INTRODUCTION: Integrating the group of the 10 main causes of death worldwide [1], cancer has stood out among the main obstacles to human health, recording 19.3 million cases and 10 million deaths in 2020 [2]. This scenario is worsened when one considers that chemotherapy drugs used in the treatment of cancer present problems of toxicity, solubility and stability, exposing the need to discover new therapeutic agents of this class [3]. To this end, organic synthesis and rational drug design have contributed to the discovery of new anticancer pharmaceutical candidates, which draw attention to promising chemical groups, such as enopyranosides: synthetic derivatives of biocompatible carbohydrates (such as glucose and galactose, for example) with interesting anticancer properties reported in scientific literature [4,5]. AIMS: Thus, the present research aimed to use computational tools for the rational design of an enopyranosidic anticancer candidate derived from D-glucose, cooperating in the prospecting of new drug candidates. METHODS: To achieve this, the first step was to determine a synthetically accessible enopyranosid structure derived from D-glucose, which led to the choice of Prop-2-yn-1-yl 2,3-dideoxy-2-enopyranoside-4-ulose. This compound was subjected to computational strategies to evaluate its drugability, using PASSonline to determine its probability of presenting antineoplastic activity,[6] SwissADME to evaluate its pharmacokinetic suitability (determination of the Lipinski and Veber rule, water solubility, cLogP and gastrointestinal absorption)[7]. Molinspiration to evaluate its possible pharmacodynamic targets (GPCR ligand, ion channel modulator, kinase inhibitor, nuclear receptor ligand, protease inhibitor and enzyme inhibitor) [8], and the Osiris property explorer to calculate druglikeness (similarity to commercial drugs) and drug-score (a value that expresses the probability of becoming a future drug) [9]. Having confirmed the in silico antineoplastic potential, enopyranoside was then synthesized starting from the D-glucose derivative 3,4,6-tri-O-acetyl-D-glucal, through its glycosylation with propargyl alcohol, followed by hydrolysis and oxidation to the formation of Prop-2-yn-1-yl 2,3-dideoxy-2-enopyranoside-4-ulose [5]. Then, the compound was characterized by 1H and 13C Nuclear Magnetic Resonance (NMR). After being synthesized and characterized, enopyranoside was subjected to MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) test to evaluate its cytotoxicity against the cancer cell line K562 (erythroleukemic cells), which was cultivated at 37 °C under atmosphere with 5% CO2 using RPMI culture medium supplemented with 10% fetal bovine serum (FBS) and 1% of antibiotics. Cytotoxicity assessment was carried out in 96-well plates containing 3x106 cells/mL at concentrations of 0.78 to 25.00 ?g/mL of enopyranoside. The tests were carried out in triplicate with repetition throughout of 72 h, adding 25 ?L MTT (5 mg/mL) at the end of this period for subsequent reading at 560 nm. Doxorubicin was used as a positive control for the tests. RESULTS AND DISCUSSION: Based on the anticancer potential of carbohydrate derivatives, especially enopyranoside derivatives [4,5], Prop-2-yn-1-yl 2,3-dideoxy-2-enopyranoside-4-ulose has been subjected to computer-aided drug design to determine its drugability and potential for in vitro testing. To this, the first step was to evaluate, using the PASSonline platform, the probability of this compound being active (Pa) and/or inactive (Pi) as antineoplastic, noting that this enopyraniside presented an 85.8% chance of being active (Pa), with only a 0.6% chance of being inactive (Pi), giving the first indications of the antineoplastic potential of this compound. Its pharmacokinetic suitability was also observed by SwissADME, demonstrating that the compound attends all requirements of Lipisnki and Veber rules [7], also demonstrating high potential for gastrointestinal absorption, good solubility in water and balanced lipophilicity (cLogP = 0.30). These results demonstrate good pharmacokinetic characteristics of this enopyranoside, which is of utmost importance, since pharmacokinetic problems are among the main causes of failure among drugs that decline in clinical trials [10], Molinspiration analyzes showed that this enopyranoside develops its antineoplastic action especially through enzymatic inhibition (score = 0.34, with scores > 0 being considered active) [8]. Additionally, a good druglikeness (1.52) was observed, since values > 0 are satisfactory, as well as a good drug-score (0.88) was observed, since values closer to 1.00 show greater pharmaceutical potential of the studied compounds. These results demonstrate the similarity of this enopyranoside with commercial drugs as well as the high probability of becoming a good drug in the future.