Multidimensional virtual screening approaches combined with drug repurposing to identify potential covalent inhibitors of SARS-CoV-2 3CL protease.

The outbreak of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has caused an unprecedented global pandemic, and new cases are still on the rise due to the absence of effective medicines. However, developing new drugs within a short time is extremely difficult. Repurposing the existing drugs provides a fast and effective strategy to identify promising inhibitors. Here we focus on the screening of drugs database for discovering potential covalent inhibitors that target 3-chymotrypsin-like protease (3CLpro), an essential enzyme mediating viral replication and transcription. Firstly, we constructed a receptor-ligand pharmacophore model and verified it through decoy set. The importance of pharmacophore features was evaluated by combining molecular dynamics simulation with interaction analyses. Then, covalent docking was used to perform further screening. According to docking score and Prime/Molecular Mechanics Generalized Born Surface Area (MM-GBSA) score, total ten compounds obtained good scores and successfully established covalent bonds with the catalytic Cys145 residue. They also formed favorable interactions with key residues in active sites and closely integrated with 3CLpro with binding modes similar to known 3CLpro inhibitor. Finally, the top four hits DB08732, DB04653, DB01871 and DB07299 were further subjected to 100 ns molecular dynamics (MD) simulation and MM-GBSA binding free energy calculations. The results suggest that the four candidates show good binding affinities for 3CLpro, which warrants further evaluation for their in-vitro/in-vivo activities. Overall, our research methods provide a valuable reference for discovering promising inhibitors against SARS-CoV-2 and help to fight against the epidemic.Communicated by Ramaswamy H. Sarma.

Synthesis and bioactivity evaluation of pachymic acid derivatives as potential cytotoxic agents.

Pachymic acid, a well-known natural lanostane-type triterpenoid, exhibits various pharmacological properties. In this study, 18 derivatives of pachymic acid were synthesized by modifying their molecular structures and evaluated for their anticancer activity against two human cancer cell lines using the CCK-8 assay. Structure-activity relationship studies according to the in vitro cytotoxicity unexpectedly found one promising derivative A17 (namely tumulosic acid, also found in Poria cocos), which had stronger anti-proliferative activity than the positive drug cisplatin against HepG2 and HSC-2 cell lines with IC50 values of 7.36 ± 0.98 and 2.50 ± 0.15 µM, respectively. Further pharmacological analysis demonstrated that A17 induced HSC-2 cell cycle arrest at the S phase, cell apoptosis, and autophagy. Western blotting confirmed the regulatory effects of A17 on cell cycle arrest-, apoptosis-, and autophagy-related proteins expression. In addition, A17 regulated the AKT and AMPK pathways in HSC-2 cells. These results demonstrated that A17 possesses great potential as an anticancer agent.

Pharmacological profiles and therapeutic applications of pachymic acid (Review).

Poria cocos is a saprophytic fungus that grows in diverse species of Pinus. Its sclerotium, called fu-ling or hoelen, has been used in various traditional Chinese medicines and health foods for thousands of years, and in several modern proprietary traditional Chinese medicinal products. It has extensive clinical indications, including sedative, diuretic, and tonic effects. Pachymic acid (PA) is the main lanostane-type triterpenoid in Poria cocos. Evidence suggests that PA has various biological properties such as cytotoxic, anti-inflammatory, antihyperglycemic, antiviral, antibacterial, sedative-hypnotic, and anti-ischemia/reperfusion activities. Although considerable advancements have been made, some fundamental and intricate issues remain unclear, such as the underlying mechanisms of PA. The present study aimed to summarize the biological properties and therapeutic potential of PA. The biosynthetic, pharmacokinetic, and metabolic pathways of PA, and its underlying mechanisms were also comprehensively summarized.

Anticancer potential of indirubins in medicinal chemistry: Biological activity, structural modification, and structure-activity relationship.

Indirubin is the crucial ingredient of Danggui Longhui Wan and Qing-Dai, traditional Chinese medicine herbal formulas used for the therapy of chronic myelocytic leukemia in China for hundreds of years. Although the monomeric indirubin has been used in China for the treatment human chronic myelocytic leukemia. However, due to low water solubility, poor pharmacokinetic properties and low therapeutic effects are the major obstacle, and had significantly limited its clinical application. Consequently, the attractive anticancer profile of indirubin has enthused numerous researchers to discover novel indirubin derivatives with improved pharmacodynamic activity as well as good pharmacokinetic property. In this paper, we comprehensively review the recent progress of anticancer potential of indirubins, structural modification and structure-activity relationship, which may provide useful direction for the further development of novel indirubins with improved pharmacological profiles for the treatment of various types of cancer.

