With the development of the research on innovative drugs in our country, first-in-class drugs are becoming a main goal for both pharmaceutical companies and scientific institutions. Discovery of first-in-class drugs require amounts of basic research, a massive investment and novel methods, acting as a beacon for the new drug development. In 2020, FDA totally approved 53 novel drugs with 38 small molecules, which still accounting for a major component. Among them, many first-in-class drugs are important including a first EZH2 inhibitor(tazemetostat) for the treatment of epithelioid sarcoma, a first attachment inhibitor(fostemsavir) with novel mechanism for the treatment of HIV, a first farnesyltransferase inhibitor(lonafarnib) for the treatment of HutchinsonGilford progeria syndrome(HGPS) and a first MC4 receptor agonist for the treatment of rare genetic diseases of obesity, etc. The research procedures of the above drugs are representative with new ideas. In this review, we outline3 of the first-in-class drugs to discuss the research background, discovery and development process as well as the therapeutic potentials to provide methods and ideas for the further drug development.

Thiazolidinediones(TZDs) are currently the only recognized insulin sensitizers available for the clinical treatment of type 2 diabetes. Although their advantages are recognized, the profiles of numerous adverse effects hinder the continued use of these drugs. Peroxisome proliferator-activated receptor γ(PPARγ) is known as a receptor for TZDs, and its underlying mechanisms of pharmacological actions and adverse effects have been deeply explored. To maximally preserve the PPARγ-mediated insulin sensitizing effects and reduce the occurrence of related adverse effects, the concept of "selective PPARγ modulators(SPPARMs)" has been proposed and developed, guiding the development of new drugs. In this review, we summarize the recent research progress in the definition of SPPARMs, the candidate classification and the molecular underpinnings, as well as present the discovery of the YR series compounds as an example, and discuss the potential application prospects of SPPARMs.

The interaction between platelets and tumor cells can not only promote the metastasis of malignant tumors, but also affect the formation of malignant tumor-related thrombus. When tumor cells enter the blood, they will immediately activate platelets to make them adhere to the surface of tumor cells, protecting tumor cells from blood flow shear force and immune system attack, thereby promoting tumor metastasis. At the same time, the massive adhesion of platelets may also lead to the formation of thrombus. In this article, we use the methods of ingenuity pathway analysis and literature integration to explore the mechanism of platelet-tumor cell interaction and potential drugs for the treatment of malignant tumor metastasis based on the platelet-tumor cell interaction. It provides a certain theoretical basis and clinical reference for the future development of new drugs targeting platelettumor cell interaction based on its mechanism of action.

Salvianolic acids are the main water-soluble active compounds of Salvia miltiorrhiza and have been widely used for the treatment of cardiovascular diseases. Based on the latest studies in China and abroad, we summarize the pharmacological effects and mechanism of salvianolic acids on ischemic heart disease by describing how salvianolic acid A and salvianolic acid B protect the vascular endothelium, relax coronary arteries, promote angiogenesis and anti-platelet aggregation, inhibit the inflammatory response, anti-cell apoptosis, and scavenge free radicals. This review provides a theoretical basis for further research on the effects of salvianolic acids on ischemic heart disease and their potential for drug development.

RAS, as a well-known proto-oncogene, is the most frequently mutated oncogene in human cancers, yet tremendous efforts over the past 30 years have failed to develop effective therapies for RAS-mutant cancer.Recently, specifically targeting the KRAS-G12C mutant, a frequently occurring KRAS mutation in human cancers, has shown promise in conquering KRAS-mutant cancers, and has inspired interest in this direction. We herein review the very recent progress achieved in the development of covalent inhibitors towards KRAS-G12C mutant, in combinational therapies and in proteolysis-targeting chimeras(PROTACs)-based approaches to disrupt KRASG12C protein. We provide insights for drug discovery against KRAS-G12C-mutated tumors and discuss the potential challenges in this field.

Alzheimer's disease(AD) is a neurodegenerative disease characterized by memory loss and cognitive impairment. To date, however, no disease-modifying strategies to prevent or cure AD exist. Synapses are involved in the connection of neurons and present as the key component for the memory and other neural activities. Synapse loss is a critical hallmark of AD pathology. In brain, glia cells, including microglia and astrocytes, are a group of highly specific cell types other than neurons. Microglia and astrocytes play a key role in maintaining the healthy neural circuit and regulating synaptic plasticity. Under development and physiological conditions, glial cells contribute to construct and maintain mature central neural networks via synaptic pruning. However, during AD pathogenesis, glial cells engulf synapses excessively, which leads to synapse loss, neuronal dysfunction, and cognitive impairment. Here, we review recent advances in our understanding of the underlying mechanisms for glia-mediated synaptic pruning in AD, and provide a novel strategy for the development of AD drugs.

