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Victor J. Hruby - One of the best experts on this subject based on the ideXlab platform.
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Delta opioid receptor selective ligands; DPLPE-deltorphin chimeric peptide analogues.
International Journal of Peptide and Protein Research, 2009Co-Authors: Aleksandra Misicka, Frank Porreca, Andrzej W. Lipkowski, Robert Horvath, Peg Davis, Henry I. Yamamura, Victor J. HrubyAbstract:: Further efforts to correlate the topography of the bioactive structures of DPDPE and the deltorphins, two delta-opioid receptor active peptide families, are reported. A number of DPLPE-deltorphin chimeric peptides have been synthesized in which the C-terminal dipeptide delta-address of the deltorphins (-Val-GlyNH2, -Nle-GlyNH2) have been linked to the highly delta-opioid selective cyclic peptides DPDPE or DPLPE. These studies demonstrate that a major structural feature determining high potency of hybrid analogues is the chirality of the amino acid residue in position 5. The radioligand binding assays have revealed a decrease in potency (compared to DPDPE) at delta-receptors when the C-terminal dipeptides were added to DPDPE. On the other hand, chimeric peptides of DPLPE with these same C-terminal dipeptides retained high delta-selectivity and affinity. Similar results were obtained using the mouse vas deferens (MVD) and guinea pig ileum (GPI) bioassays. The importance of the hydrophilicity of amino acids in positions 2 and 5 for delta-selectivity is consistent with the previous finding for DPLPE and DPDPE. On the other hand, the replacement of phenylalanine-4 with p-chlorophenylalanine-4 did not increase delta-selectivity as in DPDPE. These findings suggest that the delta-receptor interacts with hybridized enkephalins and deltorphins somewhat differently than with DPDPE.
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para substituted phenylalanine 4 analogues of l ala3 DPDPE highly selective δ opioid receptor ligands
Journal of Peptide Research, 2009Co-Authors: Ronald C Haaseth, Frank Porreca, Peg Davis, Henry I. Yamamura, Teresa Zalewska, Victor J. HrubyAbstract:Several para-substituted Phe4 analogues of the δ1-selective antagonist [l-Ala3]. DPDPE (DPADPE) were prepared and evaluated for their brain-binding and in vitro pharmacological effects. Unlike the p-haloPhe4 analogues of DPDPE and the deltorphins, similar analogues of DPADPE with electron-withdrawing groups substituted at the para-position of the Phe4 aromatic ring did not all have increased potency and selectivity for δ opioid receptors, but all retained high potency and selectivity for δ opioid receptors greater than DPDPE. © Munksgaard 1997.
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solution conformations of the peptide backbone for DPDPE and its β mephe4 substituted analogs
International Journal of Peptide and Protein Research, 2009Co-Authors: Gregory V Nikiforovich, Catherine A. Gehrig, Om Prakash, Victor J. HrubyAbstract:: The solution structures of DPDPE, a conformationally restricted pentapeptide with the sequence H-Tyr1-D-Pen2-Gly3-Phe4-D-Pen5-OH, and its four beta-MePhe4-substituted analogs were examined by a combined approach including the NMR measurements in DMSO and water as well as independent energy calculations. It was concluded that several low energy conformers of DPDPE backbone satisfy the NMR data obtained in this study as well as in previous studies by other authors. These possible solution conformers of DPDPE in both DMSO and water share virtually the same type of cyclic backbone structure, with the Gly3 residue in a conformation close to a gamma-turn, and the Phe4 residue in a conformation close to alpha-helical torsion angles. They differ in the space arrangements of the flexible Tyr1 moiety. The solution structures of the beta-MePhe4-substituted analogs of DPDPE are interesting. For analogs with an S-configuration at the C alpha atom in the Phe4 residue, the cyclic backbone conformations resemble those of DPDPE itself, whereas for analogs with an R-configuration at the C alpha atom, the backbone conformation is somewhat different. This observation is in line with the high biological potencies and selectivities displayed by the former compounds but not by the latter ones. It was noted also that as far as the peptide backbone conformers are concerned, some of the possible DPDPE conformers in water are similar to the previously suggested model for the delta-receptor-bound conformation of DPDPE, becoming virtually identical to this conformation by rotating the side chains of the Tyr1 and the Phe4 residues.
