In Vitro & In Silico Studies of Cordia myxa Phytochemicals in Contrast to Breast Cancer: A Bioinformatics Approach

Ambresh V; Gowda P

Research Article

Austin J Proteomics Bioinform & Genomics. 2022; 7(1): 1032.

In Vitro & In Silico Studies of Cordia myxa Phytochemicals in Contrast to Breast Cancer: A Bioinformatics Approach

Ambresh V* and Gowda P

Department of Biotechnology, Bangalore City College, Bangalore, India

*Corresponding author: Ambresh V, Department of Biotechnology, Bangalore City College, Bangalore, India

Received: March 17, 2022; Accepted: April 11, 2022; Published: April 18, 2022

Abstract

Cordia myxa which is rarely found in the southern part of Karnataka due to the climatic conditions, but it is mostly found in the warmer region of Karnataka which is the north part of Karnataka (Raichur, Bellary, Gulbarga, Koppal, Bijapur). The people of north Karnataka use this medicinal plant to soften their tongue blisters. We have performed the phytochemical screening and antimicrobial activity using the extracted solvents of leaf phytochemicals and broth dilution methods. We observed the excellent zone of inhibition against the E. coli and Bacillus subtitlus with the default parameters.

In the present study, we will take a step to analyze those receptors using molecular docking studies. We have selected the phytochemicals and receptors for initiate the molecular docking and will investigate the inhibitory activity of phytocompounds using in silico studies.

Keywords: Cordia myxa; Phytocompounds; Molecular docking

Introduction

This plant has its own mythological value in the history of northern Karnataka. So that led the researchers of northern part of Karnataka to select the plant and extract to know its medicinal importance and to implement its uses accordingly.

Morphology

Cordia myxa thrive in about fifty-sixty years by when its girth at the breast height is about 1-1.5 m. Its main trunk is generally straight and cylindrical at height of almost 3-4 m. The branches open out in all direction forming a beautiful inverted dome like an umbrella. The height of grown-up tree measures around 10-15 m. The growth mainly depends upon climatic conditions, in an unfavourable condition it may even end up as a dwarf shrub.

It has a greyish brown coloured bark with longevity and upright crevice. This tree can be seen from far away due to its crevice which is projecting in the bark of the main trunk of a tree approaching maturity [1].

Breast cancer (Bc)

Breast cancer is a disease where in which the tissue from breast forms of malignant cancer. Usually, a tube carries milk to nipple (duct) and towards gland that from milk. It is most common for men and women, although in male disease diagnosis is rare. It is a heterogeneous that differs by indusial age group and kind of cell in the tumor. Cancer begins when healthy cells in the breast change and grow out of control, forming a mass or sheet of cells called a tumor. A tumor can be cancerous or benign. A cancerous tumor is malignant, meaning it can grow and spread to other parts of the body. A benign tumor means the tumor can grow but will not spread. Although breast cancer most commonly spreads to nearby lymph nodes, it can also spread further through the body to areas such as the bones, lungs, liver, and brain. This is called metastatic or stage IV breast cancer and is the most advanced type of breast cancer. However, the involvement of lymph nodes alone is generally not stage IV breast cancer.

The most common cause of hereditary breast cancer is an inherited mutation in the BRCA1 or BRCA2 gene. In normal cells, these genes help make proteins that repair damaged DNA. Mutated versions of these genes can lead to abnormal cell growth, which can lead to cancer. If you have inherited a mutated copy of either genos from a parent, you have a higher risk of breast cancer.

BRCA1 AND BRCA2

BRCA1 (Breast Cancer susceptibility gene 1) and BRCA2 are tumor suppressor genes, the mutant phenotypes of which predispose to breast and ovarian cancers. Intensive research has shown that BRCA proteins are involved in a multitude of pivotal cellular processes [2]. In 1994, the breast-cancer susceptibility gene, BRCA1, was identified by positional cloning; subsequently, this gene has been the subject of intensive research efforts [3]. BRCA1 is composed of 22 coding exons distributed over 100kb of genomic DNA. This gene encodes 1863 amino acids, and more than 200 different germline mutations associated with cancer susceptibility have been identified. Many disease-predisposing alleles of BRCA1 have loss-of function mutations, the majority of which result in premature truncation of the protein. Because only 45% of familial breast cancers showed evidence of linkage to BRCA1.

