLa/Tax to these three anticancer drugs (18-56-fold decrease in IC50 value) and suppressed the expression of P-gp and MRP2. Knockdown of P-gp with small interfering RNA partially reversed MT2-induced sensitivity to PTX in HeLa/Tax cells. Moreover, MT2 directly inhibited P-gp-mediated substrate transport while interacting with membrane P-gp in non-substrate ways. MT2 was highly permeable and could not be transported in the Caco-2 cell monolayers. In nude mice bearing HeLa/Tax xenografts, the combination treatment with MT2 and PTX exerted a synergistic inhibitory effect on the growth of tumors and the expression of P-gp and MRP2 without increasing toxicity. CONCLUSION: MT2 is a potential agent for reversing MDR. It impedes membrane drug efflux pumps by suppressing P-gp and MRP2 expression, and directly inhibiting the transport function of P-gp.

Two new triterpenoids from Lysimachia heterogenea Klatt and evaluation of their cytotoxicity.

Two new 13,28-epoxy oleanane-type triterpenoids, namely heterogenoside E and F, were isolated from Lysimachia heterogenea Klatt, together with the eight known compounds: palmitic acid, ß-stigmasterol, kaempferol, quercetin, hyperin, isorhamnetin, isorhamnetin-3-O-galactopyranoside and anagallisin C. Heterogenoside F possesses acetoxyl groups at the unusual C-21 and C-22 positions of its oleanane skeleton. The cytotoxic activities of anagallisin C, heterogenoside E and F were weak.

Pharmacokinetics, plasma protein binding, and metabolism of a potential natural chemosensitizer from Marsdenia tenacissima in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Marsdenia tenacissima (Roxb.) Wight et Arn is a medicinal plant mainly distributed in southwest China. It is used in folk medicine for the treatment of tumors and is synergistic with chemotherapies. In our previous study, 11α-O-2-methybutyryl-12ß-O-tigloyl-tenacigenin B (MT2), a main steroid aglycone isolated from the total aglycones of M. tenacissima, significantly enhanced the in vivo antitumor effect of paclitaxel in mice bearing human tumor xenografts, showing its potential as a chemosensitizer. However, the pharmacokinetic characteristics, plasma protein binding rate, and metabolic profile of MT2 remain unclear. AIM OF THE STUDY: To elucidate the pharmacokinetic characteristics, plasma protein binding rate, and metabolic profile of MT2 in rats. MATERIALS AND METHODS: MT2 in rat plasma and phosphate-buffered saline was quantified using ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method, while the MT2 metabolites in rat liver microsomes were analyzed using UPLC-triple time-of-flight MS/MS. RESULTS: For intravenously administered MT2, the maximum plasma concentration and the area under the plasma concentration-time curve indicated dose dependency, while the elimination half-life time, the mean residence time, apparent volume of distribution and total apparent clearance values remained relatively unchanged in both the 5 mg/kg and 10 mg/kg groups. For orally administered MT2, the bioavailability was 1.08-1.11%. In rat plasma, MT2 exhibited a protein binding rate of 93.84-94.96%. In rat liver microsomes, MT2 was metabolized by oxidation alone or in combination with demethylation, and five MT2 metabolites were identified. CONCLUSION: MT2 has low oral bioavailability and a high plasma protein binding rate in rats. After administration, MT2 is transformed into oxidative metabolites in the liver. To achieve a high blood concentration of MT2, it should be administered intravenously. These findings would serve as a reference for further MT2-based pharmacological study and drug development.

Altered gene expression in glycolysis-cholesterol synthesis axis correlates with outcome of triple-negative breast cancer.

Identification of molecular subtypes of clinically resectable triple-negative breast cancer (TNBC) is of great importance to achieve better clinical outcomes. Inter- and intratumor metabolic heterogeneity improves cancer survival, and the interaction of various metabolic pathways may affect treatment outcome of TNBC. We speculated that TNBC can be categorized into prognostic metabolic subtype according to the expression changes of glycolysis and cholesterol synthesis. The genome, transcriptome, and clinical data were downloaded from the Cancer Genome Atlas and Molecular Taxonomy of Breast Cancer International Consortium and subsequently analyzed by integrated bioinformatics methods. Four subtypes, namely, glycolytic, cholesterogenic, quiescent, and mixed, were classified according to the normalized median expressions of the genes involved in glycolysis and cholesterol synthesis. In the four subtypes, the cholesterogenic type was correlated with the shortest median survival (log rank P = 0.044), while patients with high-expressed glycolytic genes tended to have a longer survival. Tumors with PIK3CA amplification and dynein axonemal heavy chain 2 deletion exhibited higher expressions of cholesterogenic genes than other mutant oncogenes. The expressions of mitochondrial pyruvate carrier MPC1 and MPC2 were the lowest in quiescent tumor, and MPC2 expression was higher in cholesterogenic tumor compared with glycolytic or quiescent tumor (t-test P < 0.001). Glycolytic and cholesterogenic gene expressions were related to the expressions of prognostic genes in some other types of cancers. Classification of glycolytic and cholesterogenic pathways according to metabolic characteristics provides a new understanding to previously identified subtypes of TNBC and could improve personalized treatments based on tumor metabolic profiles.

