Research Article

Lysine specific demethylase 1 as therapeutic target of cancer

Nihar Ranjan Panda1, Sudhir Ranjan Bhoi2, Rakesh M. Rawal3, Mukesh Kumar Raval1*

1Department of Chemistry, Gangadhar Meher College, Sambalpur, 768004, Odisha, India

2Department of Biotechnology & Bioinformatics, Sambalpur University, Jyoti Vihar, Burla, 768019, Odisha, India

3Department of Life Sciences, University School of Sciences, Gujarat University, Navrangpura, Ahmedabad.-380009, Gujarat, India

*For correspondence

Prof. Mukesh Kumar Raval,

Department of Chemistry, Gangadhar Meher College, Sambalpur, 768004, Odisha, India. Email:






Received: 19 April 2017

Accepted: 02 May 2017


Objective: Lysin specific demethylase 1 (LSD1) inhibits the tumor suppressor activity of p53 and facilitates the progress of tumor. In order to check the tumor growth, the activity of LSD1 enzyme is to be blunted.

Methods: Phytochemicals from naturally occurring plant-based anti-cancer compound-activity-target (NPACT) database are screened with LSD1 as target applying genetic algorithm (GA) method to study best ligand poses and free energy of binding using Argus Lab. The prediction of drug-likeness and oral toxicity of the ligands are performed by the online tools Molsoft and ProTox respectively.

Results: Calyxin H shows optimum binding affinity to both the substrate and FAD binding sites of LSD1. The LD50 value (median lethal dose) of calyxin H is more than 1000 mg/kg body weight and the toxicity class is 4.

Conclusions: Calyxin H is the inhibitor of choice against target LSD1. The lead molecule may be the future potential herbal drug for cancer treatment.

Keywords: Anti-cancer, LSD1, Calyxin H, Phytochemical, Drug-likeness, Toxicity


Cancer is a generic term for a large group of diseases that can affect any part of the body. Other terms used are malignant tumours and neoplasm. One characteristic feature of cancer is the rapid growth of abnormal cells beyond their usual boundaries. These abnormal cells can then invade adjoining parts of the body and spread to other organs. This process is referred to as metastasis, which is the major cause of death from cancer. Any failure in regulation of cell division leads to abnormal proliferation, resulting in cancer.

Lysine-specific demethylase 1 (LSD1) is associated with metastasis. It is the first identified histone demethylase, which belongs to the family of the flavin adenine dinucleotide (FAD)-dependent amine oxidases. LSD1 specifically demethylates mono- or dimethylated histone H3 lysine4 (H3K4) and H3 lysine 9 (H3K9) through a redox reaction. LSD1 plays a vital role in a wider spectrum of biological processes, including cell proliferation, adipogenesis, spermatogenesis, chromosome segregation and embryonic development.1 Furthermore, LSD1 facilitate the progress of tumor by inhibiting the tumor suppressor activity of p53 in promoting apoptosis.2 Discovery of LSD1 inhibitors has a great potential of developing anti-cancer drug.1 Inhibition of LSD1 may constitute an efficient way to interfere with tumor growth and invasiveness.3 In the present study we consider naturally occurring phytochemicals to develop an anti-cancer therapeutic agent. Phytochemicals improve the efficiency of cytotoxic agents, decrease their resistance, lower or alleviate toxic side effects, reduce the risk of tumour lysis syndrome, and detoxify the body as compare to the potential chemotherapeutic drugs.4 Here we have made a structure based search to identify and improve an inhibitor of LSD1 for further investigation.

Materials and Methods

Structures of target protein- LSD1

The 3D coordinate file of target protein LSD1 (2xag.pdb) is obtained from protein database, (

Database of herbal compounds

Naturally occurring plant-based anti-cancer compound-activity-target database (NPACT) is used ( The database consists of 3D coordinate files of phytochemicals, which exhibit anti cancerous activity.6

Identifying hit compounds

The structures of herbal compounds from the database are docked in to the subtrate and FAD binding sites of the target protein LSD1 applying genetic algorithm (GA). The parameters used are population size =50, number of genes =11, maximum generations =1000, elitism number = 5, crossover rate =0.8, mutation rate =0.2, local search rate =0.06, local search max iterations =20, converged when rmsd population fitness <1 kcal/mol pursuing upto 1000 ligand poses. The docking is done with grid resolution at 0.4 Å. The free energy of binding (ΔG) of the compounds reflect its binding affinity to the target enzyme hence the inhibitory efficiency. ArgusLab 4.0.1 is used for molecular docking (

Batch file preparation of ligand molecule

We are using Molegro Virtual Docker (MVD) as a tollkit for batch file preparation of ligand molecule (SDF Format).The site can be accessed through ( molegrovirtual-docker).

Identification of lead molecule

The list of hit molecules is screened for their drug-likeness and toxicity properties to identify lead molecules. The investigation of absorption, distribution, metabolism, excretion and toxicity (ADME/Tox) properties of a compound, is a crucial step in the drug development process.

