Bivalent Mu-Delta Opioid Ligands: Potential for Pain Control Without Tolerance or Dependence
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Transcript of Bivalent Mu-Delta Opioid Ligands: Potential for Pain Control Without Tolerance or Dependence
Bivalent Mu-Delta Opioid Ligands: Potential for Bivalent Mu-Delta Opioid Ligands: Potential for Pain Control Without Tolerance or DependencePain Control Without Tolerance or Dependence
David J DanielsDavid J DanielsDepartment of Medicinal ChemistryDepartment of Medicinal ChemistryUniversity of MinnesotaUniversity of Minnesota
Opioid AnalgesicsOpioid Analgesics
From the poppy Papaver SomniferumFrom the poppy Papaver Somniferum One of the oldest recorded medicationsOne of the oldest recorded medications Long history of use and abuseLong history of use and abuse
Morphine: Isolated from opium poppy seeds in Morphine: Isolated from opium poppy seeds in the early 1800s. the early 1800s.
Morpheus (latin) = Greek God of DreamsMorpheus (latin) = Greek God of Dreams
HO O OH
NH
CH3
(-)-MorphineOpium
Diacetylmorphine - HeroinDiacetylmorphine - Heroin
11stst Synthesized in 1874 by C.R. Alder Wright Synthesized in 1874 by C.R. Alder Wright 1897 - Bayer Pharmaceutical company 1897 - Bayer Pharmaceutical company
“re-invented” diacetylmorphine“re-invented” diacetylmorphine named it named it HeroinHeroin after the German word after the German word
heroisch heroisch “heroic” “heroic” 1898 – 1910 Bayer marketed Heroin as a 1898 – 1910 Bayer marketed Heroin as a
non-addictive morphine substitute non-addictive morphine substitute 1914 Harrison Narcotics Tax Act1914 Harrison Narcotics Tax Act 1924 US banned all use of Heroin1924 US banned all use of Heroin
Opioid Analgesics – The Holy GrailOpioid Analgesics – The Holy Grail Countless synthetic opiate compounds have Countless synthetic opiate compounds have
been synthesized over the last century in an been synthesized over the last century in an attempt to create the “perfect” analgesicattempt to create the “perfect” analgesic
History and Background
• G protein-coupled receptors• 3 Opioid Receptor Subtypes: Mu, Delta, Kappa• Approximately 60 % homology• 372 to 398 amino acid residues
Opioid Receptor Opioid Receptor Dimerization/Oligomerization?Dimerization/Oligomerization?
Opioid Receptor SubtypesOpioid Receptor Subtypes
Pharmacology predicts greater receptor subtypes than receptor cloning reveals!
Single gene-deleted mice lack all subtypes for deleted receptor
Mu1, Mu2, Mu3
Delta1, Delta2
Kappa1, Kappa2
GPCR DimerizationGPCR DimerizationContact DimerMonomer
4
6
25
7
3
1
1
23
4
5
67
4
6
25
7
3
1
1
23
4
5
67
4
6
25
7
3
1
4
6
25
7
3
1
1
23
4
5
674
6
2
7
3
1
1
2
3
4
5
67
+5
Heterodimer
4
6
25
7
3
1
1
2
3
4
5
67 4
6
25
7
3
1
1
23
4
5
67
+Homodimer
Oligomer
Opioid Receptor DimerizationOpioid Receptor Dimerization
Homodimers:Homodimers: Mu-MuMu-Mu Delta-DeltaDelta-Delta Kappa-KappaKappa-Kappa
Heterodimers:Heterodimers: Mu-DeltaMu-Delta Delta-KappaDelta-Kappa Mu-KappaMu-Kappa
Evidence for dimerization1. Pharmacological2. Coimmunoprecipitation3. BRET and FRET4. Confocal microscopy5. Atomic-force microscopy
Bivalent LigandsBivalent Ligands Pharmacological Probes Selective for Pharmacological Probes Selective for
Dimerized Opioid ReceptorsDimerized Opioid Receptors
1 2 3
4
1 2 3
4
1.066.8 ± 11.75
2.925.1 ± 5.229.434
18.93.9 ± 0.621.723
2.122.0 ± 5.014.012
2.430.2 ± 8.76.301
Rel potencybIC50 nMSpacer Lengtha ÅnCmpd no.