Cajanolactone A, a stilbenoid from Cajanus cajan, inhibits energy intake and lipid synthesis/storage, and promotes energy expenditure in ovariectomized mice.

BACKGROUND: We had reported that cajanolactone A (CLA) from Cajanus cajan dose-dependently inhibited ovariectomy-induced obesity and liver steatosis in mice, showing potential to prevent postmenopausal obesity and fatty liver. In this study, the role of CLA in the regulation of energy and lipid homeostasis was investigated. METHODS: Ovariectomized mice treated with CLA or vehicle for 12 weeks were performed a 48 h monitoring for energy metabolism and food uptake. After that, hypothalami, perigonadal (pWATs), inguinal (iWATs) and brown (BATs) adipose tissues, livers, sera, and fecal and cecal contents were collected and analyzed. FINDINGS: In CLA-treated mice, we observed reduced food uptake; increased energy expenditure; inhibited expression of orexigenic genes (ORX, ORXR2, pMCH and Gal) in the hypothalami, of lipogenic genes (CD36, SREBP-1c, ChREBP, PPARγ) in the livers, and of lipid storage proteins in the WATs (FSP27, MEST and caveolin-1) and livers (FSP27, Plin2 and Plin5); stimulated expression of metabolism-related proteins (pATGL and Echs1) in the adipose tissues and of thermogenic protein (UCP1) in the inguinal WATs; increased BAT content; increased mitochondria in the pWATs and livers; inhibited angiogenesis in the pWATs; and altered gut microbiome diversity with an increased abundance of Bacteroides. INTERPRETATION: CLA prevents ovariectomy-induced obesity and liver steatosis via regulating energy intake and lipid synthesis/storage, promoting UCP1-dependent heat production, and protecting the mitochondrial function of hepatocytes and adipocytes. The improved gut microecology and inhibited angiogenesis may also contribute to the anti-obese activity of CLA.

7-methoxyflavanone alleviates neuroinflammation in lipopolysaccharide-stimulated microglial cells by inhibiting TLR4/MyD88/MAPK signalling and activating the Nrf2/NQO-1 pathway.

OBJECTIVES: Neuroprotective potential of 7-methoxyflavanone (7MF) and its underlying mechanism was investigated. METHODS: Inhibitory effects of 7MF on microglial activation and neuroinflammation were evaluated by employment of lipopolysaccharide (LPS)-induced BV2 microglial cells. Changes in expression of genes and proteins of interest were investigated by RT-qPCR analysis and Western blot analysis. Inhibitory effects of 7MF on microglial overactivation were verified in LPS-treated C57BL/6J mice using ionized calcium-binding adaptor molecule-1 (Iba1) in the brain and interleukin-6 (IL-6) in serum as indicators. KEY FINDINGS: In BV2 cells, pretreatment with 7MF antagonized LPS-induced production of inflammatory factors IL-6, tumour necrosis factor-α (TNF-α), cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), intercellular adhesion molecule-1 (ICAM-1) and monocyte chemoattractant protein-1 (MCP-1). Mechanistic studies revealed reduced expression of Toll-like receptor 4 (TLR4), myeloid differentiation factor-88 (MyD88), phosphorylated forms of c-Jun N-terminal kinase (p-JNK) and extracellular signal-regulated kinases 1/2 (p-ERK) but increased nuclear accumulation of nuclear factor erythroid 2-related factor 2 (Nrf2) and cellular expression of NAD(P)H quinone dehydrogenase-1 (NQO-1) by 7MF. In LPS-treated mice, pretreatment with 7MF reduced the brain level of Iba1 and serum level of IL-6. CONCLUSIONS: 7-methoxyflavanone inhibited LPS-stimulated TLR4/MyD88/MAPK signalling and activated Nrf2-mediated transcription of antioxidant protein NQO-1, showing antineuroinflammatory effect, so it is a potential neuroprotective agent.

Network pharmacology-based investigation on the mechanisms of action of Morinda officinalis How. in the treatment of osteoporosis.

