Biological Activity of Selected Compounds from Annona muricata Seed as Antibreast Cancer Agents: Theoretical Study

Several natural products have been of help to humans, and its effect is noticeable in the medicinal world. Soursop with botanical name Annona muricata L. possesses antidiarrhea, anticold fever, antirheumatism, and antineuralgia properties. In this work, five selected molecular compounds were studied against type 3 of 3α-hydroxysteroid dehydrogenase (3α-HSD). Its anticancer activity was investigated using the quantum chemical method via Spartan 14 software, molecular docking via Discovery studio 2017, AutoDock Tool 1.5.6, AutoDock Vina 1.1.2, and PyMol 1.7.4.4 and the molecular dynamic simulation method via AMBER14 molecular dynamics package. Many descriptors (EHOMO, ELUMO, dipole moment, energy bandgap, area, volume, polarizability, polar surface area, Log P, hydrogen bond donor, and hydrogen bond acceptor) which describe the anticancer activity of the studied compounds were obtained. Also, the docking study revealed the inhibiting ability of the studied compound, and it was observed that compound C possesses a greater ability to inhibit than other studied compounds as well as the standard (5FU).


Introduction
e use of natural product compounds in supporting human health remains one of the vital ways of treatment globally [1]. Phytochemical compounds present in vegetation have played a crucial role in drug discovery, and their cytotoxicity has drawn the attention of researchers over the years [2]. Also, several scientists have declared plants with efficient biological importance to be source for well-being conservation [3]. As reported by Ko and Moon, the use of natural product in treating breast cancer is beneficial, and the chemical compounds present in these products have little or no negative effect compared to synthetic drug molecules [4].
Soursop with botanical name Annona muricata L. belongs to the family Annonaceae, and it is a tropical fruit-bearing tree usually found in some areas of Africa, America, Australia, and Asia [5]. It is a tree with thick leaves that are oblate. Annona muricata possesses touchy, dark green fruit [6,7]. Several researchers have reported this plant to be very useful in treating many diseases such as cancer, diabetes, inflammation, and hypertension [8]. According to Vijayameena et al., it helps in curing diabetic due to hypoglycemic and antioxidant features it contains [9]. Most parts of Annona muricata are useful in herbal medicine. Some parts, such as roots and stems, help in lowering blood pressure and also act as an antispasmodic agent [5]. Also, Annona muricata fruit plays a serious role in the medicinal world such as antidiarrhoea, anticold fever, antirheumatism, and antineuralgia and its consumption helps in amassing mother's milk after giving birth [10]. As reported by some researchers, the biological activities of extracts from Annona muricata were observed to be very toxic to cancer cells than normal human cells [11][12][13][14][15]. Also, as reported by Shashanka and Devananda, most of the molecular compounds present in this plant have biomedical significance, and further study on this work is still ongoing [16]. According to Rieser et al., it was observed that muricatacin, as well as other compounds present in Annona muricata, possesses antiproliferative characteristics against breast, colorectal, and lung cancer cell line [17]. Wu et al. reported antilung cancer, antibreast cancer, and anticolon cancer activities of annomuricin A and annomuricin B. It was shown that they are potent against the breast, lung, and colon cancer [18]. Moreover, breast cancer has caused the death of several women over the years [19]. e rate at which this malignant neoplasia increases is on the high side [20], and as reported by Keogh et al., almost 95% of women have moderately growing risk of breast cancer [21]. e prospective methods for curing cancer remain chemotherapy, surgery, and radiotherapy; these methods have been used for several years ago with victory in several cases. Nevertheless, these methods have been recorded futile in some cases such as unconventional metastatic breast cancer [22]. More so, the rate of survival among the patient with breast cancer keeps increasing, and this could be attributed to improvement in drug designing and development [23].
3α-HSD as a vital enzyme plays a serious role in breaking down of androgen [24]. In sex, 3α-HSD has been recognised in human to be a serious enzyme due to the role it played [25]. According to Aka et al., some estrogens like 17βoestradiol ignite proliferation of breast cancer cells in woman although 5α-dihydrotestosterone (5α-DHT) have the strength to shrink the growth of breast cancer in woman [26]; as reported by Penning et al., 3α-HSD can hinder 5αdihydrotestosterone, which will thereby increase the growth of breast cancer cells in woman [27]. Also, Bo et al. revealed that overriding 3α-HSD will spontaneously reduce MCF-7 cells growth [28]. erefore, this work is aimed at identifying the molecular descriptors responsible for antibreast cancer activities of selected compounds in Annona muricata seed and studies the nonbonding interactions as well as the actual binding energy between the selected compounds and 3α-HSD using docking and molecular dynamics simulation methods, respectively.

