Dawna L. Evans

Dopamine D4 versus D2 receptor selectivity of dopamine receptor antagonists: possible therapeutic implications

The dopamine D4 receptor, which is considered a close variant of the dopamine D2 receptor, has recently been cloned. Receptor binding studies demonstrated that clozapine, which is an effective antipsychotic agent but atypical in that it lacks the usual side effects of other antipsychotic agents, has high selectivity for the dopamine D4 receptor versus the dopamine D2 receptor. Comparative binding affinity studies have been carried out for a number of interesting dopaminergic agents using membranes prepared from cloned dopamine D2 and D4 receptor containing cells. It was found that clozapine is selective for the dopamine D4 vs. the D2 receptor by a factor of 2.8. Other compounds with dopamine D4 receptor selectivity were (+)-apomorphine (8.7), (+)-N-propyl-norapomorphine (NPA) (2.4) and melperone (1.3). Compounds with considerable selectivity for the dopamine D2 receptor were haloperidol (0.31), chlorpromazine (0.084), trifluoperazine (0.034) and raclopride (0.001). Overall, the results with the antipsychotic agents tested, support the concept that dopamine D4 receptor selectivity may confer clozapine-like antipsychotic efficacy and furthermore that dopamine D2 receptor selectivity may confer side effect liability (extrapyramidal side effects and tardive dyskinesia).

Rat brain binding sites for pramipexole, a clinically useful D3-preferring dopamine agonist

Pramipexole (PPX) is currently being evaluated for treatment of schizophrenia and Parkinsons disease. In studies with cloned subtypes of the dopamine (DA) D2 receptor subfamily, PPX has higher affinity for the D3 compared to the D2 and D4 subtypes; unlike 7-[3H]hydroxy-N,N-di-n-propyl-2-aminotetralin (7-OH-DPAT), it does not bind to sigma sites. Receptor binding autoradiography with [3H]PPX (5 nM, 62 Ci/mmol) was used to evaluate the distribution of PPX binding sites within the rat brain. Consistent with its preference for D3-binding sites, the highest concentrations of [3H]PPX binding sites were found in the islets of Calleja (ICj), previously reported to contain D3 but not D2 or D4 mRNA. [3H]PPX binding was also high in other mesolimbic areas such as the nucleus accumbens (N. accum), olfactory tubercle, and amygdala. [3H]PPX binding was also high in caudate (Cd), although slightly less than in mesolimbic areas. Less [3H]PPX binding sites were found in ventral tegmental area (VTA) and substantia nigra, areas rich in cell bodies for DA neurons. Thus, although PPX most potently stimulates DA autoreceptors, PPX binding sites have their highest concentrations in projection areas containing both DA terminal and postsynaptic receptors. Because of PPXs preferential affinity for the D3 receptor subtype and its resultant high mesolimbic binding, it could have a unique therapeutic profile for treatment of psychiatric and/or neurological diseases.

Dopamine D2 receptor binding properties of [3H]U-86170, a dopamine receptor agonist

U-86170F, an imidazoguinolinone, is a potent dopamine D2 agonist, binding with high affinity to the dopamine D2 receptor. A Kd of 0.99 nM was determined in membranes from Chinese hamster ovarian (CHO) cells transfected with the D2 receptor and a Kd of 1.72 nM was obtained in rat striatal homogenates. GTP sensitivity was demonstrated when its addition (300 microM) reduced [3H]U-86170 binding by 60%. This agonist ligand is especially effective in identifying agonists and partial agonists, as well as antagonists, and affords a more precise evaluation of their affinity for the dopamine D2 receptor, without the use of multiple site analysis, than does an antagonist [3H]-ligand.

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.