BACKGROUND: Osteoporosis is a systemic skeletal disease that leads to a high risk for bone fractures. Morinda officinalis How. has been used as osteoporosis treatment in China. However, its mechanism of action as an anti-osteoporotic herb remains unknown. METHODS: A network pharmacology approach was applied to explore the potential mechanisms of action of M. officinalis in osteoporosis treatment. The active compounds of M. officinalis and their potential osteoporosis-related targets were retrieved from TCMSP, TCMID, SwissTargetPrediction, DrugBank, DisGeNET, GeneCards, OMIM, and TTD databases. A protein-protein interaction network was built to analyze the target interactions. The Metascape database was used to carry out GO enrichment analysis and KEGG pathway analysis. Moreover, interactions between active compounds and potential targets were investigated through molecular docking. RESULTS: A total of 17 active compounds and 93 anti-osteoporosis targets of M. officinalis were selected for analysis. The GO enrichment analysis results indicated that the anti-osteoporosis targets of M. officinalis mainly play a role in the response to steroid hormone. The KEGG pathway enrichment analysis showed that M. officinalis prevents osteoporosis through the ovarian steroidogenesis signaling pathway. Moreover, the molecular docking results indicated that bioactive compounds (morindon, ohioensin A, and physcion) demonstrated a good binding ability with IGF1R, INSR, ESR1, and MMP9. CONCLUSION: M. officinalis contains potential anti-osteoporotic active compounds. These compounds function by regulating the proteins implicated in ovarian steroidogenesis-related pathways that are crucial in estrogen biosynthesis. Our study provides new insights into the development of a natural therapy for the prevention and treatment of osteoporosis.

Cajanolactone A, a stilbenoid from cajanus cajan, prevents ovariectomy-induced obesity and liver steatosis in mice fed a regular diet.

BACKGROUND: Visceral obesity and fatty liver are prevalent in postmenopausal women. The stilbene-rich extract of Cajanus cajan (L.) Millsp. has been reported to prevent ovariectomy-induced and diet-induced weight gain in animal models, and stilbenoids from C. cajan are thought to have the potential to prevent postmenopausal obesity and fatty liver. PURPOSE: Cajanolactone A (CLA) is the main stilbenoid from C. cajan with osteoblastogenic promoting activity. This study investigated the potential of CLA to prevent postmenopausal obesity and fatty liver. Underlying mechanisms were also investigated. METHOD: Ovariectomized C57BL/6 mice fed a regular diet were used as mimics of postmenopausal women and given 10, 20, or 40 mg/kg/d of CLA, 0.1 mg/kg/d of estradiol valerate (EV, positive control), or vehicle (OVX) orally for 16 weeks. Mice of the same age subjected to a sham operation were used as control (Sham). Body weights were recorded every 2 weeks for 16 weeks. Body compositions were analyzed via micro-CT. Serum levels of lipids, adipocytokines and aminotransferases were measured using the relevant kits. mRNA levels of genes of interest were detected by RT-qPCR. Proteomic study of perigonadal white adipose tissue (pWAT) was performed using tandem-mass-tags-based proteomic technology combined with Parallel-Reaction-Monitoring (PRM) validation. RESULTS: CLA showed potential equivalent to that of EV to prevent ovariectomy-induced overweight, obesity, dyslipidemia, liver steatosis and liver dysfunction, but did not prevent uterine atrophy. In the liver, CLA significantly inhibited ovariectomy-induced upregulation in expression of lipogenic genes SREBP-1c and ChREBP, and stimulated the mRNA expression of apolipoprotein B gene ApoB. In pWAT, CLA reversed, or partially reversed ovariectomy-induced downregulation in the expression of a number of metabolism- and mitochondrial-function-related proteins, including Ndufa3, Pcx, Pdhb, Acly, Acaca, Aldh2, Aacs and Echs1. In addition, ovariectomy-inhibited mRNA expression of Pdhb, Aacs, Acsm5, Echs1, and Aldh2 genes in pWAT was also reversed. CONCLUSION: CLA was demonstrated to be a potential non-estrogen-like drug candidate for prevention of postmenopausal obesity and fatty liver. The underlying mechanism might involve the inhibition of lipogenesis and promotion of triglycerides output in the liver, and the promotion of metabolism and mitochondrial functions of visceral white adipose tissue.

Tenacigenin B ester derivatives from Marsdenia tenacissima actively inhibited CYP3A4 and enhanced in vivo antitumor activity of paclitaxel.