The search for a second breast cancer susceptibility gene continued. In 1995, the BRCA2 gene was identified at chromosome 13q12.3Human BRCA2 protein is composed of 3418 amino acids (384 kDa). The N-terminal part of the BRCA2 protein contains a transcriptional activation domain (aa 18-105). BRCA2 exon 11 encodes eight conserved motifs termed BRC repeats. Each of these repeats is composed of about 30 residues. A DNA-binding domain has been located in the C-terminal region of the BRCA2 protein (aa 2478-3185). It is composed of a conserved helical domain and three OB folds. Two nuclear localization signals (NLS) have been identified in the C-terminal region of BRCA2.Mutations in BRCA1 and BRCA2 are not simply associated with an elevated risk of breast cancer. Mutation carriers also have increased susceptibility to ovarian, pancreatic, prostatic, and male breast cancers. Other associations may be revealed as more epidemiological information becomes available. Surprisingly, despite the inherited predisposition to cancer associated with BRCA1 and BRCA2, somatic disease-causing mutations in either of these genes are extremely rare in sporadic breast cancers. Over the last 10 years, much has been learned about the structures, functions, and unique features of BRCA gene products. In particular, research into the functions of BRCA proteins has revealed that BRCA proteins interact with a number of regulatory proteins (Table 1).

Table 1: Secondary metabolites are present in different forms in Cordia myxa with various properties.




  
    Component
    Plant uses
    Medicinal uses
    References
  
  
    Alkaloids
    Acts as plant growth regulator and used as insecticides
    Acts as antiseptic, drug, antimalarial, antiasthma, anticancer,    analgesic, antimicrobial
    [4]
  
  
    flavonoids
    Involved in floral pigmentation, symbiotic nitrogen, physiological    regulators as well as cell cycle inhibitor
    Used as anti-inflammatory, anti-allergic, antimicrobial, anticancer,    and antidiarrheal bioactivities
    [5,6] 
  
  
    Carotenoids
    Used as perfumes, provide colour to the plants
    Involved in the reduction of risk of lung cancer
    [7]
  
  
    Stigmasterol
    Acts as a precursor
    Involved in the production of the semisynthetic progesterone, and    antioxidants
    [8]
  
  
    Amino acids
    Plant growth cancer, involved in the protein synthesis, increases the    physiological activity
    Artificial sweetener, regulates, metabolic pathway and gene    expressions, cell signalling & oxidative responses
    [9]
  
  
    Beta-sitosterol
    Used as a plant sterol
    Used to reduce the blood cholesterol & benign prostatic    hyperplasia
    [10]
  
  
    Ascorbic acid
    Important role in cell division & protein modification
    Maintain the health of cartilage, skin, teeth, and blood vessels, also    used for treating tumor and scurvy.
    [11]



Table 1: Secondary metabolites are present in different forms in Cordia myxa with various properties.

Methodology

Collection of leaves

The fresh leaves of Cordia myxa were collected in the month of July from sirwar taluk, Raichur district, Karnataka. The leaves were washed with sterile distilled water to remove the dust particles and dried using hot air oven.

Preparation of Cordia myxa plant extract

Sample Collection: The plant leaves of Cordia myxa were collected from sirwar taluk, Raichur district, Karnataka.

