mohan et al., 1:10 open access scientific reports€“7230-sr-471.pdf · citation: mohan a,...

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Mohan et al., 1:10http://dx.doi.org/10.4172/scientificreports.471

Research Article Open Access

Open Access Scientific ReportsScientific Reports

Open Access

Volume 1 • Issue 10 • 2012

Keywords: Eye cancer; Retinoblastoma 1; Docking; Cyclophosphamide analogs; Absorption distribution metabolism excretion

IntroductionRetinoblastoma is a rare eye cancer that develops in retina of the

eye in early childhood before the age of five years. Incidence of 1 in 20,000 live births. Retinoblastoma 1 regulates transition of cell cycle from G2 phase to S phase of the cell cycle. The retinoblastoma 1 protein which was located on chromosome 13q14 was first cloned by friend et al in 1986. Rb1 is a nucleoprotein and it has a molecular weight of 110 kD. The retinoblastoma 1 gene is composed of 27 exons that span 180 kb of chromosome and encodes protein of 928 amino acids. The large intron spans more than 60 kb, the smallest intron has 80 bp. The complete genome sequencing of the retinoblastoma 1 gene was reported by Toguchida et al. [1-3]. The retinoblastoma 1 protein having two pocket binding sites namely pocket A and pocket B. These binding sites are mainly involved in normal function of Retinoblastoma 1. Some onco proteins mainly bind to the retinoblastoma 1 binding pocket A (amino acid 394 to 572) and pocket B (amino acid 646 to 792). These binding sites are mainly involved in normal function of Retinoblastoma 1, which causes mutation due to binding at the same site can leads to defect Retinoblastoma 1 function [4]. Rb1 bind to the E2F and form chromatin remodeling protein like histone deacetylase, histone methyltransferases is used to promote cell cycle regulation, DNA replication, DNA repair, G2/M checkpoint activation, differentiation [5]. Retinoblastoma 1 binds with E2F at G0 and G1 phase of the cell cycle. These interactions can leads to promote transcriptional activity. Some viral onco protein bind with binding pocket of retinoblastoma1 like, SV 40 large T antigen, human papilloma virus E7 protein, adenovirus E and A. This protein defects Rb1 tumour suppressor activity and cell cycle [6]. Insertion, deletions, point mutation, germ line mutation, genetic alterations, frame shift, missense mutation are important in the inactivation of retinoblastoma1 tumour suppressor activity. Inactivation of retinoblastoma 1 gene can lead to develop retinoblastoma, sporadic breast, bladder, prostate, small cell lung carcinoma, head and neck cancer, esophagus, pancreatic cancer, melanoma. Inactivation of retinoblastoma 1 can case approximately 20%

*Corresponding author: Dr. AM Lakshmi Prabha, Assistant Professor, Department of Plant Science, School of Life Sciences, Bharathidasan University, Tiruchirappalli-620024, Tamil Nadu, India, Tel: +91431 2407061; Fax: +91431 2407045; E-mail: [email protected]

Received August 20, 2012; Published October 29, 2012

Citation: Mohan A, Manickavasagam M, Ganapathi A, Lakshmi Prabha AM, Jeeva A (2012) Molecular Interaction Studies of 3D Conformers of Cyclophosphamide against Human Eye Cancer Protein Retinoblastoma1. 1:471. doi:10.4172/scien-tificreports.471

Copyright: © 2012 Mohan A, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

AbstractRetinoblastoma is a rare eye cancer form in both benign and malignant tumour especially affecting child.

Retinoblastoma 1 is a tumour suppressor gene. Functionality of the protein specified in two pockets, pocket A (amino acid 394 to 572) and pocket B (amino acid 646 to 792). This binding site is mainly involved in normal function of retinoblastoma 1 tumour suppressor activity. In the present study we have analyzed antitumour activity for human eye cancer from cyclophosphamide analogs. Totally we selected ten cyclophosphamide analogs and one currently available market drug cyclophosphamide as a reference compounds for molecular docking studies against human eye cancer protein Retinoblastoma 1 and also Absorption Distribution Metabolism Excretion properties of these compounds were carried out by using QikProp. Among the many cyclophosphamide analogs one of them (perfosfamide) are showing best glide score and more number of hydrogen bond against human eye cancer protein retinoblastoma, and it was found to be under acceptable range with predicted ADME properties. From the molecular interaction studies we are concluding that the above mentioned cyclophosphamide analogs perfosfamide have best anti tumour activity against human eye cancer.