ETHNOPHARMACOLOGICAL RELEVANCE: Marsdenia tenacissima (Roxb.) Wight et Arn is a vine distributed in southwest area of China and used in folk medicine for treatment of tumors. Recent decades of studies on this plant reveal its synergistic effects with certain anticancer drugs in cancer therapy. In our previous study, an extract ETA which contains total aglycones made from M. tenacissima significantly enhanced antitumor activity of paclitaxel in tumor-bearing mice. However, the effective constituents in ETA and the underlying mechanisms remain unclear. AIM OF THE STUDY: Reveal the active components in ETA as well as the molecular mechanism in enhancing antitumor efficacy of paclitaxel. MATERIAL AND METHODS: Main constituents in ETA were purified by chemical methods. Effects of the purified constituents on metabolic activity of CYP450 enzymes were evaluated in human liver microsomes. Ability of the constituents to enhance antitumor activity of paclitaxel were investigated in nude mice bearing HeLa tumors. Pharmacokinetic study was performed in SD rats. Molecular docking was carried out for investigation of drug-protein interactions. RESULTS: Three main C21 steroidal aglycones, 11α-O-tigloyl-12ß-O-acetyl-tenacigenin B (MT1), 11α-O-2-methylbutanoyl-12ß-O-tigloyl-tenacigenin B (MT2) and 11α-O-2-methylbutanoyl-12ß-O-acetyl-tenacigenin B (MT3), together with tenacigenin B (MT4) was prepared from ETA. Among them, MT1, MT2 and MT3 strongly inhibit the metabolic activity of CYP3A4. MT2 also showed inhibitory effects on CYP2C8, CYP2B6 and CYP2C19. In HeLa tumor xenografts, MT1, MT2 and MT3 (30 mg/kg) did not affect tumor growth themselves, but significantly enhanced paclitaxel-induced growth inhibition. In addition, coadministration of MT2 with paclitaxel resulted in significant reduction of liver CYP2C8. In pharmacokinetic study, MT2 significantly increased the blood concentration of paclitaxel with increased AUC value by 2.2-5.3 folds. Molecular docking analysis suggested hydrophobic interaction modes of tenacigenin B derivatives with CYP3A4, and also the essential roles of the C-11 and C-12 ester groups for effective interaction with CYP3A4. CONCLUSION: Our study proves that, 11α-O-tigloyl-12ß-O-acetyl-tenacigenin B, 11α-O-2-methylbutanoyl-12ß-O-tigloyl-tenacigenin B and 11α-O-2-methylbutanoyl-12ß-O-acetyl-tenacigenin B, which are the main constituents of ETA, are active inhibitors of CYP3A4 with potential to increase therapeutic efficacy of anticancer drugs that are substrates of CYP3A4. Tenacigenin B derivatives with C-11 and C-12 ester group substitutions, or at least a large part of them, are active components in ETA and M. tenacissima to enhance in vivo antitumor efficacies of paclitaxel.

Prenylated stilbenes and flavonoids from the leaves of Cajanus cajan.

Three new prenylated stilbenes, named as cajanusins A-C (1-3), and one new natural product cajanusin D (4), along with six known derivatives (5-10) were isolated from the leaves of Cajanus cajan. Their structures were fully elucidated by means of extensive spectroscopic methods and comparison with data in the reported literatures. The new compounds of 1 and 2 were evaluated for in vitro cytotoxic activities against a panel of human cancer cell lines.

Methoxylation of 3',4'-aromatic side chains improves P-glycoprotein inhibitory and multidrug resistance reversal activities of 7,8-pyranocoumarin against cancer cells.

The overexpression of P-glycoprotein (Pgp), an ATP-driven membrane exporter of hydrophobic xenobiotics, is one of the major causes of multidrug resistance (MDR) in cancer cells. Through extensive screening we have found that the extracts of Peucedanum praeruptorum Dunn. and one of the major components (+/-)-praeruptorin A (PA) may reverse Pgp-mediated multidrug resistance. Studies on novel PA derivatives have shown that (+/-)-3'-O,4'-O-dicinnamoyl-cis-khellactone (DCK) is more active than PA or verapamil and is a non-competitive inhibitor of Pgp. Here, we report that methoxylation of the cinnamoyl groups on DCK may further enhance its bioactivity. The structure-activity relationship is demonstrated by comparing two new pyranocoumarins (+/-)-3'-O,4'-O-bis(3,4-dimethoxycinnamoyl)-cis-khellactone (DMDCK) and (+/-)-3'-O,4'-O-bis(4-methoxycinnamoyl)-cis-khellactone (MMDCK). While the co-existence of 3- and 4-methoxy groups on cinnamoyl remarkably enhanced the Pgp-inhibitory activity, the lone existence of the 4-methoxy group on cinnamoyl reduced the activity. Contrary to DCK, DMDCK promoted the binding of UIC2 antibody to Pgp which signifies a conformational change of Pgp similar to that induced by transport substrates. While DCK moderately stimulated the basal Pgp-ATPase activity, DMDCK inhibited the activity. A pharmacophore search with verapamil-based template revealed that four functional groups of DMDCK could be simultaneously involved in interaction with Pgp whereas for DCK or MMDCK only three groups were involved. It is speculated that the additional 3-methoxy group on cinnamoyl allows DMDCK to interact more efficiently with Pgp substrate site(s). If DMDCK was tightly bind to Pgp substrate site(s) the complexes could be inactive with regard to transportation and ATP hydrolysis could also be inhibited.