Application Of Indocyanine Green Nanoprobe In The Integration Of Diagnosis And Treatment For Colon Cancer

Objective:Colon cancer?CC?is the common malignant tumors of the digestive system.In recent years,the incidence and mortality rate are increasing.According to the statistics,the incidence of colon cancer in China has risen to fifth of the incidence of malignant tumor.Early detection and early treatment can effectively improve the cure rate and survival rate of colon cancer patients,improve the quality of life of the patients,Accurate diagnosis of colon cancer and effective treatment has become a hot and difficult point in clinical research.Traditional methods of treatment,including surgery,chemical therapy,radiation therapy and other toxic and side effects,multidrug resistance,relapse and other limitations are still difficult to overcome.The nanoscale probe technology,which combines multidisciplinary and interdisciplinary cooperation,is expected to overcome the disadvantages of low target biological utilization,Many adverse reactions to human beings,which can combine the diagnosis and treatment of colon cancer and provide new ideas for the diagnosis and treatment of colon cancer.In this study,Indocyanine green?ICG?was used to mark human serum albumin?Human serum albumin,HSA?in clinical medical applications.Based on the photothermal properties of ICG,a biocompatible and biodegradable HSA-ICG nano probe was designed,and HSA-ICG nano probe was observed in tumor imaging.The role of diagnosis and photothermal therapy is to explore a new method for the integration of colon cancer diagnosis and treatment.Methods:1.Organic solvent precipitation technology was used to prepare reduced HSA,and HSA was mixed with ICG to form HSA-ICG photosensitive nanoparticles.The morphology of HSA-ICG was observed under transmission electron microscope.The particle size of HSA-ICG was measured by dynamic light dispersion?Dynamic light scattering,DLS?method and the near infrared laser was irradiated to test the properties and stability of the photothermal transformation of the probe.2.The colon cancer HCT116 cells were prepared,the model of subcutaneous transplantation tumor of nude mice with colon cancer was established.The colon cancer cell like liquid added to the HSA-ICG probe was irradiated with 980nm NIR laser.The survival rate of HCT116 cells added to HSA-ICG at different concentrations was detected by MTT colorimetry,and the sex of the photothermal conversion was measured.The Annexin V-FITC/PI was used to observe the apoptosis before and after irradiation.3.The FolateRsense^TM680,HSA-ICG and ICG probes were injected into the 6nude mice of the same group and the same number of nude mice with colon cancer subcutaneous tumor model,and IVIS was used for FLI to measure the average fluorescence intensity of ROI at each time point.4.To build PTT equipment,laser irradiated experimental animals for photothermal therapy of tumors.Subcutaneous tumor model nude mice were divided into 2 groups,with 3 rats in each group.HSA-ICG probe and commercial Folate Rsense^TM680 probe tail vein were injected into mice respectively.980nm subcutaneous laser was used to irradiate the subcutaneous tumor.After radiotherapy,FLI imaging was performed on the tumor to observe the change of fluorescence signal at the tumor site.After 36 hours of treatment,the tumor tissues were observed for pathological changes.The tumor targeting,fluorescence imaging and photothermal therapy were studied in nude mice with colon cancer xenograft models.Results:1.The characterization of HSA-ICG nanoparticles showed that the particle size of HSA-ICG was between 20-50nm.A high resolution transmission electron microscope was used to observe the clusters of white powder samples from the naked eyes of HSA-ICG nanoparticles.Most of them were dots or flakes,and small parts gathered together to form spheroids.2.When the concentration of HSA-ICG is 400ug/ml,the temperature of the solution increases with the prolongation of the time of the sample and the temperature of the sample solution can rise to more than 45?after 5 minutes of continuous irradiation.With the concentration of HSA-ICG solution increased echelon near infrared laser irradiation,the heating rate increase.When the concentration is greater than200ug/ml,the 4 samples can reach 42?in 5 minutes.It shows that the HSA-ICG particles have good photothermal conversion performance and the temperature of the measured HSA-ICG