Oligo-guanidyl targeted bioconjugates forming rod shaped polyplexes as a new nanoplatform for oligonucleotide delivery.
(2019) Journal of controlled release : official journal of the Controlled Release Society — Vol. 310, p. 58-73 (2019)
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- Novel bioconjugates (Agm-M-PEG-FA) for active oligonucleotide (ON) delivery have been developed by conjugating a cationic oligo-guanidyl star-like shaped "head" (Agm-M) to a polymeric "tail" (PEG) terminating with folic acid (FA) as targeting agent or methoxy group (Agm-M-PEG-FA and Agm-M-PEG-OCH, respectively). Gel electrophoresis showed that the bioconjugates completely associated with ONs at 3 nitrogen/phosphate (N/P) ratio. Studies performed with folate receptor (FR)-overexpressing HeLa cells, showed that optimal cell up-take was obtained with the 75:25 w/w Agm-M-PEG-OCH:Agm-M-PEG-FA mixture. Dynamic light scattering and transmission electron microscopy showed that the polyplexes had size <80 nm with narrow polydispersity and rod-shaped morphology. The polyplexes were stable for several hours in plasma while ON was released in the presence of heparin concentration 16-times higher than the physiological one. The polyplexes displayed negligible cytotoxicity, hemolysis and low pro-inflammatory TNF-α release. Studies performed with FR-overexpressing HeLa and MDA-MB-231 cells using siRac1 revealed that the folated polyplexes caused significantly higher gene silencing (86.1 ± 9.6%) and inhibition of cell migration (40%) than the non-folated polyplexes obtained with Agm-M-PEG-OCH only. Although cytofluorimetric analyses showed similar cell uptake for both folated and non-folated polyplexes, confocal, TEM and competition studies showed that the folated polyplexes were taken-up by lysosome escaping caveolin-mediated pathway with final polyplex localization within cytosol, while non-folated polyplexes were preferentially taken-up via clathrin-mediated pathway to localize in the lysosomes. Finally, preliminary in vivo studies carried out in mice revealed that the folated polyplexes dispose in the tumor mass.
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Malfanti, A., Scomparin, A., Pozzi, S., Gibori, H., Krivitsky, A., Blau, R., Satchi-Fainaro, R., Mastrotto, F., Caliceti, P., & Salmaso, S. (2019). Oligo-guanidyl targeted bioconjugates forming rod shaped polyplexes as a new nanoplatform for oligonucleotide delivery. Journal of controlled release : official journal of the Controlled Release Society, 310, 58-73. https://doi.org/10.1016/j.jconrel.2019.08.005 (Original work published 2019)