Design and Structure/Conformation-Activity Studies of a Prototypic Corticotropin-Releasing Factor (CRF) Antagonist: Multiple Alanine Substitutions of CRF12-41

The known physiological role(s) and proposed pathophysiological properties of the neuroendocrine peptide CRF have been previously described (for review, see 1), and CRF has been shown to exert a variety of CNS-mediated effects on behavior2,3, cardiovascular system4,5, reproduction6,7, gastrointestinal secretion8,9, motility10, and transit11. Of particular significance is that CRF may, therefore, be involved in stress stimuli-induced activation of neural/humoral pathways leading towards anxiety and depressive disorders (e.g.,depiession, panic and anorexia nervosa). Nevertheless, the molecular pharmacology and mechanisms which are involved in stress-induced behavioral, endocrine and metabolic activities are not well defined. The discovery and development of potent CRF antagonists may provide key molecular probes to investigate the biological activities of endogenous CRF in animal models as well as for studying the molecular pharmacology of CRF-receptor interactions. Such studies have been reported12,13 and have been primarily based upon synthetic modification of CRF; yet the emergence of a high affinity analog of low molecular mass (i.e., small peptide or peptidomimetic) has remained elusive to date. Nevertheless, studies14-18 on the blockade of endogenous CRF using CRF antiserum or prototypic CRF antagonists have probed the possible role that endogenous CRF may have on the effects of stress in different animal models. Of noteworthy contribution to such CRF research has been both structure-activity and structure-conformation studies12,13,19,20 to investigate CRFreceptor binding and functional properties (agonism/antagonism). These studies have culminated in the identification of prototypic CRF antagonists (or partial agonists) which were modified fragment analogs of the native peptide. Specifically, compound I (Fig. 1) has been advanced13 as a significant lead towards the development of high affinity CRF receptor antagonists. In this report we describe analogs of I to further explore the role of side-chain functionlization in CRF receptor binding using a strategy of multiple (iterative) Ala substitution with a particular focus on the central domain of this CRF analog corresponding to CRF22-31 In addition, the structure-conformation properties of these analogs were investigated by circular dichroism spectroscopy.