Frizzled receptors (FZDs) in Wnt signaling: potential therapeutic targets for human cancers.

Frizzled receptors (FZDs) are key contributors intrinsic to the Wnt signaling pathway, activation of FZDs triggering the Wnt signaling cascade is frequently observed in human tumors and intimately associated with an aggressive carcinoma phenotype. It has been shown that the abnormal expression of FZD receptors contributes to the manifestation of malignant characteristics in human tumors such as enhanced cell proliferation, metastasis, chemotherapy resistance as well as the acquisition of cancer stemness. Given the essential roles of FZD receptors in the Wnt signaling in human tumors, this review aims to consolidate the prevailing knowledge on the specific status of FZD receptors (FZD1-10) and elucidate their respective functions in tumor progression. Furthermore, we delineate the structural basis for binding of FZD and its co-receptors to Wnt, and provide a better theoretical foundation for subsequent studies on related mechanisms. Finally, we describe the existing biological classes of small molecule-based FZD inhibitors in detail in the hope that they can provide useful assistance for design and development of novel drug candidates targeted FZDs.

Macrophages promote anti-androgen resistance in prostate cancer bone disease.

Metastatic castration-resistant prostate cancer (PC) is the final stage of PC that acquires resistance to androgen deprivation therapies (ADT). Despite progresses in understanding of disease mechanisms, the specific contribution of the metastatic microenvironment to ADT resistance remains largely unknown. The current study identified that the macrophage is the major microenvironmental component of bone-metastatic PC in patients. Using a novel in vivo model, we demonstrated that macrophages were critical for enzalutamide resistance through induction of a wound-healing-like response of ECM-receptor gene expression. Mechanistically, macrophages drove resistance through cytokine activin A that induced fibronectin (FN1)-integrin alpha 5 (ITGA5)-tyrosine kinase Src (SRC) signaling cascade in PC cells. This novel mechanism was strongly supported by bioinformatics analysis of patient transcriptomics datasets. Furthermore, macrophage depletion or SRC inhibition using a novel specific inhibitor significantly inhibited resistant growth. Together, our findings elucidated a novel mechanism of macrophage-induced anti-androgen resistance of metastatic PC and a promising therapeutic approach to treat this deadly disease.

Thymosin-α1 binds with ACE and downregulates the expression of ACE2 in human respiratory epithelia.

BACKGROUND: Thymosin-α1 has been implicated into the treatment of novel respiratory virus Coronavirus Disease 2019 (COVID-19), but the underlying mechanisms are still disputable. AIM: Herein we aimed to reveal a previously unrecognized mechanism that thymosin-α1 prevents COVID-19 by binding with angiotensin-converting enzyme (ACE), which was inspired from the tool of network pharmacology. METHODS: KEGG pathway enrichment of thymosin-α1 treating COVID-19 was analyzed by Database of Functional Annotation Bioinformatics Microarray Analysis, then core targets were validated by ligand binding kinetics assay and fluorometric detection of ACE and ACE2 enzymatic activity. The production of angiotensin I, angiotensin II, angiotensin (1-7) and angiotensin (1-9) were detected by enzyme linked immunosorbent assay. RESULTS: We found that thymosin-α1 impaired the expressions of angiotensin-converting enzyme 2 and angiotensin (1-7) of human lung epithelial cells in a dose-dependent way (p < 0.001). In contrast, thymosin-α1 had no impact on their ACE and angiotensin (1-9) expressions but significantly inhibited the enzymatic activity of ACE (p > 0.05). CONCLUSION: The bioinformatic findings of network pharmacology and the corresponding pharmacological validations have revealed that thymosin-α1 treatment could decrease ACE2 expression in human lung epithelial cells, which strengthens the potential clinical applications of thymosin-α1 to prevent severe acute respiratory syndrome coronavirus 2 infection.

The Two-Way Switch Role of ACE2 in the Treatment of Novel Coronavirus Pneumonia and Underlying Comorbidities.

