The results of using a computer approach in the search for new potential biologically active compounds in a series of 9,10-anthraquinone derivatives using free online programs PASS Online, CLC-Pred (Cell Line Cytotoxicity Predictor), Acute Rat Toxicity and determining the level of binding of the studied structures of anthraquinones with target proteins using the Schrodinger software package are generalized. The directions of experimental primary assessment of antimicrobial, antiplatelet, antioxidant, antiviral, anticonvulsant, antitumor action for selected objects of research are determined. Molecular docking shows the prospects for studies of the mechanisms of anticancer and antiplatelet agents.
9,10-anthraquinone derivatives, in silico prediction, biological action
Despite the significant achievements of modern medical chemistry and pharmacology, the search
for new more effective and safer medicinal substances remains an actual problem [
The structural formulas of molecules of studied substances using computer prediction allow to find
new biologically active compounds with necessary properties [
Computer investigations are widely used to analyze the relationships "structure - biological effect"
of organic compounds [
There are a relatively small number of computer programs available for free over the Internet and predicting pKa (http://vcclab.org/lab/alogps/start.html), some physicochemical properties some types biological activity
(http://www.organic-chemistry.org/prog/peo/index.html, http://www.molinspiration.com/cgi-bin/properties). In recent years, foreign web services have appeared on the Internet that allow predicting the interaction of chemical compounds with target macromolecules
2020 Copyright for this paper by its authors. based on structural similarity (http://bioinformatics.charite.de/superpred/, http://cpi.bio-x.cn/drar/). The information available on these websites does not allow us to assess the quality of the prediction provided by these web services.
The computer program PASS Online [
Molecular docking (molecular modeling) is actively used to solve virtual screening problems [
Considering the above, an in silico approach was used in the search for new derivatives of 9,10anthraquinone using the latest resources to determine the experimental directions of research on their pharmacological activity.
The computer program PASS Online and a number of free online web services such as CLC-Pred
(Cell Line Cytotoxicity Predictor), Acute Rat Toxicity of the Way2Drug platform [
In silico evaluation of the biological properties of new functionalized 9,10-anthraquinone derivatives
(Fig. 1) was performed by the online services PASS Online, CLC-Pred, Acute Rat Toxicity of web portal
Way2Drug. As an initial information, the structural formula of the substance in MOL or SDF file format
was used to obtain prediction data in each of the mentioned programs. The connection table containing
data of the molecule valent bonds and table of atoms types of the loaded structure of compound are
basis for generation of the set of multilevel neighborhoods of atoms (MNA) structure descriptors [
CLC-Pred service allow to predict the cytotoxic action for non-transformed and cancer cell lines based on loaded molecular structure in comparison with the training set obtained from ChEMBLdb Database (https://www.ebi.ac.uk/chembldb/) [20]. Results of prediction using CLC-Pred are also presented as a list of Pa/Pi values of probable cancer cell lines (Fig. 2). Acute Rat Toxicity online service [21] allow in silico estimate probable level of toxicity using a rat model for four different routes of administration: intravenous, intraperitoneal, oral, subcutaneous (Fig. 3). Results of prediction are compared with the training sets of known compounds with determined experimental toxicity LD50 values obtained from Toxicity Database of SYMYX MDL (http://www.akosgmbh.de/accelrys/databases/symyx_dbs.htm). Output data show probable LD50 value in log10 (mmol/kg) or mg/kg and class of acute rodent toxicity.
The molecular modeling of affinity (binding) with the corresponding target protein site was used to determine probable mechanisms of action using Small-Molecule Drug Discovery Suite of Schrödinger within the test access [22] and AutoDock Vina [23] was carried out in addition to the above-mentioned free online access programs for predicting the pharmacological effects of the studied structures. The conformation of the ligand (studied chemical structure) that best interacts with the protein binding site is the result of molecular modeling. The structure of biological targets for molecular modeling were taken from database RCBS Protein DataBank (https://www.rcsb.org). For the molecular docking as target proteins were selected: receptor proteins-tyrosine kinases cyclooxygenase-1 (COX-1) - 3N8X, glycoprotein-IIb/IIIa (GPIIb /IIIa) - 2VDM, glycoprotein-VI (GP-VI) - 2G17, purine receptor P2Y12 4PXZ, prostacyclin receptor (PG-I2) - 4F8K, protein-activated receptor-1 (PAR-1) - 3VW7, antithrombin III (ATIII), factor-X (FX), factor-II (F-II), factor -IX (F-IX) and vitamin K-epoxy reductase (VKOR) - 1NQ9, 1KSN, 5JZY, 1RFN and 3KP9, c-Kit, B-Raf, EGFR (1NQL, 1IVO, 1M17, 2GS6) and PDGF (1T46, AKT1, ERK2), non-receptor tyrosine kinases SRC (1SKJ), nonspecific tyrosine kinases ABL (3OXZ, 3QRJ, 2ABL). Then, the target protein is prepared using the Protein Preparation Wizard with the removal of water molecules and the addition of missing hydrogen atoms in the structure of the target protein and the optimization of the structure of protein using the PROBKA subprogram. At the next stage, the protein structure is minimized by the OPLS-2005 subprogram. Preparation of ligand (studied chemical structure) is performed using LigPrep Wizard. Next, the ligand binding region to the receptor is generated using the Receptor Grid Generation module of the Glide Maestro and docking is performed in the XP phase. The result of docking is a set of structures in 3D with a set of numerical indicators describing their affinity Gscore (scoring function) to the active site of the protein.
