PCOS is primarily an endocrinological disorder affecting multiple aspect of women’s overall health, responsible for anovulatory related infertility. Therapeutic drugs like, metformin, spironolactone and cyproterone acetate are used to treat the PCOS but it shows side effects. Therefore, Himalayan medicinal plant ZA, phytochemicals were used to treat the fertility and reproductive health of the affected women. In this study, Molecular docking studies were carried out between receptor proteins (CYP-17, 5α-reductase and human androgen receptors) and its inhibitors (Zanthoxylum armatum, DC compounds). Before testing the phytochemical compounds of ZA and its inhibitors, a commercial drug (metformin, spironolactone and cyproterone acetate) were docked against docking proteins for comparative study. In the present study, 56 phytochemical compounds of ZA were taken for docking studies, out of which 16 compounds of ZA has shown the good binding energy. After molecular docking, these docking protein and ligands display the result with a particular docking score. Simply, least binding energy is regarded as the best mode of binding as it is stable for the ligand. i.e., lower binding energy means higher docking scores. Figure 4 is showing the docking score of phytochemical compounds of ZA and its inhibitors (CYP-17, 5α-reductase and human androgen receptor). After successfully docking of these compounds into target receptor, the final visualization of the docked structure was performed using the Biovia Discovery studio visualizer 2021. The experimental ligands of ZA were taken in this study, whose binding energy is better than the reference molecule. So, these potential lead compounds could be effective and balance the level of ovarian hormones in PCOS women.
Interaction between phytochemical compounds of Zanthoxylum armatum, DC, with CYP-17 receptor
Before the screening, we used the reference drugs for molecular docking. The target protein (CYP-17) was docked with phytochemical compounds of ZA and reference drugs (metformin, spironolactone and cyproterone acetate). The binding energy of metformin with the CYP-17 protein was − 5.2kcal/mol, -9.4kcal/mol for spironolactone and − 9.8kcal/mol for cyproterone acetate. Now virtual screening resulted in the top phytochemicals from the target showing significantly lower binding energy. The binding energy of 16phytochemical compounds (hesperidin, asarinin, isovitexin, tambuletin, chelerythrine, lupeol, vitexin, xanthyletin, kaempferol, kobusin, β-sitosterol, fargesin, eudesmin, epieudesmin, catechin and planinin) were showing lower binding energy than metformin. 4phytochemical compounds (hesperidin, asarinin, isovitexin and lupeol) were showing lower binding energy than spironolactone and 3 phytochemical compounds (hesperidin, asarinin and lupeol) were showing lower binding energy than cyproterone acetate. The more negative value of the docking score represents better docking affinity as compared to a more positive value and the best docking result of the ligand with the CYP-17 receptors were − 10.8kcal/mol for lupeol, -10.4kcal/mol for hesperidin, -9.9kcal/mol for asarinin, -9.7kcal/mol for isovitexin, -9.4kcal/mol for vitexin, -9kcal/mol for planinin and tambuletin, -8.9kcal/mol for chelerythrine, -8.8kcal/mol for kobusin and fargesin, -8.3kcal/mol for eudesmin, -8.2kcal/mol for β-sitosterol, -8.1kcal/mol for xanthyletin, -8kcal/mol for kaempferol and epieudesmin, -7.9kcal/mol for catechin. These natural ligands could be good inhibitors of the CYP-17 receptor. Figure 5 is showing the 2D and 3D interaction of cyproterone acetate (reference drug) and lupeol with the CYP-17 receptor. Hesperidin, asarinin, isovitexin, tambuletin, chelerythrine, lupeol, vitexin, kaempferol, kobusin, β-sitosterol, fargesin eudesmin, epieudesmin, catechin, xanthyletin and planinin shows better binding affinities with CYP-17 receptor as compared to reference drugs.