December 2019 saw the emergence of the coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), which has spread across the globe. The high infectivity and ongoing mortality of SARS-CoV-2 emphasize the demand of drug discovery. Angiotensin-converting enzyme II (ACE2) is the functional receptor for SARS-CoV-2 entry into host cells. ACE2 exists as a membrane-bound protein on major viral target pulmonary epithelial cells, and its peptidase domain (PD) interacts SARS-CoV-2 spike protein with higher affinity. Therefore, targeting ACE2 is an important pharmacological intervention for a SARS-CoV-2 infection. In this review, we described the two-way switch role of ACE2 in the treatment of novel coronavirus pneumonia and underlying comorbidities, and discussed the potential effect of the ACE inhibitor and angiotensin receptor blocker on a hypertension patient with the SARS-CoV-2 infection. In addition, we analyzed the S-protein-binding site on ACE2 and suggested that blocking hot spot-31 and hot spot-353 on ACE2 could be a therapeutic strategy for preventing the spread of SARS-CoV-2. Besides, the recombinant ACE2 protein could be another potential treatment option for SARS-CoV-2 induced acute severe lung failure. This review could provide beneficial information for the development of anti-SARS-CoV-2 agents via targeting ACE2 and the clinical usage of renin-angiotensin system (RAS) drugs for novel coronavirus pneumonia treatment.

Combinational applicaton of silybin and tangeretin attenuates the progression of non-alcoholic steatohepatitis (NASH) in mice via modulating lipid metabolism.

Silybin (SB) is widely used to treat chronic liver diseases, especially this compound is much efficient for the treatments of alcoholic and non-alcoholic steatohepatitis (NASH). However, low bioavailability seriously limits wide-application of SB in biomedical niche. Prior to this study, we found that tangeretin (TG) could remarkably increase the bioavailability of SB by the inhibition of efflux transporters, which encourges us to therapeutical discovery of SB and TG combitional use against NASH. Here, we revealed that TG is capable of improving hepatic-protective activity of SB in mice with NASH by interfering liver oxidative stress, inflammation, and lipid accumulation. In addition, TG was observed to enhance the exposural level of SB in the plasma and liver of mice. Our metabolome assay confirmed that amino acid metabolism and lipid biosynthesis mostly accounted for combitional use of SB and TG to teat NASH in mice, basically biosynthesis of unsaturated fatty acids was mostly affected. Notably, significant inhibitions in fatty acid generating and transporting proteins such as G6PD, FABP4, LPL and CD36/FAT, and cholesterol metabolism enzyme CYP27A1 as well as nuclear transcription factors FXR, PPAR-γ, and LXR were illustrated to decipher therapeutic mechanisms of SB and TG against experimental NASH. Taken together, the strategy based combitional use of SB and TG has a potential-capacity to treat NASH.

Anti-influenza effect and action mechanisms of the chemical constituent gallocatechin-7-gallate from Pithecellobium clypearia Benth.

Host cdc2-like kinase 1 (CLK1) is responsible for the alternative splicing of the influenza virus M2 gene during influenza virus infection and replication that has been recognized as a potential anti-influenza virus target. In this study, we showed that gallocatechin-7-gallate (J10688), a novel CLK1 inhibitor isolated from Pithecellobium clypearia Benth, exerted potent anti-influenza virus activity in vivo and in vitro. ICR mice were intranasally infected with a lethal dose of H1N1. Administration of J10688 (30 mg·kg-1·d-1, iv, for 5 days) significantly increased the survival rate of the H1N1-infected mice to 91.67% and prolong their mean survival time from 5.83 ± 1.74 days to 13.66 ± 1.15 days. J10688 administration also slowed down body weight loss, significantly alleviated influenza-induced acute lung injury, reduced lung virus titer, elevated the spleen and thymus indexes, and enhanced the immunological function. We further explored its anti-influenza mechanisms in the H1N1-infected A549 cells: as a novel CLK1 inhibitor, J10688 (3, 10, 30 µmol/L) dose-dependently impaired synthesis of the viral proteins NP and M2, and significantly downregulated the phosphorylation of splicing factors SF2/ASF and SC35, which regulate virus M2 gene alternative splicing. As a novel CLK1 inhibitor with potent anti-influenza activity in vitro and in vivo, J10688 could be a promising antiviral drug for the therapy of influenza A virus infection.

Network pharmacology-based analysis of Chinese herbal Naodesheng formula for application to Alzheimer's disease.