Mechanistic Exploration of Methionine 274 Acting as a "Switch" of the Selective Pocket Involved in HDAC8 Inhibition: An in Silico Study.

The overexpression of histone deacetylase 8 (HDAC8) causes several diseases, and the selective inhibition of HDAC8 has been touted as a promising therapeutic strategy due to its fewer side effects. However, the mechanism of HDAC8 selective inhibition remains unclear. In this study, flexible docking and in silico mutation were used to explore the structural change of methionine (M274) during HDAC8 binding to inhibitors, along with the reason for this change. Meanwhile, steered and conventional molecular dynamics simulations were employed to explore the stability of the structural change. The findings suggest that M274 acts as a "switch" to control the exposure of the HDAC8-selective pocket. The structure of M274 changes from flipped-out to flipped-in only when L-shaped inhibitors bind to HDAC8. This structural change forms a groove that allows these inhibitors to enter the selective pocket. In other HDACs, a leucine residue replaces M274 in situ, and the same structural change is not observed. The findings reveal the mechanism of selective HDAC8 inhibition and provide guidance for the development of novel selective inhibitors.

SARs of a novel series of s-triazine compounds targeting vimentin to induce methuotic phenotype.

Herein, we describe the design, synthesis and structure-activity relationships of a series of novel s-triazine compounds can induce methuotic phenotype in various types of cancer cells. (E)-1-(4-Chlorophenyl)-3-(4-((4-morpholino-6-styryl-1,3,5-triazine-2-yl)amino)phenyl)urea, compound V6, exhibited a striking methuotic phenotype with a minimal effective concentration of less than 10 nM in U87 glioblastoma cells. Based on structure-activity relationship studies, we designed and synthesized an active probe P1 that retained the full potential of V6 in inducing the methuotic phenotype in U87 glioblastoma cells. Using this probe following affinity-based proteomic profiling strategy, we identified vimentin as the specific target protein of compound V6. Molecular docking revealed that V6 can form hydrogen bonds with vimentin at 273R and 276Y in its rod domain.

Hierarchical virtual screening of the dual MMP-2/HDAC-6 inhibitors from natural products based on pharmacophore models and molecular docking.

The dual-target inhibitors tend to improve the response rate in treating tumors, comparing with the single-target inhibitors. Matrix metalloproteinase-2 (MMP-2) and histone deacetylase-6 (HDAC-6) are attractive targets for cancer therapy. In this study, the hierarchical virtual screening of dual MMP-2/HDAC-6 inhibitors from natural products is investigated. The pharmacophore model of MMP-2 inhibitors is built based on ligands, but the pharmacophore model of HDAC-6 inhibitors is built based on the experimental crystal structures of multiple receptor-ligand complexes. The reliability of these two pharmacophore models is validated subsequently. The hierarchical virtual screening, combining these two different pharmacophore models of MMP-2 and HDAC-6 inhibitors with molecular docking, is carried out to identify the dual MMP-2/HDAC-6 inhibitors from a database of natural products. The four potential dual MMP-2/HDAC-6 inhibitors of natural products, STOCK1 N-46177, STOCK1 N-52245, STOCK1 N-55477, and STOCK1 N-69706, are found. The studies of binding modes show that the screened four natural products can simultaneously well bind with the MMP-2 and HDAC-6 active sites by different kinds of interactions, to inhibit the MMP-2 and HDAC-6 activities. In addition, the ADMET properties of screened four natural products are assessed. These found dual MMP-2/HDAC-6 inhibitors of natural products could serve as the lead compounds for designing the new dual MMP-2/HDAC-6 inhibitors having higher biological activities by carrying out structural modifications and optimizations in the future studies.

Screening for the selective inhibitors of MMP-9 from natural products based on pharmacophore modeling and molecular docking in combination with bioassay experiment, hybrid QM/MM calculation, and MD simulation.

