Cytotoxic activities of alkaloid constituents from the climbing stems and rhizomes of Sinomenium acutum against cancer stem cells.

From the methanolic extract of the climbing stems and rhizomes of Sinomenium acutum, two new aporphine analogues, acutumalkaloids I and II, were isolated together with fifteen known compounds including lysicamine. The chemical structures of the isolated new compounds were elucidated based on chemical/physicochemical evidence such as NMR and MS spectra. For acutumalkaloids I and II, the absolute configurations were established by comparison of experimental and predicted electronic circular dichroism (ECD) data. We compared anti-proliferative activities of isolated compounds with reported naturally occurring Wnt/ß-catenin pathway inhibitor, nuciferine. Among the isolated compounds, we found lysicamine have anti-proliferative activity against both of HT-29 human colon cancer cell line and its cancer stem cells (CSCs). The IC50 values of lysicamine against non-CSCs and its CSCs were lower than that of nuciferine. In addition, the results of western blotting analysis suggested that lysicamine inhibited the expression of Wnt/ß-catenin pathway target protein such as survivin. These results suggested that lysicamine show cytotoxic activity via inhibition of Wnt/ß-catenin pathway.

Sinomenium acutum: A Comprehensive Review of its Botany, Phytochemistry, Pharmacology and Clinical Application.

Sinomenium acutum is the dry stem of Sinomenium acutum (Thunb.) Rehd et Wils. (S. acutum) and Sinomenium acutum (Thunb.) Rehd. et Wils. var. cinereum Rehd. et Wils and is mainly distributed in China and Japan. As a traditional Chinese medicine (TCM) for dispelling wind and dampness in China, it is widely distributed and has a long history of drug use. In recent years, with the increase of the incidence of rheumatoid disease, S. acutum has become the focus of research. This paper reviews the literature on the chemical constituents, pharmacological effects, clinical applications and pharmacokinetics and safety of S. acutum from the past 60 years. At present, more than 210 natural compounds have been isolated from S. acutum, including alkaloids, lignans, triterpenoid saponins, steroids, and other structures. Pharmacological activities of S. acutum were mainly reported on anti-inflammatory, analgesic, anti-allergic, immunosuppressive, anti-tumor, liver-protective, anti-oxidative, and other effects, and clinical applications were mainly recorded on rheumatoid arthritis, ankylosing spondylitis, and other diseases. The clinical use of SIN has fewer side effects and more safety; only a small number of gastrointestinal reactions occurred, and the symptoms disappeared after the drug stopped. The purpose of this paper is to lay a foundation and provide reference for the follow-up research and wide application of S. acutum.

Eleven undescribed alkaloids from the rhizomes of Sinomenium acutum and their IDO1 and TDO inhibitory activities.

Eleven previously undescribed alkaloids, named sinometumines A-K, along with three known alkaloids, were isolated from the rhizomes of Sinomenium acutum. The chemical structures of these unreported compounds were established using extensive spectroscopic methods (IR, UV, HRESIMS, and NMR), and their absolute configurations were determined by single crystal X-ray diffraction analyses and calculated electronic circular dichroism spectroscopy (ECD). Sinometumine D was the first aporphine-type derived alkaloid inner salt with a rearranged dibenzofuran ring backbone. Sinometumine E was a rare protoberberine-type alkaloid with a complex 6/6/6/6/6/6 hexacyclic skeleton. This was the first report of alkaloids with these two skeletons isolated from S. acutum. All isolates were evaluated for their inhibitory activities against indoleamine 2,3-dioxygenase 1 (IDO1) and tryptophan 2,3-dioxygenase (TDO). Lysicamine possessed noteworthy inhibitory activities as an IDO1/TDO dual inhibitor with IC50 values of 6.22 ± 0.26 µM and 23.76 ± 2.93 µM, respectively, and liriodenine revealed moderate dual inhibition with IC50 values of 31.65 ± 4.44 µM and 15.64 ± 0.26 µM. The intermolecular interactions and binding modes between lysicamine and IDO1/TDO were elaborated by molecular docking studies.