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Passage of a δ-opioid receptor selective enkephalin, [D-penicillamine2,5]enkephalin, across the blood-brain and the blood-cerebrospinal fluid barriers
Journal of Neurochemistry, 2002Co-Authors: Sarah A. Williams, Victor J. Hruby, Thomas J. Abbruscato, Thomas P DavisAbstract:[D-Penicillamine 2,5 ]enkephalin (DPDPE) is an enzymatically stable, δ-opioid receptor-selective peptide, which produces analgesia when given intracerebroventricularly. However, because only modest analgesic effects were seen after subcutaneous administration of DPDPE, it has been inferred that it does not cross the blood-brain barrier well. In this present study, a vascular brain perfusion technique in anesthetized rats was used to measure directly whether [ 3 H]DPDPE could cross the blood-brain and/or the blood-CSF barriers. The results indicated that the brain uptake of [ 3 H]DPDPE was significantly greater than that of [ 14 C]sucrose, a vascular marker (p < 0.01), and than that of [ 3 H]DPDPE into the CSF (p < 0.01). Furthermore, HPLC analysis confirmed the integrity of the 3 H to DPDPE and demonstrated that intact [ 3 H]DPDPE entered the brain. Although 1 mM leucine-enkephalin failed to inhibit uptake of [ 3 H]DPDPE, unlabeled DPDPE (100 μM) caused a significant inhibition of the brain uptake (p < 0.01) but not the CSF uptake of [ 3 H]DPDPE. These data provide evidence that intact [ 3 H]DPDPE enters the CNS of anesthetized rats by saturable and nonsaturable mechanisms. In addition, the saturable mechanism is likely to be found at the blood-brain barrier, with the blood-CSF barrier playing only a minor role in the brain uptake of this peptide.
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Assessment of stereoselectivity of trimethylphenylalanine analogues of δ-opioid [D-pen2,D-pen5]-enkephalin
Journal of Neurochemistry, 2001Co-Authors: Ken A Witt, Henry I. Yamamura, Victor J. Hruby, Jason D Huber, Richard D Egleton, Cheryl A. Slate, Thomas P DavisAbstract:: [D-Pen(2),D-Pen(5)]-Enkephalin (DPDPE) is an enzymatically stable delta-opioid receptor-selective peptide, which was modified by the trimethylation of the Phe(4) residue to give beta-methyl-2', 6'-dimethylphenylalanine (TMP), resulting in four conformations : (2R,3S)-beta-Phe-DPDPE, (2R,3R)-beta-Phe-DPDPE, (2R, 3S)-beta-Phe-DPDPE, and (2S,3R)-beta-Phe-DPDPE. Synthesis was by solid-phase techniques using enantiomerically pure amino acids to give the four optically pure diastereoisomer peptides. The potency and selectivity (delta- versus mu-opioid receptor) were evaluated by radioreceptor binding in rat brain, with a mu/delta ratio decrease for all TMP conformations, compared with the parent compound (DPDPE). Octanol/buffer distribution analysis showed enhanced lipophilicity of all TMP forms, with a sixfold enhancement associated with (2S,3S)-TMP. In situ vascular perfusion in anesthetized rats showed a 1.6-fold (p < 0.01) increase in the ratio of brain uptake for (2S,3S)-TMP and a 1.5-fold (p < 0.01) decrease in uptake for (2R,3R)-TMP. Saturability of (2S,3S)-TMP was shown (p < 0.01) against 100 microM unlabeled DPDPE, showing a shared nondiffusionary transport system. P-glycoprotein affinity was shown in situ for the parent and (2S,3S)-TMP (p < 0.01). Protein binding capacity of the TMP compounds in rat plasma and in situ mammalian bovine serum albumin-Ringer showed (2R,3S)-TMP and (2S,3R)-TMP with the lowest degree of protein binding (p < 0.01), and (2S,3S)-TMP and (2R,3R)-TMP with comparable affinities to DPDPE. Analgesia, via intravenous administration, showed significantly reduced (p < 0.01) end effect and time course for (2R,3R)-TMP, (2R,3S)-TMP, and (2S, 3R)-TMP as compared with DPDPE. These results demonstrate that topographical modification in a conformationally restricted peptide can significantly modulate potency and receptor selectivity, binding capacity, enzymatic stability, lipophilicity, P-glycoprotein affinity, and blood-brain barrier permeability, resulting in a change of bioavailability, and thereby provides insight for future peptide drug design.