Molecular Interaction Studies of 3D Conformers of Cyclophosphamide against Human Eye Cancer Protein Retinoblastoma 1Anbuselvam Mohan1, Markandan Manickavasagam1, Andy Ganapathi1, Azhagiya Manavalan Lakshmi Prabha2* and Anbuselvam Jeeva3

1Department of Biotechnology and Genetic Engineering, Bharathidasan University, Tiruchirappalli, Tamil Nadu, India2Department of Plant Science, Bharathidasan University, Tiruchirappalli, Tamil Nadu, India3Department of Animal Science, Bharathidasan University, Tiruchirappalli, Tamil Nadu, India

-35% of breast cancer. Heterozygosity loss of Rb1 can cause tumor. 40% of retinoblastoma is caused by hereditary and remaining 60% caused are without familial history [1,2]. In the current study we have analyzed anti-tumour activity of the compounds from ten cyclophosphamide analogs and one commercially available compound cyclophosphamide as reference compound for in silico docking studies against human eye cancer protein Retinoblastoma 1, ADME properties were carried out for these compounds by using Glide extra precision and QikProp.

Materials and MethodsCrystal structure selection for target protein Rb1

There are several X-ray crystal structures were available in the Protein Data Bank for Rb1 protein. The better quality of the retinoblastoma 1 protein structure was identified by using model validation methods PROCHECK and ERRAT [7]. The PROCHECK programs are useful for assessing the quality not only of protein structures in the process of being solved but also of existing structures and of those being modeled on known structures. The PROCHECK suite of programs provides a detailed check on the stereo chemical quality of a given protein structure. The PROCHECK was used to generate Ramachandran plot and the quality of the structure was computed in terms of % of residues in favorable regions, % of non proline, glycine residues etc. The quality of structure was also further accessed by using ERRAT [8]. ERRAT is a protein structure

http://dx.doi.org/10.4172/scientificreports.471


Citation: Mohan A, Manickavasagam M, Ganapathi A, Lakshmi Prabha AM, Jeeva A (2012) Molecular Interaction Studies of 3D Conformers of Cyclophosphamide against Human Eye Cancer Protein Retinoblastoma1. 1:471. doi:10.4172/scientificreports.471

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verification algorithm that is especially well-suited for evaluating the progress of crystallographic model building and refinement.

Preparation of protein structure

The crystal structure of Retinoblastoma 1 protein (PDB ID: 2R7G), was recovered from the Protein Data Bank [9]. After selected the protein structure, protein preparation wizard of Schrödinger suite has been used to prepare protein structure. All unwanted water molecules were removed from the protein structure, metal are treated and hydrogen atoms were added, all atom force field (OPLS-2005) charges and atom types were assigned. Energy was minimized until the average root mean square deviations of non-hydrogen atoms reached 3.0 Å.

Preparation of Cyclophosphamide and their analogs

Cyclophosphamide is currently available drug for retinoblastoma 1[10]. The 3D structures of cyclophosphamide and their ten analogs were obtained from Pubchem database [11]. All these ligands structures was prepared for molecular docking studies using ligprep, version 2.3 [12]. The ligand structure energy were minimized, partial atomic charges were computed using the OPLS-2005 force field by using

Schrödinger suite. The 2D representation of the cyclophosphamide and their ten analogs were depicted in figure 1.

Docking studies

The prepared protein was further taken into docking studies. The cyclophosphamide and their ten analogs were obtained from Pubchem database. Each ligand is separately docked with structure of the retinoblastoma 1 protein by using Glide extra precision [13]. The grid was prepared for protein with the exact same center and the size of the bounding box set on 30 Å. The grid box enclosed at the centroid of the retinoblastoma receptor. The grid box center -8.8452, -6.9348, and -15.9663 for x, y and z coordinates. The glide algorithm is based on a systematic search of positions, orientation and conformations of the ligand in the receptor binding site using funnel type approach. The cut off extra precision scoring parameters for extra precision docking were set to 0.0 Kcal/mol. The glide score and glide energy were analyzed by using glide extra precision visualizer.

Docking complex analysis

Perfosfamide 4-s-(hexane-6-01)-sulfidocyclophosphamide Trofosfamide

4-s-ethnolsulfide-cyclophosphamide Cyclophosphamide L- Cyclophosphamide

D- Cyclophosphamide 5-Bromo-Cuclophosphamide Ifosfamide

6-methyl cyclophosphamide 4,4-6,6-D4-cyclophosphamide

Figure 1: This figure represents 2D structure of cyclophosphamide and their analogs.