Soxhlet extraction: The plant leaves of Cordia myxa were washed to remove dust particles followed by drying in hot air oven at 50°C overnight. Dried leaves were then grinded to make a fine powder. The dry extract of plant leaves was subjected to Soxhlet extraction, which is used for the continuous extraction of plant extract with high efficacy. Solvent used in extraction was methanol. Extraction was carried out at 60°C for 5-6 hrs. Methanolic extract thus obtained poured in dry petri dishes and subjected to drying in hot air oven for overnight at 50°C. After drying, with the help of tooth pick, methanolic extract from petri dishes was scratched and collected in vials.

Leaves extracts preparation

In Methanol Solution: The air-dried fresh leaves of Cordia myxa was collected and pounded into a powder using pestle and mortar. 15gm of the leaf sample was mixed in 200 mL methanol and allowed to stand for 1 hour before filtering with the help of Whatman filter paper. The filtrate was overnight dried in hot air oven (40°C). The extraction yielded 6g and standard extracts obtained were kept in a refrigerator at 4°C for antibacterial activity analysis [12].

Phytochemical screening

The Phytochemical analysis for the identification of the secondary metabolites such as Flavonoids, Tannins, Saponins, Alkaloids, Proteins, and Carbohydrates were performed using the standard process by Sofowora (1993), Trease and Evans (1972) and Odebiyi and Sofowara (1979).

Antimicrobial activity

Bacterial culture preparation: To perform the antibacterial activity of given plant extract, bacterial culture of Staphylococcus aureus was obtained from MTCC (Strain number: -737). By Streak plate method, pure colonies of S. aureus were obtained. To obtain pure culture, sterile culture conditions were maintained by sterilizing every reagent and materials twice. Overnight grown culture of given bacteria was obtained in order to perform antibacterial activity.

Antibacterial assay of plant extract by Disc diffusion method/ Kirby-Bauer test: Kirby-Bauer test or disc diffusion method is used for determination of effectiveness of certain antibiotic by inhibiting the growth of particular bacteria. For the procedure of antibacterial activity, MH agar media was prepared and followed by sterilization. For sterilization purpose, various materials including petri dishes, micropipette tips, cotton swab, well borer, etc were sterilized. Sterile media was poured in petri dishes. All the procedure of antibacterial activity was performed in sterile environment of Laminar air flow cabinet. After solidification of media plates, bacterial culture of S. aureus MTCC Number 737 was spread on media plates with the help of cotton swab. It was followed by puncturing wells in media plate with the help of well borer. Afterwards, methanolic extract of Cordia myxa were added in respective wells along with positive control and negative control which were Methanol and distilled water respectively. Two media plates were taken, among which one is subjected to well boring of 4 wells and other is subjected to bore 3 wells. In petri plate containing 4 wells, plant extract was added in aliquotes of 100, 75, 50, 25 microlitre and final volume was made up to 100 microlitre by adding Methanol. In other plate containing 3 wells, 10 microlitre of pant extract was added and final volume was made up to 100 microlitre by Methanol. Rest of the wells was subjected to the addition of 100 microlitres of Methanol and distilled water respectively. In the remaining position of other plate, one disc (15mm) of standard Erythromycin antibiotic was incorporated onto the media (Table 2).

Table 2: Quantitative analysis of reagents used in antibacterial assay method.




  
    Well No.
    Volume of plant    extract (mL)
    Volume of Methanol    (mL)
    Volume of distilled    water (mL)
  
  
    1
    100
    -
    -
  
  
    2
    75
    25
    -
  
  
    3
    50
    50
    -
  
  
    4
    25
    75
    -
  
  
    5
    10
    90
    -
  
  
    6
    -
    100
    -
  
  
    7
    -
    -
    100
  
  
    Erythromycin disc    (15mm)
    -
    -
    -



Table 2: Quantitative analysis of reagents used in antibacterial assay method.

After addition of all the reagents, plates were incubated in incubator for 24hrs at 37°C.

Disease selection

Breast cancer was chosen as the target disease to investigate Cordia myxa’s anti-cancer effects. The BRCA 1 and BRCA 2 are the reason for spectra of the breast cancer.