Interaction between phytochemical compounds of Zanthoxylum armatum, DC, with 5α-reductase receptor
The target protein (5α-reductase) was docked with phytochemical compounds of ZA and reference drugs (metformin, spironolactone and cyproterone acetate). The binding energy of metformin with 5α-reductase was − 5kcal/mol, -11.2kcal/mol for spironolactone and − 10.6kcal/mol for cyproterone acetate (CPA). Now virtual screening resulted in the top phytochemicals from the target showing significantly lower binding energy. The binding energy of 16phytochemical compounds (hesperidin, asarinin, isovitexin, tambuletin, chelerythrine, lupeol, vitexin, xanthyletin, kaempferol, kobusin, β-sitosterol, fargesin, eudesmin, epieudesmin, catechin and planinin) were showing lower binding energy than metformin. 2 phytochemical compounds (hesperidin and β-sitosterol) were showing lower binding energy than spironolactone and 4 phytochemical compounds (hesperidin, asarinin, β-sitosterol and lupeol) were showing lower binding energy than cyproterone acetate. The best docking result of ligand with 5α-reductase were − 12.2kcal/mol for hesperidin, -11.4kcal/mol for β-sitosterol, -11.1kcal/mol for lupeol, -11kcal/mol for asarinin, -10.5kcal/mol for isovitexin, -10.2kcal/mol for kobusin, planinin and chelerythrine, -10kcal/mol for fargesin and tambuletin, -9.6kcal/mol for eudesmin, − 9.3kcal/mol for epieudesmin, -9.2kcal/mol for catechin and kaempferol, -8.8kcal/mol for xanthyletin, -8.7kcal/mol for vitexin. These natural ligands could be good inhibitors of 5α-reductase. Figure 6 is showing the 2D and 3D interaction of spironolactone (reference drug) and hesperidin with the 5α-reductase receptor. Hesperidin, asarinin, isovitexin, tambuletin, chelerythrine, lupeol, vitexin, kaempferol, kobusin, β-sitosterol, fargesin, eudesmin, epieudesmin, catechin, xanthyletin and planinin shows better binding affinities with the target protein as compare to reference drugs and hence act as potent inhibitors of 5α-reductase enzyme
Interaction between phytochemical compounds of Zanthoxylum armatum, DC, and human androgen receptor
The target protein (human androgen receptor) was docked with phytochemical compounds of ZA and reference drugs (metformin, spironolactone and cyproterone acetate). The binding energies withhuman androgen receptor were − 4.8kcal/mol with metformin, -6.2kcal/mol with spironolactone and − 6kcal/mol with cyproterone acetate (CPA). Now virtual screening resulted in the top phytochemicals from target showing significantly lower binding energy.The binding energy of 16phytochemical compounds (hesperidin, asarinin, isovitexin, tambuletin, chelerythrine, lupeol, vitexin, xanthyletin, kaempferol, kobusin, β-sitosterol, fargesin, eudesmin,epieudesmin, catechin and planinin) were showing lower binding energy than metformin. 15phytochemical compounds (hesperidin, asarinin, isovitexin, tambuletin, chelerythrine, vitexin, xanthyletin, kaempferol, kobusin, β-sitosterol, fargesin, eudesmin, epieudesmin, catechin and planinin) were showing lower binding energy than spironolactone and 16phytochemical compounds (hesperidin, asarinin, isovitexin, eudesmin, epieudesmin, tambuletin, chelerythrine, lupeol, vitexin, xanthyletin, kaempferol, kobusin, β-sitosterol, fargesin, Catechin and Planinin) were showing lower binding energy than cyproterone acetate. The best docking result of ligand with human androgen receptorwere − 9.8kcal/mol for asarinin, -9kcal/mol for xanthyletin, -8.8kcal/mol for β-sitosterol, -8.7kcal/mol for kobusin, -8.3kcal/mol for chelerythrine, -8.2kcal/mol for isovitexin and fargesin, -8.1kcal/mol for catechin, -7.9kcal/mol for kaempferol, -7.6kcal/mol for planinin, -7.4kcal/mol for hesperidin, -7.1kcal/mol for vitexin,-6.9kcal/mol for tambuletin, -6.8kcal/mol for eudesmin and epieudesmin, -6kcal/mol forlupeol. These natural ligands could be a good inhibitor of human androgen receptors. Figure 7 is showing the 2D and 3D interaction of spironolactone (reference drug) and asarinin with the Human androgen receptor.Hesperidin, asarinin, isovitexin, tambuletin, chelerythrine, lupeol, vitexin, xanthyletin, kaempferol, kobusin, β-sitosterol, fargesin, eudesmin, epieudesmin, catechin and planinin shows better binding affinities with the target protein as compare to reference drugs and hence act as potent inhibitors of human Androgen receptor. Figure 8A proposed model showing the inhibitory action of top 3 phytochemical compounds (asarinin, hesperidin and lupeol) of ZA with overexpressed receptor involved in steroidogenesis.