Table 1. Opioid Agonist Potencies in the GPI. aDistance from the -nitrogen of one oxymorphamine to the other -nitrogen. bBivalent potency ratio dived by monovalent potency ratio.
O HN
O O
HN
O
NH O
OH
N
OH
O
OH
NH
N
HOn n
Spacer Length
O
OH
NH
N
HO HN
O
ONH
O
CH3
5
1.066.8 ± 11.75
2.925.1 ± 5.229.434
18.93.9 ± 0.621.723
2.122.0 ± 5.014.012
2.430.2 ± 8.76.301
Rel potencybIC50 nMSpacer Lengtha ÅnCmpd no.
Table 1. Opioid Agonist Potencies in the GPI. aDistance from the -nitrogen of one oxymorphamine to the other -nitrogen. bBivalent potency ratio dived by monovalent potency ratio.
O HN
O O
HN
O
NH O
OH
N
OH
O
OH
NH
N
HOn n
Spacer Length
O
OH
NH
N
HO HN
O
ONH
O
CH3
5
0
5
10
15
20
5 15 25 35
Spacer Length (Å)
Rela
tive
pote
ncy
0
5
10
15
5 15 25 35
Spacer Length (Å)
Rela
tive
Affin
ity
Portoghese, P.S. et. al. Journal of Medicinal Chemistry, 1986, 29, 1855-1861
Purpose of our ProjectsPurpose of our Projects
Develop selective bivalent ligands to probe opioid Develop selective bivalent ligands to probe opioid receptor organizationreceptor organization
See if we can characterize putative opioid receptor See if we can characterize putative opioid receptor subtypessubtypes
Gain additional insight into the molecular Gain additional insight into the molecular organization of opioid receptorsorganization of opioid receptors
Develop novel analgesics devoid of tolerance Develop novel analgesics devoid of tolerance and physical dependenceand physical dependence
Early Evidence For Interactions Between Early Evidence For Interactions Between Mu and Delta Opioid ReceptorsMu and Delta Opioid Receptors
Potentiation of morphine with delta agonists Potentiation of morphine with delta agonists Vaught, J.; Takemori, A. Differential effects of leucine Vaught, J.; Takemori, A. Differential effects of leucine
enkephalin and methionine enkephalin on morphine-induced enkephalin and methionine enkephalin on morphine-induced analgesia, acute tolerance and dependence. analgesia, acute tolerance and dependence. J. Pharm. Exp. J. Pharm. Exp. Ther.Ther., , 19791979, 208, 86-94, 208, 86-94
Rothman,R.B.; Westfall,T.C. Allosteric coupling between Rothman,R.B.; Westfall,T.C. Allosteric coupling between morphine and enkephalin receptors in vitro. morphine and enkephalin receptors in vitro. Mol PharmacolMol Pharmacol., ., 19821982, 21, 548-557, 21, 548-557
Pharmacological Evidence for Tolerance Pharmacological Evidence for Tolerance and Dependenceand Dependence
Abdelhamid, E.E.; Sultana, M.; Portoghese, P.S.; Takemori, Abdelhamid, E.E.; Sultana, M.; Portoghese, P.S.; Takemori, A.E. Selective blockage of delta opioid receptors prevents A.E. Selective blockage of delta opioid receptors prevents the development of morphine tolerance and dependence in the development of morphine tolerance and dependence in mice. mice. J. Pharm. Exp. Ther.,J. Pharm. Exp. Ther., 19911991, 258, 299-303 , 258, 299-303