Ulcerative colitis(UC) is a chronic non-specific inflammatory bowel disease, listed as a modern refractory disease by the World Health Organization, which is difficult to recover, whereas it is easy to be attacked repeatedly. UC pathogenesis is closely related to gut microbiota dysbiosis. The gut microbiota interacts with bile acids(BAs), short-chain fatty acids(SCFAs), tryptophan, and other metabolism, immune system, intestinal barrier, etc., which regulate each other and affect the occurrence and development of UC. The active ingredients of traditional Chinese medicine(TCM), single herb and its extracts, and formulae can effectively alleviate UC symptoms by regulating the diversity, structure, composition, and metabolites of gut microbiota. In this review, the TCM based on the regulation of gut microbiota in the treatment of UC and its related mechanism for nearly three years was summarized.

Natural products and their derivatives are important components of anti-tumor drugs. Currently, anti-tumor drugs derived from natural products which are in clinical practice are mainly conventional cytotoxic or molecularly targeted drugs. Their application is limited by drug-related side effects and drug resistance. Recent studies have shown that anti-tumor natural products often act on multiple targets in tumor cells and in turn interfere with multiple processes in tumorigenesis and development. As tumor is a systemic disease induced by multiple factors, multi-targeted natural products possess unique potential in tumor therapy. However, the targets and mechanisms of the discovered multi-targeted antitumor natural products remain elusive, which limits their further development and application. This review summarized the research progress in the mechanism of action, target identification, and structure optimization of multi-targeted anti-tumor natural products exemplified by a few typical compounds. The research and development of these agents have also been proposed.

Protein kinases are intimately involved in the pathogenesis of many diseases such as cancer, inflammation, and autoimmune and neurological diseases. Therefore, kinases have been widely studied as drug targets over the past three decades. As of April, 2020, the FDA had approved 59 small molecule kinase inhibitors(SMKIs)in the emerging field of targeted drug therapy. This paper focuses on the biochemistry and pharmacology of these59 SMKIs and 121 SMKIs for which structures can be retrieved and that are now in phase Ⅱ and Ⅲ clinical trials.In addition, this paper also conducts a simple analysis of several popular targets and their inhibitors.

Transporters have a great influence on the transportation and distribution of drugs in the body. On the one hand, solute carrier transporters could transport drugs into tissues and organs, which may improve the oral bioavailability or change the tissue-distribution of the drugs. On the other hand, the ATP-binding cassette could pump some drugs out of the cell, which decreases the intracellular drug concentrations and leads to drug resistance.This paper summarizes the distribution, substrate characteristics and drug design strategies of several important drug transporters, such as improving bioavailability by prodrug design, introducing acid group to improve hepatic selectivity and adjusting the polarity of compounds to decrease efflux ratio.

With high selectivity and potency, target protein degradation technology has recently emerged as a strategy for drug discovery and design. Proteolysis-targeting chimeras(PROTAC) function as inducers for the degradation of target proteins and are a research focus in drug development. Current research on PROTAC mainly revolves around the rational design of PROTAC molecules, the discovery of new E3 ubiquitin ligase ligands and improvement in drug targeting. In this review, we focus on the PROTAC linker and its effects on the generation of the E3 enzyme-PROTAC-target protein ternary complex from three standpoints: length, binding site and chemical properties. We discuss the influences of the linker on the efficacy and the selectivity of PROTAC molecules.

Biomarkers are defined as a characteristic that is objectively measured and evaluated as an indicator of normal biological processes, pathogenic processes, or pharmacological responses to a therapeutic intervention.Biomarkers can help the decision-making process for new drug research and development, provide guidance for the early clinical development of candidate drugs and reduce the risk of failure. Therefore, as a key factor in the development of new drugs, the discovery and research on biomarkers has increased the interest of the pharmaceutical industry and regulatory agencies. Guidelines on the development and use of biomarkers have been issued by drug regulatory agencies including the EMA, FDA and ICH. Biomarkers are encouraged to be used to facilitate drug development by these relevant regulatory agencies, and also to be used to monitor the safety and efficacy of drugs in post-marketing drug surveillance. The application of biomarkers is encouraged at different stages of a drug's life cycle, including at the stage of basic science research and target identification, prototype design or discovery, preclinical development, clinical development, FDA filling/approval and launch, as well as post-marketing was reviewed. The identification, development, and application of biomarkers in pharmaceutical research is discussed.

During growth and progression, the microenvironment of tumors suffers a series of abnormal characteristics, which include hypoxia, acid pH, increased oxidative stress, excess glutathione(GSH), as well as certain overexpressed enzymes. Although affect or limit the cancer therapeutic outcomes, these factors provide possible approaches to strategies for cancer detection and novel therapy at the same time. Recently, based on these properties of the tumor microenvironment(TME), various kinds of responsive nano-platforms have been continuously developed and applied in cancer theranostics preliminarily. Thus, this review would introduce the typical features of TME firstly, then detailly summarize the design principles and research progress of corresponding hypoxiaresponsive, pH-responsive, redox-responsive, enzyme-responsive, dual-responsive and multi-responsive nanoplatforms. Finally, the challenges and the perspectives of the TME-responsive nano-platforms are briefly discussed.

