Discovery and mechanistic insights into thieno[3,2-d]pyrimidine and heterocyclic fused pyrimidines inhibitors targeting tubulin for cancer therapy.

Guided by the X-ray cocrystal structure of the lead compound 4a, we developed a series of thieno[3,2-d]pyrimidine and heterocyclic fused pyrimidines demonstrating potent antiproliferative activity against four tumor cell lines. Two analogs, 13 and 25d, exhibited IC50 values around 1 nM and overcame P-glycoprotein (P-gp)-mediated multidrug resistance (MDR). At low concentrations, 13 and 25d inhibited both the colony formation of SKOV3 cells in vitro and tubulin polymerization. Furthermore, mechanistic studies showed that 13 and 25d induced G2/M phase arrest and apoptosis in SKOV3 cells, as well as dose-dependent inhibition of tumor cell migration and invasion at low concentrations. Most notably, the X-ray cocrystal structures of compounds 4a, 25a, and the optimal molecule 13 in complex with tubulin were elucidated. This study identifies thieno[3,2-d]pyrimidine and heterocyclic fused pyrimidines as representatives of colchicine-binding site inhibitors (CBSIs) with potent antiproliferative activity.

Dual-Intermetallic Heterostructure on Hierarchical Nanoporous Metal for Highly Efficient Alkaline Hydrogen Electrocatalysis.

Constructing well-defined active multisites is an effective strategy to break linear scaling relationships to develop high-efficiency catalysts toward multiple-intermediate reactions. Here, dual-intermetallic heterostructure composed of tungsten-bridged Co3W and WNi4 intermetallic compounds seamlessly integrated on hierarchical nanoporous nickel skeleton is reported as a high-performance nonprecious electrocatalyst for alkaline hydrogen evolution and oxidation reactions. By virtue of interfacial tungsten atoms configuring contiguous multisites with proper adsorptions of hydrogen and hydroxyl intermediates to accelerate water dissociation/combination and column-nanostructured nickel skeleton facilitating electron and ion/molecule transportations, nanoporous nickel-supported Co3W-WNi4 heterostructure exhibits exceptional hydrogen electrocatalysis in alkaline media, with outstanding durability and impressive catalytic activities for hydrogen oxidation reaction (geometric exchange current density of ≈6.62 mA cm-2) and hydrogen evolution reaction (current density of ≈1.45 A cm-2 at overpotential of 200 mV). Such atom-ordered intermetallic heterostructure alternative to platinum group metals shows genuine potential for hydrogen production and utilization in hydroxide-exchange-membrane water electrolyzers and fuel cells.

Novel Medicinal Chemistry Strategies Targeting CDK5 for Drug Discovery.

Cyclin-dependent kinase 5 (CDK5) protein plays an important role not only in the central nervous system but also in the periphery, including immune response, regulation of insulin secretion, and cancer development and progression. Consequently, targeting the CDK5 protein is a potential strategy for the treatment of many diseases, especially cancer and neurodegenerative diseases. To date, numerous pan-CDK inhibitors have entered clinical trials. Nevertheless, limited clinical efficacy and severe adverse effects have prompted the application of new techniques to optimize clinical efficacy and minimize adverse events. In this Perspective, we highlight the protein properties, biofunctions, relevant signaling pathways, and associations with cancer development and proliferation of CDK5, and analyze the clinical status of pan-CDK inhibitors and the preclinical status of CDK5-specific inhibitors. In addition, CDK5-selective inhibitors, protein-protein interaction inhibitors, proteolytic-targeting chimera (PROTAC) degraders, and dual-target CDK5 inhibitors are discussed.

A multiparametric fluorescent visualization approach for detecting drug resistance in living cancer cells.

Drug resistance is a worldwide health care crisis which impedes disease treatment and increases financial burden, especially for its multifactorial nature and high complexity. Herein, we developed a multiparametric approach to visualize and detect drug resistance in living cancer cells, through the combination of DNA-templated covalent protein labeling strategy and fluorescent resonance energy transfer technique. Gefitinib resistance in non-small cell lung cancer caused by mesenchymal-epidermal transition factor (Met) overexpression and hyperactivation was investigated as a proof-of-concept. Unlike the traditional single-factor investigation, the proposed approach evaluated the contribution of three important parameters towards the resistance, including the changes of Met expression level, the homodimerization of Met with itself and the heterodimerization of Met with epidermal growth factor receptor (EGFR). A multiple regression model based on these three parameters was tentatively established for evaluation of the resistance level of laboratory-developed resistant cells and evaluation of the resistance level of patient-derived cells. Such an approach facilitates a quick identification of a drug resistance, to evaluate not only the resistance level but also the resistance mechanism.

