| PurposeThis project aims to develop a drug-loaded magnetic nanocomposite devices which carrying docetaxel and Fe3O4 nanoparticles, biodegradable and manetic mediated hyperthermia exposure under alternative magnetic field (AMF). Optimization based on the preparation of process conditions, evaluated biological safty and as mediated thermochemotherapy with sustain drug release to treat tumor in depth study. To clear devices mediated thermochemotherapy effect and establish relationship between treatment parameters and effect, determining optimal treatment parameters for the new, safe, efficient devices mediated induction thermochemotherapy.Methods1. Preparation of drug-loaded magnetic nanocomposite devices and characterization of physical and chemical properties1.1 Preparation of drug-loaded magnetic nanocomposite devices and parameter optimization:Drug loaded nanocomposite devices were prepared by the solvent casting technique with dichloromethane as the solvent.1.2 Characterization of physical and chemical properties:the morphology of magnetic nanocomposite devices was detected by scanning electron microscope (SEM). Thermal analysis used differential scanning calorimetry (DSC). The heating profiles of the nanocomposite devices with different MNPs contents under alternative magnetic field of 300kHz. Drug release in vitro was detected by high performance liquid chromatography (HPLC). Degradation was conducted in PBS in vitro and biodegradation in vivo used guinea pig.2. Biological safety evaluation of magnetic nanocomposite devices2.1 In vitro evaluation:Following the People's Republic of China T16886.5 (GB/medical instrument biology evaluation part 5:in ivtro cytotoxicity test), mice fibers L929 cell line as the research object, incubating with polymer devices, magnetic nanocomposite devices. After 2,4,7 days of incubation, check cells form and growth state with the microscope and judge relative growth rate by CCK-8 kit. Thus, evaluation biological safety of magnetic nanocomposite devices in vitro.2.2 In vivo evaluation:Following the People's Republic of China T16886.6 (GB/biological appraisement medical devices implanted in section 6:guinea pig as research object with relevant provisions after local reaction experiment, clinical medical PE film as positive counterpart. Magnetic nanocomposite devices and PE films were implanted in the back muscles in guinea pigs, take routine test of the blood and blood biochemical test to evaluate devices caused whether systemic inflammation or the influence of kidney function or not. Regularly executed after implantation, anatomic observation about surrounding tissue inflammation and whether there will be further pathologic study judgment.3. Cytotoxicity of drug-loaded magnetic nanocomposite devices based on glioma3.1 Cytotoxicity of chemotherapy by sustain release study:prepare drug loaded polymer devices and incubate with human brain U251 cells line compared with clinical injection Taxotere(?).3.2 Cytotoxicity of hyperthermia treatment to the gliomas:prepared single Fe3O4 nanoparticles loaded devices. Anti-glioma activity of the cell product was assessed using in vitro cytotoxicity assays against the Glioblastoma multiforme (GBM) cell line U251 in suspension and in adherent culture. Hyperthermia at 43℃exposure 30min, detected cell proliferation rate using CCK-8 kit, evaluated thermotherapy devices anti-tumor effect.3.3 Cytotoxicity of thermochemotherapy treatment to the gliomas:prepared drug-loaded magnetic nanocomposite devices and explored the influence of comprehensive treatment mode to U251 cell proliferation. To clear thermotherapy and chemotherapy exist collaborative gain effect or not by calculating webber coefficient.4. Anti-tumor profile of drug-loaded magnetic nanocomposite devices study4.1 Establish tumor model:the in vivo study was perfomed tumor bearing nude mice. Xenografts of human glioma cell lines would bu established by subcutaneous inoculation of U251 cells into the hind legs of Balb/c nude mice.4.2 Treatment:for animals under placebo and experimental groups, a small incision by aseptic surgery was performed on the skin and tumor was reached after which 2/3 tumor volume would be carefully excised. The magnetic nanocomposite devices would then be implanted well onto the residual tumor bed and subsequently the wound would be closed using subcutaneous suturing. Treatment was carried out on days three, six and nine after surgery using an alternating magnetic field (AMF) operating at a frequency of 300kHz and variable field strength of 110.2Gs. Through measuring nude mice tumor size after treatment, observated nude mice survival and pathological to comprehensive analysis and evaluation of magnetic nanocomposite devices anti-tumor ability.Results 1. Drug-loaded magnetic nanocomposite devices was prepared by the solvent evaporation method. Poly (D, L) lactide-trimethylene carbonate (PLA-PTMC) copolymer was employed as the film matrix. The devices is flexible, smooth and homogeneous. The desired temperature can be achieved by appropriate choosing the MNPs content or adjusting the field intensity. The cumulated release profiles of docetaxel from the devices in 35 days were investigated to be 90% in vitro. Biodegradation rate in vivo was higher than in vitro. Surprisedly, magnetic nanocomposite devices was completely degraded and absorbed by arganization.2. Biocompatibility of magnetic nanocomposite devices in vitro and in vivo were proved to be safe. Devices did not affect the cells form and proliferation. In vivo devices did not cause systemic inflammation and liver and kidney function abnormal.3. In vitro cytotoxicity study, the proliferation inhibition rate of U251 gliomas cell was highest for the thermochemotherapy groups and MIH significantly enhanced docetaxel effect to kill tumor cell as time went on. In vitro cytotoxicity results indicated the bi-modal cancer treatment approach for combined MIH and chemotherapy is more effective than the mono-modal treatment by docetaxel treatment (p<0.01).4. Tumors in the thermochemotherapy group shrunk most significantly, as compared with those of the control groups and hyperthermia treatment only. Median survival was improved significantly. The study suggested that magnetic biodegradable film implants releasing docetaxel can be promising post-surgical local delivery devices for thermochemotherapy against glioma.ConclusionThe nanocomposite devices exhibit advantageous feature for a facilitated drug delivery from the nano-carriers and the magnetic mediated heating potential is adequate for hyperthermic treatments. We thus conclude that even though further detailed investigations are still necessary, tentative use in local tumor therapies aiming at a specific chemotherapeutic release in combination with magnetic heating is promising and feasible in the long term. |
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