Silvaticusins A-D: ent-kaurane diterpenoids and a cyclobutane-containing ent-kaurane dimer from Isodon silvaticus.

Three new ent-kaurane diterpenoids, silvaticusins A-C (1-3), along with a new ent-kaurane dimer silvaticusin D (4) were isolated from the aerial parts of Isodon silvaticus. The structures of these new compounds were established mainly by comprehensive analysis of their NMR and MS data. The absolute configuration of compounds 1 and 4 were determined using a single-crystal X-ray diffraction and computational methods, respectively. Compounds 2 and 3 were found to exhibit remarkable cytotoxic effects against five human tumor cell lines (HL-60, A-549, SMMC-7721, MDA-MB-231, and SW-480), with IC50 values spanning from 1.27 ± 0.08 to 7.52 ± 0.33 µM.

Four new lanostane triterpenoids featuring extended π-conjugated systems from the stems of Kadsura coccinea.

Four new 14(13 → 12)-abeolanostane triterpenoids featuring extended π-conjugated systems, kadcoccitanes E-H (1-4), were obtained from the stems of Kadsura coccinea through using a HPLC - UV-guided approach. Their structural and configurational determination was accomplished through extensive spectroscopic analysis coupled with quantum chemical calculations. Kadcoccitanes E-H were tested for their cytotoxic activities against five human tumor cell lines (HL-60, A-549, SMMC-7721, MDA-MB-231, SW-480) but none of them exhibited activities at the concentration 40 µM.

Cytotoxic C-20 non-oxygenated ent-kaurane diterpenoids from Isodon wardii.

Twenty new ent-kaurane diterpenoids, wardiisins A-T (1-20), along with two previously undescribed artefactual compounds (21 and 22) and twelve known analogues (23-34), were isolated from the aerial part of Isodon wardii. Their structures were elucidated by comprehensive analysis of spectroscopic data and single-crystal X-ray diffraction, and most of them were found to bear unusual C-12 oxygenation. Compounds 4, 7, 8, 19, 20, 21 exhibited remarkable cytotoxicity against the cancer cell lines HL-60, SMMC-7721, A-549, MDA-MB-231, and SW480, with IC50 values ranging from 0.3 to 5.2 µM. Moreover, 7 was found to induce G2/M cell cycle arrest and promote apoptosis in SW480 cell lines.

Natural product Eriocalyxin B suppressed triple negative breast cancer metastasis both in vitro and in vivo.

Breast cancer is the most commonly diagnosed cancer among women, and its metastasis to distant organs accounts for the majority of death. Eriocalyxin B (Eri B), an ent-kaurane diterpenoid isolating from Isodon eriocalyx var. laxiflora, has previously been reported to have anti-tumor and anti-angiogenic effects in breast cancer. Here, we investigated the effect of Eri B on cell migration and adhesion in triple negative breast cancer (TNBC) cells, as well as aldehyde dehydrogenases 1 family member A1 (ALDH1A1) expression, colony- and sphere-formation in cancer stem cell (CSC) enriched MDA-MB-231 cells. The in vivo anti-metastatic activities of Eri B were determined in 3 different breast tumor-bearing mouse models. Our results indicated that Eri B inhibited TNBC cell migration and adhesion to extracellular matrix proteins, and also reduced ALDH1A1 expression and colony formation in CSC-enriched MDA-MB-231 cells. The metastasis-related pathways, such as epidermal growth factor receptor/ mitogen-activated protein kinase kinases 1/2/ extracellular regulated protein kinase signaling altered by Eri B was firstly shown in MDA-MB-231 cells. The potent anti-metastatic efficacies of Eri B were demonstrated in breast xenograft-bearing mice and syngeneic breast tumor-bearing mice. Gut microbiome analysis results revealed the change in the diversity and composition of microbiome after Eri B treatment, and the potential pathways that are involved in the anti-cancer efficacy of Eri B. In conclusion, Eri B was shown to inhibit breast cancer metastasis in both in vitro and in vivo models. Our findings further support the development of Eri B as an anti-metastatic agent for breast cancer.

