Exosomes-mediated CircRNA Regulates Lung Adenocarcinoma Cell PC9 Resistance To Gefitinib

Acquired resistance to gefitinib is one of the main reasons leading to the failure of targeted treatment for lung cancer,poor prognosis and high mortality.At present,the molecular markers that can predict gefitinib resistance are scarce and single.As a vesicle that transmits signals between cells,as research continues,it has been found that it plays an important role in tumorigenesis and development,invasion and metastasis,and drug resistance.Objective To find a new potential predictive drug resistance by analyzing high-throughput differential expression of exosomes derived from human lung adenocarcinoma cell line?PC9?and human lung adenocarcinoma gefitinib resistant strain?PC9R?.Molecular markers.Methods Cell supernatants of PC9 and PC9 R cell lines were collected,and exosomes were extracted,observed,and identified.The circular RNA?circ RNA?differential expression profiles in exosomes were established by high-throughput sequencing;the most significant differential expression was selected.Target circ RNA;peripheral blood of patients with gefitinib-sensitive lung adenocarcinoma and gefitinib-resistant patients were collected,and real-time fluorescent quantitative PCR?q RT-PCR?was used to verify the differential expression of target circ RNA in peripheral blood exosomes;The database and R language predict the mi RNA and target genes downstream of the circ RNA and conduct pathway enrichment analysis to further explore the molecular network of exosome circ RNA involved in gefitinib resistance.Results 1.We isolated extracellular particles in the supernatant of PC9 and PC9 R cells.The shape of the exosomes was circular under transmission electron microscopy;the particle size detector showed that the average diameter was 30-150 nm;Western blot test(Western?Blot?results showed that the exosome marker proteins CD9,CD63,and TSG101 were expressed at high levels,confirming that the exosomes were successfully isolated.2.Qualcomm Quantitative sequencing results showed that compared to PC9 R and PC9 cells,a significant portion of circ RNA expression in exosomes had significant differences?P <0.05?.Among them,35 circ RNA expression was significantly up-regulated and 341 circ RNA expression was significantly down-regulated.After further analysis and screening Circ KIF20B₀15.3.10 clinical samples that showed significantly different target RNAs were detected by q RT-PCR.Exosome circ KIF20B₀15 was significantly down-regulated in serum samples of drug-resistant patients,consistent with sequencing results.4.Bioinformatics analysis found that the parent This gene KIF20 B is related to tumor biological activities,and the predicted downstream target genes are related to basic biological activities such as genetic material transcriptional regulation,neurohormonal regulation,and Hippo signaling pathways.Conclusions 1.Circ RNA is differentially expressed in exosomes from PC9 R and PC9 cell supernatants.2.Clinical sample validation,exosomes Circ KIF20B₀15 is significantly down-regulated in the serum exosomes of drug-resistant patients,consistent with sequencing results.3.According to circ KIF20B₀15 parent genes and downstream target genes predicted,exosome circ KIF20B₀15 may regulate lung adenocarcinoma cells against gefitinib through downstream target genes.The drug may become a new potential biomarker for detecting drug resistance in lung adenocarcinoma.