Ligand preparation

A total of 18 ligands were selected for the present study from the plant Cordia myxa. For all the selected ligands Structure data format (SDF) files were downloaded using the PubChem database [13]. These were analyzed using Marvin view which is an advanced chemical viewer for 2D or 3D structures of chemicals and their related information. The canonical SMILES IDs of all the selected ligands were noted and estimation of the physico-chemical properties of these ligands was done using PubChem database [14]. Following this, using PyMol software the selected ligands were converted into 3D structure for the further use.

Target selection and prediction

PubChem database (https://pubchem.ncbi.nlm.nih.gov) and Protein Data Bank (www.rcsb.org/pdb) were used for retrieving the structures of the respective human protein receptors BRCA2 and BRCA1 [15]. Furthermore, the criteria for the target structure were investigated using Protein Data Bank analysis and BLAST (https:// blast.ncbi.nlm.nih.gov). Using Discovery Studio Visualizer, the ions, ligands and water molecules were removed from the protein molecules prior to molecular docking analysis [16]. Later, the hydrogen atoms were added to the protein receptors subsequently.

Homology modelling of BRCA1 and BRCA2

A protein’s three-dimensional structure contains crucial knowledge for understanding its biochemical mechanism and molecular interaction properties. The aim of homology modelling is to create 3D protein structure models from experimentally shown configurations of similar family members in the form of templates. Homology modelling is probably the most effective computational tool for generating stable structural models, and it is used in a wide range of biological applications. Many implementations use homology models, such as modelling site-directed mutagenesis tests, virtual screening and rationalizing the sequence variations results [17].

Since the 3D structure of the target proteins were not available in the Protein Data Bank, the protein sequences of BRCA2 and BRCA1 were retrieved from the NCBI databases for purpose of structure prediction. Further, the targeting of the templates was performed using a homology search through NCBI and PubChem databases. Above 90% of homology for the selected template with respect to BRCA2 and BRCA1 was observed in the terms of percent identification. This was done using the tool SWISS-MODEL (http://swissmodel.expasy. org) ]which is an automated homology modelling server of protein structures [18] which was the first automatic modelling server to be released with free access [19]. Thus, the structures obtained can be visualized using PyMOL software [20].

Drug likeness analysis

Lipinski’s rule of five or RO5 (rule of five) is a key rule for the estimation of drug likeness or to evaluate the presence of any pharmacological or biological activities of a chemical compound [21]. The Lipinski rule, that is mostly misapplied and misinterpreted, is actually developed to help in oral bioavailable drug development and was not intended for the small molecule drug development. This rule is considered as a crucial part in the insilico pre-clinical analysis where the ligands need to satisfy all the parameters included in this rule.

The five chief physiochemical parameters of the rule of five are I) Molecular weight of the compound should be lower than 500 Daltons, II) Lipophilicity have to be lower than 5 which is generally denoted as logP (the logarithm of the partition coefficient between 1-octanol and water) is signified as a quantity, III) The number of groups in the compound capable of donating hydrogen atoms to hydrogen bonds (usually the sum of amine and hydroxyl groups in a drug molecule) should be below 5, and IV) The number of groups that can accept hydrogen atoms to form hydrogen bonds is must be less than 10.

Admet analysis

ADME stands for absorption, distribution, metabolism, and excretion in pharmacology and pharmacokinetics describes the pharmaceutical compound disposition in an organism. The four parameters affect the levels of drug [22] and the rate of susceptibility of drug to tissues, and hence this exhibits impact on the compound’s performance as well as pharmacological property as a drug [23]. ADME addresses the pharmacokinetic problems that determine when a drug molecule reaches the target protein in the body and the period it remains in the bloodstream. A drug of high-quality must not only be therapeutic target effective, but should also exhibit therapeutic dosage with acceptable ADMET parameters [24].