Gerald A Young - One of the best experts on this subject based on the ideXlab platform.
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mu delta opioid interactions iii differential antagonism of DPDPE induced increases in morphine eeg and eeg power spectra by dalce and naltrindole
Peptides, 1993Co-Authors: Helen Stamidis, Gerald A YoungAbstract:Abstract In the present study, the effects of DALCE ([ d -Ala 2 ,Leu 5 ,Cys 6 ]enkephalin) and naltrindole on DPDPE ([ d -Pen 2 , d -Pen 5 ]enkephalin)-induced increases in morphine EEG and EEG power spectra were assessed. Adult female Sprague-Dawley rats were implanted with cortical EEG electrodes and permanent indwelling ICV and IV cannulae. Rats were pretreated with ICV DALCE at 15.7 nmol, ICV naltrindole at 20 nmol, or ICV sterile water. Rats were then administered ICV DPDPE at 2.5 nmol or ICV sterile water followed, 10 min later, by IV morphine at 3 mg/kg. Morphine-induced changes in EEG global (1–50 Hz) spectral parameters, the duration of morphine-induced high-voltage EEG bursts, the duration of EEG and behavioral excitation, and the latency to onset of slow-wave sleep were assessed. The DALCE pretreatment significantly decreased morphine-induced total spectral power seen in the DPDPE + morphine group. The DALCE pretreatment reversed the effects of DPDPE on the duration of morphine-induced EEG bursts and the duration of EEG and behavioral excitation. The ICV naltrindole, however, had no significant effect on DPDPE-induced increases in morphine EEG, EEG spectral parameters, and behavior. These data, therefore, suggest that DPDPE may be increasing the effects of morphine on EEG through DALCE-sensitive delta opioid receptors associated within the mu-delta opioid receptor complex.
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mu delta opioid interactions ii β fna inhibits DPDPE induced increases in morphine eeg and eeg spectral power
Peptides, 1992Co-Authors: Helen Stamidis, Gerald A YoungAbstract:Abstract In the present study, the effects of β-FNA on DPDPE-induced increases in morphine EEG and EEG power spectra were assessed. Adult female Sprague-Dawley rats were implanted with cortical EEG electrodes and permanent indwelling ICV and IV cannulae. Rats were administered ICV β-FNA at 20 nmol or ICV sterile water. Then 18–24 h later, rats were administered ICV DPDPE at 2.5 nmol or ICV sterile water followed, 10 min later, by IV morphine at 3 mg/kg. Morphine-induced changes in EEG global (1–50 Hz) spectral parameters, the duration of morphine-induced high voltage EEG bursts, the period of EEG and behavioral excitation, and the latency to onset of slow-wave sleep were statistically analyzed using a one-way analysis of variance. β-FNA pretreatment significantly decreased morphine-induced total spectral power seen in the DPDPE + morphine group. β-FNA pretreatment also significantly decreased the duration of morphine-induced EEG bursts, the period of EEG and behavioral excitation, and the latency to onset of slow-wave sleep in the DPDPE + morphine group. These data, therefore, suggest that DPDPE may be increasing the effects of morphine on EEG through delta opioid receptors associated within the mu-delta opioid receptor complex.
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mu delta opioid interactions i the delta peptide DPDPE increases morphine induced eeg and eeg spectral power
Peptides, 1992Co-Authors: Helen Stamidis, Gerald A YoungAbstract:Abstract The effects of the selective delta peptide, DPDPE ([2- d -penicillamine,5- d -penicillamine]enkephalin), on morphine-induced EEG and EEG power spectra were assessed. Adult female Sprague-Dawley rats were implanted with cortical EEG electrodes and permanent indwelling ICV and IV cannulae. Rats were pretreated with ICV sterile water or ICV DPDPE at 1.25, 2.5, or 5.0 nmol, 10 min before receiving IV morphine at 3 mg/kg. The treatments produced high voltage EEG bursts and associated behavioral stupor. The EEG data were further analyzed on a Pathfinder II computer and statistical analysis of EEG spectral parameters was performed by using a one-way ANOVA followed by Tukey's HSD. ICV DPDPE at 2.5 nmol and 5.0 nmol significantly increased EEG absolute global spectral power. Furthermore, ICV DPDPE at 2.5 and 5.0 nmol significantly increased the duration of morphine-induced high voltage EEG bursts. These data further indicate an in vivo interaction between mu and delta opioid receptors.