Suppression of macrophage migration by down-regulating Src/FAK/P130Cas activation contributed to the anti-inflammatory activity of sinomenine.

A large number of macrophages in inflamed sites not only amplify the severity of inflammatory responses but also contribute to the deleterious progression of many chronic inflammatory diseases, autoimmune diseases and cancers. Macrophage migration is a prerequisite for their entry into inflammatory sites and their participation of macrophages in the pathologic processes. Inhibition of macrophage migration is therefore a potential anti-inflammatory mechanism. Moreover, alleviation of inflammation also prevents the macrophages infiltration. Sinomenine (SIN) is an alkaloid derived from the Chinese medicinal plant Sinomenium acutum. It has multiple pharmacological effects, including anti-inflammation, immunosuppression, and anti-arthritis. However, its anti-inflammatory molecular mechanisms and effect on macrophage migration are not fully understood. The purpose of this research was to investigate the pharmacological effects and the molecular mechanism of SIN on macrophage migration in vivo and in vitro as well as to elucidate its anti-inflammatory mechanisms associated with macrophage migration. Our results showed that SIN reduced the number of RAW264.7 cells migrating into inflammatory paws and blocked lipopolysaccharide (LPS)-induced RAW264.7 cells and bone marrow-derived macrophages (BMDMs) migration in vitro. Furthermore, SIN attenuated the 3D mesenchymal migration of BMDMs. The absence of macrophage migration after circulatory and periphery macrophages depletion led to a reduction in the severity of inflammatory response. In macrophages depleted (macrophages-/-) mice, as inflammatory severity decreased, RAW264.7 cells migration was suppressed. A non-obvious effect of SIN on the inflammatory response was found in macrophages-/- mice, while the inhibitory effect of SIN on RAW264.7 cells migration was still observed. Furthermore, the migration of RAW264.7 cells pre-treated with SIN was suppressed in normal mice. Finally, Src/focal adhesion kinase (FAK)/P130Cas axis activation, which supports macrophages mesenchymal migration, and iNOS expression, NO production, integrin αV and in integrin ß3 expressions, which promote Src/FAK/P130Cas activation, were down-regulated by SIN. However, SIN had no obvious effect on the expression of the monocyte chemoattractant protein-1 (MCP-1), which is an important chemokine for macrophage migration. These results indicated that SIN significantly inhibited macrophage mesenchymal migration by down-regulating on Src/FAK/P130Cas axis activation. There was a mutual regulatory correlation between the inflammatory response and macrophage migration, and the effects of SIN on macrophage migration were involved in its anti-inflammatory activity.

Bioinformatics Based Therapeutic Effects of Sinomenium Acutum.

OBJECTIVE: To decipher the possible mechanisms of Sinomenium Acutum (SA) in treating diseases by a bioinformatics method. METHODS: SA ingredients were searched according to Chinese Pharmacopoeia, Chinese Medicine Dictionary and Traditional Chinese Medicines Database (TCMD). Active compounds and target proteins of SA were acquired through the Pubchem platform. Pathway, network and function analyses of SA were performed with ingenuity pathway analysis (IPA), a bioinformatics analysis platform. Disease, biofunction-target networks were established with Cytoscape. RESULTS: Eighteen ingredients from SA were obtained. Seven active ingredients with 31 active target proteins were acquired according to PubChem Bioassay test. By IPA analysis, 277 canonical pathways belonging to 17 function categories were collected, 23 kinds of diseases, 21 categories bio-functions were obtained. Based on P value, calculated by IPA, the top 5 significant pathway of SA targets include phosphatidylinositol 3 kinase/Akt (PI3K/Akt) signaling, prostate cancer signaling, macrophage migration inhibitory factor (MIF) regulation of innate immunity, Guanosine-binding protein coupled receptor (GPCR) signaling, and ataxia telangiectasia mutated protein (ATM) signaling. Disease and bio-function network analysis indicated that mitogen activated protein kinase 1 (MAPK1), MAPK3, p65 nuclear factor κB (RELA), nuclear factor of κB inhibitor alpha (NFκBIA), interleukin 1ß(IL-1ß), prostaglandin G/H synthase 2 (PTGS2) and tumor protein 53 (TP53) were the critical targets in various diseases treated by SA. CONCLUSION: In the different view of target, pathway, disease and bio-function, inflammation was found to be a central theme in many chronic conditions. SA could be used not only as an anti-inflammatory agent, but also for the treatment of cancers, neurological diseases, psychological disorders and metabolic diseases.

