Journal of Biomolecular Structure and Dynamics

ISSN: (Print) (Online) Journal homepage: https://www.tandfonline.com/loi/tbsd20

Dynamacophore model for breast cancer estrogen

receptor alpha as an effective lead generation
screening technique

Dhivya Shanmugarajan, Anagha Biju, Dona Sibi, Rona Sibi, Maria Shaji &
Charles David

To cite this article: Dhivya Shanmugarajan, Anagha Biju, Dona Sibi, Rona Sibi, Maria Shaji
& Charles David (2023): Dynamacophore model for breast cancer estrogen receptor alpha
as an effective lead generation screening technique, Journal of Biomolecular Structure and
Dynamics, DOI: 10.1080/07391102.2023.2203245

To link to this article: https://doi.org/10.1080/07391102.2023.2203245

Published online: 08 May 2023.

Submit your article to this journal

Article views: 22

View related articles

View Crossmark data

Full Terms & Conditions of access and use can be found at

https://www.tandfonline.com/action/journalInformation?journalCode=tbsd20
JOURNAL OF BIOMOLECULAR STRUCTURE AND DYNAMICS
https://doi.org/10.1080/07391102.2023.2203245

Dynamacophore model for breast cancer estrogen receptor alpha as an

effective lead generation screening technique
Dhivya Shanmugarajan, Anagha Biju, Dona Sibi, Rona Sibi, Maria Shaji and Charles David
Department of Biotechnology, Vignan’s Foundation for Science, Technology and Research (Deemed to be University), Guntur, Andhra
Pradesh, India
Communicated by Ramaswamy H. Sarma

ABSTRACT ARTICLE HISTORY

Regardless to overwhelming quantum of cancer research worldwide, there are few drugs on the mar- Received 3 October 2022
ket to treat disease conditions. This is owing to multiple process inferences of drug targets in inte- Accepted 11 January 2023
grated pathways for invasion, growth, and metastasis. Over the past years, the death rate due to
KEYWORDS
breast cancer has been increasing, that set the stage for improved better treatment. Therefore, there
Breast cancer; Estrogen
is a persistent and vital demand for innovative development of drugs to treat breast cancer. Many receptor-a; dynamacophore;
studies have reported that more than 60% of breast cancers are Estrogen receptor-a (ERa)-positive molecular docking; MM-
tumours and a key transcription factor, Estrogen receptor-a (ERa) was believed to promote prolifer- GBSA
ation of breast cancer cells. In this study, 150 ns of molecular dynamics was performed for protein-lig-
and complex to retrieve the potential stable conformations. The most populated dynamics cluster of
4-Hydroxytamoxifen intact with active site amino acid was selected to generate dynamacophore
model (dynamic pharmacophore). Further, internal model validation with AU-ROC values
0.93 indi-
cate the best model to screen library. The refined hits are funnelled in pharmacokinetics/dynamics,
CDOCKER molecular docking, MM-GBSA and density functional theory to identify the promising ERa
ligand candidates.

1. Introduction
Globally, one of the most common cancers and the major
cause of death in women is due to breast cancer (Kim et al.,
2022). Metastatic breast cancer is a fatal form of malignancy
that causes substantial morbidity and fatality rates (Kulkarni
& Rawtani, 2019). Compared to high-income countries, breast
cancer death rates are greater in low- and middle-income
countries (LMIC). Early identification through extensive mam-
mographic screening and accessibility to efficient adjuvant
systemic therapy for women in high-income countries has
been credited as the causes of the variations in survival rates
(Ferlay et al., 2015). There are 0.8–1% cases of breast cancer
in men as well, indicates that disease consequences is not
only for women (Fentiman et al., 2006).
As per the recent worldwide data on cancer published by
the World Health Organization, of all new cases of cancer in
2020, 11.7% cases were reported to be breast cancer (World
Health Organization: A Global report on breast cancer, 2020).
According to estimates, 627,000 women died from breast
cancer in 2018. For the year 2019, in United States alone, it
was predicted to encounter 271,270 new cases of breast can-
cer, of which 42,260 deaths were anticipated (DeSantis et al.,
2019).
Four distinct molecular sub-types of breast cancer are
based on the patterns of gene expression in breast cancer
cells, including Luminal A, which lacks the human epidermal
growth factor receptor 2 (HER2) and is positive for either the
estrogen receptor (ER) or the progesterone receptor (PR).
Luminal A-type breast tumours account for 73% of cases.
Luminal B, which is HER2 either positive or negative and ER
and/or PR positive (Anderson et al., 2014). About 70% of
breast cancer is estrogen receptor-positive (ERþ) and for the
first-time breast cancer has become the most prevalent can-
cer surpassing lung cancer (Sung et al., 2021). As cyclin D1,
Myc, B-cell lymphoma 2 (Bcl-2) and vascular endothelial
growth factor (VEGF) are all significantly involved in the cell
cycle, cell survival, and promotion of angiogenesis, the estro-
gen receptor (ER) plays a crucial role in the carcinogenesis of
breast cancer (Eeckhoute et al., 2006). Tamoxifen competes
with ER for binding sites, acting as a selective estrogen
receptor modulator (SERM) to inhibit the ER transcriptional
activity (Ali et al., 2016). Breast cancer is treated with radi-
ation therapy, chemotherapy, endocrine therapy, adjunct
therapy (which includes both radiation therapy and chemo-
therapy), and ligand-mediated therapy (Nounou et al., 2015).
Endocrine therapy (anti-estrogen) is the most ideal course
of treatment for breast cancer because it has been associ-
ated with low morbidity and mortality (Lumachi et al., 2011).
For more than 20 years, tamoxifen has been the drug of
choice for treating advanced or metastatic breast cancer
with an anti-estrogen (Memisoglu-Bilensoy et al., 2005). A
nonsteroidal tri-phenyl ethylene derivative known as

