The microbial biosynthesis of noncanonical terpenoids.

Terpenoids are a class of structurally complex, naturally occurring compounds found predominantly in plant, animal, and microorganism secondary metabolites. Classical terpenoids typically have carbon atoms in multiples of five and follow well-defined carbon skeletons, whereas noncanonical terpenoids deviate from these patterns. These noncanonical terpenoids often result from the methyltransferase-catalyzed methylation modification of substrate units, leading to irregular carbon skeletons. In this comprehensive review, various activities and applications of these noncanonical terpenes have been summarized. Importantly, the review delves into the biosynthetic pathways of noncanonical terpenes, including those with C6, C7, C11, C12, and C16 carbon skeletons, in bacteria and fungi host. It also covers noncanonical triterpenes synthesized from non-squalene substrates and nortriterpenes in Ganoderma lucidum, providing detailed examples to elucidate the intricate biosynthetic processes involved. Finally, the review outlines the potential future applications of noncanonical terpenoids. In conclusion, the insights gathered from this review provide a reference for understanding the biosynthesis of these noncanonical terpenes and pave the way for the discovery of additional unique and novel noncanonical terpenes. KEY POINTS: •The activities and applications of noncanonical terpenoids are introduced. •The noncanonical terpenoids with irregular carbon skeletons are presented. •The microbial biosynthesis of noncanonical terpenoids is summarized.

Loureirin A Promotes Cell Differentiation and Suppresses Migration and Invasion of Melanoma Cells via WNT and AKT/mTOR Signaling Pathways.

Resina Draconis is a traditional Chinese medicine, with the in-depth research, its medicinal value in anti-tumor has been revealed. Loureirin A is extracted from Resina Draconis, however, research on the anti-tumor efficacy of Loureirin A is rare. Herein, we investigated the function of Loureirin A in melanoma. Our research demonstrated that Loureirin A inhibited the proliferation of and caused G0/G1 cell cycle arrest in melanoma cells in a concentration-dependent manner. Further study showed that the melanin content and tyrosinase activity was enhanced after Loureirin A treatment, demonstrated that Loureirin A promoted melanoma cell differentiation, which was accompanied with the reduce of WNT signaling pathway. Meanwhile, we found that Loureirin A suppressed the migration and invasion of melanoma cells through the protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway. Taken together, this study demonstrated for the first time the anti-tumor effects of Loureirin A in melanoma cells, which provided a novel therapeutic strategy against melanoma.

The circHMGCS1-miR-205-5p-ErBB3 axis mediated the Sanggenon C-induced anti-proliferation effects on human prostate cancer.

Prostate cancer (PCa) is associated with a high incidence and metastasis rate globally, resulting in an unsatisfactory prognosis and a huge economic burden due to the current deficient of therapeutic strategies. As the most abundant component of Cortex Mori, Sanggenon C (SC) is well known to possess bioactivities in tumors, but its mechanism is poorly understood. Consequently, we attempted to investigate whether SC could modulate circular RNA(s) levels and hence anti-PCa development. We found that SC dramatically promoted cell apoptosis and induced G0/G1 phase arrest in PCa cell lines via the circHMGCS1-miR-205-5p-ErBB3 axis. In brief, circHMGCS1 is highly expressed in PCa and is positively correlated with the degree of malignancy. Over-expression of circHMGCS1 is not only associated with the proliferation of PCa cells but also blocks SC-induced pro-apoptotic effects. As a verified sponge of circHMGCS1, miR-205-5p is down-regulated in PCa tumors, which negatively regulates PCa cell proliferation by modulating ErBB3 expression. After miR-205-5p mimics or inhibitors were used to transfect PCa cells, the effects of circHMGCS1 OE and SC on PCa cells were completely diminished. Similar to miR-205-5p inhibitors, siErBB3 could oppose SC-triggered pro-apoptotic effects on PCa cells. All these results were confirmed in vivo. Together, SC exerts its anti-tumor effects on PCa by inhibiting circHMGCS1 expression and results in the latter losing the ability to sponge miR-205-5p. Subsequently, unfettered miR-205-5p could mostly down-regulate ErBB3 expression by binding to the 5'UTR of ErBB3 mRNA, which eventually resulted in PCa cell cycle arrest and pro-apoptosis.