Laura S Stone - One of the best experts on this subject based on the ideXlab platform.
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DPDPE-UK14,304 synergy is retained in mu opioid receptor knockout mice.
Pain, 2020Co-Authors: Carolyn A Fairbanks, Laura S StoneAbstract:When agonists to alpha(2)adrenergic receptor (AR) and delta opioid receptor (DOR) are co-administered, they act synergistically to inhibit nociceptive elicited behavior. Some previous studies of synergism have used the DOR-selective agonist [D-Pen(2),D-Pen(5)]-enkehphalin (DPDPE), however, DPDPE has been shown to be less potent in mu opioid receptor-knockout (MOR-KO) mice. It is possible, therefore, that MOR contributes to the synergism of DPDPE with the alpha(2)AR agonists. We compared the interactions of spinally administered DPDPE with an alpha(2)AR-adrenergic agonist in MOR-KO and MOR-wildtype (WT) mice. In these mice, morphine is ineffective and the potency of spinally administered DOR agonists, deltorphin II (DELT II) and DPDPE decreased 16- and 250-fold, respectively. Antagonism studies using the MOR-selective antagonist, D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Phe-Thr-NH(2) (CTOP) and the DOR-selective antagonist, naltrindole HCl (naltrindole) demonstrated that while DOR mediates DPDPE-induced antinociception in MOR-KO, both MOR and DOR participate in DPDPE antinociception in WT mice, suggesting that DPDPE is less selective for DOR than previously observed in binding studies when given in vivo. The potency of the alpha(2)AR agonist UK14,304 was equivalent in WT and MOR-KO, demonstrating that the loss of opioid-mediated antinociception in the MOR-KO was not due to generalized impairment of antinociceptive processing. Interestingly, isobolographic analysis showed that, despite substantial loss of DPDPE potency in MOR-KO, DPDPE-UK14,304 synergism is fully retained. Collectively, these experiments demonstrate that although MOR participates in DELT II- and DPDPE-mediated spinal antinociception, DOR independently participates in synergistic antinociception with alpha(2)AR. Resolution of the roles of the opioid receptor subtypes in opioid agonist-induced effects may require comparison of the effects of multiple selective agonists in knockout animals.
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DPDPE uk14 304 synergy is retained in mu opioid receptor knockout mice
Pain, 2003Co-Authors: Carolyn A Fairbanks, Laura S StoneAbstract:Abstract When agonists to α2adrenergic receptor (AR) and δ opioid receptor (DOR) are co-administered, they act synergistically to inhibit nociceptive elicited behavior. Some previous studies of synergism have used the DOR-selective agonist [ d -Pen2, d -Pen5]-enkehphalin (DPDPE), however, DPDPE has been shown to be less potent in μ opioid receptor-knockout (MOR-KO) mice. It is possible, therefore, that MOR contributes to the synergism of DPDPE with the α2AR agonists. We compared the interactions of spinally administered DPDPE with an α2AR-adrenergic agonist in MOR-KO and MOR-wildtype (WT) mice. In these mice, morphine is ineffective and the potency of spinally administered DOR agonists, deltorphin II (DELT II) and DPDPE decreased 16- and 250-fold, respectively. Antagonism studies using the MOR-selective antagonist, d -Phe-Cys-Tyr- d -Trp-Orn-Thr-Phe-Thr-NH2 (CTOP) and the DOR-selective antagonist, naltrindole HCl (naltrindole) demonstrated that while DOR mediates DPDPE-induced antinociception in MOR-KO, both MOR and DOR participate in DPDPE antinociception in WT mice, suggesting that DPDPE is less selective for DOR than previously observed in binding studies when given in vivo. The potency of the α2AR agonist UK14,304 was equivalent in WT and MOR-KO, demonstrating that the loss of opioid-mediated antinociception in the MOR-KO was not due to generalized impairment of antinociceptive processing. Interestingly, isobolographic analysis showed that, despite substantial loss of DPDPE potency in MOR-KO, DPDPE-UK14,304 synergism is fully retained. Collectively, these experiments demonstrate that although MOR participates in DELT II- and DPDPE-mediated spinal antinociception, DOR independently participates in synergistic antinociception with α2AR. Resolution of the roles of the opioid receptor subtypes in opioid agonist-induced effects may require comparison of the effects of multiple selective agonists in knockout animals.
