A Novel Bimodal Imaging Agent Targeting HER2 Molecule Of Breast Cancer

PurposeTo prepare a promising agent for near infrared fluorescence/ultrasound dual-mode imaging,we add a near-infrared fluorescent(NIRF)dye-IR783 into nanobubbles,and then conjuate biotinylated anti-ErbB2 Affibody? with IR783-NBs.Also,we detect the targeting rate of IR783-NBs-Affibody toward HER2(+)breast cancer cell and its dual-mode imaging ability.Material and methods1.To prepare targeting bimodal imaging agent agent: DPPC and DSPE-PEG 2000 were mixed and chloroform was added to dissolve the mixture,then IR783 was added into lipid to prepare IR783-NBs,via improved thin-film hydration method.Then biotinylated anti-ErbB2 Affibody? molecules was combined with IR783 loaded NBs by avidin-biotin method.2.Characteristics of IR783-NBs-Affibody: to measure the size distributions of IR783-NBs-Affibody and SonoVue microbubble,particle size analyzer was used.To further investigate the IR783-NBs-Affibody structure,transmission electron microscopy was applied.The amount of the IR783 in IR783-NBs was determined by standard curve of UV–vis absorption spectra measured by a UV–vis spectrophotometer.A CCK-8 array was used to detect the biosecurity of IR783-NBs-Affibody.Bimodal imaging of IR783-NBs-Affibody was observed by laser scanning confocal microscopy(LSCM)and ultrasound contrast imaging.To detect the stability of IR783-NBs-Affibody,its changes of size distribution and concentration were measured.3.Targeting rate: the targeting rate of IR783-NBs-Affibody towards HER2(+)breast tumor cells was measured via LSCM and flow cytometry(FCM),compared with HER2(-)breast tumor cells.To further detect its HER2 targeting ability and tumor targeting ability,IR783-NBs-Affibody labled by DiO and IR783 was co-incubated with three kinds of breast cancer cells of different HER2(+)status.Result1.The prepared IR783-NBs-Affibody presented a uniform nanoscale size around 482.7 ± 54.3 nm,good biosecurity and stability over time.The encapsulation efficiency of IR783 was 15.09% in the conjugates leading to a successful NIR fluorescence and ultrasound enhancement imaging ex vivo.2.IR783-NBs-Affibody was able to automatically accumulate on BT474 cells with a highly increased targeting rate of 85.4% compared with previous NBs-Affibody of 26.6%.While,Affibody guided HER2-binding was only found in HER2 positive cell lines(BT474 and T-47D).And IR783 guided tumor-specific targeting was found in all three breast cancer cells,no matter what the HER2 status was.ConclusionThe obtained bimodal imaging agent IR783-NBs-Affibody providing both ultrasound and near-infrared fluorescence imaging signals,is able to achieve tumor-specific and HER2-specific targeting simultaneously.IR783-NBs-Affibody presents nanoscale uniform size,a low polydispersity,and favorable stability.For ultrasound mode,IR783-NBs-Affibody shows a promising enhanced ultrasound signal in vitro similar to SonoVue does;while,for NIR fluorescence mode,IR783-NBs-Affibody treated tumor cells exhibits high fluorescence intensity.Thus,IR783-NBs-Affibody is expected to achieve a high SNR when two imaging modalities are applied in breast cancer patients,which is helpful for tumor delineation and possible SNL trace.