Strategies of Targeting CK2 in Drug Discovery: Challenges, Opportunities, and Emerging Prospects.

CK2 (casein kinase 2) is a serine/threonine protein kinase that is ubiquitous in eukaryotic cells and plays important roles in a variety of cellular functions, including cell growth, apoptosis, circadian rhythms, DNA damage repair, transcription, and translation. CK2 is involved in cancer pathogenesis and the occurrence of many diseases. Therefore, targeting CK2 is a promising therapeutic strategy. Although many CK2-specific small-molecule inhibitors have been developed, only CX-4945 has progressed to clinical trials. In recent years, novel CK2 inhibitors have gradually become a research hotspot, which is expected to overcome the limitations of traditional inhibitors. Herein, we summarize the structure, biological functions, and disease relevance of CK2 and emphatically analyze the structure-activity relationship (SAR) and binding modes of small-molecule CK2 inhibitors. We also discuss the latest progress of novel strategies, providing insights into new drugs targeting CK2 for clinical practice.

A Design Space for Surfacing Content Recommendations in Visual Analytic Platforms.

Recommendation algorithms have been leveraged in various ways within visualization systems to assist users as they perform of a range of information tasks. One common focus for these techniques has been the recommendation of content, rather than visual form, as a means to assist users in the identification of information that is relevant to their task context. A wide variety of techniques have been proposed to address this general problem, with a range of design choices in how these solutions surface relevant information to users. This paper reviews the state-of-the-art in how visualization systems surface recommended content to users during users' visual analysis; introduces a four-dimensional design space for visual content recommendation based on a characterization of prior work; and discusses key observations regarding common patterns and future research opportunities.

Visualization of Protein-Specific Glycation in Living Cells via Bioorthogonal Chemical Reporter.

Due to the lack of suitable chemical tools, probing the protein-specific glycation is highly challenging. Herein, we present a strategy based on glycation chemical reporter and proximity-induced FRET signal readout for visualizing protein-specific glycation in living cells. We first developed a bioorthogonal glucose analogue, 6-azido-6-deoxy-D-glucose (6AzGlc), as a novel glycation chemical reporter. Two types of DNA probes, glycation conversion probe and protein targeting probe, were designed to attach to glycation adducts and target proteins, respectively. After the protein was glycated by 6AzGlc, two DNA probes were sequentially applied to the target protein, triggering proximity-induced FRET signal readout. This strategy was successfully used to visualize glucose glycation of several proteins, including PD-L1 and integrin. More importantly, this strategy allowed us to analyze corresponding biological functions of glycated protein in the native environment.

Small molecule NS5B RdRp non-nucleoside inhibitors for the treatment of HCV infection: A medicinal chemistry perspective.

Hepatitis C virus (HCV) infection has become a global health problem with enormous risks. Nonstructural protein 5B (NS5B) RNA-dependent RNA polymerase (RdRp) is a component of HCV, which can promote the formation of the viral RNA replication complex and is also an essential part of the replication complex itself. It plays a vital role in the synthesis of the positive and negative strands of HCV RNA. Therefore, the development of small-molecule inhibitors targeting NS5B RdRp is of great value for treating HCV infection-related diseases. Compared with NS5B RdRp nucleoside inhibitors, non-nucleoside inhibitors have more flexible structures, simpler mechanisms of action, and more predictable efficacy and safety of drugs in humans. Technological advances over the past decade have led to remarkable achievements in developing NS5B RdRp inhibitors. This review will summarize the non-nucleoside inhibitors targeting NS5B RdRp developed in the past decade and describe their structure optimization process and structure-activity relationship.

Endothelial Biomarkers in Patients Recovered from COVID-19 One Year after Hospital Discharge: A Cross-Sectional Study.

