Study Of Doxorubicin PEG-PLA Ultrasonic-microbubbles For The Treatment Of Breast Cancer

Objective:To prepare Doxorubicin-PEG-PLA block copolymer micelles ultrasonic-microbubbles,assay the drug loading(DL)and encapsulation efficiency(EE)and average particle diameter of micelles.To establish bearing breast cancer mouse,with in vivo treatment and observed tumor growth.To study Doxorubicin-PEG-PLA block copolymer micelles ultrasonic-microbubbles for the treatment of breast cancer.Method:1.To synthesize poly(ethylene glycol)-poly(D,L-lactic acid)(PEG2000-PLA1300)block polymer by the ring-opening polymerization.2.To prepare Doxorubicin-PEG-PLA block copolymer micelles ultrasonic-microbubbles:by PEG2000-PLA1300 and film dispersion method to envelop Doxorubicin,hydration to micelles,addition perfluoropentane(PFP),by the ultrasonic wave cytoclasis to ultrasound 1 min into microbubbles.To observed the shape of the micelles before and loading drug and ultrasonic-microbubbles by transmission electron microscope(TEM).To study the micelles size,drug loading,encapsulation efficiency by laser particle size analyzer and high performance liquid chromatography.3.In vitro test:1)cell inhibition ratios assay(MTT assay):mice breast cancer cell line 4T1 and human breast cancer cell line MCF-7 were treated with 0.01?0.1?1?5?10?20?50?100?g/mL concentrations of Doxorubicin?Doxorubicin PEG-PLA micelles and Doxorubicin PEG-PLA ultrasonic-microbubbles,with the group of Doxorubicin PEG-PLA ultrasonic-microbubbles was ultrasoniced by Low Intensity Focused Ultrasound,every group incubate 0.5?24?48?72 h,control cells by MTT assay inhibition ratios.2)Flow cytometry:to evaluation the ability of get into the cell between Doxorubicin and Doxorubicin PEG-PLA ultrasonic-microbubbles.3)Confocal:4T1 cell culture in the 35 mm glass bottom Petri for 24 h,add Doxorubicin PEG-PLA ultrasonic-microbubbles(1 mg/mL)200 ?L for 10 min,evade light,suck medium,PBS wash,4%paraformaldehyde 2mL fixation 10 min,Laser Confocal observation.4.In vivo anticancer assay:after established tumor-bearing mice model,tail intravenous injected the same dose of Doxorubicin?Doxorubicin PEG-PLA micelles and Doxorubicin PEG-PLA ultrasonic-microbubbles,physiological saline,the Doxorubicin PEG-PLA ultrasonic-microbubbles group after be injected min to be treated ultrasonic,regular observation the changes of tumor scales and mice weight.39 days after inoculation,sacrifice mice.Results:The drug loading and encapsulation efficiency were 12.95%and 98.97%,average particle size of 140.4 nm.MTT assay indicated that Doxorubicin PEG-PLA ultrasonic-microbubbles could inhibit the growth of MCF-7 and 4T1 cells better.The results of in vivo test indicated that after the treatment of Doxorubicin?Doxorubicin PEG-PLA micelles?Doxorubicin PEG-PLA ultrasonic-microbubbles and saline,tumor in Doxorubicin PEG-PLA ultrasonic-microbubbles groups grew slowly,the tumor volumes were the smallest,and there was significant difference between Doxorubicin PEG-PLA ultrasonic-microbubbles treated group with Doxorubicin?Doxorubicin PEG-PLA micelles,and saline treated groups(P<0.05).Conclusion:Doxorubicin PEG-PLA ultrasonic-microbubbles have the best inhibition of the growth of breast cancer tumor.The combination of ultrasonic and microbubbles was a potential nanosale technique.