Lysine-specific demethylase 1(LSD1)plays a regulatory role in tumor immunity.LSD1 affects CD8+T cell infiltration by regulating AGO2 protein level,and it is required for normal B cell proliferation and differentiation via reducing local chromatin accessibility.LSD1 inhibition promotes NK cell cytotoxicity of cancer cells through restoring the expression of ULBPs.LSD1 affects the resistance of M1 macrophages to oxidative stress by regulating the level of SOD2;In addition,regulating the expression of LSD1 affects the anti-tumor function of CD 19 CAR-T cells and immune checkpoint blockers.The development of drugs targeting LSD1 will provide new ideas for immunotherapy of tumors.

Objective To investigate the biological function and potential mechanisms of long non-coding RNA(lncRNA)neurensin 2-antisense RNA 1(NRSN2-AS1)in liver cancer cells.Methods The Encyclopedia of RNA Interactomes(ENCORI)database was used to analyze the expression levels of NRSN2-AS1 in liver cancer tissues and normal tissues as well as its association with the prognosis of patients.Stable lncRNA NRSN2-AS1 cell lines,overexpressed or knockdown,were constructed.The effects of NRSN2-AS1 on tumor cell proliferation were explored using CCK-8 and colony formation assays.Transwell and wound healing assays were employed to examine the role of NRSN2-AS1 in tumor cell migration and invasion.The impact of NRSN2-AS1 on tumor cell aerobic glycolysis was assessed by measuring hexokinase activity,glucose uptake,ATP and etracellular lactate levels.Quantitative real-time PCR(qPCR)and Western blotting were used to evaluate the effects of NRSN2-AS1 on the mRNA and protein expression levels of hexokinase 2(HK2)in tumor cells.Results Analysis from the ENCORI database revealed that NRSN2-AS1 was upregulated in liver cancer tissues compared to normal tissues,and that high expressions of NRSN2-AS1 were closely associated with poor prognosis of patients.In vitro functional assays demonstrated that overexpression of NRSN2-AS1 promoted proliferation,migration,and invasion of liver cancer cells,and enhanced glycolysis levels while knockdown of NRSN2-AS1 inhibited these processes and suppressed glycolysis.Furthermore,overexpression of NRSN2-AS1 increased the mRNA and protein levels of HK2 while knockdown of NRSN2-AS1 decreased HK2 expression in liver cancer cells.Conclusion NRSN2-AS1 is highly expressed in liver cancer tissues,and it may promote liver cancer progression by enhancing HK2 expression and aerobic glycolysis.

Objective: To analyze the expression and clinical prognostic significance of nuclear Dbf2-related kinase 1 ( NDR1 ) in hepatocellular carcinoma ，and to investigate the biological function and regulatory mechanism of NDR1 in hepatocellular carcinoma cells． Methods: ENCORI database was used to analyze the correlation of NDR1 and c-Myc．Cycloheximide ( CHX) experiment analyzed the relationship between NDR1 and c-Myc protein stability．The expression levels of NDR1 in liver cancer tissues and normal tissues and its relationship with the survival rate of liver cancer patients were analyzed using the ENCORI database．MYC-NDR1 eukaryotic expression vector was constructed，transfected with hepatocellular carcinoma HepG2 cells，and its expression was verified by protein immuno blotting (Western blot) ; cell proliferation and migration ability were detected by CCK-8 assay，cell clone formation assay and scratch assay，respectively．The correlation between NDR1 and c-Myc expression was analyzed using the ENCORI database，and the relationship between NDR1 and c-Myc protein was investigated using a protein synthesis inhibitor CHX dosing assay. Results: The results of the ENCORI database showed that the expression of NDR1 in liver cancer tissues was higher than that in normal tissues and the overall survival rate of patients with high NDR1 expression was lower than that of patients with low NDR1 expression，and the difference was statistically significant (P＜0. 001) ．The results of the CCK-8 assay showed that the MYC-NDR1 group grew faster than the empty vector group (P＜0. 001) ．The clone formation assay showed that the number of clones in the MYC-NDR1 group was higher than that in the empty vector group (P＜0. 001) ．The cell scratch assay showed that the mean migration distance in the MYC-NDR1 group was greater than that in the empty vector group (P＜0. 001) ．ENCORI database analysis showed that NDR1 correlated with c-Myc expression (R = 0. 184，P＜0. 001) ; CHX dosing assay showed that the reduction of c-Myc protein in the MYC-NDR1 group was lower than that in the empty vector group during the same time． Conclusion NDR1 is highly expressed in hepatocellular carcinoma tissues，closely correlated with poor prognosis of hepatocellular carcinoma patients ，and positively correlated with the expression of c-Myc gene. The study successfully constructes MYC-NDR1 eukaryotic expression vector ，and the expression product MYC- NDR1 can increase the stability of c-Myc protein and promote the proliferation and migration of human hepatocellu- lar carcinoma cells.

