Xiao-Yan He

Dual-peptide-functionalized albumin-based nanoparticles with ph-dependent self-assembly behavior for drug delivery.

Drug delivery has become an important strategy for improving the chemotherapy efficiency. Here we developed a multifunctionalized nanosized albumin-based drug-delivery system with tumor-targeting, cell-penetrating, and endolysosomal pH-responsive properties. cRGD-BSA/KALA/DOX nanoparticles were fabricated by self-assembly through electrostatic interaction between cell-penetrating peptide KALA and cRGD-BSA, with cRGD as a tumor-targeting ligand. Under endosomal/lysosomal acidic conditions, the changes in the electric charges of cRGD-BSA and KALA led to the disassembly of the nanoparticles to accelerate intracellular drug release. cRGD-BSA/KALA/DOX nanoparticles showed an enhanced inhibitory effect in the growth of αvβ3-integrin-overexpressed tumor cells, indicating promising application in cancer treatments.

Biotinylated carboxymethyl chitosan/CaCO3 hybrid nanoparticles for targeted drug delivery to overcome tumor drug resistance

A tumor targeted nano-sized drug delivery system was prepared for co-delivery of an anti-tumor drug and a drug resistance inhibitor to overcome multidrug resistance (MDR) in cancer chemotherapy. The drug carrier, biotinylated carboxymethyl chitosan/CaCO3 (BCMC/CaCO3) hybrid nanoparticles with biotin as a target ligand was prepared by self-assembly in an aqueous solution. The hybrid nanoparticles exhibited a spherical shape with a hydrodynamic size less than 200 nm. An anti-cancer drug (doxorubicin hydrochloride, DOX) and a P-glycoprotein inhibitor (tariquidar, TQR) were co-loaded in the hybrid nanoparticles to obtain BCMC/CaCO3/DOX/TQR nanoparticles. For comparison, CMC/CaCO3/DOX/TQR nanoparticles without biotin ligands were also prepared. An MTT assay was employed to evaluate the cell inhibition effects of the drug loaded nanoparticles on both non-resistant cells (HeLa) and drug resistant cells (MCF-7/ADR). The in vitro study showed that BCMC/CaCO3/DOX/TQR nanoparticles exhibited obviously enhanced cell uptake and nuclear localization as compared with the free DOX and the single-drug loaded nanoparticles (BCMC/CaCO3/DOX and CMC/CaCO3/DOX) because of the efficient inhibition of P-glycoprotein (P-gp) mediated efflux. Moreover, compared with the nanoparticles without biotin moieties, the nanoparticles with biotin ligands showed stronger cell inhibition and increased cellular uptake in tumor cells. All these results indicated that the biotinylated hybrid nanoparticles have promising applications for co-delivery of multiple drugs to effectively overcome cancer drug resistance.

Fusion peptide functionalized hybrid nanoparticles for synergistic drug delivery to reverse cancer drug resistance

A facile self-assembly strategy was developed to decorate polymer/inorganic hybrid nano-sized drug delivery systems with functional peptides. To enhance drug delivery efficacy and overcome tumor drug resistance, a functional fusion peptide containing an RGD sequence for tumor targeting and an R8 sequence for cell penetration was introduced onto the surface of biotinylated carboxymethyl chitosan/CaCO3 (BCMC/CaCO3) hybrid nanoparticles through biotin-avidin interaction to obtain peptide functionalized nanoparticles (PNP). The peptide functionalization results in improved delivery efficiency and effective inhibition for drug resistant tumor cells. Co-delivery of an anti-cancerous drug (doxorubicin hydrochloride, DOX) and a cyclooxygenase-2 inhibitor (celecoxib, CXB) by PNP can further improve the therapeutic efficiency by effectively down-regulating P-gp expression to reduce P-gp mediated drug efflux and increase intracellular drug accumulation.

A Dual Macrophage Targeting Nanovector for Delivery of Oligodeoxynucleotides To Overcome Cancer-Associated Immunosuppression

To overcome cancer-associated immunosuppression, we prepared a dual-targeting vector to deliver CpG oligodeoxynucleotides (ODN) to macrophages. The dual-targeting system composed of mannosylated carboxymethyl chitosan (MCMC)/hyaluronan (HA) for macrophage targeting and protamine sulfate for ODN complexation was prepared by self-assembly. The effects of ODN delivery on immune cells was studied in J774A.1 cells. Due to the enhanced delivery efficiency, the dual-targeting delivery system exhibits a higher immune stimulatory activity compared with the monotargeting delivery system containing either MCMC or HA, resulting in a dramatically enhanced secretion of proinflammatory cytokines and a successful shift to activated macrophages (M1). Besides macrophages, the influence of the delivery system on tumor cells (MCF-7) was also investigated. In MCF-7 cells, the increased expressions of nuclear transcription factor-κB (NF-κB), PIK3R3, and phosphorylated protein kinase B (p-Akt) caused by activated NF-κB and phosphoinositide 3-kinase/Akt signalings were observed. Nevertheless, upregulated Fas as well as Fas ligand (FasL) may induce Fas/FasL-mediated apoptosis, which results in the increased expressions of caspases in tumor cells.

