Cedarville University DigitalCommons@Cedarville Pharmacy Faculty Publications School of Pharmacy 10-2008 The ffecE ts of Herkinorin, the First mu-Selective Ligand from a Salvinorin A-Derived Scaffold, in a Neuroendocrine Biomarker Assay in Nonhuman Primates Eduardo R. Butelman Szymon Rus Denise S. Simpson Cedarville University, [email protected] Angela Wolf Thomas E. Prisinzano See next page for additional authors Follow this and additional works at: http://digitalcommons.cedarville.edu/pharmacy_publications Part of the Pharmacy and Pharmaceutical Sciences Commons Recommended Citation Butelman, E. R., Rus, S., Simpson, D. S. , Wolf, A., Prisinzano, T. E. , Mary Jeanne Kreek, (2008). The Effects of Herkinorin, the First mu-Selective Ligand from a Salvinorin A-Derived Scaffold, in a Neuroendocrine Biomarker Assay in Nonhuman Primates. Journal of Pharmacology and Experimental Therapeutics, 327, 154-160. This Article is brought to you for free and open access by DigitalCommons@Cedarville, a service of the Centennial Library. It has been accepted for inclusion in Pharmacy Faculty Publications by an authorized administrator of DigitalCommons@Cedarville. For more information, please contact [email protected]. Authors Eduardo R. Butelman, Szymon Rus, Denise S. Simpson, Angela Wolf, Thomas E. Prisinzano, and Mary Jeanne Kreek This article is available at DigitalCommons@Cedarville: http://digitalcommons.cedarville.edu/pharmacy_publications/14 0022-3565/08/3271-154–160$20.00 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 327, No. 1 Copyright © 2008 by The American Society for Pharmacology and Experimental Therapeutics 140079/3381513 JPET 327:154–160, 2008 Printed in U.S.A. The Effects of Herkinorin, the First ␮-Selective Ligand from a Salvinorin A-Derived Scaffold, in a Neuroendocrine Biomarker Assay in Nonhuman Primates Eduardo R. Butelman, Szymon Rus, Denise S. Simpson, Angela Wolf, Thomas E. Prisinzano, and Mary Jeanne Kreek The Rockefeller University, New York, New York (E.R.B., S.R., M.J.K.); Division of Medicinal and Natural Products Chemistry, University of Iowa, Iowa City, Iowa (D.S.S., T.E.P.); and Department of Medicinal Chemistry, University of Kansas, Lawrence, Kansas (D.S.S., A.W., T.E.P.) Downloaded from Received April 16, 2008; accepted June 30, 2008 ABSTRACT ␮ Herkinorin is the first -opioid receptor-selective ligand from effectiveness in females and revealed a fast onset after i.v. jpet.aspetjournals.org the salvinorin A diterpenoid scaffold. Herkinorin has relative administration (e.g., by 5–15 min). Antagonism experiments ␮ Ͼ ␬ Ͼ ␦ binding selectivity, and it can act as an agonist at with different doses of nalmefene (0.01 and 0.1 mg/kg) caused both ␮- and ␬-receptors, in vitro. These studies were the first dose-dependent and complete prevention of the effect of herki- in vivo evaluation of the effects of herkinorin in nonhuman norin in females. This is consistent with a principal ␮-agonist primates, using prolactin release, a neuroendocrine biomarker effect of herkinorin, with likely partial contribution by ␬-agonist assay that is responsive to both ␮- and ␬-agonists, as well as effects. The peripherally selective antagonist quaternary nal- to compounds with limited ability to cross the blood-brain trexone (1 mg/kg s.c.) caused approximately 70% reduction in barrier. In cumulative dosing studies (0.01–0.32 mg/kg i.v.), the peak effect of herkinorin (0.32 mg/kg) in females, indicating at Cedarville Univ Lib on January 25, 2013 herkinorin produced only small effects in gonadally intact males that this effect of herkinorin is prominently mediated outside the (n ϭ 4), but a more robust effect in females (n ϭ 4). Time course blood-brain barrier. studies with herkinorin (0.32 mg/kg) confirmed this greater Salvinorin A, a plant-derived hallucinogenic diterpene, is a To date, there is no information on the in vivo effects of highly selective ␬-opioid agonist, and a novel template for herkinorin in primates. The present study focuses on an semisynthetic opioid analogs (Roth et al., 2002; Prisinzano initial evaluation of the in vivo opioid agonist effects of and Rothman, 2008). One of these novel analogs is herki- herkinorin in rhesus monkeys, using a translationally vi- norin, the first salvinorin-derived compound with ␮- over able neuroendocrine biomarker assay, release of the ante- ␬-selectivity reported in the literature (Harding et al., 2005). rior pituitary hormone prolactin. Different factors render Herkinorin has approximately 8-fold selectivity for ␮- over this neuroendocrine biomarker a practical approach for ␬-receptors and approximately 98-fold selectivity for ␮- over initial evaluation of herkinorin: 1) prolactin levels are ␦-receptors in competition binding assays (Harding et al., increased by both ␮- and ␬-agonists in mammals, including ␮ 2005). Herkinorin acts as a high-efficacy agonist at both - humans (␦-agonists seem not be to be active) (Hoehe et al., ␬ Ј and -receptors in the guanosine 5 -O-(3-thio)triphosphate 1988; Ur et al., 1997; Kreek et al., 