In silico analyzing the molecular interactions of plant-derived inhibitors against E6AP, p53, and c-Myc binding sites of HPV type 16 E6 oncoprotein

Document Type: Original article


Department of Biotechnology, Faculty of Biological Science and Technology, University of Isfahan, Isfahan, Iran


Human papillomaviruses (HPV) are a group of strong human carcinogen viruses considered to be the fourth leading cause of mortality among women in the world. HPV is the most important cause of cervical cancer, which is the second most common cancer in women living in low and middle-income countries. To date, there is no effective cure for an ongoing HPV infection; therefore, it is required to investigate anticancer drugs against this life-threatening infection. In this study, we collected more than 100 plant-derivedcompounds with anti-cancer and antiviral potentials from a variety of papers. Smile formats of these compounds (ligand), were harvested from PubChem database and examined based on the absorption, distribution, metabolism, excretion, and toxicity properties by programs such as Swiss ADME, admetSAR, and pkCSM. Twenty compounds, which were likely to be the HPV16E6 inhibitor, were selected for docking calculations. We examined these natural inhibitors against the HPV16 E6 oncogenic protein. Eventually, three of these compounds were used as the most potent inhibitors (Ginkgetin (peculiarly), Hypericin and Apigetrin) were probably used as the possible source of cancer treatment caused by E6 oncoprotein. In this research, we conducted the docking calculations by Autodock 4.2.6 software. Docking analysis showed the interaction of these plant-originated inhibitors with E6AP, p53, and Myc binding sites on the E6 oncoprotein which support the normal function of E6AP, p53, and Myc.


1. Chang Y, Moore PS, Weiss RA. Human oncogenic viruses: nature and discovery. Eng Sci B: Bio Sci 2017;20160264. 372.
2. Weiderpass E. Lifestyle and cancer risk. J Prev Med Public Health. 2010;43:459-471.
3. Anand P, Kunnumakara AB, Sundaram C, Harikumar KB, Tharakan ST, Lai OS, Sung B, Aggarwal BB. Cancer is a preventable disease that requires major lifestyle changes. Pharm Res 2008;25:2097-2116.
4. McLaughlin-Drubin ME, Munger K. Viruses associated with human cancer. Biochim Biophys Acta Mol Basis Dis 2008;1782:127-150.
5. Johari B, Ebrahimi-Rad M, Maghsood F, Lotfinia M, Saltanatpouri Z, Teimoori-Toolabi L, Sharifzadeh Z, Karimipoor M, Kadivar M. Myc decoy oligodeoxynucleotide inhibits growth and modulates differentiation of mouse embryonic stem cells as a model of cancer stem cells. Anti-Cancer Agents Med Chem 2017;17:1786-1795.
6. Johari B, Zargan J. Simultaneous targeted inhibition of Sox2‐Oct4 transcription factors using decoy oligodeoxynucleotides to repress stemness properties in mouse embryonic stem cells. Cell Biol Int 2017;41:1335-1344.
7.  Burd EM. Human papillomavirus and cervical cancer. Clin Microbiol Rev 2003;16:1-17.
8. Alemi M, Mohabatkar H, Behbahani M. In silico comparison of low-and high-risk human papillomavirus proteins. Appl Biochem Biotechnol 2014;172:188-195.
9.  Kjaer SK. Pathology of the cervix. Curr Opin Obstet Gynecol 1992;4:586-593.
10. Merkhofer C, Maslow J. Human Papilloma virus (HPV) infection and non-cervical oncogenic disease states. Virol Mycol 2015;4:2
11. Smola S. Immunopathogenesis of HPV-associated cancers and prospects for immune-therapy. J Viruses 2017;9:254.
12. Mohabatkar H. Prediction of epitopes and structural properties of Iranian HPV-16 E6 by bioinformatics methods. Asian Pac J Cancer Prev 2007;8:602-606.
13. Hoppe-Seyler K, Bossler F, Braun JA, Herrmann AL, Hoppe-Seyler F. The HPV E6/E7 oncogenes: key factors for viral carcinogenesis and therapeutic targets. Trends Microbiol 2018;26:158-168.