CONTACT Charles David microroses@gmail.com, arcda88@gmail.com

ß 2023 Informa UK Limited, trading as Taylor & Francis Group
2 D. SHANMUGARAJAN ET AL.
Tamoxifen, it is consider as the selective estrogen receptor
modulator (Hard, 1993).
Estrogen receptor-positive breast cancer can be treated
using the drug Tamoxifen, it is one of the triphenylethylene
derivative. It is the main hormone therapy for breast cancer
in both the adjuvant and metastatic situation (Braal et al.,
2020; Jordan, 2004; Morad & Cabot, 2015). It continues to be
a key therapeutic strategy for extending both recurrence-free
and overall survival (Early Breast Cancer Trialists’
Collaborative Group, 1998). Tamoxifen prevents the growth
and division of malignant cells by halting them in the G0
and G1 phases of the cell cycle (Zeng & Yee, 2007).
The active metabolite present in the anticancer drug
Tamoxifen is Endoxifen. In patients treated with similar doses
of Tamoxifen, Endoxifen has a huge inter-individual variabil-
ity (Braal et al., 2020). Although a 30% reduction in breast
cancer mortality is due to the use of Tamoxifen, resistance to
this drug is one of the fundamental issues for its clinical use
in ER-positive breast cancer. Therefore, it is indeed necessary
to overcome the resistance in breast cancer by adopting a
new therapeutic methodology to magnify Tamoxifen’s effi-
cacy (Kim et al., 2022).
Estrogen receptor (ER) gene mutation or deletion, varia-
tions in nuclear receptor coactivator expression, abnormal
cell cycle regulation, and abnormalities in cell signalling
pathways are some of the multiple factors associated with
the mechanism underneath Tamoxifen resistance, which are
overly complicated (Rani et al., 2019). Nevertheless, the
mechanism that directs this drug resistance is yet to be dis-
closed. Also, there has not been any discovery of drugs that
can efficaciously alter this resistance. The efficacy of endo-
crine therapy can be improved and a basis for better diagno-
sis and treatment can be provided by investigating the
mechanism of drug resistance, discovering drug resistance
genes, and identifying resistance reversing drugs (Zhao et al.,
2022). The main objective of this current study is to identify
the potential lead compounds using the dynamics-based
pharmacophore model.
2. Materials and methods
2.1. Nanoscale molecular dynamics and simulation
All atomic dynamics were started by applying the CHARMm
force field for apoprotein and protein-ligand bound complex
using nanoscale molecular dynamics (NAMD) 2.3 in BIOVIA
Discovery Studio (Phillips et al., 2005). Before running this
protocol, the input system must be minimized and equili-
brated, and standard dynamics cascade protocol was used.
The complex system of apoprotein without bound ligand
and protein-ligand complex was explicitly solvated in peri-
odic boundary condition with the TIP3P water model in the
ortho-rhombic box with distance of 3.0 Å. The appropriation
and the protein-ligand complex systems are neutralized by
default salt concentration of 0.145 (mol/l) of the explicitly
solvated system. The entire system is minimized conse-
quently with 500 cycles using the steepest descent and con-
jugate gradient method. The two different systems were
further heated gradually from 50 K to 300 K. Subsequently,
equilibration for 1000 ps was subjected for further stabiliza-
tion of complexes to achieve degrees of freedom. Finally, a
total of 150 ns Molecular Dynamics (MD) simulations were
performed for the apoprotein and protein-ligand complex,
where conformation coordinates were saved for 10 ps inter-
val for both biological systems to yield 15,000 frames. The
dynamic pressure and temperature were controlled by apply-
ing Langevin Piston method (Parrinello & Rahman, 1981) to
sustain 1.01325 bar and Langevin Dynamics thermostat (Bussi
et al., 2007) respectively. Further, the multiple time step algo-
rithm was integrated to study the long- and short-range
forces using impulse r-RESPA method within a cut-off inter-
action distance of 14–12 Å. Further, the same parameters are
used for dynamics simulation of top three molecules for
50,000 ps or 50 ns to obtain 5000 frames saved at 10 ps inter-
val. The output results in terms of time dependent parame-
ters were interpreted (Dhivya et al., 2018) and the graphs
were plotted using GraphPad Prism 5.0 v.
2.2. Clustering the trajectory conformations of estrogen
receptor alpha
The final output of molecular dynamics simulation ends up
with structural trajectories (s(tj)) contains atomic positions as
a function of time (f(t)). Each structural trajectories (s(tj)) has
its own 3D coordinates as a function of time, the enormous
number of frames with energy variations possess different
structural conformations during the molecular simulations.
However, studying all structural trajectories (s(tj)) is painstak-
ing task, so the frames are needed to be funnelled to eligible
number. This can be achieved by using clustering method, a
most powerful technique to reduce the frames into manage-
able size. However, 2500 potential stable frames were used
for clustering analysis.
f ðtÞ ¼ @ ðtjÞ and @ðEÞ
where, t – time, s – structure, tj – trajectories, E – energy; @
– change, f-function
A clustering algorithm applied structural trajectories (s(tj))
divides the structural frames into separate group of sets
named conformations. The data points recorded for each
frame in one cluster is ideally more like each other than that
to compare with other clusters frames (Peng et al., 2018).
The mutual root mean square deviation (RMSD) is most fre-
quently employed clustering algorithm samples all conforma-
tions sampled out from the simulation. Always, the structure
with highest population of neighbours in each of the frames
in the cluster is called as the representative clusters (Daura
et al., 1998).
Initially, the pairwise RMSD was computed for atom-pair
distances to outline the distance between two structural
frames, and then the clustering linkage approach was imple-
mented. Followed by RMSD technique was used to compute
the conformational changes in output molecular dynamics
trajectory. RMSD was calculated between pairwise conforma-
tions obtained from the 150 ns simulation trajectories of top
2500 potentially stable frames. The results of this clustering
were used to understand the protein–ligand stability