Advances in the optimization of central carbon metabolism in metabolic engineering.

Central carbon metabolism (CCM), including glycolysis, tricarboxylic acid cycle and the pentose phosphate pathway, is the most fundamental metabolic process in the activities of living organisms that maintains normal cellular growth. CCM has been widely used in microbial metabolic engineering in recent years due to its unique regulatory role in cellular metabolism. Using yeast and Escherichia coli as the representative organisms, we summarized the metabolic engineering strategies on the optimization of CCM in eukaryotic and prokaryotic microbial chassis, such as the introduction of heterologous CCM metabolic pathways and the optimization of key enzymes or regulatory factors, to lay the groundwork for the future use of CCM optimization in metabolic engineering. Furthermore, the bottlenecks in the application of CCM optimization in metabolic engineering and future application prospects are summarized.

Gamabufotalin Induces Apoptosis and Cytoprotective Autophagy through the mTOR Signaling Pathway in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a malignant tumor with a high rate of recurrence and a poor prognosis. Here, we investigated the effect and the potential antitumor mechanism of Gamabufotalin (CS-6) against HCC. Our results show that CS-6 strikingly reduced cell viability, inhibited colony formation, and promoted apoptosis in Hep3B and Huh7 cells. In vivo, CS-6 inhibited HCC xenograft tumor growth with no toxicity to normal tissues. Mechanistically, we found that CS-6 could induce cytoprotective autophagy through the mTOR-ULK1 signaling pathway through downregulation of p62 and upregulation of LC3 II/LC3 I. Meanwhile, CS-6 activated caspase-3 and PARP mediated apoptosis, and the caspase inhibitor Z-VAD-FMK blocked the CS-6-induced cell death in HCC cells. Moreover, autophagy and apoptosis were found to have antagonistic effects in Hep3B and Huh7 cells. Both the autophagy inhibitor chloroquine (CQ) and the mTOR activator MHY1485 blocked autophagy and further enhanced CS-6-induced apoptosis. Taken together, we demonstrated for the first time that CS-6 promotes apoptosis and cytoprotective autophagy through the mTOR signaling pathway in HCC, which proposes a novel strategy for HCC therapy.

Advances in the biosynthesis and metabolic engineering of rare ginsenosides.

Rare ginsenosides are the deglycosylated secondary metabolic derivatives of major ginsenosides, and they are more readily absorbed into the bloodstream and function as active substances. The traditional preparation methods hindered the potential application of these effective components. The continuous elucidation of ginsenoside biosynthesis pathways has rendered the production of rare ginsenosides using synthetic biology techniques effective for their large-scale production. Previously, only the progress in the biosynthesis and biotechnological production of major ginsenosides was highlighted. In this review, we summarized the recent advances in the identification of key enzymes involved in the biosynthetic pathways of rare ginsenosides, especially the glycosyltransferases (GTs). Then the construction of microbial chassis for the production of rare ginsenosides, mainly in Saccharomyces cerevisiae, was presented. In the future, discovery of more GTs and improving their catalytic efficiencies are essential for the metabolic engineering of rare ginsenosides. This review will give more clues and be helpful for the characterization of the biosynthesis and metabolic engineering of rare ginsenosides. KEY POINTS: • The key enzymes involved in the biosynthetic pathways of rare ginsenosides are summarized. • The recent progress in metabolic engineering of rare ginsenosides is presented. • The discovery of glycosyltransferases is essential for the microbial production of rare ginsenosides in the future.

Hernandezine Regulates Proliferation and Autophagy-Induced Apoptosis in Melanoma Cells.

