Anti-cancer Potential of Moringa oleifera on BRCA1 Gene: Systems Biology

Breast Cancer has always been a global challenge that is prevalent among women. There is a continuous increase in the high number of women mortality rates as a result of breast cancer and affecting countries at all levels of modernization. Women with high-risk factors including hereditary, obesity, and menopause have the possibility of developing breast cancer. With the advent of radiotherapy, chemotherapy, hormone therapy, and surgery in the treatment of breast cancer, there has an increased number of breast cancer survivors. Also, the design and development of drugs targeting therapeutic enzymes are helping to effectively treat the tumor cells at an early stage. However, long term use of anti-cancer drugs has been linked to severe side effects. This research aims to develop potential drug candidates from Moringa oleifera which could serve as anti-cancer agents. In silico analysis using Schrödinger Molecular Drug Discovery Suite and SWISS ADME was employed to determine the therapeutic potential of phytochemicals from M. oleifera against breast cancer via molecular docking, pharmacokinetic parameters, and drug-like properties. The result shows that Rutin, Vicenin-2, and Quercetin-3-O-glucoside have the highest binding energy of −7.522, −6.808, −6.635kcal/mol respectively in the active site of BRCA1. The essential amino acids involved in the protein-ligand interaction following active site analysis are ASN 1678, ASN 1774, GLY 1656, LEU 1657, GLN 1779, LYS 1702, SER 1655, PHE 1662, ARG 1699, GLU 1698, and VAL 1654. Thus, we propose that bioactive compounds from M. oleifera may be potential hit drug candidates against breast cancer.


INTRODUCTION
Breast cancer is the leading cause of death in women around the world. Several factors contribute significantly to the increased risk of breast cancer which includes oral contraceptives, obesity, menopause, and elevation in serum estradiol concentration [1]. Ductal carcinoma is the most common type of breast cancer which developed from the ducts. Cancerous cells developing from lobules are called lobular cells [2]. Breast cancers are mostly diagnosed by physical examination by a healthcare provider or the use of mammography [3]. High occurrence of breast cancer has been reported to be prevalent in white women within the range of forty years and above [4].
Breast cancer gene 1 (BRCA-1) also called caretaker gene is a tumor suppressor gene that functions in cell cycle regulation, DNA repair mechanism, and other metabolic processes [5] [6].
The BRCA-1 proteins interact with other essential proteins necessary in DNA replication and repairing of double-stranded DNA breaks [7]. It contains 1863 amino acid residues and helps to inhibit the proliferation of cells lining the milk ducts of the breast. Thus, BRCA-1 does not contribute to the pathogenesis of breast cancer. However, mutations in the sequence of the Breast Cancer gene can consequently increase the risk of breast cancer [8]. Mutations evolved when the genetic makeup of an individual becomes damaged via exposure to environmental factors including ultra-violet light, ionizing radiation, and genotoxic chemicals [9]. When the BRCA-1 is mutated, it cannot efficiently repair the broken DNA, thereby, the prevention of breast cancer will be hampered [10].
There are several treatment methods available for breast cancers but hormone-blocking agents, chemotherapy and monoclonal antibodies are the most commonly used [11] [12]. Hormone receptors (estrogen ER+ and progesterone PR+ receptors) are a therapeutic target in breast cancer. Drugs such as tamoxifen and anastrozole act by blocking the hormone receptors [13].
Several medicinal plants such as Camptothecan acuminate, Catharanthus roseus, Taxus brevifolia, and many others have been used as anti-cancer therapy [14].
Moringa oleifera which belongs to the family of Moringaceae has been reported to possess beneficial pharmacological properties such as anticonvulsant, antimicrobial, anticancer, and antiviral [15]. The extracts (phytochemicals) from the leaves, seeds, bark, and flowers of M.oleifera have been used in the treatment of several chronic diseases including hypercholesterolemia, high blood pressure, diabetes, insulin resistance, non-alcoholic liver disease, cancer, and inflammation [16]. Bioactive compounds of M. oleifera shows inhibitory potential against cancerous cell line by inhibiting proliferation of carcinoma cells and malignant astrocytoma cells [17] [18]. In this study, in silico analysis via: molecular docking and pharmacokinetic profiles were employed to screen the library of bioactive compounds from M.oleifera to determine their anticancer property.

LIGAND PREPARATION
The phytochemicals of M. oleifera were retrieved from published literature [15] and their crystal structures were downloaded from the PubChem database (https://pubchem.ncbi.nlm.nih.gov/).

PROTEIN PREPARATION
The crystal structures of the BRCA-1 (PDB ID: 4OFB) was retrieved from Protein Data Bank (https://www.rcsb.org/) in complex with co-crystallized ligands. The protein was prepared using ProteinPrep Wizard of Maestro interface (11.5) by adding missing hydrogen atoms. Furthermore, the metal ionization was corrected to ensure formal charge and force field treatment. The protein was optimized and refined for docking analysis [20] [21].

MOLECULAR DOCKING
The docking analysis was conducted using the Glide tool from Schrodinger molecular drug discovery suite (version 2017-1). The grid was generated using the receptor grid generation module of the Glide tool. The coordinate (x, y, z) of the grid was centered to -9.07, 27.02, and -0.91 respectively. The refined M.oleifera ligands were docked into the active site of BRCA-1.
The energy calculation was achieved using the scoring function of the Glide tool. The drug-like properties of the compounds were evaluated using the QikProp module and SWISS ADME Web tool following Lipinski's rule of five [22].

RESULTS AND DISCUSSION
Molecular docking was employed to carry out the virtual screening of the library of phytochemicals from Moringa oleifera against the targeted protein (BRCA-1). The phytocompounds of M.oleifera were ranked according to their binding poses and energy calculations.
The compounds were further subjected to pharmacokinetic study to predict their drug-able properties. The molecular docking analysis which includes: binding affinity (Kcal/mol) predication, the interaction of the ligands within the binding pocket of BRCA-1, and their pharmacokinetic study was shown (Table 1). Each ligand was analyzed using Lipinski's rule of five (ROF), the result confirms that the ligands ROF with few violations. The ligand docking shows how the phyto-compounds bind effectively with BRCA-1. Visualization of the proteinligand complex was carried out using the surface module of the Glide tool ( Figure 1). The interaction between the compounds and BRCA-1 identified the amino acid residues involved in the interaction as well as the position of each amino acid residues in their ligand-binding site.
The interaction was associated with a structure-based drug design depicting protein-ligand interaction.  properties. Thus, we proposed that the phytochemicals from M. oleifera may be potential BRCA-1 inhibitors. Further biochemical analysis such as invitro and invivo study is required to establish the pharmacological properties of the compounds.