Scopariusicides D-M, ent-clerodane-based isomeric meroditerpenoids with a cyclobutane-fused γ/δ-lactone core from Isodon scoparius.

Scopariusicides D-M (1-10), ten new ent-clerodane-based meroditerpenoids with a cyclobutane-fused γ/δ-lactone core, were isolated from Isodon scoparius. Their structures were determined by comprehensive analysis of spectroscopic data, single-crystal X-ray diffraction, chemical transformation, and TDDFT ECD calculation. A plausible biosynthetic pathway of 1-10 was proposed in which the asymmetrical cyclobutane ring was formed via a crossed "head-to-tail" intermolecular [2 + 2] cycloaddition in anti/syn facial approaches between an ent-clerodane lactone and a cis-4-hydroxycinnamic acid. Bioactivity evaluation manifested that 5 exhibited significant neuroprotective effect against corticosterone-induced injury in PC12 cells, while 6 and 7 exhibited moderate immunosuppressive activity against human T cell proliferation stimulated by anti-CD3/anti-CD28 mAb.

Structurally diverse diterpenoids from Isodon oresbius and their bioactivity.

Twelve new diterpenoids, isoresbins A-L (1-12), together with twenty-eight known ones, were isolated from the aerial parts of Isodon oresbius. Their diverse structures included 6,7-seco-ent-kaurane, 7,20-epoxy-ent-kaurane, 6,7:8,15-diseco-ent-kaurane, and abietanes skeletons, which were elucidated by spectroscopic data interpretation, single-crystal X-ray diffraction, and quantum chemical calculation. Isoresbins A (1) and B (2) possessed a new rearranged 15(8 â†’ 11)-abeo-6,7-seco-ent-kaurane skeleton. 1 and 5 promoted lysosomal function, which was evaluated by LysoTracker Red staining and DQ-ovalbumin dequenching assay. 1 showed cytotoxicity against six human tumor cell lines with IC50 values in 2.07-4.04 µM range. Moreover, 1 induced damage of mitochondrial membrane potential, G2/M cell cycle arrest and apoptosis in SW480 cells.

Discovery of ent-kaurane diterpenoids, characteristic metabolites of Isodon species, from an endophytic fungal strain Geopyxis sp. XY93 inhabiting Isodon parvifolia.

Isogeopyxins A-C (1-3), three new diterpenoids with ent-kaurane, ent-pimarane, and ent-abietane scaffolds, respectively, along with six known ent-kauranoids, were isolated from the fermentation culture of Geopyxis sp. XY93 inhabiting the leaves of Isodon parvifolia. Their structures were elucidated by interpretation of spectroscopic data, and single crystal X-ray diffraction. It marks the first time that ent-kauranoids, characteristic metabolites of Isodon species, have been isolated from an associated endophytic fungus.

Cordyceps polysaccharide marker CCP modulates immune responses via highly selective TLR4/MyD88/p38 axis.

Cordyceps, one of the most expensive natural health supplements, is popularly used to modulate immune function. However, little is known regarding the underlying mechanism of its immunomodulatory activity. We newly reported a Cordyceps quality marker CCP (Mw 433.778 kDa) which was characterized as a 1,4-α glucan by chemical and spectral analysis and is able to induce significant immune responses of macrophages. Herein, we further investigated the molecular mechanism of CCP's immunomodulatory effects. The results indicate that CCP modulates the TLR4/MyD88/p38 signaling pathway of macrophages, where TLR4 plays a crucial role as verified on TLR4-deficient (TLR4-/-) bone marrow-derived macrophages (BMDMs) and TLR4-/- mice. These findings provide a precise understanding of the molecular mechanism of Cordyceps' immunomodulatory benefits.