Hernandezine is isolated from an herbal medicine that selectively inhibits multidrug resistance and improves the efficacy of drugs for cancer treatment. To date, no studies on hernandezine in melanoma have been conducted. In this study, hernandezine was found to inhibit proliferation and induce apoptosis in melanoma A375 cells and B16 cells. In hernandezine-treated melanoma cells, G0/G1 cycle arrest occurred accompanied by significantly downregulated levels of phosphorylated JAK2 and STAT3. In addition, the cycle arrest could be enhanced by AG490 (JAK2 inhibitor), suggesting that the JAK2/STAT3 pathway is involved in cell cycle regulation in hernandezine-treated melanoma cells. Hernandezine-treated melanoma cells exhibited autophagy-specific structures, autophagy markers (LC3II/LC3-I), and autophagic flow over time. Moreover, 3-MA (autophagy inhibitor) significantly inhibited apoptosis, indicating that hernandezine promotes apoptosis by inducing autophagy. Combined with differential expression of P-AMPK, P-ACC (downstream targets of adenine monophosphate activated protein kinase, AMPK), and P-p70S6K (downstream targets of mammalian target of rapamycin, mTOR) and significant inhibition of apoptosis by AMPK inhibitor complex C (CC) in hernandezine-treated melanoma cells suggested that hernandezine could induce autophagy via the AMPK-mTOR pathway, thereby inducing apoptosis. This study first analyzed the effect of melanoma cells by hernandezine and provided a theory for hernandezine in the treatment of melanoma.

Sanggenon C Suppresses Tumorigenesis of Gastric Cancer by Blocking ERK-Drp1-Mediated Mitochondrial Fission.

Sanggenon C is a flavonoid extracted from the root bark of white mulberry, which is a traditional Chinese medicine with anti-inflammatory, antioxidative, and antitumor pharmacological effects. In this study, sanggenon C was found to inhibit human gastric cancer (GC) cell proliferation and colony formation, induce GC cell cycle arrest in the G0-G1 phase, and promote GC cell apoptosis. Moreover, sanggenon C was found to decrease the level of mitochondrial membrane potential in GC cells and inhibit mitochondrial fission. Mechanistically, RNA sequencing, bioinformatics analysis, and a series of functional analyses confirmed that sanggenon C inhibited mitochondrial fission to induce apoptosis by blocking the extracellular regulated protein kinases (ERK) signaling pathway, and constitutive activation of ERK significantly abrogated these effects. Finally, sanggenon C was found to suppress the growth of tumor xenografts in nude mice without obvious side effects to the vital organs of animals. This study reveals that sanggenon C could be a novel therapeutic strategy for GC treatment.

Picropodophyllin Inhibits the Proliferation of Human Prostate Cancer DU145 and LNCaP Cells via ROS Production and PI3K/AKT Pathway Inhibition.

The reactive oxygen species (ROS) and phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway play critical roles in the pathogenesis of prostate cancer by modulating cell proliferation. Picropodophyllin (PPP), an inhibitor of the insulin-like growth factor 1 receptor (IGF-1R), exerts significant antitumor effects via the PI3K/AKT signaling pathway. However, the effects of PPP on prostate cancer via ROS production and the PI3K/AKT signaling pathway remain elusive. Herein, we focused on examining the antitumor effects of PPP on DU145 and LNCaP human prostate cancer cells to determine the possible molecular mechanism. Our data indicated that the inhibitory effect of PPP on the proliferation of DU145 and LNCaP human prostate cancer cells was mediated by apoptosis induction and cell cycle blockade. Furthermore, PPP significantly influenced the expression of apoptosis-related, cell cycle, ROS production, and PI3K/AKT signaling proteins. These findings suggest that PPP can induce cell cycle arrest and apoptosis via the production of ROS and inhibition of PI3K/AKT signaling pathway, thereby suppressing the proliferation of prostate cancer cells.

Exploring the variation of bacterial community and nitrogen transformation functional genes under the pressure of heavy metals in different coastal mariculture patterns.

