(±)-hypermonanones A-G, seven pairs of monoterpenoid polyprenylated acylphloroglucinol enantiomers from Hypericum monanthemum.

Seven pairs of undescribed monoterpenoid polyprenylated acylphloroglucinol enantiomers [(±)-hypermonanones A-G (1-7)], together with three known analogues, were identified from the whole plant of Hypericum monanthemum Hook. The structures of these compounds were determined by analyses of their UV, HRESIMS, 1D/2D NMR spectroscopic data, and NMR calculations. The absolute configurations of these compounds were assigned by ECD calculations after chiral HPLC separation. Diverse monoterpene moieties were fused at C-3/C-4 of the dearomatized acylphloroglucinol core, which led to 3,4-dihydro-2H-pyran-integrated angular or linear type 6/6/6 tricyclic skeletons in 1-7. Compounds (-)-2 and (+)-2 exhibited significant NO inhibitory activity against LPS induced RAW264.7 cells with the IC50 values of 7.07 ± 1.02 µM and 11.39 ± 0.24 µM, respectively.

Design, synthesis and biological evaluation of CDC20 inhibitors for treatment of triple-negative breast cancer.

The involvement of CDC20 in promoting tumor growth in different types of human cancers and it disturbs the process of cell division and impedes tumor proliferation. In this work, a novel of Apcin derivatives targeting CDC20 were designed and synthesized to evaluate for their biological activities. The inhibitory effect on the proliferation of four human tumor cell lines (MCF-7, MDA-MB-231, MDA-MB-468 and A549) was observed. Among them, compound E1 exhibited the strongest inhibitory effect on the proliferation of MDA-MB-231 cells with an IC50 value of 1.43 µM, which was significantly superior to that of Apcin. Further biological studies demonstrated that compound E1 inhibited cancer cell migration and colony formation. Furthermore, compound E1 specifically targeted CDC20 and exhibited a higher binding affinity to CDC20 compared to that of Apcin, thereby inducing cell cycle arrest in the G2/M phase of cancer cells. Moreover, it has been observed that compound E1 induces autophagy in cancer cells. In 4T1 Xenograft Models compound E1 exhibited the potential antitumor activity without obvious toxicity. These findings suggest that E1 could be regarded as a CDC20 inhibitor deserved further investigation.

Structurally diverse triterpenoid alkaloids from Buxus rugulosa.

Four new nortriterpenoid alkaloids, namely buxrugulines E-H (1-4), along with five known ones (5-9), were isolated from the twigs and leaves of Buxus rugulosa. Their structures were identified based on extensive NMR data and MS spectroscopic analyses. Our bioassays revealed that compounds 5, 6 and 8 exhibited potent cytotoxicity in vitro against MCF-7 cell lines, with IC50 values ranging from 6.70 to 11.00 µM, respectively.

Withanolides from Physalis angulata var. villosa and the Relative Configurational Revision of Some Known Analogs.

Physalis angulata var. villosa is a plant possessing abundant withanolides, but in-depth research is lacking. In our ongoing study of P. angulata var. villosa, 15 previously undescribed withanolides (1-15), along with 21 known analogs (16-36), were isolated from the whole plant. The structures of the withanolides (1-15) were elucidated based on analysis of their 1D and 2D NMR, HRESIMS, and ECD data. Additionally, the application of γ-gauche effects with the help of ROESY correlations led to the formulation of empirical rules for withanolides with 14-OH/15-OAc to rapidly determine the 14-OH orientations, making it possible to propose configurational revisions of 19 previously reported analogs (1'-19'). Withanolides 1, 4-6, and 10 showed potent cytotoxic activities against three human cancer cell lines (HCT-116, MDA-MB-231, and A549).

Nickel Nanoparticles Protruding from Molybdenum Carbide Micropillars with Carbon Layer-Protected Biphasic 0D/1D Heterostructures for Efficient Water Splitting.