Background: COVID-19 is characterized by endothelial dysfunction and is presumed to have long-term cardiovascular sequelae. In this cross-sectional study, we aimed to explore the serum levels of endothelial biomarkers in patients who recovered from COVID-19 one year after hospital discharge. Methods: In this clinical follow-up study, 345 COVID-19 survivors from Huanggang, Hubei, and 119 age and gender-matched medical staff as healthy controls were enrolled. A standardized symptom questionnaire was performed, while electrocardiogram and Doppler ultrasound of lower extremities, routine blood tests, biochemical and immunological tests, serum soluble vascular cell adhesion molecule-1(VCAM-1), intercellular cell adhesion molecule-1(ICAM-1), P-selectin, and fractalkine were measured by enzyme-linked immunosorbent assays (ELISA). Results: At one year after discharge, 39% of recovers possessed post-COVID syndromes, while a few had abnormal electrocardiogram manifestations, and no deep vein thrombosis was detected in all screened survivors. There were no significant differences in circulatory inflammatory markers (leukocytes, neutrophils, lymphocytes, C-reactive protein and interleukin-6), alanine aminotransferase, estimated glomerular filtration rate, glucose, triglycerides, total cholesterol and D-dimer observed among healthy controls with previously mild or severe infected. Furthermore, serum levels of VCAM-1, ICAM-1, P-selectin, and fractalkine do not significantly differ between survivors and healthy controls. Conclusions: SARS-CoV-2 infection may not impose a higher risk of developing long-term cardiovascular events, even for those recovering from severe illness.

Insulin-Like Growth Factor-1 Enhances Motoneuron Survival and Inhibits Neuroinflammation After Spinal Cord Transection in Zebrafish.

Insulin-like growth factor-1 (IGF-1) is a neurotrophic factor produced locally in the central nervous system which can promote axonal regeneration, protect motoneurons, and inhibit neuroinflammation. In this study, we used the zebrafish spinal transection model to investigate whether IGF-1 plays an important role in the recovery of motor function. Unlike mammals, zebrafish can regenerate axons and restore mobility in remarkably short period after spinal cord transection. Quantitative real-time PCR and immunofluorescence showed decreased IGF-1 expression in the lesion site. Double immunostaining for IGF-1 and Islet-1 (motoneuron marker)/GFAP (astrocyte marker)/Iba-1 (microglia marker) showed that IGF-1 was mainly expressed in motoneurons and was surrounded by astrocyte and microglia. Following administration of IGF-1 morpholino at the lesion site of spinal-transected zebrafish, swimming test showed retarded recovery of mobility, the number of motoneurons was reduced, and increased immunofluorescence density of microglia was caused. Our data suggested that IGF-1 enhances motoneuron survival and inhibits neuroinflammation after spinal cord transection in zebrafish, which suggested that IGF-1 might be involved in the motor recovery.

Activating Transcription Factor 6 Contributes to Functional Recovery After Spinal Cord Injury in Adult Zebrafish.

Spinal cord injury (SCI) is one of the most common devastating injuries, with little possibility of recovery in humans. However, zebrafish efficiently regenerate functional nervous system tissue after SCI. Therefore, the spinal cord transection model of adult zebrafish was applied to explore the role of ATF6 in neuro-recovery. Activating transcription factor 6 (ATF6) is a type-II transmembrane protein in the endoplasmic reticulum (ER). ATF6 target genes could improve ER homeostasis, which contributes to cytoprotection. Herein, we found that the ATF6 level increased at 12 h and 3 days post SCI, and returned to sham levels at 7 days post SCI. ATF6-expressing motor neurons were present in the central canal of the spinal cord and increased at 12 h post SCI. ATF6 morpholino treatment showed that inhibition of ATF6 delayed locomotor recovery and hindered neuron axon regrowth in SCI zebrafish. Furthermore, we investigated the role of both binding immunoglobulin protein (Bip) and C/EBP homologous transcription factor protein (CHOP), the two target genes of ATF6. We found that Bip expression significantly increased in the spinal cord at 7 days after SCI, which served as a pro-survival chaperone. Our results also showed that CHOP expression significantly decreased in the spinal cord at 7 days after SCI, which was identified as a protein involved in apoptosis. Taken together, our data demonstrate that ATF6 may contribute to the functional recovery after SCI in adult zebrafish, via up-regulation of Bip and down-regulation of CHOP to restore the homeostasis of ER.

