Evaluation Of The Correlation And Molecular Mechanism Between ATAD2 Expression Of Lung Adenocarcinoma And Glycometabolism In ¹⁸F-FDG PET Imaging

Objectives:Lung cancer is caused by a common malignant tumor whose incidence and mortality has increased year by year.It is also a major malignancy which affects the health of people living in China.Lung adenocarcinoma is the most common pathological type of lung cancer and is typically treated by surgery,chemotherapy and biologically targeted therapy.Lung adenocarcinoma progresses rapidly and often leads to metastasis.The challenge for treating lung adenocarcinoma is its resistance to chemotherapy,while the tumor molecular marker,which can reflect the biological behavior of the tumor,may have a good value for treatment.Glycometabolism works through two main ways:oxidative phosphorylation and glycolysis.The main way to produce ATP in normal cells is through glucose oxidative phosphorylation,in which glycolysis is inhibited,while in malignant tumor cells,even if there is sufficient oxygen,glycolysis is active.Both glucose uptake and lactate metabolism will increase significantly.This is known as the Warburg effect and is characteristic of most malignant tumors.ATPase family AAA domain-containing protein 2?ATAD2?,which is a member of the AAA+ATPase family of proteins,contains both an ATPase domain and a bromodomain.Research has shown that ATAD2 is highly expressed in several types of tumors.This study aimed to investigate the correlation between ATAD2 and lung adenocarcinoma glycometabolism as well as the mechanism of its effects on glycometabolism.Methods:1.Patients who undergone ¹⁸F-Fluorodeoxyglucose(¹⁸F-FDG)positron emission tomography/computed tomography?PET/CT?imaging before surgery and who had a pathological diagnosis of lung adenocarcinoma were included.The participant groups contained 66 cases of lung adenocarcinoma tissues and 52 cases of para-carcinoma lung tissues.The expression of ATAD2 in lung adenocarcinoma and para-carcinoma lung tissues,as well as glucose transporter 1?GLUT1?and hexokinase 2?HK2?in lung adenocarcinoma tissues were detected using immunohistochemical staining.In order to investigate the correlation between the ATAD2 expression of lung adenocarcinoma and the glucose metabolism,the correlation between the ATAD2 expression and the semi-quantitative indexes of¹⁸F-FDG PET/CT imaging,including maximum standard uptake value(SUV_max),total lesion glycolysis?TLG?,metabolic tumor volume?MTV?,GLUT1 expression and HK2 expression were analyzed.The relationship between ATAD2 expression and the clinical characteristics of lung adenocarcinoma patients was also analyzed.2.The human lung adenocarcinoma cell line A549 was selected,while the ATAD2expression was silenced using siRNA.The human lung adenocarcinoma cell line H1299 was selected,and plasmid used for overexpression of ATAD2.LY294002 was used to inhibit PI3K/AKT pathway.Quantitative real-time PCR?qRT-PCR?analysis was then utilized to detect the mRNA expression of GLUT1 and HK2 after ATAD2was changed,while Western blot analysis was utilized to locate the protein expression of the phosphorylated protein kinase B?pAKT?,GLUT1 and HK2 after ATAD2 had changed.The proliferation and migration capacity of human lung adenocarcinoma cells was measured after ATAD2 changed.¹⁸F-FDG uptake was detected utilizing gamma counter.Both ATP production and lactic acid in the culture medium were detected after ATAD2 changed.3.A BALB/c-nude mouse model with human lung adenocarcinoma was established,while the expression of ATAD2 was silenced by shRNA intratumoral injection.¹⁸F-FDG micro PET imaging was performed.qRT-PCR analysis was used to locate the mRNA expression levels of GLUT1 and HK2,while Western blot and immunohistochemical staining analyses were utilized to detect the protein expression levels of GLUT1,HK2 and pAKT following the down-regulated expression of ATAD2.Results:1.According to the results of immunohistochemical staining,the expression of ATAD2 in 66 cases of lung adenocarcinoma tissues was significantly higher than that in the 52 cases of adjacent normal lung tissues?P<0.01?.The expression level of ATAD2 in lung adenocarcinoma showed a significant positive correlation with the semi-quantitative indexes of ¹⁸F-FDG PET/CT imaging,including SUV_max and TLG?rho=0.849,0.681,P<0.01?,while there was no significant correlation found between MTV and the expression level of ATAD2 in lung adenocarcinoma?rho=0.168,P>0.05?.The expression level of ATAD2 in lung adenocarcinoma demonstrated a significant positive correlation with the glucose metabolism indexes,such as GLUT1and HK2?rho=0.851,0.903,P<0.01?.Overexpression of ATAD2 protein was found to be closely related to lymph node metastasis and poor differentiation in lung adenocarcinoma?P<0.01?.However,it had had no significant relationship with age,gender or tumor size?P>0.05?.2.The transfection efficiency of siRNA-ATAD2 and plasmid-ATAD2 in the human lung adenocarcinoma cell lines?A549 and H1299?was detected using qRT-PCR method.The results of qRT-PCR and Western blot analyses showed that both mRNA and protein expression levels of GLUT1 and HK2 were significantly down-regulated following ATAD2 interference?P<0.01?,while,the mRNA and protein expression levels of GLUT1 and HK2 were increased following ATAD2 up-regulation?P<0.05?.Down-regulation of ATAD2 expression inhibited proliferation,migration,¹⁸F-FDG uptake and lactate production of A549 cells?P<0.05?,while,overexpression of ATAD2 expression promoted proliferation,migration,¹⁸F-FDG uptake and lactate production of H1299 cells?P<0.05?.The expression level of pAKT was significantly reduced in siATAD2-transfected A549 cells,while,pAKT expression was increased after ATAD2 overexpression in H1299 cells?P<0.01?.It was found that LY294002?20?M?treatment significantly reduced the expression levels of GLUT1,HK2 and pAKT,proliferation,migration,¹⁸F-FDG uptake and lactate production?P<0.01?.However,the expression level of ATAD2 was not being affected by LY294002?P>0.05?.Production of ATP did not alter markedly even though ATAD2 had been changed?P>0.05?.3.The SUV_max of ¹⁸F-FDG micro PET imaging,mRNA expression levels of GLUT1and HK2 and protein expression levels of GLUT1,HK2 and pAKT were downregulated following ATAD2 interference in nude mice with human lung adenocarcinoma.This difference was found to be statistically significant?P<0.01?.Conclusions:The expression of ATAD2 in lung adenocarcinoma tissues was significantly higher than that in adjacent normal lung tissues.The overexpression of ATAD2 protein was closely related to lymph node metastasis as well as poor differentiation in lung adenocarcinoma.The expression level of ATAD2 in lung adenocarcinoma showed a significant positive correlation with the SUV_max and TLG as well as the expression of GLUT1 and HK2.ATAD2 affected the glucose metabolism of lung adenocarcinoma through the AKT-GLUT1/HK2 pathway.The changes to ATAD2 expression also affected the uptake of glucose as well as lactic acid production of lung adenocarcinoma cells.The expression of ATAD2 was related to the proliferation and migration of lung adenocarcinoma cells.In the BALB/c-nude mouse model of lung adenocarcinoma,tumor growth was inhibited following ATAD2interference.The expression of ATAD2 is associated with glycolysis,which has potential to be a target for the treatment of anti-tumor energy metabolism.