The Effect Of Bortezomib Combined With Autophagy Inhibitor In The Treatment Of Adenoid Cystic Carcinoma

Objective:Adenoid cystic carcinoma(Adenoid Cystic Carcinoma,ACC)is one of the most common malignant tumors of salivary glands.At present,the main clinical treatment of ACC is still mainly combined with surgery and radiotherapy.Its biological behavior is relatively inert,and ACC is not sensitive to conventional chemotherapy.Therefore,the long-term prognosis is poor,resulting in local recurrence or high incidence of distant metastasis.Therefore,it is necessary to explore new drugs with inhibitory and killing effects on this tumor.Bortezomib is the first small molecule proteasome inhibitor approved by the United States for clinical trials.It is a highly selective proteasome inhibitor that has been approved by the FDA.It is used in the treatment of many cancers,the most representative of which are multiple myeloma and mantle cell lymphoma,which have not been reported in adenoid cystic carcinoma cells.In this experiment,by observing the effect of bortezomib on the proliferation and apoptosis of adenoid cystic carcinoma cells,the aim is to explore new molecular targeted drugs for adenoid cystic carcinoma,in order to provide a certain experimental reference for clinical treatmentMaterials and Methods:In this study,we conducted a cell viability counting kit(Cell-counting kit 8,CCK8)to determine the effect of bortezomib on the proliferation of adenoid cystic cancer cells and analyzed the changes of apoptosis by flow cytometry.We applied Western Blot(Western Blot,WB)and real-time quantitative polymerase chain reaction(Quantitative Real-time,RT-PCR)to observe the expression of Beclin-1,LC3 and P62 levels.Observe the levels of autophagosomes and autophagy by Transmission Electron Microscope(TEM)and Immunofluorescence(IF)staining.Results:1.According to the results of CCK8,the proteasome inhibitor bortezomib reduced the activity of adenoid cystic carcinoma cell ACC-83 in a concentration-dependent manner.Cell flow cytometry analysis showed that bortezomib induced the adenoid cystic carcinoma cell ACC-83.Apoptosis.2.It can be observed by transmission electron microscope that the number of autophagosomes increased in the experimental group after adding bortezomib;immunofluorescence staining showed that,compared with the blank control group,the number of fluorescent spots connected to LC3 in the experimental group increased,indicating that Bortezomib can induce autophagy.Furthermore,the protein and mRNA levels of P62,Beclin-1,and LC3 were quantitatively analyzed by WB and RT-PCR.The results showed that with the increase of bortezomib concentration,the expression of P62 and LC3-Ⅰ protein was down-regulated,and LC3-Ⅱ,Beclin-1 The protein expression level increased,and RT-PCR also confirmed this conclusion.3.After adding the autophagy inhibitor 3-MA(3-Methyladenine),by observing the results of cell viability detection and cell flow cytometry analysis,it can be seen that the proteasome inhibitor bortezomib has an effect on the proliferation of adenoid cystic carcinoma cell ACC-83.The inhibitory effect is enhanced,and the induction of apoptosis is enhanced.Conclusion:Our research shows that bortezomib inhibits the proliferation of adenoid cystic carcinoma cell ACC-83 and induces apoptosis and autophagy.In addition,the combination of autophagy inhibitor 3-MA and bortezomib can enhance the apoptosis of adenoid cystic carcinoma cell ACC-83.Targeted proteasome synergistic inhibition of the autophagy pathway may be one of the effective treatments for the treatment of adenoid cystic carcinoma.