complex to select and generate structure-based pharmaco-
phore model. All 2500 frames were studied using analyse tra-
jectory of simulation tool present in Discovery Studio to
calculate both pairwise and clustering conformations.
2.3. Sampling trajectory for pharmacophore mapping
The RMSD stabilized trajectory of the ligand-protein complex
generated major clusters within linkage distance of 0.47 Å
(Figure 1) among top clusters having the largest number of
trajectories with least RMSD snapshots were sampled for
structure-based pharmacophore modelling and screening.
The root mean square fluctuations of active site amino
acid backbone atom C-alpha were calculated for both ligand
complex and apoprotein. This forms the simulated trajectory
output, because the stability of active site is considered as a
crucial factor to generate the best pharmacophore model.
2.4. Structure based pharmacophore modelling with
validation
The virtual screening of structure-based pharmacophore
model approach search using interaction generation pharma-
cophore (Meslamani et al., 2012) in BIOVIA Discovery Studio.
The Structure-Based Pharmacophore (SBP) modelling of pro-
tein-ligand complex uses surrounding the crystal bound or
docked ligand 3D space of binding site or active site amino
acids residues to generate the chemical space environment
to screen the compounds. Therefore, the generated pharma-
cophore had potential to capture more detailed information
about regions that are available to the ligand. The dynamic
frames generated pharmacophore models were named as
Figure 1. Overall workflow of the dynamacophore modelling study.
JOURNAL OF BIOMOLECULAR STRUCTURE AND DYNAMICS 3
dynamacophore (molecular dynamics frames-based pharma-
cophore model) with validation of the generated pharmaco-
phore models using active/inactive ligands. The Receiver
Operating Characteristic (ROC) curves and the optimal cut-off
values were reported. The pharmacophore model by review-
ing the area of the ROC curve (AUC) was evaluated. A value
closer to 1.0 indicates good selectivity, whereas a value
closer to 0.5 indicates random selection. The cut-off indicates
the best pharmacophore fit value which gives the highest
accuracy as calculated by the following equation:
ðTP þ TNÞ=ðTP þ TN þ FP þ FNÞ
On this context, frames of 4-hydroxytamoxifen in complex
with estrogen receptor alpha conformations that were
obtained from the top three populated representative cluster
were used as the input to upload to generate pharmaco-
phore feature identification and library search.
2.5. Virtual screening, docking and binding energy
MM/GBSA
The compounds from medicinal plant Cardiospermum halicaca-
bum (http://bidd.group/CMAUP/plant.php?plant=NPO21096),
LCMS and Immuno-Oncology Library from Asinex (https://www.
asinex.com/immuno-oncology). As a customized library, scPDB
(7622), MiniMaybridge (2000) compounds were chosen to be
screened by the dynamacophore model. The hits from three
pharmacophore models were populated and refined by the
ADMET and TopKatV. Finally, molecular docking was performed
R
screened compounds with estrogen receptor alpha using
BIOVIA Discovery studio CDOCKER. The compounds with nega-
tive CDOCKER energy and CDOCKER interaction energy with
4 D. SHANMUGARAJAN ET AL.
calculating binding energies using MM/GBSA (molecular
mechanics-generalized born surface area). In the process of vir-
tual screening MM/GBSA or MM/PBSA plays a significant role to
investigate the binding affinity of the compounds (Ercheng
et al., 2019). This technique can be implemented for docked
complex to screen best compound poses, also this method can
be applied to all frames/conformations generated during
molecular dynamics and it is calculated using the below equa-
tion (Poli et al., 2020; Shafi et al., 2021; Shanmugarajan et al.,
2020)
DGbinding ¼ DGcomplex  ðDGprotein þ DGligand Þ  equation 1:
2.6. DFT calculations
Density Functional Theory (DFT) analysis was performed to
determine the electrostatic properties of top three com-
pounds also, to investigate potential drug interactions which
demonstrate suitability (Hiteshi et al., 2019; Maryam et al.,
2020). The frontier orbital energies, i.e. Highest Occupied
Molecular Orbital (HOMO) and Lowest Unoccupied Molecular
Orbital (LUMO) energies were calculated using Becke’s three
parameter (Becke, 1993) exchange functionals (B3LYP) of
Dmol3 package of BIOVIA Discovery studio 2022. The HOMO
and LUMO energy profile describe the electron donor and
acceptor properties of any compounds which provides the
idea on the reactivity of the compounds to their receptors
(Gogoi et al., 2017). In this study, by analysing the HOMO
and LUMO energy the energy gap of top three compounds
and standard (OHT) using the following formula. The overall
workflow of the study is depicted in Figure 2.
EgapðevÞ ¼ ELUMO  EHOMO :
3. Results
The 4-hydroxytamoxifen molecule has three major features
including alkylamino side chain, triphenylethylene and small lipo-
philic chain (Figure 3). The crystal structure of the estrogen
receptor alpha in complex with 4-hydroxytamoxifen was solved
(Heldring et al., 2007) with 2.10 Å resolution. There are more
than 300 crystal structures available for estrogen receptor and

14 proteins in RCSB (Research Collaboratory for Structural
Bioinformatics) protein databank which can bind with drug
Tamoxifen. The crystal structure (2JF9) has the missing loop
region found between residues A: THR460 and A: GLU470 in the
ligand binding domain region also, disordered residues and alter-
native conformers were resolved by protein preparation tools in
BIOVIA Discovery Studio. The final loop-built structure without
any protein residue disorder is used for the further analysis.
3.1. RMSD of structural conformations of estrogen
receptor alpha
The Root mean square deviations (RMSD) of conformations
captured at the intervals of every 10 ps as change in time
were carried out for the apo and 4-hydroxytamoxifen-bound
receptor to understand structural stability. MD trajectory of
the simulations generates 15,000 frames as the output coor-
dinates. The structural divergence and convergence of the
systems from molecular dynamics simulation trajectory and
RMSD plots for both apo and ligand-complex structures were
analysed. The RMSD plots (Å) of estrogen receptor-alpha
apoprotein (black) and 4-hydroxytamoxifen bound complex
(red) for the protein backbone atoms in 150 ns MD simula-
tion is as shown in Figure 4(a) and the ligand conformation
changes were depicted in Figure 4(b). A trajectory tool for
analysis was utilized to study and ensure the stability of the
structure. We investigated the conformational changes of
backbone atom in dynamics simulation trajectory for both
with ligand bound and without ligand bound structures.
In RMSD plot, initially both apo and ligand bound complex
deviations were gradually increases at the beginning of the
dynamics with slight convergence and divergence. After 50 ns
apoprotein plummeted from the trajectory path till
60 ns with
a least deviation of <1 Å. In contrast, 4-hydroxytamoxifen
bound complex plummeted at 70 ns with deviation of 1.9 Å
runs up to 80 ns with slight deviations, after which it had
increased to attain its stability over a time. At the end of the
dynamics, both the complexes travel in parallel with RMSD vari-
ation of <2 Å (apo) and <3 Å (4-hydroxytamoxifen bound com-
plex) to maintain its structural stability with least fluctuations.
Overall, the average RMSD deviations of apoprotein, 4-
hydroxytamoxifen bound complex and 4-hydroxytamoxifen is
1.257 ± 0.3183, 2.198 ± 0.2229 and 1.988 ± 0.2344 respectively.
Moreover, the results of RMSD were compared with previ-
ously performed molecular dynamics of long scale dynamics
deviations which are within 3 Å for 4-hydroxytamoxifen
bound complex concert this study. Also, the RMSD deviations
of 4-hydroxytamoxifen deviations are within 1.5 Å to 2 Å.
3.2. RMSF of estrogen receptor alpha
RMSF plots drawn from the backbone Ca atoms have been ana-
lysed throughout the MD simulation and the atomic fluctuation
of the overall simulations can be used to understand the flexibil-
ity and stability of the different protein residues positioned. The
higher RMSF of the residues indicate more flexibility of the
amino acid and the lower fluctuations indicate the limited move-
ment throughout the MD simulation. The total protein fluctu-
ation observed above 0.5 Å and some of the residues were
showing higher fluctuations. The RMSF range of the active site
amino acid residues interacting with the ligand (4-hydroxytamox-
ifen) fluctuated between 0.572–2.226 Å were as shown in
Figure 4(c).
Overall, a secondary structure turns, and coils show higher
fluctuations than helix region of protein. Apoprotein shows
maximum deviation in the loop region of 415–424 amino
acid residues, more specifically highest fluctuation was
observed for amino acids Met421 and Phe461 with values of
2.3 Å and 2.2. Å. In contrast, helix-turn-coil region (332–352)
of 4-hydroxytamoxifen complex system shows least fluctu-
ation (1.15–0.659 Å) and higher fluctuations at C-terminal res-
idues (481–504) helix-turn-turn secondary structure as
compared to the apo structure. Furthermore, backbone Ca
atoms and side chain of the active site residues interacting

