Badwaik V.D.,Multi Disciplinary Cancer Research Facility |
Aicart E.,Multi Disciplinary Cancer Research Facility |
Mondjinou Y.A.,Multi Disciplinary Cancer Research Facility |
Johnson M.A.,Indiana University |
And 2 more authors.
Biomaterials | Year: 2016
Nanoparticle-mediated siRNA delivery is a promising therapeutic approach, however, the processes required for transport of these materials across the numerous extracellular and intracellular barriers are poorly understood. Efficient delivery of siRNA-containing nanoparticles would ultimately benefit from an improved understanding of how parameters associated with these barriers relate to the physicochemical properties of the nanoparticle vectors. We report the synthesis of three Pluronic®-based, cholesterol end-capped cationic polyrotaxanes (PR+) threaded with 2-hydroxypropyl-β-cyclodextrin (HPβCD) for siRNA delivery. The biological data showed that PR+:siRNA complexes were well tolerated (∼90% cell viability) and produced efficient silencing (>80%) in HeLa-GFP and NIH 3T3-GFP cell lines. We further used a multi-parametric approach to identify relationships between the PR+ structure, PR+:siRNA complex physical properties, and biological activity. Small angle X-ray scattering and cryoelectron microscopy studies reveal periodicity and lamellar architectures for PR+:siRNA complexes, whereas the biological assays, ζ potential measurements, and imaging studies suggest that silencing efficiency is influenced by the effective charge ratio (ρeff), polypropylene oxide (PO) block length, and central PO block coverage (i.e., rigidity) of the PR+ core. We infer from our findings that more compact PR+:siRNA nanostructures arising from lower molecular weight, rigid rod-like PR+ polymer cores produce improved silencing efficiency relative to higher molecular weight, more flexible PR+ vectors of similar effective charge. This study demonstrates that PR+:siRNA complex formulations can be produced having higher performance than Lipofectamine® 2000, while maintaining good cell viability and siRNA sequence protection in cell culture. © 2015 Elsevier Ltd Source
Badwaik V.,Multi Disciplinary Cancer Research Facility |
Mondjinou Y.,Multi Disciplinary Cancer Research Facility |
Kulkarni A.,Multi Disciplinary Cancer Research Facility |
Liu L.,Multi Disciplinary Cancer Research Facility |
And 2 more authors.
Macromolecular Bioscience | Year: 2016
A family of cationic Pluronic-based polyrotaxanes (PR+), threaded with 2-hydroxypropyl-β-cyclodextrin (HPCD), was synthesized for pDNA delivery into multiple cell lines. All PR+ formed highly stable, positively charged pDNA complexes that were < 250 nm in diameter. The cellular uptake and pDNA transfection efficiencies of the PR+:pDNA complexes was enhanced relative to the commercial transfection standards L2K and bPEI, while displaying similar or lower toxicity profiles. Charge density and threading efficiency of the PR+ agent significantly influenced the colloidal stability and physical properties of the complexes, which impacted their intracellular transfection efficiencies. Taken together, our results suggest that HPCD: Pluronic PR+ can be used as potent vectors for pDNA-based therapeutics. Dried α, ω-bis-tris (2-aminoethyl) amine Pluronic triblock copolymer (0.08 mmol) and 2-hydroxypropyl-β-cyclodextrin (using the ratio of CD:PPG unit = 1:2 for each type of Pluronic) are dissolved (or suspended) in 60 mL hexane and the mixtures are vortexed before vigorously stirring for 2 h. The mixtures are then gently stirred for 72 h at 20 C and cholesteryl chloroformate (12 equiv) is used to cap the polyrotaxanes before purification and lyophilization. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. Source