SwissADME (http://www.swissadme.ch) was used for the ADMET analysis which is a web platform that provides unlimited access to a swift and accurate predictive model for drug-likeness, medicinal chemistry friendliness, physicochemical properties, and pharmacokinetics. Besides this, it also includes techniques such as the iLOGP, Bioavailability Radar, and BOILED-Egg. SwissADME is also a login-free web platform that provides user-friendly interface enabling easy and effective interpretation of the results, which can also be used by non-expert users in CADD. It calculates the key pharmacokinetic, drug likeness and associated parameters for more than one molecule [25].

Active binding site analysis

The molecular docking process generally starts with determination of active binding site of the selected proteins [26] which indicates the particular proteins restricted region [27]. In virtual screening, prediction of ligand binding sites on the surface of proteins with the aid of swift, accurate and automated tool is a major challenge [28]. Recognition of active binding sites was identified using Biovia discovery tool. In this study blind docking method was performed between target proteins and selected ligands.

Molecular docking analyses

An important step in virtual screening is estimation and finding the correct target for the successful docking. Using the available threedimensional structure of target proteins, application of appropriate docking tool for the search of best binding site of ligand and target protein can be performed [29]. Several docking tools are available for the docking-based virtual screening (DBVS). In this study, PyRx (https://pyrx.sourceforge.io/) which is an open-access and userfriendly tool is being used. PyRx includes various other software’s like open babel, AutoDock and AutoDock vina [30]. The analysis of the docked proteins will be done once the docking is completed [31]. Thus, after the docking the results will be saved and based on the dock score function the ranking is done [32].

Results

Phytochemicals test

See Table 3.

Table 3: Following table gives brief idea about phytochemical screening of Cordia myxa.




  
    Biochemical Test
    Observation
    Result
  
  
    Mayer’s test 
    Yellow coloured precipitate
    +
  
  
    Fehling’s test 
    Red coloured precipitate
    +
  
  
    Tannin’s test
    Bluish black coloured precipitate
    +
  
  
    Xanthoproteic test 
    Yellow coloured precipitate
    +
  
  
    Ferric chloride test 
    No bluish black coloured precipitate
    -
  
  
    Saponin’s test 
    Foam formation
    +



Table 3: Following table gives brief idea about phytochemical screening of
Cordia myxa.

Biochemical assay of leaf extract of Cordia myxa

See Figure 1-6.

Figure 1: Mayer’s test.

    
    
    Figure 1: Mayer’s test.

Figure 2: Fehling’s test.

    
    
    Figure 2: Fehling’s test.

Figure 3: Tannin’s test.

    
    
    Figure 3: Tannin’s test.

Figure 4: Xanthoproteic test.

    
    
    Figure 4: Xanthoproteic test.

Figure 5: Ferric chloride test.

    
    
    Figure 5: Ferric chloride test.

Figure 6: Saponin’s test.

    
    
    Figure 6: Saponin’s test.

Organism

Among different pathogens, Staphylococcus aureus is a potential microscopic organism causing diseases. There are different antimicrobial agents which are utilized to treat Staphylococcus aureus infectious diseases. Among them, we have concentrated on Erythromycin. We performed the antibacterial activity of plant extract; Cordia myxa, in correlation with Standard Erythromycin disc (15mm) (Figure 7).

Figure 7: Preparation of Methanolic extract of Cordia myxa.

    
    
    Figure 7: Preparation of Methanolic extract of Cordia myxa.

We saw that the given plant extricate was displaying antibacterial action against S. aureus as it was indicating zone of restraint or zone of inhibition around the well on MH (Mueller Hinton) agar plate. We did investigation of crude plant concentrate and erythromycin so as to explain how much compelling a plant concentrate could be in counteraction of diseases brought about by S. aureus. Results were demonstrated that well containing plant extricate was indicating zone of inhibition of diameter 24.33 mm in examination with standard Erythromycin plate which was demonstrating 29 mm of zone of inhibition (Figure 8 and 9).