Sodium Butyrate Exacerbates Parkinson's Disease by Aggravating Neuroinflammation and Colonic Inflammation in MPTP-Induced Mice Model.

The abnormal production of short chain fatty acid (SCFAs) caused by gut microbial dysbiosis plays an important role in the pathogenesis and progression of Parkinson's disease (PD). This study sought to evaluate how butyrate, one of SCFAs, affect the pathology in a subacute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride (MPTP) treated mouse model of PD. Sodium butyrate (NaB; 165 mg/kg/day i.g., 7 days) was administrated from the day after the last MPTP injection. Interestingly, NaB significantly aggravated MPTP-induced motor dysfunction (P < 0.01), decreased dopamine (P < 0.05) and 5-HT (P < 0.05) levels, exacerbated declines of dopaminergic neurons (34%, P < 0.05) and downregulated expression of tyrosine hydroxylase (TH, 47%, P < 0.05), potentiated glia-mediated neuroinflammation by increasing the number of microglia (17%, P < 0.05) and activating astrocytes (28%, P < 0.01). In vitro study also confirmed that NaB could significantly exacerbate pro-inflammatory cytokines expression (IL-1ß, 4.11-fold, P < 0.01; IL-18, 3.42-fold, P < 0.01 and iNOS, 2.52-fold, P < 0.05) and NO production (1.55-fold, P < 0.001) in LPS-stimulated BV2 cells. In addition, NaB upregulated the expression of pro-inflammatory cytokines (IL-6, 3.52-fold, P < 0.05; IL-18, 1.72-fold, P < 0.001) and NLRP3 (3.11-fold, P < 0.001) in the colon of PD mice. However, NaB had no effect on NFκB, MyD88 and TNF-α expression in PD mice. Our results indicate that NaB exacerbates MPTP-induced PD by aggravating neuroinflammation and colonic inflammation independently of the NFκB/MyD88/TNF-α signaling pathway.

Nongenetic engineering strategies for regulating receptor oligomerization in living cells.

Cell surface receptors are important proteins that mediate communication between the cells and their outside environment, and also play essential roles in the control of a wide variety of biological processes, such as cell cycle, proliferation, communication, migration and apoptosis. Receptor oligomerization is an essential signal transduction mechanism that cell surface receptors use to transmit extracellular signals into the internal cytosol cellular machinery. Therefore, regulating receptor oligomerization provides an opportunity to customize cellular signaling and to direct cellular behavior in a user-defined manner. Some techniques have been developed for receptor oligomerization regulation, such as chemically induced dimerization (CID) and optogenetics, which involve traditional genetic engineering. However, the process of genetic manipulation is time-consuming, unpredictable and inefficient. Thus, development of nongenetic strategies for precisely regulating receptor oligomerization remains a desirable goal. Recently, along with the utilization of DNA, protein, small molecules and stimuli-responsive materials-based nongenetic engineering strategies, various receptor oligomerization and multiple cellular behaviors could be regulated, including migration, proliferation, apoptosis, differentiation and immune responses, etc. In this review, we aim to systematically introduce advances in the development of nongenetic engineering strategies for regulating receptor oligomerization, and provide insights into the existing challenges and future perspectives of this field.

Sodium butyrate causes α-synuclein degradation by an Atg5-dependent and PI3K/Akt/mTOR-related autophagy pathway.

Aggregation of α-Synuclein is central to the pathogenesis of Parkinson's disease (PD). However, these α-Synuclein inclusions are not only present in brain, but also in gut. Enteroendocrine cells (EECs), which are directly exposed to the gut lumen, can express α-Synuclein and directly connect to α-Synuclein-containing nerves. Dysbiosis of gut microbiota and microbial metabolite short-chain fatty acids (SCFAs) has been implicated as a driver for PD. Butyrate is an SCFA produced by the gut microbiota. Our aim was to demonstrate how α-Synuclein expression in EECs responds to butyrate stimulation. Interestingly, we found that sodium butyrate (NaB) increases α-Synuclein mRNA expression, enhances Atg5-mediated autophagy (increased LC3B-II and decreased SQSTM1 (also known as p62) expression) in murine neuroendocrine STC-1 cells. Further, α-Synuclein mRNA was decreased by the inhibition of autophagy by using inhibitor bafilomycin A1 or by silencing Atg5 with siRNA. Moreover, the PI3K/Akt/mTOR pathway was significantly inhibited and cell apoptosis was activated by NaB. Conditioned media from NaB-stimulated STC-1 cells induced inflammation in SH-SY5Y cells. Collectively, NaB causes α-Synuclein degradation by an Atg5-dependent and PI3K/Akt/mTOR-related autophagy pathway.