Prediction of drug-likeness

Drug-likeness score is computed from the molecular properties, i.e. molecular weight, number of hydrogen bond donors, number of hydrogen bond acceptors, polar surface area, MolLogP, MolLogS, and number of stereo centers by Molsoft through online server (

Prediction of toxicity

ProTox is an online web server for prediction of oral toxicities of small molecules in rodent.8 The medial lethal dose (LD50) and toxicity class is calculated based on their similarities on toxic compound. The toxic fragments generated by ROTBOND and RECAP methods.9 Based on severity the compounds are classified into six different groups (Class) where class 6 is the most non toxic group.

Lead optimization

Lead optimization is carried out through structural alteration of lead compound with the help of Hyperchem software, which is a versatile tool for molecular modelling. It provides a simple way to produce 3D molecular structures on screen, geometry optimization techniques to search for stable structures, and molecular dynamics techniques to carry out conformational searching and to investigate structural changes.10 Hyperchem is commercially available from

The strategy for docking applied is as follows

LSD1 has two binding sites: cofactor (FAD) and substrate binding sites. The inhibitor para-bromo-cis-2-phenylcyclopropyl-1-amine (TCF) binds to subtrate binding sites. The ligand molecules are docked on FAD and TCF binding sites under four situations.

  1. FAD BS (+TCF) - Docking of ligand on FAD binding site when TCF binding site is occupied.
  2. FAD BS (-TCF) - Docking of ligand on FAD binding site when TCF binding site is unoccupied.
  3. TCF BS (+FAD) - Docking of ligand on TCF binding site when FAD binding site is occupied.
  4. TCF BS (-FAD) - Docking of ligand on TCF binding site when FAD binding site is unoccupied.

A list of top five hit molecules based on ligand pose energies is prepared.

Results and Discussion

Short listing of phytochemical inhibitors of LSD1

The best five ligand molecules from the phytochemical database are selected in four situations on the basis of their binding energies.

The binding of calyxin H to the FAD and TCF binding sites in LSD1 under situations a and c are depicted in Figure 1A and 2A. The interactions of inhibitor withe residues of LSD1 are presented in Figure 1B and 2B.

Prediction of drug-likeness of lead compound by Molsoft

Calyxin H has good affinity to both the binding sites of the LSD1 enzyme on different four situations. Hence it is considered as a lead compund. The drug-likeness score of calyxin H is determined by Molsoft online tool (Table 2).

Prediction of toxicity of lead compound by ProTox

Toxicity of the lead molecule predicted using ProTox online tool (Table 3).

Lead optimization

The structure of calyxin H, the lead molecule, is modified to investigate the improvement in drug-likeness, toxicity, or binding energy.

In silico derivatives of calyxin H

Different derivatives of calyxin H are designed by alteration the structure in silico through build module of Hyperchem software. The IUPAC name of different derivative are listed in Table 4. The 2D structures of derivatives are depicted in Figure 3.

The binding free energies of these in silico derivatives with FAD and TCF binding sites of LSD1 are obtained by docking and the scores are depicted in Table 5.

Prediction of drug-likeness of derivatives of the lead molecule

All the derivative are analysed through Molsoft and ProTox online tool for drug-likeness test and the scores are listed in Table 6.

Several reports on inhibition of LSD1 to suppress the tumor growth rate, and cellular proliferation are available.1 Many molecules are in pre-clinical and clinical phases of trials, acting as LSD1 inhibitors. In this report we have suggested new phytochemicals that specifically inhibit LSD1. It is reported that monoamine oxidase inhibitors inhibit LSD1 activity.11 The lead molecule, calyxin H, and its derivatives show good drug-likeness scoring as well as tolerable toxicity behaviour.

Table 1: Binding free energies (kcal/mol) of top five hits on FAD and TCF binding site of LSD1.

FAD -8.154 FAD -7.993
Calyxin H -13.222 Calyxin A -15.902
Calyxin L -11.858 Calyxin H -14.991
Epicalyxin B -10.857 Secofoveogline -14.488
22-epicalamistrin -10.845 Calyxin L -13.814
Calyxin A -10.668 Blepharocalyxin C -13.262
TCF -6.453 TCF -7.014
Calyxin H -14.631 Deoxycalyxin A -14.123
Calyxin A -11.907 Calyxin L -13.652
Epicalyxin B -11.617 Calyxin H -13.626
Secofoveogline -11.534 Epicalyxin B -13.170
Deoxycalyxin A -11.397 22-epicalamistrin -12.356

Figure 1: Calyxin H bound to FAD binding site of enzyme LSD1 when TCF binding site is occupied. A=Surface binding locations, B=interactions with binding site residues of calyxin H. 

Figure 2: Calyxin H bound on TCF binding site of enzyme LSD1 when FAD binding site is occupied. A=Surface binding locations, B=interactions with binding site residues of calyxin H.

The derivative-5 and 4 show better binding energies as compared to the lead towards the FAD binding site in presence and absence of TCF respectively while derivative-1 and 4 in case of TCF binding site in presence and absence of FAD respectively.