Matrix metalloproteinase-9 (MMP-9) has been considered as an attractive target involving cancer therapy. In this study, the 3D QSAR pharmacophore model of MMP-9 inhibitors is built, and its reliability is subsequently validated based on different methods. The built pharmacophore model consists of the four chemical features, including two hydrogen bond acceptors (HBA), one hydrophobic (HY), and one ring aromatic (RA). Among them, both HY and RA are found to be especially important features because they involve the interactions of inhibitors with the S1' pocket of MMP-9, which determines the selectivity of MMP-9 inhibitors. By combining pharmacophore model with molecular docking, the virtual screening is carried out to identify the selective MMP-9 inhibitors from natural products. The four potential selective MMP-9 inhibitors of natural products are found. One of them was used to carry out the bioassay experiment inhibiting MMP-9, and the estimated IC50 value of only 26.94 µM clearly shows its strongly inhibitory activity; besides, both the hybrid quantum mechanics/molecular mechanics (QM/MM) calculation and the molecular dynamics simulation are performed to examine the reliability regarding the binding mode of this inhibitor with MMP-9 active sites predicted by molecular docking. All the screened four natural products are found to well bind with the MMP-9 active sites by different kinds of interactions. Finally, the ADMET properties of screened four natural products are assessed. These screened MMP-9 inhibitors of natural products could be used as the lead compounds to perform structural modifications and optimizations in the future work. Communicated by Ramaswamy H. Sarma.

Synthesis and biological evaluation of novel 2-arylvinyl-substituted naphtho[2,3-d]imidazolium halide derivatives as potent antitumor agents.

Two series of novel 2-arylvinyl-naphtho[2,3-d]imidazol-3-ium iodide derivatives and 2-arylvinyl-naphtho[2,3-d]imidazol-3-ium bromide derivatives were designed and synthesized by the structural combination of YM155 with stilbenoids. All compounds were tested for anti-proliferative activity against PC-3, A375 and HeLa human cancer cell lines. Two of the compounds were selected for further investigation: 12b, which showed potent cytotoxicity against the three tested cell lines with IC50 values in the range of 0.06-0.21 µM, and 7l, which displayed excellent selectivity for PC-3 cells with an IC50 of only 22 nM. Western blot analysis results indicated that both 12b and 7l suppress the expression of Bcl-2 and Survivin proteins, which helps induce apoptosis. As determined by the percent of Annexin V-FITC-positive apoptotic cells, 12b was not only significantly more effective than 7l at a concentration of 100 nM in PC-3 cells but also induced apoptosis in a dose-dependent manner with more potency than 7l at a concentration of 1000 nM in A375 cells. Therefore, compound 12b was chosen for further in-depth studies investigating the mechanism of apoptosis. The results showed that it could activate caspase-3, hydrolyze PARP, and even inactivate ERK. Moreover, 12b arrested A375 cells at S phase in a time-dependent and dose-dependent manner, while having a visible effect on microtubule dynamics. In addition, (E)-2-(2-(1H-indol-3-yl)vinyl)-1-benzyl-3-(2-methoxyethyl)-4,9-dioxo-4,9-dihydro-1H-naphtho[2,3-d]imidazol-3-ium bromide (12b) exhibited significant antitumor activity when evaluated in a subcutaneous solid tumor model. Our study reveals that 2-arylvinyl-substituted naphtho[2,3-d]imidazolium scaffolding is a promising new entity for the development of multi-target anticancer drugs.

Glioma and microenvironment dual targeted nanocarrier for improved antiglioblastoma efficacy.

Drug delivery systems based on nanoparticles (nano-DDS) have aroused attentions for the treatment of glioblastoma (GBM), the most malignant brain cancer with a dismal prognosis. However, there are still numerous unmet challenges for traditional nano-DDS, such as the poor nanoparticle penetration, short retention in the GBM parenchyma and low glioma targeting ability. Herein, we used Pep-1 and CREKA peptides to construct a novel multifunctional GBM targeting nano-DDS (PC-NP). Pep-1 was used to overcome the blood-brain tumor barrier (BBTB) and home to glioma cells via interleukin-13 receptor-α2-mediated endocytosis, and CREKA was used to bind to fibrin-fibronectin complexes abundantly expressed in tumor microenvironment for enhanced retention in the GBM. Biological studies showed that the cellular uptake of PC-NP by U87MG cells was significantly enhanced compared with the non-targeting NP. Furthermore, CREKA modification increased the binding capacity of PC-NP to fibrin-fibronectin complexes as confirmed by the competition experiment. In accordance with the increased cellular uptake, PC-NP remarkably increased the cytotoxicity of its payload paclitaxel (PTX) against U87MG cells with an IC50 of 0.176 µg/mL. In vivo fluorescence imaging and antiglioma efficacy evaluation further confirmed that PC-NP accumulated effectively and penetrated deeply into GBM tissue. PC-NP-PTX exhibited a median survival time as long as 61 days in intracranial GBM-bearing mice. In conclusion, our findings indicated PC-NP as a promising nano-DDS for GBM targeting delivery of anticancer drugs.