Figure 2. (a) Cluster dendrogram of top 2500 potentially stable frames and similarities between conformations/frames/snapshots, the closely related frames are
grouped closer together in rectangular view. The frames are represented by the terminal nodes of the tree. (b) Circular view of Cluster dendrogram, (c) Unrooted
view of cluster dendrogram.
Figure 3. The structure of 4-hydroxytamoxifen.
with standard drug 4-hydroxytamoxifen and apoprotein
RMSF deviations in Å are shown in Table 1. There was not
much deviation in the active site residues except Met421
which shows elevated RMSF value of above 2 Å and 2.5 Å of
Ca atoms and side chain, respectively. The 4-hydroxytamoxi-
fen RMSF indicate the 2-(dimethylamino) ethan-1-ol aliphatic
side chain had a fluctuation of around 3.083 Å–1.845 Å, the
aromatic trunk triphenylethylene group atomic fluctuation
observed around 0.4 Å–0.5 Å, the small lipophilic tail has
0.6 Å of the fluctuation and hydroxyl group (OH) was moved
with 0.52–0.59 Å atomic fluctuation.
JOURNAL OF BIOMOLECULAR STRUCTURE AND DYNAMICS 5
3.3. Radius of gyration
Radius of gyration is the most crucial parameter to study the
compactness of protein folding during molecular dynamics. The
structural compactness, close pack and stability is more for
alpha þ beta and alpha/beta and lesser for alpha-helix which can
be studied using radius of gyration. The estrogen receptor alpha
receptor secondary structure composition details were obtained
from secondary structure server (https://2struc.cryst.bbk.ac.uk/sub-
mit/) using DSSP method shows that ER-alpha is made up of
60.8% helical and 3.4% beta and 35.8% turns.
The Alpha class with Orthogonal Bundle architecture and
Retinoid X Receptor topology of nuclear receptor ligand-binding
domain, the helical domain is involved in binding and respon-
sible for various biological process in the cell. From Figure 5, it
was observed that apoprotein deviations are less than 17.5 Å
while ligand bound complex shows more structural deviations at
17.5 Å at the beginning of dynamics but overlaps at certain times
and merged slightly after 70 ns dynamics. This indicates that
both apoprotein and ligand bound complex protein structure
attains stability at the end of dynamics (Lobanov et al., 2008).
3.4. Trajectories clustering of potentials stable
snapshots
The 2500 structures were analysed using analyse trajectory
clustering tool to find the close neighbouring clusters. 4-
hydroxytamoxifen interactions with the active site of the pro-
tein with four main functional pharmacophores such as
6 D. SHANMUGARAJAN ET AL.

Figure 4. (a) RMSD deviation of apoprotein and OHT bound estrogen receptor alpha, (b) RMSD deviations of ligand (OHT) during 150 ns molecular dynamics simu-
lation. (c) Root mean square fluctuation of estrogen receptor alpha and yellow colour highlights the fluctuations.

Table 1. Root mean square fluctuation of apoprotein and 4-hydroxytamoxifen active site residues.
Aminoacids Met343 LEU346 THR347 ALA350 ASP351 GLU353 TRP383
Apo 1.491 1.149 0.902 0.629 0.799 0.678 0.621
4-OHT 1.041 0.72 0.759 0.55 0.578 0.804 0.521
Aminoacids LEU384 ARG394 PHE404 MET421 LEU428 GLY521 HIS524
Apo 0.525 0.521 0.788 1.684 0.506 1.001 1.491
4-OHT 0.475 0.519 1.192 1.735 0.491 1.366 1.395

Figure 5. Radius of gyration of Apoprotein and OHT bound estrogen receptor
alpha.
number 348, 1260 and 2175 are 2.90 Å, 3.34 Å, and 3.386 Å,
respectively. In the estrogen receptor alpha crystal structure
(PDB_ID:2JF9), two hydrogen bonds between 4-hydroxyl moi-
ety attached with aromatic trunk triphenylethylene in tam-
oxifen with ARG394 and GLU353. During the 150 ns
simulations, the molecules slightly moved, and the 4-
hydroxyl group lost the hydrogen bond with NH2 moiety of
ARG394 ring. The GLU353 retained its hydrogen bond with
the 4-hydroxytamoxifen OH group throughout the
simulations.

3.5. Structure based pharmacophore modelling of

aromatic parent triphenylethylene scaffold, hydroxyl group,
2-(dimethylamino)ethan-1-ol and small lipophilic tail.
To identify the conserved active site residues and to
understand their interactions with 4-hydroxytamoxifen in the
crystal structure and apoprotein, the largest top three cluster
snapshots were reviewed to elucidate the specific frames
348, 1260 and 2175. The superimposed model of these three
frames is depicted in Figure 6. The RMSD between the initial
protein structure of the apoprotein to different frame num-
bers 348, 1260 and 2175 were 1.994, 1.927, and 2.021 Å,
respectively. Whereas 4-hydroxytamoxifen bound protein
RMSD value from its initial protein frame to specific frame
frames of estrogen receptor alpha
4-hydroxytamoxifen based pharmacophore generation was
performed on the three snapshot representative complexes
from the MD simulations. Pharmacophore models were gen-
erated with statistical validation (Table 2) of from cluster rep-
resentative frames 348, 1260 and 2175 with three features
including hydrogen bond acceptor vector features (pink,
with head-2.2 Å and tail 1.6 Å radius), hydrophobic (cyan,
with 1.6 Å radius), and hydrophobic aromatic (cerulean blue,
with 1.6 Å radius). The frame 1260 and 2175 has three hydro-
phobic aromatic and one hydrogen bond acceptors
 JOURNAL OF BIOMOLECULAR STRUCTURE AND DYNAMICS 7

pharmacophore features. Whereas cluster conformation
frame 348 has three hydrophobic aromatic features, one
hydrogen bond acceptors and one hydrophobic feature
3.6. Lead optimization and molecular docking
Furthermore, all the compounds are combined to screen for
ADMET and TopKatV to explore the pharmacokinetics and
R

(Figure 7). The standard drug 4-hydroxytamoxifen bound

with Estrogen receptor alpha shows different pharmacophore
features and was mapped based on functional groups. A
hydrogen (H) in 4-hydroxyl group as hydrogen bond
acceptor, aromatic triphenylethylene moiety as hydrophobic
aromatic, and small lipophilic tail as hydrophobic. The gener-
ated pharmacophore model as input for structural based
pharmacophore screening of leads in four different data-
bases retrieved hit counts of 118, 35, and 45 specifically for
three molecular dynamics conformation frames 348, 1260
and, 2175, respectively.
Figure 6. Superimposed model of frames 348-Cyan, Reddish pink – 1260 and
Purple – 2175 and its interacting residues at the active site.
dynamics properties among 9 compounds obey ADMET and
lessen toxicity profile (Table 3). All the compounds predicted
solubility levels are low (2) to optimal (4) with high penetra-
tion (1) and remarkably high penetration (0) to cross the
blood brain barrier. Except compound 11166301 (3(poor)) for
all other compounds, the human intestinal absorption is
good (0) and non-inhibitor of metabolizing enzyme CYP2D6.
Also, all the compounds are found to be non-toxic, non-car-
cinogen in both gender and non-mutagen.
Before docking the validation of docking was carried out
for parameter optimization of CDOCKER protocol with slight
modification in pose cluster radius to achieve the RMSD
deviation of native pose <2.5 Å. Finally docking of nine com-
pounds using molecular dynamics docking results are favour-
able all compounds at the active site in the ranking order of
top three molecule BDD23570620 > BDC 22439020 > 670995
are tabulated in Table 4 with binding energy. In depth
atomic level binding interaction analysis shows that top
three compounds commonly form H-bond with GLU353 and
hydrophobic interactions with ALA350 and LEU387. Apart,
specific type of p-p stacking interaction was uniquely
observed in 670995 with amino acid PHE404. Three hydro-
phobic interactions such as MET421, ILE424, LEU525 are in
for compound BDC 22439020 and BDD23570620 docking
results with standard drug OHT are presented in Figure 8.
Interestingly, 670995 (isomer of Estrone) and 5870 (Estrone),
the small atomic and configuration changes have impact in
binding and interaction. Even though, these two structures
are structurally like each other, the slight change in structural
confirmation shows variation in docking scores. Eventually,
during drug designing atomic position, angle, Cartesian
space, and coordinate of the compound and active site/bind-
ing site has direct impact on the binding, scoring function
and biological activity experimentally.

