A direct radiolabeling of 99Tcm cyclic RGD peptide, an antagonist of αvβ3 receptor

Objective: It has been reported that Cys-Asp-Cys-Arg-Gly-Asp-Cys-Lys-Cys (RGD) peptides, as integrin αvβ3 receptor antagonists, might inhibit angiogenesis of tumor. The aim of the present study was to investigate the feasibility of direct labeling of 99Tcm to a cyclic nine peptide containing RGD sequence (RGD-4CK), to observe the effects of different conditions on labeling efficiency, and to evaluate the radiochemical property of 99Tcm-RGD-4CK. Methods: The peptide RGD-4CK contained the sequence of Cys-Asp-Cys-Arg-Gly-Asp-Cys-Lys-Cys. It was labeled directly by 99Tcm with 'pretinning' method. The Rf value of 99Tcm-RGD-4CK and 99TcmO4 were determined in 3 MM paper chromatography. The labeling efficiency and specific activity were calculated. The influences of various conditions to the labeling rate were studied. High performance liquid chromatography (HPLC) analysis and Sep-Pak C18 chromatography were used to assess the property of radiolabeled 99Tcm-RGD-4CK. The stability in vitro, the cysteine replacement and the serum protein combination were also tested. Results: The Rf values of 99Tcm-RGD-4CK were 0 and 0.8-0.9 respectively in mobile phase of acetone and V(NH4OH): V(ethanol): V(water) = 1: 2: 5. The radiochemical yield was (97.8±0.4)% and the specific activity (11.90±0.05) TBq/mmol in the basic condition of labeling. The labeling efficiency remained over 95% under the condition of 150-300 μg stannous tartrate, pH value 2.0-3.5, temperature 60 degree C and incubation time over 6 h in the process of pretinning reaction, the activity of 99TcmO4- was 37-185 MBq within volume of 200 μl. With the labeling temperature of 95 degree C in 30 min, the retention of 99Tcm-RGD-4CK was in accordance with the major peak of time-activity curve from the analytical HPLC. The radiochemical purity of 99Tcm-RGD-4CK remained above 95% after 6 h at room temperature. The amount of 99TcmO4-; increased only by 0.5% as measured with Sep-Pak C18 chromatography. The free 99TcmO4- from the labeled product increased by 1.88% after incubation with 300 mmol/L cysteine for 1 h at 37 degree C. There was no obvious binding of 99Tcm-RGD-4CK to serum protein. Conclusions: This study indicated that direct 'pretinning' techniques was practical in preparing 99Tcm- RGD-4CK with acceptable labeling efficiency and radiochemical characteristics, and might be potentially useful in future preparation of 99Tcm-RGD-4CK kit. (authors)