Potential use of compounds from sea cucumbers as MDM2 and CXCR4 inhibitors to control cancer cell growth
- Authors:
- Published online on: August 7, 2018 https://doi.org/10.3892/etm.2018.6588
- Pages: 2985-2991
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Copyright: © Wargasetia et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
Abstract
Introduction
The prevalence of cancer is increasing rapidly (1) and research is focusing on the exploration of novel anticancer treatments. Although the field is growing rapidly, very few cancer drugs are able to pass clinical trials (2). There are numerous types of cancer cells, which are characterized based on either the source of the cell or the development of cells due to gene mutations (3). Various types of drugs may be required for treatments based on the particular characteristics of the cancer cells. As a result, novel anticancer treatments from plants and marine invertebrates, including sea cucumbers were explored.
In general, in vitro and in vivo studies involving sea cucumbers have primarily focused on the toxicity of active compounds on cancer cells by induction of apoptosis or cell cycle arrest (4). Compounds from sea cucumbers exhibiting anticancer properties have been reported (5), including colochiroside A from Colochirus anceps (6), ds-echinoside A from Pearsonothuria graeffei (7), philinopside E from Pentacta quadrangularis (8), sphingosine from Stichopus variegatus (9) and stichoposide C from Thelenota anax (10). However, the mechanisms of action controlling cancer cell growth at a molecular level remain unclear. The current study analyzed the potential of these compounds as inhibitors of mouse double minute 2 homolog (MDM2) and C-X-C chemokine receptor type 4 (CXCR4). The inhibition of these two targets simultaneously may induce a synergistic effect, increasing treatment efficacy.
MDM2 serves a role in binding pro-apoptotic tumor protein 53 (p53) and degrading it (11). Inhibiting the activity of MDM2 may increase p53 levels in the cell, which are necessary for apoptosis (11,12). CXCR4 belongs to the G-protein-coupled receptor family that is involved in several pathways associated with cancer and serves a role in controlling cell proliferation (13). CXCR4 promotes survival of various cell types (14) and serves a critical role in tumorigenesis (15). Furthermore, it acts as receptor for the C-X-C motif chemokine ligand 12 (CXCL12) which serves a role in signal transduction for calcium uptake and enhances the activity of mitogen-activated protein kinase (MAPK)1/MAPK3 (15,16). CXCR4 was reported to be a potent inducer of apoptosis in acute myeloid leukemia cell lines (14). The protein has been a target in drug development (17) and cancer treatment (18,19), and anti-CXCR4 antibodies were demonstrated to induce apoptosis in hematologic malignancies (15). The current study describes the potential of compounds from sea cucumbers as MDM2 and CXCR4 inhibitors, aiming to reveal novel insight into the mechanisms of inhibiting cancer cell growth.
Materials and methods
Preparation of molecule structures and codes
The ligands used for docking analysis were colochiroside A, ds-echinoside A, philinopside E, sphingosine, stichoposide C, 1-(5-chloro-2-methylphenyl)-5-(3-chlorophenyl)-2-(3-methylphenyl)-1H-imidazole-4-carboxylic acid, a tetra-substituted imidazole (an MDM2 inhibitor) and chalcone-4 (a CXCR4 inhibitor). SMILES codes of the compounds were converted to 3D structures in Protein Data Bank (PDB) format using BIOVIA Discovery Studio 4.5 (20). These structures were used for ligand docking. The 3D structure for chalcone-4 was obtained from the binding database (https://www.bindingdb.org/bind/index.jsp) (21) and the 3D structure for the substituted imidazole was obtained from the PDB (PDB ID, 4OQ3). The receptor structures were retrieved from the PDB for MDM2 (PDB ID, 4OQ3) and CXCR4 (PDB ID, 3OE6). The proteins then were prepared by BIOVIA Discovery Studio.
Ligand docking studies
Interactions between receptors and ligand were analyzed by AutoDock Vina integrated in PyRx 0.8 (https://pyrx.sourceforge.io) (22,23). The docking method was used to evaluate binding affinities and to elucidate molecular mechanisms, and was performed according to previous literature (24). Docking was performed by setting receptors as rigid molecules and ligands as flexible molecules within the active site. Results of docking and bonding interactions were analyzed by BIOVIA Discovery Studio (20).