Compared with normal tissues and cells, the tumor microenvironment has significant differences.For example, glutathione-related metabolic enzymes and reactive oxygen species are highly expressed in different subcellular structures, resulting in an unbalanced redox state. Aiming at the specific redox state in tumor tissues and cells, a series of small molecule prodrug self-assembled nanoparticles can be designed and connected by intelligent response linkers including disulfide bonds, sulfide bonds, and selenium bonds, thioketal bonds, etc. The in vitro and in vivo efficiency and metabolic mode of these nanoparticles are related to the type of linker. This review will summarize the tumor redox microenvironment, the design of intelligent responsive small molecule prodrug nanoparticles, and the metabolic pathways of small molecule prodrug nanoparticles with different connecting linkers and their relationship with drug efficacy.

Nanoparticles have better applicability in the detection, treatment of cancer and various difficult diseases, but mononuclear phagocytosis system can seriously shorten the time of nanoparticles in vivo circulation, reduce the drug efficacy. The protein crown formed on the surface of the nanoparticle after entering the body can change its surface properties, interfere with the recognition of phagocytes, and thus affect its circulation time in vivo.This article outlines the general composition and formation process of protein crowns. It also summarizes the influence of the physical and chemical properties of nanoparticles, such as particle size, surface charge, hydrophilicity and surface materials on the formation of protein crowns. The protein crown affects the circulation of nanoparticles in vivo, mainly because the adsorbed opsonic protein promotes cell phagocytosis. Therefore, we also introduce the method of using protein crowns to promote the long circulation of nanoparticles in vivo. By designing appropriate physical and chemical properties, surface modification, and directed design of protein crowns, the adsorption of proteins on the surface of nanoparticles can be reduced. Therefore, it can reduce the clearance of nanoparticles in the mononuclear phagocytic system(mainly the phagocytes of the liver and spleen), and achieve the goal of long circulation of nanoparticles in the body.

Antibody-drug conjugates(ADCs) are one of the most important classes of anticancer therapeutics.Human epidermal growth factor receptor-2(HER2), which is highly expressed in many types of aggressive cancers including breast and ovarian cancer, has been approved as an ideal target for ADCs. Lidamycin(LDM), developed by Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences, is an enediyne-containing antibiotic with potent anti-tumor activity. LDM is a promising payload for ADCs. In the present research, using a special site-directed conjugating technology, we made a novel ADC(607-LDM) with a drug-to-antibody ratio(DAR) of 2 and composed of the anti-HER2 antibody 607 and LDM. The new ADC exhibited potent antitumor activity against human ovarian cancer SKOV3 and breast cancer BT-474 cells. It also induced apoptosis and G2/M arrest.In nude mice with SKOV3 xenografts and a tumor volume of 150-200 mm³, a single intravenous injection 607-LDM at 1 mg·kg^-1 induced tumor growth inhibition of 72.4%, which was significant compared to either LDM(50.6%) or antibody(30.2%) treatment alone, or both in combination(50.1%, P < 0.05). All animal experiments were performed in accord with National Regulations and approved by the Animal Experiments Ethical Committee of College of Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences. The novel ADC designed in this study, 607-LDM, is a promising candidate for the treatment of HER2-positive cancers.

Drug use during pregnancy is unavoidable. Therefore, it is vitally important for medical workers to help pregnant women take drugs correctly to reduce the incidence of spontaneous abortion, premature birth, and low birth weight. In our study, drug screening model with induced pluripotent stem cells(iPSCs) was used to find some improper drugs which will result in woman's abortion. With 3D culture in vitro, iPSCs can form embryoid bodies(EBs) and cerebral organoids, which simulated in vitro development of early embryos, from inner cell mass to germ-layer differentiation. In the experiment, EBs were exposed to mifepristone(RU486), and three experimental groups were divided randomly. They were control group(without RU486), low-dose group(L-RU486, 10 μg·mL^-1), and high-dose group(H-RU486, 20 μg·mL^-1). After mifepristone exposure, EBs were observed at days 5, 8, and 11, including size of EB, cell apoptosis, and differentiation of germ layers, by using inverted optical microscope, TUNEL assay, and immunofluorescent staining. The results showed that through 3D culture, iPSCs could develop into embryoid bodies, neural rosettes, and finally cerebral organoids. After mifepristone exposure, EBs' sizes were decreased(P < 0.01); the levels of cell apoptosis in EBs were increased after mifepristone exposure(P < 0.01); the development of EBs' germ layer was affected. Mifepristone exposure could inhibit the proliferation of embryonic stem cells, reduce the differentiation of ectoderm(P < 0.01) and promote the development of mesoderm(P < 0.05).In conclusion, iPSCs can be used as a screening model for abortion drug, and EBs' diameter, cell apoptosis, and differentiation changes of the germ layers can serve as criteria of abortion drug screening.