It remains a tremendous challenge to achieve high-efficiency bifunctional electrocatalysts for both the hydrogen evolution reaction (HER) and the oxygen evolution reaction (OER) for hydrogen production by water splitting. Herein, a novel hybrid of 0D nickel nanoparticles dispersed on the one-dimensional (1D) molybdenum carbide micropillars embedded in the carbon layers (Ni/Mo2C@C) was successfully prepared on nickel foam by a facile pyrolysis strategy. During the synthesis process, the nickel nanoparticles and molybdenum carbide were simultaneously generated under H2 and C2H2 mixed atmospheres and conformally encapsulated in the carbon layers. Benefiting from the distinctive 0D/1D heterostructure and the synergistic effect of the biphasic Mo2C and Ni together with the protective effect of the carbon layer, the reduced activation energy barriers and fast catalytic reaction kinetics can be achieved, resulting in a small overpotential of 96 mV for the HER and 266 mV for the OER at the current density of 10 mA cm-2 together with excellent durability in 1.0 M KOH electrolyte. In addition, using the developed Ni/Mo2C@C as both the cathode and anode, the constructed electrolyzer exhibits a small voltage of 1.55 V for the overall water splitting. The novel designed Ni/Mo2C@C may give inspiration for the development of efficient bifunctional catalysts with low-cost transition metal elements for water splitting.

Discovery and optimization of indirubin derivatives as novel ferroptosis inducers for the treatment of colon cancer.

Glutathione peroxidase 4 (GPX4) is an essential antioxidant enzyme that negatively regulates ferroptosis. To exploit novel GPX4 inhibitors, we designed and synthesized 32 indirubin derivatives. Compound 31 exhibited the strongest antitumor activity against HCT-116 cells (IC50 = 0.49 ± 0.02 µM). Further studies suggested that 31 could induce ferroptosis in colon cancer cells and its cytotoxic activity could be reversed by ferroptosis inhibitors. Mechanism research showed that 31 promoted the degradation of GPX4, causing the accumulation of lipid ROS to induce ferroptosis. Animal experiments also proved that 31 could inhibit the growth of colon cancer cells in vivo and reduce the expression of GPX4 in tumor tissues. These results indicated that compound 31 had potential as a novel ferroptosis inducer agent for colon cancer.

Anti-Inflammatory Lindenane Sesquiterpenoid Dimers from the Roots of Sarcandra glabra.

Sarglaroids A-H (1-8), eight new lindenane dimers, and a monomer sarglaroid I (9), along with fourteen known analogues (10-23), were isolated from the roots of Sarcandra glabra. The planar structures and the absolute configurations were elucidated by HR-MS, NMR, ECD calculations, and X-ray diffraction crystallography. Sarglaroid A (1) was identified as a rare 8,9-seco lindenane dimer with a unique 5/5/5 tricyclic system. The biological evaluation showed that compounds 1 and 13 potently inhibited NO production with IC50 values at 19.8 ± 1.06 and 10.7 ± 0.25 µM, respectively, and had no cytotoxicity to RAW264.7 cells. Compound 6 significantly inhibited the LPS-/ATP-induced IL-1ß release by inactivating the NLRP3 inflammasome through inhibiting the initiation and assembly by affecting the K+ efflux. Compounds 2 and 3 inhibited the proliferation of MCF-7 and MDA-MB-231 with IC50 values ranging from 5.4 to 10.2 µM.

Sanguinarine induces apoptosis in osteosarcoma by attenuating the binding of STAT3 to the single-stranded DNA-binding protein 1 (SSBP1) promoter region.