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and acceptor, log p (octanol/water), log p (octanol/gas), log p (water/gas), log p (hexadecane/gas), log p Caco cell permeability, log pMDCK, serum binding protein, Lipinski’s rule of five, percentage of human oral absorption, which is essential for rational drug design [15-17].

Results and DiscussionSeveral crystal structures are available for the retinoblastoma

1 protein in protein Data Bank. We have analysis structure quality checking by using PROCHECK and ERRAT to identify the structure which is the best one. The results shows the 2R7G crystal structure was good quality for molecular docking studies based on the quality factor and stereochemistry analysis and structure resolution value from PDB among ten Retonoblastoma1 crystal structures. The Ramachandran plot shows most favored region 94.4% residues, in additional allowed region 5.1% residues, in generously allowed region 0.3% residues and in disallowed region 0.2% residues (Figure 2). The structure resolution value 1.67 Å was obtained from Protein Data Bank. The overall quality factor of the protein 98.946 shows in ERRAT plot (Figure 3). The PROCHECK, ERRAT and structure resolution value were reported in table 2.

Protein-ligand complex interaction analysis

Docking of retinoblastoma 1 with perfosfamide: Docked simulation of perfosfamide into Retinoblastoma 1 resulted in the formed five hydrogen bonds and observed the chain A ASN 522 protein molecule of the hydrogen atom interacted with oxygen atom of the Ligand molecule with bond length (1.8 Å). The protein molecule of the hydrogen atom, act as hydrogen bond donor. The oxygen atom of the ligand molecule, act as hydrogen bond acceptor. The protein molecule of the chain C ARG 418 hydrogen atom interacted with oxygen atom of the Ligand with bond distance (2.4 Å). The hydrogen atom of the protein molecule, act as hydrogen bond donor whereas the ligand molecule of

Protein-ligand complex were further taken into PyMol. PyMol is a molecular visualization tool. PyMol viewer is used to analyse the type of interaction between ligand and protein. To identify the residues located in binding pocket, protein structure were enlarge around the bound ligand [14]. The hydrogen bond interaction and their distance were analyzed were depicted in figure 4-14. The glide score, glide energy, number of hydrogen bonds and their distance were reported in table 1.

ADME properties analysis

The QikProp program was used to predict the ADME properties of the compounds. The QikProp predicts both pharmaceutically relevant and physically significant descriptor. The predicted ADME properties such as number of molecular weight, hydrophophic SASA, hydrophilic SASA, number of rotatable bonds, number of hydrogen bond donor

Ramachandran Plot2R7G

PROCHECK

180

135

90

45

0

-45

-90

-135

-180 -135 -90 -45 0 45 90 135 180Phi (degrees)

Psi (

degr

ees)

GLU 746 (C)

∼b

∼b

∼b

∼b

∼a

∼pp

∼1

a

b

B

b

b

1

Figure 2: The Ramachandran Plot describes 94.4% of residues fall in the most favored region.

Compound ID Compound name Glide score Glide energy No. of H bond Interacting residues

Distance H bond donor H bond acceptor

38347 Perfosfamide -6.50 -44.10 5 ASN 522ARG 418

ASN 480 (2)

ASNN 480

1.82.42.22.11.7

A:ASN522:(H)HD21C:ARG418:(H)HE

Ligand: H C:ASN480: (H)HD22

A:ASN480: (H)

Ligand: OLigand: O

C: ASN480:(O)OD1Ligand: OLigand: O

129828 4-s-(hexane-6-01)-sulfidocyclophosphamide

-5.44 -71.06 5 ASN 478SER 414

ASN 480 (2)

ASN 480

2.02.12.11.21.9

Ligand: H A:SER414: (H)HG

A:ASN480: (H)HD22Ligand: H

C:ASN 480: (H)HD22

A:ASN478: (O)OD1Ligand: OLigand: O

A:ASN480: (O)OD1Ligand: O

65702 Trofosfamide -5.20 -49.38 1 ASN 480 2.1 A:ASN480: (H)HD22 Ligand: O128314 4-s-ethnolsulfide-

cyclophosphamide-5.09 -53.52 1 ASN522 2.1 Ligand: H A: ASN522: (O)

2907 Cyclophosphamide -5.04 -41.87 2 ASN 480LEU 521

2.02.3

C:ASN 480: (H)HD22Ligand: H

Ligand: OC:LEU521: (O)

