| BackgroundAt present,the incidence of prostate cancer in our country is increasing year by year,which now becomes a serious threat to men’s health.As we all know,early diagnosis and treatment of prostate cancer is beneficial to the patients’ prognosis.However,current technologies to diagnose early stage of prostate cancer are of low sensitivity and weak specificity.Prostate cancer is an androgen-dependent cancer,so in most cases,advanced prostate cancer uses androgen as an effective treatment.However,among those cases,most of them will develop into the androgen-independent prostate cancer soon,when patients are receiving the androgen treatment.At present,the methods for treating androgenindependent prostate cancer are insufficient.Due to characteristics of the incompleteness and the wide vascular endothelial gaps in tumors,we designed a nanoscaled ultrasound contrast agent which can go through the tumor vessel to target cancer cells at the cellular or molecular level.Prostate specific membrane antigen(PSMA)is one of specific antigens,which appears on the surface of cell membrane in prostate cancer.As a significance target for prostate cancer,it has now been widely used to diagnose and treat prostate cancer.Doxorubicin is an anti-tumor drug which wildly used in clinic,but it also has the side effects on health cells at the same time.Hence,while applying doxorubicin at the local site to increase the inhibition effect,the side-effect of the other normal cells will be significantly decreased.PSMA-targeted nanobubbles had been made in previous research and its imaging in prostate cancer is pretty good.Thus,this research is to prepare a kind of PSMA-targeted nanobubbles loaded with doxorubicin.This nanobubbles can image prostate cancer,release the doxorubicin destroyed by the ultrasound mechanical destruction.And then the concentration of doxorubicin is increased around the prostate tumor.This novel agent could provide a new method to diagnose and treat prostate cancer.ObjectiveOur study is to prepare PSMA-targeted nanobubbles whose shells load doxorubicin and investigate its basic characteristics,contrast-enhanced effect in vivo and the effectiveness under ultrasound irradiation towards prostate cancer cells.MethodsDoxorubicin-loading nanobubbles were prepared with mechanical vibration method,After that,the anti-PSMA monoclonal antibody was attached to the surface of doxorubicin-loaded nanobubbles with streptavidin-biotin method.The characteristics of nanobubbles were observed by light microscope and transmission electron microscopy,and a fluorescence microscope was used to observe doxorubicin and PSMA monoclonal antibodies.The size and electric potential were analyzed by Marlvern Zetasizer Nano ZS90.A microplate reader was used to investigate the drug concentration in the nanobubbles.To explore the best intensity and time of irradiation in ultrasonic therapy apparatus,the drug-releasing rate was detected under irradiation of ultrasonic therapy apparatus at the sound intensity of 1.0 W/cm2.Two contrast agents,including targeted-drug nanobubbles and ordinary microbubbles,were utilized in livers and kidneys of the laboratory mice,and the contrast-enhanced effects were analyzed.The therapeutic effectiveness of doxorubicin-loading nanobubbles on prostate cancer cells was conducted,and the prostate cancer cells with drug-loaded nanobubbles under ultrasonic irradiation was observed by scanning electron microscopeResultsThe doxorubicin-loaded and PSMA targted nanobubbles have proper sizes and were evenly distributed.The shells of the nanobubbles showed red fluorescence because of doxorubicin.The surface of targeted nanobubbles showed green fluorescence because of its connection with FITC-IgG polyclonal antibody.The average diameter of the nanobubbles was(793.80±92.05)nm,and the electric potential was(8.74±2.41)mV.The doxorubicin concentration in the nanobubbles was(1.18±0.11)mg/ml.The best radiation intensity and time of ultrasonic therapeutic apparatus were 1 W/cm2 and 18 s.The doxorubicin release efficiency was(48.65 ± 3.99)% under ultrasonic irradiation.The contrast-enhanced ultrasound showed that the peak time,ascending time and contrast duration of the nanobubbles were significantly longer than that of ordinary microbubbles(P<0.05),but their peak intensities have no statistical difference(P>0.05).The therapeutic effect of the doxorubicin-loading and PSMA targted nanobubbles group on C4-2 and PC-3 was stronger than that of single doxorubicin group,blank nanobubbles group,targeted nanobubbles group and blank control group,but the difference between doxorubicin-loading nanobubbles and targted doxorubicin-loading nanobubbles was not significant.The surface of C4-2 group without irradiation was smooth with no channel on it,while the surface of C4-2 group after irradiation was rough with some channels.ConclusionThe stable doxorubicin-loading and PSMA targeted nanobubbles are successfully prepared,which have a good effect on prostate cancer treatment and a nice contrast-enhanced effect in vivo. |
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