In vitro transfection potential of fluorinated G5 PAMAM dendrimers for miRNA delivery to MRC-5 cells

Year 2018, Volume: 4 Issue: 2, 92 - 100, 04.04.2018

Ali Öztuna , Hasan Nazır

https://doi.org/10.18621/eurj.346612

Cited By: 2

Abstract

Objectives. MicroRNAs (miRNAs) are involved in the regulation of
most biological processes and also contribute to many types of disease.
Fibroblast cells, such as MRC-5, are often used in biological researches
utilizing cell transfection methods due to their difficult to transfect nature.
Cells can be genetically engineered by using viral and non-viral methods.
Poly(amidoamine) (PAMAM) dendrimers are very promising alternative as a
delivery vehicle due to their well-defined
characteristics. In this study, in vitro
transfection potential of cystamine core generation five (G5) PAMAM dendrimers
fluorinated with 2,3,4,5,6-pentafluorobenzoic acid (PFB) and pentafluoropropionic
acid (PFP) for miRNA delivery to MRC-5 cells was examined. Methods. Spectroscopic
techniques were used in the characterization of the prepared dendrimers. miRNA
binding and condensation capability of dendrimers was examined by gel
retardation assay. Characterization of dendriplexes was made by zeta potential,
particle size measurements and transmission electron microscopy. Transfection
efficiencies of the dendriplexes were determined by flow cytometry and
intracytoplasmic distribution of the dendriplexes was shown by laser scanning
confocal microscopy. Also, quantitative structure-activity relationship and molecular
docking calculations were used to be able to discuss transfection efficiencies
of the dendriplexes into the cell. Results. While high level of viability
on MRC-5 cells was observed for dendriplexes prepared with PFB and PFP,
transfection efficiency with PFP was higher than PFB. Transfection efficiency
difference between these two compounds was attributed to their molecular
structures. Conclusions. Obtained results hold promise for the usage of
these compounds as a transfection reagent at MRC-5 cells. Further studies are
needed to support these findings. 

Keywords

Cystamine-G5-PAMAM, pentafluorobenzoic acid, pentafluoropropionic acid, miRNA, MRC-5

References

• [1] Bartel DP. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 2004;116:281-97.
• [2] Shah MY, Ferrajoli A, Sood AK, Lopez-Berestein G, Calin GA. microRNA therapeutics in cancer - An emerging concept. EBioMedicine 2016;12:34-42.
• [3] Gary DJ, Puri N, Won YY. Polymer-based siRNA delivery: perspectives on the fundamental and phenomenological distinctions from polymer-based DNA delivery. J Control Release 2007;121:64-73.
• [4] Abbasi E, Aval SF, Akbarzadeh A, Milani M, Nasrabadi HT, Joo SW, et al. Dendrimers: synthesis, applications, and properties. Nanoscale Res Lett 2014;9:247.
• [5] Daneshvar N, Abdullah R, Shamsabadi FT, How CW, Mh MA, Mehrbod P. PAMAM dendrimer roles in gene delivery methods and stem cell research. Cell Biol Int 2013;37:415-19.
• [6] Kaur D, Jain K, Mehra NK, Kesharwani P, Jain NK. A review on comparative study of PPI and PAMAM dendrimers. J Nanopart Res 2016;18:146.
• [7] van der Aa LJ, Vader P, Storm G, Schiffelers RM, Engbersen JF. Optimization of poly(amido amine)s as vectors for siRNA delivery. J Control Release 2011;150:177-86.
• [8] Kesharwani P, Gajbhiye V, Jain NK. A review of nanocarriers for the delivery of small interfering RNA. Biomaterials 2012;33:7138-50.
• [9] Kesharwani P, Banerjee S, Gupta U, Amin MCIM, Padhye S, Sarkar FH, et al. PAMAM dendrimers as promising nanocarriers for RNAi therapeutics. Materials Today 2015;18:565-72.
• [10] Oupický D, Li J. Bioreducible polycations in nucleic acid delivery: past, present, and future trends. Macromol Biosci 2014;14:908-22.
• [11] Liu X, Li G, Su Z, Jiang Z, Chen L, Wang J, et al. Poly(amido amine) is an ideal carrier of miR-7 for enhancing gene silencing effects on the EGFR pathway in U251 glioma cells. Oncol Rep 2013;29:1387-94.
• [12] Rahbek UL, Nielsen AF, Dong M, You Y, Chauchereau A, Oupicky D, et al. Bioresponsive hyperbranched polymers for siRNA and miRNA delivery. J Drug Target 2010;18:812-20.
• [13] Wang M, Liu H, Li L, Cheng Y. A fluorinated dendrimer achieves excellent gene transfection efficacy at extremely low nitrogen to phosphorus ratios. Nat Commun 2014;5:3053.
• [14] Liu H, Wang Y, Wang M, Xiao J, Cheng Y. Fluorinated poly(propylenimine) dendrimers as gene vectors. Biomaterials 2014;35:5407-13.
• [15] Tomalia DA, Huang B, Swanson DR, Brothers II HM, Klimash JW. Structure control within poly(amidoamine) dendrimers: size, shape and regio-chemical mimicry of globular proteins. Tetrahedron 2003;59:3799-813.
• [16] Duhovny D, Nussinov R, Wolfson HJ. Efficient unbound docking of rigid molecules. In: Guigó R, Gusfield D, eds. Algorithms in Bioinformatics. WABI. Lecture Notes in Computer Science. Springer 2002;2452:185-200.
• [17] Schneidman-Duhovny D, Inbar Y, Nussinov R, Wolfson HJ. PatchDock and SymmDock: servers for rigid and symmetric docking. Nucleic Acids Res 2005;33 (Web Server issue):W363-7.
• [18] Gao H, Shi W, Freund LB. Mechanics of receptor-mediated endocytosis. Proc Natl Acad Sci USA 2005;102:9469-74.
• [19] Kim SH, Jeong JH, Lee SH, Kim SW, Park TG. PEG conjugated VEGF siRNA for anti-angiogenic gene therapy. J Control Release 2006;116:123-9.
• [20] Li W, Szoka FC Jr. Lipid-based nanoparticles for nucleic acid delivery. Pharm Res 2007;24:438-49.
• [21] Dobrovolskaia MA, Patri AK, Simak J, Hall JB, Semberova J, De Paoli Lacerda SH, et al. Nanoparticle size and surface charge determine effects of PAMAM dendrimers on human platelets in vitro. Mol Pharm 2012;9:382-93.
• [22] Ferenc M, Pedziwiatr-Werbicka E, Nowak KE, Klajnert B, Majoral JP, Bryszewska M. Phosphorus dendrimers as carriers of siRNA-characterisation of dendriplexes. Molecules 2013;18:4451-66.
• [23] Fischer D, Li Y, Ahlemeyer B, Krieglstein J, Kissel T. In vitro cytotoxicity testing of polycations: influence of polymer structure on cell viability and hemolysis. Biomaterials 2003;24:1121-31.
• [24] Hunter AC. Molecular hurdles in polyfectin design and mechanistic background to polycation induced cytotoxicity.Adv Drug Deliv Rev 2006;58:1523-31.
• [25] Khan W, Muthupandian S, Domb AJ. Cationic polymers for the delivery of therapeutic nucleotides. In: Peer D, ed. Nanotechnology for the Delivery of Therapeutic Nucleic Acids. Singapore: Pan Stanford Publishing; 2013:27-56.