Osteoprotegerin(OPG), secreted by osteoblasts, is a marker of bone turnover. OPG can inhibit osteoclastic differentiation by binding receptor activator of nuclear factor-κB ligand(RANKL). In this study, we found that rutaecarpine(RUT) had the up-regulating OPG activity, and it could significantly increase OPG protein levels in both mouse embryonic osteogenic precursor MC3 T3-E1 and human osteosarcoma U-2OS cells. Osteoblastogenic differentiation calcified nodules staining results showed that RUT significantly promoted the osteogenic differentiation of MC3 T3-E1 cells. Osteoclastic differentiation tartrate resistant acid phosphatase(TRAP) staining results showed that RUT obviously inhibited the osteoclast differentiation of mouse macrophages RAW264.7 induced by RANKL. In vivo studies showed that low-dose RUT group(5 mg·kg^-1·day^-1) and high-dose RUT group(45 mg·kg^-1·day^-1) treatments for 3 months significantly increased bone density in ovariectomized(OVX) rats; calcein double labeling experiment and toluidine blue staining results indicated that low-dose RUT group promoted bone formation and decreased bone loss in vivo; immunohistochemistry results showed that low-dose RUT group increased the expression of OPG in rat femur. All animal procedures were performed in accordance with the regulations of the Institutional Animal Care and Use Committee of Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences. In summary, this study demonstrated that RUT could up-regulate OPG expression and had promoting osteoblastic differentiation and inhibiting osteoclastic differentiation effects in vitro and in vivo.

We investigated the inhibitory effect and mechanism of action of bruceantin (BCT) on the proliferation, invasion and migration of non-small cell lung cancer (NSCLC) cells. The cytotoxic activity of BCT was measured by MTT assay; a colony forming assay, wound healing assay, and a Transwell assay were used to investigate the anti-proliferative, anti-migration, and anti-invasion effects, respectively; immunoblotting and RT-qPCR were used to detect the expression of related proteins, miRNA, and mRNA, respectively, that were involved in cell proliferation, migration, and invasion. Two gene prediction websites were used to predict the downstream target gene of miRNA. Our results show that BCT has a potent cytotoxic effect on NSCLC cell lines, with a half maximal inhibitory concentration (IC₅₀) of BCT against H1299, PC-9, and A549 of 0.12 ± 0.02, 0.31 ± 0.20, and 2.07 ± 0.70 μmol·L^-1, respectively. When H1299 cells were treated with 0.03, 0.15, and 0.75 μmol·L^-1 BCT for 24 h, the proliferation, migration, and invasive ability were inhibited in a concentration-dependent manner. It is worth noting that the expression level of miRNAs related to cell migration and invasion, such as miR-29a-3p, miR-21-3p, miR-183-5p, and miR-34b-5p increased with the concentration of BCT, especially for miR-29a-3p. Using the two gene prediction websites, we predict that integrin β1 (ITGB1) may be the target gene of miR-29a-3p; immunoblot results further show that a variety of proteins related to cell proliferation, migration, and invasion, such as various proteins of the integrin family, β-catenin, p-Src, and vascular endothelial growth factor, all decreased in a concentration-dependent manner, among which the reduction of ITGB1 protein was the most obvious. RT-qPCR results showed that there was no change in ITGB1 mRNA expression. We speculate that BCT might inhibit the expression of ITGB1 protein by up-regulating miR-29a-3p independent of its mRNA level. The in-depth mechanism needs to be further explored. This study suggests that BCT has the potential for further development in the treatment of NSCLC.

The antidepressant effect of Xiaoyaosan has been demonstrated. It is of value to explore the biological mechanism of Xiaoyaosan in the treatment of depression from the perspective of functional modules by using the method of functional module division of the metabolic network. The differential metabolites and related enzymes and proteins regulated by Xiaoyaosan were identified in the database. Pathway enrichment analysis and crosstalk pathway analysis of Xiaoyaosan regulated metabolites was carried out. A network of differentially regulated metabolites and their enzymes and proteins was constructed by using the STRING tool. The CNM decomposition algorithm was used to extract the functional modules of the network and enrichment analysis of functional modules was carried out. The results show that Xiaoyaosan regulates 97 differential metabolites, 234 related enzymes and258 depression-related proteins. The pathways crosstalk analysis was divided into two sub-networks, one of which is related to the neural system and cell signal transduction, the other is related to the endocrine system and metabolic pathways. KEGG pathway enrichment analysis of the network and 9 functional modules extracted by the CNM algorithm shows that module 1 and module 3 belong to the pathways that can be enriched into more pathways with fewer proteins. The corresponding functions of these pathways include the endocrine system, amino acid metabolism, the nervous system and signal transduction. In this study, pathway crosstalk analysis and metabolic network module division strategies were used to explain the biological mechanism of Xiaoyaosan in the treatment of depression, providing ideas and methods for in-depth study of the pharmacological mechanism of this traditional Chinese medicine from the perspective of metabolic regulation.