Equilibrium in microbial dynamics and nitrogen transformation in the sediment is critical for maintaining healthy mariculture environment. However, our understanding about the impact of heavy metals on the bacterial community and nitrogen transformation functional genes in different mariculture patterns is still limited. Here, we analyzed 30 sediment samples in the vertical distribution from three different mariculture patterns mainly include open mariculture zone (K), closed mariculture pond (F) and pristine marine area (Q). Illumina MiSeq Sequencing was applied to investigate the bacterial community and structure in the sediment. Quantitative polymerase chain reaction (qPCR) was used to determine the effect of heavy metals on nitrogen transformation functional genes. Results showed that bacterial community and structure varied greatly in different mariculture patterns. Chloroflexi, Proteobacteria and Desulfobacterota were predominant phyla in the coastal mariculture area. High concentrations of heavy metals mainly enriched in the up layer (5-40 cm) of the sediment in the mariculture zone. The abundance of functional genes in the closed mariculture pond was much higher than the open mariculture zone and pristine marine area. And the high abundance of nitrification and denitrification functional genes mainly accumulated at the depth from 5 cm to 40 cm. Heavy metals content such as Fe, Cr, Mn, Ni, As, Cd, Pb and nutrient content NH4+-N, NO3--N and NO2--N were highly associated with bacterial community and nitrogen transformation functional genes. This study comprehensively elaborated the effect of heavy metals on the bacterial community and nitrogen transformation functional genes in different coastal mariculture patterns, indicating the possible role of closed mariculture pond in reducing nitrogen transformation efficiency, which will provide useful information for preventing pollution risk in the mariculture area.

[Protective effect of safflower yellow injection against rat MIRI by TLR-NF-κB inflammatory pathway].

This study was to investigate the mechanism of safflower yellow injection for regulating inflammatory response against myocardial ischemia-reperfusion injury( MIRI) in rats. Male Wistar rats were randomly divided into sham operation group,model group,Hebeishuang group,safflower yellow injection high,medium and low dose groups. MIRI model was established by ligating left anterior descending coronary artery. Myocardial histopathological changes were observed by HE staining; myocardial infarct size was detected by TTC staining; content and changes of tumor necrosis factor-α( TNF-α) and interleukin-6( IL-6),serum creatine kinase( CK),aspartate aminotransferase( AST),and lactate dehydrogenase( LDH) were detected by biochemical method or enzyme-linked immunosorbent assay( ELISA). Western blot assay was used to detect the protein expression of Toll-like receptor 4( TLR4) and nuclear factor-κB( NF-κB p65) in myocardial tissues. The results showed that as compared with the sham operation group,the myocardial arrangement of the model group was disordered,with severe edemain the interstitial,significantly increased area of myocardial infarction,increased activities of AST,CK and LDH in serum,and significantly increased contents of TNF-α and IL-6; the expression levels of TLR4 and NF-κB( p65) protein in myocardial tissues were also increased. As compared with the model group,the myocardial tissues were arranged neatlyin the Hebeishuang group and safflower yellow injection high,medium and low dose groups; the edema was significantly reduced; the myocardial infarct size was significantly reduced; the serum AST,CK,LDH activity and TNF-α,IL-6 levels were significantly decreased,and the expression levels of TLR4 and NF-κB( p65) protein in myocardial tissues were decreased. As compared with the Hebeishuang group,the myocardial infarct size was larger in the safflower yellow injection high,medium and low dose groups; the activities of AST,CK and LDH in serum and the contents of TNF-α and IL-6 in serum were higher,but there was no statistically significant difference in the expression levels of TLR4 and NF-κB( p65) protein in tissues. It is suggested that safflower yellow injection has a significant anti-MIRI effect,and its mechanism may be related to the regulation of TLR-NF-κB pathway to inhibit inflammatory response.

Deoxyschizandrin Loaded Liposomes on the Suppression Lipid Accumulation in 3T3-L1 Adipocytes.

Deoxyschizandrin (DS) is a bioactive benzocyclooctadiene lignan found in the fruit of Schisandra chinensis. However, poor bioavailability and non-specificity of DS frequently caused low therapeutic efficacy. In the present study, DS-liposome (DS-lipo) was implemented to enhance the hepatic targeting and inhibition effects on adipocyte differentiation in 3T3-L1 cells. The formulations enabled encapsulation of as much as 24.14% DS. The DS-lipo prepared was about 73.08 nm, as measured by laser light scattering (LLS) morphology. In the visual field of a scanning electron microscope (SEM), the liposomes were spherical with similar size and uniform dispersion. Fluorescence live imaging study exhibited hepatic targeting of liposomes in vivo. Furthermore, High-Content Analysis (HCS) imaging microassay analyses revealed DS-lipo and DS reduced cytoplasmic lipid droplet in 3T3-L1 adipocytes, with the IC50 value of 8.68 µM and 31.08 µM, respectively. The lipid droplet accumulation inhibition rate of 10 µM DS-lipo was above 90%, which was even superior to the effect of 30 µM DS solution. The current findings suggest that DS-lipo was a therapeutic strategy for alleviating lipid-associated diseases and nonalcoholic fatty liver disease (NAFLD).