Table 2. Statistical validation of structure-based pharmacophore modelling of different frames.

Frames
Pharmacophore 348 1260 2175
Number of Features 5 4 4
Features DHHaromHaromHarom DHaromHaromHarom DHaromHaromHarom
Statitical Validation
Total actives 44 44 44
Total Inactives 19,503 19,503 19,503
True Positives 40 41 41
True Negatives 16,182 11,790 13,365
False Positives 3321 7713 6138
False Negatives 4 3 3
Sensitivity 0.90909 0.93182 0.93182
Specificity 0.82972 0.60452 0.68528
AUC ROC 0.931 0.935 0.931
Model quality Excellent Excellent Excellent
Mapped hits fit value >0.7
Cardiospermum halicacabum compounds 4 2 2
Asinex IM-ON 61 17 22
Minimay bridge 22 7 6
scPDB 31 9 15
Total hits 118 35 45
 IM-ON – Immuno-Oncology, D – Hydrogen bond donor, Harom – Hydrophobic aromatic, H – Hydrophobic.
8 D. SHANMUGARAJAN ET AL.

Figure 7. Structure-based pharmacophore models of frames and its AU-ROC validation graph (a) 348, (b) 1260 and (c) 2175.

Table 3. Prediction of ADME/Toxicity properties of compounds.

NTP NTP
Rat Rat Ames
Name Solubility Level BBB Level CYP2D6 Hepato toxic Absorption Level Female Male Prediction
5870 2 1 NI NT 0 NC NC NM
670995 2 1 NI NT 0 NC NC NM
11166301 4 4 NI NT 3 NC NC NM
ASF 17525180 2 1 NI NT 0 NC NC NM
BDC 22439020 2 1 NI NT 0 NC NC NM
BDD 23570620 2 1 NI NT 0 NC NC NM
BDD 23570672 2 1 NI NT 0 NC NC NM
BDD 23570769 2 1 NI NT 0 NC NC NM
BDH 32328306 2 1 NI NT 0 NC NC NM
OHT 1 0 NI T 1 NC NC NM
NTP – National Toxicity program, NM – Non-mutagen, NC – Non-carcinogen, NI – Non-Inhibitor, NT – Non-toxic.

Table 4. CDOCKER scoring of top ranked compounds and its binding energy. are within
4 Å (Figure 9) alike standard drug tamoxifen (Figure
-CDOCKER 3(a)) which validates the rigid conformation of the drug and
-CDOCKER interaction Binding energy screened lead candidate complexes. Further, average MM/GBSA
Molecule energy energy GBSA(kcal/mol)
results of 2JF9_670995 (-31.523 kcal/mol), 2JF9_BDC22439020
5870 13.5651 46.1465 16.3366
670995 19.4085 47.61 17.2139 (-32.88 kcal/mol) and 2JF9_BDD23570620 (-43.4061 kcal/mol)
11166301 2.70019 37.6345 10.0256 correlates with docked complex binding energies concerts the
ASF 17525180 18.1867 45.9056 16.1465
BDC 22439020 21.3226 54.2778 21.9448 lead candidate’s potential.
BDD 23570620 24.0417 57.1656 33.5721
BDD 23570672 3.90171 42.0454 15.8578
BDD 23570769 2.80003 42.1819 14.8333
BDH 32328306 11.3874 46.8249 15.1961 3.8. Density functional theory for top ranked
OHT 29.5092 62.948 38.6737 compounds
DFT is the QM (Quantum Mechanical) method gained its
importance in the drug designing process in the recent
3.7. Dynamics and simulation of top compounds years, because it is considered as a significant competent
The simulation of the molecular dynamics of the docked com- method in the field of pharmaceutical studies (Shaheena,
plex served to validate the docking studies, and to understand 2022). It aids to understand properties of drugs/leads/hits
the degree of stability of the top three docked complex trajec- and their interaction with their specific receptors (Inoue
tory path as f(t). All the complexes gradually increase its devia- et al., 1998; Mavri & Hadzi, 2001). DFT analysis reveals that
tions at the beginning of the dynamics and attain its state of the smaller gap between the highest occupied molecular
equilibrium f(t). Also, three screened compounds viz., 2JF9_ orbital (HOMO) and Lowest unoccupied molecular orbital
670995, 2JF9_BDC22439020 and 2JF9_BDD23570620 deviations (LUMO) indicates lower kinetic stability and higher chemical
 JOURNAL OF BIOMOLECULAR STRUCTURE AND DYNAMICS 9

Figure 8. Receptor-ligand interaction of top three ranked compounds with standard drug (OHT).

was close to the standard drug OHT energy gap of

0.1717 eV. While BDC 22439020 and 670995 energy gaps are
0.1954 eV, and 0.2086 eV was 1.061 and 1.164 folds higher
than the standard. Compounds with least energy gaps tend
to show optimum binding affinity which is required for good
biological activities (Table 5).

