In Silico Pharmacokinetic Profiling of the Identified Bioactive Metabolites of Pergularia tomentosa L. Latex Extract and In Vitro Cytotoxic Activity via the Induction of Caspase-Dependent Apoptosis with S-Phase Arrest

Abstract

The in vitro cytotoxic efficacy of plant latex from Pergularia tomentosa L. was studied using five human cancer cell lines: HeLa cells (cervical carcinoma cells), A-549 (lung carcinoma), Panc-1 (pancreatic carcinoma cells), MDA-MB-231 (metastatic mammary adenocarcinoma), and MRC-5 (lung fibroblast cell line) cells. The phytonutrient content of plant latex was identified using the liquid chromatography/mass spectra-quadrupole time of flight (LC/MS-QTOF) technique. In silico studies of polyphenols were carried out to clarify the potential mode of action of the plant latex's constituents. The treatment of different tumor cell lines with different concentrations of plant latex revealed a potent efficacy on the human lung carcinoma cell line (A-549) (IC₅₀ = 3.89 µg/mL) compared with that with vinblastine as a positive control (IC₅₀ = 7.12 µg/mL). The effect of the potent concentration of plant latex on the A-549 cell line induced cell arrest, upregulated the expression of pre-apoptotic markers, and downregulated the expression of antiapoptotic markers. Seven identified polyphenols were selected for the in silico study. A docking assessment using the epidermal growth factor receptor kinase (EGFRk) and eltronib as a positive control showed a higher affinity for the enzyme receptor of the selected polyphenols, except for methyl orsellinate and ginkgotoxin. The ADMET assessment demonstrated the inhibitory effect of the polyphenols on CYP450, except for ouabagenin and xanthyletine. The selected polyphenols obey Lipinski's drug-likeness with no significant toxicity effect. In conclusion, the plant latex of P. tomentosa L. showed cytotoxic activity on the A-549 cell line, and the selected polyphenols showed a promising prodrug agent with a low profile of toxicity in the study.

Keywords: Pergularia tomentosa L. latex; apoptosis; cytotoxicity; docking; pharmacokinetic.

Figure 1

Demonstrated dose response profile of the plant latex from P. tomentosa L. on the cell viability of A-495, Panc-1, MDA-MB-231, MRC-5, and HeLa cell lines following incubation for 24 h. All data are represented as the mean ± SD, n = 3.

Figure 2

Photograph of the flow cytometry analysis shows the effect of 3.8 µg/mL of the plant latex of P. tomentosa L. compared to the negative control on the phases of the cell cycle of the lung cancer cell line (A-549). (A) negative control and (B) plant latex of P. tomentosa L.

Figure 3

Demonstrated effect of P. tomentosa L. latex (IC₅₀ = 3.8 ug/mL) (A) on the percentage of Annexin V-FITC-positive staining monolayer A-549 cells versus the control (B) using flow cytometry.

Figure 4

The figure showed the protein expression of (A) caspase-3, (B) Bcl-2, (C) Bax, and (D) p53, relative to (E) β-actin-mediated apoptosis in lung cancer cell lines (A-549) following treatments with P. tomentosa L. latex (IC₅₀ = 3.8 µg/mL).

Figure 5

Base peak chromatogram of the identified bioactive metabolites in P. tomentosa L. latex: methyl orsellinate (1), ginkgotoxin (2), furaneol 4-glucoside (3), ouabagenin (4), prenyl arabinosyl-(1->6)-glucoside (5), corchoroside-A (6), and xanthyletine (7).

Figure 6

(A) Protein-ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ouabagenin with the active site of EGFRK. (B) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of corchoroside A. with the active site of EGFRK. (C) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of xanthyletine with the active site of EGFRK. (D) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of prenyl arabinosyl-(1->6)-glucoside with the active site of EGFRK. (E) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of furaneol 4-glucoside with the active site of EGFRK. (F) Proteinx–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of methyl orsellinate with the active site of EGFRK. (G) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ginkgotoxin with the active site of EGFRK.

Figure 6

(A) Protein-ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ouabagenin with the active site of EGFRK. (B) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of corchoroside A. with the active site of EGFRK. (C) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of xanthyletine with the active site of EGFRK. (D) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of prenyl arabinosyl-(1->6)-glucoside with the active site of EGFRK. (E) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of furaneol 4-glucoside with the active site of EGFRK. (F) Proteinx–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of methyl orsellinate with the active site of EGFRK. (G) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ginkgotoxin with the active site of EGFRK.

Figure 6

(A) Protein-ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ouabagenin with the active site of EGFRK. (B) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of corchoroside A. with the active site of EGFRK. (C) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of xanthyletine with the active site of EGFRK. (D) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of prenyl arabinosyl-(1->6)-glucoside with the active site of EGFRK. (E) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of furaneol 4-glucoside with the active site of EGFRK. (F) Proteinx–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of methyl orsellinate with the active site of EGFRK. (G) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ginkgotoxin with the active site of EGFRK.