Kaempferide Protects against Myocardial Ischemia/Reperfusion Injury through Activation of the PI3K/Akt/GSK-3ß Pathway.

The aim of this study is to investigate both the efficacy and mechanism of action of kaempferide (Kae) as a therapy for the treatment of cardiovascular disease. A rat model of myocardial ischemia/reperfusion (I/R) injury was established by ligation of the left anterior descending coronary artery for 30 min followed by a 2 h perfusion. In our study, we show that Kae remarkably improved cardiac function, alleviated myocardial injury via a decrease in myocardial enzyme levels, and attenuated myocardial infarct size in a dose-dependent manner. In addition, preconditioning treatment with Kae was found to significantly decrease serum TNF-α, IL-6, C-reactive protein (CRP), MDA, and ROS levels, while it was found to increase serum levels of SOD. Nuclear factor erythroid 2-related factor 2 (Nrf2) and cleaved caspase-3 expression levels were observed to be downregulated, while phospho-Akt (p-Akt) and phospho-glycogen synthase kinase-3ß (p-GSK-3ß) expression levels were upregulated. However, cotreatment with LY294002 (a PI3K inhibitor) or TDZD-8 (a GSK-3ß inhibitor) was found to abolish the above cardioprotective effects observed with the Kae treatment. The data presented in this study provides evidence that Kae attenuates I/R-induced myocardial injury through inhibition of the Nrf2 and cleaved caspase-3 signaling pathways via a PI3K/Akt/GSK 3ß-dependent mechanism.

[Research on whole blending end-point evaluation method of Angong Niuhuang Wan based on QbD concept].

The blending end-point determination of Angong Niuhuang Wan (AGNH) is a key technology problem. The control strategy based on quality by design (QbD) concept proposes a whole blending end-point determination method, and provides a methodology for blending the Chinese materia medica containing mineral substances. Based on QbD concept, the laser induced breakdown spectroscopy (LIBS) was used to assess the cinnabar, realgar and pearl powder blending of AGNH in a pilot-scale experiment, especially the whole blending end-point in this study. The blending variability of three mineral medicines including cinnabar, realgar and pearl powder, was measured by moving window relative standard deviation (MWRSD) based on LIBS. The time profiles of realgar and pearl powder did not produce consistent results completely, but all of them reached even blending at the last blending stage, so that the whole proposal blending end point was determined. LIBS is a promising Process Analytical Technology (PAT) for process control. Unlike other elemental determination technologies such ICP-OES, LIBS does not need an elaborate digestion procedure, which is a promising and rapid technique to understand the blending process of Chinese materia medica (CMM) containing cinnabar, realgar and other mineral traditional Chinese medicine. This study proposed a novel method for the research of large varieties of traditional Chinese medicines..

Neuroprotective effects of liquiritin on cognitive deficits induced by soluble amyloid-ß1-42 oligomers injected into the hippocampus.

This study assessed the modulating effects of liquiritin against cognitive deficits, oxidative damage, and neuronal apoptosis induced by subsequent bilateral intrahippocampal injections of aggregated amyloid-ß1-42 (Aß1-42). This study also explored the molecular mechanisms underlying the above phenomena. Liquiritin was orally administered to rats with Aß1-42-induced cognitive deficits for 2 weeks. The protective effects of liquiritin on the learning and memory impairment induced by Aß1-42 were examined in vivo by using Morris water maze. The rats were then euthanized for further studies. The antioxidant activities of liquiritin in the hippocampus of the rats were investigated by biochemical and immunohistochemical methods. The apoptosis of the neurons was assessed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling assay. Liquiritin at doses of 50-100 mg/kg significantly improved the cognitive ability, restored the abnormal activities of glutathione peroxidase and superoxide dismutase, and decreased the levels of malondialdehyde，8-hydroxy-2'-deoxyguanosine and protein carbonyl in the hippocampus of rats with Alzheimer's disease. Moreover, neural apoptosis in the hippocampus of Aß1-42-treated rats was reversed by liquiritin. Liquiritin can significantly ameliorate Aß1-42-induced spatial learning and memory impairment by inhibiting oxidative stress and neural apoptosis.

