Physical-chemical measurement method development for self-assembled, core-shell nanoparticles.

Improvements in dimensional metrology and innovations in physical-chemical characterization of functionalized nanoparticles are critically important for the realization of enhanced performance and benefits of nanomaterials. Toward this goal, we propose a multi-technique measurement approach, in which correlated atomic force microscopy, dynamic light scattering, high performance liquid chromatography and mass spectroscopy measurements are used to assess molecular and structural properties of self-assembled polyplex nanoparticles with a core-shell structure. In this approach, measurement methods are first validated with a model system consisting of gold nanoparticles functionalized with synthetic polycationic branched polyethylenimine macromolecules. Shell thickness is measured by atomic force microscopy and dynamic light scattering, and the polyelectrolyte uptake determined by chromatographic separation and mass spectrometric analysis. Statistical correlation between size, structure and stability provide a basis for extending the methods to more complex self-assembly of nucleic acids and macromolecules via a condensation reaction. From these size and analytical chemical measurements, we obtain a comprehensive spatial description of these assemblies, obtain a detailed interpretation of the core-shell evolution, and identify regions of the parameter space where stable, discrete particle formation occurs.

A nanoparticle carrying the p53 gene targets tumors including cancer stem cells, sensitizes glioblastoma to chemotherapy and improves survival.

Temozolomide (TMZ)-resistance in glioblastoma multiforme (GBM) has been linked to upregulation of O(6)-methylguanine-DNA methyltransferase (MGMT). Wild-type (wt) p53 was previously shown to down-modulate MGMT. However, p53 therapy for GBM is limited by lack of efficient delivery across the blood brain barrier (BBB). We have developed a systemic nanodelivery platform (scL) for tumor-specific targeting (primary and metastatic), which is currently in multiple clinical trials. This self-assembling nanocomplex is formed by simple mixing of the components in a defined order and a specific ratio. Here, we demonstrate that scL crosses the BBB and efficiently targets GBM, as well as cancer stem cells (CSCs), which have been implicated in recurrence and treatment resistance in many human cancers. Moreover, systemic delivery of scL-p53 down-modulates MGMT and induces apoptosis in intracranial GBM xenografts. The combination of scL-p53 and TMZ increased the antitumor efficacy of TMZ with enhanced survival benefit in a mouse model of highly TMZ-resistant GBM. scL-p53 also sensitized both CSCs and bulk tumor cells to TMZ, increasing apoptosis. These results suggest that combining scL-p53 with standard TMZ treatment could be a more effective therapy for GBM.

Nanoimmunoliposome delivery of superparamagnetic iron oxide markedly enhances targeting and uptake in human cancer cells in vitro and in vivo.

To circumvent the problem of reduction of the supermagnetic properties of superparamagnetic iron oxide (SPIO) nanoparticles after chemical modification to conjugate targeting molecules, we have adapted a tumor-targeting nanoimmunoliposome platform technology (scL) to encapsulate and deliver SPIO (scL-SPIO) in vitro and in vivo without chemical modification. Scanning probe microscopy, confocal microscopy, and Prussian blue staining were used to analyze the scL-SPIO and assess intracellular uptake and distribution of SPIO in vitro. In vivo targeting and tumor-specific uptake of scL-SPIO was examined using fluorescent-labeled SPIO. We demonstrated that SPIO encapsulation in the scL complex results in an approximately 11-fold increase in SPIO uptake in human cancer cells in vitro, with distribution to cytoplasm and nucleus. Moreover, the scL nanocomplex specifically and efficiently delivered SPIO into tumor cells after systemic administration, demonstrating the potential of this approach to enhance local tumor concentration and the utility of SPIO for clinical applications.

Materializing the potential of small interfering RNA via a tumor-targeting nanodelivery system.

The field of small interfering RNA (siRNA) as potent sequence-selective inhibitors of transcription is rapidly developing. However, until now, low transfection efficiency, poor tissue penetration, and nonspecific immune stimulation by in vivo administered siRNAs have delayed their therapeutic application. Their potential as anticancer therapeutics hinges on the availability of a vehicle that can be systemically administered, safely and repeatedly, and will deliver the siRNA specifically and efficiently to the tumor, both primary tumors and metastases. We have developed a nanosized immunoliposome-based delivery complex (scL) that, when systemically administered, will preferentially target and deliver molecules useful in gene medicine, including plasmid DNA and antisense oligonucleotides, to tumor cells wherever they occur in the body. This tumor-targeting nanoparticle delivery vehicle can also deliver siRNA to both primary and metastatic disease. We have also enhanced the efficiency of this complex by the inclusion of a pH-sensitive histidine-lysine peptide in the complex (scL-HoKC) and by delivery of a modified hybrid (DNA-RNA) anti-HER-2 siRNA molecule. Scanning probe microscopy confirms that this modified complex maintains its nanoscale size. More importantly, we show that this nanoimmunoliposome anti-HER-2 siRNA complex can sensitize human tumor cells to chemotherapeutics, silence the target gene and affect its downstream pathway components in vivo, and significantly inhibit tumor growth in a pancreatic cancer model. Thus, this complex has the potential to help translate the potent effects of siRNA into a clinically viable anticancer therapeutic.