CHITOSAN NANOPARTICLES MEDIATED DELIVERY OF MIR-106B-5P TO BREAST CANCER CELL LINES MCF-7 AND T47D

  • LEONNY DWI RIZKITA Department of Pharmacology and Therapy, Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Yogyakarta 55281, Indonesia, Master in Biomedical Sciences Program, Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Yogyakarta 55281, Indonesia
  • YSRAFIL Department of Pharmacology and Therapy, Faculty of Medicine, Public Health and Nursing, UniversitasGadjahMada, Yogyakarta 55281, Indonesia, Department of Pharmacy, Health Polytechnic of Gorontalo, Ministry of Health, Gorontalo, Indonesia
  • RONNY MARTIEN Department of Pharmaceutical Chemistry, Faculty of Pharmacy, UniversitasGadjahMada, Yogyakarta 55281, Indonesia
  • INDWIANI ASTUTI Department of Pharmacology and Therapy, Faculty of Medicine, Public Health and Nursing, UniversitasGadjahMada, Yogyakarta 55281, Indonesia

Abstract

Objective: The development of nanomedicine, such as miRNA transfection to cancer cells,has widely gained interest in the past decade. Unfortunately, miRNA tends to decay easily by the cellular enzymatic process and requires a carrier. As a cationic biopolymer, chitosan is widely known as a non-viral vector. However, research about chitosan as a miRNA delivery system remains limited. This study aimed to investigate the effect and characters of synthetic miRNA loaded chitosan nanoparticles on breast cancer cell lines.


Methods: To obtain the nanocomplex, chitosan-antimiR-106b-5p was formulated using natriumtripolyphosphate through ionic gelation methods. The nanochitosan formula was characterized by using gel electrophoresis; Nano Quant for encapsulation of entrapment quantification; morphology appearance as viewed by Scanning Electron Microscope (SEM), nanochitosan size analysis; in vitro analysis using MCF-7 and T47D breast cancer cell lines; in silico prediction of possible gene target; polymerase chain reaction analysis and gel electrophoresis for E2F1/GAPDH expression.


Results: The efficiency entrapment value was 96.7%, particle size analysis was 458±11.79 nm, and polydispersity index (PDI) was 0.65±0.07, with spherical morphology as viewed in SEM. There was no significant difference between the nanochitosan supplemented group and the control group in MCF-7 cells (p=0.067). However, the ratio of E2F1 to GAPDH was significantly lower than the control group after nanochitosan antimiR-106b-5p was loaded at concentration 140 nmol (p=0.022) and 35 nmol (p=0.016).


Conclusion: Our nanochitosan formula is non-toxic to use in MCF-7 cell lines. Most importantly, as the formula was conjugated to synthetic antimiR-106b-5p, the E2F1 expression decreased.

