| Obj ective(s):The lung cancer incidence and mortality in the Xuanwei and neighboring region,Yunnan,China,is among the highest in China,which has the unique characteristics including a high incidence in women,the age of onset to incline to the young and familial aggregation.However,little is known about its pathogenesis.The purpose of this study was to screen and identify potential novel "drivers" in Xuanwei lung cancer(XWLC),and to explore its molecμlar mechanism.Methods:1.The Genome-Wide DNA copy number changes(CNVs)were detected by Human CGH Array chip in 24 XWLC and matched para-carcinoma tissues,and the copy number variation hotspots were screened out.2.The differentially expressed genes(DEGs)were identified by expression microarray in 24 XWLC and matched para-carcinoma tissues and genes related to pathological grade were screened out.3.Thecandidate "driver genes" involved in the progression of XWLC were identified by integrated analysis of Genome-Wide copy number alterations and gene expression profiling;4.The gene and protein expression levels of candidate genes were confirmed in XWLC by real-time quantitative polymerase chain reaction,Western blot and immunohistochemistry,and analyzed the relationship with clinicopathological features;5.The effects of candidate "driver genes" on the basic biological behavior of XWLC cell lines were carried out in vitro;6.The candidate "driver gene" interaction protein network was constructed and the pathway pathway analysis was performed to screen out the possible signal transduction pathways.Western blotting was used to identify the influence of candidate "driver genes" on candidate pathway-related protein expression.Resμlts:1.A large number of CNVs were detected in XWLC,Overall,the CNVs featured more gains than losses.The higher frequency of CNVs amplification is mainly located on chromosomes 7,5,14,18,17,20,10,11,12,and 16 and the deletion is mainly located on chromosomes 8,10,and 5.Recurrent CNVs was found in 5p11-pl5.33,7p11.2-p22.3 and 1q21.1-q44;2.XWLC harboured abundant DEGs,of which 9444 were up-regμlated and 3369 were down-regμlated.GO analysis found that most of them involved in chromosome assembly,DNA replication,mitosis,and protein heterodimer formation;the path pathway enrichment analysis suggested that it mainly involved cell signaling,cell cycle regplation,and protein amino acid phosphorylation.Associated with pathological stage,1018,2047and 1391 DEGs were found in "stage Ⅰ,Ⅱ Ⅲ,respectively.Notably,461 dysregμlated genes including 461 up-regμlated were closely linked to stage Ⅰ-Ⅲ;3.The up-regμlated genes associated with pathological staging maped to chromosomes 1,5 and 7,and 40 candidate genes were obtained;The 40 candidate genes were located three recurrent CNVs regions,and finally 12 genes were output.TRIP 13 was a potential "driver gene" due to the highest expression level and the highest sample homogeneity;4.TRIP 13 was up-regμlated and increased protein levels in tissues;and high expression of TRIP13 in cancer tissues was positively correlated with pathological grade and clinical stage(p>0.05);5.In vitro experiments confirmed that stable over-expressed TRIP 13 can inhibit the proliferation,migration and invasion of XWLC cells as well as promoting G2/M phase transition and inhibiting cell apoptosis;and knockdown of TRIP 13 will have a completely opposite trend;6.Overexpression of TRIP 13 attenuated the expression of the downstream effector molecμle of p-catenin and inhibited the Wnt/β-catenin pathway.However,TRIP 13 knockdown showed that c-jun only was up-regμlated.Conclusion(s):TRIP 13,a potential new driver gene in xuanwei lung cancer,china,was identified by integrating CNVs and DEGs arrays,cell function experiment and molecular assays;The role of TRIP13 was preliminarily discussed,and to lay the foundation for the molecular diagnosis and effective treatment of Xuanwei lung cancer. |
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