Study On Preparing Technique Of Hydroxycamptothecin Thermal Magentoliposomes And Pharmacodynamics Evaluation

ObjectHigh intensity focused ultrasound (HIFU) as a new technology of heat treatment, which is used in malignant tumor with non-invasive treatment. Not only has small side effects, and the treatment effect is obvious. Thermal magnetic liposome as the drug carrier which can play the role of heat treatment and chemotherapy in recent year. Compared with ordinary liposome, the thermal magnetic liposome has stronger organization targeting and controlled release characteristics. Combining both of the technology in the treatment of tumor, on one hand, most of tumor cells can be killed by the HIFU treatment, on the other hand, the thermal magnetic liposome can release in heating of HIFU and realize second-kill to the tumor cells, achieve the best effect of cancer treatment.(Hydroxycamptothecin, HCPT) as the main medicine, explore the preparation technology of Hydroxycamptothecin thermal magentoliposomes,and evaluate related quality projects of the thermal magentoliposomes, such as the targeted, thermal sensitive and cell toxicity characteristics, and joint the new heat treatment-high intensive focused ultrasound, examine the anti-tumor effect of two technologies. MethodsThis research includes four sections:1Establishment of analytical method of samplesMethod of RP-HPLC with ultraviolet detector was applied to detect the content of the Lactone structure and Carboxylate structure. The chromatographic condictions were optimized and the methodology was investigated.2The preparation and the evaluation of Hydroxycamptothecin thermal magentoliposom.esUse the appropriate methods and material to prepare the thermal magentoliposomes by the use of film dispersion-ultrasonic method, and evaluate the envelopment, the drug loading, the meaning diameter and the targeted, thermal sensitive characteristics of the thermal magento-liposomes.3The cell toxicity test investigation of the Hydroxycamptothecin thermal magentoliposomesBecause of HCPT has good treatment effect to the liver cancer, so take the Huh-7cell as the study object, use the MTT colorimetric method, compared with the HCPT thermal magentoliposomes group and HCPT injection group on the inhibition of cancer cell, calculate the inhibition rate, investigate the antitumor activity of two groups and the different concentrations among them.4Investigation of the treatment of Hydroxycamptothecin thermal magentoliposomes joint HIFU technique for tumor-burdened mice.Implant the liver cancer cells H22in the mice, and then establish hypodermic tumor model. Set up four groups as follow:(1) blank group;(2) HCPT injection group;(3) HIFU treatment group:tumor without magnetic field;(4) HCPT thermal magentoliposomes group: add iron boron on the tumor surface magnets as additional magnetic field;(5) HCPT thermal magentoliposomes joint with HIFU group. Give medicine by the tail vein, measure the tumor size after giving medicine every day, observe the growth of the tumor, and evaluate the efficacy of the treatment.Result1Establishment of analytical method of samplesAccording to the nature of the drug, set up a chromatographic conditions to test the open loop structure and closed loop structure of HCPT.The chromatographic conditions were optimized and the methodology was investigated.The results show that the Lactone structure and the Carboxylate structure of HCPT under the condition of chromatographic separation is good, good symmetry, the linear range and precision, stability, recovery, etc which conform to the requirements of experiments.2The preparing and evaluation of Hydroxycamptothecin thermal magentoliposomesSelecting the appropriate preparation method and material, to prepare hydroxycamptothecin thermal magentoliposomes with phospholipids and cholesterol as raw materials, by the use of film dispersion-ultrasonic method,,a reasonable criterion of accurate evaluation was established.And the meaning diameter of liposomes was268nm,envelopment was41.8%and the drug loading is1.32%.The release of the thermosensitive liposomes is more than50%in60min and has obvious targeting.The optimized prearation condition for hydroxycamptothecin thermal magentoliposomes were obtained as follows:the ratio of lipid to the drug was1:30.The ratio of phospholipid to cholesterol was80:30. The temperature of preparation was42℃. The medium of the liposomes was phosphate buffer (pH6.8).The dosage of Fe3O4is HCPT:magnetic particle (w/w) for1:1-1.5. 3The cell toxicity test investigation of the Hydroxycamptotltecin thermal magentoliposomesUsing two different dosage form of HCPT as the experimental groups, and compared the tumor inhibition rate among the three groups at different concentration levels.The result of the tumor inhibition rate show as HCPT magnetic thermosensitive liposomes> HCPT injection group.Basically conformed to the principle that the Lactone structure of HCPT is better than the Carboxylate structure in the tumor inhibition4Investigation of the treatment of Hydroxycamptothecin thermal magentoliposomes joint HIFU technique for tumor-burdened mice.Implanting the liver cancer cells H22in the mice with appropriate cell number and implanted volume, can establish the uniform size of the animal model. While investigating the antitumor activity of the Hydroxyca-mptothecin thermal magentoliposomes, we can found that the liposome group has obvious effect compare with the blank groups.The mice tumor diameter the of the given medicine groups is small and inhibits the growth of tumors.The two groups with the HIFU treatment appear gradually death, and can’t examine its final therapeutic effect.Conclusion1Study on preparing technique of HCPT liposomesThe HCPT liposomes in phosphate buffer storage process, the liposomes is more stable in4℃environment than in the room temperature and the liposomes appear statified phenomenon in the room temperature. The leakage rate of the HCPT liposomes large in short time,and then slow down gradually. So we must pay attention to the environmental factors on the stability of the storage of liposomes. 2Study on preparing technique of the thermal liposomeThe natural thermal can be used as the heat-sensitive material to prepare the thermal liposome by the use of film dispersion-ultrasonic method. Investigate the influence factors of liposome phase transition temperature, also can get better thermal liposomes.3Study on preparing technique of magentoliposomesThe magnetic Fe3O4surface modification can be finished by the use of sodium oleate, which can improve the lipophilic.And the magnetic powder can combine with the surface of the liposome better,establish a stable magnetic liposomes suspension system.4The study of pharmacodynamics with thermal magentoliposomes of HCPT in vitroIn the cell toxicity test investigation, the Lactone structure of the HCPT have better antitumor activity than the Carboxylate structure, closed loop structure of the HCPT was encapsulated in liposome, so in test can find that HCPT liposome and HCPT injection which in the same concentration, the cancer inhibition rate of the former is better than the latter.5The study of antitumor with thermal magentoliposomes of HCPT combines with HIFUIn the test of chemotherapy treatment combined with HIFU, because of the HIFU treatment is suitable for person in clinical and the mice is selected to use in the test, so when using the HIFU treatment, in order to reduce the cause of death by treatment and promote the treatment effect, the treatment range of positioning and energy must be control.