| Objective:The unfavorable prognostic features includes age(?18 mo at presentation),stage(distant metastases in lymph nodes,cortical bone,bone marrow,and liver),and molecular pathology(MYCN oncogene amplification and chromosomal loss of 1p36).Therefore,the purpose of this study was to evaluate the prognostic value of 18-F-fluorodeoxyglucose(18F-FDG)positron emission tomography/computed tomography(PET/CT)in pediatric patients with neuroblastoma(NB);to investigate the imaging value of 18F-4-FGln in neuroblastoma compared to 18F-FDG PET imaging.Methods:1.Forty-seven pediatric patients with newly diagnosed neuroblastoma who underwent 18F-FDG PET/CT were retrospectively reviewed.Clinicopathological factors and metabolic parameters including maximum standardized uptake value(SUVmax),metabolic tumor volume(MTV),total lesion glycolysis(TLG)and bone marrow uptake(BMU)patterns on PET were compared to predict recurrence-free survival(RFS)and overall survival(OS)by univariate and multivariate analyses.2.MYCN amplification of NB cells IMR-32,SK-N-BE(2)and non-MYCN amplification NB cell SH-SY5Y as in vitro tumor model cell lines.The uptake of 18F-4-FGln and18F-FDG in NB three cell lines were measured by gamma counting meter,and the cells proliferation were monitored by CCK8 kits.In vitro IMR-32,SK-N-BE(2)and SH-SY5Y xenograft tumors were stablished,before the uptake of 18F-4-FGln and 18F-FDG were measured through a small animal PET,separately;Western blot(WB)and immunofluorescence(IF)were performed to analyze the expression of MYCN and ASCT2 in three tumor cells or tissues.3.ASCT2 knockdown cell and tumor models were established by sh RNA,and gene expression knockdown was detected by fluorescence quantitative PCR and WB.The uptake of 18F-4-FGln and 18F-FDG were measured in Sh ASCT2-IMR-32 and Sh Control-IMR-32 cell and mouse tumor models in vivo and in vitro.WB was performed to detect the level of ASCT2 and LAT1 after ASCT2 knockdown;weight and volum of tumors were measured;the expression of ASCT2,ki-67 and c-caspase-3 in tumor tissues was detected by IF assay.4.IMR-32 and SK-N-BE(2)the subcutaneous transplantation tumor models were established before intraperitoneal injection of GPNA and dimethyl sulfoxide(DMSO)for 3 weeks.Small animal PET imaging and IF were performed to compare uptake of tumors and to evaluate the expression ASCT2,KI-67,c-Caspase 3 before and after the treatment.The tumor size and tumor volume were measured after the treatment.Results:1.MTV(P=0.001),TLG(P=0.004)and BMU patterns(P=0.025)remained significant predictive factors for tumor recurrence,along with tumor size,histology,stage,lactate dehydrogenase(LDH),other distant metastasis(except bone metastasis).Univariate analyses showed that histology,stage,tumor size(>37.25cm),other distant metastasis,MTV(>88.10cm~3)and TLG(>1045.2g)and BMU patterns correlated with both RFS and OS.On multivariate analyses,TLG remained to be the only independent prognostic factor for RFS(P=0.016)and OS(P=0.012),and BMU patterns and MTV were statistic significantly for OS(P=0.024 and P=0.038,respectively).2.The uptake peak of IMR-32 and SK-N-BE(2)were significantly higher than that of SH-SY5Y and the difference was statistically significant(IMR-32 vs SH-SY5Y,P<0.001;SK-N-BE(2)vs SH-SY5Y,P<0.001).GPNA could significantly inhibit the uptake of 18F-4-FGln,while glutamine deactivation and CB839 increased the uptake in MYCN amplification NB cell line cells.Compared with 18F-FDG,the peak uptake of 18F-4-Fgln in IMR-32and SK-N-BE(2)cell lines were much higher.The levels of ASCT2 protein among IMR-32,SK-N-BE(2)and SH-SY5Y were statistically significant(IMR-32 vs SK-N-BE(2),P<0.01;IMR-32 vs SH-SY5Y,P<0.001;SK-N-BE(2)vs SH-SY5Y,P<0.01).3.The uptake peak of 18F-4-FGln appeared at 60-minute after intravenous injection in vivo,and the uptake of IMR-32,SK-N-BE(2)and SH-SY5Y tumors were6.62±0.36%ID/g,5.62±0.25%ID/g,3.75±0.15%ID/g,respectively;18F-4-FGln small animal PET imaging showed that the uptake of IMR-32,SK-N-BE(2)and SH-SY5Y tumors were 3.71,2.63 and 1.54,respectively,while the ratios of 18F-FDG were 1.90,1.21 and 1.23.18F-4-FGln PET/CT imaging showed no uptake in the inflammatory model.4.Sh ASCT2 RNA knockdown cells and tumor models of IMR-32 were established and the uptake of 18F-4-FGln was significantly lower than that of sh Control-IMR-32(P<0.01).Tumor growth slowed down(P<0.01),the expression level of ASCT2 and KI-67 protein decreased,and the c-caspase-3 water skin increased in sh ASCT2-IMR-32 tumor.The uptake of 18F-4-FGln was 5.71±2.38%ID/g,5.16±2.52%ID/g and 3.62±2.54%ID/g,3.87±1.84%ID/g(P=0.0017,P=0.0070),before and after the treatment of GPNA.The expression level of ASCT2 was decreased in MYCN amplification NB cells after the therapy(IMR-32,P<0.001;SK-N-BE(2),P<0.01).Conclusion:1.Patients with high MTV,TLG and focal bone marrow(unifocal and multifocal)uptake on PET/CT may have inferior outcome during the subsequent treament.2.18F-4-FGln PET imaging can evaluate the NB glutamine metabolism,and may provide a reliable way for clinical diagnosis,risk stratification and therapeutic effect evaluation in NB;3.ASCT2 is the most important transporter in the pathway of glutamine metabolism for NB,which is closely related to the growth and proliferation of tumors,and can provide a new therapeutic target for clinical treatment. |
PDF Full Text Request