Naodesheng (NDS) formula, which consists of Rhizoma Chuanxiong, Lobed Kudzuvine, Carthamus tinctorius, Radix Notoginseng, and Crataegus pinnatifida, is widely applied for the treatment of cardio/cerebrovascular ischemic diseases, ischemic stroke, and sequelae of cerebral hemorrhage, etc. At present, the studies on NDS formula for Alzheimer's disease (AD) only focus on single component of this prescription, and there is no report about the synergistic mechanism of the constituents in NDS formula for the potential treatment of dementia. Therefore, the present study aimed to predict the potential targets and uncover the mechanisms of NDS formula for the treatment of AD. Firstly, we collected the constituents in NDS formula and key targets toward AD. Then, drug-likeness, oral bioavailability, and blood-brain barrier permeability were evaluated to find drug-like and lead-like constituents for treatment of central nervous system diseases. By combining the advantages of machine learning, molecular docking, and pharmacophore mapping, we attempted to predict the targets of constituents and find potential multi-target compounds from NDS formula. Finally, we built constituent-target network, constituent-target-target network and target-biological pathway network to study the network pharmacology of the constituents in NDS formula. To the best of our knowledge, this represented the first to study the mechanism of NDS formula for potential efficacy for AD treatment by means of the virtual screening and network pharmacology methods.

[Network pharmacology study of Xiaoxuming Decoction based on vasodilatory and vasoconstrictory related GPCR targets].

In this study, bioinformatics methods such as molecular docking and network pharmacology were adopted to establish Xiaoxuming Decoction (XXMD) "compound-vasodilatory and vasoconstrictory related G protein-coupled receptors (GPCR) targets" network, then the vascular function regulatory effective components and the potential targets of XXMD were analyzed. Based on the XXMD herb sources, the chemical structures of the compounds were retrieved from the national scientific data sharing platform for population and health pharmaceutical information center, TCMSP database and the latest research literatures. The chemical molecular library was established after class prediction and screening for medicinal and metabolic properties. Then, five kinds of vasodilatory and vasoconstrictory related GPCR crystal structure including 5-HT receptors (5-HT1AR, 5-HT1BR), AT1R, ß2-AR, hUTR and ETB were retrieved from RCSB Protein Data Bank database or constructed by homology modeling of Discovery Studio 4.1 built-in modeling tools. After virtual screening by Libdock molecular docking, the highest rated 50 compounds of each target were collected and analyzed. The collected data were further used to construct and analyze the network by Cytoscape 3.4.0. The results showed that most of the chemical composition effects were associated with different vasodilatory and vasoconstrictory related GPCR targets, while a few effective components could be applied to multiple GPCR targets at the same time, therefore forming synergies and vasorelaxant effects of XXMD.

Actinomycin D synergistically enhances the cytotoxicity of CDDP on KB cells by activating P53 via decreasing P53-MDM2 complex.

The aim of this study is to investigate the synergism of low dose of actinomycin D (LDActD) to the cytotoxicity of cisplatin (CDDP) on KB cells. The role of P53 reactivation by LDActD in the synergism and its mechanism were further studied. Cell viability was determined by MTT assay. Apoptosis was determined by AnnexinV-FITC/PI staining. Mitochondrial membrane potential (MMP) was detected by JC-1 staining. Expression of proteins was detected by Western blotting (WB) and/or immunofluorescence (IF). Molecular docking of actinomycin D (ACTD) to Mouse double minute 2 homolog (MDM2) and Mouse double minute 2 homolog X (MDMX). MDMX was analyzed by Discovery Studio. The content of P53-MDM2 complex was detected by ELISA assay. The cytotoxicity of CDDP was increased by the combination of LDActD in kinds of cancer cells. Molecular docking showed strong interaction between ACTD and MDM2/MDMX. Meanwhile, LDActD significantly decreased P53-MDM2 complex. Significant increase of the apoptotic activity by the combination therapy in KB cells is P53 upregulated modulator of apoptosis (PUMA) dependent. In addition to the decrease in MMP, LDActD increased P53 regulated protein and decreased BCL-XL in KB cells. LDActD efficiently enhanced the cytotoxicity of CDDP in cancer cells and induced P53-PUMA-dependent and mitochondria-mediated apoptosis in KB cells. The reactivation of P53 was probably achieved by disturbing the interaction of P53 and MDM2/MDMX.