Protein-protein interactions and networks
Proteins that interact with CXCR4 were identified using BioGRID database (https://thebiogrid.org/) (25). Protein-protein interaction networks were examined using STRING (https://string-db.org/) (26).
Pathway analysis
Pathway analysis for CXCR4 was performed using Kyoto Encyclopedia Gene and Genome (KEGG; http://www.genome.jp/kegg/) (27). The role of CXCR4 proteins in various molecular pathways was identified using KEGG pathways databases with STRINGdb 10.5 software. The database covers a range of pathways that have been used as references for the determination of gene or protein function within a cell (28).
Results
Ligand docking analysis
The results of the docking between CXCR4 or MDM2 with the five compounds identified in sea cucumbers revealed that four of the compounds (Ds-echinoside A, Philinopside E, Stichoposide C and Colochiroside A, with values of −9.0, −8.5, −9.2 and −8.5 kcal/mol, respectively) exhibited higher binding affinities to CXCR4 compared with a known inhibitor (chalcone; −7.1 kcal/mol; Table I). Additionally, two compounds (Ds-echinoside A and Philinopside E) were identified to potentially bind to MDM2, with binding affinities of −7.1 and −7.5 kcal/mol, respectively. The compounds (Ds-echinoside A and Philinopside E) were predicted to inhibit MDM2 and to exhibit binding energies higher than its inhibitor, imidazole. However, the binding energies of these two molecules to MDM2 were similar to those of chalcone bound to CXCR4 (−7.1 kcal/mol), but lower when compared with protease bound to its inhibitor (−7.0 kcal/mol) (29) and to the coline receptor bound to its ligand (−6.0 kcal/mol) (30). Sphingosine exhibited lowest binding affinities for CXCR4 and MDM2 (−5.8 kcal/mol and −5.2 kcal/mol, respectively).
Table I.Binding affinity between compounds from sea cucumbers and CXCR4 (PDB ID, 3OE6) or MDM2 (PDB ID, 4OQ3). |
Further analysis focused on evaluating the orientation of the compounds when interacting with the active site of MDM2. This analysis describes a critical part in assessing the potential of a compound for inhibiting MDM2. It was demonstrated that two compounds (Ds-echinoside A and Philinopside E) from sea cucumbers bound to the active site of MDM2 in a similar position to the known inhibitor, a substituted imidazole (Fig. 1). The data suggested that philinopside E and ds-echinoside A, extracted from Pentacta quadrangularis and Pearsonothuria graeffei, respectively, may have potential as MDM2 inhibitors.
The binding of ds-echinoside A, philinopside E, stichoposide C and colochiroside A to CXCR4 were compared to the binding of chalcone-4, a known CXCR4 inhibitor, to CXCR4 (Fig. 2). It was demonstrated that all compounds bound to the active site of CXCR4 in a similar position to chalcone-4. The data indicated that these compounds may have potential as CXCR4 inhibitors.
Protein interaction and pathway analysis
Furthermore, the binding of proteins to CXCR4 were investigated. The data obtained using BioGRID revealed over 40 proteins interacting with CXCR4 (Fig. 3). This analysis is essential to map and resolve functions of proteins that interact with CXCR4. The data may be used in further pathway analysis to help understand the role of CXCR4 in the mechanism of cancer cell growth regulation. The results of the analysis are summarized as a map of proteins interacting with CXCR4. Identified proteins may serve a role in the pathways involved in the pathomechanism of cancer. The proteins are involved in two pathways associated with cancer cell signaling, including the chemokine and Janus kinase (JAK) signal transducer and activator of transcription (STAT) signaling pathway (Fig. 4). A minimum of 15 proteins participates in these pathways, including protein tyrosine kinase 2 (PTK2), C-C chemokine receptor type 5, JAK2, JAK3, β-adrenergic receptor kinase 2, CXCR5, CXCL12, CXCR4, protein tyrosine phosphatase (PTP)6, PTPN11, suppressor of cytokine signaling 3, interleukin-28 α receptor and signal transducing adapter molecule 1. These pathways serve a role in cell proliferation, angiogenesis, cell growth, and metastasis (31). Inhibition of these two pathways may inhibit cancer cell growth and induce apoptosis.