Neuroprotection of Fasting Mimicking Diet on MPTP-Induced Parkinson's Disease Mice via Gut Microbiota and Metabolites.

Parkinson's disease (PD) is strongly associated with life style, especially dietary habits, which have gained attention as disease modifiers. Here, we report a fasting mimicking diet (FMD), fasting 3 days followed by 4 days of refeeding for three 1-week cycles, which accelerated the retention of motor function and attenuated the loss of dopaminergic neurons in the substantia nigra in 1-methyl-4-phenyl-1,2,3,6-tetrathydropyridine (MPTP)-induced PD mice. Levels of brain-derived neurotrophic factor (BDNF), known to promote the survival of dopaminergic neurons, were increased in PD mice after FMD, suggesting an involvement of BDNF in FMD-mediated neuroprotection. Furthermore, FMD decreased the number of glial cells as well as the release of TNF-α and IL-1ß in PD mice, showing that FMD also inhibited neuro-inflammation. 16S and 18S rRNA sequencing of fecal microbiota showed that FMD treatment modulated the shifts in gut microbiota composition, including higher abundance of Firmicutes, Tenericutes, and Opisthokonta and lower abundance of Proteobacteria at the phylum level in PD mice. Gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry revealed that FMD modulated the MPTP-induced lower propionic acid and isobutyric acid, and higher butyric acid and valeric acid and other metabolites. Transplantation of fecal microbiota, from normal mice with FMD treatment to antibiotic-pretreated PD mice increased dopamine levels in the recipient PD mice, suggesting that gut microbiota contributed to the neuroprotection of FMD for PD. These findings demonstrate that FMD can be a new means of preventing and treating PD through promoting a favorable gut microbiota composition and metabolites.

Neuroprotective effects of Astilbin on MPTP-induced Parkinson's disease mice: Glial reaction, α-synuclein expression and oxidative stress.

Astilbin (AST), a dihydro-flavonol glycoside, is a major bioactive ingredient in Astilbe thunbergii, Engelhardia roxburghiana, Smilax corbularia and Erythroxylum gonocladum, and has been shown to have anti-inflammatory, antioxidative and neuroprotective effects, suggesting potential therapeutic value in the treatment of Parkinson's disease (PD). We explored the neuroprotective effects of AST in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced Parkinson's disease mice. Mice were administered with MPTP (30 mg/kg, i.p) daily for 5 days, to establish a subacute Parkinson's disease model, followed by daily treatment with AST or saline for 7 days. Pole and traction tests showed that AST ameliorated the impaired motor functions in MPTP-induced Parkinson's disease mice. High performance liquid chromatography analysis revealed that AST treatment prevented MPTP-induced decreases in striatal dopamine levels. Immunofluorescence assays showed that AST reduced the loss of dopaminergic neurons and the activation of microglia and astrocytes in the substantia nigra. Western blot analyses revealed that AST suppressed α-synuclein overexpression and activated PI3K/Akt in the striatum following MPTP treatment. AST also prevented the MPTP-induced reduction in total superoxide dismutase and glutathione activity in the striatum. AST exerts neuroprotective effects on MPTP-induced PD mice by suppressing gliosis, α-synuclein overexpression and oxidative stress, suggesting that AST could serve as a therapeutic drug to ameliorate PD.

Aucubin alleviates glial cell activation and preserves dopaminergic neurons in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced parkinsonian mice.