Figure 8: Zone of inhibition on MH agar plate; Well no.1) 100ul extract, 2) 75ul extract+25ul methanol, 3) 50ul extract+50ul methanol, 4) 25ul extract+75ul methanol, 5) 10ul extract+ 90ul methanol, 6) 100ul methanol, 7) 100ul d/w, 8) No well; Erythromycin disc (15mm).

    
    
    Figure 8: Zone of inhibition on MH agar plate; Well no.1) 100ul extract, 2) 75ul
extract+25ul methanol, 3) 50ul extract+50ul methanol, 4) 25ul extract+75ul
methanol, 5) 10ul extract+ 90ul methanol, 6) 100ul methanol, 7) 100ul d/w, 8)
No well; Erythromycin disc (15mm).

Figure 9: Comparative analysis of Cordia myxa plant extract and Standard Erythromycin disc with respect to diameter of zone of inhibition.

    
    
    Figure 9: Comparative analysis of Cordia myxa plant extract and Standard
Erythromycin disc with respect to diameter of zone of inhibition.

In silico analysis

Ligand preparation: For all the selected ligands Structure data format (SDF) files were downloaded using the PubChem database and were analyzed using Marvin view which is an advanced chemical viewer. The energy minimization of all the ligands was done prior to the molecular docking analysis.

Phytochemical structures: 3-dimensional structures of all the ligands were retrieved using PubChem database and were analyzed using Marvin sketch.

Target selection and prediction: The target proteins were selected using Protein Data Bank. The criteria for the target structures of human protein receptors BRCA1 and BRCA2 were investigated using PDB analysis and BLAST. After the retrieval of 3D structure of the target proteins, Discovery Studio Visualizer tool was used for the removal of ions, ligands and water molecules from the proteins prior to the docking and addition of polar hydrogen atoms.

Homology modelling of BRCA1 and BRCA2 proteins: The target proteins modelling was done using SWISS-MODEL server and the structures obtained were visualized using PyMOL software (Figure 10 and 11).

Figure 10: 3D structure of BRCA1 protein.

    
    
    Figure 10: 3D structure of BRCA1 protein.

Figure 11: 3D structure of BRCA2 protein.

    
    
    Figure 11: 3D structure of BRCA2 protein.

Drug likeness analysis

See Table 4.

Table 4: Lipinski filter analysis.




  
    Ligands 
    Molecular formula 
    Hydrogen Bond Acceptor 
    Hydrogen Bond Donor 
    cLogP 
    Molar Refractivity 
  
  
    Cordiagenol 
    C30H50O4 
    4
    4 
    4.73 
    138.37 
  
  
    Cordiagenin 
    C30H50O4
    6 
    6 
    3.17 
    140.69 
  
  
    Cordianol 
    C30H50O4
    5 
    5 
    4 
    139.53 
  
  
    Catechol 
    C6H6O2 
    2 
    2 
    0.97 
    30.49 
  
  
    Tetradecanoic acid 
    C14H28O2 
    2 
    1 
    4.45 
    71.18 
  
  
    Squalene 
    C30H50 
    0
    0 
    9.38 
    143.48 
  
  
    Nerolidol 
    C15H26O 
    1
    1 
    4.19 
    74 
  
  
    Stigmasterol 
    C29H48O 
    1
    1 
    6.96 
    132.75 
  
  
    Cedrene V6 
    C15H24 
    0
    0 
    4.38 
    68.52 
  
  
    Lupeol 
    C30H50O 
    1
    1 
    7.31 
    135.14 
  
  
    1-Docosene 
    C22H44 
    0 
    0 
    8.67 
    107.39 
  
  
    Beta amyrin 
    C30H50O 
    1 
    1 
    7.18 
    134.88 
  
  
    Isophytol 
    C20H40O 
    1
    1 
    6.18 
    98.98 
  
  
    Cordiasaponin II 
    C42H70O14 
    14
    10 
    0.93 
    203.13 
  
  
    Gymnemic acid II 
    C43H68O14 
    7
    14 
    2.97 
    207.58 
  
  
    Gymnemic acid III 
    C41H66O13 
    8
    13 
    2.64 
    197.84 
  
  
    Gymnemic acid IV 
    C41H64O13 
    8
    13 
    2.55 
    197.37



Table 4: Lipinski filter analysis.