Optimization and Molecular
Docking. Five molecular compounds selected from Annona muricata seed were screened against 3α-HSD via an in silico method. e crystal structure was retrieved from protein data bank [29] with the accession code 4XO6 [28]. e selected compounds were optimized using Spartan 14 [30]. Also, the docking study was executed using a series of software (Discovery studio 2017, AutoDock Tool 1.5.6, AutoDock Vina 1.1.2, and EduPyMol version 1.7.4.4), and the calculated binding affinities were reported.
is docking study involves the preparation of 3α-HSD and the studied ligands using PyMol. e binding site in the studied receptor was located using AutoDock Tool, and the docking calculation was performed via AutoDock Vina. More so, the nonbonding interaction was viewed through Discovery studio 2017. e observed grid box was as follows: center (X � 12.444, Y � −0.171, and Z � 2.943) and size (X � 62, Y � 60, and Z � 74), and the spacing was set to be 1.

Molecular Dynamic Simulation Studies.
e compound with better binding affinity was selected from the compounds docked, and chloroquine was used as a standard; both were subjected to molecular dynamics simulation on AMBER14 molecular dynamics package [32]. Hydrogen atom was added to the studied complexes using a leap segment of AMBER14. e protein was described with AMBER force field 99SB while the ligand with general AMBER force field [33,34]. To neutralize the studied complexes before solvation, the required number of counterions was added. e solvation was implemented in a condensed octahedral cell of TIP3P [35] water molecules, and 12Å was stretched outside the protein on each side. e complexes were minimized using 5000 frames of steepest descent minimization and 10000 frames of conjugated gradient minimization to eliminate bad atom interactions. e entire systems were heated at 300 K and 50000 ps. Molecular dynamics simulation was accomplished for the studied complex at 300 K and 1 atm via the particle mesh Ewald method [36]. CPPTRAJ module [33] implemented in AMBER14 software was used for molecular dynamics trajectories analysis.

Density Functional eory Calculations.
Series of molecular descriptors obtained from carefully selected five molecular compounds in Annona muricata seed via density functional theory (DFT) are highest occupied molecular orbital energy (E HOMO ), lowest unoccupied molecular orbital energy (E LUMO ), dipole moment, energy bandgap, area, volume, polarizability, polar surface area, log P, hydrogen bond donor, and hydrogen bond acceptor (Table 1). In this work, the contribution of dipole moment to the biological activity of studied compounds was observed. According to David et al., it was reported that the presence of dipole moment as a nonbonded interaction in the drug-like compound-protein complex is vital [37]. e accepted range for dipole moment is 3-5 kJ/mol; thus, compound E was the only compound that falls within the set standard, so it is expected to have strong nonbonded connections 3α-HSD.
Furthermore, the calculated log P reveals the dissolving capacity of drug-like molecules in lipophilic media [38]. As reported by Meanwell 2011, there may be complications in oral absorption if the calculated log P is greater than the standard value for log P (Log P ≤ 5) [36]. erefore, compound E is expected to dissolve well in nonaqueous substance and possesses the ability to be administered through the human mouth. More so, the calculated E HOMO which describes the tendency of drug-like molecules to donate electron to the neighbouring compounds was calculated to be −6.74 (A), −6.69 (B), −6.66 (C), −6.75 (D), and −7.17 (E). As shown in Table 1, compound C is expected to inhibit than other studied compounds [39]. e calculated E LUMO shows the capability of compounds to accept electrons from nearby molecules [40]. In this regard, compound C with −1.15 eV is expected to possess a greater ability to accept electron from the nearby compounds than other studied compounds. Other studied calculated descriptors are displayed in Table 1.
More so, the calculated band gap shows the level of reactivity of compounds with drug potentials. Also, many reports from several researchers have proved that level of reactivity of molecular compounds is a function of lower bandgap; thus, compound C is expected to react better than other studied compounds [41]. Figure 2 shows the E HOMO -E LUMO overlay.
Furthermore, chiral information of some stereogenic centers of the studied compounds as displayed in Figure 1 was investigated. In compound A, seven (7) chiral centers were observed (4R and 3S), and seven chiral centers were also observed in compound B with 7R and 6R as the chiral center was observed in compound C. More so, seven (7) and two (2) chiral centers were detected in compounds D and E, respectively.

