Peptide-mediated Nano-scaled Drug Delivery System For Target Treatment Of Glioma

Glioma is the most common and fatal primary brain tumor, andone of the ten main cauces of death by cancer.All of the common treatment such as surgical debulking, radiation and chemotherapy could not achieve ideal therapeutic effect. Most of the drug possessing promising future can not accumulate in the glioma site due to the limitation of blood-brain barrier (BBB) or the lack of targetability. Hence, the treatment of glioma has always been recognized as worldwide problem.Targeted nanotechnology has shown the special advances and great clinical prospect in oncotherpy. In this study, two strategies about drug delivery and oncotherpy were designed according to the varying physiological characteristic of early or late phase of glioma.1. In the early phase of glioma, the integrity of BBB is still retaining, the drugs have to cross BBB to accumulate in the glioma. The peptides with high affinity to both BBB and glioma may endow the drug delivery system with BBB-glioma dual targeting function. The outstanding characteristic of glioma is invasive growth that leads to there is no obvious boundary between the normal brain tissue and glioma. Hence, it’s necessary to employ the drug which could recognize andkill off the glioma cells specifically without damaging normal brain cells.2. In the late phase of glioma, the integrity of BBB is disrupted, and the glioma possesses the EPR effect and tumor microenvironment as most of tumors. The drug delivery system could passively accumulate in the glioma tissue. And after that, the drug delivery system modified with CPP that possessing efficient internalization could drive the drug internalize into the glioma cells. As glioma is a multi-factor, multi-link, multi-stage complex disease, so it’s necessary and meaningful to apply the multi-target treatment.According to the above two strategiesand the interdisciplinary application ofpharmaceutics, macromolecular chemistry, pathology and biology, two kind of peptide-mediated nano-scaled drug delivery system for target treatment of glioma were constructed. The first system was dual targeting effect of Angiopep-mediated, DNA-loaded nanoparticles. Polyamidoamine (PAMAM) was applied as the main macromolecular gene vector.Angiopep, a BBB-gliomadual targeting peptide, was modified to the gene vector with polyethyleneglycol (PEG) as a spacer to form the nanocarrier, PAMAM-PEG-Angiopep. Then the nanocarrier was complexed with the plasmid pORF-TRAIL that expressing TRAIL protein to form the dual targeting nanoparticles, PAMAM-PEG-Angiopep/pORF-TRAIL.The second system was tumor targeting effect of dtACPP-mediated, co-delivery nanoparticles. Dendrigraft poly-L-lysine (DGL) was applied as the main macromolecular gene vector.dtACPP, a tumor microenvironment-responsive CPP, was modified to the gene vector, with polyethyleneglycol (PEG) as a spacer to form the nanocarrier,dtACPP-PEG-DGL (dtACPPD). Inserting DOX into shVEGF to form the shVEGF-DOX complex, and then complexed with the dtACPPD to form the co-delivery system, dtACPPD/shVEGF-DOX.In the first part, PAMAM-PEG-Angiopep/pORF-TRAIL, the dual targeting nanoparticles were constructed andboth of the corresponding in vitro and in vivo characteristics were evaluated. MTT results confirmed that the nanocarrier was a secure platform for drug delivery. And the confocal results showed that PAMAM-PEG-Angiopep/pORF-TRAIL could effectively internalize into the glioma cells via the endosome/lysosome pathway and further get into the nuclei. According to the results of pharmacodynamics, PAMAM-PEG-Angiopep/pORF-TRAIL treated group could induce apoptosis in the interior of the glioma, and the corresponding median survival time was up to61days, obviously longer than the Temozolomide (49days) or PAMAM-PEG/pORF-TRAIL (30days) treadted ones. The reason might be attributed to Angiopep mediated the nanoparticles to cross the BBB to accumulate in the glioma, and further internalized into the glioma cells to express TRAIL protein. All of results confirmed that the BBB-glioma dual targeting effect of Angiopep and the selective apoptosis glioma cells inducing by TRAIL, and further showed that PAMAM-PEG/pORF-TRAIL was an efficient dual target nano-scaled drug delivery system for glioma.In the first section of the second part,tumor microenvironment responsive CPP, dtACPP, and the corresponding nanocarrier, dtACPP, were constructed, and both of the corresponding in vitro and in vivo characteristics were evaluated. SDS-PAGE results showed that both dtACPP and dtACPPD could deshield to expose CPP well in the mimetic tumor microenvironment. The cellular uptake from different cell lines showed that, the cellular uptake efficiency of dtACPP-mediated nanocarricer was similar to the CPP-mediated group. The in vivo distribution imaging confirmed that that dtACPP-mediated nanocarrier possessed good tumor targetability in different tumor-bearing mice.In the second section of the second part,dtACPP-mediated tumor targeting nanoparticles,dtACPPD/DNA, were constructed, and both of the corresponding in vitro and in vivo characteristics were evaluated.The cellular uptake from different cell lines showed that, the cellular uptake efficiency of dtACPP-mediated naonparticles was similar to the CPP-mediated group. The in vivo distribution imaging confirmed that that dtACPP-mediated nanocarrier possessed good tumor targetability in different tumor-bearing mice. And the section results further confirmed that the modification of dtACPP could promote the accumulation of the nanoparticles in the tumor site, and further forward the nanoparticles’internalization into the cells and neclei for efficient gene expression.In the third part, tumor targeting effect of dtACPP-mediated, co-delivery nanoparticles,dtACPPD/shVEGF-DOX,were constructed, and both of the corresponding in vitro and in vivo characteristics were evaluated. The results showed that basing on the delivery of dtACPPD, shVEGF could effectively inhibit the generation of blood vessels by down-regulated the expression of endogenous VEGF of glioma, and induce the apoptosis of glioma indirectly. Additionally, after combined with DOX, the apoptosis phenomenon was more obvious. Themedian survival time of the combination therapy,dtACPPD/shVEGF-DOX treated group, was up to58.5days, obviously longer than dtACPPD/shVEGF (38days) or dtACPPD/pControl-DOX (35days) treadted ones, confirming that the co-delivery nanoparticles could effectively prolong the survival timr of the glioma-bearing mice. Meanwhile, the combination therapy group exhibited a gentlest loss in body weight. All of the results showed that dtACPPD/shVEGF-DOX could effectively cure the glioma by implementing the combination ofantiangiogenesis and apoptosis simultaneously.