Brain-targeted CRISPR/Cas9 nanomedicine for effective glioblastoma therapy.

CRISPR/Cas9 gene-editing technology shows great potential for treating a variety of diseases, such as glioblastoma multiforme (GBM). However, CRISPR components suffer from inherent delivery challenges, such as poor in vivo stability of Cas9 protein and gRNA, low blood-brain barrier (BBB) permeability and non-specific tissue or cell targeting. These defects have limited the application of Cas9/gRNA ribonucleoprotein (RNP) complexes for GBM therapy. Here, we developed a brain-targeted CRISPR/Cas9 based nanomedicine by fabricating an angiopep-2 decorated, guanidinium and fluorine functionalized polymeric nanoparticle with loading Cas9/gRNA RNP for the treatment of GBM. The guanidinium and fluorine domains of our polymeric nanoparticles were both capable of interacting with Cas9/gRNA RNP to stabilize it in blood circulation, without impairing its activity. Moreover, by leveraging angiopep-2 peptide functionality, the RNP nanoparticles efficiently crossed the BBB and accumulated in brain tumors. In U87MG cells, we achieved approximately 32% gene knockout and 67% protein reduction in the targeted proto-oncogene polo-like kinase 1 (PLK1). This was sufficient to suppress tumor growth and significantly improved the median survival time of mice bearing orthotopic glioblastoma to 40 days, while inducing negligible side or off-target effects. These results suggest that the developed brain-targeted CRISPR/Cas9 based nanomedicine shows promise for effective human glioblastoma gene therapy.

Polymeric nanoparticle mediated inhibition of miR-21 with enhanced miR-124 expression for combinatorial glioblastoma therapy.

Glioblastoma (GBM) is the most common and fatal form of malignant brain tumor. Despite intensive effort, there is still no effective GBM treatment. Therefore, novel and more effective GBM therapeutic approaches are highly desired. In this study, we combined polymeric nanotechnology with microRNA (miRNA) regulation technology to develop a targeted polymeric nanoparticle to co-deliver anti-miR-21 and miR-124 into the brain to effectively treat GBM. The polymeric nanoparticle decorated with Angiopep-2 peptide not only can encapsulate miRNA via triple-interaction (electrostatic, hydrogen bond and hydrophobic bonding) to protect miRNA against enzyme degradation in the blood, but also is capable of crossing blood brain barrier (BBB) and allowing targeted delivery of miRNAs to GBM tissue due to the dual-targeting function of Angiopep-2. Moreover, the co-delivered anti-miR-21 and miR-124 simultaneously regulated the mutant RAS/PI3K/PTEN/AKT signaling pathway in tumor cells, consequently achieving combinatorial GBM therapy. This combinatorial effect was confirmed by our results showing that these miRNA nanomedicines can effectively reduce tumor cell proliferation, migration and invasion as well as reducing tumor angiogenesis. Consequently, effective suppression of tumor growth and significantly improved medium survival time are observed when these miRNA nanomedicines were assessed in an orthotopic GBM xenograft model. This work indicated that our new polymeric nanoparticles successfully mediate inhibition of miR-21 and miR-124 supplementation to significantly reduce tumorigenesis, and may have strong potential in GBM therapy.

Evaluation of nanomechanical properties of hyperbranched polyglycerols as prospective cell membrane engineering block.

Owing to the excellent biocompatibility, hyperbranched polyglycerols (hbPGs) are one of the most promising polymers and widely employed in drug delivery. Presented as an excellent bioinert coating material, hbPGs can significantly improve the biosafety of biomedical nanomaterials. However, it is still unclear what specific properties of hbPGs are the key effectors to bioinertness. Here, atomic force microscopy was employed to test the Young's modulus and adhesion of hbPGs, spin-coated onto mica substrate. High Young's modulus indicated that the hbPGs cannot be further compressed and low adhesion implied that it is not easy to form hbPGs aggregators. This could owe to the intramolecular hydrogen bond. Morphology characterization of hbPGs self-assembled monolayer onto Si(100) substrate, confirmed the lower adhesion among different hbPGs and indicated their biofouling properties. Further confocal laser microscopy of cell membrane modified with alkyl chain (C18)-modified hbPGs and hbPGs-NH2, confirmed that the antifouling properties of hbPGs are determined by terminal glycerol units. Our findings demonstrated that only hbPGs with entire terminal surface can be used as perspective cell membrane modification skeleton.