Evaluation of the in vitro Antitumor Activity of Nanostructured Cyclotides in Polymers of Eudragit® L 100-55 and RS 30 D

Author(s): Osmar N. Silva, Michelle F.S. Pinto, Juliane F.C. Viana, Camila G. Freitas, Isabel C.M. Fensterseifer, David J. Craik, Octavio L. Franco*.

Journal Name: Letters in Drug Design & Discovery

Volume 16 , Issue 4 , 2019

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Abstract:

Background: Cancer is a major cause of mortality and morbidity and given the limitations of many current cancer drugs, there is great need to discover and develop novel treatments. An alternative to the conventional drug discovery path is to exploit new classes of natural compounds such as cyclotides. This peptide family is characterized by linked C- and N-termini and a structural fold called the cyclic cystine knot (CCK). The CCK fold is responsible for the exceptional enzymatic, chemical and thermal stability of cyclotides.

Methods: In the present study, an alternative to traditional cancer treatments, involving new nanomaterials and nanocarriers allowing efficient cyclotide delivery, is proposed. Using the polymers Eudragit® L 100-55 and RS 30 D, the cyclotides kalata B2 and parigidin-br1 (PBR1) were nanocapsulated, and nanoparticles 91 nm and 188 nm in diameter, respectively, were produced.

Results: An encapsulation rate of up to 95% was observed. In vitro bioassays showed that the nanostructured cyclotides were partially able to control the development of the colorectal adenocarcinoma cell line CACO2 and the breast cancer cell line MCF-7.

Conclusion: Data reported herein indicate that nanoformulated cyclotides exhibit antitumor activity and sustained drug release. Thus, the system using Eudragit® nanocapsules seems to be efficient for cyclotide encapsulation and probably could be used to target specific tumors in future studies.

Keywords: Antitumor activity, cyclotide, drug release, nanocapsules, methacrylate, adenocarcinoma.