Sinomenine hydrochloride sensitizes cervical cancer cells to ionizing radiation by impairing DNA damage response.

The use of plant­based compounds derived from traditional medicine to improve human diseases has been gaining momentum, due to their high bioavailability and moderate adverse effects. Sinomenine is one such biomonomer alkali compound derived from Sinomenium acutum and is known for its anti­inflammatory and antitumor effects. However, the molecular mechanism(s) of its antitumor properties are not fully characterized. In the present study, we evaluated the radiosensitizing effects of the water­soluble sinomenine, sinomenine hydrochloride (SH) in human cervical cancer cell line (HeLa). SH sensitized HeLa cells to ionizing radiation (IR) by promoting accumulation of IR­induced DNA double­strand breaks (DSBs) and also by interfering with DNA damage checkpoint activation. We then investigated the molecular mechanisms underlying the SH­mediated cellular sensitization to IR and found that SH inhibited the expression of DNA damage response (DDR) factors Ku80 and Rad51 at the transcription level. Finally, the radiosensitizing activity of SH was confirmed in a cervical cancer mouse xenograft model. The combinatorial treatment of SH and IR significantly slowed the tumor growth rate compared with IR alone. Collectively, our study not only provides molecular insights into the novel role of SH in cellular response to IR, but also suggests a therapeutic potential of SH as a radiosensitizer in cervical cancer therapy.

Sinomenine alleviates dextran sulfate sodium­induced colitis via the Nrf2/NQO­1 signaling pathway.

Sinomenine (SIN), a pure alkaloid isolated from Sinomenium acutum, has been widely used in arthritis for its anti­inflammatory effect, but little is known about the effect of SIN on human ulcerative colitis (UC). In the present study, the effect and mechanism of SIN was examined in a dextran sulfate sodium (DSS)­induced murine colitis model, which mimics human UC. Oral administration of SIN significantly suppressed the elevated disease activity index and ameliorated colonic histological damage in a DSS­induced colitis model. Tumor necrosis factor­α, interleukin­6 and inducible nitric oxide synthase levels were also reduced as detected by reverse transcription­quantitative polymerase chain reaction. In addition, SIN reversed the decreased colon length and colonic superoxide dismutase activity. Furthermore, western blot analysis revealed that nuclear factor­erythroid 2­related factor 2 (Nrf2) and its downstream genes, heme oxygenase­1 and NADP(H) quinone oxidoreductase 1 (NQO­1), were markedly activated by SIN. The current results indicated that SIN alleviated DSS­induced colitis in mice, which may be due to its antioxidant properties and was at least in part dependent on the Nrf2/NQO­1 signaling pathway. Therefore, SIN may have potential applications as a protective drug for patients with UC.

Anti-osteoclastogenic effects of isoquinoline alkaloids from the rhizome extract of Sinomenium acutum.

A phytochemical investigation for the rhizome extract from Sinomenium acutum (Menispermaceae) resulted in the isolation of several active principles responsible for the anti-osteoclastogenic property of the extract, together with related isoquinoline alkaloids (1-13) including two new compounds, 1 and 2. Among isolated compounds, salutaridine (7), dauricumine (10), cheilanthifoline (12), and dauriporphine (13) were observed to give significant inhibitions on receptor activator of nuclear factor-κB ligand-induced differentiation of mouse bone marrow-derived macrophages into multinucleated osteoclasts, respectively. The chemical structures of two newly isolated compounds, 1 and 2 were established as 8-demethoxycephatonine (1) and 7(R)-7,8-dihydrosinomenine (2), by spectroscopic analyses including 2D NMR experiments.