M701 is a bispecific CD3/EpCAM T-cell engager antibody for the treatment of malignant ascites. We developed a population pharmacokinetic/pharmacodynamic (PK/PD) model to quantitatively describe and predict the antitumor effect of M701 in human colorectal cancer xenograft mice. We developed the M701 PK model based on plasma concentration data after i.v. administration. A tumor growth model for human colorectal cancer xenograft was developed to evaluate the antitumor effect of M701. We additionally simulated the inhibitory effect of M701 on tumor volume under different dose regimens based on a PK/PD model. A two-compartment model was developed to predict the PK in human colorectal cancer xenograft mice. The relationship between the M701 concentration and tumor growth inhibition was characterized by a combined Simeoni tumor growth/transit compartment model. The estimated pharmacodynamic parameters were related to the tumor growth characteristics λ₀ (0.212 d^-1) and λ₁ (0.044 7 cm³·d^-1), to the drug potency k₂ (0.071 5 mL·ng^-1·d^-1), and to the kinetics of tumor cell death k₁ (2×10^-5 d^-1). A model visual predictive check showed that both the PK model and the tumor growth model closely fit the observed data. Simulated tumor growth after administration of M701 (0.5 mg·kg^-1 every 6 days and 0.25 mg·kg^-1 every 3 days) could be effectively inhibited. This population PK/PD model of M701 provides insight into the antitumor effect of M701 and supports the further therapeutic development of M701.

The purpose of this study was to discover novel inhibitors of sirtuin-1(SIRT1) that could be used in the treatment of acute myeloid leukemia(AML).Eight potential SIRT1 inhibitors were identified from 231 511 natural drug-like molecules by virtual screening-based molecular docking and molecular mechanics-generalized Born surface area(MM-GBSA) calculation of binding free energies.Using existing SIRT1 inhibitor molecules as training and test sets, a series of quantitative structure-activity relationship models were established, and the best quantitative structure-activity relationship(QSAR) model was used to predict the IC₅₀ of these 8 potential inhibitor molecules for SIRT1.Subsequently, molecular dynamics simulations were performed to verify the binding mode and stability of these complexes of potential inhibitors and SIRT1 protein.Finally, the activity of these potential SIRT1 inhibitors was verified by cell proliferation assays of OCI-AML2, OCI-AML3 and MV4-11 cells and SIRT1 enzyme activity assays, and it was found that 5 compounds could inhibit AML cell proliferation.Among them, the most active compound, ZINC000001774455, had an IC₅₀ of 2.29 ± 0.09 μmol·L^-1 with OCI-AML2 cells, and at a concentration of 1 μmol·L^-1, the inhibitory ratio of this compound on SIRT1 protein activity was 65.33%.ZINC000001774455 can be used as a lead compound for the development of new AML treatments.

(±)-Bicoryanhunine B(1), a new dimeric benzylisoquinoline alkaloid was isolated from the dried tubers of Corydalis yanhusuo by various chromatographic methods, including silica gel, Sephadex LH-20, reverse phase C18, and semi-preparative HPLC.Its structure was determined by spectroscopic methods, including UV, IR, ESI-MS, HR-ESI-MS and 1D/2D NMR.(±)-Bicoryanhunine B(1) was a moderate PD-1/PD-L1 interaction inhibitor with an IC₅₀ value of 7.80±0.49 μmol·L^-1.In addition, 1 exhibited potent inhibitory activities against LPS-induced NO production in RAW 264.7 macrophages with an IC₅₀ value of 4.83 ± 2.21 μmol·L^-1.