[1]
Siegel, R.L.; Miller, K.D.; Jemal, A. Cancer Statistics, 2015. CA Cancer J. Clin., 2015, 65(1), 5-29.
[2]
Islami, F.; Torre, L.A.; Jemal, A. Global trends of lung cancer mortality and smoking prevalence. Transl. Lung Cancer Res., 2015, 4(4), 327-338.
[3]
Siegel, R.; DeSantis, C.; Virgo, K.; Stein, K.; Mariotto, A.; Smith, T.; Cooper, D.; Gansler, T.; Lerro, C.; Fedewa, S. Cancer treatment and survivorship statistics, 2012. CA Cancer J. Clin., 2012, 62(4), 220-241.
[4]
Brock, A.; Krause, S.; Ingber, D.E. Control of cancer formation by intrinsic genetic noise and micro-environmental cues. Nat. Rev. Cancer, 2015, 15(8), 499-509.
[5]
Coburn, J.M.; Kaplan, D.L. Engineering biomaterial-drug conjugates for local and sustained chemotherapeutic delivery. Bioconjug. Chem., 2015, 26(7), 1212-1223.
[6]
Colgrave, M.L.; Craik, D.J. Thermal, chemical, and enzymatic stability of the cyclotide kalata B1: The importance of the cyclic cystine knot. Biochemistry, 2004, 43(20), 5965-5975.
[7]
Craik, D.J.; Swedberg, J.E.; Mylne, J.S.; Cemazar, M. Cyclotides as a basis for drug design. Expert Opin. Drug Discov., 2012, 7(3), 179-194.
[8]
Puttamadappa, S.S.; Jagadish, K.; Shekhtman, A.; Camarero, J.A. Backbone dynamics of cyclotide MCoTI-I free and complexed with trypsin. Angew. Chem. Int. Ed. Engl., 2010, 49(39), 7030-7034.
[9]
Daly, N.L.; Thorstholm, L.; Greenwood, K.P.; King, G.J.; Rosengren, K.J.; Heras, B.; Martin, J.L.; Craik, D.J. Structural insights into the role of the cyclic backbone in a squash trypsin inhibitor. J. Biol. Chem., 2013, 288(50), 36141-36148.
[10]
Craik, D.J.; Mylne, J.S.; Daly, N.L. Cyclotides: Macrocyclic peptides with applications in drug design and agriculture. Cell. Mol. Life Sci., 2010, 67(1), 9-16.
[11]
Gran, L.; Sandberg, F.; Sletten, K. Oldenlandia affinis (R&S) DC: A plant containing uteroactive peptides used in african traditional medicine. J. Ethnopharmacol., 2000, 70(3), 197-203.
[12]
Huang, Y-H.; Colgrave, M.L.; Daly, N.L.; Keleshian, A.; Martinac, B.; Craik, D.J. The biological activity of the prototypic cyclotide kalata B1 is modulated by the formation of multimeric pores. J. Biol. Chem., 2009, 284(31), 20699-20707.
[13]
Gerlach, S.L.; Rathinakumar, R.; Chakravarty, G.; Göransson, U.; Wimley, W.C.; Darwin, S.P.; Mondal, D. Anticancer and chemosensitizing abilities of cycloviolacin 02 from viola odorata and psyle cyclotides from psychotria leptothyrsa. Biopolymers, 2010, 94(5), 617-625.
[14]
Pinto, M.F.S.; Silva, O.N.; Viana, J.C.; Porto, W.F.; Migliolo, L.; Da Cunha, N.B.; Gomes, N.; Fensterseifer, I.C.M.; Colgrave, M.L.; Craik, D.J. Characterization of a bioactive acyclotide from palicourea rigida. J. Nat. Prod., 2016, 79(11), 2767-2773.
[15]
Troeira, H.S.; Huang, Y-H.; Chaousis, S.; Wang, C.K.; Craik, D.J. Anticancer and toxic properties of cyclotides are dependent on phosphatidylethanolamine phospholipid targeting. ChemBioChem, 2014, 15(13), 1956-1965.
[16]
Huang, Y-H.; Henriques, S.T.; Wang, C.K.; Thorstholm, L.; Daly, N.L.; Kaas, Q.; Craik, D.J. Design of substrate-based BCR-ABL kinase inhibitors using the cyclotide scaffold. Sci. Rep., 2015, 5, 12974.
[17]
Ji, Y.; Majumder, S.; Millard, M.; Borra, R.; Bi, T.; Elnagar, A.Y.; Neamati, N.; Shekhtman, A.; Camarero, J.A. In vivo activation of the p53 tumor suppressor pathway by an engineered cyclotide. J. Am. Chem. Soc., 2013, 135(31), 11623-11633.
[18]
Gerlach, S.L.; Burman, R.; Bohlin, L.; Mondal, D.; Göransson, U. Isolation, characterization, and bioactivity of cyclotides from the micronesian plant Psychotria leptothyrsa. J. Nat. Prod., 2010, 73(7), 1207-1213.
[19]
Thakral, N.K.; Ray, A.R.; Bar-Shalom, D.; Eriksson, A.H.; Majumdar, D.K. The quest for targeted delivery in colon cancer: Mucoadhesive valdecoxib microspheres. Int. J. Nanomedicine, 2011, 6, 1057-1068.
[20]
Henriques, S.T.; Craik, D.J. Cyclotides as templates in drug design. Drug Discov. Today, 2010, 15(1-2), 57-64.
[21]
