Victor J. Hruby

Differentiation between rat brain and mouse vas deferens δ opioid receptors

Certain enkephalin analogues, including those which contain the conformationally restricted amino acid E-(2R,3S)-cyclopropylphenylalanine [2R,3S)-delta E Phe), have been shown to have high affinity for brain delta opioid receptors but are much less active in mouse vas deferens bioassays. To investigate whether there are differences between delta opioid receptors in brain and mouse was deferens, the ability of a selective delta opioid compound, [D-Pen2,pCl-Phe4,D-Pen5]enkephalin (pCl-DPDPE), and [D-Ala2,(2R,3S)-delta E Phe4,Leu5]enkephalin methyl ester (CP-OMe), to inhibit [3H]pCl-DPDPE binding in both rat brain and mouse vas deferens were measured. pCl-DPDPE recognized brain and mouse vas deferens binding sites with equal affinity, however, CP-OMe showed 33 fold lower affinity in mouse vas deferens compared to brain. This suggests that mouse vas deferens delta opioid receptors may be distinct from brain delta opioid receptors.

CHAPTER 13:Peptide Therapeutics: Neuropeptides

The development of neuropeptide therapeutics poses several difficulties, including stability, bioavailability, receptor selectivity, and the ability (or not) to cross membrane barriers. In this chapter we discuss briefly the efforts that are being made to address these problems using selective examples for the melanocortin, bradykinin and opioid systems. The use of conformational constraint, topographical constraint, and computational approaches to ligand design, including ligand–receptor homology modeling are discussed as an approach to enhance the properties of peptides for the development of peptide therapeutics.

Effects of Methyl and Halogens in Analgesic Peptide Molecules for Their Potent Agonist Activities at MOR/DOR and Antagonist Activity at NK1R

Treating pain has been always challenging, especially when it becomes chronic in nature. Current drugs (e.g. opioid drugs) cannot treat this problem effectively. In addition, constant use of these drugs has deadly side effects including drowsiness and mental clouding, nausea and emesis, and constipation [1]. Development of analgesic tolerance and hyperalgesia in many patients as the results of taking these drugs are also a serious concern. Overexpression of substance P and its receptor has been observed during prolonged pain states. Studies revealed that co-administration of cocktails of drugs containing μ/δ opioid agonist and NK1 antagonist can provide better analgesic effects in rat model while reducing side effects [2-4]. Herein, we report design, synthesis and SARs of few multifunctional ligands having μ/δ opioid agonist (μ-preferring) and NK1 antagonist activities. The detail of this study has already been communicated to J. Med. Chem. for publication [5]. We anticipate that these ligands will show better analgesia while reducing the adverse side effects. Our drug design principle is based on overlapping and adjacent pharmacophores (Figure 1) and we considered our previously published μ-selective (binding) ligand TY012 (H-Tyr-D-Ala-Gly-Phe-Pro-Leu-Trp-NH-Bn(3ʹ,5ʹ-(CF3)2)) [6].

Design of Multivalent Ligands that Cross the Blood Brain Barrier for the Treatment of Neuropathic Pain Without Toxicities

Pain, especially prolonged and neuropathic, is the most ubiquitous and expensive disease in the U.S. and worldwide and treatments are inadequate or ineffective and often lead to enhanced pain sensitivity, addiction and poor quality of life. New modalities for treatment are urgently needed but are not forthcoming from pharma. We have proposed that a new approach to drug design is needed which takes into account the changes in the expressed genome that accompany these disease states [1]. In particular, in prolonged and neuropathic pain there is up-regulation of neurotransmitters and their receptors in ascending and descending pain pathways that are stimulatory and therefore enhance the pain perception. From these and other findings, we have hypothesized that the design of a multivalent ligand that has agonist activity at the mu and delta opioid receptors (balanced or favoring μ or δ receptors) with antagonist activities at the up-regulated stimulatory receptor (e.g. ligands for neurokinin-1 (NK-1) receptor) all in single ligand, would have potent analgesic activities but without the toxicities of current opioid drugs and without the development of tolerance or addiction.

Differential Potencies for Endogenous Dynorphins Indicate Functional Selectivity at the Delta Opioid Receptor

Opioid receptors (ORs) are G-protein Coupled Receptors (GPCRs), which mediate analgesia, tolerance, withdrawal, GI transit. Classically, ORs couple inhibitory Gi/o proteins and recruit arrestin – a multifaceted scaffold molecule implicated in opioid mediated effects including tolerance, constipation, dysphoria and naseua [1,2]. Upon activation -arrestin and Gi/o induce downstream signaling responses such as reduced cAMP levels. Recent drug discovery efforts identified several functionally selective exogenous opiates which prefer certain signaling pathways at a given receptor – such as G stimulation – to others – such as -arrestin recruitment and generate desired pharmacological properties [3,4]. Noting that most of the 20+ endogenous opioid peptides are nonselective and some opiates display functional selectivity, two important points emerge. First, endogenous and exogenous ligands, such as those used during studies, do not necessarily generate the same effects. Second, two different endogenous opioid peptides may differentially activate a given receptor. Dynorphin A (DynA) and Dynorphin B (DynB) are considered OR agonists, despite binding to the OR at 1.29nM and 3.39 nM [1], respectively. The Dynorphins start with the 5 amino acid Leuenkephalin (Leu-Enk) sequence – YGGFL – traditionally considered a OR agonist followed by distinct C-terminal sequences. Thus, we ask: Do Dynorphin A (1-17), Dynorphin B (1-13) and Leuenkephalin induce functionally selective signaling at the OR?