ADMET analysis: The analysis of the ADMET parameters was performed using the tool SwissADME and results were estimated for various parameters. The results for this analysis were shown in the Table 5 and 6.

Table 5: Physico-chemical properties of the ligands.




  
    Ligands 
    Molecular formula 
    Molecular weight 
    Heavy atom count 
    TPSA 
  
  
    Allointoin 
    C4H6N4O3 
    158.121g/mol
    34 
    80.9 Å2 
  
  
    a-amyrin 
    C30H50O 
    426.7g/mol 
    36 
    121 Å2 
  
  
    Apigenin 
     C15H10O5 
    270g/mol 
    35 
    101 Å2 
  
  
    Β-sitosterol glycoside 
    C35H60O6 
    576.85g/mol 
    8 
    40.5 Å2 
  
  
    Hentricontanol 
    C31H64 
    436.85g/mol 
    16 
    37.3 Å2 
  
  
    Hesperidine 
    C28H34O15 
    610.1898g/mol
    30 
    0 Å2 
  
  
    Octasanol 
    C28H58O 
    410g/mol
    16 
    20.2 Å2 
  
  
    Lupeol 
    C30H50O 
    426.72g/mol
    30 
    20.2 Å2 
  
  
    Robonine 
    C17H26O10 
    390.4g/mol
    15 
    0 Å2 
  
  
    Rutin 
     C27H30O16 
    610.517g/mol
    31 
    20.2 Å2 
  
  
    Betulin 
    C30H50O2 
    442.72g/mol 
    22 
    0 Å2 
  
  
    Caffeic acid 
    C9H8O4 
    180.16g/mol 
    31 
    20.2 Å2 
  
  
    Carotenoid 
    C40H56 
    536.9g/mol
    21 
    20.2 Å2 
  
  
    Chlorogenic acid 
    C16H18O9 
    354.31g/mol
    56 
    239 Å2 
  
  
    Datiscoside 
    C38H54O12 
    702.8g/mol
    57 
    230 Å2 
  
  
    Dihydrorobenetin 
    C19H29NO3 
    319.4g/mol
    54 
    224 Å2 
  
  
    Β-sitosterol 
    C29H50O 
    414.71g/mol
    54 
    224 Å2



Table 5: Physico-chemical properties of the ligands.

Table 6: Admesar analysis.




  
    Ligands 
    Blood brain barrier  
    GI absorption 
    Permeability glycoprotein Substrate 
    Log S (scale Insoluble < -10<Poorly<-6< Moderately 

      <- 4<Soluble<-2Very<0< Highly) [Water solubility]
  
  
    Allointoin 
    No 
    High
    Yes 
    -5.20 (Moderately soluble) 
  
  
    a-amyrin 
    No 
    High 
    Yes 
    -3.55 (soluble) 
  
  
    Apigenin 
    No 
    High 
    Yes 
    -4.37 (Moderately soluble) 
  
  
    Β-sitosterol glycoside 
    Yes 
    High 
    No 
    -1.18 (soluble) 
  
  
    Hentricontanol 
    Yes 
    High 
    No 
    -4.51 (Moderately soluble) 
  
  
    Hesperidine 
    No 
    Low
    No 
    -7.48 (poorly soluble) 
  
  
    Octasanol 
    Yes 
    High
    No 
    -3.15 (soluble) 
  
  
    Lupeol 
    No 
    Low
    No 
    -5.47 (Moderately soluble) 
  
  
    Robonine 
    No 
    Low
    No 
    -4.09 (Moderately soluble) 
  
  
    Rutin 
    No 
    Low
    No 
    -6.74 (poorly soluble) 
  
  
    Betulin 
    No 
    Low 
    Yes 
    -8.39 (poorly soluble) 
  