Pinto, M.F.S.; Fensterseifer, I.C.M.; Migliolo, L.; Sousa, D.A.; de Capdville, G.; Arboleda-Valencia, J.W.; Colgrave, M.L.; Craik, D.J.; Magalhães, B.S.; Dias, S.C. Identification and structural characterization of novel cyclotide with activity against an insect pest of sugar cane. J. Biol. Chem., 2012, 287(1), 134-147.
[22]
Jennings, C.V.; Rosengren, K.J.; Daly, N.L.; Plan, M.; Stevens, J.; Scanlon, M.J.; Waine, C.; Norman, D.G.; Anderson, M.A.; Craik, D.J. Isolation, solution structure, and insecticidal activity of kalata B2, a circular protein with a twist: do möbius strips exist in nature? Biochemistry, 2005, 44(3), 851-860.
[23]
Saúde, A.C.M.; Ombredane, A.S.; Silva, O.N.; Barbosa, J.A.R.G.; Moreno, S.E.; Araujo, A.C.G.; Falcão, R.; Silva, L.P.; Dias, S.C.; Franco, O.L. Clavanin bacterial sepsis control using a novel methacrylate nanocarrier. Int. J. Nanomedicine, 2014, 9(1), 5055-5069.
[24]
Mahmoudi, M. Debugging nano-bio interfaces: Systematic strategies to accelerate clinical translation of nanotechnologies. Trends Biotechnol., 2018, 36(8), 755-769.
[25]
Momenzadeh, S.; Sadeghi, A.; Vatandoust, N.; Salehi, R. Evaluation of in vivo transfection efficiency of eudragit coated nanoparticles of chitosan-DNA: A PH-sensitive system prepared for oral DNA delivery. Iran. Red Crescent Med. J., 2015, 17(4), e16761.
[26]
Mulcahy, L.A.; Pink, R.C.; Carter, D.R.F. Routes and mechanisms of extracellular vesicle uptake. J. Extracell. Vesicles, 2014, 3.
[27]
Benet, L.Z.; Cummins, C.L.; Wu, C.Y. Unmasking the dynamic interplay between efflux transporters and metabolic enzymes. Int. J. Pharm., 2004, 277(1-2), 3-9.
[28]
Silva, O.N.; Fensterseifer, I.C.M.; Rodrigues, E.A.; Holanda, H.H.S.; Novaes, N.R.F.; Cunha, J.P.A.; Rezende, T.M.B.; Magalhães, K.G.; Moreno, S.E.; Jerônimo, M.S.; Bocca, A.L.; Franco, O.L. Clavanin A improves outcome of complications from different bacterial infections. Antimicrob. Agents Chemother., 2015, 59(3)
[29]
Greish, K. Enhanced Permeability and Retention (EPR) effect for anticancer nanomedicine drug targeting. Methods Mol. Biol., 2010, 624, 25-37.
[30]
Yadav, K.S.; Jacob, S.; Sachdeva, G.; Chuttani, K.; Mishra, A.K.; Sawant, K.K. Long circulating PEGylated PLGA nanoparticles of cytarabine for targeting leukemia. J. Microencapsul., 2011, 28(8), 729-742.
[31]
Li, S-D.; Huang, L. Pharmacokinetics and biodistribution of nanoparticles. Mol. Pharm., 2008, 5(4), 496-504.
[32]
Panyam, J.; Williams, D.; Dash, A.; Leslie-Pelecky, D.; Labhasetwar, V. Solid-State solubility influences encapsulation and release of hydrophobic drugs from PLGA/PLA nanoparticles. J. Pharm. Sci., 2004, 93(7), 1804-1814.
[33]
Barry, D.G.; Daly, N.L.; Clark, R.J.; Sando, L.; Craik, D.J. Linearization of a naturally occurring circular protein maintains structure but eliminates hemolytic activity. Biochemistry, 2003, 42(22), 6688-6695.
[34]
Garcia, A.E.; Osapay, G.; Tran, P.A.; Yuan, J.; Selsted, M.E. Isolation, synthesis, and antimicrobial activities of naturally occurring theta-defensin isoforms from baboon leukocytes. Infect. Immun., 2008, 76(12), 5883-5891.
[35]
Chan, L.Y.; Craik, D.J.; Daly, N.L. Dual-Targeting anti-angiogenic cyclic peptides as potential drug leads for cancer therapy. Sci. Rep., 2016, 6, 35347.
[36]
Malagón, D.; Botterill, B.; Gray, D.J.; Lovas, E.; Duke, M.; Gray, C.; Kopp, S.R.; Knott, L.M.; McManus, D.P.; Daly, N.L. Anthelminthic activity of the cyclotides (Kalata B1 and B2) against schistosome parasites. Biopolymers, 2013, 100(5), 461-470.
[37]
Craik, D.J. Circling the enemy: Cyclic proteins in plant defence. Trends Plant Sci., 2009, 14(6), 328-335.
[38]
Paharia, A.; Yadav, A.K.; Rai, G.; Jain, S.K.; Pancholi, S.S.; Agrawal, G.P. Eudragit-Coated pectin microspheres of 5-fluorouracil for colon targeting. AAPS PharmSciTech, 2007, 8(1), 12.
[39]
Mirhosseini, H.; Tan, C.P.; Hamid, N.S.A.; Yusof, S. Effect of arabic gum, xanthan gum and orange oil contents on ζ-potential, conductivity, stability, size index and ph of orange beverage emulsion. Collo. Sur. A Physio. chem. Eng. Asp., 2008, 315(1-3), 47-56.