Figure 9. RMSD Plot of top three compounds Receptor–ligand interaction

molecular dynamics simulation. 4. Discussion
Tamoxifen (TamoGelTM) in a native form is a prodrug that
Table 5. Density functional QM calculation of top three compounds using
Hybrid functional B3LYP method. gets converted into active 4-hydroxytamoxifen (4-(1-[4-
Compounds 670995 BDC 22439020 BDD 23570620 OHT (Dimethylaminoethoxy)phenyl]-2-phenyl-1-butenyl) phenol)
Binding Energy 62.03557834 101.4577958 99.88006206 88.5792 (Zhong et al., 2015) metabolite after its metabolism with
(Ha) cytochrome P450 isoforms such as CYP2D6 and CYP3A4
HOMO Energy 0.22165623 0.22365736 0.18200186 0.18419
(Ha)
enzymes secreted in the liver. This drug is known for its use
LUMO Energy 0.01302126 0.02819712 0.00709031 0.01247 in the treatment of metastatic breast cancer, ductal carcin-
(Ha) oma, ovarian cancer, and Adjuvant therapy (Chew, 2001). It is
Energy 0.20863497 0.19546024 0.17491155 0.17172
gap (eV) also known as selective estrogen receptor modulators (SERM)
Dipole Mag 0.68117811 3.52093486 1.44140799 1.195993 which possess mixed estrogenic and anti-estrogenic activity
Total Energy 901.4071772 1488.605818 1470.26022 1287.51 of the estrogen receptors and therefore can be used as
(Ha)
agonist and antagonist on selective tissues (Archer, 2011).
The 4-hydroxytamoxifen molecule competes with estro-
gen to bind with estrogen receptors, this metabolite acts as
reactivity of the compounds in terms of molecular interac- an ER antagonist in breast cancer tissues and blocks the tran-
tions or charge transfer interactions of compounds with tar- scription of estrogen-responsive genes thereby no cell prolif-
geted receptor enzyme. This is a computational quantum eration occurs (Shanmugarajan & David, 2022; Smith &
mechanical modelling method used to examine electronic O’Malley, 2004). However, from the in silico ADMET study, it
structure as well as to study the interactions involved was observed that the drug 4-hydroxytamoxifen can causes
between receptors and ligands (Mavri & Hadzi, 2001). DFT hepatotoxic (Gao et al., 2016) and it was correlated with an
calculations performed with B3LYP functional showed that experimental study performed in mice at the early stage dur-
the BDD 23570620 has HOMO-LUMO energy gap 0.1749 eV ing endocrine therapy (Blackburn et al., 1984; Floren et al.,
10 D. SHANMUGARAJAN ET AL.
1998; Gao et al., 2016). Thus, an alternative compound with
hepatotoxic free is required to treat estrogen receptor-posi-
tive breast cancer.
This study attempted to introduce the dynamacophore
(dynamics-based pharmacophore) model, this method accel-
erates to identify of the binding interaction maintained on
several configurations of the active site amino acids and to
define a complementary model based on those regions.
Screening of compounds in static pharmacophore position
ends may lead to generating false positives compounds and
increases uncertainty in the experimental method. Whereas
the dynamics-based pharmacophore generation at various
time positions of the drug target complex and its screening
at different time points and Cartesian space will reduce the
uncertainty in a biological assay. To bolster the study the
previous reports on the significant difference between static
and dynamic pharmacophore models of HIV-1 integrase.
Further, in the same study of HIV-1 integrase the library of
molecules was screened in both models shows that the
results of the dynamic model are likely to inhibit the drug
target (HIV-1 integrase) more efficiently than the static
pharmacophore model (Carlson et al., 2000).
In addition, the potent PPAR-gamma lead candidate was
screed using a multi-conformation-based structure pharma-
cophore model (Prabitha et al., 2022). Similarly, the study on
100 ns of estrogen receptor –alpha conformations structure-
based pharmacophore models were retained based on
potential energy and binding interaction leads to generate
the potent lead candidates from medicinal plant
Andrographis paniculata (Shanmugarajan & David, 2022).
Hence, in the rational-based drug discovery process, a new
“paradigm shift” is required in pharmacophore-based screen-
ing to generate the lead candidates with therapeutical
potential.
5. Conclusion
OHT bound with estrogen receptor alpha was used to gener-
ate multi-conformation and structure-based pharmacophore.
The molecular dynamics and simulation of 150 ns is to
understand the stable equilibria of protein-ligand complex.
The active site residues interaction of OHT plays an essential
role in biological activity generating the static pharmaco-
phore model using computational biology tools will increase
the uncertainty in drug discovery process. Thus, dynamaco-
phore based screening of compounds with potential stable
frames was considered as effective method for lead optimiza-
tion and library screening. The generated three pharmaco-
phore model based on OHT in three potential stable cluster
representative complexes with internal validation using AU-
ROC values above 0.9 lead to build five major pharmaco-
phore features including hydrogen binding acceptor, hydro-
phobic and aromatic hydrophobic. Further screening with
four diverse set databases resulted a set of 198 molecules.
Lead optimization for pharmacokinetics and dynamics ended
up with nine candidates, further receptor-ligand interaction was
more favourable for compounds BDD23570620 > BDC
22439020 > 670995. Moreover, all these compounds are non-
mutagen and carcinogen with high penetration across the
blood brain barrier, but OHT was hepatoxic with moderate solu-
bility. Thus, these compound BDD23570620 and BDC 22439020
Asinex, Immune Oncology library and 670995 compound from
medicinal plant, C. halicacabum shows least HUMO-LUMO
energy that can act as a lead compound against estrogen recep-
tor alpha and can be developed as a therapeutic solution for
treating hormone dependent breast cancer through ESR1
(Estrogen Receptor Alpha). Overall, dynamacophore based
pharmacophore model was a promising technique to develop
more estrogen receptor alpha ligand candidates to help the
drug discovery process to combat cancer.
Acknowledgements
The First author, Ms. Dhivya is grateful to Altem Technologies,
Bengaluru, India, for their extended support in performing this research
work.
Disclosure statement
No potential conflict of interest was reported by the author(s).
Funding
The author(s) reported there is no funding associated with the work fea-
tured in this article.
References
Ali, S., Rasool, M., Chaoudhry, H., N Pushparaj, P., Jha, P., Hafiz, A.,
Mahfooz, M., Abdus Sami, G., Azhar Kamal, M., Bashir, S., Ali, A., &
Sarwar Jamal, M. (2016). Molecular mechanisms and mode of tamoxi-
fen resistance in breast cancer. Bioinformation, 12(3), 135–139. https://
doi.org/10.6026/97320630012135
Anderson, W. F., Rosenberg, P. S., Prat, A., Perou, C. M., & Sherman, M. E.
(2014). How many etiological subtypes of breast cancer: Two, three,
four, or more? Journal of the National Cancer Institute, 106(8), 1–13.
https://doi.org/10.1093/jnci/dju236
Archer, D. F. (2011). The gynecologic effects of lasofoxifene, an estrogen
agonist/antagonist, in postmenopausal women. Menopause (New York,
N.Y.), 18(1), 6–7. https://doi.org/10.1097/gme.0b013e318203a46b
Becke, A. D. (1993). Density-functional thermochemistry. III. The role of
exact exchange. Journal of Chemical Physics. 98(7), 5648–5652. https://
doi.org/10.1063/1.464913
Blackburn, A. M., Amiel, S. A., Millis, R. R., & Rubens, R. D. (1984).
Tamoxifen and liver damage. British Medical Journal (Clinical Research
ed.), 289(6440), 288. https://doi.org/10.1136/bmj.289.6440.288
Braal, L., Jager, A., Lommen, K. M., Oomen-de Hoop, E., de Bruijn, P.,
Vastbinder, M. B., van Rossum-Schornagel, Q. C., Thijs-Visser, M. F.,
van Alphen, R., Struik, E. M., Zuetenhorst, J. M., Mathijssen, R. H., &
Koolen, S. L. (2020). Therapeutic drug monitoring of tamoxifen to
improve adjuvant treatment of hormone sensitive breast cancer: The
TOTAM study. Annals of Oncology, 31(4), S319. https://doi.org/10.
1016/j.annonc.2020.08.313
Bussi, G., Donadio, D., & Parrinello, M. (2007). Canonical sampling
through velocity rescaling. The Journal of Chemical Physics, 126(1),
014101. https://doi.org/10.1063/1.2408420
Carlson, H. A., Masukawa, K. M., Rubins, K., Bushman, F. D., Jorgensen,
W. L., Lins, R. D., Briggs, J. M., & McCammon, J. A. (2000). Developing
a dynamic pharmacophore model for HIV-1 integrase. Journal of
Medicinal Chemistry, 43(11), 2100–2114. https://doi.org/10.1021/
jm990322h