The results of the current study indicated that two out of the five chosen compounds, philinopside E and ds-echinoside A, may inhibit MDM2 and CXCR4 (Fig. 5). Two other compounds, stichoposide C and colochiroside A, were predicted to inhibit CXCR4. The data suggested that philinopside E and ds-echinoside A may exhibit higher efficiencies due to inhibiting two targets simultaneously. Inhibition of MDM2 may trigger apoptosis through p53 activation (32) and inhibition of CXCR4 may affect cell proliferation and growth through JAK2/3-STAT and PTK2 signalling pathways.
Discussion
Previous studies have demonstrated that sea cucumbers contain compounds which exhibit anticancer properties and are described as beneficial agents for human health (33). However, mechanisms of action explaining the anticancer properties remain unclear. The current study analyzed the anticancer mechanisms of compounds from five species of sea cucumber (Table II) using an in silico approach. These compounds included colochiroside A, ds-echinoside A, philinopside E, sphingosine and stichoposide C. Molecular docking was conducted to examine the binding affinity between these compounds and MDM2 or CXCR4. MDM2 has been used as a target in cancer therapy (32). It serves a role in the degradation of p53, a pro-apoptotic protein (32). CXCR4 has also been described as a target in the development of cancer treatments (13). The protein is a receptor that regulates cell cycle, cell proliferation, metastasis and angiogenesis (34).
Details on mechanisms of action for compounds from sea cucumbers as anticancer treatments remain limited. An in-depth docking analysis was conducted to evaluate the potential of compounds from sea cucumbers as anticancer treatments through inhibition of MDM2 and CXCR4. The findings indicated that four out of the five chosen compounds from sea cucumbers are predicted to inhibit CXCR4 and two of these further inhibit MDM2. However, based on the in silico analysis performed in the current study, sphingosine may not be a suitable inhibitor for CXCR4 or MDM2.
The data obtained from pathway analysis suggested that the studied compounds may inhibit cancer cell growth through the chemokine and JAK-STAT signaling pathway or p53 pathway (4). These three pathways serve a central role in the process of controlling cell cycle, migration and apoptosis (13,31,32). Therefore, the data indicated that the anticancer mechanism of the active compound of sea cucumber occurs through inference with the JAK-STAT and Chemokine signaling pathways.
CXCR4 is a receptor located on the cell surface, functioning as a communicator between cells and their environment (14). The receptor binds chemokines and other growth factors, which then transmit signals into cells through multiple pathways, including JAK2/3 and PTK2, which regulate cell division (35). By interrupting the signal transmission through CXCR4, inhibition of cancer cell growth may occur. CXCR4 is known to bind CXCL12, which influences calcium uptake and enhances MAPK1/MAPK3 (15,16) and may be a suitable target in cancer treatment (17,18).
In conclusion, the current study suggested that several compounds from sea cucumbers may have potential as MDM2 or CXCR4 inhibitors. Philinopside E and ds-echinoside A were predicted as MDM2 and CXCR4 inhibitors, while stichoposide C and colochiroside A were predicted as CXCR4 inhibitors. The compounds may be able to inhibit MDM2 and CXCR4 and induce apoptosis in cancer cells. Further research should be conducted in vitro to validate the activity of the studied compounds.
Acknowledgements
The authors would like to thank the Ministry of Research, Technology and Higher Education for supporting the present study.
Funding
The present study was funded by the Ministry of Research, Technology and Higher Education, Republic of Indonesia (grant no. 1598/K4/KM/2017).
Availability of data and materials
All data generated or analyzed during this study are included in this published article.
Authors' contributions
SP and NW designed the study. TWL and NW conducted the research and prepared the manuscript.
Ethics approval and consent to participate
Not applicable.
Patient consent for publication
Not applicable.
Competing interests
The authors declare that they have no competing interests.
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