9554282 L- Cyclophosphamide -5.04 -41.87 2 ASN 480LEU 521

2.02.3


Ligand: OC:LEU521: (O)

9554281 D- Cyclophosphamide -5.03 -39.92 2 ASN 480LEU 521

2.12.1

A:ASN480: (H)HD22Ligand: H

Ligand: OA: LEU521:(O)

320153 5-Bromo-Cuclophosphamide

-5.01 -40.86 3 ASN 480 (2)

ASN 480

1.9,2.82.1

A:ASN480: (H)HD22A:ASN480: (H)HD22C:ASN 480: (H)HD22

Ligand: OLigand: OLigand: O

3690 Ifosfamide -4.80 -36.37 3 ASN 480 (2)

ASN 480

2.1,2.12.1


A:ASN 480: (H)HD22

Ligand: OC:ASN480: (O)OD1

Ligand: O84041 6-methyl

cyclophosphamide-4.19 -36.99 2 LEU 521

ASN5222.12.1

Ligand: HA:ASN: (H)HD21

A:LEU521: (O)Ligand: O

3035226 4,4-6,6-D4-cyclophosphamide

-2.20 -36.44 2 ASN522LEU 521

2.12.1

A:ASN: (H)HD21Ligand: H

Ligand: OA:LEU521: (O)

Table 1: Docking results for the cyclophosphamide and their ten analogs against human Retinoblastoma1.



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the oxygen atom, act as hydrogen bond acceptor. The protein molecule of the chain C ASN 480 formed two hydrogen bonds with ligand. We observed the distance (2.2 Å, 2.1 Å). The oxygen atom of the protein molecule interacted with hydrogen atom of the ligand molecule. The hydrogen atom of the ligand molecule, act as hydrogen bond donor. The protein molecule of the oxygen atom, act as hydrogen bond acceptor. The chain C, ASN 480 protein molecule of the hydrogen atom is well interacted with oxygen atom of the ligand molecule with bond length (2.1 Å). The hydrogen atom of the protein molecule, act as hydrogen bond donor. The oxygen atom of the ligand molecule, act as hydrogen bond acceptor. The oxygen atom of the ligand molecule interacted with the chain A ASN 480 protein molecule of hydrogen atom with bond distance (1.7 Å).The protein molecule of the hydrogen atom act as hydrogen bond donor. The ligand molecule of oxygen atom, act as hydrogen bond acceptor (Figure 4), and the observed glide score glide energy for perfosfamide were -6.50 and -44.10 kcal/mol respectively.

Docking of retinoblastoma 1 with 4-s-(hexane-6-01)-sulfido-cyclophosphamide: Docked simulation of 4-s-(hexane-6-01)-sulfido-cyclophosphamide into retinoblastoma1 resulted in the formation of five hydrogen bonds and observed the chain A ASN 478 oxygen atom

400 420 440 460 480 500 520 540 560 580 600 620 640 660 680Residue # (Window center)

Chain#: 1Overall quality factor**: 98.946

99%

95%Err

or v

alu*

Figure 3: The overall quality factor of the errat plot is 98.946 which indicate that the structure become more reliable.

Figure 4: This figure shows the docked structure of the target protein Retinoblastoma 1 and the ligand Perfosfamide.

protein molecule well interacted with hydrogen atom of the ligand molecule with bond length (2.0 Å).The hydrogen atom of the ligand molecule of the act as hydrogen bond donor. The oxygen atom of the protein molecule, act as hydrogen bond acceptor. The oxygen atom of the Ligand molecule interacted with chain C SER 414 protein molecule of hydrogen atom with bond distance (2.1 Å). The protein molecule of chain A SER 414 hydrogen atom act as hydrogen bond donor, the oxygen atom ligand molecule act as hydrogen bond acceptor. The protein molecule of the chain C ASN 480 formed two hydrogen bonds with ligand. The ASN 480 hydrogen atom interacted with oxygen atom of the ligand molecule with bond length (2.1 Å). The ASN 480 present in chain A. The ligand molecule of oxygen atom, act as hydrogen bond acceptor. The protein molecule of the hydrogen atom, act as hydrogen bond donor. The hydrogen atom ligand molecule interacted with oxygen atom of protein molecule ASN chain A with bond length (1.2 Å). The ligand molecule of hydrogen atom, act as hydrogen bond donor. The chain A, ASN 480 protein molecule of oxygen atom act as hydrogen bond acceptor. The chain C protein molecule of hydrogen atom interacted with oxygen atom of the ligand molecule. The hydrogen atom of the protein molecule, act as hydrogen bond donor. The ligand molecule of oxygen atom act as hydrogen bond acceptor with bond length (1.9 Å) (Figure 5). The observed glide score glide energy for 4-s-(hexane-6-01)-sulfido-cyclophosphamide was -5.44 and -71.06 kcal/mol respectively.

