Study Of LncRNA AC012073.1 And AC073352.1 In The Metastasis Of Breast Cancer And Their Clinical Value

Objective:Breast cancer(BC)is the most common malignancy in women worldwide and has become a serious threat to the health of women.Patients with BC metastasis have poor prognosis and the mechanisms of BC metastasis are complex.Long non-coding RNAs(lncRNAs),are known as a class of noncoding transcripts RNA with a length of more than 200 nucleotides,which have been shown to have crucial roles in BC progression.However,the underlying mechanisms by which IncRNA-drive BC metastasis remains unclear.In this study,we aim to explore the biological functions and molecular mechanisms of novel lncRNAs in BC,and to assess their diagnostic in clinical samples.Methods:1.The lncRNA microarray and TCGA database analysis were used to screen the candidate IncRNAs that were highly expressed in BC tissues and associated with poor prognosis,and their expression levels were verified using the online database or Tissue microarray and in situ hybridization(ISH).2.BC cells were transfected with AC012073.1 and AC073352.1 siRNAs or plasmid and further investigated by gain and loss of functions assays.3.The downstream genes of AC012073.1 were predicted by TargetScan and miRDB database,its ceRNA regulatory network was mapped by Cytoscape software.The regulatory mechanism of AC073352.1 was analyzed by RNA pulldown,Western blot,RNA immunoprecipitation(RIP)and rescue experiments.4.Exosome identification was detected by transmission electron microscopy(TAE)and nanoparticle tracking analysis(NTA).Co-culture and exosome labeling experiments were used to assess exosomal AC073352.1 transferred by BC cells into endothelial cells.The effect of exosomal AC073352.1 in angiogenesis was further investigated by tube formation.5.The expression of AC012073.1 and AC073352.1 was detected in BC serum by quantitative real-time PCR(qRT-PCR),and receiver operating curve(ROC)was using to evaluate their diagnostic values.Results:1.Two novel lncRNAs AC012073.1 and AC073352.1 were screened and significantly upregulated in BC tissue and were associated with poor prognosis.The clinicopathological analysis showed that a high expression level of AC073352.1 was associated with TNM stage and lymph node metastasis of BC.2.In vitro function experiments showed that knockdown of AC012073.1 or AC073352.1 significantly inhibited BC cell migration and invasion,while AC012073.1 or AC073352.1 overexpression could enhance the above ability noticeably.In addition,vivo functional experiments showed that AC073352.1 knockdown could inhibit lung metastasis of BC.3.Mechanistically,AC012073.1 may act as an oncogene by affecting ERK1/ERK2,Wnt and other tumor classical signaling pathways.We also elucidated that AC073352.1 interacts with YBX1 and stabilized its protein expression.Knockdown of YBX1 reduced cell migration and invasion,and could partially reverse BC metastasis driven by AC073352.1.4.Moreover,the results of co-culture and tube formation indicated that AC073352.1 might be packaged into exosomes by binding to YBX1 in BC cells resulting in angiogenesis.5.qRT-PCR and ROC analysis suggested that AC012073.1 and AC073352.1 were relatively high expressed in BC serum and were expected to be diagnostic markers of BC.Conclusions:1.AC012073.1 and AC073352.1 are highly expressed in BC and are associated with poor prognosis.2.AC012073.1 promotes the migration and invasion of breast cancer and may be involved in the regulation of tumor-related signaling pathways.3.AC073352.1 can interact with YBX1 to promote metastasis and angiogenesis of BC.4.Serum AC012073.1 and AC073352.1 may be biomarkers for the diagnosis of BC.