Antitumor Effect Of Dextran-magnetic Layered Double Hydroxide-fluorouracil Targeted Liposomes In Nude Mice With MGC-803Solid Cancer Model

Fluorouracil (FU) is one of the commonly used chemotherapy drugs of gastrointestinalcancer in clinic treatment. In order to overcome the gastrointestinal side effects and thedefects of short half-life of FU, we have prepared dextran-magnetic layered doublehydroxide-fluorouracil (DMF) drug delivery system by intercalation5-Fu into the laminatesof a magnetic layered double hydroxide and further modification with dextran. In this paper,by combining the active targeting of liposomes with the sustained release and magnetictargeting of DMF drug delivery system, we prepared a magnetic nanometer liposomes(DMFL) as injection. Using tumor-burdened mice as experimental animals, we observed theinhibition effect of DMFL on solid cancer in vivo with the integrated monitoring, tissuemorphology, cell biological detection and immunohistochemical techniques.A solid tumor nude mouse model of human MGC-803gastric cancer have beenestablished successfully by subcutaneous infection,10d after implantation, mice wereassigned randomly to four groups (n=7) and were treated intraperitoneally every day withone of the following treatments:(i)saline,(ii)5-Fu (20mg5-Fu·kg-1),(iii) DMFL no magneticfield (20mg5-Fu·kg-1),(iv) DMFL (20mg5-Fu·kg-1) with an external magnetic field. Micewere weighed and the length (L) and width (W) of the tumor formed at the transplanted sitewere measured every other day, and the relative tumor weight was calculated as W2×L×1/2.Mice received17doses were sacrificed, tumor mass were weighed and the tumor inhibitionrate were calculated with the formula (1-tumor weight of treatment group/tumor weight of control group)×100%. The affect of DMFL on important visceral function was also inspectedwith HE staining method. Results show that the MGC-803gastric cancer model are easy toestablish subcutaneously with shorter period and higher success rate, so we used the model tostudy antitumor function of DMFL. DMFL showed good slow-releasing and magnetictargeting property, as5-Fu in DMFL could move directionally in the body under an externalmagnetic field, and maintain an effective cure concentration in tumor for a longer period,which enhances the killing effects of5-Fu on cancer cells and reduces damage to thesurrounding normal tissue. The mice in DMFL group (with applied magnetic field) was ingood condition, without obvious weight losing, and tumor grew slowly with a significantdifferences in mass from other treatment groups. DMFL did not show obvious harm toimportant organs compared with other groups.As a classical chemotherapeutics drug, FU can prevent the formation of pyrimidinenucleotide to induce apoptosis of tumor cells. To confirm whether FU-loaded DMFL drugsystem would more availably promote apoptosis or necrosis of tumor cells, we investigatedfurther cells in the tumor tissue by qualitative (HE staining, transmission electron microscope)and quantitative (flow cytometry) analysis methods in this paper. Results showed that in thecure group of DMFL under an external magnetic field, a large number of cells in the tumortissue showed obvious apoptosis characteristics, and organelles also appeared the lesionsymptom such as cavitation and atrophy; at the same time, a large number of DMF particlescould be found in the cytoplasm and main organelles such as the nucleus and mitochondria bytransmission electron microscope. The TEM morphology of DMF particles in tumor tissuewas same as that in pure solution with hexagonal flake and spherical shape, and the apoptoticand necrotic cells of DMFL group under an external magnetic field were more obvious thanfluorouracil group and DMFL group without magnetic field. The result of flow cytometryanalyzer showed that the DMFL were more effective to promote apoptosis of tumor cell andinhibit growth of tumor cell under an external magnetic field. The inhibitory effect of DMFL under an external magnetic field was the strongest under the same fluoride doses, whichportends that DMFL could be used in tumor treatment with smaller doses to achieve bettereffect in the future.To probe into the mechanism for DMFL delivery system to inhibit tumor growth in vivo,the protein expressions of vascular endothelial growth factor (VEGF) and microvasculardensity (MVD) were assayed with immunohistochemistry staining, and the proteinexpressions of Ki-67were determined by immunofluorescence staining. The results ofimmunohistochemistry indicated that the protein expression of VEGF and MVD under anexternal magnetic field were reduced significantly (P <0.05) with a positive correlationbetween the protein expression of VEGF and MVD. The results of immunofluorescencemethod indicated that the protein expression of Ki-67in DMFL group under an externalmagnetic field was also reduced significantly (P <0.05). These results demonstrated thatDMFL liposomes could not only effectively block the transforming of deoxidization uracilnucleotide into thymine nucleotide, but also weaken the effect of angiogenesis and suppressthe formation of new blood vessels by inhibiting the expression or decreasing actin activity ofVEGF, consequently inducing apoptosis of tumor cell and inhibiting proliferation of cancercell. In other words, in addition to kill the tumor cell directly, DMFL also could increase thenecrosis risk of tumor tissue due to ischemia.