Docking of retinoblastoma 1 with trofosfamide: Only one hydrogen bond was formed between docked simulations of retinoblastoma 1 into trofosfamide. The glide score glide energy was noted for trofosfamide were -5.20 and -49.38 kcal/mol respectively. The ligand molecule of oxygen atom well interacted with chain A ASN 480 protein molecule of hydrogen with bond length (2.1 Å). The protein molecule of the hydrogen atom, act as hydrogen bond donor whereas oxygen atom of the ligand molecule, act as hydrogen bond acceptor (Figure 6).

Docking of retinoblastoma 1 with 4-s-ethnolsulfide-cyclophosphamide: The docked simulation Retinoblastoma 1 into 4-s-ethnolsulfide-cyclophosphamide formed only one hydrogen bond. The glide score, glide energy of complex was observed with -5.09 and -53.42 kcal/mol respectively. The hydrogen atom of the ligand molecule well interacted with side chain oxygen atom of the chain A ASN 522 protein molecule of hydrogen with bond distance (2.1 Å). The oxygen atom of the protein molecule, act as hydrogen bond acceptor whereas the hydrogen atom of the ligand molecule, act as hydrogen bond donor (Figure 7).

Docking of retinoblastoma 1 with cyclophosphamide: The docked simulation retinoblastoma 1 into cyclophosphamide formed

S. No PDB ID Most favored region (%) Additional allowed region (%) Generously allowed region Disallowed region Errat Resolution (Å)

1 2R7G 94.4 5.1 0.3 0.2 98.946 1.67

2 1GUX 91.7 7.7 0.7 0.0 96.764 1.853 2QDJ 92.8 7.2 0.0 0.0 96.341 2.004 1N4M 85.2 11.4 2.7 0.7 87.408 2.205 1AD6 89.0 9.8 0.6 0.6 94.350 2.306 3POM 89.2 10.5 0.3 0.0 97.561 2.507 2AZE 89.7 8.4 1.9 0.0 89.020 2.558 1O9K 88.0 10.6 1.3 0.1 89.302 2.609 1GH6 75.6 20.0 3.2 0.2 84.471 3.20

10 3N5U 79.8 17.9 1.2 1.2 94.056 3.20

Table 2: Over all ramachandran plot results and structure resolution value for selected PDB ID’s.



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two hydrogen bonds with bond distance (2.0 Å, 2.3 Å). The protein molecule of chain C ASN 480 hydrogen atom interacted with oxygen atom of ligand molecule. The protein molecule of hydrogen atom, act as hydrogen bond donor whereas the oxygen atom of the ligand molecule, act as hydrogen bond acceptor. The side chain oxygen atom of the protein molecule LEU 521 interacted with hydrogen atom of the ligand molecule. The hydrogen atom of the ligand molecule, act as hydrogen bond donor. The side chain oxygen atom of the protein molecule, chain C LEU 521 act as hydrogen bond acceptor (Figure 8). The glide score and glide energy calculated by glide extra precision for Cyclophosphamide was -5.04 and -41.87 kcal/mol respectively.

Docking of retinoblastoma 1 with L- cyclophosphamide: The docked simulation retinoblastoma 1 into L- cyclophosphamide formed two hydrogen bonds with bond distance (2.0 Å, 2.3 Å). The interaction energies calculated for L- Cyclophosphamide are -5.04 kcal/mol of

glide score and -41.87 kcal/mol of glide energy. The protein molecule of chain C ASN 480 hydrogen atom interacted with oxygen atom of ligand molecule. The protein molecule of hydrogen atom, act as hydrogen bond donor whereas the oxygen atom of the ligand molecule, act as hydrogen bond acceptor. The side chain oxygen atom of the protein molecule C LEU 521 interacted with hydrogen atom of the ligand molecule. The hydrogen atom of the ligand molecule, act as hydrogen bond donor. The side chain oxygen atom of the protein molecule chain C LEU 521 act as hydrogen bond acceptor (Figure 9).