Chew, H. K. (2001). Adjuvant therapy for breast cancer: Who should get
what? The Western Journal of Medicine, 174(4), 284–287. https://doi.
org/10.1136/ewjm.174.4.28
Daura, X., Jaun, B., Seebach, D., Van Gunsteren, W. F., & Mark, A. E.
(1998). Reversible peptide folding in solution by molecular dynamics
simulation. Journal of Molecular Biology, 280(5), 925–932. https://doi.
org/10.1006/jmbi.1998.1885
DeSantis, C. E., Ma, J., Gaudet, M. M., Newman, L. A., Miller, K. D., Goding
Sauer, A., Jemal, A., & Siegel, R. L. (2019). Breast cancer statistics. CA:
A Cancer Journal for Clinicians, 69(6), 438–451. https://doi.org/10.3322/
caac.21583
Dhivya, S., Suresh Kumar, C., Bommuraj, V., Janarthanam, R., Chandran,
M., Usha, T., & Middha, S. K. (2018). A study of comparative modelling,
simulation, and molecular dynamics of CXCR3 receptor with lipid
bilayer. Journal of Biomolecular Structure & Dynamics, 36(9), 2361–
2372. https://doi.org/10.1080/07391102.2017.1354783
Early Breast Cancer Trialists’ Collaborative Group. (1998). Tamoxifen for
early breast cancer: An overview of the randomised trials. The Lancet,
351(9114), 1451–1467. https://doi.org/10.1016/S0140-6736(97)11423-4
Eeckhoute, J., Carroll, J. S., Geistlinger, T. R., Torres-Arzayus, M. I., &
Brown, M. (2006). A cell type-specific transcriptional network required
for estrogen regulation of cyclin D1 and cell cycle progression in
breast cancer. Genes & Development, 20(18), 2513–2526. https://doi.
org/10.1101/gad.1446006
Ercheng, W., Huiyong, S., Junmei, W., Zhe, W., Hui, L., John, Z. H. Z., &
Tingjun, H. (2019). End-point binding free energy calculation with
MM/PBSA and MM/GBSA: Strategies and applications in drug design.
Chemical Reviews, 119(16), 9478–9508. https://doi.org/10.1021/acs.
chemrev.9b00055
Fentiman, I. S., Fourquet, A., & Hortobagyi, G. N. (2006). Male breast can-
cer. Lancet (London, England), 367(9510), 595–604. https://doi.org/10.
1016/s0140-6736(06)68226-3
Ferlay, J., Soerjomataram, I., Dikshit, R., Eser, S., Mathers, C., Rebelo, M.,
Parkin, D. M., Forman, D., & Bray, F. (2015). Cancer incidence and mor-
tality worldwide: Sources, methods, and major patterns in GLOBOCAN
2012. International Journal of Cancer, 136(5), E359–E386. https://doi.
org/10.1002/ijc.29210
Floren, L. C., Hebert, M. F., Venook, A. P., Jordan, V. C., Cisneros, A., &
Somberg, K. A. (1998). Tamoxifen in liver disease: Potential exacerba-
tion of hepatic dysfunction. Annals of Oncology: Official Journal of the
European Society for Medical Oncology, 9(10), 1123–1126. https://doi.
org/10.1023/a:1008269025294
Gao, F. F., Lv, J. W., Wang, Y., Fan, R., Li, Q., Zhang, Z., & Wei, L. (2016).
Tamoxifen induces hepatotoxicity and changes to hepatocyte morph-
ology at the early stage of endocrinotherapy in mice. Biomedical
Reports, 4(1), 102–106. https://doi.org/10.3892/br.2015.536
Gogoi, D., Chaliha, A. K., Sarma, D., Kakoti, B. B., & Buragohain, A. K.
(2017). Identification of potential type 4 cAMP phosphodiesterase
inhibitors via 3D pharmacophore modeling, virtual screening, DFT
and structural bioisostere design. Medicinal Chemistry Research, 26(11),
3000–3014. https://doi.org/10.1007/s00044-017-1998-3
Hard, G. C. (1993). Major difference in the hepatocarcinogenicity and
DNA adduct forming ability between toremifene and tamoxifen in
female Crl: CD (BR) rats. Cancer Res, 53(19), 4534–4541.
Heldring, N., Pawson, T., McDonnell, D., Treuter, E., Gustafsson, J. A., &
Pike, A. C. (2007). Structural insights into corepressor recognition by
antagonist-bound estrogen receptors. The Journal of Biological
Chemistry, 282(14), 10449–10455. https://doi.org/10.1074/jbc.
M611424200
Hiteshi, T., Tanmoy, C., & Vandana, S. (2019). A brief review on import-
ance of DFT in drug design. Res Med Eng Sci, 7(4), 791-795. https://
doi.org/10.31031/RMES.2019.07.00068
Inoue, Y., Sugio, S., Andzelm, J., & Nakamura, N. (1998). A density func-
tional study of the receptor  ligand interaction: Stabilization energy
in ammonium salt  aromatic interactions. The Journal of Physical
Chemistry A, 102(3), 646–648. https://doi.org/10.1021/jp972661x
Jordan, V. C. (2004). Selective estrogen receptor modulation: Concept
and consequences in cancer. Cancer Cell, 5(3), 207–213. https://doi.
org/10.1016/S1535-6108(04)00059-5
JOURNAL OF BIOMOLECULAR STRUCTURE AND DYNAMICS 11
Kim, Y. J., Jang, S. K., Hong, S. E., Park, C. S., Seong, M. K., Kim, H. A.,
Park, K. S., Kim, C. H., Park, I. C., & Jin, H. O. (2022). Knockdown of
YAP/TAZ sensitizes tamoxifen resistant MCF7 breast cancer cells.
Biochemical and Biophysical Research Communications, 601(2022), 73–
78. https://doi.org/10.1016/j.bbrc.2022.02.083
Kulkarni, P., & Rawtani, D. (2019). Application of box-Behnken design in
the preparation, optimization, and in vitro evaluation of self-assembly
based tamoxifen- and doxorubicin-loaded and dual drug loaded nio-
somes for combinatorial breast cancer treatment. Journal of
Pharmaceutical Sciences, 108(8), 2643–2653. https://doi.org/10.1016/j.
xphs.2019.03.020
Lobanov, M. Y., Bogatyreva, N. S., & Galzitskaya, O. V. (2008). Radius of
gyration as an indicator of protein structure compactness. Molecular
Biology, 42(4), 623–628. https://doi.org/10.1134/S0026893308040195
Lumachi, F., Luisetto, G., Basso, S. M. M., Basso, U., Brunello, A., &
Camozzi, V. (2011). Endocrine therapy of breast cancer. Current
Medicinal Chemistry, 18(4), 513–522. https://doi.org/10.2174/
092986711794480177
Maryam, A. J., Oluwakemi, E., Nkululeko, D., & Michael, S. (2020). Virtual
screening, molecular docking studies and DFT calculations of FDA
approved compounds similar to the non-nucleoside reverse tran-