A quantitative analytical method based on HPLC coupled with the charged aerosol detector (CAD) for quantitative analysis of multi-components with a single marker (QAMS) was established for simultaneous determinations of astragaloside Ⅰ, astragaloside Ⅱ, astragaloside Ⅳ, calycosin-7-O-β-D-glucoside, formononetin and 7, 2'-dihydroxy-3', 4'-dimethoxyisoflavan in Astragalus membranaceus. The separation was performed on an Agilent SB-C18 (150 mm×4.6 mm, 3.5 μm), with gradient elution using the mobile phase consisting of 0.05% formic acid solution and 0.05% formic acid acetonitrile at the flow rate of 1.0 mL·min^-1. The column temperature was 35 ℃, and the injection volume was 20 μL. For CAD, the drift tube temperature was at 50 ℃. The contents of six components in A. membranaceus were determined by both external standard method (ESM) and QAMS, and then were compared. The results showed that chromatographic peaks were separated well and the linear ranges of astragaloside Ⅰ, astragaloside Ⅱ, astragaloside Ⅳ, calycosin-7-glucoside, formononetin and 7, 2'-dihydroxy-3', 4'-dimethoxyisoflavan were 0.113-2.250 mg·mL^-1, 0.012-0.240 mg·mL^-1, 0.004-0.080 mg·mL^-1, 0.065-1.300 mg·mL^-1, 0.005-0.100 mg·mL^-1 and 0.007-0.150 mg·mL^-1, respectively. The content ranges of astragaloside Ⅰ, astragaloside Ⅱ, astragaloside Ⅳ, calycosin-7-glucoside, formononetin and 7, 2'-dihydroxy-3', 4'-dimethoxyisoflavan were 0.306-0.922 mg·g^-1, 0.053-0.183 mg·g^-1, 0.015-0.092 mg·g^-1, 0.069-0.823 mg·g^-1, 0-0.098 mg·g^-1 and 0.020-0.107 mg·g^-1 in 20 batches of A. membranaceus, respectively. Using astragaloside Ⅱ as an internal reference, the relative correlation factors of astragaloside Ⅰ, astragaloside Ⅳ, calycosin-7-O-β-D-glucoside, formononetin, and 7, 2'-dihydroxy-3', 4'-dimethoxyisoflavan were calculated as 0.561, 0.835, 0.299, 0.796, and 0.799, respectively. The results were compared with those obtained by the external standard method to verify the feasibility, rationality and repeatability of QAMS method, and there was no significant difference in assay results between the two methods. In conclusion, the QAMS method is accurate and feasible, and could be used to determine the contents such as astragaloside Ⅰ, astragaloside Ⅱ, astragaloside Ⅳ, calycosin-7-glucoside, formononetin and 7, 2'-dihydroxy-3', 4'-dimethoxyisoflavan, and it can be used for quality control of A. membranaceus.

The bioactivity of a working reference standard was determined by replicate bioassays with calibration against a primary reference standard. In this study the number of bioassay replicates needed for calibration first was calculated theoretically, and if the mean value of the experimental bioassay replicates fell within the predefined bioactivity level the bioactivity of the working reference was defined as 100%. Our results showed that when the total intermediate precision of the bioassay method was at 11.66% and the predefined bioactivity level was set at 95%-105% with a confidence level of 95%, 21 bioassay replicates should be carried out for calibration.The average value of the 22 experimental bioassay replicates was 101.96%, so the bioactivity of the working reference standard was consistent with that of the primary reference standard at 100%. The results suggest that a strategy of first calculating the number of bioassay replicates needed for calibration and then determining whether the resulting experimental mean value is within the predefined bioactivity level will be of value to the biopharmaceutical industry.

Four salts of ticagrelor, ticagrelor-3, 5-dinitrobenzoic acid, ticagrelor-pyrazinamide, ticagrelor-Dproline and ticagrelor-L-proline were prepared by solvent suspension and liquid-assisted grinding to improve the solubility of ticagrelor. The compounds were characterized by powder X-ray diffraction, Fourier transform infrared spectroscopy, differential scanning calorimetry, nuclear magnetic resonance spectroscopy, elemental analysis, and the intermolecular salt-bonding forces were analyzed. The equilibrium solubility of salts and pure drug in hydrochloride buffer pH 1.2 and phosphate buffer pH 6.8 were measured by high-performance liquid chromatography.Ticagrelor was salted with 3, 5-dinitrobenzoic acid, pyrazinamide, D-proline, L-proline all in a stoichiometric ratio of 1∶1; with the exception of ticagrelor-D-proline, the solubility of the other three salts provided significantly improved solubility in hydrochloride buffer pH 1.2, and the equilibrium solubility of ticagrelor-3, 5-dinitrobenzoic acid was increased by approximately 1.7 folds as compared to pure drug. Salt-forming technology is convenient and can improve the solubility of ticagrelor.

Sacubitril valsartan sodium(LCZ696) is an ionic cocrystal drug. The purpose of this study was to explore the cocrystal features of LC696 by establishing a variety of characterization methods, and thus provide basic research data for effective quality control. The cocrystal characteristics of LCZ696 and its tablets were identified by applying analytical means including powder X-ray diffraction(PXRD), fourier transform infrared spectroscopy(FTIR), Raman spectra(RM), differential scanning calorimetry(DSC) and solid-state nuclear magnetic resonance spectroscopy(ssNMR). The crystalline water and hygroscopicity of LCZ696 were analyzed by thermogravimetric analysis(TGA), dynamic vapor sorption(DVS), hygroscopicity test and Karl Fischer reaction method.The results show that PXRD, FTIR, DSC and ssNMR can effectively distinguish the features of LCZ696 cocrystal, sacubitril monomer, valsartan monomer, and sacubitril-valsartan(1∶1) mixture. RM can be used as a supplementary approach. Combined with the analysis by TGA, DVS, hygroscopicity test and Karl Fischer reaction method results, LCZ696 contains 2.5 crystalline water molecules and is very hygroscopic; we recommend that LCZ696 be stored in an environment with a relative humidity below 60%. By characterizing the crystal features we can establish quality control measure and evaluate the stability of the drug tablets. This study provides data in support for the establishment of the LCZ696 quality standard.