Intra-articular delivery of sinomenium encapsulated by chitosan microspheres and photo-crosslinked GelMA hydrogel ameliorates osteoarthritis by effectively regulating autophagy.

Reduced expression of autophagy regulators has been observed in pathological cartilage in humans and mice. The present study aimed to investigate the synergistic therapeutic effect of promotion of chondrocyte autophagy via exposure to sinomenium (SIN) encapsulated by chitosan microspheres (CM-SIN) and photo-crosslinked gelatin methacrylate (GelMA) hydrogel, with the goal of evaluating CM-SIN as a treatment for patients with osteoarthritis. First, we fabricated and characterized GelMA hydrogels and chitosan microspheres. Next, we measured the effect of SIN on cartilage matrix degradation induced by IL1-ß in chondrocytes and an ex vivo model. SIN ameliorated the pathological changes induced by IL1-ß at least partially through activation of autophagy. Moreover, we surgically induced osteoarthritis in mice, which were injected intra-articularly with CM-SIN and GelMA. Cartilage matrix degradation and chondrocyte autophagy were evaluated 4 and 8 weeks after surgery. Treatment with the combination of CM-SIN and GelMA retarded the progression of surgically induced OA. SIN ameliorated cartilage matrix degradation at least partially by inducing autophagy in vivo. Our results demonstrate that injection of the combination of GelMA hydrogel and CM-SIN could be a promising strategy for treating patients with osteoarthritis.

Anti-inflammatory effects of novel sinomenine derivatives.

Sinomenine is an isoquinoline-type alkaloid found in Sinomenium acutum (Thunb.) Rehd. et Wils and S. acutum (Thunb.) Rehd. et Wils var. cinereum Rehd. et Wils. When used as a medicine, this compound exhibits anti-inflammatory properties; however, sinomenine's use as a medication is limited by side effects, a short half-life, and low efficacy. Owing to these limits, attempts have been made to synthesize sinomenine derivatives with enhanced efficacy. In this study, the anti-inflammatory effects of novel sinomenine derivatives (S1a-S1f) were examined on the basis of lipopolysaccharide-induced inflammatory factor expression in Raw264.7 cells, dimethylbenzene-induced ear oedema, and Evan's blue leakage in mice, and carrageenan-induced paw oedema in rats. Compared with sinomenine, the derivatives significantly inhibited the expression of the inflammatory factors IL-1ß and IL-6 at the transcriptional and translational levels. Topical application of 3.250mg/kg of the derivatives also alleviated ear oedema. Compared with the vehicle, the derivatives significantly inhibited carrageenan-induced rat paw oedema after 6h. Among the derivatives, S1a exhibited the most potent anti-inflammatory activity. S1a also significantly increased the sinomenine-induced inhibition of Evan's blue leakage. Thus, S1a may elicit the strongest anti-inflammatory effects of the tested compounds. Based on these results, further development of this compound may be warranted.

Lignan glucosides from Sinomenium acutum rhizomes.

The new lignan glucoside, acutumoside (1), was isolated from Sinomenium acutum rhizomes together with nine known compounds (2-10). The structure of 1 was elucidated on the basis of extensive spectroscopic analyses, including two-dimensional nuclear magnetic resonance and chemical reactions. Compounds 2, 7, 8, and 10 displayed potential antiproliferative activity against A549, SK-OV-3, SK-MEL-2, and HCT-15 cell lines, while compound 1 showed weak activity against these human tumor cells.

Sinomenine protects against lipopolysaccharide-induced acute lung injury in mice via adenosine A(2A) receptor signaling.