Claes Wahlestedt - One of the best experts on this subject based on the ideXlab platform.
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supraspinal antinociceptive response to d pen2 5 enkephalin DPDPE is pharmacologically distinct from that to other δ agonists in the rat
Journal of Pharmacology and Experimental Therapeutics, 2000Co-Authors: Graeme Fraser, Amynah A Pradhan, Paul B S Clarke, Claes WahlestedtAbstract:The cloned δ-opioid receptor (DOR) is being investigated as a potential target for novel analgesics with an improved safety profile over μ-opioid receptor agonists such as morphine. The current study used antisense techniques to evaluate the role of DOR in mediating supraspinal antinociception in rats. All of the opioid agonists tested (δ-selective: deltorphin II, DPDPE, pCl-DPDPE, SNC80; μ-selective: DAMGO; i.c.v . ) provided significant, dose-dependent antinociception in the paw pressure assay. Administration of a phosphodiester antisense oligonucleotide (i.c.v . ) targeted against DOR inhibited antinociception in response to SNC80, deltorphin II, and pCl-DPDPE compared with mismatch and saline-treated controls. However, antisense treatment did not inhibit the response to DPDPE or DAMGO. In contrast, the highly selective μ-antagonist CTOP blocked antinociception in response to ED 80 concentrations of DAMGO and DPDPE, reduced the response to pCl-DPDPE, and did not alter the response to deltorphin II or SNC80. In total, these data suggest that DOR mediates the antinociceptive response to deltorphin II, SNC80, and pCl-DPDPE at supraspinal sites and further demonstrates that the DOR-mediated response to deltorphin II and SNC80 is independent of μ-receptor activation. Conversely, supraspinal antinociception in response to DPDPE is mediated by a receptor distinct from DOR; this response is directly or indirectly sensitive to μ-receptor blockade. The distinct pharmacological profile of DPDPE suggests that either this prototypical δ-agonist mediates antinociception by a direct, nonselective interaction at μ-receptors or DPDPE interacts with a novel δ-subtype that, in turn, indirectly activates μ-receptors in the brain.
Thomas P Davis - One of the best experts on this subject based on the ideXlab platform.
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Passage of a δ-opioid receptor selective enkephalin, [D-penicillamine2,5]enkephalin, across the blood-brain and the blood-cerebrospinal fluid barriers
Journal of Neurochemistry, 2002Co-Authors: Sarah A. Williams, Victor J. Hruby, Thomas J. Abbruscato, Thomas P DavisAbstract:[D-Penicillamine 2,5 ]enkephalin (DPDPE) is an enzymatically stable, δ-opioid receptor-selective peptide, which produces analgesia when given intracerebroventricularly. However, because only modest analgesic effects were seen after subcutaneous administration of DPDPE, it has been inferred that it does not cross the blood-brain barrier well. In this present study, a vascular brain perfusion technique in anesthetized rats was used to measure directly whether [ 3 H]DPDPE could cross the blood-brain and/or the blood-CSF barriers. The results indicated that the brain uptake of [ 3 H]DPDPE was significantly greater than that of [ 14 C]sucrose, a vascular marker (p < 0.01), and than that of [ 3 H]DPDPE into the CSF (p < 0.01). Furthermore, HPLC analysis confirmed the integrity of the 3 H to DPDPE and demonstrated that intact [ 3 H]DPDPE entered the brain. Although 1 mM leucine-enkephalin failed to inhibit uptake of [ 3 H]DPDPE, unlabeled DPDPE (100 μM) caused a significant inhibition of the brain uptake (p < 0.01) but not the CSF uptake of [ 3 H]DPDPE. These data provide evidence that intact [ 3 H]DPDPE enters the CNS of anesthetized rats by saturable and nonsaturable mechanisms. In addition, the saturable mechanism is likely to be found at the blood-brain barrier, with the blood-CSF barrier playing only a minor role in the brain uptake of this peptide.