Puerarin(PUE), as an isoflavone component, has a wide range of pharmacological activities, while its poorly aqueous solubility limits the development of solid oral dosage forms. In this study, PUE along with nicotinamide(NIC) were prepared into the coamorphous system by solvent-evaporation method and characterized by powder X-ray diffraction(PXRD), differential scanning calorimetry(DSC) and Fourier transform infrared spectroscopy(FT-IR). In addition, its dissolution behavior and solubilization mechanism were also investigated.PUE-NIC coamorphous was a single homogeneous binary system, with a single glass transition temperature at 35.1 ℃. In comparison to crystalline PUE, during the dissolution process, coamorphous PUE-NIC not only exhibited the "liquid-liquid phase separation"(LLPS) phenomenon, but the formation of A_p type complexation(1∶1 and 1∶2)between PUE and NIC molecules was also verified, which significantly improved the solubility of PUE and prolonged the supersaturation time, and would benefit its absorption.

The therapeutic effect of tumor photodynamic therapy is severely limited by the hypoxic tumor microenvironment. Inhibiting tumor celloxygen consumption is a more effective way than increasing its oxygen supply to overcome the tumor hypoxia and enhance photodynamic therapy. To carry out this strategy, the supramolecular nanoparticles VER-ATO-SMN loaded with photosensitizer verteporfin(VER), oxygen-consuming inhibitor atovaquone(ATO), and stabilizer polyvinylpyrrolidone(PVP)-K30 were prepared by the nanoprecipitation method, and the optimal prescription was screened and optimized by single factor experiments. The results showed that the optimal prescription for VER-ATO-SMN was ATO∶VER(w/w) = 1∶1, PVP-K30 = 100 mg, N, N-dimethylformamide∶water(v/v) = 1∶10. The morphology, particle size, particle dispersion index and encapsulation efficiency of supramolecular nanoparticles were characterized. The VER-ATO-SMN showed a spherical morphology and was well dispersed. The hydrodynamic size of VER-ATO-SMN was 101.21 ± 4.30 nm as determined by dynamic light scattering(DLS). The encapsulation efficiencies of VER and ATO in VER-ATO-SMN prepared with the optimal prescription were 70.86% and 77.52%, respectively. The VER-ATO-SMN exhibited good laser stability and also showed high stability in conditions which simulated the physiological solution. Compared with free VER and VER liposome, VER-ATO-SMN performed enhanced therapeutic effect at the cell level. The mechanism was that VERATO-SMN could effectively incorporate into cells and improving the intracellular oxygen concentration by reducing the oxygen consumption of tumor cells could increase the amount of reactive oxygen species generated by VER mediated photodynamic therapy. The in vivo anticancer efficacy results of tumor-bearing mice suggested that VER-ATO-SMN could effectively inhibit the tumor growth or even completely eliminate the tumor. All animal experiments were performed in line with national regulations and approved by the Animal Experiments Ethical Committee of 900 Hospital of the Joint Logistics Team.

After entering the physiological environment, proteins and other biomolecules bind to the nanoparticles' surface, called protein corona. The corona establishes a new bio-interface that affects its physicochemical properties and biological behaviors. Variations in types and contents of human plasma proteins during the different physiological states can substantially change the composition and effects of the corona. With folic acid(FA)-modified polylactic acid-polyglycolic acid copolymer(PLGA) nanoparticles, the formation of protein coronas and their influence on the targeting capability are studied in healthy and ovarian human plasma. All human plasma samples were collected at the Peking University Third Hospital and this study protocol has been approved by Peking University Third Hospital Medical Science Research Ethics Committee(2019-409-1). Dynamic light scattering measurements demonstrated a 10-40 nm increase in their size distributions and a 30 mV decreased in their absolute zeta-potential since protein corona-coated PLGA-PEG and PLGA-FA were formed. The SDS-PAGE analysis showed the composition of the protein coronas from ovarian and healthy plasma in PLGA-FA were markedly distinct, particularly for proteins with molecular weight of 45, 110 and > 180 kDa. Flow cytometry indicated that the absorption of ovarian plasma in PLGA-FA led to a lower cellular uptake by SKOV3 cells. Our results suggest that in vitro formed ovarian plasma protein corona could shield targeting molecules and reduced receptor-mediated internalization. The results of this pilot study will provide evidence of the effectiveness of active targeting nanoparticles under pathologic conditions. Additionally, the protein corona in different diseases is emerging as a key point; thus, a comprehensive understanding could accelerate clinical translation of functionalized nanoparticles.