BACKGROUND AND PURPOSE: Osteosarcoma, a primary malignant bone tumour prevalent among adolescents and young adults, remains a considerable challenge despite protracted progress made in enhancing patient survival rates over the last 40 years. Consequently, the development of novel therapeutic approaches for osteosarcoma is imperative. Sanguinarine (SNG), a compound with demonstrated potent anticancer properties against various malignancies, presents a promising avenue for exploration. Nevertheless, the intricate molecular mechanisms underpinning SNG's actions in osteosarcoma remain elusive, necessitating further elucidation. EXPERIMENTAL APPROACH: Single-stranded DNA-binding protein 1 (SSBP1) was screened out by differential proteomic analysis. Apoptosis, cell cycle, reactive oxygen species (ROS) and mitochondrial changes were assessed via flow cytometry. Western blotting and quantitative real-time reverse transcription PCR (qRT-PCR) were used to determine protein and gene levels. The antitumour mechanism of SNG was explored at a molecular level using chromatin immunoprecipitation (ChIP) and dual luciferase reporter plasmids. KEY RESULTS: Our investigation revealed that SNG exerted an up-regulated effect on SSBP1, disrupting mitochondrial function and inducing apoptosis. In-depth analysis uncovered a mechanism whereby SNG hindered the JAK/signal transducer and activator of transcription 3 (STAT3) signalling pathway, relieved the inhibitory effect of STAT3 on SSBP1 transcription, and inhibited the downstream PI3K/Akt/mTOR signalling axis, ultimately activating apoptosis. CONCLUSIONS AND IMPLICATIONS: The study delved further into elucidating the anticancer mechanism of SNG in osteosarcoma. Notably, we unravelled the previously undisclosed apoptotic potential of SSBP1 in osteosarcoma cells. This finding holds substantial promise in advancing the development of novel anticancer drugs and identification of therapeutic targets.

Structure of the Major G-Quadruplex in the Human EGFR Oncogene Promoter Adopts a Unique Folding Topology with a Distinctive Snap-Back Loop.

EGFR tyrosine kinase inhibitors have made remarkable success in targeted cancer therapy. However, therapeutic resistance inevitably occurred and EGFR-targeting therapy has been demonstrated to have limited efficacy or utility in glioblastoma, colorectal cancer, and hepatocellular carcinoma. Therefore, there is a high demand for the development of new targets to inhibit EGFR signaling. Herein, we found that the EGFR oncogene proximal promoter sequence forms a unique type of snap-back loop containing G-quadruplex (G4), which can be targeted by small molecules. For the first time, we determined the NMR solution structure of this snap-back EGFR-G4, a three-tetrad-core, parallel-stranded G4 with naturally occurring flanking residues at both the 5'-end and 3'-end. The snap-back loop located at the 3'-end region forms a stable capping structure through two stacked G-triads connected by multiple potential hydrogen bonds. Notably, the flanking residues are consistently absent in reported snap-back G4s, raising the question of whether such structures truly exist under in vivo conditions. The resolved EGFR-G4 structure has eliminated the doubt and showed distinct structural features that distinguish it from the previously reported snap-back G4s, which lack the flanking residues. Furthermore, we found that the snap-back EGFR-G4 structure is highly stable and can form on an elongated DNA template to inhibit DNA polymerase. The unprecedented high-resolution EGFR-G4 structure has thus contributed a promising molecular target for developing alternative EGFR signaling inhibitors in cancer therapeutics. Meanwhile, the two stacked triads may provide an attractive site for specific small-molecule targeting.

Discovery of a potent inhibitor of chaperone-mediated autophagy that targets the HSC70-LAMP2A interaction in non-small cell lung cancer cells.

BACKGROUND AND PURPOSE: Chaperone-mediated autophagy (CMA) is a selective type of autophagy targeting protein degradation and maintains high activity in many malignancies. Inhibition of the combination of HSC70 and LAMP2A can potently block CMA. At present, knockdown of LAMP2A remains the most specific method for inhibiting CMA and chemical inhibitors against CMA have not yet been discovered. EXPERIMENTAL APPROACH: Levels of CMA in non-small cell lung cancer (NSCLC) tissue samples were confirmed by tyramide signal amplification dual immunofluorescence assay. High-content screening was performed based on CMA activity, to identify potential inhibitors of CMA. Inhibitor targets were determined by drug affinity responsive target stability-mass spectrum and confirmed by protein mass spectrometry. CMA was inhibited and activated to elucidate the molecular mechanism of the CMA inhibitor. KEY RESULTS: Suppression of interactions between HSC70 and LAMP2A blocked CMA in NSCLC, restraining tumour growth. Polyphyllin D (PPD) was identified as a targeted CMA small-molecule inhibitor through disrupting HSC70-LAMP2A interactions. The binding sites for PPD were E129 and T278 at the nucleotide-binding domain of HSC70 and C-terminal of LAMP2A, respectively. PPD accelerated unfolded protein generation to induce reactive oxygen species (ROS) accumulation by inhibiting HSC70-LAMP2A-eIF2α signalling axis. Also, PPD prevented regulatory compensation of macroautophagy induced by CMA inhibition via blocking the STX17-SNAP29-VAMP8 signalling axis. CONCLUSIONS AND IMPLICATIONS: PPD is a targeted CMA inhibitor that blocked both HSC70-LAMP2A interactions and LAMP2A homo-multimerization. CMA suppression without increasing the regulatory compensation from macroautophagy is a good strategy for NSCLC therapy.