Figure 6

(A) Protein-ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ouabagenin with the active site of EGFRK. (B) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of corchoroside A. with the active site of EGFRK. (C) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of xanthyletine with the active site of EGFRK. (D) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of prenyl arabinosyl-(1->6)-glucoside with the active site of EGFRK. (E) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of furaneol 4-glucoside with the active site of EGFRK. (F) Proteinx–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of methyl orsellinate with the active site of EGFRK. (G) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ginkgotoxin with the active site of EGFRK.

Figure 6

(A) Protein-ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ouabagenin with the active site of EGFRK. (B) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of corchoroside A. with the active site of EGFRK. (C) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of xanthyletine with the active site of EGFRK. (D) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of prenyl arabinosyl-(1->6)-glucoside with the active site of EGFRK. (E) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of furaneol 4-glucoside with the active site of EGFRK. (F) Proteinx–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of methyl orsellinate with the active site of EGFRK. (G) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ginkgotoxin with the active site of EGFRK.

Figure 6

(A) Protein-ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ouabagenin with the active site of EGFRK. (B) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of corchoroside A. with the active site of EGFRK. (C) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of xanthyletine with the active site of EGFRK. (D) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of prenyl arabinosyl-(1->6)-glucoside with the active site of EGFRK. (E) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of furaneol 4-glucoside with the active site of EGFRK. (F) Proteinx–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of methyl orsellinate with the active site of EGFRK. (G) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ginkgotoxin with the active site of EGFRK.

Figure 6

(A) Protein-ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ouabagenin with the active site of EGFRK. (B) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of corchoroside A. with the active site of EGFRK. (C) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of xanthyletine with the active site of EGFRK. (D) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of prenyl arabinosyl-(1->6)-glucoside with the active site of EGFRK. (E) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of furaneol 4-glucoside with the active site of EGFRK. (F) Proteinx–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of methyl orsellinate with the active site of EGFRK. (G) Protein–ligand interactions of identified bioactive metabolites of P. tomentosa L. latex with the active site of EGFRK. Diagrams representing 3D and 2D protein–ligand interactions of ginkgotoxin with the active site of EGFRK.

Figure 7

A boiled egg diagram (A) and bioavailability radar (B) for compounds 1: ouabagenin; 2: corchoroside A; 3: xanthyletine; 4: prenyl arabinosyl-(1->6)-glucoside; 5: furaneol 4-glucoside; 6: methyl orsellinate; and 7: ginkgotoxin. A boiled egg diagram represents the passive gastrointestinal absorption (HIA) and brain penetration (BBB) in the function of the position of the molecules in the WLOGP-versus-TPSA referential. The white region is for a high probability of passive absorption by the gastrointestinal tract, and the yellow region (yolk) is for a high probability of brain penetration. Yolk and white areas are not mutually exclusive. In addition, the points are colored in blue if they are predicted as actively effluxed by P-gp (PGP+) and in red if they are predicted as a non-substrate of P-gp (PGP−). The bioavailability radar represents the drug-likeness of a molecule, with the pink area representing the optimal range for each property (lipophilicity: XLOGP3 between −0.7 and +5.0; size: MW between 150 and 500 g/mol; polarity: TPSA between 20 and 130 Å2; solubility: log S not higher than 6; saturation: fraction of carbons in the sp3 hybridization that is no less than 0.25; and flexibility: no more than nine rotatable bonds for compounds).

Figure 7

A boiled egg diagram (A) and bioavailability radar (B) for compounds 1: ouabagenin; 2: corchoroside A; 3: xanthyletine; 4: prenyl arabinosyl-(1->6)-glucoside; 5: furaneol 4-glucoside; 6: methyl orsellinate; and 7: ginkgotoxin. A boiled egg diagram represents the passive gastrointestinal absorption (HIA) and brain penetration (BBB) in the function of the position of the molecules in the WLOGP-versus-TPSA referential. The white region is for a high probability of passive absorption by the gastrointestinal tract, and the yellow region (yolk) is for a high probability of brain penetration. Yolk and white areas are not mutually exclusive. In addition, the points are colored in blue if they are predicted as actively effluxed by P-gp (PGP+) and in red if they are predicted as a non-substrate of P-gp (PGP−). The bioavailability radar represents the drug-likeness of a molecule, with the pink area representing the optimal range for each property (lipophilicity: XLOGP3 between −0.7 and +5.0; size: MW between 150 and 500 g/mol; polarity: TPSA between 20 and 130 Å2; solubility: log S not higher than 6; saturation: fraction of carbons in the sp3 hybridization that is no less than 0.25; and flexibility: no more than nine rotatable bonds for compounds).