Molecular docking of fisetin as a multi-target drug in the treatment of multiple sclerosis

Encephalomyelitis disseminate also known as Multiple Sclerosis is an autoimmune disorder caused due to demyelination, inflammation, and axonal loss in the central nervous system(Leray et al. , 2016). The French neurologist Jean-Martin Charcot was the first scientist to describe MS by the appearance of developed glial scars or lesions on the white matter of the brain and spinal cord. As the disease occurs from the age of 20 to 60 years the symptoms differ from one and another, but at the peak, it results in physical and mental instability, blurred or double vision, loss of coordination, muscle weakness, etc. A close up to one million estimated by The National Sclerosis Society is being diagnosed with MS although the women with MS outnumber men. The diagnosis is been done by tools like MRI imaging, evoked potential test, and spine fluid analysis. Contrasting to the present study, the National Multiple Sclerosis Society and Multiple Sclerosis International Federation have introduced 4 types of MS such as CIS (Clinically isolated syndrome), RRMS (Relapsing-remitting MS), PPMS (Primary Progressive MS), SPMS (Secondary progressive MS). Though there is a lack of evidence to cure Multiple sclerosis, technology, and Research scientists still pertain us with hope to find alternative treatments like rehabilitation and physical therapy which intend to slow the progression of Multiple sclerosis.

One of the ways to understand and overcome MS is by inhibiting the enzyme that is potentially causing damage, especially to the myelin sheath, limb function.

Many studies have shown the enzyme belonging to the Protease family play a critical role in the development of MS pathologies(Scarisbrick, 2008).

In the current study, we have focused on three proteins that have been shown to have an elevated level that causes various effects ultimately that leads to the development or progression of MS.

Caspase-1 a protease enzyme that is responsible for the inflammatory response of IL-1 Beta, many studies show blocking caspase-1 potentially decreases the injury in various tested models(McKenzie et al. , 2018; Ming et al. , 2002).

Oligodendrocytes help in the production of myelin sheath in the CNS. Oligodendrocytes cell death caused by cytokines results in MS has been observed through in-vitro studies.

From these cytokines, tumor necrosis factor (TNF) mediates the damage of Oligodendrocytes in MS which is mediated by the Caspase-1 pathway (Hisahara et al. , 2000).

Calpain-1 is another major enzyme found to be responsible for the breakdown of all main myelin proteins, the elevated activity & expression of this proteinase may play a significant role in myelinolysis (Shields et al. , 1999).

Relating to modern studies, Fisetin a natural flavonoid having anti-inflammatory, anti-oxidative and neuroprotective properties could play a huge role in treating neurodegenerative diseases like MS (Ahmad et al. , 2019).

The objective of the present study is to focus on docking the therapeutic targets proteins of Multiple Sclerosis with the plant flavone fisetin which further computationally analyzed.

MATERIALS AND METHODS

Ligand Generation:

Ligand fisetin having Molecular formula C15H10O6 and a molecular weight of 286.2. The log P value for fisetin is 2.282. The ligand structure was extracted from 1/) and downloaded in SDF format which needs to be converted into PDB format for docking; this was done using Open Babel Software. SDF file was converted into PDB format.

Preparation of therapeutic target proteins:

The selected docking macromolecules (target enzymes) for Multiple sclerosis are Caspase 1, Calpain-1, and Cathepsin B. Protein Data Bank is a global repository for 3D structural data of proteins from which, structures of all our three target proteins were extracted.

The 3D crystal structures were downloaded from (ww.rscb.org) in a Protein Data bank format (PDB) having the following ids Caspase 1 (PDB Id: 1lBC), Calpain-1(PDB Id: 2ARY), and Cathepsin B (PDB Id: 1GMY) respectively. The extracted protein structures had ligand interaction which all were extracted out and a clean structure of the protein was put through energy minimization using a Swiss PDB viewer.

RESULTS AND DISCUSSION

Continuous and rapid advances of computational techniques in science have led to various discoveries. It is entirely based on the prediction of binding affinities of the target molecule with crystallographic X-ray structures as well as potential drugs used for docking analysis. The surface grid-based docking method was used to analyze the different linkages in the active site of the therapeutic target proteins Caspase 1(PDB Id: 1lBC), Calpain-1(PDB Id: 2ARY), and Cathepsin B (PDB Id: 1GMY) respectively that were subjugated by the fisetin.

The following results have been discussed in Tables 1 and 2.

The five best minimum binding energy of the ligand (fisetin) with the 3 therapeutic target proteins are mentioned in the following table.

The lowest binding energy conformation of the ligand Fisetin with the three therapeutic target proteins

Calpain-1 (PDB Id: 2ARY), Caspase 1 (PDB Id: 1lBC), and Cathepsin B (PDB Id: 1GMY) were shown in the following figures 1-3. Visualization of these structures was done using the chimera 11- program that is used for analyzing 3D protein structures.