Spiro ent-Clerodane Dimers: Discovery and Green Approaches for a Scalable Biomimetic Synthesis.

Scospirosins A (1) and B (2), two unprecedented spiro ent-clerodane dimers with 6/6/10/6 and 6/6/6/6/6 ring systems, respectively, were isolated from Isodon scoparius. Their structures were unambiguously established by spectroscopic, X-ray crystallographic, and chemical approaches. A bioinspired protecting-group-free strategy for their synthesis was achieved on a gram scale and featured the application of green methods, including neat reaction, sensitized photooxygenation, and electrochemical oxidation. 2 exhibited selective immunosuppressive activity against the proliferation of T lymphocytes (IC50 = 1.42 µM).

Neuroprotective schinortriterpenoids from Schisandra neglecta collected in Medog County, Tibet, China.

Schisdilactones K-U (1-11), a series of previously unreported 16,17-secopreschisanartane-type schinortriterpenoids (SNTs), were isolated from the leaves and stems of Schisandra neglecta A. C. Smith. Their structures were mainly established through analysis of their spectroscopic data. Besides, schisdilactones K (1), O (5) and R (8) were confirmed by single-crystal X-ray crystallographic analysis, and the configurations of schisdilactones T and U (10 and 11) were elucidated via quantum chemical calculation of their NMR chemical shifts and electronic circular dichroism (ECD) spectra. Schisdilactones L-S (2-8) and U (11) were found to exhibit moderate protective activities against corticosterone-induced apoptosis of PC12 cells at 20 µM, with cell viability in the range of 62.95-66.97%.

High-content screening of diterpenoids from Isodon species as autophagy modulators and the functional study of their antiviral activities.

Autophagy is a conserved lysosomal degradation process, and abnormal autophagy has been associated with various pathological processes, e.g., neurodegeneration, cancer, and pathogen infection. Small chemical modulators of autophagy show the potential to treat autophagy-associated diseases. Diterpenoids, nature products found in various plants, exhibit a wide range of bioactivity, and we have recently isolated and characterized over 150 diterpenoids from Isodon species distributed in China. Here, we applied a high-content fluorescence imaging-based assay to assess these diterpenoids' ability to affect autophagic flux in HeLa cells. We found that enanderinanin J, an ent-kauranoid dimer, is an autophagy inhibitor, manifested by its ability to increase lysosomal pH and inhibit the fusion between autophagosomes and lysosomes. Autophagy has been shown to be either positively or negatively involved in the life cycle of Zika virus (ZIKV), Japanese encephalitis virus (JEV), Dengue virus (DENV), and enterovirus-A71 (EV-A71). We found that enanderinanin J significantly inhibited the infection of ZIKV, DENV, JEV, or EV-A71. Interestingly, although ATG5 knockdown inhibited ZIKV or JEV infection, enanderinanin J further inhibited the infection of ZIKV or JEV in ATG5-knockdown cells. Taken together, our data indicate that enanderinanin J inhibits autophagosome-lysosome fusion and is a potential antiviral agent.

A review on the relationship of mast cells and macrophages in breast cancer - Can herbs or natural products facilitate their anti-tumor effects?

Breast cancer is an inflammation-related cancer whose tumor microenvironment is largely infiltrated by inflammatory cells. These inflammatory cells including mast cells and macrophages have been elucidated to be vital participants in breast tumor proliferation, survival, invasion and migration. However, the functions of mast cells and macrophages in breast cancer are quite distinct based on recent data. Mast cells exhibit both anti-tumoral and pro-tumoral functions on breast cancer, while high number of tumor-associated macrophages (TAMs) are strongly correlated with poor prognosis and higher risk of distant metastasis in breast cancer patients. Besides, many natural products/extracts have been reported to regulate mast cells and macrophages. In this review, the roles of mast cells and macrophages play in breast cancer are discussed and a summary of those natural products/herbs regulating the functions of mast cells or macrophages is also presented.

