Polyplex-microbubble hybrids for ultrasound-guided plasmid DNA delivery to solid tumors

Abstract

Microbubble ultrasound contrast agents are being developed as image-guided gene carriers for targeted delivery in vivo. In this study, novel polyplex-microbubbles were synthesized, characterized and evaluated for systemic circulation and tumor transfection. Branched polyethylenimine (PEI; 25 kDa) was modified with polyethylene glycol (PEG; 5 kDa), thiolated and covalently attached to maleimide groups on lipid-coated microbubbles. The PEI-microbubbles demonstrated increasingly positive surface charge and DNA loading capacity with increasing maleimide content. The in vivo ultrasound contrast persistence of PEI-microbubbles was measured in the healthy mouse kidney, and a two-compartment pharmacokinetic model accounting for free and adherent microbubbles was developed to describe the anomalous time-intensity curves. The model suggested that PEI loading dramatically reduced free circulation and increased nonspecific adhesion to the vasculature. However, DNA loading to form polyplex-microbubbles increased circulation in the bloodstream and decreased nonspecific adhesion. PEI-microbubbles coupled to a luciferase bioluminescence reporter plasmid DNA were shown to transfect tumors implanted in the mouse kidney. Site-specific delivery was achieved using ultrasound applied over the tumor area following bolus injection of the DNA/PEI-microbubbles. In vivo imaging showed over 10-fold higher bioluminescence from the tumor region compared to untreated tissue. Ex vivo analysis of excised tumors showed greater than 40-fold higher expression in tumor tissue than non-sonicated control (heart) tissue. These results suggest that the polyplex-microbubble platform offers improved control of DNA loading and packaging suitable for ultrasound-guided tissue transfection.

New chitosan nanobubbles for ultrasound-mediated gene delivery: preparation and in vitro characterization

“…Sonoporation is being studied as an effective means of promoting extravasation of large macromolecules, such as plasmid DNA, to improve their delivery to tissues and targeted release. 6 In particular, much research has been directed towards combined use of ultrasound and microbubbles with the aim of improving transfection efficiency. 7,8 Several formulations of microbubbles with shells composed of lipids, proteins, polymers, and surfactants have been designed as DNA carriers.…”

Polyplex-microbubbles for ultrasound-mediated gene therapy

“…Results were originally presented in (Sirsi et al, 2012). Figure 1 shows the total DNA loading capacity per unit surface area of the PEI-microbubbles.…”

“…Generation and characterization of polyplex-microbubbles was described in (Sirsi et al, 2012). Briefly, the lipids used to encapsulate the microbubbles, 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-distearoylsn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)-2000] (DSPE-PEG2K), and 1,2-distearoyl-snglycero-3-phosphoethanolamine-N-[maleimide(polyethylene glycol)-2000] (DSPE-PEG2K-Mal), were obtained from Avanti Polar Lipids (Alabasater, AL).…”

Quantification of transient increase of the blood–brain barrier permeability to macromolecules by optimized focused ultrasound combined with microbubbles

“…Figure 1 shows the size distribution curve of the MBs generated by our technique, which have the optimal size of ~2 μm as an ultrasound contrast agent in in vivo ultrasound imaging and treatment applications. 7,10,11,26,[32][33][34] The MB concentration prepared for injection was ~1.7×10 6 bubbles/mL, which is in an optimal concentration range for reversible disruption of the BBB based on the study by Yang et al 7 animal preparation…”