Cathepsin B and Calpain-1, showed the best interaction with Fisetin having the lowest binding energy with No. of hydrogen bonds: 20 and 18 respectively.

Molecular Properties of fisetin was analyzed using molsoft

Lipinski's benchmark rule of five (RO5)5 defines desirable drug candidate physicochemical property space as MW < 500 Da, log P < 5, HBD < 5, and HBA < 10.

The Biological Activity and Toxicity of the ligand fisetin were analyzed by Way2Drug Bio tool.

The biological activity and toxicity of Fisetin were analyzed using the way2drug bio tool, where pa [pharmacologically active] relates to the probability of the activity (pa >0.7).

Table 1: Minimum Binding Energy of fisetin with the 3 drug targets of the top hits

1GMY - Cathepsin B
S.no	Run	Minimum binding energy (kcal/mol)
1	4	-10.01
2	9	-9.35
3	8	-9.3
4	3	-9.32
5	10	-9.29

2ARY : Calpain-1
S.NO	Run	Minimum binding energy (kcal/mol)
1	5	-9.95
2	2	-8.92
3	1	-8.91
4	4	-8.91
5	3	-8.89

lBC - Caspase 1
S. No	Run	Minimum binding energy (kcal/mol)
1	10	-8.18
2	2	-8.11
3	4	-7.99
4	1	-7.99
5	8	-7.96

Table 2: Lowest binding energy of fisetin with the 3 drug targets of the top hits

S.No.	Protein	Final Intermolecular Energy (vdW + Hbond + desolv Energy+Electrostatic Energy) (kcal/mol)	Final      Total Internal Energy (kcal/mol)	Torsional Free Energy (kcal/mol)	Unbound System's Energy (kcal/mol)
1	Calpain-1	-10.23	-0.52	0.3	-0.5
2	Cathepsin B	-10.01	-0.56	0.3	-0.31
3	Caspase 1	-8.54	-0.46	0.3	-0.52

Table 3: Analysis score

Analysis of the score for Fistein

Log P: 1.35

HBD: 4

HBA: 6

Table 4: Biological activity

Sr.NO	Pa	Biological activity
1	0.966	Membrane integrity agonist
2	0.959	Aryl-alcohol dehydrogenase (NADP+) inhibitor
3	0.950	Kinase inhibitor
4	0.865	2 Enoate reductase inhibitor
5	0.844	UGT 1A substrate
6	0.808	CYP2B5
7	0.725	vasoprotector

Table 5: Toxicity

Sr.no	Pa	Toxicity
1	0.755	Vascular toxic
2	0.713	Aphthous ulceration

Figure 1: Calpain1. The oxygen atom in the image is signified as red in color in the ligand structure fisetin. The blue lines signify the hydrogen bonds between the ligand fisetin and the therapeutic target protein (calpain1). No. of hydrogen bond = 18

Figure 2: Capase 1. The oxygen atom in the image is signified as red in color in the ligand structure fisetin. The blue lines signify the hydrogen bonds between the ligand fisetin and the therapeutic target protein (capase1). No. of hydrogen bond = 23.

Figure 3: Cathepsin B. The oxygen atom in the image is signified as red in color in the ligand structure fisetin. The blue lines signify the hydrogen bonds between the ligand fisetin and the therapeutic target protein (cathepsin B). No. of hydrogen bond = 20.

Molecular Properties and Drug-likeness.

Molecular formula: CIS H10 06

Molecular weight: 286.05

Number of HE A: 6

Number of HBD: 4

MolLogP: 1.35

MolLogS : -2.41 (in Loglinolesll) 1121.11 (in mg 11

MolPSA: 86.06A*

MolVol: 271.53 A3

pKe of most Basic/Acidir group : 0. 6 86

[HA]

pK_M = - log,₀ K_a = log_la

[A ]|H + ]

Number of stereo centers: 0

Drug-likeness model score: 0 46

4.00

6.00

____J2 -2.00    0.W

Figure 4: Molecular properties & drug-likeness.

CONCLUSION

In the present study, the effectiveness of fisetin showing the characteristic binding ability to the therapeutic target proteins and their effective progressiveness against the disease multiple sclerosis was assessed.

To the three therapeutic target proteins caspase 1 (PDBId:1IBC), Calpain-1 (PDB id:2ARY), and cathepsin B(PDB Id: 1GMY) fisetin, exhibited satisfactory docking output to all of them.

The primary cause for the disease MS as known is due to the formation of lesions in CNS and destruction in the myelin sheath that leads to multiple pathological defects, from tremors to complete loss of coordination. As fisetin can interact with the target proteins of Multiple sclerosis, this can be a potent candidate for treating and reducing the symptoms of Multiple Sclerosis.

ACKNOWLEDGMENTS

The authors would like to thank the support rendered by Kristu Jayanti College, Autonomous.

CONFLICTS OF INTEREST

The authors declare that they have no potential conflicts of interest.