14. Rabelo-Santos SH, Termini L, Boccardo E, Derchain S, Longatto-Filho A, Andreoli MA, Costa MC, Nunes RAL, Ângelo-Andrade LAL, Villa LL. Strong SOD2 expression and HPV-16/18 positivity are independent events in cervical cancer. Oncotarget 2018;9:21630.
15. Haedicke J, Iftner T. Human papillomaviruses and cancer. Radiother Oncol 2013;108:397-402.
16. de Martel C, Plummer M, Vignat J, Franceschi S. Worldwide burden of cancer attributable to HPV by site, country and HPV type. Int J Cancer 2017;141:664-670.
17. LaVigne AW, Triedman SA, Randall TC, Trimble EL, Viswanathan AN. Cervical cancer in low and middle income countries: addressing barriers to radiotherapy delivery. Gynecol Oncol Rep 2017;22:16-20.
18. Esmaeili M, Mohabatkar H, Mohsenzadeh S. Using the concept of Chou's pseudo amino acid composition for risk type prediction of human papillomaviruses. J Theor Biol 2010; 263:203-209.
19. Moosavi F, Mohabatkar H, Mohsenzadeh S. Computer-aided analysis of structural properties and epitopes of Iranian HPV-16 E7 oncoprotein. Interdiscip Sci Comput Life Sci 2010;2:367-372.
20. Pinidis P, Tsikouras P, Iatrakis G, Zervoudis S, Koukouli Z, Bothou A, Galazios G, Vladareanu S. Human papilloma virus’ life cycle and carcinogenesis. Maedica 2016;11:48.
21. Pańczyszyn A, Boniewska-Bernacka E, Głąb G. Telomeres and telomerase during human papillomavirus-induced carcinogenesis. Mol Diagn Ther 2018;22:421-430.
22. Beaudenon S, Huibregtse JM. HPV E6, E6AP and cervical cancer. BMC Biochem 2008; 9:S4.
23. Scheffner M, Werness BA, Huibregtse JM, Levine AJ, Howley PM. The E6 oncoprotein encoded by human papillomavirus types 16 and 18 promotes the degradation of p53. Cell 1990;63:1129-1136.
24. Kgatle MM, Spearman CW, Kalla AA, Hairwadzi HN. DNA oncogenic virus-induced oxidative stress, genomic damage, and aberrant epigenetic alterations. Oxid Med Cell Longev 2017;2017:3179421.
25. Howie HL, Katzenellenbogen RA, Galloway DA. Papillomavirus E6 proteins. Virology 2009;384:324-334.
26. Mahesh R, Langat D, Langat D, Barbara E, Cheng A. Protein E6 in high risk human Papillomaviruses. FASEB J 2018;32(Supplement 1):652.23-.23.
27.   Filippova M, Filippov VA, Kagoda M, Garnett T, Fodor N, Duerksen-Hughes PJ. Complexes of human papillomavirus type 16 E6 proteins form pseudo-death-inducing signaling complex structures during tumor necrosis factor-mediated apoptosis. J Virol 2009;83:210-227.
28.   Wongworawat YC, Filippova M, Williams VM, Filippov V, Duerksen-Hughes PJ. Chronic oxidative stress increases the integration frequency of foreign DNA and human papillomavirus 16 in human keratinocytes. Am J Cancer Res 2016;6:764.
29.   Williams VM, Filippova M, Filippov V, Payne KJ, Duerksen-Hughes P. Human papillomavirus type 16 E6* induces oxidative stress and DNA damage. J Virol 2014; 88:6751-6761.
30.   Yim E-K, Park J-S. The role of HPV E6 and E7 oncoproteins in HPV-associated cervical carcinogenesis. Cancer Res Treat 2005;37:319.
31.   Tungteakkhun SS, Duerksen-Hughes PJ. Cellular binding partners of the human papillomavirus E6 protein. Arch Virol 2008;153:397.