Alternol induces an S-phase arrest of melanoma B16F0 cells.

Alternol is a novel compound purified from the fermentation products of a microorganism in the yew tree bark. This study looks at the effects of alternol on the proliferation and cell cycle distribution of mouse melanoma cells. The inhibition of cell proliferation and changes in cell cycle distribution were analysed by sulforhodamine B and flow cytometry assays, respectively. mRNA expression of cyclin A, cyclin-dependent kinase 2 (CDK2), proliferating cell nuclear antigen (PCNA) and CDK inhibitor1A (p21) were measured by real-time reverse transcription PCR (RT-PCR). The protein levels of cyclin A, CDK2 and PCNA were analysed by Western blot analysis. p21 was measured by ELISA. Alternol treatment caused a significant decrease in the proliferation rate of B16F0 and B16F10 cells, which were significantly arrested in S phase, but this treatment had less effect on normal human embryonic kidney 293T cells. The mechanism by which alternol inhibits B16F0 proliferation in vitro may be associated with the inhibition of CDK2 and PCNA, and the activation of p21.

Treatment with lutein provides neuroprotection in mice subjected to transient cerebral ischemia.

Lutein is known to be a nonprovitamin A carotenoid found in broccoli and spinach. The aim of present study was to investigate whether lutein can protect brain against ischemic injury by reducing oxidative stress. Male ICR mice were randomly divided into five experimental groups: model group, sham group, lutein high, middle, and low-dose groups (30, 15, and 7.5 mg/kg). Mice were subjected to a 2-h middle cerebral artery occlusion followed by reperfusion for 22 h. The reduced glutathione/oxidized glutathione (GSH/GSSG) ratio, antioxidant enzyme activities, malondialdehyde (MDA), and the carbonyl content in oxidatively modified proteins in brain tissue were determined with colorimetric method. The 8-hydroxy deoxyguanosine (8-OHdG) expression was measured by immunohistochemistry assay, and the neuron apoptosis was detected by TdT-mediated dUTP nick end labeling assay. Then, the neurological deficit scores were measured at last. Treatment of lutein significantly elevated the ratio of GSH/GSSG as well as activities of superoxide dismutase, glutathione peroxidase, and catalase and obviously decreased the contents of MDA, brain carbonyl, the expression of 8-OHdG, the number of apoptotic cells, and neurological deficit scores. Our results demonstrate that administration of lutein affords strong neuroprotective effect against transient cerebral ischemic injury and that the effect might be associated with its antioxidant property.

Research on drug treatment and the novel signaling pathway of chronic atrophic gastritis.

BACKGROUND: Chronic atrophic gastritis (CAG) is a global digestive system disease and one of the important causes of gastric cancer. The incidence of CAG has been increasing yearly worldwide. PURPOSE: This article reviews the latest research on the common causes and future therapeutic targets of CAG as well as the pharmacological effects of corresponding clinical drugs. We provide a detailed theoretical basis for further research on possible methods for the treatment of CAG and reversal of the CAG process. RESULTS: CAG often develops from chronic gastritis, and its main pathological manifestation is atrophy of the gastric mucosa, which can develop into gastric cancer. The drug treatment of CAG can be divided into agents that regulate gastric acid secretion, eradicate Helicobacter. pylori (H. pylori), protect gastric mucous membrane, or inhibit inflammatory factors according to their mechanism of action. Although there are limited specific drugs for the treatment of CAG, progress is being made in defining the pathogenesis and therapeutic targets of the disease. Growing evidence shows that NF-κB, PI3K/AKT, Wnt/ ß-catenin, MAPK, Toll-like receptors (TLRs), Hedgehog, and VEGF signaling pathways play an important role in the development of CAG.

A novel polysaccharide isolated from Coriolus versicolor polarizes M2 macrophages into an M1 phenotype and reversesits immunosuppressive effect on tumor microenvironment.