Keywords: Chitosan, Nanoparticle, miR-106b-5p, Breast cancer, E2F1

References

1. Kaban K, Salva E, Akbuga J. In vitro dose studies on chitosan nanoplexes for microRNA delivery in breast cancer cells. Nucleic Acid Ther 2017;27:45–55.
2. Sabit H, Cevik E, Tombuloglu H, Farag K, Said OAM, Shaimaa E, et al. miRNA profiling in MCF-7 breast cancer cells: seeking a new biomarker. J Biomed Sci 2019;8:1–9.
3. Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A. Global cancer statistics 2018:GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin 2018;68:394–424.
4. American Cancer Society. Global cancer facts andfig. 4th edition; 2018.
5. World Health Organization. Breast cancer; 2018. Available from: https://www.who.int/cancer/prevention/diagnosis-screening/breast-cancer/en/ [Last accessed on 26 Aug 2020]
6. Mollaei H, Safaralizadeh R, Rostami Z. MicroRNA replacement therapy in cancer. J Cell Physiol 2019;2:1–16.
7. Nigussie Mekuria A, Degaga Abdi A, Mishore KM. MicroRNAs as a potential target for cancer therapy. J Cancer Sci Ther 2018;10:152–61.
8. Ahmad J, Hasnain SE, Siddiqui MA, Ahamed M, Musarrat J. microRNA in carcinogenesis and cancer diagnostics: new paradigm biogenesis of miRNA. Indian J Med Res 2013;137:680–94.
9. Peng B, Chen Y, Leong KW. microRNA delivery for regenerative medicine. Adv Drug Delivery Rev 2015;88:108–22.
10. Karimi M, Avci P, Ahi M, Gazori T, Hamblin MR, Naderi Manesh H. Evaluation of chitosan-tripolyphosphate nanoparticles as a p-shRNA delivery vector: formulation, optimization and cellular uptake study. J Nanopharmaceutical Drug Delivery 2013;1:1–28.
11. Arya G, Mankamna Kumari R, Sharma N, Gupta N, Chandra R, Nimesh S. Polymeric nanocarriers for site-specific gene therapy. In: Drug targeting and stimuli sensitive drug delivery systems; 2018. p. 689–714.
12. Wicki A, Witzigmann D, Balasubramanian V, Huwyler J. Nanomedicine in cancer therapy: challenges, opportunities, and clinical applications. J Controlled Release 2015;200:138–57.
13. Fluhmann B, Ntai I, Borchard G, Simoens S, Mühlebach S. Nanomedicines: the magic bullets reaching their target? Eur J Pharm Sci 2019;128:73–80.
14. Cao Y, Tan YF, Wong YS, Liew MWJ, Venkatraman S. Recent advances in chitosan-based carriers for gene delivery. Mar Drugs 2019;17:1–21.
15. Martien R, K Irianto ID, Farida V, Purwita Sari D. Perkembangan teknologi nanopartikel sebagai sistem penghantaran obat. Maj Farm 2012;8:133–44.
16. Patra JK, Das G, Fraceto LF, Campos EVR, Rodriguez Torres MDP, Acosta-Torres LS, et al. Nano based drug delivery systems: recent developments and future prospects. J Nanobiotechnol 2018;16:1–33.
17. Bahrami B, Hojjat Farsangi M, Mohammadi H, Anvari E, Ghalamfarsa G, Yousefi M, et al. Nanoparticles and targeted drug delivery in cancer therapy. Immunol Lett 2017;190:64–83.
18. Xu X, Ho W, Zhang X, Bertrand N, Farokhzad O. Cancer nanomedicine: from targeted delivery to combination therapy. Trends Mol Med. 2015;21:223–32.
19. Aftab S, Shah A, Nadhman A, Kurbanoglu S, Ays?l Ozkan S, Dionysiou DD, et al. Nanomedicine: an effective tool in cancer therapy. Int J Pharm 2018;540:132–49.
20. Ishii T, Okahata Y, Sato T. Mechanism of cell transfection with plasmid/chitosan complexes. Biochim Biophys Acta Biomembranes 2001;1514:51–64.
21. Gascon R, del Pozo Rodriguez A, Solinis M. Non-viral delivery systems in gene therapy. In: Gene therapy-tools and potential applications; 2013. p. 3–34.
22. Santos Carballal B, Aaldering LJ, Ritzefeld M, Pereira S, Sewald N. Physicochemical and biological characterization of chitosan-microRNA nanocomplexes for gene delivery to MCF-7 breast cancer cells. Nat Publ Gr 2015;5:13567.
23. Naskar S, Koutsu K, Sharma S. Chitosan-based nanoparticles as drug delivery systems: a review on two decades of research. J Drug Target 2018;27:1-41.
24. Mohammed MA, Syeda JTM, Wasan KM, Wasan EK. An overview of chitosan nanoparticles and its application in non-parenteral drug delivery. Pharmaceutics 2017;9:1–26.
25. Ding Y, Shen SZ, Sun H, Sun K, Liu F, Qi Y, et al. Design and construction of polymerized-chitosan coated Fe3O4 magnetic nanoparticles and its application for hydrophobic drug delivery. Mater Sci Eng C 2015;48:487–98.
26. Denizli M, Aslan B, Mangala LS, Jiang D, Rodriguez Aguayo C, Lopez Berestein G, et al. Chitosan nanoparticles for miRNA delivery. In: Bindewald E, Shapiro B, editors. RNA nanostructures: methods and protocols, methods in molecular Biology. 1st ed. Humana Press; 2017. p. 219–30.
27. Ysrafil Y, Astuti I, Anwar LS, Martien R, Sumadi FAN, Wardhana T, et al. microRNA-155-5p diminishes in vitro ovarian cancer cell viability by targeting HIF1? expression. Adv Pharm Bull 2020;10:630–7.