32.   Yeo-Teh NS, Ito Y, Jha S. High-risk human papillomaviral oncogenes E6 and E7 target key cellular pathways to achieve oncogenesis. Int J Mol Sci.2018;19(6):1706.
33.   Messa L, Celegato M, Bertagnin C, Mercorelli B, Nannetti G, Palù G, Loregian A. A quantitative LumiFluo assay to test inhibitory compounds blocking p53 degradation induced by human papillomavirus oncoprotein E6 in living cells. Sci Rep 2018;8:1-11.
34.   Miller DM, Thomas SD, Islam A, Muench D, Sedoris K. c-Myc and cancer metabolism. Clin Cancer Res 2012;18:5546‐5553
35.   Katzenellenbogen R. Telomerase induction in HPV infection and oncogenesis. Viruses 2017;9:180.
36.   Katzenellenbogen RA. Activation of telomerase by HPVs. Virus. Res 2017;231:50-55.
37.   Zhang Y, Dakic A, Chen R, Dai Y, Schlegel R, Liu X. Direct HPV E6/Myc interactions induce histone modifications, Pol II phosphorylation, and hTERT promoter activation. Oncotarget 2017;8:96323.
38.   Pettersen EF, Goddard TD, Huang CC, Couch GS, Greenblatt DM, Meng EC, Ferrin TE. UCSF Chimera- a visualization system for exploratory research and analysis. J Comput Chem 2004;25:1605-1612.
39.   Wallace AC, Laskowski RA, Thornton JM. LIGPLOT: a program to generate schematic diagrams of protein-ligand interactions. Protein Eng Des Sel 1995;8:127-134.
40.   Steffen C, Thomas K, Huniar U, Hellweg A, Rubner O, Schroer A. TmoleX—a graphical user interface for TURBOMOLE. J Comput Chem 2010;31:2967-2970.
41.   Forli S, Botta M. Lennard-Jones potential and dummy atom settings to overcome the AUTODOCK limitation in treating flexible ring systems. J Chem Inf Model 2007;47:1481-1492.
42.   Wang Y, Xiao J, Suzek TO, Zhang J, Wang J, Bryant SH. PubChem: a public information system for analyzing bioactivities of small molecules. Nucleic Acids Res 2009;37(Suppl-2):W623-W633.
43.   Pires DE, Blundell TL, Ascher DB. pkCSM: predicting small-molecule pharmacokinetic and toxicity properties using graph-based signatures. J Med Chem 2015;58:4066-4072.
44.   Daina A, Michielin O, Zoete V. SwissADME: a free web tool to evaluate pharmacokinetics, drug-likeness and medicinal chemistry friendliness of small molecules. Sci Rep 2017;7:42717.
45.   Cheng F, Li W, Zhou Y, Shen J, Wu Z, Liu G, Lee PW, Tang Y. admetSAR: a comprehensive source and free tool for assessment of chemical ADMET properties. J Chem Inf Model 2012;52:3099‐3105.
46.   Kelley LA, Sternberg MJ. Protein structure prediction on the Web: a case study using the Phyre server. Nat Protoc 2009;4:363.
47.   Wiederstein M, Sippl MJ. ProSA-web: interactive web service for the recognition of errors in three-dimensional structures of proteins. Nucleic Acids Res 2007;35(Suppl-2):W407-W410.
48.   Lüthy R, Bowie JU, Eisenberg D. Assessment of protein models with three-dimensional profiles. Nature 1992;356:83-85.
49.   Wallner B, Elofsson A. Can correct protein models be identified? Protein Sci 2003;12: 1073-1086.
50.   Laskowski RA, MacArthur MW, Moss DS, Thornton JM. PROCHECK: a program to check the stereochemical quality of protein structures. J Appl Crystallogr 1993;26:283-291.
51.   Colovos C, Yeates TO. Verification of protein structures: patterns of nonbonded atomic interactions. Protein Sci 1993;2:1511-1519.