[9] All these results encouraged the synthesis and characterization of Prop-2-yn-1-yl 2,3-dideoxy-2-enopyranoside-4-ulose, which was obtained with a yield of 94% for the glycosylation step, 98% for hydrolysis and 89% for the oxidation reaction, demonstrating a spectroscopic characterization compatible with that reported in the literature, attesting to the success in its synthetic obtainment [5]. The evaluation using the MTT method confirmed the cytotoxic potential observed in the computational stage, showing that enopyranoside was able to inhibit the growth of neoplastic cells at all tested concentrations, reducing the viability of this cancer cell line by 95.1% at the highest concentration (25.0 ?g/mL) and recording an IC50 of 0.997 ?g/mL (with confidence interval - 95% of 0.924 to 1.081 ?g/mL). According to the literature, compounds that demonstrate IC50 values < 5 µg/mL are classified as highly active, demonstrating sufficient anticancer potential to continue in testing for the discovery of new chemotherapy drugs [11]. CONCLUSION: The present results demonstrated the good drugability of Prop-2-in-1-yl 2,3-dideoxy-2-enopyranoside-4-ulose, especially its good physicochemical, pharmacokinetic and pharmacodynamic properties, as well as its high antineoplastic potential in silico. After obtaining it satisfactorily and in line with the literature, in vitro cytotoxicity studies confirmed the antineoplastic potential identified in the computational stage of this work, classifying this compound as highly active. These results encourage future rational design studies involving new derivatives of this class, as well as the continued anticancer exploration of Prop-2-yn-1-yl 2,3-dideoxy-2-enopyranoside-4-ulose, contributing to the discovery of new anticancer drug candidates with adequate pharmacodynamic and pharmacokinetic characteristics. ACKNOWLEDGEMENT: The authors are grateful for the postgraduate scholarship provided by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES). REFERENCES: [1] OMS. Organização Mundial da Saúde. The top 10 causes of death. OMS; 2020. Available in: https://www.who.int/news-room/fact-sheets/detail/the-top-10-causes-of-death. Access at: 25 Jul. 2023. [2] SUNG, H. et al. Global cancer statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA: a cancer journal for clinicians, v. 71, n. 3, p. 209-249, 2021. [3] KAUR, J. et al., Recent advances in developing polymeric micelles for treating cancer: Breakthroughs and bottlenecks in their clinical translation. Drug Discovery Today, v. 27, p. 1495–1512, 2022. [4] SILVA, A. J. F. S. et al. Synthesis and Cytotoxic Activity of New Alkyl 2-Azido-2, 3-Dideoxy-?-D-Lixo-Hexopyranosides From ?, ?-Unsaturated Sugar Enones and Sodium Azide. Revista Virtual de Quimica, v. 11, n. 6, p. 1823-1834, 2019. [5] DANTAS, Claudio R. et al. Stereoselective synthesis and antitumoral activity of Z-enyne pseudoglycosides. Organic & Biomolecular Chemistry, v. 14, n. 28, p. 6786-6794, 2016. [6] DASH, Linu et al. Prediction of anticancer activity of potential anticancer compounds using pass online software. Plant Archives, v. 20, n. 2, p. 2808-2813, 2020. [7] DAINA, A.; MICHIELIN, O.; ZOETE, V. SwissADME: a free web tool to evaluate pharmacokinetics, druglikeness and medicinal chemistry friendliness of small molecules. Scientific reports, v. 7, n. 1, p. 1-13, 2017. [8] Molinspitation. Molinspiration cheminformatics software. Available in: www.molinspiration.com. Access at: 25 Jul. 2023. [9] Organic Chemistry Portal. Osiris Properety Explorer. Available in: https://www.organic-chemistry.org/prog/peo/. Access at: 25 Jul. 2023. [10] ROSENBAUM, S. E. (Ed.). Basic pharmacokinetics and pharmacodynamics: An integrated textbook and computer simulations. John Wiley & Sons, 2016. [11] CHIMPLEE, S. et al. Anti?breast cancer potential of frullanolide from Grangea maderaspatana plant by inducing apoptosis. Oncology Letters, v. 17, n. 6, p. 5283-5291, 2019.
Comissão Organizadora
Francisco Mendonça Junior
Pascal Marchand
Teresinha Gonçalves da Silva
Isabelle Orliac-Garnier
Gerd Bruno da Rocha
Comissão Científica
Ricardo Olimpio de Moura
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