Introduction: Targeted delivery of anticancer drugs to brain tumors, especially glioblastoma multiform, which is the most frequent and aggressive types, is one of the important objectives in nanomedicine. Vascular endothelial growth factor (VEGF) and its receptor type II (VEGFR2) are promising targets for brain tumors, because they are overexpressed by not only core cells but also by migrated glioma cells, which are responsible for resistance and rapid progression of brain tumors. The purpose of this research was to develop VEGF- and VEGFR2-targeted liposomes for targeted delivery of cisplatin to intracranial glioblastomas.
Materials and Methods: We designed cisplatin-loaded stealth liposomes by the film hydration method. Antibodies against the native form of VEGF or VEGFR2 were conjugated to liposomes via PEG-linkers with high retained immunochemical activity (about 80% of initial affinity), as detected by ELISA. Particle size, loading capacity, drug release and stability of drug-loaded nanocontainers were evaluated. Glioma C6 and U-87 cells were used to study the cellular uptake of cisplatin-loaded liposomes as well as their cytotoxicity. We used rats with intracranial glioma C6[1] to evaluate the accumulation of DiI-labeled targeted liposomes in the brain tumor after intravenous injection. Using fluorescence microscopy and imaging system IVIS Spectrum-CT, we analyzed the accumulation and distribution of liposomes in the brain. We compared distribution of free CDDP and CDDP-loaded liposomes in rats with intracranial glioma C6 after single i.v. injection using ICP-MS.
Results and Discussion: The main obstacle impeding further advancement of liposomal CDDP formulations is low cisplatin solubility in water, leading to insufficient loading capacity (LC) of these nanocontainers[2]. We prepared stable negatively charged liposomes with a high LC (25-28% w/w). All formulations displayed a pH-depended sustained drug release profile without an initial burst release. The fluorescent-labeled targeted liposomes showed enhanced internalization in comparison with untargeted nanocontainers or non-specific IgG-targeted nanocontainers in glioma cells. This led to a considerable increase of cytotoxicity of the CDDP-loaded targeted liposomes in VEGF-overexpressed cells. Using fluorescence microscopy, we analyzed appropriate brain sections with additional staining of the astroglial shaft by mAb to GFAP that separates the tumor from the parenchyma. This allowed us to detect the accumulation of nanoparticles, both in the glioma and the peritumoral space. The comparative analysis of brain sections revealed the substantial distinction in the accumulation of different liposomes in the tumor tissue at 48 h after single i.v. injection. We observed the sharp difference between the malignant and normal tissues due to the selective accumulation of VEGF-targeted liposomes. We demonstrated that VEGF-targeted nanoparticles might penetrate through abnormal blood vessels of the disturbed blood brain barrier (BBB) into the extracellular space of the glioma, where antibodies selectively recognize expressed VEGF.
Conclusions: We designed the cisplatin-loaded liposomes conjugated with monoclonal antibodies to VEGF or VEGFR2 for targeted delivery of cisplatin to the intracranial glioma. These nanocontainers may lead to efficient accumulation of drug in brain tissues and an enhanced therapeutic effect of liposomes loaded with anticancer drugs.
RSF grant 14-15-00698, RFBR grants 12-04-31731 and 13-04-01383
References:
[1] Chekhonin VP, Baklaushev VP, Yusubalieva GM, Pavlov KA, Ukhova OV, Gurina OI. Modeling and immunohistochemical analysis of C6 glioma in vivo. Bull Exp Biol Med 2007; 143(4): 501–509.
[2] Oberoi HS, Nukolova NV, Kabanov AV, Bronich TK. Nanocarriers for delivery of platinum anticancer drugs, Advanced Drug Delivery Reviews, 2013, 65 (13), 1667–1685