The Molsoft scores of the derivative-1 and 4 are comparable with calyxin H. The derivatives of calyxin H show better ProTox scores than calyxin H in terms of LD50 and toxicity class. The molecular weight of all the derivatives less than 500 which is a favourable drug like value.

Table 2: Prediction of drug-likeness of lead compound by Molsoft.

Molsoft score of lead molecule
Name of ligand Molecular weight (≤500) H bond acceptors (≤10) H bond donors (≤5) logP (octanol / water) (-4.0 to 5.6) Polar surface area (0-150Å2 ) Drug likeness score
Calyxin H 566.23 7 5 6.99 104.18 1.73

Table 3: Prediction of toxicity of lead compound by ProTox.

Toxicity prediction using ProTox
Name of ligand LD50 (mg/kg body weight ) Toxicity class (1-6) Toxycity target Average Pharmacophore Fit Average similarity known Ligand
Calyxin H 1000 4 Amine Oxidase A 33.45% 0.00
Prostaglandin G/H Synthase 1 40.01% 77.31%

Table 4: Listing of calyxin H derivative with their molecular formula and IUPAC name

In Silico derivative of Calyxin H Molecular formula IUPAC name
Calyxin H C35H34O7 (E)-1-[2,4-dihydroxy-3-[ (E,1S,5S)-5-hydroxy-1- (4-hydroxyphenyl)-7-phenylhept-2-enyl]-6-methoxyphenyl]-3- (4-hydroxyphenyl)prop-2-en-1-one
Derivative-1 C28H28O7 (2E)-1-{2,4-dihydroxy-3-[ (2E)-5-hydroxy-1- (4-hydroxyphenyl)hex-2-en-1-yl]-6-methoxyphenyl}-3- (4-hydroxyphenyl)prop-2-en-1-one
Derivative-2 C27H26O7 (2E)-1-{2,4-dihydroxy-3-[ (2E)-5-hydroxy-1- (4-hydroxyphenyl)pent-2-en-1-yl]-6-methoxyphenyl}-3- (4-hydroxyphenyl)prop-2-en-1-one
Derivative-3 C27H26O6 (2E)-1-{2,4-dihydroxy-3-[ (2E)-5-hydroxy-1-phenylpent-2-en-1-yl]-6-methoxyphenyl}-3- (4-hydroxyphenyl)prop-2-en-1-one
Derivative-4 C28H26O8 4-[ (2E)-1-{2,6-dihydroxy-3-[ (2E)-3- (4-hydroxyphenyl)prop-2-enoyl]-4-methoxyphenyl}-5-hydroxypent-2-en-1-yl]benzoic acid
Derivative-5 C24H20O7 4- ({2,6-dihydroxy-3-[ (2E)-3- (4-hydroxyphenyl)prop-2-enoyl]-4-methoxyphenyl}methyl)benzoic acid

Table 5: Binding free energies (kcal/mol) of calyxin H derivatives in the binding sites of LSD1 under four different situations.

Calyxin H -13.222 -14.991 -14.631 -13.626
Derivative-1 -8.545 -11.132 -14.754 -9.926
Derivative-2 -10.286 -12.856 -10.506 -11.599
Derivative-3 -10.968 -10.364 -10.299 -10.871
Derivative-4 -11.692 -15.365 -11.215 -14.259
Derivative-5 -14.824 -10.645 -9.848 -10.895

Table 6: Molsoft and ProTox score of calyxin H and its derivatives.

MolSoft and Protox scores of calyxin H and its derivatives
  Mol. Weight MolSoft LD50 (in mg/kg body weight) Toxicity Class (1-6)
Calyxins H 566.23 1.73 1000 4
Derivative-1 476.18 1.64 3800 5
Derivative-2 462.17 1.47 3800 5
Derivative-3 446.17 1.31 3800 5
Derivative-4 490.16 1.64 3800 5
Derivative-5 420.12 1.40 3800 5

Figure 3: The 2D structures of calyxin H and its derivatives (A= Calyxin H, B= Derivative-1, C= Derivative-2, D= Derivative-3, E= Derivative-4, F= Derivative-5).

Diarylhepatanoids (calyxins) in the ethanolic extract of seeds of Alpinia blepharocalyx have been reported to have anti-proliferative action. However, mechanism of the activity is still not clear.12 It is proposed to be associated with the antioxidant property of the calyxins.12 The present report provides a clue to the antiproliferative acivity of calyxin H by inhibition of the enzyme LSD1 associated with metastasis.

The docking scores suggest that the lead molecule may decrease the catalytic activity of LSD1 by competitive binding to the active site of the native substrate. The binding of these small ligands may prohibit the substrate binding and thereby decreasing the rate of catalysis. This eventually may block the cell proliferation. Our work thus suggests calyxin H and its derivatives may serve as future possible drug molecules for therapeutic inhibition of LSD1.

Funding: No funding sources Conflict of interest: None declared


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