toxicity study result
In-silico toxicity was done to confirm Lipinski’s rule of five (Ro5). This rule emphasises about 4 points to predict the potential toxicity of any drug molecule. These are-
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A molecular mass of less than 500 Daltons.
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No more than 5 H-bond donors.
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No more than 10 H-bond acceptors.
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M Log P not greater than 4.5.
Molecular weight, number of H-bond donors and acceptors are inversely proportional to drug permeability, and lipophilicity LogP is directly proportional to drug permeability [26], [27].The Swiss ADME software results have shown that 10phytochemicals of ZA, namely asarinin, chelerythrine, xanthyletin, kaempferol, kobusin, fargesin, eudesmin, epieudesmin, catechin and planinin may follow 4 rules out of 4. Lupeol, β-sitosterol, may follow 3 rules out of 4. Isovitexin and vitexin may follow 2 rules out of 4. Hesperidin and tambuletin may follow 1 rule out of 4.If a compound does follow two or more violations of Lipinski’s rule of five (Ro5), then the probability of a compound being an oral drug is reduced [28]. Thus, phytochemicals of ZA may be studied further for in-vitro screening for a potential anti-retroviral drug. The results of Swiss ADME are shown in the following (Table 1).
Table 1
Showing the Lipinski’s rule of five for ADME analysis of inhibitors (ligands) using Swiss ADME online server
Lipinski’s Rule of five (Ro5) |
S.No. | Name | Molecular weight (g/mol) | Lipophilicity (MLogP) | H-bond donors | H-bond acceptors | No. Of rule violations | Drug-Likeness Lipinski’s |
Less than 500 dalton | Less than 4.5 | Less than 5 | Less than 10 | Less than 2 violations | Rule Follows |
1. | Hesperidin | 610.56 | -3.04 | 8 | 15 | 3 | No |
2. | Asarinin | 354.35 | 1.98 | 0 | 6 | 0 | Yes |
3. | Isovitexin | 432.38 | -2.02 | 7 | 10 | 2 | No |
4. | Tambuletin | 508.43 | -2.64 | 7 | 13 | 3 | No |
5. | Chelerythrine | 348.37 | 2.53 | 0 | 4 | 0 | Yes |
6. | Lupeol | 426.72 | 6.92 | 1 | 1 | 1 | Yes |
7. | Vitexin | 432.38 | -2.02 | 7 | 10 | 2 | No |
8. | Xanthyletin | 228.24 | 2.37 | 0 | 3 | 0 | Yes |
9. | Kaempferol | 286.24 | -0.03 | 4 | 6 | 0 | Yes |
10. | Kobusin | 370.40 | 1.79 | 0 | 6 | 0 | Yes |
11. | β-sitosterol | 414.71 | 6.73 | 1 | 1 | 1 | Yes |
12. | Fargesin | 370.40 | 1.79 | 0 | 6 | 0 | Yes |
13. | Eudesmin | 386.44 | 1.61 | 0 | 6 | 0 | Yes |
14. | Epieudesmin | 386.44 | 1.61 | 0 | 6 | 0 | Yes |
15. | Catechin | 290.27 | 0.24 | 5 | 6 | 0 | Yes |
16. | Planinin | 370.40 | 1.79 | 0 | 6 | 0 | Yes |
17. | Metformin | 129.16 | -0.56 | 3 | 2 | 0 | Yes |
18. | Spironolactone | 416.57 | 3.58 | 0 | 4 | 0 | Yes |
19. | Cyproterone acetate | 416.94 | 3.71 | 0 | 4 | 0 | Yes |