Application And Mechanism Of CuS-MnS₂ Nanoflowers As A Theranostic Agent In Ovarian Cancer

Background:Ovarian cancer keeps a low early diagnosis rate and a high mortality,which seriously endangers women's health.The treatment is mainly surgery + chemotherapy.In recent years,new diagnostic techniques such as tumor-related gene sequencing and molecular targeted therapy have emerged,while the effect is quite limited,and the early diagnosis rate and five-year survival rate of ovarian cancer patients have not been significantly improved.As nanotechnology developing rapidly,Scholars put forward the concept of theranostic in recent years,which means integration of diagnosis and therapy.The imaging function of nanomaterials can help diagnosis,guidance and monitoring the treatment.The photothermal conversion efficiency combined with NIR irradiation provides targeted,visible and manageable photothermal therapy.At present,there is little report of nanomaterials applied in obstetrics and gynecology.This study is aimed to evaluate the theranostic efficacy of newly produced nanomaterials for ovarian cancer,and explore its mechanism,expecting to provide new diagnosis method and treatment for ovarian cancer.Objectives:1.Synthesize CuS-MnS2 nanoflower and define its characterization;2.Evaluate the toxicity of CuS-MnS2 nanoflower;3.Investigate the PPT/PDT effect of CuS-MnS2 on ovarian cancer cells;4.Explore the mechanism of CuS-MnS2+ NIR killing ovarian cancer cells.Methods:1.The CuS-MnS2 nanoflowers were synthesized by chemical bath deposition.The crystal structure was observed by X-ray diffractometer.The morphology,size and element distribution were observed by transmission electron microscopy(TEM),scanning electron microscopy(SEM),and energy dispersive spectroscopy.The UV-Vis-NIR absorption spectrum of CuS-MnS2 nanoflower is detected using a spectrophotometer.To evaluate its photothermal performance,CuS-MnS2 nanoflower solution were irradiated by near infrared laser irradiation(NIR)at 808 nm,at the same time infrared thermal imaging was collected,temperature was recorded to mapping heating curves.The materials were configured into agarose gel solutions to different concentration,then tested MRI imaging efficacy.2.Human ovarian cancer cells were co-cultured with CuS-MnS2 nanoflowers,and the cell viability was tested by CCK-8 kit.Human red blood cell suspension were coincubated with different concentrations of CuS-MnS2 nanoflower,in order to evaluate and the erythrocyte toxicity of CuS-MnS2 by hemolysis.3.Pre-cultured human ovarian cancer cells added CuS-MnS2 nanoflowers,were irradiated by near-infrared laser to observe the photothermal ablation.A2780 cells were inoculated into mice,and the tumor-bearing mice were injected with CuS-MnS2 and PBS respectively,then irradiated with NIR to verify the vivo cancer cell ablation.4.A2780 cells with or without CuS-MnS2 were irradiated and then detected the apoptosis-related protein,anti-apoptosis-related protein,and necrosis-related protein by Western Blot.Results:1.The material is a flower-like CuS-MnS2 composite,which possesses a photothermal conversion efficiency(?)of estimated 67.5%and quite good photothermal stability,and also T1 enhanced MRI imaging function.2.In human ovarian cancer A2780 cells co-cultured with different concentrations of CuS-MnS2 nanoflower,The survival rate of A2780 cells decreases slightly at 0.5 mg/mL concentration of the nanoflower,but still higher than 75%.In red blood cells co-incubated with different concentrations of CuS-MnS2 nanoflowers,mild hemolysis was overseved at a lower concentration,while not obvious was found at the concentration of 0.5,1.0 mg/mL.3.With concentration above 0.5 mg/mL,CuS-MnS2 combined laser irradiation can cause significant A2780 cell death.Temperature of the tumor in tumor-bearing mice treated with CuS-MnS2+NIR increased to above 55 0C rapidly during irradiation,while the mice treated with PBS+NIR below 40 °C.With H&E staining,the tumor tissue in CuS-MnS2+NIR group was found obvious cell damage.4.CuS-MnS2+NIR can induce intracellular reactive oxygen species.Down-regulatiorn of apoptosis-associated protein,up-regulation of anti-apoptosis-associated proteins,and up-regulation of necrosis-associated protein were detected by WB experiment.Conclusions:1.CuS-MnS2 nanoflowers possess a high photothermal conversion efficiency and photothermal stability,also T1 enhanced MRI imaging efficacy.2.CuS-MnS2 nanoflowers hold a low cytotoxicity.In the hemolysis test,CuS-MnS2 nanoflower cause mild hemolysis at a lower concentration of,while no obvious hemolysis at increased concentrations.We consider that products of hemolysis such as hemoglobin and iron ions may be adsorbed by CuS-MnS2 nanoflowers.The blood compatibility of CuS-MnS2 nanoflowers calls for further study.3.CuS-MnS2+NIR perform good in photothermal therapy for ovarian cancer and have photodynamic effect.4.Activating the programmed necrotic pathway of cells is the main mechanism of CuS-MnS2+NIR inducing cell death.