28. Rahiemna A, Megafitrah M, Ramadhani P, Mustikawaty A, Martien R. Formulasi nanopartikel kitosan-PGV-0 dengan metode ionik gelasi. J Saintifika Gadjah Mada 2011;3:17–22.
29. Adhyatmika A, Martien R, Ismail H. Preparasi nanopartikel senyawa pentagamavunon-0 menggunakan matriks polimer kitosan rantai sedang dan pengait silang natrium tripolifosfat melalui mekanisme gelasi ionik sebagai kandidat obat antiinflamasi. Maj Farm 2018;13:65.
30. Suardi R, Ysrafil Y, Sesotyosari S, Martien R, Wardana T, Astuti I, et al. The effects of combination of mimic miR-155-5p and antagonist miR-324-5p encapsulated chitosan in ovarian cancer SKOV3. Asian Pacific J Cancer Prev 2020;21:2603–8.
31. Nguyen MA, Wyatt H, Susser L, Geoffrion M, Rasheed A, Duchez AC, et al. Delivery of microRNAs by chitosan nanoparticles to functionally alter macrophage cholesterol efflux in vitro and in vivo. ACS Nano 2019;13:6491–505.
32. Riss T, Moravec R, Niles A, Duellman S, Benink H, Worzella T, et al. Cell viability assays. In: Markossian S, Sittampalam GS, Grossman A, Brimacombe K, Arkin M, Auld D, et al. editors. Assay guidance manual. Medical Bethesda (MD): Eli Lilly and Company and the National Center for Advancing Translational Sciences; 2004.p. 295–320.
33. Danaei M, Dehghankhold M, Ataei S, Hasanzadeh Davarani F, Javanmard R, Dokhani A, et al. Impact of particle size and polydispersity index on the clinical applications of lipidic nanocarrier systems. Pharmaceutics 2018;10:1–17.
34. Ragelle H, Vanvarenberg K, Vandarmeulen G, Preat VP. Chitosan nanoparticles for siRNA delivery in vitro. In: Walker JM. editor. Methods in molecular biology (Clifton, NJ). Humana Press; 2016. p. 143–50.
35. Katas H, Alpar HO. Development and characterization of chitosan nanoparticles for siRNA delivery. J Controlled Release 2006;115:216–25.
36. Villegas Peralta Y, Lopez Cervantes J, Madera Santana TJ, Sanchez Duarte RG, Sánchez Machado DI, Martinez Macias M del R, et al. Impact of the molecular weight on the size of chitosan nanoparticles: characterization and its solid-state application. Polym Bull 2020. DOI:10.1007/s00289-020-03139-x
37. Yang HC, Hon MH. The effect of the molecular weight of chitosan nanoparticles and its application on drug delivery. Microchem J 2009;92:87–91.
38. Al-Nemrawi NK, Alsharif SSM, Dave RH. Preparation of chitosan-tpp nanoparticles: the influence of chitosan polymeric properties and formulation variables. Int J Appl Pharm 2018;10:60–5.
39. Hassan NAFA, Sahudin S, Hussain Z, Hussain M. Self-assembled chitosan nanoparticles for percutaneous delivery of caffeine: preparation, characterization and in vitro release studies. Int J Appl Pharm 2018;10:172–85.
40. Katas H, Raja MAG, Lam KL. Development of chitosan nanoparticles as a stable drug delivery system for protein/siRNA. Int J Biomater 2013;4:1–9.
41. Morris GA, Castile J, Smith A, Adams GG, Harding SE. The effect of prolonged storage at different temperatures on the particle size distribution of tripolyphosphate (TPP)-chitosan nanoparticles. Carbohydr Polym 2011;84:1430–4.
42. Kean T, Thanou M. Biodegradation, biodistribution and toxicity of chitosan. Adv Drug Delivery Rev 2010;62:3–11.
43. Stender JD, Frasor J, Komm B, Chang KCN, Kraus WL, Katzenellenbogen BS. Estrogen-regulated gene networks in human breast cancer cells: involvement of E2F1 in the regulation of cell proliferation. Mol Endocrinol 2007;21:2112–23.
44. Laine A, Westermarck J. Molecular pathways: harnessing E2F1 regulation for prosenescence therapy in p53-defective cancer cells. Clin Cancer Res 2014;20:3644–51.
45. Petrocca F, Visone R, Onelli MR, Shah MH, Nicoloso MS, de Martino I, et al. E2F1-regulated microRNAs impair TGF?-dependent cell-cycle arrest and apoptosis in gastric cancer. Cancer Cell 2008;13:272–86.
46. Denechaud PD, Fajas L, Giralt A. E2F1, a novel regulator of metabolism. Front Endocrinol (Lausanne) 2017;8:1–8.
47. Wei CY, Tan QX, Zhu X, Qin QH, Zhu FB, Mo QG, et al. Expression of CDKN1A/p21 and TGFBR2 in breast cancer and their prognostic significance. Int J Clin Exp Pathol 2015;8:14619–29.
48. Louie MC, McClellan A, Siewit C, Kawabata L. Estrogen receptor regulates E2F1 expression to mediate tamoxifen resistance. Mol Cancer Res 2010;8:343–52.
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RIZKITA, L. D., YSRAFIL, MARTIEN, R., & ASTUTI, I. (2021). CHITOSAN NANOPARTICLES MEDIATED DELIVERY OF MIR-106B-5P TO BREAST CANCER CELL LINES MCF-7 AND T47D. International Journal of Applied Pharmaceutics, 13(1), 129-134. https://doi.org/10.22159/ijap.2021v13i1.39749
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