Docking of retinoblastoma 1 with D- cyclophosphamide: The docked simulation retinoblastoma 1 into D- cyclophosphamide formed two hydrogen bonds. The calculated glide score was -5.03 kcal/mol for both the interactions formed for protein molecule of chain C. The hydrogen atom of protein molecule ASN 480 interacts with oxygen atom of ligand molecule with bond length (2.1 Å). The protein molecule acts as hydrogen bond donor, the ligand molecule act as hydrogen bond acceptor. The hydrogen atom of ligand molecule interacted with side chain protein molecule of LEU 521 with bond distance (2.1 Å). The hydrogen atom of the ligand molecule, act as hydrogen bond donor. The LEU 521 protein molecule of oxygen atom, act as hydrogen bond acceptor (Figure 10).

Docking of retinoblastoma 1 with 5-Bromo-cyclophosphamide: Three hydrogen bonds were formed between Retinoblastoma 1 and 5-Bromo-Cuclophosphamide. The protein molecule of ASN 480 residues formed two hydrogen bonds with bond distance (1.9Å, 2.8Å). The oxygen atom of the ligand molecule well interacted with protein molecule of ASN 480 residues. The ASN 480 residues present in chain A. The oxygen atom of the ligand, act as hydrogen bond acceptor. The hydrogen atom of the protein molecule, act as hydrogen bond donor.

Figure 5: This figure shows the docked structure of the target protein Retinoblastoma 1 and the ligand 4-s-(hexane-6-01)-sulfidocyclophosphamide.

Figure 6: This figure shows the docked structure of the target protein Retinoblastoma 1 and the ligand Trofosfamide.

Figure 7: This figure shows the docked structure of the target protein Retinoblastoma 1 and the ligand 4-s-ethnolsulfide-cyclophosphamide.

Figure 8: This figure shows the docked structure of the target protein Retinoblastoma 1 and the ligand Cyclophosphamide.

Figure 9: This figure shows the docked structure of the target protein Retinoblastoma 1 and the ligand L- Cyclophosphamide.



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The oxygen atom of the ligand interacted with hydrogen atom of the chain C ASN 480 protein molecule with bond distance (2.1 Å) (Figure 11). The glide score of the complex was calculated as -5.01 kcal/mol.

Docking of retinoblastoma 1 with ifosfamide: Docked simulation of retinoblastoma 1 into ifosfamide formed three hydrogen bonds. The hydrogen atom of the protein molecule ASN 480 residues interacted with oxygen atom of the ligand with bond length (2.1 Å). The hydrogen atom of the protein molecule, act as hydrogen bond donor. The oxygen atom of the ligand molecule, act as hydrogen bond acceptor. The oxygen atom of the protein molecule ASN 480 residues interacted with hydrogen atom of the ligand molecule with bond distance (2.1 Å). The hydrogen atom of the ligand, act as hydrogen bond donor whereas oxygen atom of the protein molecule ASN 480 act as hydrogen bond acceptor. The ASN 480 residues present in chain C. The hydrogen atom of the protein molecule chain A ASN 480 residues interacted with oxygen atom of the ligand with bond distance (2.1 Å). The hydrogen atom of the residues, ASN 480 acts as hydrogen bond donor. The ligand molecule of oxygen atom, act as hydrogen bond acceptor (Figure 12). The glide score and glide energy were calculated (-4.80, -36.37).

Docking of retinoblastoma 1 with 6-methyl cyclophosphamide: Two hydrogen bonds were identified between Rb1 with 6-methyl cyclophosphamide with distance (2.1 Å, 2.1 Å). The oxygen atom of LEY 521 residues interacted with hydrogen atom of the ligand molecule. The oxygen atom of ligand interacted with hydrogen atom of the ASN 522 residues. The hydrogen atom of the ASN 521, act as hydrogen bond donor (Figure 13). This complex was formed with glide score of -4.19 kcal/mol and glide energy of 36.99 kcal/mol.

Docking of Retinoblastoma 1 with 4,4-6,6-D4-cyclophosphamide: Two hydrogen bond were formed between Rb1 with 4, 4-6, 6-D4-cyclophosphamide. The glide score of this complex was -2.20 kcal/mol. The hydrogen atom of the protein molecule of ASN 522 residues interacted with oxygen atom of the ligand with bond distance (2.1 Å). The oxygen atom of the LEU 521 residue interacted with hydrogen atom of the ligand with bond length (2.1 Å). The hydrogen atom of the ligand, act as hydrogen bond donor (Figure 14).