Converting M2 macrophages into an M1 phenotype in the tumor microenvironment, provides a new direction for tumor treatment. Here, we further report CVPW-1, a new polysaccharide of 1.03 × 106 Da that was isolated from Coriolus versicolor. Its monosaccharide was composed of mannose, glucose, and galactose at a ratio of 1.00:8.73:1.68. The backbone of CVPW-1 was composed of (1 â†’ 3)-linked α-D-Glcp residues and (1 â†’ 3,6)-linked α-D-Glcp residues that branched at O-6. The branch consisted of (1 â†’ 6)-linked α-D-Glcp residues and (1 â†’ 4)-linked α-D-Glap, and some branches were terminated with (1→)-linked ß-D-Manp residues according to the results of HPLC, FT-IR, GC-MS, 1D and 2D NMR. Meanwhile, CVPW-1 could polarize M2 macrophages to M1 phenotypein vitro by binding to TLR4 and inducing the activation of Akt, JNK and NF-κB. This process involved reversing the functional inhibition of CD8+ T lymphocytes by inhibiting the expression of TREM2 in M2 macrophages. The in vivo experiments showed that oral administration of CVPW-1 could inhibit the growth of tumor in mice and polarize TAMs to M1 phenotype. Thus, the novel polysaccharide CVPW-1 from Coriolus versicolor might activate a variety of immune cells and then play an anti-tumor role. These results demonstrated that CVPW-1 could be developed as a potential immuno-oncology treatment reagent.

Hernandezine acts as a CDK4 suppressor inhibiting tumor growth by the CDK4/PKM2/NRF2 axis in colon cancer.

BACKGROUND: The cyclin-dependent kinase 4 (CDK4) interacts with its canonical and non-canonical substrates modulating the cell cycle in tumor cells. However, the potential substrates and the beyond-cell-cycle-regulated functions of CDK4 in colon cancer (CC) are still unknown. Hernandezine (HER) is previously verified to induce G0/G1 phase arrest and autophagic cell death in human cancer cells, which implies that HER might target G0/G1 phase-related proteins, including CDK4. PURPOSE: The present study tried to investigate the glycolytic metabolism and oxidative stress functions of CDK4 in colon cancer. Furthermore, the inhibitory effects and potential binding sites of HER on CDK4, as well as its anti-tumor activity were investigated in CC cells. METHODS: The mass spectrometry assay was performed to identify potential endogenous substrates of CDK4 and the correlation between glycolytic metabolic rate and CDK4 level in COAD patient tissues. Meanwhile, after inhibiting the activity or the expression of CDK4, the binding capacity of CDK4 to PKM2 and NRF2 and the latter two protein distributions in cytoplasm and nucleus were detected in CC cells. In vitro, the regulatory effects of the CDK4-PKM2-NRF2 axis on glycolysis and oxidative stress were performed by ECAR, OCR, and ROS assay. The inhibitory effect of HER on CDK4 activity was explored in CC cells and the potential binding sites were predicted and testified in vitro. Furthermore, tumor growth inhibition of HER by suppressing the CDK4-PKM2-NRF2 axis was also investigated in vitro and in vivo. RESULTS: PKM2 and NRF2 were identified as endogenous substrates of CDK4 and, high-expressed CDK4 was associated with low-level glycolysis in COAD. In vitro, inactivated CDK4 facilitated CDK4-PKM2-NRF2 complex formation which resulted in 1) inhibited PKM2 activity and retarded the glycolytic rate; 2) cytoplasm-detained NRF2 failed to transcript anti-oxidative gene expressions and induced oxidant stress. Additionally, as a CDK4 inhibitor, HER developed triple anti-tumor effects including induced G0/G1 phase arrest, suppressed glycolysis, and disrupted the anti-oxidative capacity of CC cells. CONCLUSION: The results first time revealed that CDK4 modulated glycolytic and anti-oxidative capacity of CC cells via bound to its endogenous substrates, PKM2 and NRF2. Additionally, 140Asp145Asn amino acid sites of CDK4 were potential targets of HER. HER exerts anti-tumor activity by inhibited the activity of CDK4, promoted the CDK4-PKM2-NRF2 complex formation in the CC cells.