solution is basically consistent with the 980nm near infrared laser irradiation.It shows that the studied HSA-ICG nanoparticles have good photothermal stability.According to the cytotoxicity test results,the HSA-ICG cells are at different concentrations at different concentrations.There was no significant difference between survival rate and ICG,and no significant toxicity was found.It is proved that the synthesized HSA-ICG has reliable biosafety.3.The near infrared laser continued to irradiate the HCT116 colon cancer cell culture medium with HSA-ICG particles.It is concluded that the temperature of the sample medium increases with the increase of the concentration of the added HSA-ICG solution,but it will not continue to rise after the temperature rises to a certain extent,indicating that the photothermal conversion properties of HSA-ICG in the colon cancer cells are stable,and the flow detection,after the 980nm near infrared laser irradiation,shows that HSA-The cell survival rate of the ICG+HCT116+laser group was 32.2+2.3%,indicating a significant photothermal killing effect?P<0.05?.4.Colon cancer xenograft model in nude mice were injected with HSA-ICG,FolateRsense^TM680,ICG.The FLI after irradiation showed that the constructed HSA-ICG nanoprobe and commercial fluorescent probe had the same strong fluorescent signal in the nude mouse model of colon cancer subcutaneous transplantation tumor,and a good signal-to-noise ratio.The peak time of concentration of FolateRsense^TM680and HSA-ICG in the model of colon cancer of nude mice was 1h and 24h,respectively.The ICG as the control was especially strong in the liver,and there was no particular tumor aggregation in the tumor site.5.The two groups of mice were injected with Folate Rsense^TM680 and HSA-ICG,after 0.6W/cm² laser irradiation for 5 minutes,injection of HSA-ICG nano probe in nude mice,the tumor location can reach the highest temperature 50 degrees Celsius,the temperature will cause irreversible damage to tumor tissue.As the control group,the temperature of the tumor site of the injection of Folate Rsense^TM680 did not change,and remained at about 30?and the tumor could not damage the tumor.After injection of HSA-ICG,nude mice were subjected to FLI imaging and white light imaging after PTT.The results showed that tumors could be significantly reduced after each treatment,and the tumor treatment effect was obvious,showing obvious tumor necrosis and cavity.After FLI imaging of tumor after photothermal therapy,we can also find that the fluorescence signal gradually weakened,and the fluorescence signal disappeared in the tumor necrosis area,which is consistent with the white light and the naked eye.Pathological results confirmed that the necrosis of tumor site after injection of HSA-ICG was a typical feature of thermal injury and the imaging effect of FLI had a good correlation with its pathological findings.However,the tumor tissue injected with FolateRsense^TM680 after laser irradiation had no damage to tumor tissues and tumor tissue structure was intact.The pathological results after light also confirmed that FolateRsense^TM680 had no photothermal therapy effect on tumors.Conclusion:1.The successfully constructed HSA-ICG has an ideal size,good photothermal conversion stability,low toxic side effects and reliable biological safety.After modifying and granulation of HSA-ICG,the particle size of ICG can be reduced and the EPR effect of the tumor can be used to achieve specific aggregation of tumor and produce tumor targeting.2.The target imaging effect of HSA-ICG nano probe FLI is similar to that of commercial Folate Rsense^TM680.The ability of labeling tumor cells is strong,the intensity of fluorescence signal and signal to noise ratio are good.It has a good ability to detect colon cancer.3.The photothermal effect of the HSA-ICG nano probe after NIR irradiation can produce obvious ablation effect on colon cancer cells,significantly kill cancer cells and have good effect of photothermal treatment.4.The near infrared fluorescence diagnosis of albumin encapsulated ICG and PTT process are organically integrated.After labeling colon cancer cells by molecular imaging technology,laser irradiation based on diagnostic results is applied to the diagnosis of PTT,avoiding damage normal tissues and to improve the efficiency of diagnosis and treatment for colon cancer.It provides a new train of thought for the integrative diagnosis and treatment of colon cancer.