Neuroprotective schinortriterpenoids with diverse scaffolds from Schisandra henryi.

Eleven undescribed schinortriterpenoids (SNTs) and one known analogue (12) were isolated from the stems and leaves of Schisandra henryi. Their diverse structures included preschisanartane (1), 18-norschiartane (2-5, 12), schiartane (6 and 7), and schisanartane (8-11) skeletons, which were elucidated by comprehensive NMR, MS, electronic circular dichroism analyses, single crystal X-ray diffraction and biogenetic considerations. 1 was the first case of preschisanartane-type SNT with six-membered lactone ring, and 2 was one of the most highly oxygenated 18-norschiartane SNTs. Three types of the highly oxygenated SNTs, 1, 4, 10 and 11, effectively prevent apoptosis induced by corticosterone in PC12 cells. In addition, 11 showed neurite outgrowth-promoting activity.

Isorugosiformins A-F, six ent-kaurane diterpenoids from Isodon rugosiformis.

Six new ent-kaurane diterpenoids, isorugosiformins A-F (1-6), were isolated from the aerial parts of Isodon rugosiformis Hand.-Mazz. Hara. Their structures were elucidated by spectroscopic data interpretation, single crystal X-ray diffraction, and quantum chemical calculation of NMR parameters. The absolute configuration of 5 as 6R was the first case in the known 6,7:8,15-diseco-7,20-olide-6,8-cyclo-ent-kaurane diterpenoids.

Discovery of isopenicin A, a meroterpenoid as a novel inhibitor of tubulin polymerization.

Microtubules are involved in celluar processes of movement, intracellular trafficking and mitosis, thus microtubule-targeting agents have been widely used in cancer therapy. Herein, we report isopenicin A, a novel meroterpenoid isolated from the plant endophytic fungus of Penicillium sp. sh18, as a novel microtubule binding molecule that efficiently depolymerizes microtubule polymerization to evoke G2/M cell cycle arrest and subsequent cell apoptosis, contributing to proliferation inhibition of human tumor cell lines. The discovery of isopenicin A provides a new chemotype for discovery and development of promising microtubule inhibitors.

Validation of Cadherin HAV6 Peptide in the Transient Modulation of the Blood-Brain Barrier for the Treatment of Brain Tumors.

The blood-brain barrier (BBB) poses a major obstacle by preventing potential therapeutic agents from reaching their intended brain targets at sufficient concentrations. While transient disruption of the BBB has been used to enhance chemotherapeutic efficacy in treating brain tumors, limitations in terms of magnitude and duration of BBB disruption exist. In the present study, the preliminary safety and efficacy profile of HAV6, a peptide that binds to the external domains of cadherin, to transiently open the BBB and improve the delivery of a therapeutic agent, was evaluated in a murine brain tumor model. Transient opening of the BBB in response to HAV6 peptide administration was quantitatively characterized using both a gadolinium magnetic resonance imaging (MRI) contrast agent and adenanthin (Ade), the intended therapeutic agent. The effects of HAV6 peptide on BBB integrity and the efficacy of concurrent administration of HAV6 peptide and the small molecule inhibitor, Ade, in the growth and progression of an orthotopic medulloblastoma mouse model using human D425 tumor cells was examined. Systemic administration of HAV6 peptide caused transient, reversible disruption of BBB in mice. Increases in BBB permeability produced by HAV6 were rapid in onset and observed in all regions of the brain examined. Concurrent administration of HAV6 peptide with Ade, a BBB impermeable inhibitor of Peroxiredoxin-1, caused reduced tumor growth and increased survival in mice bearing medulloblastoma. The rapid onset and transient nature of the BBB modulation produced with the HAV6 peptide along with its uniform disruption and biocompatibility is well-suited for CNS drug delivery applications, especially in the treatment of brain tumors.

Isoforrethins A-D, four ent-abietane diterpenoids from Isodon forrestii var. forrestii.

