| Background Head and neck squamous cell carcinoma is one of the most common malignant tumors in the world,with high morbidity,high mortality and poor prognosis.Due to the secrecy of the structure of the head and neck tissue,the patient is often in the advanced stage of medical treatment.At present,the treatment of head and neck cancer is still mainly surgical resection.Head and neck on the function of many organs and nervous,basically complete breathing,swallowing,the physiological function such as sound,surgical resection with physiological function damage,seriously reduce the patient's quality of life,and a variety of complications including pharyngeal fistula,postoperative but also nots allow to ignore.Induced chemotherapy for head and neck cancer treatment in recent years,more and more research of the new project,can be carried out after induction chemotherapy in patients with subsequent treatment options,such as sensitive to chemotherapy patients can choose the way of chemotherapy treatment,to avoid the surgical removal of the postoperative complications,this treatment can be a great deal of retention in patients with organ function,greatly improve the survival quality of patients with head and neck cancer.But not the target of chemotherapy drugs makes it also has a role to normal tissue,at the same time of treatment is often accompanied by strong side effects,such as application in the treatment of head and neck cancer of platinum drugs,the gastrointestinal side effects and side effects such as cardiac toxicity,limits its application in clinical treatment of head and neck cancer.Therefore,according to the situation of the treatment of head and neck cancer,to develop good targeting drug carrier for targeted treatment of head and neck cancer,in order to improve the los platinum chemotherapy drugs in patients with head and neck cancer treatment efficiency and reduce the side effects of chemotherapy drugs.Exosomes,a saucer shape of phospholipid bilayer vesicle,diameter size of about 30-150 nm,carried RNA,proteins,such as signal substance,take important roles in the intercellular communication,in recent years,exosomes as organism,signal transmission and the angel of targeting drug carrier research emerge in endlessly.A number of research results show that the biological cells in the body such as stem cells,dendritic cells,exosomes can stable exist in organism body fluids such as blood,breast milk,saliva,secrete body due to the nature of its own makes it have excellent biological stability,more and more researchers choose modified exosomes makes its can pack many substances such as RNA,which have effect of treatment of protein,chemotherapy drugs can be used for the treatment of many diseases.In this study,we selected exosomes as a targeted transport vehicle,and used near-infrared fluorescence molecular imaging technology to diagnose and treat head and neck cancer tumors.Using independent TRP-PK1 peptides can insert the characteristics of the lipid bilayer,combines iRGD in TRP-PK1,form the TRP-PK1-iRGD polypeptide,exosomes outside to decorate,make its have targeted high expression of ?v?3 integrin tumor tissue.Platinum drugs package load to target by means of total hatch outside the body of the secretion,and the modified version of through intravenous drug delivery,accurate targeting tumor location,release chemotherapy drugs to kill cancer cells,achieve maximum extent reduce chemotherapy drugs on normal tissue damage,reduce the side effects,improve the treatment efficiency.In this study,through the modified natural exosomes outside loading tumor,chemotherapy drugs to targeted therapy in improving the efficiency of chemotherapy drugs at the same time,reduce the side effects of chemotherapy drugs,provide a new method for head and neck cancer treatment.Objective 1.Through TRP-PK1-iRGD polypeptide modified exosomes to enable it to target the head and neck cancer tissue,and through molecular imaging technology to detect and transform the exosomes and coupling near-infrared dye in the body distribution and metabolism characteristics.2.After modification exosomes bag outside los platinum chemotherapy drug targeting role in tumor tissue,improve the lobaplatin chemotherapy drugs in head and neck squamous cell carcinoma of the treatment effect,reduce the side effects on lobaplatin chemotherapy drugs.Method 1.Experimental materials Exosomes,iRGD peptide,TRP-PK1 peptide,IRDYE800CW-NHS-easter,injection of LBP.2.Biomaterials Head and neck cancer cell line HN4-luc cells,human embryo kidney HEK293 T cell line,cell culture medium,Balb/c nude mouse,biological matrix glue,luciferase substrate enzyme.3.Extraction of exosomes.To extract the exosomes,exosomes are used in the experiment are from one embryo kidney cell culture supernatant on HEK293 T cell line of human embryonic kidney,Human embryonic kidney HEK293 T cells were cultured with DMEM containing 10% FBS.Free FBS DMEM was used for the extraction of exocrine for 48 h,the cells culture supernatant was collected after treatment,and the exosomes were extracted by means of ultracentrifugation.4.Experimental equipment Ultracentrifuge,malven potentiometer,inverted fluorescence microscope and confocal microscopy,transmission electron microscopy(TEM),Inverted fluorescence microscope,enzyme standard instrument,small animal anesthesia machine,IVIS small animal in vivo imaging system. 5.Cell culture and tumor planting Head and neck cancer HN4-luc cells cells,human embryonic kidney HEK293 T cells were cultured under standard conditions.Balb/c rats are purchased from the animal center of Beijing.Male Balb/c nude mice were used for the establishment of subcutaneous tumor model of nude mice,1:1 mixed biostromal gel and head and neck cancer HN4-luc cells cell suspension.To establish a model of nude mouse skin head and neck cancer transplantation tumor,a total of 2 x 106 head and neck cancer HN4-luc cells cell suspension was inoculated in the axillary region of each nude mouse.6.The biological distribution of exosomes after transformation The tissue distribution of exosomes in nude mice was determined by the fluorescent distribution of IRDYE800CW-NHS-easter by IVIS,a small animal fluorescence imaging device.After the modification,the exosomes were injected into the mice.Through the real-time detection of IVIS,the biological distribution and metabolic pathway of the modified exosomes in the body were observed,and the time point and record of the enrichment of exosomes in the tumor site were recorded.7.The therapeutic effect of the exosomes after the reconstruction in vivo After intravenous injection of different groups,the external secretion was performed,and the weight of the nude mice was weighed every two days,and the tumor size and survival time were measured to evaluate the therapeutic effect.Result Cell and animal level results show that the TRP-PK1 peptides and iRGD peptides can be anchored exosomes and gives out the role of integrin targeted tumor cells,can guarantee that exosomes lobaplatin drugs targeted tumor therapy effect into full play.In addition,EXO(TRP-PK1-iRGD)+LBP was effective in inhibiting the tumor growth rate compared with the free LBP group,according to the results of intravenous modified treatment group EXO(TRP-PK1-iRGD)+ LBP compared with intravenous free lobaplatin drugs group,after modification exosomes of lobaplatin drug treatment group(TRP-PK1-iRGD)+ LBP weight of nude mice in treatment process is relatively stable,not present weight fell sharply.The experimental results show that the modified exosomes can improve the therapeutic effect of the chemotherapy drugs,and significantly reduce the toxic and side effects of the chemotherapy drugs.Conclusion After transforming the exosomes carrying lobaplatin chemotherapy drugs can be targeted to kill cancer cells effectively,not only can achieve good treatment effect,and in treatment at the same time also can reduce the lobaplatin chemotherapy drug damage to normal tissue.Therefore,we hypothesized that,in the follow-up study,it could improve the efficiency of the drug delivery and improve the therapeutic effect significantly.After transforming exosomes the target determines the side effects of chemotherapy drugs,therefore,we suspect that if can improve the secrete body outside the target,will most likely reduce exosomes the various side effects of chemotherapy drugs. |
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