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Ethnopharmacological relevance Dendrobium chrysotoxum Lindl is a cultivation of Dendrobium which belongs to the family of Orchidaceae. D. chrysotoxum Lindl is a traditional Chinese medicine with a wide range of clinical applications including tonic, astringent, analgesic and anti-inflammatory properties as early as the 28th century B.C. Erianin is a representative index component for the quality control of the D. chrysotoxum Lindl, which is included in the Pharmacopoeia of the People's Republic of China (2020 version). Aim of the study To clarify the anti-tumour mechanisms of erianin in vitro and in vivo. Materials and methods We detected the anti-tumour activity of erianin using in vitro HeLa cell models and in vivo cervical cancer xenograft models. We performed MTT, western blot, RT-PCR, homology modeling, flow cytometry, and immunoprecipitation assays to study the proteins, genes, and pathways related to erianin's anti-tumour activity. LysoTracker Red staining was performed to detect lysosome function. Transwell, wound healing, tube formation, colony formation and EdU labelling assays were performed to determine cell proliferation, migration and invasion abilities, respectively. Cytotoxic T lymphocytes ability was confirmed using HeLa/T-cell co-culture model. Results Experimental data demonstrated that erianin inhibited PD-L1 expression and induced the lysosomal degradation of PD-L1. Erianin suppressed HIF-1α synthesis through mTOR/p70S6K/4EBP1 pathway, and inhibited RAS/Raf/MEK/MAPK-ERK pathway. Immunoprecipitation experiments demonstrated that erianin reduced the interaction between RAS and HIF-1α. Experiments using a co-cultivation system of T cells and HeLa cells confirmed that erianin restored cytotoxic T lymphocytes ability to kill tumour cells. Erianin inhibited PD-L1–mediated angiogenesis, proliferation, invasion and migration. The anti-proliferative effects of erianin were supported using in vivo xenotransplantation experiments. Conclusions Collectively, these results revealed previously unknown properties of erianin and provided a new basis for improving the efficacy of immunotherapy against cervical cancer and other malignant tumours through PD-L1.

Ethnopharmacological relevance The use of Panax ginseng C.A.Mey. in traditional Chinese medicine dates back to about 5000 years ago thanks to its several beneficial and healing properties. Panaxadiol is a triterpenoid sapogenin monomer found in the roots of Panax ginseng C.A.Mey. and has been proven to have various bio-activities such as anti-inflammatory, anti-tumour and neuroprotective effects. Aim of the study The present study focuses on investigating the inflammation inhibitory effect and mechanism of panaxadiol by regulating zinc finger protein 91-regulated activation of non-canonical caspase-8 inflammasome and MAPKs in macrophages. Materials and methods In vitro, the underlying mechanisms by which panaxadiol inhibits ZFP91-regulated IL-1β expression were investigated using molecular docking, western blotting, RT-PCR, ELISA, immunofluorescence, and immunoprecipitation assays. In vivo, colitis was induced by oral administration of DSS in drinking water, and peritonitis was induced by an intraperitoneal injection of alum. Recombinant adeno-associated virus (AAV serotype 9) vector was used to establish ZFP91 knockdown mouse. Results We confirmed that panaxadiol inhibited IL-1β secretion by suppressing ZFP91 in macrophages. Further analysis revealed that panaxadiol inhibited IL-1β secretion by suppressing ZFP91-regulated activation of non-canonical caspase-8 inflammasome. Meanwhile, panaxadiol inhibited IL-1β secretion by suppressing ZFP91-regulated activation of MAPKs. In vivo, prominent anti-inflammatory effects of panaxadiol were demonstrated in a DSS induced acute colitis mouse model and in an alum-induced peritonitis model by suppressing ZFP91-regulated secretion of inflammatory mediators, consistent with the results of the AAV-ZFP91 knockdown in mice. Conclusions We report for the first time that panaxadiol inhibited IL-1β secretion by suppressing ZFP91-regulated activation of non-canonical caspase-8 inflammasome and MAPKs, providing evidence for anti-inflammation mechanism of panaxadiol treatment for inflammatory diseases.