DL0410, a candidate for anti-Alzheimer disease with multiple targets in multiple pathways / 中国药理学与毒理学杂志

OBJECTIVE To find a promising candidate for anti- Alzheimer disease (AD) with multiple targets in multiple pathways. METHODS A series of classifiers were constructed for predicting the active compounds against 51 key targets toward Alzheimer disease (AD) using the multitarget-quantitative structure- activity relationships (mt- QSAR) method. While drug screening assays were established to evaluate the predicted active molecules. In addition, various cellular models and animal models related with AD were set up to further study the effects of the active compounds. RESULTS A system for the discovery of Multitarget-Directed Ligands against AD was set up and applied, the predicted active compounds were validated by the drug screening assays, and several active compounds with multiple targets were discovered. Among them, DL0410 exerted high activity on H3R, α7nAChR, AChE and ERα, also displayed the most significant effect in improving the ability of memory and learning in several AD animal models. The study on its action mechanisms showed that it's effect may partially through increasing neurotransmitter, inhibiting oxidative emergency, inhibiting the expression of APP, and promoting long- term potentiation. Besides, DL0410 is of more safety than the first- line clinical medicines. CONCLUSION DL0410 is a promising candidate for further development for AD treatment.

Effects of the Nrf2 Protein Modulator Salvianolic Acid A Alone or Combined with Metformin on Diabetes-associated Macrovascular and Renal Injury.

Nuclear factor E2-related factor 2 (Nrf2) is considered a promising target against diabetic complications such as cardiovascular diseases and diabetic nephropathy. Herein, we investigated the effects of a potential Nrf2 modulator, salvianolic acid A (SAA), which is a natural polyphenol, on diabetes-associated macrovascular and renal injuries in streptozotocin-induced diabetic mice. Given that lowering glucose is the first objective of diabetic patients, we also examined the effects of SAA combined with metformin (MET) on both complications. Our results showed that SAA significantly increased the macrovascular relaxation response to acetylcholine and sodium nitroprusside in diabetic mice. Interestingly, treatment with SAA alone only provided minor protection against renal injury, as reflected by minor improvements in impaired renal function and structure, despite significantly reduced oxidative stress observed in the diabetic kidney. We demonstrated that decreased oxidative stress and NF-κB p65 expression were associated with SAA-induced expression of Nrf2-responsive antioxidant enzymes heme oxygenase-1 (HO-1), NAD(P)H dehydrogenase (quinone) 1 (NQO-1), and glutathione peroxidase-1 (GPx-1) in vivo or in vitro, which suggested that SAA was a potential Nrf2 modulator. More significantly, compared with treatment with either SAA or MET alone, we found that their combination provided further protection against the macrovascular and renal injury, which was at least partly due to therapeutic activation of both MET-mediated AMP-activated protein kinase and SAA-mediated Nrf2/antioxidant-response element pathways. These findings suggested that polyphenol Nrf2 modulators, especially combined with drugs activating AMP-activated protein kinase, including hypoglycemic drugs, are worthy of further investigation to combat diabetic complications.

[Network pharmacology study of effective constituents of traditional Chinese medicine for Alzheimer's disease treatment].

This study aims to investigate the network pharmacology of Chinese medicinal formulae for treatment of Alzheimer's disease.Machine learning algorithms were applied to construct classifiers in predicting the active molecules against 25 key targets toward Alzheimer's disease(AD).By extensive data profiling, we compiled 13 classical traditional Chinese medicine(TCM) formulas with clinical efficacy for AD. There were 7 Chinese herbs with a frequency of 5 or higher in our study. Based on the predicted results, we built constituent-target, and further construct target-target interaction network by STRING(Search Tool for the Retrieval of Interacting Genes/Proteins) and target-disease network by DAVID(Database for Annotation,Visualization and Integrated Discovery) and gene disease database to study the synergistic mechanism of the herbal constituents in the Chinese traditional patent medicine. By prediction of blood-brain penetration and validation by TCMsp (traditional Chinese medicine systems pharmacology) and Drugbank, we found 7 typical multi-target constituents which have diverse structure. The mechanism uncovered by this study may offer a deep insight into the action mechanism of TCMs for AD. The predicted inhibitors for the AD-related targets may provide a good source of new lead constituents against AD.

