Page - (000314) - in Biomedical Chemistry: Current Trends and Developments

the double nature of EM-2 (Fichna, 2007; Kruszynski, 2005; Okada, 2003). In the Ala scan of EM-2, substitution of the internal phenylalanine was accepted (12), but truncation down to a single phenylalanine resulted in 27 with no binding affinity (Ki of 5028 nM). The binding features of the new dipeptide lead compound H-Phe- Phe-NH2 were further explored via the synthesis of peptides 28–32 (Table 3.3.4). As observed for SP1–7 and EM-2, the C-terminal function should be a primary amide. The corresponding carboxylic acid was devoid of activity (cf. 26 with 28). All four stereoisomers of H-Phe-Phe- NH2 (26, 30, 31 and 32) were synthesized and evaluated. The natural l- Phe-l-Phe isomer (26) was found to be preferred, followed by the d,d compound (31), although with a 40-fold lower affinity. As mentioned above, incorporation of d-amino acids into neuropeptides can change their biological function from an agonist to an antagonist, e.g. d-SP1–7, which makes it interesting to evaluate the analogs 30, 31 and 32 in animal studies concerning their functional activities. Due to the smaller size of 26 in comparison to the heptapeptide SP1–7, lower selectivity can be expected. Moreover, 26 resembles ligands for the NK3 receptor (Boden, 1995; 1994). Hence, the possible binding affinity of 26 to the human neurokinin receptors NK1 and NK3 was studied. Binding was evaluated in agonist radioligand binding assays relying on the displacement of [Sar9, Met(O2)11]-SP from NK-1 receptors, and [MePhe7]-NKB from NK-3 receptors (Anthes, 2002; Heuillet, 1993), 26 was tested at a concentration of 10 µM, but showed no affinity for any of the receptors. Table 3.3.4: Ki values of Phe-Phe analogs for inhibition of [3H]-SP1–7 binding to rat spinal cord membrane. Compound Sequencea Ki ± SEM (nM) 26 1.5 ± 0.1