Basic Science Investigation
Preparation of 99Tcm-CP3-peptide and in vitro and in vivo study of lung adenocarcinoma cell lines
Kong Qi, Liu Yan, Liu Zhiguo, Wang Xiaohui, Yang Guoren
Published 2019-01-25
Cite as Int J Radiat Med Nucl Med, 2019,43(1): 40-46. DOI: 10.3760/cma.j.issn.1673-4114.2019.01.008
Abstract
ObjectiveTo preparation a 99Tcm labeled caspase -3 polypeptide apoptosis molecular probe(99Tcm-CP3-peptide) based on aspartic glutamic acid-valine-aspartic acid(DEVD) as the core and study the biodistribution and SPECT imaging of lung adenocarcinoma cell line(A549) after chemotherapy.
Methods99Tcm-cp3-peptide was synthesized by bifunctional chelate method, and the radiochemical purity was detected by HPLC at the time points of 0, 1, 2, 4, 6 hours after the labeling. An in vitro binding assay was performed in A549 cells treated with paclitaxel.In the control group, without chemotherapy, the intracellular and extracellular radioactive ratio(Cin/Cout) was measured, and the cell apoptosis rate was detected by flow cytometry. 99Tcm-CP3-peptide was injected via tail vein of tumor bearing nude mice treated with paclitaxel(3.7 MBq for each mouse), to detect the biological distribution of 5, 15, 30, 60, 120, 240 min respectively. Static SPECT imaging was performed, ROI outlined, and the ratio of tumor/contralateral normal muscle tissue was calculated. HE staining was performed to observe the morphological changes of tumor tissues after imaging. Test was used for comparison between the two groups, and bivariate correlation analysis was used for correlation study, P<0.05 was considered statistically significant.
ResultsThe radiochemical purity of 99Tcm-CP3- peptide at 0, 1, 2, 4 and 6 h was greater than 97%. The synthesis rate was(64.5+5.2)%. An in vitro investigation showed that the Cin/Cout in the tumor cell chemotherapy group was 10.27±2.02 vs. 1.09±0.03 in control group 24 h after paclitaxel chemotherapy. The chemotherapy group of Cin/Cout was 7.3 times as high as that of the control group. Flow cytometric detection of tumor cells in the same state, the rate of apoptosis in the chemotherapy group was(75.62±2.57)% vs.(3.42±0.32)% in contorl group. The ratio of Cin/Cout in the chemotherapy group and the control group was positively correlated with the percentage of apoptotic cells detected by flow cytometry(r=0.970, P<0.05). In vivo distribution of tumor bearing mice showed that: 99Tcm-CP3-peptide is rater fast in blood clearance, and mainly metabolized through kidney. The values of radioactive uptake were low for important organs such as heart, spleen and lung etal, but it is slightly higher in the liver and slower in metabolism. The value of tumor tissue uptake reached a peak(4.26±1.03)%ID/g after injection of 1 h. Clear images of the tumor bearing mice can be obtained 1h after injection of drugs. The ROI showed that the ratio of T/NT in the chemotherapy group was 3.83±0.11, which was significantly higher than that of the control group(1.57±0.09, t=16.19, P<0.05). After imaging, HE staining of tumor tissue showed that a large number of nuclear retraction, nuclear fragmentation and apoptotic cells were found in the tumor tis-sue of the chemotherapy group and only a small amount of that were in control group.
Conclusions99Tcm-CP3-peptide, a kind of imaging agent targeting caspase-3 activity,which has good biological distribution, and can be applied to apoptosis imaging of animal models, it has a potential clinical value in monitoring tumor cell apoptosis after chemotherapy.
Key words:
Isotope labeling; Peptides; Apoptosis; Tomography, emission-computed, single-photon; Tumor
Contributor Information
Kong Qi
School of Medicine and Life Sciences, University of Jinan-Shandong Academy of Medical Sciences, Jinan 250000, China; Department of Nuclear Medicine, Shandong Cancer Hospital Affiliated to Shandong University, Jinan 250117, China
Liu Yan
Department of Nuclear Medicine, Qingdao Municipal Hospital, Qingdao 266011, China
Liu Zhiguo
Department of Nuclear Medicine, Shandong Cancer Hospital Affiliated to Shandong University, Jinan 250117, China
Wang Xiaohui
Department of Nuclear Medicine, Shandong Cancer Hospital Affiliated to Shandong University, Jinan 250117, China
Yang Guoren
Department of Nuclear Medicine, Shandong Cancer Hospital Affiliated to Shandong University, Jinan 250117, China