A Sequential Targeting Theranostic Nanoplatform For Multi-modal Imaging And Synergistic Therapy Of Anaplastic Thyroid Carcinoma

Anaplastic thyroid carcinoma(ATC),as an extremely malignent tumor,lacks a satisfactory therapeutic method and a diagnostic strategy.With the development of nanotechnology,multifunctional nanoprobes have been promoted in the flied of theranostics applications.The difficulty point of nanotechnology is the effective accumulation of nanoparticles in tumor sites.By breaking the limitation of traditional target,such as low efficacy and frequent recurrence as the results of low accumulation,s,we successfully synthesized a sequential targeting nanoplatform(IR825@Bev-PLGA-PFP NPs)with theranostic functions.This nanoplatform specifically binds to the tumor cell surface and further cascade into mitochondria to achieve enough accumulation of nanoparticles in the tumor regions.On the one hand,by the assistant of targeting capability,tumors have been completely eradicated without any side effect and residual tissue under the external laser(808 nm)irradiation.On the other hand,comprehensive imaging information could be detected by multimodal molecular imaging to achieve the purpose of early diagnosis and real-time monitoring.Objectives: In this study,we synthesized a Bev-modified multifunctional nanoplatform IR825@B-PPNs loaded with IR825 and PFP for sequential targeting and multimodal imaging-guided synergistic therapy.To observe their characterization and physicochemical properties,to detect their US/PA/FL imaging capability,targeting ability and to evaluate the antitumor synergistic therapy effect in vitro and in vivo.Methods: Polylactic acid-glycolic acid(PLGA-COOH)modified by carboxyl group was used as carrier.IR825@PLGA-PFP NPs(which are termed IR825@PPNs)encapsulated with PFP and IR825 were prepared using a double emulsion method.Bevacizumab was covalently connected on the shell of IR825@PPNs using the carbodiimide method.The characterization and physicochemical properties of IR825@B-PPNs,including particle size,zeta potential,uniformity,morphology,US-vis absorption spectrum,in vitro photothermal effect,ODV,stability,antibody linking efficiency,encapsulation efficiency and loading capability,were evaluated.The targeting ability of nanoparticles was detected in vitro and in vivo.And the distribution of NPs in the tumor sites was detected to confirm the optimal time for tumor diagnosis and treatment.The multi-model imaging of the NPs in vitro and in vivo was tested,and ultrasound imaging(including B-mode and CUES mode),photoacoustic imaging and fluorescence were performed respectively.As for the therapeutic effect,cck-8 method was used to analyzed in vitro cytotoxicity assay with different NPs samples on C643/LO2 cells.And the cells viability under laser irradiation based on the above results.The effect of synergistic therapy on cells apoptosis was analyzed by flow cytometry.And the staining dead/live cells were observed by CLSM after different treatment.In addition,western blotting assay was detected to analyze the effect of antiangiogenesis on the downstream pathway factors.In vivo assay,including biosafety assay,synergistic tumor therapy,histologic and immunohistochemical examination were detected.Results: IR825@B-PPNs,a multifunctional sequential targeting nanoplatform,was successfully prepared.IR825@B-PPNs exhibited a spherical morphology,a typical shell-core structure,a uniform particle size distribution,a negative zeta potential,a good long-term stability,and successfully modified with Bev on the surface.The NPs could specifically target VEGF on the C643 cell surface,and further accumulate into the mitochondria due to the encapsulation of IR825.The peak accumulation of the NPs in the tumor region occurred in 3 h after injection.As an excellent PTA with mitochondria targeting ability,IR825 can be applied as an outstanding PA/FL imaging agent for NPs in vitro/vivo.The laser-induced photothermal effect triggered the liquid-gas phase transition of PFP in IR825@B-PPNs,which performed US imaging.Based on the excellent targeting capability,sufficient nanoparticles were accumulated in the tumor site.IR825@B-PPNs loaded Bev and IR825 in single nanosystem achieved PTT in combined with anti-angiogenesis significantly,which improved the therapeutic efficiency in completely eliminating tumors without recurrence.Conclusion:In this study,a multifunctional sequential targeting nanoplatform(IR825@B-PPNs)was successfully engineered for achieving theranositic of anaplastic thyroid carcinoma.This nanoplatform has good biocompatibility and biodegradability.The sequential targeting ability promotes efficient accumulation in tumor sites,achieving synergistic antiangiogenic photothermal therapy(PTT)and multimodal molecular imaging.It provides a new lighthouse for ATC's theranostic strategy.