The Study Of LncRNA Involved In Liver Metastasis Of Colorectal Cancer

Objective:1.Aim to study differentially expressed lnc RNAs during colorectal cancer liver metastasis,we performed Lnc RNA-seq in LM2 HCT116 via second-generation sequencing.2.We investigate the five lnc RNAs(LOC100507487,BISPR,PHKA2-AS1,LINC02525 and HM13-AS1)that are up-regulated in the liver metastasis,and explore their role inregulating colorectal cancer.3.We study the effect of lnc RNA LOC100507487 on the migration and proliferation of colorectal cancer cells,and further explore its interaction proteins.Methods:1.The colorectal cancer cell HCT116 primary cell(M0)was subjected to spleen injection in nude mice and resected.After liver metastasis occurred in the tumor,liver metastatic tumor cells were extracted and cultured to obtain LM1(liver metastasis)cells.Then LM1 cells were injected into spleen of nude mice.After resection,liver metastasiswas expected to occur.and liver metastatic tumor cells are extracted and cultured to obtain LM2 cells.Existing data show that liver metastatic tumor cells have more proliferation and metastasis than primary tumor cells.LM2 cells and MO cells were sequenced to compare the lnc RNA expression profiles.2.We analyzed differential lnc RNA expression profiles and screened lnc RNAs with significantly differential expression and high expression levels via QPCR in HCT116 cells.The expression of these lnc RNAs in colorectal cancer cells HCT116 was knocked down by si RNA.then we performed Transwell experiments to knock down detect the effect of these lnc RNAs on cell migration.3.We examined the cell migration ability through Transwell experiments with stable transfected cell lines,which were knocked down or overexpressed of LOC100507487.And cell proliferation was detected via MTS experiments to explore the effect of knockdown or overexpression LOC100507487.Further,we performed RNA pulldown experiments and RIP experiments to explore related proteins that may bind to LOC100507487.Results:1.A series of lnc RNAs related to liver metastasis were screened by performing lnc RNA transcriptome analysis on two liver metastatic colorectal cancer cells HCT116.At the same time,we performed GO functional enrichment analysis and KEGG pathway analysis on differentially expressed m RNA and predicted lnc RNA target genes.2.Transwell experiments verified that knockdown of lnc RNA(LOC100507487,BISPR,HM13-AS1)played an inhibitory role in cell migration of colorectal cancer,which was consistent with the results of sequencing expression profiles.3.Transwell experiments have verified that knocking down LOC100507487 could inhibit the migration of colorectal cancer cells,and overexpression of LOC100507487 could promote the migration of colorectal cancer.MTS experiment results showed that downregulated LOC100507487 can inhibit the proliferation of colorectal cancer cells,overexpression of LOC100507487 can promote the proliferation of colorectal cancer.The results of RNA pulldown and RIP showed that LOC100507487 binding protein was analyzed by mass spectrometry,and it was found that LOC100507487 may have the potential of binding to metabolism-related proteins APMAP and SLC2A1.Conclusion:1.We have screened out multiple lnc RNAs that are differentially expressed during colorectal cancer liver metastasis.2.Lnc RNA(LOC100507487,BISPR,HM13-AS1)is up-regulated during colorectal cancer liver metastasis,and knocking down these lnc RNAs can inhibit colorectal cancer cell migration.3.Knockdown of lnc RNA LOC100507487 can inhibit the proliferation and migration of colorectal cancer cells,and overexpression of LOC100507487 can promote the proliferation and migration of colorectal cancer cells.LOC100507487 may bind with the metabolism-related proteins SLC2A1.