Analysis of prediction data for all new structures of functionalized 9,10-anthraquinone derivatives 1-12 (Fig. 1) showed that the main, expected, direction of experimental studies is the investigation of the antitumor effect (Pa in the range 0.3-0.6). Since the studied structures 1-12 are new objects for the PASS Online training set, the Pa values for many structures are in the range 0.3-0.6. Taking into account results of anticancer activity prediction, cytotoxicity was evaluated in silico against cancer cell lines using the web service CLC-Pred [20]. The analysis of the obtained results showed that the most probable effect of the studied compounds 1-12 (Pa=0.3-0.8) is on cancer cell lines of the lungs, lymphoid tissue, glandular tissue of the breast, and for some compounds is on also tissues of the cervix, brain, and colon.
The introduction of new biophore fragments into the 9,10-anthraquinone molecule allowed to expand the range of experimental in vitro and in vivo studies, in addition to the study of the antitumor effect predicted using PASS Online, in the following areas. In particular, antimicrobial and antioxidant activities were predicted for N-acylamino-9,10-anthraquinones 1, amino acid derivatives of 2-chloroN-(9,10-dioxy-9,10-dihydroanthracen-1-yl) acetamide 2 (Pa=0.3-0.6). 1,2,3-Substituted guanidine 3 would be interesting subjects to test for antianginal (Pa=0.3-0.4), anti-ischemic (Pa=0.3-0.35), cardiotonic (Pa=0.3-0.45) effects. The direction of research on hypoglycemic activity would be interesting for 5-arylidene derivatives 4 (Pa=0.3-0.35). Iminothiazoles 5, 1,2,4-triazoles 6 and tetrazoles 7 can be studied also for antimicrobial (Pa=0.3-0.4), anti-inflammatory (Pa=0.3), anti-allergic effects (Pa=0.3-0.4) and effects on vascular processes (Pa=0.3-0.33). An additional area of testing for pyrrolylantracenediones 8 is to determine the level of antibacterial and antifungal activity (Pa=0.4-0.5). The antitumor activity of the dithiocarbamate derivatives 9, 10 was supplemented by antimicrobial, antioxidant, antiplatelet, antiviral, anticonvulsant effects at Pa=0.3-0.55 according to Pa prediction data. Predicted Pa values (Pa=0.3-0.4) for the dithiocarbamates 9, 10 and hydrazone derivatives 11, 12 showed perspective for their further investigations for antiviral properties.
The analysis of the prediction results showed that, in the overwhelming majority, the Pa values for the studied structures are in the range Pa=0.3-0.6. Taking into account the categorization of Pa values, such low Pa values can be explained by the novelty of molecular structures for the PASS Online training set. The predicted data became the basis for the experimental study of the above mentioned antraquinone derivatives 1-12.
The probable acute toxicity of LD50 using the online resource Acute Rat Toxicity using in silico rat model with four different routes of administration: intraperitoneal, intravenous, oral and subcutaneous [21] for promising compounds, selected by the experimental tests, allows to estimate the level probable toxicity of anthaquinone derivatives 1-12. The results showed that the studied structures of compounds can be classified as medium-, low- and non-toxic compounds.
As outcome result of the molecular docking, the values of Gscore scoring functions were determined and the anthraquinone derivatives with high, medium and low levels of binding to a specific target protein PDGF (1T46), VKOR (3KP9), c-Kit and B-Raf associated with anticancer and antiplatelet actions with Gscore in the range from -6.7 to -11.8 were found among the studied molecular structures 112 [24]. Examples of visualization of binding of hit compounds among triazole 6, dithiocarbamate 9,10 and triazinone 7 anthraquinones in the corresponding active zone of the sites of previously mentioned proteins are given below (Fig. 2-7).
It should be noted that the dithiocarbamate derivatives of anthraquinone of type 9,10, which had the highest values of the scoring function Gscore= -10.92…-11.2 are promising objects for experimental study of the mechanisms their antioxidant, anticancer and antiplatelet action.
Therefore, the results of predicted by the online programs PASS Online, CLC-Pred, Acute Rat Toxicity regarding the probable manifestation of antimicrobial, antiplatelet, antioxidant, antiviral, anticonvulsant, antitumor actions and predicted values of acute toxicity, as well as molecular docking data obtained by Small-Molecule Drug Discovery Suite Schrödinger and AutoDock Vina have been verificated by experimental in vitro and in vivo investigations.
In this article, we present the generalized results of using in silico tools in the search for new potential pharmacologically active compounds in the series of 9,10-anthraquinone derivatives. The used computer approach carried out by free online programs PASS Online, CLC-Pred, Acute Rat Toxicity allowed to determine the directions of the experimental primary assessment of antimicrobial, antiplatelet, antioxidant, antiviral, anticonvulsant, antitumor action for the selected objects of study. Molecular docking data on biotargets of pathological processes allowed to define compounds with a high degree of affinity and showed the prospect of studying the mechanisms of anticancer and antiplatelet agents from this class of compounds by modifying the ligand with various pharmacophore fragments. The results of in silico approach allowed experimentally identify promising compounds in the series of 9,10-anthraquinone.
This work (Project number: 0119U002252) is supported by the Ministry of Education and Science of Ukraine.
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