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The drug pair consisting of Sophora flavescens Aiton (Sophorae flavescentis radix, Kushen) and Coptis chinensis Franch. (Coptidis rhizoma, Huanglian), as described in Prescriptions for Universal Relief (Pujifang), is widely used to treat laxation. Matrine and berberine are the major active components of Kushen and Huanglian, respectively. These agents have shown remarkable anti-cancer and anti-inflammatory effects. A mouse model of colorectal cancer was used to determine the most effective combination of Kushen and Huanglian against anti-colorectal cancer. The results showed that the combination of Kushen and Huanglian at a 1:1 ratio exerted the best anti-colorectal cancer effect versus other ratios. Moreover, the anti-colorectal cancer effect and potential mechanism underlying the effects of matrine and berberine were evaluated by the analysis of combination treatment or monotherapy. In addition, the chemical constituents of Kushen and Huanglian were identified and quantified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). A total of 67 chemical components were identified from the Kushen–Huanglian drug pair (water extraction), and the levels of matrine and berberine were 129 and 232 µg/g, respectively. Matrine and berberine reduced the growth of colorectal cancer and relieved the pathological conditions in mice. In addition, the combination of matrine and berberine displayed better anti-colorectal cancer efficacy than monotherapy. Moreover, matrine and berberine reduced the relative abundance of Bacteroidota and Campilobacterota at phylum level and that of Helicobacter, Lachnospiraceae_NK4A136_group, Candidatus_Arthromitus, norank_f_Lachnospiraceae, Rikenella, Odoribacter, Streptococcus, norank_f_Ruminococcaceae, and Anaerotruncus at the genus level. Western blotting results demonstrated that treatment with matrine and berberine decreased the protein expressions of c-MYC and RAS, whereas it increased that of sirtuin 3 (Sirt3). The findings indicated that the combination of matrine and berberine was more effective in inhibiting colorectal cancer than monotherapy. This beneficial effect might depend on the improvement of intestinal microbiota structure and regulation of the RAS/MEK/ERK-c-MYC-Sirt3 signaling axis.

BackgroundThe activation of peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α) stimulates the transcription of the downstream target proteins, mitochondrial transcription factor A (TFAM) and nuclear respiratory factor 1 (NRF1), which induces mitochondrial biogenesis and promotes colorectal tumorigenesis. Agrimol B (Agr) is a constituent of Agrimonia pilosa Ledeb. that exerts anticancer effects. Herein, we aimed to investigate the antitumor activity of Agr and its mechanism of action.MethodsThe interaction between Agr and PGC-1α was predicted by molecular docking. After the treatment with different concentrations of Agr (0, 144, 288, and 576 nM), the cell viability, migration rate, proliferation rate, and apoptosis rate of human colon cancer HCT116 cells were determined. Mitochondrial activity, cellular reactive oxygen species (ROS), and mitochondrial membrane potential were assessed to measure the regulatory effect of Agr on mitochondrial function. Western blotting (WB) assay was used to examine the expression of PGC-1α, NRF1, and TFAM, as well as of the pro-apoptotic proteins, Bax and Caspase-3, and the antiapoptotic protein (Bcl-2). Finally, subcutaneous tumor xenograft model mice were used to evaluate the effect of Agr on colorectal cancer (CRC) in vivo.ResultsThe molecular docking results revealed a high likelihood of Agr interacting with PGC-1α. Agr inhibited the proliferation and migration of HCT116 cells, promoted ROS production and mitochondrial oxidative stress, inhibited mitochondrial activity, and decreased mitochondrial membrane potential. Agr induced cell apoptosis and, in combination with PGC-1α, impaired mitochondrial biogenesis and suppressed the expression of NRF1 and TFAM. Agr also suppressed the expression of Bcl-2 and Cleaved-Caspase-3 and increased the expression of Bax and Caspase-3. In addition, the in vivo antitumor effect and mechanism of Agr were confirmed by using a subcutaneous tumor xenograft mouse model.ConclusionsOur findings demonstrated that Agr regulates the expression of PGC-1α, thereby inducing mitochondrial dysfunction and promoting tumor cell apoptosis. This work highlights the potential of Agr as a promising therapeutic candidate in CRC.