Prediction of ADMET Properties.
Series of obtained ADMET properties using admetSAR server [31] were reported. As shown in Table 2, it was observed that compounds A, B, D, and E had better human intestinal absorption than the standard used (5FU) except compound C since drug-like compound with the higher value of human intestinal absorption can be absorbed well in the intestine [42]. e values obtained for the blood-brain barrier for compounds A-E showed that compound E has a very closer value to that of 5FU. Also, the blood-brain barrier value for compounds A-D falls within the same ranges shown in Table 2. e obtained P-glycoprotein report (nonsubstrate and noninhibitor) for all the studied compounds was similar to those of 5FU. Also, to know the level of mutagenicity of the studied compounds, AMES was considered. us, in terms Journal of Chemistry of AMES study, all the studied compounds were negative, and this also correlated with the standard (5FU). As shown in Table 2, all the studied compounds were not carcinogenic and these were similar to the similar used. e calculated cytochrome P450 showed that it could be inhibited by some of the studied compounds.

Molecular Docking and Scoring
Studies. Series of selected molecular compounds from Annona muricata seed were docked into the active site of 3α-HSD to observe the nonbonding interaction that was involved. To establish the accuracy of the engaged docking method, the docked ligand retrieved from the crystal structure of 3α-HSD was docked again into the active site of the 3α-HSD to determine the similarity of the best conformation of the redocked ligand (i.e., compound with the lowest energy) to the position of the native ligand ( Figure 3). Hence, the similarity index of the native drug-like compound retrieved from the downloaded receptor to the redocked drug compound was similar to 1. In this work, the docking method used is observed to be dependable. In this study, it was observed that compound C has a promising ability to inhibit than other studied compounds. Also, the correlation between the molecular descriptors was observed and it was discovered that E HOMO , E LUMO , and bandgap as displayed in Table 1 greatly contributed to the promising inhibiting activity of compound C than other studied compounds. e docking score, as well as the binding affinity obtained for the studied complexes, was calculated to be negative, and this is a sign that the studied compounds could serve as the starting point for developing effective drugs targeting 3α-HSD, thereby reducing breast cancer in woman globally. Also, as shown in Table 3

Root of Mean Square Deviation (RMSD)
. RMSD of 3α-HSD backbone atoms in connection to the initial molecular structure of the studied enzyme in forming complexes with compounds C and 5FU during the 100 ns MD simulation is displayed in Figure 5. is was accomplished to investigate the extent of deviation from the initial structure upon binding and the steadiness of the simulated complexes. us, the structures shown in Figure 5 proved that the conformation of the studied complexes was stable and the average RMSD value for compounds C-3α-HSD and 5FU-3α-HSD was 1.17Å and 1.47Å, respectively. e systems were well equilibrated and stable after 60 ns of the simulation time.

Root of Mean Square
Fluctuation. e calculated root mean square fluctuation (RMSF) reveals the flexibility of residues upon binding of the two compounds tested during simulation time.
e studied RMSF helped to know the flexibility of the residue present in the studied enzymes during 50000 ps simulation time ( Figure 6). As displayed in Figure 5, there is a similarity in the trend at which the observed residue vacillates; however, little variation was observed between compounds C-3α-HSD and 5FU-3α-HSD. e pattern shown in Figure 5 proved that compound C-3α-HSD exhibited a minor degree of deviation and this shows a higher level of affinity for compound C to inhibit 3α-HSD.

Binding Free Energy.
e binding energy for compounds C-3α-HSD and 5FU-3α-HSD was calculated using the MM/GBSA method ( Table 4).
As shown in Table 4, it was observed that compound C possesses a better ability to inhibit 3α-HSD than the standard used. e calculated free energy includes van der Waal energy (−87.98 kcal/mol), electrostatic energy (−10.94 kcal/ mol), polar solvation energy (34.36 kcal/mol), and nonpolar solvation energy (−11.16 kcal/mol). is specifies that van der Waal energy, electrostatic energy, and nonpolar solvation energy are favourable; however, polar solvation energy was not favourable for the molecular binding of compound C to 3α-HSD.

Conclusion
Breast cancer is an incessantly growing menace worldwide. Despite several ways to combat this deadly disease by many scientists/researchers, its rate of operation among women keeps increasing. Also, 3α-HSD contributes to the existence of breast cancer in woman, and inhibiting 3α-HSD will impulsively decrease MCF-7 cells growth. In this work, an  investigation was carried out on Annona muricata L. seed as an anticancer agent via density functional theory, docking study, and molecular dynamics simulation studies. All the studied compounds proved to be active as an antibreast cancer agent. Also, the molecular descriptors that described the antihuman breast cancer activities in the studied compounds and their contribution to antihuman breast cancer of the studied compounds were identified. More so, compound C possesses a better ability to inhibit 3α-HSD than other studied compounds and the standard (5FU). erefore, this study shows the potential ability of Annona muricata L. seed as an effective antibreast cancer agent.

Data Availability
All data used to support the findings of the study are available from the corresponding author on reasonable request.

Conflicts of Interest
e authors declare that they have no conflicts of interest.