Radioactive Immuno-imaging And Radioimmunotherapy Of ¹³¹I-Labeled Human Single-chain Variable Fragment Antibodies Against Anaplastic Thyroid Cancer In Tumor-bearing Nude Mice

Objective:Anaplastic thyroid carcinoma has a high degree of malignancy and lacks effective treatment,as the tumor tissue does not respond to radioactive iodine therapy.With the continuous development of immunotherapy,we investigated the use of 131 iodine-labeled antibodies,to explore a new treatment method.Methods:The human single chain variable fragments antibodies against anaplastic thyroid carcinoma was successfully received.TEC cells and ARO cell were used as antigen and then four rounds of’adsoption-elution-amplification’ from the human phage display library against ATC.E.coli HB2151 was infected to induce the expression of soluble scFv by IPTG.ELISA was used to detect the expression of scFv.HiTrapTM Anti E-tag column was used for purification of soluble scFv.TT cells,ARO cells,Hep G2 cells and PBS blank control group were set up to identify specific of soluble scFv,the absorbance of A450nm was detected by microplate reader respectively.The expression and relative molecular mass of soluble scFv were tested by SDS-PAGE and Western blot.The chloramine T method was used to label scFv with 131I.Labeling rate,radiochemical purity and the distribution of 131I-scFv in body tissues and organs were analyzed.SPECT imaging was performed at 12,24,48,and 72 h after injection to observe the intratumoral accumulation of radioactivity.SPECT/CT image fusion was performed when the tumor tissues were clearly visible.Therapy group A:intravenous injection of saline 100 μL;group B intravenous injection of unlabeled 131I scFv 100 μL;group C intravenous injection of 131I-scFv 100 μL;group D:intratumoral injection of 131I-scFv 100 μL.The tumor volume change was observed at 1 day and then every 4 days to 25 days and the tumor inhibition rate was calculated.The survival time of each mouse was also recorded.The tumor tissue was removed from each mouse within 1 h after death,and fixed for histological analysis after hematoxylin and eosin staining;while the remaining part was fixed in glutaraldehyde for observation by transmission electron microscopy.Results:The purification and expression of soluble antibody scFv were completed.Phage ELISA results showed that specific binding with ARO cells.SDS-PAGE and Western blot said that the relative molecular mass of soluble scFv was about 29kD.The labeling rate was 91.66%and the radiochemical purity was 93.1 ± 0.32%.SPECT imaging showed that 131I-scFv selectively accumulated in tumor tissue.The target/non-target(T/NT)value was highest and clearest at 48 h,and SPECT/CT image fusion results were in agreement with the biodistribution results.The survival times of the four groups of tumor-bearing nude mice were 39.5,43.0,54.5,and 55.0 d.In the latter two groups,there were some treatment effects,and the outcomes were significantly better than in group B;however the treatment effect was not significantly different(P>0.05)between the two groups.Conclusion:We conclude that 131I-scFv selectively accumulated in tumor tissue and provided a more effective diagnosis and treatment method for the clinical treatment of anaplastic thyroid carcinoma.