期刊
PHARMACOLOGICAL REPORTS
卷 65, 期 4, 页码 836-846出版社
SPRINGER HEIDELBERG
DOI: 10.1016/S1734-1140(13)71064-8
关键词
in vivo analgesia; PK20 metabolite; [S-35]GTP gamma S; antinociception; opioid receptor binding
资金
- European Grant Normolife [LSHC-CT-2006-037733]
- Fund for Scientific Research-Flanders (FWO-Vlaanderen) [G.000.08]
- European Social Fund, Human Capital Operational Programme for the execution of the project Support for bio tech med scientists in technology transfer [UDA-POKL.08.02.01-14-041/09]
Background: Recently, we presented a novel compound (PK20, Dmt-D-Lys-Phe-Phe-Lys-Lys-Pro-Phe-Tle-Leu-OH) that targets single entity opioid and neurotensin pharmacophores. This endomorphin-2-like opioid peptide was introduced as a highly active analgesic because it elicited a strong dose- and time-dependent antinociceptive response when administered centrally and peripherally. Its pain-relieving activity was observed as rapidly as 5 min after drug injection. Such promising results led us to perform further studies, such as determining the resistance to enzymatic degradation, which resulted in obtaining a very stable opioid pharmacore PK20 metabolite. Methods: The synthesis of PK20 and its N-terminal tetrapeptide fragment has been accomplished using solid phase peptide chemistry. The biological stability of peptides has been measured in human serum and analyzed by HPLC/MS. Peptides were pharmacologically characterized in in vitro MOP and DOP receptor binding as well as [S-35]GTP gamma S receptor binding assays. Antinociceptive properties of compounds were measured by in vivo assays in C(57)Bl(6) mice after intravenous or intrathecal applications. Results: Dmt-D-Lys-Phe-Phe-OH (PK20M), an N-terminal tetrapeptide metabolite of the opioid-neurotensin hybrid peptide PK20, is characterized by a long duration of action, as demonstrated by a preserved, long-lasting analgesic effect even 2 h post-injection (average % MPE = 69.33). In rat brain membranes, PK20M efficiently displaced both the MOP and DOP receptor selective radio-probes [H-3]DAMGO and [H-3]DIDI (pK(i) of 9.52 and 7.86, respectively) and potently stimulated [S-35]GTP gamma S binding, proving full agonism at both receptor types. In the [S-35]GTP gamma S assay, which measured the agonist-mediated G protein activation, PK20M together with PK20 and Met-enkephalin were potent stimulators of the regulatory G proteins. The relative affinities of PK20M for the mu and delta receptor subtypes revealed mu-receptor selectivity. Conclusion: The novel MOP receptor selective metabolite has been shown to possess opioid subtype receptor selectivity, high potency, and effective analgesic activities as measured in various bioassays.
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