The overexpression of doublecortin-like kinase 1 (DCLK1) is closely related to the occurrence and development of various malignant tumors. Discovery of novel anti-tumor agents targeting DCLK1 represents a hot spot in this field. So far, several DCLK1 small molecule inhibitors with excellent anti-tumor activity in vitro and in vivo have been designed and synthesized, which is expected to provide a new strategy for tumor therapy. This article reviews the research progress in the discovery, structure type, structural optimization, biological activity and mechanism of action of DCLK1 small molecule inhibitors, and provides research basis for the development of new anti-tumor small molecule inhibitors targeting DCLK1.

The epidemic of Zika virus (ZIKV) raises critical public health and safety problems. However, there are currently no vaccines or drugs that are clinically approved for ZIKV infections. Since RNA-dependent RNA polymerase (RdRp) plays an important role in replication and transcription of ZIKV and is absent in human beings, it is a potential drug screening target of anti-ZIKV agents. According to the fluorescence-based alkaline phosphatase-coupled polymerase assay method, we established the NS5 RdRp inhibitor screening model. Through screening from an anti-infection compound library, we found a compound octenidine dihydrochloride (OCT) that could inhibit ZIKV RdRp activity with a half maximal inhibitory concentration (IC₅₀) of 5.43 μmol·L^-1. Biolayer interferometry (BLI) assay showed that OCT could bind to ZIKV RdRp and had a strong affinity. Moreover, OCT exhibited an inhibitory effect on ZIKV replication with a half maximal effective concentration (EC₅₀) of 29.94 μmol·L^-1. All these results indicated that OCT had the anti-ZIKV activity by targeting ZIKV RdRp, and it is likely to be a promising lead compound against ZIKV.

In this study, network pharmacology research and animal experiments were used to predict and validate the potential targets of ShengMaiSan (SMS) in the treatment of diabetic cardiomyopathy. The active components of SMS were obtained through TCMSP and BATMAN databases. The potential targets of the active components and diabetic cardiomyopathy were predicted by Swiss Target Prediction and GeneCards databases, respectively. The protein-protein interaction (PPI) network was constructed by String database. Cytoscape software was adopted to perform topological analysis to select the core action target. Gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analyses were performed using Metascape platform. To validate the potential targets, a type I diabetic rat model which induced by intraperitoneal injection of streptozotocin (STZ) was prepared. Rats were divided into sham group, model group, SMS group and trimetazidine (TMZ) group. Left ventricular hemodynamics was detected after 4 weeks administrated of SMS or TMZ. Myocardial contraction and calcium transients were detected synchronously in cardiomyocytes, as well as sarcoplasmic reticulum calcium content, calcium leak level and ryanodine receptor 2 (RyR2) expression were detected. Based on network pharmacology, 1 288 targets of SMS, 1 066 targets of diabetic cardiomyopathy, and 180 overlapped targets were obtained. The 39 core targets were screened, and 159 pathways including calcium signaling pathway were screened by KEGG pathway analysis. According to the previous studies and focusing on the contractility of diabetic cardiomyopathy, this study was involved calcium signaling regulation pathway in the SMS protection mechanism. The results showed that, compared with sham group, the systolic function of left ventricular and myocardial cells were decreased, and the calcium transport was in disorder in model group; compared with model group, both SMS group and TMZ group increased the maximum systolic pressure of left ventricle and the maximum systolic rate of left ventricular contraction. In addition, SMS group and TMZ group increased the contraction amplitude of cardiomyocytes, decreased the diastolic calcium concentration, the sarcoplasmic reticulum calcium leak and decreased the phosphorylation level of RyR2. There was no significant difference between SMS and TMZ groups. In summary, SMS could reduce the calcium leak of the sarcoplasmic reticulum and enhance the myofilament sensitivity of calcium to increasing contractile function of diabetic rats. The animal welfare and experiment procedures of this study were in accordance with the regulations of the Experimental Animal Ethics Committee of Experimental Research Center, China Academy of Chinese Medical Sciences.

