Tocopheryl Pullulan-based Self-assembled Nanomicelles For Anti-cancer Drug Delivery

Three a-tocopherol succinate (TOS)-modified pullulan amphiphilic polymers, PUTC1, PUTC2and PUTC3were synthesized by changing the mole ratio of TOS to pulullan. The structure of PUTC was confirmed by’HNMR and FTIR analysis. The CMC values of the PUTCs were determined to be38.0,8.0,4.3mg/L for PUTC1, PUTC2and PUTC3respectively by a fluorescence probe technique with pyrene as a probe. Transmission electron microscope (TEM) showed that the PUTC self-assembled nanomicelles prepared with dialysis method were dispersed uniformly with regular spherical shape. The particle sizes of PUTC self-assembled micelles determined by DLS were around160nm.10-HydroxyCamptothecin (HCPT), which was employed as a model drug, was loaded into PUTC nanomicelles by dialysis and ultrasound method. When the mass ratio of drug to carriers was1:10, the loading capacities of PUTC1, PUTC2, PUTC3were6.86%,8.14%,8.60%respectively, and the encapsulating efficiencies were73.68%,88.7%,94.10%respectively. In vitro release of HCPT from PUTC micelles demonstrated sustained release over80h in PBS buffer (pH7.4) at37℃. The release rate of drug from nanomicelles was different at varying pH, and when the pH was lower, the release rate was higher, demonstrating that PUTC micelles have pH-sensitivity to some extent.MTT assays demonstrated that the blank PUTC micelles were non-toxic to T lymphocyte, HEK293cells, and COS-7cells. HCPT-loaded PUTC nanomicelles showed similar cytotoxicity to that of free HCPT in SMMC-7721and MCF-7cells. The results of flow cytometry revealed that both free HCPT and HCPT-PUTC affected the S phase of cell cycle, and HCPT-PUTC induced more cells to apoptosis. The uptake of DOX-PUTC in MCF-7and SMMC-7721cells could be visualized quickly after cells were incubated with DOX-PUTC. The quantitative uptake of DOX-PUTC assayed by flow cytometry displayed that both MCF-7and SMMC-7721cells can uptake more DOX-PUTC than free DOX, and SMMC-7721cells uptake more PUTC-DOX than that by MCF-7cells. implying that the PUTC nanomicelles have liver-targeting.The images of confocal laser scanning microscopy (CLSM) showed that DOX-loaded PUTC nanomicells accumulated around nuclei of cells rapidly. The cellular uptake of micelles was found to be an energy-dependent and actin polymerization-associated endocytic process by the experiments of endocytosis inhibition.