scriptase inhibitor (NNRTI) efavirenz. Heliyon, 6(8), e04642. https://doi.
org/10.1016/j.heliyon.2020.e04642
Mavri, J., & Hadzi, D. (2001). Modelling of ligand–receptor interactions:
Ab-initio and DFT calculations of solvent reaction field effects on
methylated ammonium–p and–acetate complexes. Journal of
Molecular Structure: THEOCHEM, 540(1–3), 251–255. https://doi.org/10.
1016/S0166-1280(00)00733-8
Memisoglu-Bilensoy, E., Vural, I., Bochot, A., Renoir, J. M., Duchene, D., &
Hincal, A. A. (2005). Tamoxifen citrate loaded amphiphilic b-cyclodex-
trin nanoparticles: In vitro characterization and cytotoxicity. Journal of
Controlled Release: Official Journal of the Controlled Release Society,
104(3), 489–496. https://doi.org/10.1016/j.jconrel.2005.03.006
Meslamani, J., Li, J., Sutter, J., Stevens, A., Bertrand, H. O., & Rognan, D.
(2012). Protein-ligand-based pharmacophores: Generation and utility
assessment in computational ligand profiling. Journal of Chemical
Information and Modeling, 52(4), 943–955. https://doi.org/10.1021/
ci300083r
Morad, S. A. F., & Cabot, M. C. (2015). Tamoxifen regulation of sphingo-
lipid metabolism—Therapeutic implications. Biochimica et Biophysica
Acta, 1851(9), 1134–1145. https://doi.org/10.1016/j.bbalip.2015.05.001
Nounou, M. I., ElAmrawy, F., Ahmed, N., Abdelraouf, K., Goda, S., & Syed-
Sha-Qhattal, H. (2015). Breast cancer: Conventional diagnosis and
treatment modalities and recent patents and technologies. Breast
Cancer: Basic and Clinical Research, 9(Suppl 2), 17–34. https://doi.org/
10.4137/bcbcr.s29420
Parrinello, M., & Rahman, A. (1981). Polymorphic transitions in single
crystals: A new molecular dynamics method. Journal of Applied
Physics, 52(12), 7182–7190. https://doi.org/10.1063/1.328693
Peng, J., Wang, W., Yu, Y., Gu, H., & Huang, X. (2018). Clustering algo-
rithms to analyze molecular dynamics simulation trajectories for com-
plex chemical and biological systems. Chinese Journal of Chemical
Physics, 31(4), 404–420. https://doi.org/10.1063/1674-0068/31/
cjcp1806147
Phillips, J. C., Braun, R., Wang, W., Gumbart, J., Tajkhorshid, E., Villa, E.,
Chipot, R., Skeel, R. D., Kale, L., & Schulten, K. (2005). Scalable molecu-
lar dynamics with NAMD. Journal of Computational Chemistry, 26(16),
1781–1802. https://doi.org/10.1002/jcc.20289
Poli, G., Granchi, C., Rizzolio, F., & Tuccinardi, T. (2020). Application of
MM-PBSA methods in virtual screening. Molecules, 25(8), 1971. https://
doi.org/10.3390/molecules25081971
Prabitha, P., Dhivya, S., Durai, A. K. T., & Prashantha Kumar, B. R. (2022).
Multi-conformational frame from molecular dynamics as a structure-
based pharmacophore model for mapping, screening, and identifying
ligands against PPAR-c: A new protocol to develop promising candi-
dates. Journal of Biomolecular Structure & Dynamics, 40(6), 2663–2673.
https://doi.org/10.1080/07391102.2020.1841677
Rani, A., Stebbing, J., Giamas, G., & Murphy, J. (2019). Endocrine resist-
ance in hormone receptor positive breast cancer-from mechanism to
12 D. SHANMUGARAJAN ET AL.
therapy. Frontiers in Endocrinology, 10, 245. https://doi.org/10.3389/
fendo.2019.00245
Shafi, M., Mohammad, A. R. U., Gobindo, K. P., Sharmin, S. S., Saiful, I.,
Ekhtiar, R., Ariful, I., Mohammed, S. I., Maria, M. P., Talha, B. E., &
Mohammed, A. S. (2021). Virtual screening and molecular dynamics
simulation study of plant-derived compounds to identify potential
inhibitors of main protease from SARS-CoV-2. Briefings in
Bioinformatics, 22(2), 1402–1414. https://doi.org/10.1093/bib/bbaa428
Shaheena, R. (2022). Role of DFT in drug design: A mini review. Drug
Designing, 11, 216.
Shanmugarajan, D., & David, C. (2022). Estrogen receptor potentially sta-
ble conformations from molecular dynamics as a structure-based
pharmacophore model for mapping, screening, and identifying
ligands-a new paradigm shift in pharmacophore screening. Journal of
Biomolecular Structure and Dynamics. 11, 1–10. https://doi.org/10.
1080/07391102.2022.2074543
Shanmugarajan, D., Prabitha, P., Kumar, B. R. P., & Suresh, B. (2020).
Curcumin to inhibit binding of spike glycoprotein to ACE2 receptors:
Computational modelling, simulations, and ADMET studies to explore
curcuminoids against novel SARS-CoV-2 targets. RSC Advances, 10(52),
31385–31399. https://doi.org/10.1039/d0ra03167d
Smith, C. L., & O’Malley, B. W. (2004). Coregulator function: A key to
understanding tissue specificity of selective receptor modulators.
Endocrine Reviews, 25(1), 45–71. https://doi.org/10.1210/er.2003-0023
Sung, H., Ferlay, J., Rebecca, L., Siegel, Laversanne, M., Soerjomataram, I.,
Jemal, A., & Bray, F. (2021). Global Cancer Statistics 2020: GLOBOCAN
estimates of incidence and mortality worldwide for 36 cancers in 185
countries. CA: A Cancer Journal for Clinicians, 71(3), 209–249. https://
doi.org/10.3322/caac.2166
Zeng, X., & Yee, D. (2007). Insulin-like growth factors and breast cancer
therapy. Advances in Experimental Medicine and Biology, 608, 101–112.
https://doi.org/10.1007/978-0-387-74039-3_7
Zhao, X., Jiang, Y., Xu, M., Hu, J., Feng, N., Deng, H., Lu, C., & Huang, T.
(2022). Indoleamine 2,3-dioxygenase 1 regulates breast cancer
Tamoxifen resistance through interleukin-6/signal transducer and acti-
vator of transcription. Toxicology and Applied Pharmacology,
440(2022), 115921. https://doi.org/10.1016/j.taap.2022.115921
Zhong, Q., Zhang, C., Zhang, Q., Miele, L., Zheng, S., & Wang, G. (2015).
Boronic prodrug of 4-hydroxytamoxifen is more efficacious than tam-
oxifen with enhanced bioavailability independent of CYP2D6 status.
BMC Cancer, 15(1), 625. https://doi.org/10.1186/s12885-015-1621-2