As a key gene in the regulation of long-chain fatty acid biosynthesis, 3-ketoacyl-CoA synthase (KCS) plays an important role in the growth and development of Coix lacryma-jobi L. In this study, the KCS gene was cloned from cDNA of Coix lachryma-Jobi L. and bioinformatics analysis was performed. Results showed that the full length KCS gene was 1 548 bp encoding 515 amino acids. Bioinformatics analysis indicated that the gene encoded a 58 608.12 Da protein with an isoelectric point of 9.20 containing two transmembrane helical structure domains and lacking a signal peptide, with a likely subcellular localization in main plastid membranes. The results of multiple sequence comparisons and evolutionary tree analysis revealed that KCS had three identical conserved sequences and was closely related to KCS from monocotyledons such as Sorghum bicolor, Zea mays, Setaria italica, Panicum miliaceum, Oryza brachyantha, Hordeum vulgare, Aegilops tauschii subsp. Tauschii. We speculated that the evolution of the gene was similar among these plants of the same family. In addition, gene expression analysis showed that the KCS gene was significantly different in Coix lacryma-jobi L. isolates having different lipid content. This work will facilitate further study of the regulatory mechanism of this enzyme in fatty acid synthesis.

Bupleurum L.(Apiaceae) is an economically important genus, in which many species are of medicinal value. In this study, the complete plastid genomes(plastomes) of B. chinense DC. and B. boissieuanum H. Wolff were sequenced and their characteristics were investigated. Comparative and phylogenetic analyses were conducted with other published Bupleurum plastomes. The complete plastomes of B. chinense and B. boissieuanum were 155 458 and 155 800 bp in length, and both exhibited the typical quadripartite circular structure consisting of a large single copy region(LSC, 85 343 and 85 804 bp), a small single copy region(SSC, 17 495 and 17 410 bp), and a pair of inverted repeat regions(IRa/b, 26 310 and 26 293 bp), respectively. A total of 129 genes, including84 protein-coding genes, 37 transfer RNA(tRNA) genes, and eight ribosomal RNA(rRNA) genes were identified from each of the two plastomes. Repeat sequences detected were similar in types and distribution patterns, but the numbers were slightly different. Comparative analyses revealed that the Bupleurum plastomes were highly conserved in length, structure, the guanine and cytosine(GC) content, and gene content and order, both intraspecifically and interspecifically, and no obvious expansion or contraction of the inverted repeat regions occurred. Sequence variation was lower within the same species than among different species, noncoding sequences(including intergenic regions and introns) showed a higher divergence than the protein-coding sequences, and sequences in the LSC and SSC regions were more divergent than those in the IR regions. In addition, 11 sequences with higher nucleotide diversity among species were detected in the LSC and SSC regions. All studied Bupleurum species were inferred forming a monophyletic group with a 100% bootstrap value. Bupleurum chinense and B. boissieuanum were phylogenetically closest to B. commelynoideum and B. falcatum, separately, with all three B. chinense accessions clustered into a distinct clade. These results provide genetic information for further species identification, phylogenetic resolution, and will assist in exploration and utilization of medicinal Bupleurum species.

Chalcone isomerases (CHIs) play an essential role in the biosynthesis of flavonoids important in plant self-defense. Based on the transcriptome data of Aquilaria sinensis Calli, a full-length cDNA sequence of CHI1 (termed as AsCHI1) was cloned by reverse transcription PCR. AsCHI1 contains a complete open frame (ORF) of 654 bp. The deduced protein is composed of 217 amino acids, with a predicted molecular weight of 23.11 kDa. The sequence alignment and phylogenetic analysis revealed that AsCHI1 has conserved most of the active site residues in type I CHIs, indicating a close relationship with the CHI from Gossypium hirsutum. The recombinant AsCHI1 protein was obtained by heterologous expression of AsCHI1 in E. coli BL21(DE3). The purified AsCHI1 protein exhibited CHI activity by catalyzing the production of naringenin from naringenin chalcone. Remarkably, AsCHI1 expression in A. sinensis Calli treated with various abiotic stresses including salt, mannitol, cold, and heavy metals could be markedly increased, and plant hormones such as abscisic acid (ABA), gibberellin (GA3), and salicylic acid (SA) could also increase the expression of AsCHI1, suggesting that AsCHI1 might play an important role in plant self-defense. The results expand our understanding of the biosynthesis of flavonoids in A. sinensis and give further insight into the defensive responses of A. sinensis to abiotic and biotic stresses.