Breast cancer is one of the most common malignancies that seriously threaten women's health. In the process of the malignant transformation of breast cancer, metabolic reprogramming and immune evasion represent the two main fascinating characteristics of cancer and facilitate cancer cell proliferation. Breast cancer cells generate energy through increased glucose metabolism. Lipid metabolism contributes to biological signal pathways and forms cell membranes except energy generation. Amino acids act as basic protein units and metabolic regulators in supporting cell growth. For tumor-associated immunity, poor immunogenicity and heightened immunosuppression cause breast cancer cells to evade the host's immune system. For the past few years, the complex mechanisms of metabolic reprogramming and immune evasion are deeply investigated, and the genes involved in these processes are used as clinical therapeutic targets for breast cancer. Here, we review the recent findings related to abnormal metabolism and immune characteristics, regulatory mechanisms, their links, and relevant therapeutic strategies.
Breast Neoplasms ; Cell Proliferation ; Cell Transformation, Neoplastic ; Energy Metabolism ; Female ; Humans ; Lipid Metabolism ; Signal Transduction

Objective To construct the eukaryotic expression vector of PRDX3 labeled with FLAG tag and to study its localization in human tongue cancer cell line SCC15.Methods PRDX3 gene was obtained from the breast library by PCR and cloned into PCDH vector to construct PCDH-FLAG-PRDX3.The plasmid was transiently transfected into 293T cells and the expression was detected by Western blot.Subcellular localization was detected by cellular immunofluorescence.Results The result of double digestion and sequencing showed that PCDH-FLAG-PRDX3 eukaryotic expression vector was constructed.The expression of FLAG-PRDX3 in human 293T cells was positively confirmed by Western blotting.In human tongue cancer cell line SCC15, the result of cellular immunofluorescence showed FLAG-PRDX3 was located in the cytoplasm rather than in the nucleus.Conclusion PRDX3 eukaryotic expression vector labeled with FLAG tag is constructed successfully, which is located in cytoplasm in human SCC15 cells.Construction and identification of PRDX3 could shed light on the function and mechanism of PRDX3 in tongue cancer.

Objective To purify and prokaryotically express the histone methyltransferase SET7.Methods The coding sequence of full length SET7 was amplified from breast library by PCR and cloned into the pGEX-KG vector.The correct recombinant plasmid was introduced into E.coli.The expressed protein was identified by SDS-PAGE.Results DNA sequencing indicated the SET7 was constructed.GST-SET7 fusion protein was identified by SDS-PAGE.Conclusion The prokaryotically expressed protein of GST-SET7 is obtained,which will falilitate further study of SET7 function.

Objective To detect the effect of E6AP on gastric cancer cell proliferation and migration.Methods The expression of E6AP in different gastric cancer cell lines and normal gastric mucosa epithelial cell lines was detected by Western blotting.Gastric cancer cells BGC-823 stably expressing E6AP short hairpan RNA(shRNA) were obtained by lentiviral vector of E6AP.The effect of E6AP on BGC-823 cell growth and migration was determined by CCK-8 kit, Tran-swell and wound healing assay.Results Gastric cancer cell line BGC-823 in which E6AP was stably knocked down was established.Knockdown of E6AP inhibited the proliferation and migration of BGC-823 cells.Conclusion E6AP plays a key role in gastric cancer proliferation and migration.

Objective To construct the eukaryotic expression vector of human telomerase RNA component ( hTR) and study its biological function tentatively .Methods hTR Gene was obtained by PCR from cDNA template , which was reverse transcribed from 293T mRNA and cloned into pCDNA3.0 vector.The recombinant plasmid and empty vector were trans-fected into 293T cells, and hTR expression was identified by qRT-PCR.HepG2 cells that stably transfected with pCDNA3.0-hTR were constructed and identified by qRT-PCR.These cells were used to assess the interaction of hTR with human telomerase revese transcriptase ( hTERT ) and dyskerin .Telomerase activity was also detected in HepG 2 cells transfected with pCDNA3.0-hTR.Results pCDNA3.0-hTR eukaryotic expression vector was successfully constructed by double digestion identification .The inserted fragment was confirmed by sequencing .The expression of hTR in human 293T cells and HepG2 pCDNA3.0-hTR stable cell line was identified.In addition, qRT-PCR and Western blotting results showed that hTR could interact with hTERT and dyskerin , while hTR overexpression could not regulate the telomerase activity in HepG2 cells.Conclusion The eukaryotic expression vector of pCDNA 3.0-hTR is successfully constructed and expressed.This study will contribute to the further study of cancer therapy targeting hTR .

Objective To construct the lentiviral vector of RNA interference(RNAi) for DEK,and to detect its effect on breast cancer cell growth.Methods The DEK siRNA was designed and constructed based on DEK sequence using a lentiviral vector.The lentivirul vector containing DEK siRNA was named PSIH-H1-DEK as confirmed by PCR and sequenceing.PSIH-H1-DEK was then packaged with accessory plasmids into lentivirus in 293T cells and selected for 2 weeks with puromycin ( puro ) before the mixed colonies stably expressing DEK siRNA were obtained and the DEK expression was detected by real time PCR( RT-PCR) and Western blotting.The effect of DEK siRNA on ZR75-1 cell growth was determined by cell counting kit.Results Western blot and RT-PCR showed that PSIH-H1-DEK siRNA could suppress DEK gene expression.Suppression of DEK could markedly inhibit the growth of ZR75-1 cells.Conclusion The lentivirus-mediated DEK siRNA is obtained,which will facilitate further research on DEK function in breast cancer development.

Objective To construct PES1 shRNA stable expression cell lines in tongue squamous cell carcinoma ( TSCC) cells and to study the effect of knockdown of PES 1 on the growth of TSCC cells .Methods Recombinant lentivirus carrying PES1 shRNA was packaged and obtained in 293T cells.TSCC cells (Tca8113, SCC6 and SCC15) were infected with the lentivirus and selected for stable cells .PES1 expression was identified by Western blot .The effect of inhibition of PES1 on the growth and cell cycle of TSCC cells was detected by growth curve and flow cytometry .Results TSCC cells stably expressing PES1 shRNA were constructed.Knockdown of PES1 inhibited cell proliferation and induced cell cycle ar-rest at G0/G1 phase.Knockdown of PES1 inhibited expression of cyclin D1 in TSCC cells.Conclusion Inhibition of PES1 results in reduced cell proliferation , cell cycle arrest at G 0/G1 phase and reduction of cyclin D 1 expression in TSCC cells . PES1 may be a target for TSCC gene therapy .