Synthesis of salicylic acid phenylethyl ester (SAPE) and its implication in immunomodulatory and anticancer roles
The reaction for the production of SAPE is illustrated in Fig. The ADME- toxicity properties are directly related to the biological effect of drugs and their fate in an organism, and in silico methods can predict these properties at an early stage in drug design20. Initially, the prediction of the potential ligand binding site (active site) on COX-2 displayed the cavity volume to be 56. [i] (A) Protein–ligand interactions between SAPE and active site residues of COX-2 enzyme (B) Binding mode of SAPE at the active site residues of COX-2 enzyme (C) Protein–ligand interactions between ibuprofen and active site residues of COX-2 enzyme (D) Binding mode of ibuprofen at the active site residues of COX-2 enzyme (E) Protein–ligand interactions between indomethacin and active site residues of COX-2 enzyme (F) Binding mode of indomethacin at the active site residues of COX-2 enzyme. The molecular docking scores (Table 1) revealed that SAPE could be docked at the active site of the COX-2 protein with the favourable docking scores; which is an indication that the enzyme usually favours SAPE compared to ibuprofen and indomethacin. In order to examine the molecular interaction, the ligand–protein interaction analysis for the docking hits SAPE, ibuprofen and indomethacin using the MVD Ligand Energy Inspector were assessed, and Table 2 shows the details of molecular interactions of the docking hits. The MD simulation was conducted only for the docking hit (SAPE-COX-2, ibuprofen-COX-2 and indomethacin-COX-2 docked complexes). The HOMO and LUMO energies of SAPE are illustrated in Fig. The results for the membrane stability assay in terms of percentage of hemolysis are shown in Fig. (A) Membrane stability assay of SAPE in erythrocytes and (B), (C) and (D) Results of Cell viability assay of SAPE at 48 h on cells of (A) PBMC, (B) Caco2 and (C) HepG2 cells with doses of (25, 50 and 100 µg/ml) or with 0. The results for MTT assay of SAPE on PBMC, human intestinal cell lines derived from colon carcinoma (CaCo-2) and liver cancer cell lines (HepG-2) are shown in Fig.
COX-2 is an inducible isoform of cyclooxygenase that is mainly produced in inflamed tissues and effectively absent in healthy tissue. (A) The inhibition of COX-2 production in inflamed CaCo-2 cells, and (B) the direct relative inhibition of COX-2 by phenyl ethyl alcohol (PEA), salicylic acid (SA), salicylic acid phenyl ethyl ester (SAPE) and indomethacin (IM). From the results it becomes evident that the esterfication of salicylic acid considerably increases the anti-inflammatory activity of salicylic acid and these results corroborates with the previous findings5,13,14. To determine the cytotoxic effect of SAPE on MCF-7 cells, cell viability assay was performed using the MTT (Fig. (a) Results of cell viability assay of SAPE on MCF-7 cells measured by MTT based method. The MCF-7 cell line is an ER-positive and luminal subtype human breast carcinoma29. (A) Cell cycle analysis of SAPE on MCF-7 cells with doses of 25, 50, 100 µg/mL or with 0. The level of ROS in the cells was analyzed in order to address the role played by ROS in SAPE induced apoptosis. The loss in MMP was analyzed, since a loss in Δψm is directly associated with disruption of balance between the expressions of pro- and anti-apoptotic proteins within the Bcl-2 family. Apoptosis or type I programmed cell death is an inbuilt mechanism of the body to destroy cells that represent a threat to the physiological integrity of an organism. (A) Representative pictures of apoptosis assay of SAPE on MCF-7 with different doses (25, and100 µg/mL) or with 0.
Accessing the integrity of DNA is a useful marker for screening potential anticancer agent since the phenomena of apoptosis is expressed by cells after suffering DNA damage31. Autophagy or type II programmed cell death is an evolutionarily conserved catabolic mechanism of cell death involving degradation of damaged proteins or organelles. (A) Fluorescence microscopy for analysis of acidic vesicles and autophagosomes against SAPE treated MCF-7 cells (B) Analysis of the degree of acidity against SAPE (25, 50, 100 µg/mL) treated MCF7 cells for 48 h. The results obtained for the test of acute inflammatory process by paw oedema model are presented in Table 3. Excised intestines of the different groups of rats after treatment and euthanization, and their SEM images are shown in Fig. Excised intestines and SEM images of the intestinal section from (A) RG-CF (colitis free control group without any treatment) rats, (B) RG-CI (colitis induced group without any treatment) rats, (C) RG-SP (colitis induced group treated with SAPE) rats, RG-IM (colitis induced group treated with indomethacin) rats. Inflammation induced oxidative damage is aggravated by the decrease in antioxidant enzymes activities such as catalase (CAT), glutathione S-transferase (GST) and gamma-glutamyltransferase (GGT) which act as free radical scavengers in conditions associated with oxidative stress36. (a) Malondialdehyde (MDA) content; (b) catalase (CAT) content; (c) γ-glutamyl transferase (GGT) activity; (d) glutathione S-transferase (GST) activity; (e) tumor necrosis factor alpha (TNF-α) content and (f) interleukin 6 (IL-6) content in the intestinal tissue extract for different treatment groups of rats (RG-CI: colitis induced group without any treatment; RG-CF: colitis free control group without any treatment; RG-SP: colitis induced group treated with SAPE; Group RG-IM: colitis induced group treated with indomethacin). GGT contributes to the extracellular catabolism of glutathione (GSH), as it catalyzes the transfer of the gamma-glutamyl moiety from conjugated GSH to acceptors such as amino acids and dipeptides. The cytokines are stimulators of the production of acute phase proteins that are produced during inflammatory processes. IL-6 is a pleiotropic cytokine produced at the site of inflammation and plays a key role in the acute phase response, regulation of immune responses and hematopoiesis49.
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