Pharmacokinetics properties prediction

We have analyzed 44 physically significant descriptor and Figure 10: This figure shows the docked structure of the target protein Retinoblastoma 1 and the ligand D- Cyclophosphamide.

Figure 11: This figure shows the docked structure of the target protein Retinoblastoma 1 and the ligand 5-Bromo-Cuclophosphamide.

Figure 12: This figure shows the docked structure of the target protein Retinoblastoma 1 and the ligand Ifosfamide.

Figure 13: This figure shows the docked structure of the target protein Retinoblastoma 1 and the ligand 6-methyl cyclophosphamide.

Figure 14: This figure shows the docked structure of the target protein Retinoblastoma 1 and the ligand 4,4-6,6-D4-cyclophosphamide.



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pharmaceutically relevant properties of the cyclophosphamide and their ten cyclophosphamide analogs including molecular weight, hydrogen bond donor, hydrogen bond acceptor, log p (octanol/water), log p (octanol/gas), log p (water/gas), log p (hexadecane/gas), log p Caco cell permeability, log pMDCK, serum binding protein, Lipinski’s rule of five, percentage of human oral absorption were reported in table 3. Lipinski’s rule of five is a rule of thumb to evaluate drug likeness, or determine if a chemical compound with a certain pharmacological (or) biological activity has properties that would make it a like orally active drug in human, The rule describes molecular properties that are important in the drug is pharmacokinetics in the human body, including its ADME. These compounds were further evaluated for their drug-like behavior through analysis of pharmacokinetics parameters required for absorption, distribution, metabolism, excretion (ADME) by using QikProp. The cyclophosphamide and their ten analogs were in the acceptable range of Lipinski’s rule of five, hydrophobic SASA, hydrophilic and partition coefficient (QP log p (octanol/water)), that defines their liphophilic nature and quality these compounds for proper absorption and permeability. While the cell permeability (QPlog pCaco), a key factor governing drug metabolism and its access to biological membranes, ranged from 687 to 6216. QPPMDCK ranges from -3.070 to 10000. Overall, the percentage human oral absorption for the compounds ranged from 74 to 100%. All the pharmacokinetics parameters are within the acceptable range defines for human use, there by indicates their potential use as drug-like molecules.

ConclusionIn the present study molecular interaction studies were performed

for ten cyclophosphamide analogs and commercially available drug cyclophosphamide. The following ligand perfosfamide having better glide score (-6.50) and more number of hydrogen bond interaction good binding affinity with Rb1 protein (PDB ID: 2R7G) when compare to the commercially available drug cyclophosphamide and their analogs. From the molecular interaction studies we concluded that the following ligand perfosfamide could a potential inhibitor for human eye cancer protein Rb1 in human.

References

1. Hung CC, Lin SY, Lee CN, Chen CP, Lin SP, et al. (2011) Low penetrance of retinoblastoma for p.V654L mutation of the RB1 gene. BMC Med Genet 12: 76.

2. Sivakumaran TA, Shen P, Wall DP, Do BH, Kucheria K, et al. (2005) Conservation of the RB1 gene in human and primates. Human Mutat 25: 396-409.

3. Mateu E, Sánchez F, Nájera C, Beneyto M, Castell V, et al. (1997) Genetics of retinoblastoma: a study. Cancer Genet Cytogenet 95: 40-50.

4. Saijo M, Sakai Y, Kishino T, Niikawa N, Matsuura Y, et al. (1995) Molecular cloning of a human protein that binds to the retinoblastoma protein and chromosomal mapping. Genomics 27: 511-519.

5. Nead MA, Baglia LA, Antinore MJ, Ludlow JW, McCance DJ (1998) Rb binds c- Jun and activates transcription. EMBO J 17: 2342-2352.

