*Article* **Role of Persistent Organic Pollutants in Breast Cancer Progression and Identification of Estrogen Receptor Alpha Inhibitors Using In-Silico Mining and Drug-Drug Interaction Network Approaches**

**Bibi Zainab <sup>1</sup> , Zainab Ayaz <sup>1</sup> , Umer Rashid <sup>2</sup> , Dunia A. Al Farraj <sup>3</sup> , Roua M. Alkufeidy <sup>3</sup> , Fatmah S. AlQahtany <sup>4</sup> , Reem M. Aljowaie <sup>3</sup> and Arshad Mehmood Abbasi 1,5,\***


**Simple Summary:** The role of persistent organic pollutants (POPs) in breast cancer progression and their bioaccumulation in adipose tissue has been reported. We used a computational approach to study molecular interactions of POPs with breast cancer proteins and identified natural and synthetic compounds to inhibit these interactions. Moreover, for comparative analysis, standard drugs and screened compounds were also docked against estrogen receptor alpha (ERα) and identification of the finest inhibitor was performed using in-silico mining and drug-drug interaction (DDI) network approaches. Based on scoring values, short-chained chlorinated paraffins demonstrated strong interactions with ERα compared to organo-chlorines and PCBs. Synthetic and natural compounds demonstrating strong associations with the active site of the ERα protein could be potential candidates to treat breast cancer specifically caused by POPs and other organic toxins and can be used as an alternative to standard drugs.

**Abstract:** The strong association between POPs and breast cancer in humans has been suggested in various epidemiological studies. However, the interaction of POPs with the ERα protein of breast cancer, and identification of natural and synthetic compounds to inhibit this interaction, is mysterious yet. Consequently, the present study aimed to explore the interaction between POPs and ERα using the molecular operating environment (MOE) tool and to identify natural and synthetic compounds to inhibit this association through a cluster-based approach. To validate whether our approach could distinguish between active and inactive compounds, a virtual screen (VS) was performed using actives (627 compounds) as positive control and decoys (20,818 compounds) as a negative dataset obtained from DUD-E. Comparatively, short-chain chlorinated paraffins (SCCPs), hexabromocyclododecane (HBCD), and perfluorooctanesulfonyl fluoride (PFOSF) depicted strong interactions with the ERα protein based on the lowest-scoring values of −31.946, −18.916, −17.581 kcal/mol, respectively. Out of 7856 retrieved natural and synthetic compounds, sixty were selected on modularity bases and subsequently docked with ERα. Based on the lowest-scoring values, ZINC08441573, ZINC00664754, ZINC00702695, ZINC00627464, and ZINC08440501 (synthetic compounds), and capsaicin, flavopiridol tectorgenin, and ellagic acid (natural compounds) showed incredible interactions with the active sites of ERα, even more convening and resilient than standard breast cancer drugs Tamoxifen, Arimidex and Letrozole. Our findings confirm the role of POPs in breast cancer progression and suggest that natural and synthetic compounds with high binding affinity could be

**Citation:** Zainab, B.; Ayaz, Z.; Rashid, U.; Al Farraj, D.A.; Alkufeidy, R.M.; AlQahtany, F.S.; Aljowaie, R.M.; Abbasi, A.M. Role of Persistent Organic Pollutants in Breast Cancer Progression and Identification of Estrogen Receptor Alpha Inhibitors Using In-Silico Mining and Drug-Drug Interaction Network Approaches. *Biology* **2021**, *10*, 681. https://doi.org/10.3390/ biology10070681

Academic Editors: Shibiao Wan, Yiping Fan, Chunjie Jiang, Shengli Li and Lucia Mangone

Received: 2 June 2021 Accepted: 8 July 2021 Published: 19 July 2021

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**Copyright:** © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

more efficient and appropriate candidates to treat breast cancer after validation through in vitro and in vivo studies.

**Keywords:** breast cancer; estrogen receptor alpha; persistent organic pollutants; drug-drug interaction networks; molecular docking

#### **1. Introduction**

POPs are the most common synthetic, lipophilic, toxic, bio-accumulative, and persistent pollutants in the environment. Most POPs are of anthropogenic origin, but some substances, i.e., dioxins and furans, are also produced naturally during volcanism. POPs are also used intentionally in pesticides and other industrial products and may be released accidentally as a by-product from industrial processes or fuel combustion, such as dioxins and furans [1]. POPs release in the environment through industrial and agricultural effluents, drainage systems, urban effluents and landfill leachate [2,3]. Contaminated soil, water, air, dust and processed goods like textiles and packaging materials contain considerable amounts of POPs. Importantly, at ambient temperatures, POPs have a tendency to enter the gas phase; as a result they may volatilize from soils, plants, and water bodies into the atmosphere. They preferentially partition to solids, particularly organic materials in aquatic systems and soils, avoiding the aqueous phase. Being hydrophobic in nature [4], rather than entering the aqueous milieu of cells, some major types of POPs, such as polychlorinated dibenzo-p-dioxins and furans (PCDD/PCFs), polychlorinated biphenyl (PCBs), organo-chlorinated pesticides (OCPs), perfluorooctane sulfonate (PFOS) and pentadecafluorooctanoic acid (PFOAs) are hydrophobic and accumulate in the fatty tissues of the living host. POPs may accumulate in food chains [5] and, from contaminated food such as fruits, vegetables, chicken, meat, milk and fish etc., may enter humans and other living organisms [6,7]. As a result, predatory species like humans often have the highest concentration of POPs, and their presence in humans, i.e., in adipose tissue and human milk, is associated with the up-regulation of hormone-dependent breast cancers [2].