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Pluronic P85 block copolymer enhances opioid peptide analgesia
Journal of Pharmacology and Experimental Therapeutics, 2002Co-Authors: Ken A Witt, Jason D Huber, Richard D Egleton, Thomas P DavisAbstract:Peptide-based drug development is a rapidly growing field within pharmaceutical research. Nevertheless, peptides have found limited clinical use due to several physiological and pathological factors. Pluronic block copolymers represent a growing technology with the potential to enhance efficacy of peptide therapeutics. This investigation assesses Pluronic P85 (P85) and its potential to enhance opioid peptide analgesia. Two opioid peptides, [d-Pen 2 ,d-Pen 5 ]-enkephalin (DPDPE) and biphalin, were examined as to the benefits of P85 coadministration, above (1.0%) and below (0.01%) the critical micelle concentration, with morphine as a nonpeptide control. P85 was examined in vitro to assess blood-brain barrier uptake in association with P-glycoprotein effect, DPDPE and morphine being P-glycoprotein substrates. P85 coadministration with DPDPE and biphalin showed increased ( p p 125 I-DPDPE and [ 3 H]morphine at 0.01% ( p p p 125 I-DPDPE and [ 3 H]morphine increased cellular uptake ( p p 125 I-DPDPE and [ 3 H]morphine coadministered with 0.01% P85 and cyclosporin-A increased cellular uptake compared with control ( p p p 125 I-DPDPE and [ 3 H]morphine uptake. In our examination, we determined that P85 enhanced the analgesic profile of biphalin, DPDPE, and morphine, both above and below the critical micelle concentration.
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Assessment of stereoselectivity of trimethylphenylalanine analogues of δ-opioid [D-pen2,D-pen5]-enkephalin
Journal of Neurochemistry, 2001Co-Authors: Ken A Witt, Henry I. Yamamura, Victor J. Hruby, Jason D Huber, Richard D Egleton, Cheryl A. Slate, Thomas P DavisAbstract:: [D-Pen(2),D-Pen(5)]-Enkephalin (DPDPE) is an enzymatically stable delta-opioid receptor-selective peptide, which was modified by the trimethylation of the Phe(4) residue to give beta-methyl-2', 6'-dimethylphenylalanine (TMP), resulting in four conformations : (2R,3S)-beta-Phe-DPDPE, (2R,3R)-beta-Phe-DPDPE, (2R, 3S)-beta-Phe-DPDPE, and (2S,3R)-beta-Phe-DPDPE. Synthesis was by solid-phase techniques using enantiomerically pure amino acids to give the four optically pure diastereoisomer peptides. The potency and selectivity (delta- versus mu-opioid receptor) were evaluated by radioreceptor binding in rat brain, with a mu/delta ratio decrease for all TMP conformations, compared with the parent compound (DPDPE). Octanol/buffer distribution analysis showed enhanced lipophilicity of all TMP forms, with a sixfold enhancement associated with (2S,3S)-TMP. In situ vascular perfusion in anesthetized rats showed a 1.6-fold (p < 0.01) increase in the ratio of brain uptake for (2S,3S)-TMP and a 1.5-fold (p < 0.01) decrease in uptake for (2R,3R)-TMP. Saturability of (2S,3S)-TMP was shown (p < 0.01) against 100 microM unlabeled DPDPE, showing a shared nondiffusionary transport system. P-glycoprotein affinity was shown in situ for the parent and (2S,3S)-TMP (p < 0.01). Protein binding capacity of the TMP compounds in rat plasma and in situ mammalian bovine serum albumin-Ringer showed (2R,3S)-TMP and (2S,3R)-TMP with the lowest degree of protein binding (p < 0.01), and (2S,3S)-TMP and (2R,3R)-TMP with comparable affinities to DPDPE. Analgesia, via intravenous administration, showed significantly reduced (p < 0.01) end effect and time course for (2R,3R)-TMP, (2R,3S)-TMP, and (2S, 3R)-TMP as compared with DPDPE. These results demonstrate that topographical modification in a conformationally restricted peptide can significantly modulate potency and receptor selectivity, binding capacity, enzymatic stability, lipophilicity, P-glycoprotein affinity, and blood-brain barrier permeability, resulting in a change of bioavailability, and thereby provides insight for future peptide drug design.