Cerebrovascular diseases have the characteristics of high morbidity, high disability, high mortality and high recurrence rate, which seriously harm human health and increase the national health economic burden. 3-n-Butylphthalide (NBP) is a new drug commonly used in clinical treatment of cerebrovascular diseases, and it is also one of the main active components in traditional Chinese medicine such as Angelica sinensis and Chuanxiong. In this review, the pharmacological effects of NBP were systematically summarized. Studies have shown that NBP has pharmacological effects such as antiplatelet aggregation, anti-thrombosis, inhibiting neuronal apoptosis, anti-oxidation, anti-cerebral ischemia, anti-brain injury, and anti-vascular dementia. In clinical practice, it is often combined with edaravone, alteplase, fasudil, Xingnaojing injection, and compound Danshen injection to treat cerebral vascular diseases such as stroke, vascular dementia, and cerebral vasospasm, and plays a good synergistic effect. This summary could provide support for the rational clinical application of NBP, and also provide basis for the in-depth study of the interaction of its drug combination.

The blood-brain barrier (BBB) plays an important role in maintaining the homeostasis of the central nervous system. BBB is disrupted in many neurological disorders such as ischemic stroke and Alzheimer's disease. Traditional Chinese medicine has great potential to prevent ischemic stroke, but the lack of clinically relevant models has been a challenge. We adapted a BBB organoid model formed by human brain microvascular endothelial cells (HBMEC), human astrocytes (HA), and human brain vascular pericytes (HBVP), and established conditions for oxygen-glucose deprivation/reoxygenation (OGD/R) on the cell vitality, barrier permeability, as well as BBB signature marker expression. The protective effect of Guanxinning injection (GXNI) on OGD/R-induced BBB dysfunction was then investigated in the organoid model. The results showed that OGD/R decreased BBB organoid cell viability, increased permeability (leakage), decreased the level of tight junction proteins zonula occludens-1 (ZO-1), claudin-5, occludin and P-glycoprotein (P-gp). GXNI significantly prevented OGD/R-induced BBB disruption such as the decreased cell viability and increased permeability. This study provides a new human cell-derived 3D ischemic brain disease model for central nervous system-targeted drug research and development and demonstrates that a Chinese injection medicine GXNI effectively protects BBB dysfunction in vitro.

Ultra-high performance liquid chromatography-tandem quadrupole time-of-flight mass spectrometry (UHPLC/Q-TOF-MS) was applied to rapidly identify the phospholipids in human plasma and explore the mass spectrometric fragmentation pattern. An acquity UHPLC^TM BEH C18 column (50 mm × 2.1 mm, 1.7 μm) was utilized and eluted with a gradient system; the mobile phase consisted of 10 mmol·L^-1 ammonium formate aqueous solution-0.1% formic acid aqueous solution (A) and acetonitrile-isopropanol (1∶1) organic solution (B) containing 10 mmol·L^-1 ammonium formate-0.1% formic acid. The flow rate was 0.3 mL·min^-1 and the column temperature was set at 50 ℃. An electrospray ionization (ESI) source was used to collect mass spectra in positive and negative ion mode. Based on the precise relative molecular weight and elemental composition calculated by Masslynx 4.1 software, comparison with references, and secondary mass spectrometry fragment ions and lipid databases, a total of 82 plasma lipids were identified, including 14 lysophosphatidylcholines (LysoPCs), 39 phosphatidylcholines (PCs), 17 sphingomyelins (SMs), 7 ceramides (Cers), 4 phosphatidylethanolamines (PEs), and 1 phosphatidylinositol (PI). A simple, efficient, fast and stable analytical method was established in this study for the qualitative analysis of phospholipids in human plasma, and the fragmentation regularity of the main phospholipids was determined. This work provides a good foundation for further metabolomics studies of plasma phospholipids. This study was approved by the Second Affiliated Hospital of Guangzhou Medical University Clinical Research and Application Institutional Review Board Approval (No. 2020-hs-07).

