IAJPS 2017, 4 (05), 1141-1167 Patel et al ISSN 2349-7750 CODEN (USA): IAJPBB ISSN: 2349-7750 INDO AMERICAN JOURNAL OF PHARMACEUTICAL SCIENCES Available online at: http://www.iajps.com Research Article - PREDICTIVE MODELS FOR SYSTEM XC ANTIPORTER INHIBITION BASED ON STRUCTURALLY DIVERSE MOLECULES Dhaval Patel1, Mukesh Nandave1 and Prashant S. Kharkar2* 1Department of Pharmacology SPP School of Pharmacy and Technology Management, SVKM's NMIMS, Mumbai, India 2Department of Pharmaceutical Chemistry, SPP School of Pharmacy and Technology Management, SVKM's NMIMS, Mumbai, India Abstract: The morbidity and mortality throughout the world is increasing day by day due to cancer. Several molecular targets - have been identified and being targeted for treatment of cancer cells. System xc , an amino acid antiporter, is one such potential target. With the uptake of one molecule of cystine and release of one molecule glutamate, over expressed - system xc manipulates the redox status within cancer cells and protects them. Simultaneously, released glutamate helps in growth and metastasis of cancer cells. Few researches have synthesized and screened structurally diverse molecules - against system xc antiporter. Amongst these, few molecules like erastin analogues, amino acid analogues, iso-oxazole analogues, hydantoin analogues and sulfasalazine analogues exhibited potent inhibitory activity. It is possible to - identify desirable molecular properties required for system xc inhibition using information from above mentioned molecules. In context, we developed different predictive models using above mentioned analogues using SAS software using regression and decision tree analysis mainly. The score ranking overlay plots showed moderate to good fit of data for the training and validation data sets. These predictive models may further be used for the design and - development of potent system xc inhibitors. - Key words: System xc antiporter, cystine, Glutamate, SAS software, Predictive modelling Corresponding author: Dr. Prashant S. Kharkar, QR code SPP School of Pharmacy and Technology Management, SVKM's NMIMS, Mumbai, India Phone: +91-22-42332000 Ext. 2028 E-mail: [email protected] - Please cite this article in press as Patel et al, Predictive Models for System Xc Antiporter Inhibition Based On Structurally Diverse, Indo Am. J. P. Sci, 2017; 4(05). www.iajps.com Page 1141 IAJPS 2017, 4 (05), 1141-1167 Patel et al ISSN 2349-7750 INTRODUCTION: Overall, erastin series looks more promising over SSZ - - - System xc (Sxc ) antiporter is an amino acid transporter series as Sxc inhibitors. localised on several cell types of the human body [1]. Several other groups synthesized and evaluated - The basic function of Sxc (as observed in several in isoxazole (ISO) [16-18], hydantoin [19] and amino - vitro experiments) is to release glutamate from the acid [20] analogues for Sxc inhibition and reported cells with uptake of cystine simultaneously in promising inhibitory activities for these molecules. The equimolar concentration [2]. By doing so, it initial work on ISO analogues was based on amino contributes in preserving redox status of the cell methylisoxazole propionic acid (AMPA) and amino-3- through the production of glutathione (GSH) [3]. carboxy-5-methylisoxazole propionic acid (ACPA). - Moreover, in brain, it participates in maintaining These molecules were competitive inhibitors of Sxc - neural plasticity through release of an excitatory [16]. The homology model of Sxc and the - neurotransmitter glutamate [4]. Structurally, Sxc computational analysis of the binding mode of consists of two polypeptide chains, the light chain previously reported isoxazole hydrazide inhibitors was called xCT and the heavy chain called 4F2hc. The key elucidated in a subsequent paper [17]. In a related - role of Sxc is performed by the light chain xCT while paper, Newell, et al. reported diaryl isoxazoles as - heavy chain helps in trafficking of light chain [5]. noncompetitive inhibitors of Sxc [18]. This was an However, it was observed in one of the in vitro extension of the previous work based on ACPA experiments that the activity of xCT is lost if heavy analogues. The lead molecules were the most potent - chain is removed [6]. Since the redox status of cancer inhibitors of Sxc discovered so far. cells is manipulated by producing GSH in abundant In an attempt to discover inhibitors for vesicular quantity to protect the cancer cell, the involvement of glutamate transporter (VGLUT), Ahmed, et al. tested - - Sxc in such phenomenon can be outlined in cancer hydantoin analogues for VGLUT and obligate Sxc pathophysiology. It is also evident through several in inhibition. Majority of the inhibitors were potent - vitro experiments that, Sxc is over- expressed