Aucubin (AUC) is a major bioactive ingredient in Eucommia ulmoides, Plantain asiatica, and Aucuba japonica, and has been shown to exert anti-inflammatory, antioxidative, and neuroprotective effects. We explore the neuroprotective effects of AUC in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced parkinsonian mice. Mice were administered MPTP (30 mg/kg) daily for 5 days, followed by treatment with AUC for 7 days. Measurement of dopamine levels was performed by high-performance liquid chromatography and tyrosine hydroxylase expression was assessed by western blot. Our results showed that AUC treatment improved mobility in the pole descent test and the traction test, and reduced the loss of dopaminergic neurons in MPTP-induced parkinsonian mice. AUC treatment rescued the decreased dopamine and tyrosine hydroxylase levels in the striatum of parkinsonian mice. Furthermore, AUC treatment reduced both microglia and astrocyte activation in the substantia nigra of parkinsonian mice. These findings suggest that AUC exerts neuroprotective effects, in part by reducing inflammation and preserving dopaminergic neurons. Possible protection mechanisms involved in MPTP-induced parkinsonian mice need to be clarified further.

Neuroprotective effects of fecal microbiota transplantation on MPTP-induced Parkinson's disease mice: Gut microbiota, glial reaction and TLR4/TNF-α signaling pathway.

Parkinson's disease (PD) patients display alterations in gut microbiota composition. However, mechanism between gut microbial dysbiosis and pathogenesis of PD remains unexplored, and no recognized therapies are available to halt or slow progression of PD. Here we identified that gut microbiota from PD mice induced motor impairment and striatal neurotransmitter decrease on normal mice. Sequencing of 16S rRNA revealed that phylum Firmicutes and order Clostridiales decreased, while phylum Proteobacteria, order Turicibacterales and Enterobacteriales increased in fecal samples of PD mice, along with increased fecal short-chain fatty acids (SCFAs). Remarkably, fecal microbiota transplantation (FMT) reduced gut microbial dysbiosis, decreased fecal SCFAs, alleviated physical impairment, and increased striatal DA and 5-HT content of PD mice. Further, FMT reduced the activation of microglia and astrocytes in the substantia nigra, and reduced expression of TLR4/TNF-α signaling pathway components in gut and brain. Our study demonstrates that gut microbial dysbiosis is involved in PD pathogenesis, and FMT can protect PD mice by suppressing neuroinflammation and reducing TLR4/TNF-α signaling.

[Study on different doses of mercury-containing preparations on acute toxicity in rabbits].

OBJECTIVE: To observe the effect of single administration of mercury- containing preparation Jiuyi Dan (calcined gypsum-Shengdan 9: 1) and Shengdan on acute toxicity of rabbits, in order to assess the safety of tested drugs. METHOD: The rabbits were randomly divided into 4 groups: the calcined gypsum group (excipient control), the Jiuyi Dan group, the 90 mg Shengdan group and the 180 mg Shengdan group. After 270 mg of calcined gypsum, 300 mg of Jiuyi Dan, 90 mg of Shengdan, and 180 mg of Shengdan were used on the surface of wounds (5 cm x 5 cm) on two sides of rabbit back for 5 h, the surfaces of wound were washed by water. The bloods were taken from the rabbit hearts before and after the drug administration for 24 h, 72 h, 7 d and 14 d for determining Hg level in blood and liver & kidney function indicators (ALT, AST, CREAT, and BUN). The rabbits were dissected after the drugs treatment for 14 d, and pathological tests were made for their livers and kidneys. RESULT: Compared with the calcined gypsum group, the 90 mg Shengdan group and the 180 mg Shengdan group showed significant increase (P < 0.01 or P < 0.05), as evidenced by increase in CREAT for 24 h and 72 h and increase in BUN for 24 h and on 7 d. AST is significantly increased as well (P < 0.01) for 24 h and 72 h compared to that of the group before drug treatment. The Hg level in blood was significantly enhanced (P < 0.01) after the rabbits were administrated with drugs for 24 h to 72 h. The pathological changes in livers and kidneys of rabbits were observed in the two doses of Shengdan treatment groups. CONCLUSION: The Hg blood levels were increased significantly in an obvious dose-effect relationship in all drugs treatment groups. Liver & kidney function indicators were influenced by Shengdan treatment to some extent. Meanwhile, pathological changes in rabbit livers and kidneys were also caused by Shengdan, while Jiuyi Dan has no significantly effect on livers and kidneys.