Zhu, Y.; King, M.A.; Schuller, A.G.; Nitsche, J.F.; Reidl, M.; Zhu, Y.; King, M.A.; Schuller, A.G.; Nitsche, J.F.; Reidl, M.; Elde, R.P.; Unterwald, E.; Pasternak, G.W.; Pintar, J.E. Elde, R.P.; Unterwald, E.; Pasternak, G.W.; Pintar, J.E. Retention of supraspinal morphine-like analgesia and loss of Retention of supraspinal morphine-like analgesia and loss of morphine tolerance in delta opioid receptor knockout mice. morphine tolerance in delta opioid receptor knockout mice. Neuron,Neuron, 1999 1999 24, 243-252 24, 243-252
Recent Evidence For Interactions Recent Evidence For Interactions Between Mu and Delta Opioid ReceptorsBetween Mu and Delta Opioid Receptors Cultured cellsCultured cells
George, S.R.; Fan, T.; Xie Z.; Tse R.; Tam, V.; Varghese, G.; O’Dowd, George, S.R.; Fan, T.; Xie Z.; Tse R.; Tam, V.; Varghese, G.; O’Dowd, B.F. Oligomerization of mu and delta-Opioid Receptors. B.F. Oligomerization of mu and delta-Opioid Receptors. Journal of Journal of Biological Chemistry, Biological Chemistry, 20002000,, 275, 26128-26135 275, 26128-26135
Gomes, I.; Jordan, B.A.; Gupta, A.; Trapaidze, N.; Nagy, V.; Devi, L.A. Gomes, I.; Jordan, B.A.; Gupta, A.; Trapaidze, N.; Nagy, V.; Devi, L.A. Heterodimerization of mu and delta Opioid Receptors: A Role in Opiate Heterodimerization of mu and delta Opioid Receptors: A Role in Opiate Synergy, Synergy, The Journal of NeuroscienceThe Journal of Neuroscience, , 20002000, 20, 1-5, 20, 1-5
Living Cells - BRETLiving Cells - BRET Mu luciferase and delta YFP cotransfected into living cellsMu luciferase and delta YFP cotransfected into living cells
Spinal cord membranesSpinal cord membranes Coimmunoprecipitation experiments with mouse abs to mouse mu and Coimmunoprecipitation experiments with mouse abs to mouse mu and
mouse delta opioid receptors in WT and delta “knockout” mice mouse delta opioid receptors in WT and delta “knockout” mice Gomes, I.; Gupta, A.; Filipovska, J.; Szeto, H. H.; Pintar, J. E.; Devi, L. A. Gomes, I.; Gupta, A.; Filipovska, J.; Szeto, H. H.; Pintar, J. E.; Devi, L. A. A role for heterodimerization of mu and delta opiate receptors in A role for heterodimerization of mu and delta opiate receptors in enhancing morphine analgesia. enhancing morphine analgesia. Proc. Natl. Acad. Sci. U.S.A. Proc. Natl. Acad. Sci. U.S.A. 20042004, 101, , 101, 5135-5139.5135-5139.
Mu-Delta Bivalent Ligand ProjectMu-Delta Bivalent Ligand Project
M D A N - #
MU
Delta
Agonist
Antagonist
# of Atoms ofthe spacer
Agonist
Antagonist
Daniels, D.J.; Lenard, N.R.; Etienne, C.L.; Law, P.Y.; Roerig, S.C.; Portoghese, P.S. Opioid-induced tolerance and dependence in mice is modulated by the distance between pharmacophores in a bivalent ligand series. Proc. Natl. Acad. Sci. U.S.A. 2005, 102, 19208-19213.