in many VGLUT inhibitors whereas only few compounds - cancer cell lines including glial [7], head and neck [8], exhibited Sxc inhibition slightly over 50%. A similar lung [9], breast [10], gastrointestinal [11], pancreatic study, Etoga et al., reported confromationally [12], ovarian [13] and colon [11] cancers. Moreover, constrained amino acid analogues bearing distal - - Sxc is also being explored for its role in CNS sulfonic acid reduced glutamate uptake at Sxc by 70- disorders like addiction, depression, epilepsy, 75% with no effect on glutamate uptake at VGLUT. - schizophrenia, Parkinson’s disease etc. [1]. Overall, These structurally diverse series of Sxc inhibitors can - Sxc is a promising target for the treatment of several greatly aid the design and development of more potent ailments. and selective analogues with the help of conventional - Pharmacological inhibition of Sxc may result in medicinal and computational chemistry approaches disturbed redox status of cancer cells with increased such as quantitative structure-activity relationship reactive oxygen species (ROS) (produced through (QSAR) and others. metabolism) inside the cell causing cell death [2]. In The QSAR is a tool to predict the desirable molecular glioblastoma, the overexpressed Sxc- may release properties of molecules required to exhibit biological elevated glutamate causing excitotoxicity followed by response. Different variables are used in QSAR for the the death of neurons [7]. From the studies it was prediction of biological activity for the designed observed that, two FDA approved drugs sulfasalazine molecules. This potentially helps in rationalizing the (SSZ) and sorafenib were potent inhibitors of Sxc . synthesis of new molecules. Working on the similar Additionally, erastin also exhibited potent activity lines, we began this study with the aim of - against Sxc [14]. However, the enzymatic breakdown understanding the structural features and molecular - of SSZ in stomach, safety issues associated with properties required for Sxc inhibition. Such insights are sorafenib and unavailability of in vivo data for erastin of potential benefit for the design and development of have created hurdles for these potent inhibitors to newer agents devoid of the problems associated with move into higher phases of drug development. Indeed, the earlier molecules. We hereby present our efforts in few researches have synthesized analogues of erastin this direction to develop a robust QSAR model using - - [14] and SSZ [15] and evaluated them against Sxc structurally diverse group of structurally diverse Sxc which showed equivalent or superior potency. The inhibitors. metabolic liability in SSZ, i. e., the diazo group, was mitigated by the relatively stable alkyne group. In MATERIALS AND METHODS: addition, suitable alternative fragments for the distal Data set selection sulfapyridine end were discovered to reduce the The data set used in the present study was prepared - toxicity of SSZ [15]. On the similar lines, extensive from the published literature on Sxc inhibitors. All the structure-activity relationship (SAR) of erastin led to inhibitors belonging to five series - SSZ analogues - the discovery of very potent Sxc inhibitors [14]. [14], erastin derivatives [15], isooxazole analogues Erastin and its analogues, representing altogether a [16-18], hydantoin derivatives [19] and amino acid different chemotype (quinazolin-4-one), are more analogues [20] were compiled (Tables 1-5) for the drug-like than SSZ. These analogues are likely to QSAR study. The inhibitory activities of all included penetrate blood-brain barrier (BBB) unlike SSZ. compounds were measured in terms of their ability to www.iajps.com Page 1142 IAJPS 2017, 4 (05), 1141-1167 Patel et al ISSN 2349-7750 - inhibit cystine/glutamate uptake at Sxc . In case of Various predictive modelling methods were tried for erastin derivatives (Table 1) and SSZ analogues (Table each of the data sources. Since the target (here activity) 5), 50% of inhibitory concentrations (IC50) of is interval target (e. g., pIC50, logit, etc.), appropriate - cystine/glutamate uptake at Sxc in the presence of settings were used for the corresponding nodes in respective inhibitors have been reported. While the SAS® Enterprise Miner™ 13.2. Further details can be inhibitory activity of isooxazole analogues (Table 4), found in the Results and Discussion section. The hydantoin derivatives (Table 3) and amino acid model development process involved mostly trial-and- analogues (Table 2) were reported as % inhibition of error with respect to node settings. Figure 1 depicts this - uptake of cystine/glutamate at Sxc considering 100% process for erastin analogues. Similar diagrams for uptake in untreated controls. The respective