Page - (000309) - in Biomedical Chemistry: Current Trends and Developments

existence of a specific receptor for the N-terminal partial sequences of SP in mouse spinal cord was discussed at the beginning of the 1980s (Piercey, 1982) and in 1990 Igwe et al. characterized the specific binding site of SP1–7 in mouse brain and spinal cord (Igwe, 1990). Binding was found to be specific, saturable and reversible, which strongly supported the existence of an N-terminal-directed SP receptor. In 2006, Nyberg and coworkers demonstrated the presence of specific binding sites for SP1–7 in rat spinal cord in accordance with previously reported results from the mouse spinal cord and brain (Botros, 2006; Igwe, 1990). A binding assay was developed using spinal cord tissue homogenate and measurements of the binding affinity for various compounds by displacement of tritiated SP1–7 ([3H]-SP1–7), which was a prerequisite for the medicinal chemistry program which will be described below. Initially, ligands for opioid and neurokinin receptors, as well as various N-terminal SP fragments, were screened against the SP1–7 binding site. In short, none of these ligands did bind efficiently to this binding site. However, interestingly, the high-affinity µ-opioid receptor agonists endomorphin-2 (EM-2) and endomorphin-1 (EM-1) were shown to interact differentially with the binding site of SP1–7. Thus, EM-2 had only a 10-fold lower affinity than SP1–7, whereas EM-1 had a 1,400-fold lower affinity. To rule out involvement of the µ-opioid receptor, the binding affinity of SP1–7 to the µ-opioid receptor and the ability of the heptapeptide to activate it were further investigated (Botros, 2006). However, no specific binding or any activation of the µ-opioid receptor was observed, which is indicative of a specific target protein for SP1–7, identical to neither the tachykinin receptors nor the µ-opioid receptor. 3.3.2.2 SAR and Truncation Studies of SP1–7 and EM-2 3.3.2.2.1 Strategy A SAR study of the two peptide leads, SP1–7 and EM-2, was initiated, with the intention of identifying the pharmacophoric groups. This design strategy included Ala scans, truncations and C- and N-terminal modifications of the two target peptides (Fig. 3.3.9). Thus, a series of