Meso-Hannokinol inhibits breast cancer bone metastasis via the ROS/JNK/ZEB1 axis.

Distal metastases from breast cancer, especially bone metastases, are extremely common in the late stages of the disease and are associated with a poor prognosis. EMT is a biomarker of the early process of bone metastasis, and MMP-9 and MMP-13 are important osteoclastic activators. Previously, we found that meso-Hannokinol (HA) could significantly inhibit EMT and MMP-9 and MMP-13 expressions in breast cancer cells. On this basis, we further explored the role of HA in breast cancer bone metastasis. In vivo, we established a breast cancer bone metastasis model by intracardially injecting breast cancer cells. Intraperitoneal injections of HA significantly reduced breast cancer cell metastasis to the leg bone in mice and osteolytic lesions caused by breast cancer. In vitro, HA inhibited the migration and invasion of breast cancer cells and suppressed the expressions of EMT, MMP-9, MMP-13, and other osteoclastic activators. HA inhibited EMT and MMP-9 by activating the ROS/JNK pathway as demonstrated by siJNK and SP600125 inhibition of JNK phosphorylation and NAC scavenging of ROS accumulation. Moreover, HA promoted bone formation and inhibited bone resorption in vitro. In conclusion, our findings suggest that HA may be an excellent candidate for treating breast cancer bone metastasis.

Hyperxylones A and B, two polycyclic polyprenylated acylphloroglucinols with a benzoyl substituted bicyclo[3.2.1]octane core from Hypericum beanii.

Two new polycyclic polyprenylated acylphloroglucinols (PPAPs) possessing a rare benzoyl substituted bicyclo[3.2.1]octane core, hyperxylones A (1) and B (2), along with three new dearomatized isoprenylated acylphloroglucinols (DIAPs), hyperxylones C - E (3-5), were isolated from the roots of Hypericum beanii. The structures of 1-5 were determined by high-resolution electrospray ionization mass spectroscopy (HRESIMS) and 1D/2D nuclear magnetic resonance (NMR) spectroscopic analyses, gauge-independent atomic orbital (GIAO) NMR calculations, and electronic circular dichroism (ECD) calculations. Compounds 1 and 2 were biomimetically semi-synthesized starting from 5 and 4, respectively, enabling the correct stereochemical assignment of 5 and 4. Moreover, compounds 1 and 2 showed anti-nonalcoholic steatohepatitis (NASH) activity by inhibiting lipid deposition in L02 cells; compounds 3 and 5 exhibited nitric oxide (NO) inhibitory activity in lipopolysaccharides (LPS)-induced RAW264.7 cells.

Single-cell sequencing reveals the landscape of the tumor microenvironment in a skeletal undifferentiated pleomorphic sarcoma patient.

Skeletal undifferentiated pleomorphic sarcoma (SUPS) is an invasive pleomorphic soft tissue sarcoma with a high degree of malignancy and poor prognosis. It is prone to recur and metastasize. The tumor microenvironment (TME) and the pathophysiology of SUPS are barely described. Single-cell RNA sequencing (scRNA-seq) provides an opportunity to dissect the landscape of human diseases at an unprecedented resolution, particularly in diseases lacking animal models, such as SUPS. We performed scRNA-seq to analyze tumor tissues and paracancer tissues from a SUPS patient. We identified the cell types and the corresponding marker genes in this SUPS case. We further showed that CD8+ exhausted T cells and Tregs highly expressed PDCD1, CTLA4 and TIGIT. Thus, PDCD1, CTLA4 and TIGIT were identified as potential targets in this case. We applied copy number karyotyping of aneuploid tumors (CopyKAT) to distinguish malignant cells from normal cells in fibroblasts. Our study identified eight malignant fibroblast subsets in SUPS with distinct gene expression profiles. C1-malignant Fibroblast and C6-malignant Fibroblast in the TME play crucial roles in tumor growth, angiogenesis, metastasis and immune response. Hence, targeting malignant fibroblasts could represent a potential strategy for this SUPS therapy. Intervention via tirelizumab enabled disease control, and immune checkpoint inhibitors (ICIs) of PD-1 may be considered as the first-line option in patients with SUPS. Taken together, scRNA-seq analyses provided a powerful basis for this SUPS treatment, improved our understanding of complex human diseases, and may afforded an alternative approach for personalized medicine in the future.