A Novel Synthesis of the Efficient Anti-Coccidial Drug Halofuginone Hydrobromide.

Background: Halofuginone hydrobromide (1) is recognized as an effective drug against several species of Eimeria (E.) in poultry. In this paper, we describe a convenient and low cost preparation method for the compound, as well as primary validation of its activity. Methods: First, 7-bromo-6-chloroquinazolin-4(3H)-one (2) was prepared from m-chlorotoluene by a conventional process, and then chloroacetone was creatively introduced in two steps. Finally, halofuginone hydrobromide (1) was obtained from 7-bromo-6-chloro-3-(3-cholroacetonyl) quinazolin-4(3H)-one (4) by a four-step reaction sequence including condensation, cyclization, deprotection and isomerization. The structures of the relative intermediates and target compound were characterized by melting point, IR, MS and ¹H-NMR. Besides, the protective effect of compound 1-supplemented chicken diet at doses of 6, 3 and 1.5 mg per 1 kg were evaluated on chickens infected with E. tenella, by reduction in mortality, weight loss, fecal oocyst excretion and gut pathology, respectively. Results: Halofuginone hydrobromide (1) was prepared successfully by and improved and innovative method based on traditional research. Moreover, the synthesized halofuginone hydrobromide significantly exhibited an anti-coccidial property. Conclusions: The fruitful work described in this Communication has resulted in halofuginone hydrobromide, which has a good pharmaceutical development prospects, becoming more available for large-scale production.

Multiple receptor-ligand based pharmacophore modeling and molecular docking to screen the selective inhibitors of matrix metalloproteinase-9 from natural products.

Matrix metalloproteinase-9 (MMP-9) is an attractive target for cancer therapy. In this study, the pharmacophore model of MMP-9 inhibitors is built based on the experimental binding structures of multiple receptor-ligand complexes. It is found that the pharmacophore model consists of six chemical features, including two hydrogen bond acceptors, one hydrogen bond donor, one ring aromatic regions, and two hydrophobic (HY) features. Among them, the two HY features are especially important because they can enter the S1' pocket of MMP-9 which determines the selectivity of MMP-9 inhibitors. The reliability of pharmacophore model is validated based on the two different decoy sets and relevant experimental data. The virtual screening, combining pharmacophore model with molecular docking, is performed to identify the selective MMP-9 inhibitors from a database of natural products. The four novel MMP-9 inhibitors of natural products, NP-000686, NP-001752, NP-014331, and NP-015905, are found; one of them, NP-000686, is used to perform the experiment of in vitro bioassay inhibiting MMP-9, and the IC50 value was estimated to be only 13.4 µM, showing the strongly inhibitory activity of NP-000686 against MMP-9, which suggests that our screening results should be reliable. The binding modes of screened inhibitors with MMP-9 active sites were discussed. In addition, the ADMET properties and physicochemical properties of screened four compounds were assessed. The found MMP-9 inhibitors of natural products could serve as the lead compounds for designing the new MMP-9 inhibitors by carrying out structural modifications in the future.

Facile access to 2,2-disubstituted indolin-3-ones via a cascade Fischer indolization/Claisen rearrangement reaction.

An efficient approach for the synthesis of 2,2-disubstituted indolin-3-ones is described. From readily accessible aryl hydrazines and allyloxyketones, 2,2-disubstituted indolin-3-ones could be obtained in good to excellent yields under mild reaction conditions via a cascade Fischer indolization/Claisen rearrangement process. This protocol provides a facile entry to 2,2-disubstituted indolin-3-ones, which have been applied in the construction of the benzofuroindoline framework related to Phalarine.

Ru(II)-Catalyzed Direct C(sp(2))-H Activation/Selenylation of Arenes with Selenyl Chlorides.

A new ruthenium catalytic system was developed for the construction of a C(sp(2))-Se bond with the assistance of directing groups. This protocol features mild reaction conditions, wider substrate scope, and convenient late-stage selenylation of bioactive molecules.

Scaffold Hopping Approach to a New Series of Pyridine Derivatives as Potent Inhibitors of CDK2.