Aptamer-functionalized albumin-based nanoparticles for targeted drug delivery

Proteins have been extensively explored as versatile nanocarriers for drug delivery due to their complete biocompatibility, ease of surface modification, and lack of toxicity and immunogenicity. In this study, a facile strategy was used to construct aptamer-functionalized albumin-based nanoparticles for effective drug delivery and targeted cancer therapy. A hydrophobic drug, doxorubicin (DOX) was employed to trigger the self-assembly of bovine serum albumin (BSA) to from stable nanoparticles via hydrophobic interaction, and then a tumor targeting aptamer AS1411 was incorporated to the surface of DOX loaded BSA. Due to the specific recognition between AS1411 and its receptor over-expressed on tumor cells, the aptamer-modified nanoparticles show higher cellular uptake and stronger cell inhibitory efficacy against cancerous MCF-7 cells as compared with the nanoparticles without aptamer modification. In addition, DOX loaded aptamer-functionalized nanoparticles can induce more significant down-regulation of Bcl-2 and PCNA as well as up-regulation of pRB, PARP and Bax in MCF-7 cells compared with unmodified nanoparticles, indicating the aptamer modification can induce cell apoptosis more effectively. Besides, aptamer-modified nanoparticles exhibit a significantly improved capability in up-regulating p16, p21 and E-cadherin, and down-regulating EpCAM, vimentin, Snail, MMP-9, CD44 and CD133, implying the favorable effects of drug delivery on the prevention of tumor progression and metastasis.

A multi-functional macrophage and tumor targeting gene delivery system for the regulation of macrophage polarity and reversal of cancer immunoresistance

To achieve effective tumor eradication using anti-tumor immunotherapies, a fusion peptide functionalized gene delivery system for macrophage and tumor targeting delivery of the plasmid DNA encoding the IL-12 gene (pDNA IL-12) was prepared for macrophage re-polarization as well as reversal of cancer immunosuppression. A fusion peptide containing the tuftsin sequence that can interact with Fc receptors and neuropilin-1, and hyaluronic acid (HA) that can interact with CD44 were introduced into the delivery system by self-assembly to form peptide/hyaluronic acid/protamine/CaCO3/DNA nanoparticles (PHNP) with both macrophage targeting and tumor targeting capabilities. PHNP provides an efficient immunoregulation on J774A.1 cells to shift the anti-inflammatory M2 phenotype to the anti-tumor M1 phenotype with enhanced secretion of pro-inflammatory cytokines and increased expression of M1 markers. Owing to the improved delivery efficiency caused by the fusion peptide and HA, the transfection mediated by multi-functional PHNP can up-regulate IL-12 as well as down-regulate IL-10 and IL-4 more effectively as compared with the nanoparticles without HA and/or peptide decoration. More importantly, the gene delivery system can also deliver pDNA IL-12 to targeted cancerous HeLa cells to realize the secretion of IL-12. PHNP not only enables tumorous cells to produce pDNA IL-12, but also down-regulates CD47 and up-regulate CD80 and HLA-1 in the malignant cells, indicating that the gene delivery system can effectively reverse tumor induced immunosuppression.

Tumor targeted genome editing mediated by a multi-functional gene vector for regulating cell behaviors

ABSTRACT For effective regulation of cell behaviors and prevention of tumor development by genome editing, we constructed multi‐functional self‐assembled nanoparticles based on natural polymers to deliver CRISPR‐Cas9 plasmid to tumorous cells. The CRISPR based gene editing plasmid to knockout CDK11 gene was complexed with protamine sulfate, and then the complex was decorated by a multi‐functional outer layer composed of an endosomolytic peptide (KALA) and aptamer AS1411 incorporated carboxymethyl chitosan. The resultant multi‐functional nanoparticles, which exhibit significantly enhanced delivery efficiency, can specifically deliver the plasmid into tumor cell nuclei owing to the favorable effects of KALA in cellular uptake and endosomal escape, together with the cancer cell and cell nucleus targeting capability of AS1411 ligands. The genome editing mediated by the nanoparticles leads to a dramatic decrease (>75%) in CDK11 expression, which results in further modulation of cancer cells with significant down‐regulation of the proteins (MMP‐9 and VEGF) involved in tumor development and metastasis as well as up‐regulation of the tumor suppressor protein p53. More importantly, the detection of immune‐related proteins after genome editing shows that the significantly enhanced Fas, CD80, MICA, MICB, and HLA‐1 expression and decreased CD47 and MUC1 expression, indicating the genome editing is favorable for reversal of tumor‐induced immunosuppression and prevention of tumor development.

A Dual-Targeting Delivery System for Effective Genome Editing and In Situ Detecting Related Protein Expression in Edited Cells

One of critical steps in genome editing by CRISPR-Cas9 is to deliver the CRISPR-Cas9 system into targeted cells. In this study, we developed a dual-targeting delivery system based on polymer/inorganic hybrid nanoparticles to realize highly efficient genome editing in targeted tumor cells as well as in situ detection on the related protein expression in edited cells. The CRISPR-Cas9 plasmid for CDK11 knockout was encapsulated in the core of the delivery system composed of protamine sulfate, calcium carbonate, and calcium phosphate by coprecipitation, and functional derivatives of carboxymethyl chitosan (biotinylated carboxymethyl chitosan with biotin ligands and aptamer-incorporated carboxymethyl chitosan with AS1411 ligands) were decorated on the nanovector surface by electrostatic interactions to form the dual-targeting delivery system. On the basis of the tumor cell targeting capability of biotin and AS1411 ligands as well as the nuclear targeting of AS1411, the dual-targeting system can deliver the CRISPR-Cas9 plasmid into the nuclei of tumor cells to realize highly efficient genome editing, resulting in a dramatic decrease (>90%) in CDK11 protein together with the significant downregulation of other proteins involved in tumor development, including an ∼90% decrease in MMP-9, >40% decrease in VEGF, and ∼70% decrease in survivin. Using the same vector, molecular beacons can be easily delivered to edited cell nuclei to in situ detect the mRNA level of related proteins (p53 and survivin as typical examples) and mRNA distribution in subcellular organelles. Our strategy can realize effective genome editing and in situ detection on related protein expression simultaneously.