Mu Receptor Delta Receptor
Approximate Distance Between recognition sites = 17 Å
Model of a Mu-Delta Heterodimer
MDAN-19 through MDAN-21 Spacer Distance = 23 -26 Å
Mu-Delta Bivalent Ligand ProjectMu-Delta Bivalent Ligand Project
MDAN Seriesn = 2 - 7
MA Seriesn = 2 - 7
DN-20
Measure of analgesia/antinociceptionMeasure of analgesia/antinociception
Mouse Tail Flick Assay
Rat
Antinociceptive activity in the mouse tail-flick Antinociceptive activity in the mouse tail-flick assay after acute I.C.V administrationassay after acute I.C.V administration
10 100 10000
25
50
75
100
MA-16MA-17
MA-20MA-21
MA-19
Oxymorphone
Drug ED50 (95% CI) pmolOxy 43 (34-52)MA-16 39 (32-47)MA-17 40 (33-46)MA-19 40 (29-50)MA-20 37 (29-45)MA-21 43 (39-48)
Drug Dose (pmol)
0.01 0.1 1 100
25
50
75
100
MDAN-18MDAN-17
MDAN-19
MDAN-21MDAN-20
MDAN-16
Drug Dose (nmol)
Model for Testing Tolerance and Model for Testing Tolerance and Physical Dependence in MicePhysical Dependence in Mice
Osmotic minipump infuses constant rate of Osmotic minipump infuses constant rate of ligand via cannula i.c.v. for 3 daysligand via cannula i.c.v. for 3 days
Day 4, naloxone (1 mg/kg s.c.) injected; Day 4, naloxone (1 mg/kg s.c.) injected; the number of jumps in ten minutes is the number of jumps in ten minutes is recordedrecorded
4 hours laters: “chronic” ED50 value with 4 hours laters: “chronic” ED50 value with tail-flick assaytail-flick assay
Tolerance determined by comparing saline Tolerance determined by comparing saline infusion vs. chronic infusioninfusion vs. chronic infusion
Effect of Spacer Length on Effect of Spacer Length on Tolerance and Dependence in Mice Tolerance and Dependence in Mice
CompoundSaline ED50 nmol
(95% C.I.)Chronic ED50 nmol
(95% C.I.) Fold Diff # of Jumps
(SEM)
MDAN - 16 1.62 (1.35 – 1.89) 4.72 (3.47 – 5.91) 2.8 30 (23)
MDAN - 17 1.54 (0.89 – 2.20) 5.61 (4.39 – 6.83) 3.6 0.9 (0.7)
MDAN - 18 1.29 (0.97 – 1.61) 4.75 (3.50 – 6.00) 3.7 8.9 (3.0)
MDAN - 19 0.42 (0.37 – 0.47) 0.40 (0.33 – 0.47) 1.0 3.6 (1.7)
MDAN - 20 0.17 (0.15 – 0.20) 0.17 (0.13 – 0.21) 1.0 0.4 (0.4)
MDAN - 21 0.10 (0.09 – 0.11) 0.10 (0.09 – 0.11) 1.0 3.5 (1.7)
MA - 19 0.04 (0.03 – 0.05) 0.22 (0.19 – 0.26) 5.5 83 (13)
MA-19 + DN-19 0.04 (0.03 – 0.04) 0.03 (0.02 – 0.05) 9.4 29 (8)
Morphine 4.54 (3.51 – 5.56) 26.8 (20.8 – 32.8) 6.0 100 (15)
Tolerance Dependence
Model for the role of heterodimers in tolerance and dependence
Model for the role of heterodimers in tolerance and dependence
Tolerance &Dependence
CAMKII ??
Analgesia
Analgesia Analgesia
Opioid receptor organization may be responsible for separate mechanisms leading to tolerance, dependence, and other unwanted side effects
Conditioned Place PreferenceConditioned Place PreferenceA Model of AddictionA Model of Addiction
Adapted from Feldman, R.S.; Meyer, J.S. and Quenzer, L.F. Principles of NeuropsychopharmacologySunderland, MA, Sinauer Associates, 1997.
Lenard, N.R.; Daniels, D.J.; Portoghese, P.S.; Roerig, S.C. Absence of conditioned placepreference or reinstatement with bivalent ligands containing mu-opioid agonist and delta-opioid receptor antagonist pharmacophores. Eur. J. Pharmacol. 2007, Accepted and in press.