A scaffold hopping approach was exploited to guide the discovery of a series of pyridine derivatives as novel cyclin-dependent kinase (CDK2) inhibitors. These new compounds were designed, synthesized, and evaluated as CDK2 inhibitors. Most of the compounds showed potent inhibition against CDK2, and preliminary structure-activity relationship trends were revealed. A docking study on the most potent compound 6g implied the structural basis for potent CDK2 inhibition. All of the synthesized compounds were also evaluated for their cytotoxicity against the H522 and U87 cancer cell lines. The most potent and drug-like compound 6g was further tested against a normal cell line (L02), demonstrating that this compound is selectively toxic toward cancer cell lines. The results provide the foundation for further improving the potency of this series of compounds.

Enantioselective Bromo-oxycyclization of Silanol.

Relying on the nucleophilicity of silanol for building up silicon-incorporated scaffold with an enantiopure tetrasubstituted carbon center remains elusive. In this report, asymmetric bromo-oxycyclization of olefinic silanol by using chiral anionic phase-transfer catalyst is described. This protocol provided a facile entry to a wide arrangement of enantiopure benzoxasilole in moderate to excellent enantioselectivities depending on the unique reactivity of bromine/N-benzyl-DABCO complex.

An Effective Synthesis Method for Tilorone Dihydrochloride with Obvious IFN-α Inducing Activity.

Tilorone dihydrochloride (1) has great potential for inducing interferon against pathogenic infection. In this paper, we describe a convenient preparation method for 2,7-dihydroxyfluoren-9-one (2), which is a usual pharmaceutical intermediate for preparing tilorone dihydrochloride (1). In the novel method, methyl esterification of 4,4'-dihydroxy-[1,1'-biphenyl]-2-carboxylic acid (4) was carried out under milder conditions with higher yield and played an important role in the preparation of compound 2. The structures of the relative intermediates and target compound were characterized by melting point, IR, MS, and ¹H-NMR. Furthermore, the synthesized tilorone dihydrochloride exhibited an obvious effect on induction of interferon-α (IFN-α) in mice within 12 h, and the peak level was observed until 24 h. This fruitful work has resulted in tilorone dihydrochloride becoming available in large-scale and wide application in clinics, which has a good pharmaceutical development prospects.

Novel tetracyclic benzo[b]carbazolones as highly potent and orally bioavailable ALK inhibitors: design, synthesis, and structure-activity relationship study.

Four series of tetracyclic benzo[b]carbazolone compounds possessing more rotatable bonds and higher molecular flexibility were designed by either inserting a linker within the C8-side chain or by opening the middle ketone ring on the basis of compound 5 (Alectinib, CH5424802). Compound 15b was identified showing nearly identical high potency against both wild-type and the gatekeeper mutant ALK kinase (3.4 vs 3.9 nM). This compound has favorable PK profile with an oral bioavailability of 67.1% in rats. Moreover, compound 15b showed significant growth inhibition against ALK driven cancer cells and KARPAS-299 xenograft model.

N-(1-Oxy-2-picolyl)oxalamic acids as a new type of O,O-ligands for the Cu-catalyzed N-arylation of azoles with aryl halides in water or organic solvent.

N-(1-Oxy-pyridin-2-ylmethyl)oxalamic acids (L3-L5) were identified as novel efficient ligands for copper-catalyzed C-N cross-coupling of azoles and aryl halides in water. The N-arylation of imidazoles, indoles and pyrazoles proceeded with moderate to excellent yields and complete selectivity over aromatic amines and phenols. Moreover, L5, which is also efficient in organic solvent with low catalyst loading, can be used to promote the N-arylation reactions with water-sensitive materials. The catalytic mechanism was proposed based on the results of several verification experiments which indicated that the ligands as new-type chelators may coordinate to Cu(I) with two oxygen atoms of N-oxide and amide in the coupling process.

Discovery and SAR study of c-Met kinase inhibitors bearing an 3-amino-benzo[d]isoxazole or 3-aminoindazole scaffold.

A series of 3-amino-benzo[d]isoxazole-/3-aminoindazole-based compounds were designed, synthesized and pharmacologically evaluated as tyrosine kinase c-Met inhibitors. The SAR study was conducted leading to identification of nine compounds (8d, 8e, 12, 28a-d, 28h and 28i) with IC50s less than 10nM against c-Met. Compound 28a stood out as the most potent c-Met inhibitor displaying potent inhibitory effects both at enzymatic (IC50=1.8 nM) and cellular (IC50=0.18 µM on EBC-1 cells) levels. In addition, 28a had a relatively good selectivity compared to a panel of our in-house 14 RTKs.