Sinomenine (SIN) is a bioactive alkaloid extracted from the Chinese medicinal plant Sinomenium acutum, which is widely used in the clinical treatment of rheumatoid arthritis (RA). However, its role in acute lung injury (ALI) is unclear. In this study, we investigate the role of SIN in lipopolysaccharide (LPS)-induced ALI in mice. After ALI, lung water content and histological signs of pulmonary injury were attenuated, whereas the PaO2/FIO2 (P/F) ratios were elevated significantly in the mice pretreated with SIN. Additionally, SIN markedly inhibited inflammatory cytokine TNF-α and IL-1ß expression levels as well as neutrophil infiltration in the lung tissues of the mice. Microarray analysis and real-time PCR showed that SIN treatment upregulated adenosine A(2A) receptor (A(2A)R) expression, and the protective effect of SIN was abolished in A(2A)R knockout mice. Further investigation in isolated mouse neutrophils confirmed the upregulation of A(2A)R by SIN and showed that A(2A)R-cAMP-PKA signaling was involved in the anti-inflammatory effect of SIN. Taken together, these findings demonstrate an A(2A)R-associated anti-inflammatory effect and the protective role of SIN in ALI, which suggests a potential novel approach to treat ALI.

The involvement of magnoflorine in the sedative and anxiolytic effects of Sinomeni Caulis et Rhizoma in mice.

The present study seeks to evaluate the sedative and anxiolytic effects of the 70% ethanol extract of Sinomeni Caulis et Rhizoma (SR). The extract was orally administered to mice at dosages of 25, 50, 100, 200 or 400 mg/kg. The mice were then subjected to an array of behavioral tests to assess the sedative (open-field, rota-rod, and thiopental sodium-induced sleeping test) and anxiolytic (elevated plus maze test) effects of the substance. SR (100, 200 mg/kg) significantly reduced locomotor activity, decreased rota-rod performance, and potentiated thiopental sodium-induced sleeping in mice, all indicative of its sedative effects. SR (50, 100 mg/kg) also produced anxiolytic effects, as shown by an increase in entries and staying time on the open arm of the plus maze. SR's sedative and anxiolytic effects were comparable to that of the benzodiazepine, diazepam. Moreover, to identify SR's probable mechanism of action, intracellular Cl⁻ ion influx was observed in cultured human neuroblastoma cells. SR dose-dependently increased Cl⁻ influx, which was blocked by co-administration of the GABAA receptor competitive antagonist, bicuculline. Among the major constituents of SR, only magnoflorine showed a similar increment in Cl⁻ influx, which was also blocked by bicuculline. Altogether, the present results suggest that SR has sedative and anxiolytic effects, probably mediated by magnoflorine through a GABAergic mechanism of action.

Sinoscrewtine, an alkaloid with novel skeleton from the roots of Sinomenium acutum.

An alkaloid with novel skeleton, sinoscrewtine (1), has been isolated from the roots of Sinomenium acutum. Its structure was established by spectral analysis and X-ray crystallographic study, and its possible biosynthetic pathway was delivered. In vitro experiments, 1 showed weak injurious effects against H(2)O(2)/Aß(25-35) induced oxidative injury in PC-12 cells and DPPH radical scavenging activity with IC(50) of 32.6µM.

New alkaloids and cytotoxic principles from Sinomenium acutum.

Two new alkaloids, 2-demethyl-oxypalmatine (1) and 5-ethoxycarbonylsinoracutine (2), were isolated from the rhizomes of Sinomenium acutum, along with thirty-four known compounds. Cytotoxicity of the isolated compounds was examined for the MCF-7, H460, HT-29, and CEM human cancer cell lines. Dauriporphine (16), 6-O-demethylmenisporphine (17), bianfugecine (18), menisporphine (19), and 6-O-demethyldauriporphine (20) showed differential effects in their cytotoxic activity on the target cancer cell lines. Significant angiogenesis inhibitions of 16 and 19 were also observed.

Two new morphinane alkaloids from Sinomenium acutum.