6. Dannenberg JH, Schuijff L, Dekker M, van der Valk M, te Riele H (2004) Tissue-

3035226

261.087

437.667

238.155

54.493

3.000

1.000

8.500

6.906M

12.697M

10.369M

0.729

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0.295

1

-3.070

10000

3014

0

93

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275.114

472.326

273.452

54.869

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1.000

8.500

7.412M

13.536M

10.455M

1.080

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0.271

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-3.368

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2989

0

95

3690

261.087

455.642

276.585

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1.000

8.500

6.822M

12.493M

10.260M

0.992

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0.472

2

-3.389

10000

4930

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100

320153

339.984

467.772

196.897

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8.500

7.831M

13.745M

10.464M

1.307

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0.458

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-3.347

10000

3043

0

97

9554281

261.087

453.298

254.020

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6.970M

12.843M

10.403M

0.827

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0.284

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-3.337

10000

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94

9554282

261.087

437.669

238.160

54.492

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1.000

8.500

6.906M

12.697M

10.369M

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-0.974

0.295

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-3.070

10000

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261.087

437.669

238.160

54.492

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6.906M

12.697M

10.369M

0.729

-0.974

0.295

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-3.070

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93

128314

337.200

535.210

265.270

91.937

7.000

2.000

10.700

9.249M

17.078M

13.619M

0.862

-0.979

-0.246

2

-3.762

6361

1330

0

88

65702

323.586

520.007

281.762

21.336

4.000

0.000

9.000

8.327M

13.523M

9.174M

1.487

-1.336

0.666

2

-3.647

10000

6216

0

100

129828

393.308

632.695

369.243

99.608

11.000

2.000

10.700

11.218M

19.174M

13.019M

1.986

-0.704

-0.640

2

-4.134

4439

1125

0

93

38347

293.086

463.079

195.857

122.191

5.000

2.000

11.900

8.029M

16.648M

15.263M

-0.616

-1.273

-0.402

1

-3.253

2055

687

0

74

Predicted properties and compound ID

Molecular weight(130.0/725.0)

Total SASA (300.0/1000.0)

Hydrophobic SASA(0.0/750.0)

Hydrophilic SASA(7.0/330.0)

No. of rotatable bond(0.0/15.0)

Hydrogen bond donor(0.0/6.0)

Hydrogen bond acceptor(2.0/20.0)

QPLogP Hexadecane/gas(4.0/18.0)

QPLogP octanol/gas(8.0/35.0)

QPLogP water/gas(4.0/45.0)

QPLogP octanol/water(-2.0/6.5)

Serum binding protein(-1.5/1.5)

QPLogBlood/brain(-3.0/1.2)

No. of primary metabolite

HERG channel blockage (below -5)

Apparent MDCK Permeability (<25 poor,>500 great)

Apparent Caco-2 Permeability (<25 poor,>500 great)

Lipinski Rule of Five (Maximum 4)

% of human oral absorption(<25 poor)

Table 3: Predicted ADME properties for cyclophosphamide and their ten analogs.


http://www.ncbi.nlm.nih.gov/pubmed/21615945







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specific tumor suppressor activity of retinoblastoma gene homologs p107 and p130. Genes Dev 18: 2952-2962.

7. Laskowski RA, MacArthur MW, Moss DS, Thornton JM (1993) PROCHECK: a program to check the stereochemical quality of protein structures. J Appl Cryst 26: 283-291.

8. Colovos C, Yeates TO (1993) Verification of protein structures: Patterns of nonbonded atomic interactions. Protein Sci 2: 1511-1519.

9. Liu X, Marmorstein R (2007) Structure of the retinoblastoma protein bound to adenovirus E1A reveals the molecular basis for viral oncoprotein inactivation of a tumor suppressor. Genes Dev 21: 2711-2716.

10. http://www.cancer.gov/cancertopics/druginfo/retinoblastoma.

11. Wang Y, Xiao J, Suzek TO, Zhang J, Wang J, et al. (2009) Pubchem: a public

information system for analyzing bioactivities of small molecules. Nucleic Acids Res 37: W623-W633.

12. Ligprep, version 2.3, Schrodinger, LLC, New York, NY, 2011.

13. Glide, version 5.5, Schrodinger, LLC, New York, NY, 2009.

14. http://www.pymol.org/

15. Duffy EM, Jorgensen WL (2000) Prediction of properties from simulations: Free Energies of Solvation in Hexadecane, Octanol, and Water. J Am Chem Soc 122: 2878-2888.

16. Lipinski CA, Lombardo F, Dominy BW, Feeney PJ (2001) Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings. Adv Drug Deliv Rev 46: 3-26.

17. Qikprop, version 3.3, Schrödinger, LLC, New York, NY, 2010.



http://scripts.iucr.org/cgi-bin/paper?gl0276





http://www.schrodinger.com/citations/44/10

https://www.schrodinger.com/products/14/5

http://www.pymol.org/

http://pubs.acs.org/doi/abs/10.1021/ja993663t


https://www.schrodinger.com/products/14/17

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