52.   Lovell SC, Davis IW, Arendall III WB, De Bakker PI, Word JM, Prisant MG, Richardson JS, Richardson DC. Structure validation by Calpha geometry: phi, psi and Cbeta deviation. Proteins 2003;50:437‐450.
53.   Lipinski C. a; Lombardo, F.; Dominy, BW; Feeney, PJ Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings. Adv Drug Deliv Rev 2001;46:3.
54.   Zanier K, Charbonnier S, Sidi AOMhO, McEwen AG, Ferrario MG, Poussin-Courmontagne P, Cura V, Brimer N, Babah KO, Ansari T. Structural basis for hijacking of cellular LxxLL motifs by papillomavirus E6 oncoproteins. Science 2013;339:694-698.
55.   Martinez-Zapien D, Ruiz FX, Poirson J, Mitschler A, Ramirez J, Forster A, Cousido-Siah A, Masson M, Pol SV, Podjarny A. Structure of the E6/E6AP/p53 complex required for HPV-mediated degradation of p53. Nature 2016;529:541-545.
56.   Pol SBV, Klingelhutz AJ. Papillomavirus E6 oncoproteins. Virology 2013;445:115‐137.
57.   Tomaić V. Functional roles of E6 and E7 oncoproteins in HPV-induced malignancies at diverse anatomical sites. Cancers 2016;8:95.
58.   Luo GG, Ou J-hJ. Oncogenic viruses and cancer. Virol Sin 2015;30:83‐84.
59.   Mamgain S, Sharma P, Pathak RK, Baunthiyal M. Computer aided screening of natural compounds targeting the E6 protein of HPV using molecular docking. Bioinformation 2015;11:236‐242.
60.   Kumar S, Jena L, Sahoo M, Nayak T, Mohod K, VARMA AK. The in Silico Approach to Identify a Unique Plant-Derived Inhibitor Against E6 and E7 Oncogenic Proteins of High-Risk Human Papillomavirus 16 and 18. Avicenna J Med Biochem 2016;4:9.
61.   Rizvi SMD, Shakil S, Haneef M. A simple click by click protocol to perform docking: AutoDock 4.2 made easy for non-bioinformaticians. EXCLI J 2013;12:831.
62.   Guedes IA, de Magalhães CS, Dardenne LE. Receptor–ligand molecular docking. Biophys Rev 2014;6:75-87.
63.   Baek SH, Lee JH, Ko JH, Lee H, Nam D, Lee SG, Yang WM, Um JY, Lee J, Kim SH. Ginkgetin blocks constitutive STAT3 activation and induces apoptosis through induction of SHP‐1 and PTEN tyrosine phosphatases. Phytother Res 2016;30:567-576.
64.   Jeon YJ, Jung SN, Yun J, Lee CW, Choi J, Lee YJ, Han DC, Kwon BM. Ginkgetin inhibits the growth of DU− 145 prostate cancer cells through inhibition of signal transducer and activator of transcription 3 activity. Cancer Sci 2015;106:413-420.
65.   Cao J, Tong C, Liu Y, Wang J, Ni X, Xiong MM. Ginkgetin inhibits growth of breast carcinoma via regulating MAPKs pathway. Biomed Pharmacother 2017;96:450-458.
66.   Pan LL, Wu WJ, Zheng GF, Han XY, He JS, Cai Z. Ginkgetin inhibits proliferation of human leukemia cells via the TNF-α signaling pathway. Z Naturforsch C J Biosci 2017; 72:441-447.
67. Lou JS, Bi WC, Chan GK, Jin Y, Wong CW, Zhou ZY, Wang HY, Yao P, Dong TT, Tsim KW. Ginkgetin induces autophagic cell death through p62/SQSTM1-mediated autolysosome formation and redox setting in non-small cell lung cancer. Oncotarget 2017;8:93131.
68. Park Y, Woo SH, Seo SK, Kim H, Noh WC, Lee JK, Kwon BM, Min KN, Choe TB, Park IC. Ginkgetin induces cell death in breast cancer cells via downregulation of the estrogen receptor. Oncol Lett 2017;14:5027-5033.