Two new morphinane alkaloids, 1-hydroxy-10-oxo-sinomenine (1) and 4,5-epoxy-14-hydroxy sinomenine N-oxide (2), have been isolated from the stems of Sinomenium acutum. Their structures were established by various spectral analyses, especially 2D NMR experiments. The structure of 2 was confirmed by single crystal X-ray diffraction. The absolute configurations of 1 and 2 were deduced by comparison of CD spectra with the known alkaloid sinomenine (3). Compound 1 was tested for DPPH inhibition and gave IC(50) of 27.9 µM. Compound 2 was tested for neuroprotective effect and showed significant activity against ß-amyloid(25-35)-induced oxidative injury (*P < 0.05) at 10 µM in PC-12 cells.

Morphinane alkaloid dimers from Sinomenium acutum.

Two new morphinane alkaloid dimers, 2,2'-disinomenine (1) and 7',8'-dihydro-1,1'-disinomenine (2), and known 1, 1'-disinomenine (3), were isolated from ethanol extracts of stems of Sinomenium acutum. Their structures were elucidated on the basis of spectroscopic methods. The absolute configuration of alkaloids 1-3 was determined by direct comparison of their CD spectra with the known alkaloid sinomenine. The isolated alkaloids were tested for cytotoxicity against A549, P388, and HeLa cell lines, and 1 and 3 showed weak inhibition against A549 and Hela cells.

Sinomenine promotes differentiation but impedes maturation and co-stimulatory molecule expression of human monocyte-derived dendritic cells.

Dendritic cells (DC) excel at presenting antigen to T cells and thus make a key contribution to the induction of primary and secondary immune responses. Sinomenine has been used for centuries in the treatment of patients with autoimmune diseases as it possesses immunosuppressive and anti-inflammatory activities. However, the effect of sinomenine on the differentiation, maturation, and functionality of DC derived from monocytes has not been studied. We show here that DC differentiation is promoted when monocytes are treated with GM-CSF and IL-4 (IL-4) in the presence of sinomenine (200 microg/ml), as evidenced by the upregulation of CD1a while CD14 was decreased. In addition, incubation of immature DC with sinomenine significantly blunted lipopolysaccharide (LPS)-induced DC maturation, as shown by the reduction of expression of the maturation marker CD83 and co-stimulatory molecules, including CD86, B7-H1, and CD40. Moreover, sinomenine also prevented decreases in antigen (FITC-Dextran or Lucifer Yellow) uptake by LPS-treated DC. Mixed lymphocyte reactions (MLRs) revealed that sinomenine-treated DC impede the secretion of the cytokines IL-2 and IFN-gamma by co-cultured CD4(+) T cells. Therefore, modulation of DC differentiation, maturation, and functionality by sinomenine is of potential relevance to its immunomodulatory effects in controlling specific immune responses in autoimmune diseases, transplantation, and other immune-mediated conditions.

Aporphine alkaloids and their reversal activity of multidrug resistance (MDR) from the stems and rhizomes of Sinomenium acutum.

Chromatographic separation of the MeOH extract from the stems and rhizomes of Sinomemium acutum led to the isolation of nine alkaloids and a lignan. Their structures were determined to be dauriporphine (1), bianfugecine (2), dauriporphinoline (3), menisporphine (4), (-)-syringaresinol (5), N-feruloyltyramine (6), acutumine (7), dauricumine (8), sinomenine (9), and magnoflorine (10) by spectroscopic means. These compounds were examined for their P-gp mediated MDR reversal activity in human cancer cells. Compound 1 showed the most potent P-gp MDR inhibition activity with an ED50 value 0.03 microg/mL and 0.00010 microg/mL in the MES-SA/DX5 and HCT15 cells, respectively.

Angiogenesis: from plants to blood vessels.

Angiogenesis is a major pathological component of diseases such as cancer and coronary heart disease. Although major advances have been made and encouraging clinical results obtained, safer and more effective approaches are required. The identification of new drugs from plants has a long and successful history, and certain proangiogenic and antiangiogenic plant components have been used in traditional Chinese medicine (TCM) for thousands of years. Similar to Western combination therapy, TCM uses mixtures of plant extracts, termed fufang, to maximize efficacy and minimize adverse effects or toxicity. More evidence-based research and chemical optimization of these compounds could further enhance the effectiveness of these plant-based medicines in angiotherapy.