DL0410 can reverse cognitive impairment, synaptic loss and reduce plaque load in APP/PS1 transgenic mice.

Cholinesterase inhibitors are first-line therapy for Alzheimer's disease (AD). DL0410 is an AChE/BuChE dual inhibitor with a novel new structural scaffold. It has been demonstrated that DL0410 could improve memory deficits in both Aß1-42-induced and scopolamine-induced amnesia in mice. In the present study, the therapeutic effect of DL0410 and its action mechanism were investigated in APP/PS1 transgenic mice. Six-month old APP/PS1 transgenic mice were orally administered with DL0410 (3, 10, 30 mg/kg/day). After 60 days, several behavioural tests, including the Morris water maze and step-down tests, were used to investigate the effects of DL0410 on mice behaviours. All the behavioural experimental results showed that DL0410 significantly ameliorated memory deficits. Meanwhile, DL0410 attenuated neural cell damage and reduced senile plaques significantly in the hippocampus of APP/PS1 transgenic mice. In addition, DL0410 significantly decreased Aß plaques, while increasing the number of synapses and the thickness of PSD in the hippocampus. We also found DL0410 decreased the expression of APP, NMDAR1B and the phosphorylation level of NMDAR2B, and increased the phosphorylation level of CAMKII and the expression of PSD-95. In this study, the results of behavioural tests demonstrated for the first time that DL0410 could improve learning and memory dysfunction in APP/PS1 transgenic mice. The mechanism of its beneficial effects might be related to cholinesterase inhibition, Aß plaques inhibition, improvement of synapse loss by regulating of expression of proteins related to synapses. As a result, DL0410 could be considered as a candidate drug for the therapy of AD.

Ameliorative effects of baicalein in MPTP-induced mouse model of Parkinson's disease: A microarray study.

Baicalein, a flavonoid from Scutellaria baicalensis Georgi, has been shown to possess neuroprotective properties. The purpose of this study was to explore the effects of baicalein on motor behavioral deficits and gene expression in N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mice model of Parkinson's disease (PD). The behavioral results showed that baicalein significantly improves the abnormal behaviors in MPTP-induced mice model of PD, as manifested by shortening the total time for climbing down the pole, prolonging the latent periods of rotarod, and increasing the vertical movements. Using cDNA microarray and subsequent bioinformatic analyses, it was found that baicalein significantly promotes the biological processes including neurogenesis, neuroblast proliferation, neurotrophin signaling pathway, walking and locomotor behaviors, and inhibits dopamine metabolic process through regulation of gene expressions. Based on analysis of gene co-expression networks, the results indicated that the regulation of genes such as LIMK1, SNCA and GLRA1 by baicalein might play central roles in the network. Our results provide experimental evidence for the potential use of baicalein in the treatment of PD, and revealed gene expression profiles, biological processes and pathways influenced by baicalein in MPTP-treated mice.

In vitro antiviral effects and 3D QSAR study of resveratrol derivatives as potent inhibitors of influenza H1N1 neuraminidase.

The anti-influenza virus activities of 50 resveratrol (RV: 3, 5, 4'-trihydroxy-trans-stilbene) derivatives were evaluated using a neuraminidase (NA) activity assay. The results showed that 35 compounds exerted an inhibitory effect on the NA activity of the influenza virus strain A/PR/8/34 (H1N1) with 50% inhibitory concentration (IC50) values ranging from 3.56 to 186.1 µm. Next, the 35 RV derivatives were used to develop 3D quantitative structure-activity relationship (3D QSAR) models for understanding the chemical-biological interactions governing their activities against NA. The comparative molecular field analysis (CoMFA r2=0.973, q2=0.620, qtest2=0.661) and the comparative molecular similarity indices analysis (CoMSIA r2=0.956, q2=0.610, qtest2=0.531) were applied. Afterward, molecular docking was performed to study the molecular interactions between the RV derivatives and NA. Finally, a cytopathic effect (CPE) reduction assay was used to evaluate the antiviral effects of the RV derivatives in vitro. Time-of-addition studies demonstrated that the RV derivatives might have a direct effect on viral particle infectivity. Our results indicate that the RV derivatives are potentially useful antiviral compounds for new drug design and development for influenza treatment.