Acquisition of Place PreferenceAcquisition of Place Preference Day 1: Exposure to novel environment (15 min)Day 1: Exposure to novel environment (15 min) Day 2: PreconditioningDay 2: Preconditioning
The time spent in each side of the box in 15 min recordedThe time spent in each side of the box in 15 min recorded Days 3 – 5: Injected with saline and confined to one Days 3 – 5: Injected with saline and confined to one
side of box (30 min); Later injected with ligand and side of box (30 min); Later injected with ligand and confined to other side of box (30 min)confined to other side of box (30 min)
Day 6: Determine place preference (15 min)Day 6: Determine place preference (15 min) Place Preference = Percent change in time spent on Place Preference = Percent change in time spent on
drug-paired sidedrug-paired side Positive = Place PreferencePositive = Place Preference Negative = Place AversionNegative = Place Aversion
MA-19MA-19 + DN-20
DN-20Saline
Saline
MDAN-16
MDAN-19
MDAN-21
ConditionedPlace PreferenceResults
MonovalentLigands
BivalentLigands
Primining injection-induced Primining injection-induced reinstatement of morphine CPPreinstatement of morphine CPP
Parenteral BioavailabilityParenteral Bioavailability
Comparison of Intravenous to Intracerebral Ventricular Administration Potencies for MDAN-21, MA-19 and Morphine
Ligandi.c.v. ED50 (95%
C.I.) nmoli.v. ED50 (95% C.I.)
nmoli.v. / i.c.v.
ratio
MA-19 0.04 (0.03 – 0.05) 1.61 (1.29 – 1.92) 40.3MDAN-21 0.08 (0.06 – 0.10) 3.3 (3.0 – 3.6) 41.3Morphine 4.1 (3.7 – 4.8) 168 (146 – 178) 41.0
Project SummaryProject Summary
Clinical ImplicationsClinical Implications Potent, efficacious analgesicsPotent, efficacious analgesics No toleranceNo tolerance No physical dependenceNo physical dependence No drug seeking behaviorNo drug seeking behavior
Molecular ImplicationsMolecular Implications Tolerance and dependence mediated through Tolerance and dependence mediated through
associated mu-delta opioid receptors??associated mu-delta opioid receptors??
Future DirectionsFuture Directions University of Minnesota is in the process of University of Minnesota is in the process of
patenting the MDAN seriespatenting the MDAN series Several pharmaceutical companies have Several pharmaceutical companies have
expressed interestexpressed interest
Other laboratories are in the process of Other laboratories are in the process of designing compounds with mixed designing compounds with mixed -agonist/-agonist/--antagontist activityantagontist activity
AcknowledgementsAcknowledgements
Philip PortoghesePhilip Portoghese Sandy RoerigSandy Roerig
Natalie LenardNatalie Lenard Ping LawPing Law Chris EtienneChris Etienne
1. Work Supported by NIH project Grant DA15092. NIDA Postdoctoral Fellowship DA18028 (to N.L.)
Funding:
Distance between recognition sites in Mu-Mu Distance between recognition sites in Mu-Mu opioid receptor models using two TM helices as opioid receptor models using two TM helices as
the interface between dimersthe interface between dimersDimer Interface Distance Å Dimer Interface Distance Å
