Combined delivery of temozolomide and the thymidine kinase gene for treatment of glioblastoma.

Glioblastoma is the most malignant form of brain tumor. In this study, combination therapy with temozolomide (TMZ) and the herpes simplex virus thymidine kinase (HSVtk) gene was evaluated in glioblastoma models. The R7L10 peptide was used as a carrier of TMZ and the HSVtk gene. TMZ was loaded into R7L10 micelles using the oil-in-water emulsion/solvent evaporation method. The TMZ-loaded R7L10 (R7L10-TMZ) micelles formed a complex with the HSVtk gene, pHSVtk. The formation of the R7L10-TMZ/pHSVtk complex was confirmed by gel retardation and heparin competition assays. An in vitro transfection assay demonstrated that the transfection efficiency of R7L10-TMZ was similar to that of R7L10 in C6 glioblastoma cells. R7L10-TMZ had greater anti-tumor effects than TMZ alone in C6 cells in vitro, suggesting that R7L10 is an efficient carrier of TMZ. The in vivo efficacy of the R7L10-TMZ/pHSVtk complex was evaluated in the intracranial glioblastoma model. HSVtk expression in tumors was confirmed by immunohistochemistry. Furthermore, a greater anti-tumor effect was observed in the R7L10-TMZ/pHSVtk group compared with the TMZ or R7L10/pHSVtk single injection group. In conclusion, combined delivery of TMZ and the HSVtk gene using R7L10 peptides may be useful for the treatment of glioblastoma.

Peptide micelle-mediated curcumin delivery for protection of islet ß-cells under hypoxia.

Islet transplantation is one of many therapeutic approaches for the treatment of diabetes. During transplant procedures, the isolated islets are subjected to hypoxic conditions, and undergo the apoptotic process. Curcumin has a cytoprotective effect, and may therefore be useful for the protection of islets under hypoxia. However, curcumin is hydrophobic, and an efficient curcumin carrier is required for effective treatment. In this study, R3V6 peptide micelles, composed of a 3-arginine stretch and 6-valine stretch, were evaluated as a curcumin carrier to INS-1 insulinoma cells. Curcumin was loaded into R3V6 micelles at a weight ratio of 10:3 (R3V6:curcumin). The size and surface charge of the curcumin-loaded R3V6 micelles (R3V6-curcumin) were approximately 250 nm and 17.49 mV, respectively. R3V6-curcumin delivered curcumin to the INS-1 cells more efficiently than either curcumin alone or a simple mixture of R3V6 and curcumin. MTT assay indicated that under hypoxia, R3V6-curcumin protected INS-1 cells more efficiently than curcumin alone. TUNEL and reactive oxygen species (ROS) assays suggested that R3V6-curcumin reduced INS-1 cell apoptosis under hypoxia. These results demonstrate that R3V6 peptide micelles are an effective carrier of curcumin, and that R3V6-curcumin may improve the viability of pancreatic ß-cells in islet transplantation.

Peptide micelle-mediated delivery of tissue-specific suicide gene and combined therapy with avastin in a glioblastoma model.

Bevacizumab (Avastin) is an angiogenesis inhibitor used as a treatment for various cancers. In this study, the combination therapy of Avastin and glioblastoma-specific thymidine kinase gene [pEpo-NI2-SV-herpes simplex virus thymidine kinase(HSVtk)] was evaluated in a glioblastoma animal model. The R7L10 peptide was used as a gene carrier of pEpo-NI2-SV-HSVtk. Gel retardation assays confirmed that R7L10 formed stable complexes with pEpo-NI2-SV-HSVtk. R7L10 protected DNA from nuclease digestion. R7L10 had lower transfection efficiency than polyethylenimine (PEI; 25 kDa). However, the in vitro and in vivo toxicity assays showed that R7L10 had lower cytotoxicity than PEI, suggesting that R7L10 is safer than PEI. For the combination therapy, Avastin was injected intravenously and the pEpo-NI2-SV-HSVtk/R7L10 complexes were injected intratumorally in the glioblastoma animal model. Tumor growth was most effectively inhibited by the combination therapy of Avastin and the gene. The immunostaining results confirmed that the HSVtk genes were expressed in the groups with the pEpo-NI2-SV-HSVtk/R7L10 complex. The terminal deoxynucleotidyl transferase dUTP nick end labeling assay showed a higher level of apoptotic cells in the combination group than the pEpo-NI2-SV-HSVtk/R7L10 complex or Avastin group. In conclusion, the combination of Avastin and the glioblastoma-specific HSVtk gene has a higher antitumor effect than single therapy of Avastin or HSVtk after intratumoral administration in glioblastoma animal model.

Thymidine kinase gene delivery using curcumin loaded peptide micelles as a combination therapy for glioblastoma.

PURPOSE: The effect of the combination therapy of curcumin and the herpes simplex virus thymidine kinase (HSVtk) gene using R7L10 as a carrier was evaluated in a glioblastoma animal model. METHODS: Curcumin was loaded into the cores of R7L10 peptide micelles using an oil-in-water emulsion/solvent evaporation method to generate curcumin loaded R7L10 micelles (R7L10-Cur), which were used as a carrier to deliver the HSVtk gene. The plasmid DNA (pDNA)/R7L10-Cur complex was confirmed by gel retardation, heparin competition, and dynamic light scattering analyses. Transfection efficiency and cytotoxicity were measured using luciferase, MTT, and TUNEL assays. Intracellular delivery of curcumin was determined by fluorescence and absorbance. In the glioblastoma animal model, the effects of the intratumoral delivery of curcumin and the HSVtk gene were evaluated according to tumor size, immunohistochemistry, and TUNEL assays. RESULTS: R7L10-Cur delivered pDNA into the cells more efficiently than PLL and R7L10. In addition, R7L10-Cur delivered curcumin into the cells more efficiently than curcumin alone. The pHSVtk/R7L10-Cur complex induced cell death efficiently both in vitro and in vivo. Likewise, the combination of curcumin and the HSVtk gene using the pHSVtk/R7L10-Cur complex reduced tumor size more efficiently than the pHSVtk/PEI and pHSVtk/R7L10 complexes in a glioblastoma animal model. CONCLUSION: R7L10 is an efficient carrier for delivery of curcumin and the HSVtk gene, which may be a useful combination therapy for glioblastoma.