Structural insight into the bulge-containing KRAS oncogene promoter G-quadruplex bound to berberine and coptisine.

KRAS is one of the most highly mutated oncoproteins, which is overexpressed in various human cancers and implicated in poor survival. The G-quadruplex formed in KRAS oncogene promoter (KRAS-G4) is a transcriptional modulator and amenable to small molecule targeting. However, no available KRAS-G4-ligand complex structure has yet been determined, which seriously hinders the structure-based rational design of KRAS-G4 targeting drugs. In this study, we report the NMR solution structures of a bulge-containing KRAS-G4 bound to berberine and coptisine, respectively. The determined complex structure shows a 2:1 binding stoichiometry with each compound recruiting the adjacent flacking adenine residue to form a "quasi-triad plane" that stacks over the two external G-tetrads. The binding involves both π-stacking and electrostatic interactions. Moreover, berberine and coptisine significantly lowered the KRAS mRNA levels in cancer cells. Our study thus provides molecular details of ligand interactions with KRAS-G4 and is beneficial for the design of specific KRAS-G4-interactive drugs.

Diverse polycyclic polyprenylated acylphloroglucinols with anti-neuroinflammatory activity from Hypericum beanii.

A phytochemical investigation on the roots of Hypericum beanii resulted in the isolation of six new polycyclic polyprenylated acylphloroglucinols (PPAPs), hyperberlones A-F, along with fourteen known analogues. The structural characterization of these compounds was carried out by analyzing the HRESIMS data, 1D and 2D NMR spectroscopic data, electronic circular dichroism (ECD) calculations, and gauge-independent atomic orbital (GIAO) NMR calculations. Hyperberlone A (1) was a caged PPAP with a rare tricyclo[4.3.1.03,8]decane carbon skeleton. It was deduced to be biosynthetically generated from hyperbeanol C (8) through key Paternò-Büchi reaction, radical cascade cyclizations, and retro-aldol reaction. Compounds 4, 6, 7, 9, 14, and 16 exhibited significant nitric oxide (NO) production inhibitory effects in lipopolysaccharide (LPS)-induced BV-2 microglial cells with IC50 values of 6.11-25.28 µM. Moreover, compound 4 significantly decreased the expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in LPS-induced BV-2 microglia, as well as the phosphorylation of JNK.

Design, synthesis, and biological evaluation of novel diphenylamine derivatives as tubulin polymerization inhibitors targeting the colchicine binding site.

A novel series of diphenylamine derivatives were designed and synthesized, and their biological activities were evaluated. The anti-proliferative activities of the derivatives were tested against five human cancer cell lines (MCF-7, MDA-MB-231, A549, HeLa and HT29). Among them, compound 5f exhibited the promising anti-proliferative activity against HT29 cell lines with the IC50 value of 23 nM. Further biological studies depicted that compound 5f inhibited cancer cell migration, colony formation and angiogenesis. Besides, dynamics studies and molecular docking studies revealed that compound 5f inhibited tubulin polymerization which may be a result of the compound binding to the colchicine site of tubulin. Furthermore, compound 5f arrested HT29 cell cycle at G2/M phase, and induced HT29 cell apoptosis by upregulating cyclin B1, Bcl-2, Bax, Cleaved-caspase9, Cleaved-caspase3, PARP, Cleaved-PARP proteins, and downregulating p-cdc25c (S216), p-cdc2 (T15) proteins. Mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) were also determined to confirm the cell apoptosis process. Finally, compound 5f greatly inhibited the tumor growth in HT29 xenograft mice by 75.5% at 10 mg/kg. Meanwhile, compound 5f owned the good pharmacokinetic properties. All the results promised that 5f is of potential to act as an antitumor candidate and worthy of further investigation.