TM1,2 – TM2,1 41.6 TM3,4 –TM5,4 37.5
TM1,2 – TM3,2 41.0 TM3,4 –TM6,5 29.5
TM1,2 – TM4,3 38.6 TM3,4 –TM7,6 26.8
TM1,2 – TM5,4 39.8 TM3,4 –TM1,7 29.5
TM1,2 – TM6,5 31.8 TM4,5 –TM5,4 33.8
TM1,2 – TM7,6 28.2 TM4,5 –TM6,5 29.2
TM1,2 – TM1,7 38.3 TM4,5 –TM7,6 30.8
TM2,3 –TM3,2 36.7 TM4,5 –TM1,7 30.2
TM2,3 –TM4,3 36.1 TM5,6 –TM6,5 22.2TM2,3 –TM5,4 38.7 TM5,6 –TM7,6 19.2TM2,3 –TM6,5 30.0 TM5,6 –TM1,7 18.7TM2,3 –TM7,6 27.0 TM6,7 –TM7,6 17.4TM2,3 –TM1,7 38.0 TM6,7 –TM1,7 24.2
TM3,4 –TM4,3 36.4 TM7,1 –TM1,7 32.4
LigandSpacer Length
(Å)a ED50b (95% C.I.) nmol
MA-16 19.1 0.039 (0.032 – 0.046)
MA-17 20.4 0.040 (0.033 – 0.046)
MA-19 22.9 0.040 (0.023 – 0.050)
MA-20 24.1 0.037 (0.029 – 0.045)
MA-21 25.4 0.044 (0.039 – 0.048)
MDAN-16 19.1 1.79 (1.54 - 2.04)
MDAN-17 20.4 1.49 (1.04 - 1.95)
MDAN-18 21.6 0.95 (0.68 - 1.23)
MDAN-19 22.9 0.43 (0.36 - 0.50)
MDAN-20 24.1 0.17 (0.15 - 0.19)
MDAN-21 25.4 0.08 (0.06 - 0.10)
MA-19 + DN-20 0.037 (0.031 – 0.043)
Oxymorphone 25.4 0.043 (0.034 – 0.052)
Antinociceptive activity in the mouse tail-flick Antinociceptive activity in the mouse tail-flick assay after acute I.C.V administrationassay after acute I.C.V administration
NTI PretreatmentNTI PretreatmentEffect of Pretreatment with Naltrindole on Acute Antinociceptive Potency of Selected - Bivalent Ligands Administered i.c.v.
ED50 (95% C.I.) nmol
Ligand No PretreatmentNTI (50 pmol) Pretreatment
MDAN-16 1.70 (1.50 – 2.00) 0.30 (0.19 – 0.43)
MDAN-19 0.43 (0.36 – 0.50) 0.05 (0.05 – 0.07)
MDAN-21 0.08 (0.06 – 0.10) 0.06 (0.05 – 0.07)
MA-19 0.04 (0.023 – 0.045) 0.08 (0.065 – 0.085)
Model for Negative Modulation of AntinociceptionModel for Negative Modulation of Antinociception
Other Approaches to Avoid Tolerance Other Approaches to Avoid Tolerance and Dependenceand Dependence
Ananthan, S.; Khare, N.K.; Saini, S.K.; Seitz, L.E.; Bartlett, J.L.; Davis, P.; Dersch, C.M.; Ananthan, S.; Khare, N.K.; Saini, S.K.; Seitz, L.E.; Bartlett, J.L.; Davis, P.; Dersch, C.M.; Porreca, F.; Rothman, R.B.; Bilsky, E.J. Identification of opioid ligands possessing Porreca, F.; Rothman, R.B.; Bilsky, E.J. Identification of opioid ligands possessing mixed mu agonist/delta antagonist activity among pyridomorphinans derived from mixed mu agonist/delta antagonist activity among pyridomorphinans derived from naloxone, oxymorphone, and hydromorphone. naloxone, oxymorphone, and hydromorphone. J. Med. Chem.J. Med. Chem. 20042004, 47, 1400-1412 , 47, 1400-1412
DIPP-NH2[]
1
1
2
2SoRI 9409
Schiller, P.W.; Fundytus, M.E.; Merovitz, L.; Weltorski, G.; Nguyen, T.M.D.; Lemieux, C.; Schiller, P.W.; Fundytus, M.E.; Merovitz, L.; Weltorski, G.; Nguyen, T.M.D.; Lemieux, C.; Chung, N.N.; Coderre, T. The opioid mu agonist/delta antagonist DIPP-NH(2)[Psi] Chung, N.N.; Coderre, T. The opioid mu agonist/delta antagonist DIPP-NH(2)[Psi] produces a potent analgesic effect, no physical dependence, and less tolerance than produces a potent analgesic effect, no physical dependence, and less tolerance than morphine in rats. morphine in rats. J. Med. Chem. J. Med. Chem. 19991999, 42, 3520-3526., 42, 3520-3526.