[40]
Sajja, H.K.; East, M.P.; Mao, H.; Wang, Y.A.; Nie, S.; Yang, L. Development of multifunctional nanoparticles for targeted drug delivery and noninvasive imaging of therapeutic effect. Curr. Drug Discov. Technol., 2009, 6(1), 43-51.
[41]
El-Kamel, A.H.; Sokar, M.S.; Al Gamal, S.S.; Naggar, V.F. Preparation and evaluation of ketoprofen floating oral delivery system. Int. J. Pharm., 2001, 220(1-2), 13-21.
[42]
Yadav, S.K.; Mishra, S.; Mishra, B. Eudragit-based nanosuspension of poorly water-soluble drug: Formulation and In vitro-in vivo evaluation. AAPS PharmSciTech, 2012, 13(4), 1031-1044.
[43]
Salvador-Morales, C.; Zhang, L.; Langer, R.; Farokhzad, O.C. Immunocompatibility properties of lipid-polymer hybrid nanoparticles with heterogeneous surface functional groups. Biomaterials, 2009, 30(12), 2231-2240.
[44]
Moustafine, R.I.; Zaharov, I.M.; Kemenova, V.A. Physicochemical characterization and drug release properties of eudragit E PO/Eudragit L 100-55 interpolyelectrolyte complexes. Eur. J. Pharm. Biopharm., 2006, 63(1), 26-36.
[45]
Tang, J.; Xu, N.; Ji, H.; Liu, H.; Wang, Z.; Wu, L. Eudragit nanoparticles containing genistein: Formulation, development, and bioavailability assessment. Int. J. Nanomedicine, 2011, 6, 2429-2435.
[46]
She, X.; Chen, L.; Velleman, L.; Li, C.; Zhu, H.; He, C.; Wang, T.; Shigdar, S.; Duan, W.; Kong, L. Fabrication of high specificity hollow mesoporous silica nanoparticles assisted by eudragit for targeted drug delivery. J. Colloid Interface Sci., 2015, 445, 151-160.
[47]
Fontana, M.C.; Beckenkamp, A.; Buffon, A.; Beck, R.C.R. Controlled release of raloxifene by nanoencapsulation: Effect on In vitro antiproliferative activity of human breast cancer cells. Int. J. Nanomedicine, 2014, 9, 2979-2991.
[48]
Sun, H.; Liu, D.; Li, Y.; Tang, X.; Cong, Y. Preparation and In vitro/in vivo characterization of enteric-coated nanoparticles loaded with the antihypertensive peptide VLPVPR. Int. J. Nanomedicine, 2014, 9, 1709-1716.
[49]
Petros, R.A.; DeSimone, J.M. Strategies in the design of nanoparticles for therapeutic applications. Nat. Rev. Drug Discov., 2010, 9(8), 615-627.
[50]
Liu, F.; Lizio, R.; Meier, C.; Petereit, H-U.; Blakey, P.; Basit, A.W. A novel concept in enteric coating: A double-coating system providing rapid drug release in the proximal small intestine. J. Control. Release, 2009, 133(2), 119-124.
[51]
Liechty, W.B.; Peppas, N.A. Expert opinion: Responsive polymer nanoparticles in cancer therapy. Eur. J. Pharm. Biopharm., 2012, 80(2), 241-246.
[52]
Gratton, S.E.A.; Ropp, P.A.; Pohlhaus, P.D.; Luft, J.C.; Madden, V.J.; Napier, M.E.; DeSimone, J.M. The effect of particle design on cellular internalization pathways. Proc. Natl. Acad. Sci. USA, 2008, 105(33), 11613-11618.
[53]
Akhgari, A.; Tavakol, A. Prediction of Optimum combination of eudragit RS/eudragit RL/ethyl cellulose polymeric free films based on experimental design for using as a coating system for sustained release theophylline pellets. Adv. Pharm. Bull., 2016, 6(2), 219-225.
[54]
Lindholm, P.; Göransson, U.; Johansson, S.; Claeson, P.; Gullbo, J.; Larsson, R.; Bohlin, L.; Backlund, A. Cyclotides: A novel type of cytotoxic agents. Mol. Cancer Ther., 2002, 1(6), 365-369.
[55]
Tang, J.; Wang, C.K.; Pan, X.; Yan, H.; Zeng, G.; Xu, W.; He, W.; Daly, N.L.; Craik, D.J.; Tan, N. Isolation and characterization of cytotoxic cyclotides from viola Tticolor. Peptides, 2010, 31(8), 1434-1440.
[56]
Peer, D.; Karp, J.M.; Hong, S.; Farokhzad, O.C.; Margalit, R.; Langer, R. Nanocarriers as an emerging platform for cancer therapy. Nat. Nanotechnol., 2007, 2(12), 751-760.


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Article Details

VOLUME: 16
ISSUE: 4
Year: 2019
Page: [437 - 445]
Pages: 9
DOI: 10.2174/1570180815666180801115526
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