Four new ent-abietane diterpenoids, isoforrethins A-D (1-4), were isolated from the aerial parts of Isodon forrestii var. forrestii by a variety of chromatographic techniques. Their structures were elucidated on the basis of spectroscopic data interpretation, single crystal X-ray diffraction, and quantum chemical calculation. All of these compounds were evaluated for their cytotoxicity against five human tumor cell lines (HL-60, SMMC-7721, A-549, MCF-7 and SW-480), and compound 4 showed significant inhibitory activities against SMMC-7721, A-549, MCF-7, and SW-480.

Adenanthin, a Natural ent-Kaurane Diterpenoid Isolated from the Herb Isodon adenantha Inhibits Adipogenesis and the Development of Obesity by Regulation of ROS.

Adenanthin, a natural ent-kaurane diterpenoid extracted from the herb Isodon adenantha, has been reported to increase intracellular reactive oxygen species in leukemic and hepatocellular carcinoma cells. However, the function and mechanism of the compound in adipogenesis and the development of obesity is still unknown. In this study, we demonstrated that adenanthin inhibited adipogenesis of 3T3-L1 and mouse embryonic fibroblasts, and the underlying mechanism included two processes: a delayed mitotic clonal expansion via G0/G1 cell cycle arrest by inhibiting the RB-E2F1 signaling pathway and a reduced C/EBPß signaling by inhibiting the expression and activity of C/EBPß during mitotic clonal expansion. Furthermore, adenanthin significantly reduced the growing body weight and adipose tissue mass during high-fat diet-inducing obesity of mice, indicating the beneficial effects of adenanthin as a potential agent for prevention of obesity.

Acetyl-macrocalin B suppresses tumor growth in esophageal squamous cell carcinoma and exhibits synergistic anti-cancer effects with the Chk1/2 inhibitor AZD7762.

Natural products derived from herbal medicines have become a major focus of anti-cancer drug discovery studies. Acetyl-macrocalin B (A-macB) is an ent-diterpenoid isolated from Isodon silvatica. This study aimed to examine the effect and molecular action of A-macB in esophageal squamous cell carcinoma (ESCC) and explore possible drug synergistic modalities. A-macB induced cellular reactive oxygen species (ROS) generation, initiated the p38 mitogen-activated protein kinase (MAPK) signaling pathway, and triggered the caspase-9-dependent apoptosis cascade in ESCC cells. The ROS scavenger N-acetylcysteine (NAC) and the specific p38 inhibitor SB203580 reversed the effects of A-macB on the p38 network and thus rescued ESCC cells from apoptosis. The cellular ROS increase was at least partially due to the suppression of glutathione-S-transferase P1 (GSTP1) by A-macB. A-macB also upregulated the Chk1/Chk2-Cdc25C/Cdc2/Cyclin B1 axis to induce G2/M phase arrest. The cell growth inhibition induced by A-macB was further enhanced by AZD7762, a specific Chk1/Chk2 inhibitor, with a combination index (CI) of <1. Moreover, A-macB efficiently suppressed xenograft growth without inducing significant toxicity, and AZD7762 potentiated the effects of A-macB in the suppression of tumor growth in vivo. Taken together, A-macB is a promising lead compound for ESCC and exerts synergistic anti-cancer effects with AZD7762.

Isopenicins A-C: Two Types of Antitumor Meroterpenoids from the Plant Endophytic Fungus Penicillium sp. sh18.

Isopenicins A-C (1-3), three novel meroterpenoids possessing two types of unprecedented terpenoid-polyketide hybrid skeletons, were isolated from the cultures of Penicillium sp. sh18. Their structures were determined through synergetic use of extensive spectroscopic analysis, quantum-chemical calculation with ANN-PRA analysis, and X-ray crystallographic analysis. Additionally, the inhibitory activities of these compounds on the Wnt/ß-catenin signaling pathway were evaluated, and 1 was identified as a potent inhibitor of the Wnt signaling pathway.