  
    Caffeic acid 
    No 
    Low 
    Yes 
    -7.16 (poorly soluble) 
  
  
    Carotenoid 
    No 
    Low
    Yes 
    -5.5 (Moderately soluble) 
  
  
    Chlorogenic acid 
    No 
    Low
    Yes 
    -1.54 (soluble) 
  
  
    Datiscoside 
    No 
    Low
    Yes 
    -3.21 (soluble) 
  
  
    Dihydrorobenetin 
    No 
    Low
    Yes 
    -2.62 (soluble) 
  
  
    Β-sitosterol 
    No 
    Low
    Yes 
    -2.26 (soluble)



Table 6: Admesar analysis.

Molecular docking analysis

The energy minimization of all the selected ligands was done and was converted into pdbqt file format. The binding affinities of all the ligands were estimated using PyRx tool (Table 7). After the docking process, the docked structures were saved and visualized using the Biovia Discovery Studio tool. The 2D diagram of receptorligand interactions were analyzed of all the docked structures (Figure 12a-12c).

Table 7: Binding affinity of selected phytochemicals for BRCA1 and BRCA2 proteins.




  
    Phytochemicals 
    Binding affinity (Kcal/mol) for BRCA2 
    Binding affinity (Kcal/mol) for BRCA1 
  
  
    Allointoin 
    -8.4 
    -9.3 
  
  
    a-amyrin 
    -9.1 
    -8.2 
  
  
    Apigenin 
    -8.4 
    -8.9 
  
  
    Β-sitosterol glycoside 
    -5.2 
    -5.5 
  
  
    Hentricontanol 
    -5.2 
    -6.1 
  
  
    Hesperidine 
    -7.4 
    -8.5 
  
  
    Octasanol 
    -6.6 
    -6.3 
  
  
    Lupeol 
    -9.7 
    -9.8 
  
  
    Robonine 
    -7.7 
    -7.6 
  
  
    Rutin 
    -8.9 
    -10.3 
  
  
    Betulin 
    -6.2 
    -5.9 
  
  
    Caffeic acid 
    -8.9 
    -9.9 
  
  
    Carotenoid 
    -6.3 
    -7 
  
  
    Chlorogenic acid 
    -9.3 
    -10.6 
  
  
    Datiscoside 
    -9.2 
    -9.3 
  
  
    Dihydrorobenetin 
    -15.4 
    -15.7 
  
  
    Β-sitosterol 
    -15.2 
    -15.7



Table 7: Binding affinity of selected phytochemicals for BRCA1 and BRCA2
proteins.

Figure 12a: Docked Phytochemicals with BRCA2 Protein. Docked and 2D diagram of receptor-ligand interactions

    
    
    Figure 12a: Docked Phytochemicals with BRCA2 Protein. Docked and 2D diagram of receptor-ligand interactions

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Citation: Ambresh V and Gowda P. In Vitro & In Silico Studies of Cordia myxa Phytochemicals in Contrast to Breast Cancer: A Bioinformatics Approach. Austin J Proteomics Bioinform & Genomics. 2022; 7(1): 1032.

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In Vitro & In Silico Studies of Cordia myxa Phytochemicals in Contrast to Breast Cancer: A Bioinformatics Approach

Abstract

Introduction

Morphology

Breast cancer (Bc)

BRCA1 AND BRCA2

Methodology

Collection of leaves

Preparation of Cordia myxa plant extract

Leaves extracts preparation

Phytochemical screening

Antimicrobial activity

Disease selection

Ligand preparation

Target selection and prediction

Homology modelling of BRCA1 and BRCA2

Drug likeness analysis

Admet analysis

Active binding site analysis

Molecular docking analyses

Results

Phytochemicals test

Biochemical assay of leaf extract of Cordia myxa

Organism

In silico analysis

Drug likeness analysis

Molecular docking analysis

Discussion

Conclusion

Acknowledgement

References