The prevalence of breast cancer, one of the most common types of cancers, specifically in females, is increasing worldwide, which cannot be explained solely by the emergence of mammography screening [8]. In 2018, about two million cases of breast cancer were reported in women globally [9]. The survival rate was up to 26% in cases where distant metastases were present. About 25% of breast cancers have been reported in developed countries; furthermore, it is one of the leading causes of death in Western countries. Deregulation of estrogen balance is known to promote breast cancer, and in Asia, over 60% of breast cancer cases have been diagnosed as estrogen receptor alpha-positive (ERα) cancers [10]. The estrogen receptor 1 (ESR1) gene encodes the ERα protein, a ligand regulated transcription factor, which plays a central role in the proliferation of breast cancer. Production of testosterone enhances the synthesis of progesterone and estrogen receptors in breast glands. Particularly, ERα expressed in the mammary glands and uterus of women has binding ability with DNA and contributes significantly to apoptosis, homeostasis, metabolism, and in breast cancer. An estimated 60% pre- and 75% postmenopausal women are suffering from estrogen-dependent breast cancer [11]. Through disturbing the functioning of adipose tissue, POPs affect the production of estrogens by stimulating genotoxic enzymes and leading to cross-generational epigenetic modifications by modifying the epigenome [12]. Many in vitro studies have shown that certain POPs promote the development of estrogen-positive breast cancer cells by receptor (ER). Exposure to certain POPs, particularly in perinatal studies, can enhance the development of breast cancer and sensitivity to carcinogens and cancerous breast tumors in animal studies. Chemotherapy, hormone therapy, immunotherapy, radiotherapy, and surgery are among the common methods for breast cancer treatment [13], which eventually have multiple

side effects. Therefore, it is necessary to find better natural and synthetic compounds for treatment.

In this context, extensive use of anticancer drugs and potential inhibitors with increasing resistance together with numerous side effects highlights an urgent need for novel cancer treatment methods. Therefore, VS methods including negative image-based screening, molecular docking and the pharmacophore hypothesis could be effective tools for identification and screening of the ligands against ER-α receptor. Recent studies have demonstrated that VS methods have the ability to provide structural insights into complex interactions for repositioning and remediation [14], specifically using natural and synthetic compounds [15]. At present, in-silico methods for drug designing, receptor mapping, molecular modeling, and homology modulation etc. are gaining tremendous popularity in drug development, molecular biology, nanotechnology and biochemistry domains. In addition, these methods are used to complement in vitro and in vivo toxicity assessments, particularly to reduce the need for animal monitoring, costs, and time [16]. Furthermore, in-silico cancer modelling opens up new avenues for research into oncogenesis in different biological dimensions and systems. These approaches can assist in expediting the development of diagnostic and therapeutic technologies for clinical care. With reliable digital representations of cancer, the consequences of therapeutic treatments at both the molecular and surgical scales may be anticipated in silico without exposing patients to danger. Previously, an in-silico drug discovery technique exposed that a potential ligand, 1,2,3,4,6-penta-O-galloyl-β-D-glucopyranose, which is a naturally occurring tannin, can inhibit the activity of Ror1 (protein) that contributes significantly to cancer growth and proliferation [17].

Many complementary resources, including microarray, protein-protein interaction, and protein complexes, are being used to discover enriched biological processes and pathways. One example of this is graph theory, which is being used to analyze the lung cancer protein-protein interaction network (PPIN), and to discover highly dense modules which are potential cancer-associated protein complexes [18]. Previously, flavonoids have been proven as potential anticancer agents by virtue of molecular binding to some key targets such as aromatase, fatty acid synthase, xanthine oxidase, cyclooxygenase, lipoxygenase, ornithine decarboxylase, protein tyrosine kinase, phosphoinositide 3-kinase, protein kinase C, topoisomerase II (ATP binding site), ATP binding cassette (ABC) transporter, and phospholipase A2 [19]. The present study was conducted with the aim of determining the molecular interactions between ERα (target) and POPs which were considered as key factors in breast cancer progression. Moreover, for comparative analysis, standard drugs and screened compounds were docked against ERα and the finest inhibitors (natural and synthetic compounds) were identified using in-silico mining and DDI network approaches.

## **2. Materials and Methods**
