In vitro studies of glypican-3 targeting pretargeting technology for molecular MRI of hepatocellular carcinoma / 中华放射学杂志

ABSTRACT

Objective To explore the value of pretargeting technology in vitro MRI of L5 peptide guided streptavidin-conjugated and polyethylene glycol modification protected ultra-small superparamagnetic iron oxide(SA-PEG-USPIO) to hepatocellular carcinoma(HCC) via glypican-3(GPC3) receptor.Methods Direct immumofluorescence assay with carboxyfluorescein(FAM) labeled L5 and competitive inhibition was performed in HepG2 and HL-7702 cells.Imaging was obtained from fluorescent microscope.Immunoassay fluorescence images were carried out to determine the expression of GPC3 in HepG2 cell.PEG-USPIO conjugated with streptavidin was made by carbodiimide reaction,and the hydrodynamic diameters,Zeta potential and magnetic relaxivity of SA-PEG-USPIO and PEG-USPIO were measured.HL7702 cells were used for evaluate cells viability of SA-PEG-USPIO and PEG-USPIO.HepG2 and HL-7702 cells were used as experimental and control group respectively.Each of the two cell lines were further divided into three groupsL5-BT united SA-PEG-USPIO group,SA-PEG-USPIO group and control group.Prussian blue staining and MRI was preformed to observe the targeting efficacy of SA-PEG-USPIO respectively,and normalized T2 signal was recorded.The significant changes of normalized T2 signal intensity among groups was deterumine by using One-way analysis of variance.Results There were much more fluorescences on the membrane and cytoplasm of HepG2 cells than those on HL-7702 cells and cells of competition group.And indirect immunofluorescence images show the obvious expression of GPC3 in HepG2 cell.The SA-PEG-USPIO and PEG-USPIO nanoparticles had hydrodynamic diameters of (22.73 ± 3.31) and (35.97±5.19)nm,Zeta potential of them were (4.22±0.53) and (-7.91± 1.22)mV and magnetic relaxivity were 0.139 4× 103 and 0.103 9 × 103 mM-1s1.Although the highest concentration of SA-PEG-USPIO and PEG-USPIO was 2.4 mmol/L,cells viability was greater than 80％.The most iron particle was observed in L5-BT united SA-PEG-USPIO group of HepG2 cells.In vitro MR,the normalized T2 signal intensity of HepG2 cells in L5-BT united SA-PEG-USPIO group,SA-PEG-USPIO group and control group were 39±7,77 ± 12 and 93 ± 4.There was significant difference among those three groups (F=23.96,P＜0.01).The normalized T2 signal intensity of HL-7702 cells in each of three groups were 69± 11,78±8 and 95±5.There was no significant difference among those three groups (F=2.86,P＞0.05).Conclusion By the pretargeting method,L5 peptide guided SA-PEG-USPIO has effective targeting ability to HepG2 cells in vitro.