1999; Bowen et al., 2002; assay (Harding et al., 2005), with greater relative potency at Butelman et al., 2007); and 2) effects of these opioids are at ␮ -receptors. Herkinorin also displays some unique features least partially mediated by opioid receptors outside the ␮ in its interactions at the -receptor, such as decreased ago- blood-brain barrier (e.g., in particular hypothalamic nuclei) nist-induced internalization (Groer et al., 2007; Xu et al., (Merchenthaler, 1991; Butelman et al., 2004, 2008; Zheng et 2007). al., 2005), and they may therefore be used to determine the pharmacodynamic effects of compounds with limited ability This work was funded by National Institutes of Health-National Institute on Drug Abuse Grants DA017369 (to E.R.B.), DA018151 (to T.E.P.), and to cross the blood-brain barrier. DA05130 (to M.J.K.). This study presents the first direct evaluation of the phar- Article, publication date, and citation information can be found at http://jpet.aspetjournals.org. macodynamic effects of herkinorin in male and female non- doi:10.1124/jpet.108.140079. human primates, and it also investigates potential ␮-versus ABBREVIATIONS: ANOVA, analysis of variance. 154 Herkinorin Effects on a Neuroendocrine Biomarker 155 ␬-receptor effects of this novel compound, as well as activity occurred 15 min after each injection. Herkinorin was studied herein outside the blood-brain barrier. up to the largest dose that could be administered due to solubility limitations (0.32 mg/kg). Herkinorin was then compared with salvi- norin A (its structurally related parent compound, a selective ␬-ag- Materials and Methods onist) and loperamide (a peripherally selective ␮-agonist), under Subjects similar conditions. A vehicle control study was also carried out, with four consecutive vehicle injections under identical timing conditions. Four captive-bred male and four female rhesus monkeys (Macaca Time Course Procedure. After baseline sample collection, a mulatta; age range, approximately 8–12 years; weight range, 7.0– single agonist dose (e.g., of herkinorin) was administered i.v., fol- 12.5 kg) were used. All subjects were gonadally intact. Unless oth- lowed by sampling at 5, 15, 30, 60, 90, and 120 min after adminis- ϭ erwise stated, each study was carried out with an n 4 of either tration. In antagonism experiments, a single dose of antagonist (s.c. males or females. Females were studied in the follicular phase, nalmefene or quaternary naltrexone) was administered 30 min be- estimated as days 1 to 12 from the onset of visible menses. Monkeys fore an agonist, followed by testing as described above. In these were singly housed in colony rooms maintained at 20 to 22°C, with antagonism experiments, a single sample was also taken 20 min controlled humidity and a 12:12-h light/dark cycle (lights on at 7:00 after administration of the antagonist alone (i.e., during the pre- AM). Monkeys were fed approximately 12 jumbo primate chow bis- treatment period). cuits, adjusted individually (PMI Feeds, Richmond, VA). Subjects also received appetitive treats, and multivitamins plus iron. An Design environmental enrichment plan was in place (e.g., music and videos). Water was freely available in home cages, via a waterspout. Cumulative Dose-Effect Curve Studies. Cumulative dose-ef- Downloaded from Subjects had complex history of pharmacological exposure (pri- fect curve studies were carried out in males and females for herki- marily opioid), but they had no history of any chronic or high- norin (0.01–0.32 mg/kg i.v.) compared with repeated vehicle injec- frequency exposure to any agent. Consecutive experiments in the tion, for control purposes. The effects of loperamide [0.01–0.32 mg/kg same subject were separated by at least 72 h; the order of experi- (males and females)] and salvinorin A [0.001–0.032 mg/kg (males) ments was unsystematic among subjects. All studies were carried and 0.00032–0.01 mg/kg (females), based on prior studies; Butelman et al., 2007, 2008] were also studied under identical cumulative out over the course of several months while all subjects were in jpet.aspetjournals.org stable colony rooms. Studies were reviewed by the Rockefeller Uni- dosing procedures. versity Animal Care and Use Committee, in accordance with the Time Course Studies. The time course effects of herkinorin were Guide for the Care and Use of Animals (National Academies Press; studied in male subjects, at the largest dose that could be adminis- Washington, DC, 1996). tered, as limited by solubility (0.32 mg/kg i.v., compared with vehicle i.v.). This was followed by a similar determination of the effects of Procedure for Neuroendocrine Experiments this dose in females. Because males demonstrated only a small effect of herkinorin up Chair-trained monkeys were tested after repeated exposure to the to the largest dose that could be studied, antagonism experiments at Cedarville Univ Lib on January 25, 2013 experimental situation. Monkeys were chaired and transferred to described below were only carried out in females.