Ethnopharmacological relevance Glycyrrhiza glabra L., a traditional medicinal, has a history of thousands of years. It is widely used in clinic and has been listed in Chinese Pharmacopoeia. Licochalcone A is a phenolic chalcone compound and a characteristic chalcone of Glycyrrhiza glabra L. It has many pharmacological activities, such as anti-cancer, anti-inflammatory, anti-viral and anti-angiogenic activities. Aim of the study In this study, we explored the anti-tumor activity and potential mechanism of licochalcone A in vitro and in vivo. Materials and methods In vitro, the mechanism of licochalcone A at inhibiting PD-L1 expression was investigated by molecular docking, western blotting, RT-PCR, flow cytometry, immunofluorescence and immunoprecipitation assays. The co-culture model of T cells and tumor cells was used to detect the activity of cytotoxic T lymphocytes. Colony formation, EdU labelling and apoptosis assays were used to detect changes in cellular proliferation and apoptosis. In vivo, anti-tumor activity of licochalcone A was assessed in a xenograft model of HCT116 cells. Results In the present study, we found that licochalcone A suppressed the expression of programmed cell death ligand-1 (PD-L1), which plays a key role in regulating the immune response. In addition, licochalcone A inhibited the expressions of p65 and Ras. Immunoprecipitation experiment showed that licochalcone A suppressed the expression of PD-L1 by blocking the interaction between p65 and Ras. In the co-culture model of T cells and tumor cells, licochalcone A pretreatment enhanced the activity of cytotoxic T lymphocytes and restored the ability to kill tumor cells. In addition, we showed that licochalcone A inhibited cell proliferation and promoted cell apoptosis by targeting PD-L1. In vivo xenograft assay confirmed that licochalcone A inhibited the growth of tumor xenografts. Conclusion In general, these results reveal the previously unknown properties of licochalcone A and provide new insights into the anticancer mechanism of this compound.

Ethnopharmacological relevance Curcuma wenyujin is a Chinese traditional herbal medicine that is commonly used as an anti-oxidant, anti-proliferative, and anti-tumorigenic agent. Curcumol is a representative index component for the quality control of the essential oil of Curcuma wenyujin, which is currently used as an anti-cancer drug, and is included in the State Pharmacopoeia Commission of the People's Republic of China (2005). However, the mechanisms of action and molecular functions of curcumol are not yet fully elucidated. Aim of the study This study aimed to identify new effects of curcumol from the perspective of cancer immunotherapy. Materials and methods The underlying mechanism of the inhibition of programmed cell death-ligand 1 (PD-L1) activation by curcumol was investigated in vitro via homology modeling, molecular docking experiments, luciferase reporter assays, MTT assays, RT-PCR, western blotting, and immunofluorescence assays. Changes in cellular proliferation, angiogenesis, and the tumor-killing activity of T-cells were analyzed via EdU labeling, colony formation, flow cytometry, wound-healing, Matrigel Transwell invasion, tube formation, and T-cell killing. The anti-tumor activity of curcumol was assessed in vivo in a murine xenograft model using Hep3B cells. Results Curcumol reduced the expression of phosphorylated signal transducer and activator of transcription 3 (p-STAT3) via JAK1, JAK2, and Src pathways and inhibited hypoxia-inducible factor-1α (HIF-1α) protein synthesis via mTOR/p70S6K/eIF4E and MAPK pathways. Furthermore, we revealed crosstalk between STAT3 and HIF-1α pathways, which collaboratively regulated PD-L1 activation, and that curcumol played a role in this regulation. Curcumol inhibited cell proliferation, S-phase progression, tube formation, invasion, and metastasis by inhibiting PD-L1. In addition, curcumol restored the activity of cytotoxic T-cells and their capacity for tumor cell killing by inhibiting PD-L1. In vivo experiments confirmed that curcumol inhibited tumor growth in a xenograft model. Conclusions These results illustrated that curcumol inhibits the expression of PD-L1 through crosstalk between HIF-1α and p-STAT3 (T705) signaling pathways in hepatic cancer. Thus, curcumol might represent a promising lead compound for the development of new targeted anti-cancer therapeutics.