Photoaffinity Probe Reveals the Potential Target of Harringtonolide for Cancer Cell Migration Inhibition.

Harringtonolide (HO, 1) is a bioactive diterpenoid tropone isolated from Cephalotaxus harringtonia with antiproliferation activity. Until now there have been no reports to elucidate its anticancer mechanism. Herein we report the synthesis of HO-derived probes (10, 11, and 12) to identify the possible target of HO. As a result, the application of a novel photoaffinity alkyne-tagged probe from HO (compound 12) showed direct engagement between HO and receptor for activated C kinase 1 (RACK1). Furthermore, HO could suppress the epithelial-mesenchymal transition (EMT) process and inhibit activation of the FAK/Src/STAT3 signaling pathway in A375 cells. This study provides a groundwork for HO as an effective antitumor agent that targets RACK1 to suppress cancer cell migration.

Hyperpatulones C-G, new spirocyclic polycyclic polyprenylated acylphloroglucinols from the leaves of Hypericum patulum.

Five new spirocyclic polycyclic polyprenylated acylphloroglucinols, Hyperpatulones C-G (1-5), were obtained from the leaves of Hypericum patulum. Their structures were characterized by the comprehensive analysis of their IR, NMR, CD spectra and HRESIMS data. All the new compounds were evaluated for the α-glycosidase inhibitory activities. Among them, compounds 3-5 showed α-glucosidase inhibitory activities, with IC50 values of 14.06-37.69 µM.

Cephalotaxine-type alkaloids from the seeds of Cephalotaxus fortunei and their cytotoxic activities.

Six new Cephalotaxus alkaloids, including five cephalotaxine-type alkaloids, and one homoerythrina-type alkaloid, along with six known analogues, were isolated from the seeds of Cephalotaxus fortunei. Their structures were elucidated by combination of spectroscopic data analyses, time-dependent density functional theory (TDDFT) ECD calculation, and single-crystal X-ray diffraction. Cephalofortine B represents the first example of C-5 epi-cephalotaxine-type alkaloid. All isolated compounds were tested for cytotoxicities against HCT-116, A375, and SK-Mel-28 cell lines. Cephalofortine E showed moderate activity against HCT-116 cell line, with an IC50 value of 7.46 ± 0.77 µM.

Avicularin suppresses cartilage extracellular matrix degradation and inflammation via TRAF6/MAPK activation.

BACKGROUND: Osteoarthritis (OA) is an intractable degenerative disease of the whole joint, which is characterized by synovitis inflammation, cartilage damage, and chronic pain. Tumor necrosis factor receptor (TNFR)-associated factor 6 (TRAF6) performs an important role in OA. PURPOSE: We aim to investigate avicularin to protect cartilage extracellular matrix degradation (ECM) and suppresses inflammation both in rat and human chondrocytes. METHODS: 5-Ethynyl-2'-deoxyuridine (EdU) staining, Quantitative real-time PCR, TRAF6 plasmid transfection, Western blot, Measurement of nitric oxide (NO), ROS detection and Immunofluorescence were utilized in vitro. micro-CT scanning, Safranin O-Fast Green, toluidine blue and immunohistochemistry staining were performed in vivo. RESULTS: In vitro, avicularin attenuates the degradation of ECM and inflammation, which could inhibit the activation of TRAF6/MAPK pathway via targeting TRAF6. Increased MMP3 and MMP13 expressions and decreased Aggrecan and Collagen Ⅱ levels were observed in anterior cruciate ligament transection (ACLT) induced osteoarthritic rats. Interestingly, intra-articular injection of avicularin attenuates this phenomenon. CONCLUSIONS: Taken together, our results indicate that avicularin suppresses cartilage extracellular matrix degradation and inflammation via TRAF6/MAPK activation by targeting TRAF6. These observations identify TRAF6 as a relevant drug target, and avicularin may as a potential therapeutic agent in osteoarthritis.