131I-labelled c(RGDyk)-targeting doxorubicin thermosensitive nanoliposome and preparation and application thereof
A nano-liposome and doxorubicin technology, applied in the field of medicine, can solve the problems of limiting the application of hyperthermia, inability to ensure constant temperature, uneven distribution of thermal effects, etc., achieve good anti-tumor effect, good biological safety, and reduce damage Effect
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Embodiment 1
[0041] Example 1: Adriamycin thermosensitive nanoliposome (DPPC-DOX-Fe 3 O 4 liposome) preparation and characterization
[0042] Accurately weigh 2mg of cholesterol, 16mg of DPPC, and 2mg of PEG-DSPE in an eggplant-shaped bottle. Slowly add 3ml of methanol and 6ml of chloroform. After a short period of time, the mixture is quickly mixed with ultrasound, and then it is rotary evaporated into a round, white, uniform lipid film. Add 30mg Fe to 10mL0.3mol / L citric acid solution (pH4.0) 3 O 4 , After a short time ultrasonic mixing, the lipid membrane is hydrated to prepare liposomes. Add adriamycin hydrochloride to the prepared liposome and use 1mol / LNa 2 HPO 4 The pH value was adjusted to 7.0, and the magnetic thermosensitive adriamycin liposome was prepared in a water bath at 55°C for 30 minutes.
[0043] The liposomes were diluted 10 times and spotted on copper mesh. After drying, the characteristics and particle size of liposomes were observed by transmission electron microscope (TE...
Embodiment 2
[0045] Example 2: 131 I-labeled c (RGDyk) targeting doxorubicin thermosensitive nanoliposomes ( 131 I-RGD-DPPC-DOX-Fe 3 O 4 liposome) preparation and labeling conditions optimization
[0046] Weigh 1.5mgc (RGDyk) and dissolve it in 5μL methanol, add 45μL PBS, and mix with ultrasound for a short time; weigh 1mg chloramine-T and partial sodium sulfite to prepare a 10mg / mL corresponding solution. Take 1mCi 131 I. Add to the mixture of RGD cyclic peptide solution and chloramine-T, react at room temperature for 5 minutes, and add sodium metabisulfite to terminate the reaction. Mix the product with 15ml water through the C18 column, and then rinse the C18 column 3 times with 15ml water, use 300μL hydrochloric acid ethanol as the eluent, and recover the mark 131 I's c (RGDyk).
[0047] Accurately weigh 1.5 mg DSPE-PEG-NHS and dissolve it in freshly distilled anhydrous and ammonia-free DMF. Add DSPE-PEG-NHS solution dropwise 131 The I-cRGD solution was stirred at room temperature and prot...
Embodiment 3
[0048] Example 3: Different concentrations 131 I-labeled c (RGDyk) targeting doxorubicin thermosensitive nanoliposomes ( 131 I-RGD-DPPC-DOX-Fe 3 O 4 liposome) impact on cells
[0049] MCF-7 cells were added to the 96-well plate and divided into four groups. The liposomes prepared in Examples 1 and 2, free DOX and the control group were added respectively, and the DOX concentrations were added at 5μg / mL, 10μg / mL, and 20μg / mL, 30μg / mL, 40μg / mL corresponding liposomes and free drugs, add 20μL of MTT solution to each well after 48 hours, continue to incubate for 4 hours and then aspirate the supernatant, add 150μl of DMSO to each well, shake and dissolve for 10 minutes. Measure the OD value at 492nm with a microplate reader. According to the formula "cell inhibition rate (%) = (control group OD value-each experimental group OD value) / control group OD value × 100%" to calculate the cell inhibition rate, see the results Figure 4 . The IC50 of free DOX is about 25μg / mL. The result...
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