In this study, we investigated the pharmacological effect and possible molecular mechanism of higenamine (HG) in isoproterenol (ISO)-induced myocardial infarction (MI). All procedures were approved by the Institutional Animal Care and Use Committee of the Guangzhou University of Chinese Medicine. ISO was used to induce MI model in rats and H9c2 cells. The effects of HG on biomarkers and cardiac function in MI rats were evaluated by enzyme linked immunosorbent assay (ELISA), echocardiography and hematoxylin-eosin staining (HE). The expression of apoptosis and autophagy related proteins were detected by Western blot in myocardial tissue and H9c2 cells, as well as methyltransferase-like 3 (METTL3) and transcription factor EB (TFEB) protein expression. Molecular docking was used to evaluate the interaction between HG and METTL3. The results showed that HG significantly improved cardiac function and pathologic changes in ISO-induced MI, and inhibited the levels of MI-related biomarkers such as creatine kinase Mb (CK-MB), creatine kinase (CK) and lactate dehydrogenase (LDH). Mechanism studies showed that HG inhibited the expression of apoptosis-related proteins (Bax/Bcl2, caspase3, cleaved-caspase3). Interestingly, HG up-regulated the expression of autophagy related protein Beclin1, promoted autophagy flux, and decreased the ratio of light chain 3B-I/light chain 3B-II (LC-3B-I/LC-3B-II). Further studies found that HG increased the autophagy regulator TFEB and inhibited METTL3 expression. Molecular docking results showed that HG had a good interaction with METTL3. Taken together, HG has a potential anti-MI effect via regulating METTL3/TFEB signaling pathway-mediated autophagy.

Influenza virus is an RNA virus that classified into 4 types, A, B, C, and D, where influenza A and B virus infection may cause human acute respiratory tract infection and nearly 0.3 million deaths annually. The life cycle of influenza virus infection is highly dependent on the host response, demonstrating an important strategy of developing anti-influenza agents that target the host factors. This research utilized a transcriptome signature reversion (TSR) strategy to discover a list of multi-host-factor-target anti-influenza agents and determined their anti-influenza activities in vitro. BIX02189 was discovered and exhibited broad spectrum anti-influenza activity, with half maximal effective concentration (EC₅₀) of 17.1 μmol·L^-1 against influenza A virus H1N1 (A/Puerto Rico/8/1934) and 9.4 μmol·L^-1 for influenza B virus (B/Jiangxi Xinjian/BV/39/2008). The anti-influenza A virus activity of BIX01289 is stronger than the positive control ribavirin with EC₅₀ of 97.9 μmol·L^-1 for influenza A virus H1N1 (A/Puerto Rico/8/1934). According to the unsupervised transcriptomic profile similarity clustering analysis, BIX02189 was considered to inhibit viral protein synthesis and release of influenza virus mainly through inhibiting the Raf/MEK/ERK cascade, revealing its potential mechanism of inhibiting influenza virus infection.

Inosine 5′-monophosphate dehydrogenase (IMPDH) is a key enzyme catalyzing the rate-limiting step of de novo nucleotide synthesis in vivo. In recent years, it has become a therapeutic target for anti-virus, anti-bacterial, anti-cancer, anti-parasitic and other diseases. IMPDH inhibitors have been shown to inhibit viral prolife-ration in host cells by depleting guanosine 5′-monophosphate (GMP), the raw material required for viral replication in host cells, with broad-spectrum antiviral properties. In order to find novel anti-coronavirus drugs, this study screened 22 potential IMPDH inhibitors from 70 000 natural small molecule libraries based on IMPDH protein structure using molecular docking and ROC calculation for virtual screening. With ribavirin as the positive control drug, Huh7 cell and H460 cell models were used to verify the anti-coronavirus HCoV-229E and HCoV-OC43 activities of 22 selected target compounds. Among them, compounds 11, 12, 15 and 16 showed inhibitory activity against coronavirus HCoV-229E. The compounds 4, 12, 13 and 15 showed inhibitory activities against coronavirus HCoV-OC43. 12 and 15 showed significant inhibitory activity against both two coronaviruses, and their efficacy was similar to ribavirin at the same dose, which can be further studied as a lead compound for IMPDH inhibitors.

For rapid discovery of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease (M^pro) inhibitors from a natural product library, a novel colorimetric screening assay was developed. According to the colorimetric principle, the synthetic peptide TSAVLQ-para-nitroanilide (pNA) was used as the M^pro hydrolysis substrate. Subsequently, the working concentration of pNA substrate, M^pro working concentration, hydrolysis time and DMSO tolerance were optimized for the development of a simple and robust colorimetric screening assay. Through these systematic optimizations, we selected 0.4 μmol·L^-1 M^pro and 100 μmol·L^-1 pNA substrate as the optimal working concentrations in this colorimetric screening assay, and a high Z' factor of 0.9 was achieved. Using this screening assay, natural product ginkgolic acid C13:0 (GA13:0) was identified as a novel competitive M^pro inhibitor in vitro. Taken together, we have successfully developed a simple and optimized colorimetric screening assay, which will be vital for the discovery of novel SARS-CoV-2 M^pro inhibitors.