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pharmacodynamic and pharmacokinetic characterization of poly ethylene glycol conjugation to met enkephalin analog d pen2 d pen5 enkephalin DPDPE
Journal of Pharmacology and Experimental Therapeutics, 2001Co-Authors: Ken A Witt, Henry I. Yamamura, Jason D Huber, Richard D Egleton, Michael James Roberts, Michael D Bentley, Thomas P DavisAbstract:Poly(ethylene glycol), or PEG, conjugation to proteins and peptides is a growing technology used to enhance efficacy of therapeutics. This investigation assesses pharmacodynamic and pharmacokinetic characteristics of PEG-conjugated [d-Pen2,d-Pen5]-enkephalin (DPDPE), a met-enkephalin analog, in rodent (in vivo, in situ) and bovine (in vitro) systems. PEG-DPDPE showed increased analgesia (i.v.) compared with nonconjugated form ( p < 0.01), despite a 172-fold lower binding affinity for the δ-opioid receptor. [125I]PEG-DPDPE had a 36-fold greater hydrophilicity ( p < 0.01) and 12% increase in the unbound plasma protein fraction ( p < 0.01), compared with [125I]DPDPE. [125I]PEG-DPDPE had a 2.5-fold increase in elimination half-life ( p < 0.01), 2.7-fold decrease in volume of distribution ( p < 0.01), and a 7-fold decrease in plasma clearance rate ( p < 0.01) to [125I]DPDPE. Time course distribution showed significant concentration differences ( p < 0.01) in plasma, whole blood, liver, gallbladder, gastrointestinal (GI) content, GI tract, kidneys, spleen, urine, and brain (brain, p < 0.05), between the conjugated and nonconjugated forms. Increased brain uptake of [125I]PEG-DPDPE corresponded to analgesia data. [125I]PEG-DPDPE in brain was shown to be 58.9% intact, with 41.1% existing as [125I]DPDPE (metabolite), whereas [125I]DPDPE was 25.7% intact in the brain (at 30 min). In vitro P-glycoprotein affinity was shown for [125I]DPDPE ( p < 0.01) but not shown for [125I]PEG-DPDPE. In vitro saturable uptake, with 100 μM DPDPE, was shown for [125I]PEG-DPDPE ( p < 0.05). In this study, PEG-conjugated DPDPE seems to act as a prodrug, enhancing peripheral pharmacokinetics, while undergoing hydrolysis in the brain and allowing nonconjugated DPDPE to act at the receptor.
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Insulin enhancement of opioid peptide transport across the blood-brain barrier and assessment of analgesic effect.
Journal of Pharmacology and Experimental Therapeutics, 2000Co-Authors: Ken A Witt, Jason D Huber, Richard D Egleton, Thomas P DavisAbstract:Insulin crosses the blood-brain barrier (BBB) via receptor-mediated transcytosis and has been suggested to augment uptake of peripheral substances across the BBB. The δ-opioid receptor-selective peptided-penicillamine2,5 (DPDPE), a Met-enkephalin analog, produces analgesia via a central nervous system-derived effect. In vitro ( K cell, μl · min−1 · mg−1) and in situ ( K in, μl · min−1· g−1) analyses of DPDPE transport ( K cell = 0.56 ± 0.15; K in = 0.28 ± 0.03) revealed significant ( P < .01) increases in DPDPE uptake by the BBB with 10 μM insulin ( K cell = 1.61 ± 0.25; K in = 0.48 ± 0.04). In vitro cellular uptake was significantly increased ( P < .05) at 1 μM insulin, whereas no significant uptake was observed with CTAP (a somatostatin opioid peptide analog) or sucrose (a paracellular diffusionary marker). No significant change in uptake was seen with DPDPE, CTAP, or sucrose in the presence of holo-transferrin (0–100 μM), indicating that the effect of insulin on DPDPE was not a generalized effect of receptor endocytosis. Insulin did not affect P-glycoprotein efflux, a mechanism that has shown affinity for DPDPE. A similar uptake of DPDPE into the brain (64% increase) was seen with the in situ brain perfusion model. Analgesic assessment revealed a significant decline in DPDPE (i.v.)-induced analgesia with increasing concentrations of insulin (i.v., i.c.v., s.c.) in a dose-dependent manner. Thus, insulin significantly increases DPDPE uptake across the BBB by a specific mechanism. The analgesic effect seen with DPDPE and insulin coadministration was shown to decrease, indicating that insulin reduces the analgesic effect within the central nervous system rather than at the BBB.