The Role And Mechanism Of NNMT In Nasopharyngeal Carcinoma

NPC is a malignant tumor of head and neck originating from nasopharyngeal epithelium.With the development of radiotherapy and chemotherapy technology,the clinical treatment effect of NPC is improving continuously.However,some patients still have recurrence and distant metastasis after radiotherapy and chemotherapy.Therefore,a comprehensive understanding of the molecular mechanisms underlying NPC progression and metastasis may help to develop more effective targeted treatment strategies.NNMT is a methyltransferase,which is highly expressed in a variety of tumors and it is closely related to cell metabolism and epigenetic regulation of tumors.This study focuses on the function and mechanism of NNMT in NPC.Purpose: This study aims to explore the function and molecular mechanism of NNMT in NPC progression.Methods:1.The relationship between the expression of NNMT and the prognosis of HNSCC patients was analyzed by using TCGA database,and the expression of NNMT in normal nasopharyngeal epithelial tissues and NPC tissues was detected by immunohistochemical assay.2.NNMT was knocked down or overexpressed in NPC cells,and the effects of knockdown or overexpression of NNMT on proliferation,cell cycle and invasion ability of NPC cells were detected by CCK-8,clony formation,flow cytometry and Transwell assay,respectively.3.The subcutaneous tumor model and lung metastasis model in nude mice were established to observe the effects of NNMT on tumor growth and metastasis in vivo.4.Gene Card and JASPAR websites were used to predict the upstream transcription factor and its binding site of NNMT.q PCR and Western Blot were used to detect the expression changes of NNMT m RNA and protein levels after knockdown or overexpression of NNMT upstream transcription factor.The binding of transcription factor to the NNMT promoter region was detected by Ch IP assay.5.Stable cell lines 5-8F-control and 5-8F-sh NNMT were established in NPC cell line 5-8F,and RNA sequencing analysis was performed to screen downstream target genes of NNMT.The biological function of downstream target gene was detected.Western Blot method and Ch IP assay were used to explore the mechanism of NNMT regulating downstream target gene expression.The rescue assay was used to analyze whether the downstream target gene could recover the effects of NNMT expression changes on the phenotype of NPC cells.Results:1.The results of bioinformatics analysis with TCGA datasets showed that high expression of NNMT shortened PFS and OS of HNSCC patients.Immunohistochemical results showed that NNMT was lowly expressed in normal nasopharyngeal epithelial tissues and highly expressed in NPC tissues.2.Knockdown or overexpression of NNMT could respectively inhibit or promote the proliferation and invasion of NPC cells,and block or promote the process of NPC cell cycle.3.In vivo experiments showed that NNMT could promote the tumorigenicity and metastasis of NPC cells in nude mice.4.The expression of NNMT was positively regulated by STAT3 at the transcriptional level.5.We found that NNMT positively regulated the expression of BCAT1 through affecting the level of H3K27me3 in the promoter region of BCAT1,thereby promoting the malignant progression of NPC.Conclusions:1.NNMT is highly expressed in NPC tissues,and its high expression is associated with poor prognosis of HNSCC patients.2.NNMT can promote NPC cell proliferation,invasion,cell cycle progression,tumor growth and metastasis in vitro and in vivo.3.Under the positive regulation of upstream transcription factor STAT3,NNMT plays a significant role in promoting NPC progression by positively regulating the expression of BCAT1 through affecting the level of H3K27me3 in the promoter region of BCAT1.