Hybrid polypeptide micelles loading indocyanine green for tumor imaging and photothermal effect study.

Indocyanine green (ICG) is a near-infrared (NIR) fluorescence dye for extensive applications; however, it is limited for further biological application due to its poor aqueous stability in vitro, concentration-dependent aggregation, rapid elimination from the body, and lack of target specificity. To overcome its limitations, ICG was encapsulated in the core of a polymeric micelle, which self-assembled from amphiphilic PEG-polypeptide hybrid triblock copolymers of poly(ethylene glycol)-b-poly(l-lysine)-b-poly(l-leucine) (PEG-PLL-PLLeu), with PLLeu as the hydrophobic core and PEG as the hydrophilic shell. The ICG was associated with the hydrophobic core via hydrophobic interaction and also the hydrophilic heads through electrostatic attractive interaction. Compared with free ICG, PEG-PLL-PLLeu-ICG micelles significantly improved quantum yield and fluorescent stability. The cellular uptake experiments showed that PEG-PLL-PLLeu-ICG micelles have a high cellular uptake rate. And the in vivo experiments revealed the excellent passive tumor targeting ability and long circulation time of PEG-PLL-PLLeu-ICG. The above results indicated the broad prospects of PEG-PLL-PLLeu-ICG application in the fields of tumor diagnosis and imaging. In addition, temperature measurements under NIR laser irradiation and in vitro photothermal ablation studies proved the potential application of PEG-PLL-PLLeu-ICG in tumor photothermal therapy.

Self-assembled cationic micelles based on PEG-PLL-PLLeu hybrid polypeptides as highly effective gene vectors.

Developing safe and effective nonviral gene vector is highly crucial for successful gene therapy. In the present study, we designed a series of biodegradable micelles based on hybrid polypeptide copolymers of poly(ethylene glycol)-b-poly(l-lysine)-b-poly(l-leucine) (PEG-PLL-PLLeu) for efficient gene delivery. A group of amphiphilic PEG-PLL-PLLeu hybrid polypeptide copolymers were synthesized by ring-opening polymerization of N-carboxyanhydride, and the chemical structure of each copolymer was characterized by (1)H NMR and FT-IR spectroscopy measurement. The PEG-PLL-PLLeu micelles were positively charged with tunable sizes ranging from 40 to 90 nm depending on the length of PLL and PLLeu segment. Compared with PEG-PLL copolymers, PEG-PLL-PLLeu micelles demonstrated significantly higher transfection efficiency and less cytotoxicity. Furthermore, the transfection efficiency and biocompatibility of the micelles can be simultaneously improved by tuning the length of PLL and PLLeu segments. The transfection efficiency of PEG-PLL-PLLeu micelles in vivo was two to three times higher than that of PEI(25k), which was attributable to their capability of promoting DNA condensation and cell internalization as well as successful lysosome escape. Hence well-defined PEG-PLL-PLLeu micelles would serve as highly effective nonviral vectors for in vivo gene delivery.

Polypeptide cationic micelles mediated co-delivery of docetaxel and siRNA for synergistic tumor therapy.

Combination of two or more therapeutic strategies with different mechanisms can cooperatively impede tumor growth. Co-delivery of chemotherapeutic drug and small interfering RNA (siRNA) within a single nanoparticle (NP) provides a rational strategy for combined cancer therapy. Here, we prepared polypeptide micelle nanoparticles (NPs) of a triblock copolymer poly(ethylene glycol)-b-poly(l-lysine)-b-poly(l-leucine) (PEG-PLL-PLLeu) to systemically codeliver docetaxel (DTX) and siRNA-Bcl-2 for an effective drug/gene vector. The hydrophobic PLLeu core entrapped with anticancer drugs, while the PLL polypeptide cationic backbone allowed for electrostatic interaction with the negatively charged siRNA. The resulting micelle NP exhibited very stable, good biocompatible and excellent passive targeted properties. The micelle complexes with siRNA-Bcl-2 effectively knocked down the expression of Bcl-2 mRNA and protein. Moreover, the co-delivery system of DTX and siRNA-Bcl-2 (DTX-siRNA-NPs) obviously down-regulation of the anti-apoptotic Bcl-2 gene and enhanced antitumor activity with a smaller dose of DTX, resulting the significantly inhibited tumor growth of MCF-7 xenograft murine model as compared to the individual siRNA and only DTX treatments. Our results demonstrated well-defined PEG-PLL-PLLeu polypeptide cationic micelles with the excellent synergistic effect of DTX and siRNA-Bcl-2 in combined cancer therapy.

Effect of peptides and their introduction methods on target gene transfer of gene vector based on disulfide-containing polyethyleneimine.

To evaluate the effect of different peptides as well as their introduction methods on target gene transfer of gene vectors based on disulfide-containing polyethyleneimine (SS-PEI), a series of peptides including N(3)-GRGDSF, GRGDSF, and EEEEEEEEGRGDSF (E(8)GRGDSF) were prepared. N(3)-GRGDSF was conjugated to SS-PEI by click chemistry and SS-PEI-GRGDSF was obtained. GRGDSF was non-covalently introduced into SS-PEI/DNA mainly through hydrogen bonding to obtain SS-PEI/DNA/GRGDSF complexes, whereas E(8)GRGDSF was further non-covalently introduced to SS-PEI/DNA through electrostatic force to obtain SS-PEI/DNA/E(8)GRGDSF complexes. Transfection efficiency of all complexes with peptides was lower than that of SS-PEI/DNA in COS-7 cells due to the fact that nonspecific endocytosis was prohibited after peptide introduction. Whereas in HeLa cells, transfection activity of SS-PEI-GRGDSF/DNA and SS-PEI/DNA/E(8)GRGDSF at certain w/w ratios was higher than that of SS-PEI/DNA. But the transfection efficiency of SS-PEI/DNA/E(8)GRGDSF at peptide/DNA w/w ratios higher than 30 dropped due to targeted binding interactions between surplus E(8)GRGDSF and the integrins in HeLa cells, which would prohibit specific endocytosis of E(8)GRGDSF in complexes. Transfection activity of SS-PEI/DNA/GRGDSF was lower than or comparable to that of SS-PEI/DNA because of loose complexes constructed by hydrogen bonding between GRGDSF and SS-PEI/DNA.

Poly(ß-amino amine) cross-linked PEIs as highly efficient gene vectors.

To increase the release of DNA into the cytoplasm and further improve transgene expression of nucleic acid novel polymeric gene carriers were prepared which would be biodegradable under the reducing conditions in the cytoplasm. Disulfide-containing poly(ß-amino amine)s were first synthesized and then used to cross-link low molecular weight polyethyleneimine (1800 Da) through Michael addition to obtain SS-PBAA-PEIs as the final gene carriers. The physicochemical characteristics of SS-PBAA-PEI/DNA complexes were characterized. In vitro transfection mediated by the SS-PBAA-PEIs under serum conditions was carried out. Cell uptake of the gene delivery systems was observed by confocal laser scanning microscopy. The results of the physicochemical characterisation demonstrated that the SS-PBAA-PEIs could efficiently condense DNA. In vitro transfection under serum conditions showed that SS-PBAA-PEIs had comparable or even higher transfection efficiencies than 25 kDa PEI. And SS-PBAA-PEIs showed much lower cytotoxicity compared with 25 kDa PEI. In summary, the SS-PBAA-PEIs possess great potential as non-viral gene vectors and exhibit high transfection efficiency under serum conditions.