AimsYes-associated protein (YAP), a downstream protein in the Hippo signaling pathway, plays an important role in tumor proliferation, including in hepatocellular carcinoma (HCC). α-hederin, a monodesmosidic triterpenoid saponin isolated from Fructus akebiae, displayed anti-cancer effects on several cancer cell lines but the precise mechanism has not been ascertained. In the present study, we explored the effects of α-hederin on cell proliferation and apoptosis in human HCC cell lines and the underlying mechanisms.Main MethodCell proliferation and apoptosis were assessed using 5-ethynyl-2’-deoxyuridine staining, colony formation, flow cytometry. The expression patterns of components of Hippo signaling pathway and apoptotic genes were further examined via RT-qPCR and immunoblotting. A xenograft tumor model in nude mice was used to evaluate the anti-HCC effects of α-hederin in vivo.Resultsα-hederin promoted the apoptosis and inhibited the proliferation of SMMC-7721 and HepG2 cells in vitro, and remarkably inhibited the tumor size and weight in the xenograft mouse model. Additionally, α-hederin increased the expression of pro-apoptosis proteins and suppressed the expression of anti-apoptosis proteins. Moreover, α-hederin treatment upregulated the expression of Hippo signaling pathway-related proteins and genes, while, effectively reduced the level of nuclear YAP, which resulted in the inhibition of proliferation and the induction of apoptosis of HCC cells. Finally, the effects of α-hederin on HCC cell proliferation and apoptosis were alleviated by XMU-MP-1, a Mst1/2 inhibitor in vitro.SignificanceWe identified α-hederin is a novel agonist of Hippo signaling pathway and possesses an anti-HCC efficacy through inhibiting YAP activity.

BackgroundExosomes are extracellular vesicles secreted by most cells to deliver functional cargoes to recipient cells. MicroRNAs (miRNAs) constitute a significant part of exosomal contents. The ease of diffusion of exosomes renders them speedy and highly efficient vehicles to deliver functional molecules. Cancer cells secrete more exosomes than normal cells. Reports have showed that exosomal miRNAs of cancer cells facilitate cancer progression. Yet the complexity of cancer dictates that many more functional exosomal miRNAs remain to be discovered.MethodsIn this study, we analyzed miRNA expression profiles of tissue and plasma exosome samples collected from 10 colorectal cancer (CRC) patients and 10 healthy individuals. We focused on hsa-miR-101-3p (101-3p), a profoundly up-regulated miRNA enriched in plasma exosomes of patients bearing CRC. We performed target analysis of 101-3p and pursued functional studies of this microRNA in two colorectal cancer cell lines, namely HCT116 and SW480.ResultsOur results indicated that inhibiting 101-3p slowed cell growth and retarded cell migration in vivo in two colorectal cancer cell lines. Target analysis showed that Homeodomain-interacting protein kinase (HIPK3) is a target of miR-101-3p. HCT116 and SW480 cells stably overexpressing HIPK3 showed increased level of phosphorylated FADD, as well as retarded cell growth, migration, and increased sensitivity to 5-FU. In-depth analysis revealed increased mitochondrial membrane potential upon HIPK3 overexpression along with increased production of reactive oxygen species, number of mitochondria, and expression of respiratory complexes. Measurements of glycolytic parameters and enzymes revealed decreased level of glycolysis upon HIPK3 overexpression in these two cell lines. Xenograft model further confirmed a profoundly improved potency of the synergistic treatment combining both 5-FU and 101-3p inhibitor compared to 5-FU alone.ConclusionThis study unraveled an oncogenic nature of the exosomal 101-3p and suggested a relationship between the 101-3p-HIPK3 axis and metabolic homeostasis in colorectal cancer. Expression level of 101-3p is positively correlated with glycolytic capacity in CRC and therefore 101-3p itself is an oncomiR. Combining 101-3p inhibitor with chemotherapeutic agents is an effective strategy against CRC.

MAPKAPK2 (MK2), the direct substrate of p38 MAPK, has been well-acknowledged as an attractive drug target for cancer therapy. However, few studies have assessed the functions of it in multiple myeloma (MM). In the present study, MK2 expression of MM patients was analyzed by gene expression profiling (GEP) and array-based comparative genomic hybridization (aCGH). Several experiments in vitro including MTT assay, Western blot and flow cytometry analysis were performed to identify the function of MK2 in MM. In addition, we conducted mouse survival experiments to explain the effects of MK2 on MM in vivo. mRNA level of MK2 and chromosomal gain of MK2 locus in MM cells significantly increased compared to normal samples. Furthermore, MM patients with high expression of MK2 were associated with a poor outcome. Follow-up studies showed that MK2 exerted a remarkably positive effect on MM cell proliferation and drug-resistance. Further exploration focusing on MK2 inhibitor IV revealed its inhibitory action on MM growth and drug-resistance, as well as improving survival in mouse models. In addition, a combination of MK2 inhibitor IV and the key MM therapeutic agents including bortezomib, doxorubicin, or dexamethasone facilitated curative effects on inhibiting MM cell proliferation. Taken together, our study reveals the clinical relevance of MK2 inhibition in MM and demonstrates that targeting MK2 may afford a new therapeutic approach to MM.