Preparation and characterization of [99TcO] apcitide: a technetium labeled peptide.

[ 9 9 m TcO] apcitide ( 9 9 m TcO(P246)), the technetium complex of the 13 amino acid, apcitide, cyclc-(D-Tyr-Apc-Gly-Asp-Cys)-Gly-Gly-Cys(Acm)-Gly-Cys(Acm)-Gly-Gly-Cys-NH2, where Apc is L-[S-(3-aminopropyl)]cysteine (an arginine mimetic) and Acm is the acetamidomethyl protecting group, has high affinity and selectivity for the GPIIb/IIIa receptor that is expressed on the membrane surface of activated platelets and plays an integral role in platelet aggregation and thrombus formation. Bibapcitide, a 26 amino acid, bis-succinimidomethyl ether-linked dimer of the peptide apcitide has been formulated as a single-vial, lyophilized kit having the trade name AcuTect. When sterile, nonpyrogenic sodium pertechnetate ( 9 9 m TcO 4 -) in 0.9% sodium chloride is added to the AcuTect radiopharmaceutical kit and the resulting kit is heated, [ 9 9 m TcO] apcitide forms. This is the first radiopharmaceutical to target acute deep vein thrombosis (DVT) in the lower extremities. We report here the preparation, purification, and isolation of the 9 9 Tc complex of apcitide and its characterization to determine the mode of binding of Tc to apcitide. [ 9 9 TcO] apcitide was prepared, on the macroscopic level, by reaction of [ 9 9 TcOCl 4 ] - with apcitide, purified by preparative HPLC and isolated as a trifluoroacetate salt. [ 9 9 TcO] apcitide can also be formed from the reaction of bibapcitide and 9 9 TcO 4 in the presence of Sn(II) and glucoheptonate at 80 °C, conditions that mimic the radiopharmaceutical kit preparation. FTIR data show a Tc=O stretch at 961.2 cm - 1 , in the range observed for anionic [Tc v O] 3 + amide thiolate complexes. The mass spectral data is in agreement with the formula, [C 5 1 H 7 3 O 2 0 N 1 7 S 5 Tc] - , consistent with retention of Acm groups and the Tc binding in the Gly 1 1 -Gly 1 2 -Cys 1 3 region of the peptide. Despite significant spectral overlap due to numerous similar amino acids, all protons of apcitide and [ 9 9 TcO] apcitide were unambiguously assigned. The observation of two nonequivalent Acm groups and the observation of only 10 NH-CH cross-peaks in the TOCSY and COSY spectra of [ 9 9 TcO] apcitide (NH-CH cross-peaks were absent for Gly 1 1 -Gly 1 2 -Cys 1 3 ), compared to all 13 cross-peaks found in apcitide, provided compelling evidence to support the 9 9 Tc binding to the terminal Gly 1 1 -Gly 1 2 -Cys 1 3 region of apcitide.