Synthesis and radiolabeling of (S)-4-amino-5-iodo-2-methoxy-N-(1-azabicyclo[2.2.2]oct-3-yl)benzamide, the active enantiomer of [125I]iodozacopride, and re-evaluation of its 5-HT3 receptor affinity.

ABSTRACT

We report an improved synthesis of unlabeled (S)-iodozacopride, the radiolabeling of (S)-[125I]iodozacopride via deschloro-(S)-zacopride, and a re-evaluation of its affinity for the 5-HT3 receptor. Unlabeled (S)-iodozacopride was prepared in seven steps from 4-aminosalicylic acid via alkaline hydrolysis of its 4-acetamide derivative. Catalytic hydrogenation of (S)-iodozacopride gave deschloro-(S)-zacopride, identical to that obtained from (S)-3-amino-quinuclidine and 4-amino-2-methoxybenzoic acid via its corresponding 1-imidazole derivative. Radioiodination to produce (S)-[125I]iodozacopride was accomplished by treatment of deschloro-(S)-zacopride with 5 mCi sodium 125iodide and chloramine-T in hydrochloric acid. Purification of the reaction products using an HPLC system capable of detecting chlorinated side-products revealed a mixture of 2.1 mCi (1.3 nmol) (S)-[125I]iodozacopride and (S)-zacopride (1.5 nmol). Saturation analysis of the binding of the purified (S)-[125I]iodozacopride to whole rat brain homogenates gave an estimated KD of 1.10 +/- 0.07 nM. As anticipated, this is approximately half the KD reported for binding of racemic [125I]iodozacopride, and differs from the previously reported value by an order of magnitude. Analysis of the apparent binding affinity of a 11 mixture of (S)-[125I]iodozacopride and (S)-zacopride suggests that the previous result may have been confounded by contamination of the product with unlabeled (S)-zacopride. Competition analysis of the displacement of (S)-[125I]iodozacopride binding by unlabeled (S)-iodozacopride and (S)-zacopride gave Ki values of 0.95 and 0.21 nM, respectively.