Roques BP, Garbay-Jaureguiberry C, Oberlin R, Anteunis AK & … · 2019. 10. 2. · RB101(S), a dual inhibitor of enkephalinases does not induce antinociceptive tolerance, or cross-tolerance

2

Roques BP, Garbay-Jaureguiberry C, Oberlin R, Anteunis AK & Lala AK, Nature (1976) 262, 778-779.

Conformation of Met5-enkephalin determined by high fieldPMR spectroscopy

(DPLPE)

(DPDPE)

Tyr-D-Ser-Gly-Phe-Leu-Thr (DSLET)

Tyr-D-Thr-Gly-Phe-Leu-Thr (DTLET)

Potency of opioid peptides to inhibit the binding of [3H]DAGO and [3H]DTLETfrom rat brain tissue at 35°C.

3

From Besse D, Lombard M.C., Perrot S. & Besson J.M., Neurosci. (1992) 50 921-933.

The four different examples correspond to rats with: asham operation on the left sciatic nerve (sham), a looseligation of the right sciatic nerve and a sham operation onthe left one (lig.), a section of the right sciatic nerve (sec.)and a right T13-S2 dorsal rhizotomy (rhiz.).

Individual examples of autoradiograms showing the binding of 3 nM [3H]DAMGO (MOR) and 3 nM[3H]DTLET (DOR) at two weeks post-lesion in the L4 lumbar segment of rat spinal cord

Time-related modifications in 3 nM [3H]DAMGO and 3 nM[3H]DTLET specific binding in laminae I and II of the L4segment for intact rats, shamA rats with a right sciatic nervesection.

4

Autoradiographic study of µ and ∂ opioid binding sites andneutral endopeptidase-24.11 in rat dorsal root rhizotomy

From J.M. Zajac, M.C. Lombard, M. Peschanski, J.M. Besson & B.P. Roques. Brain Research (1989) 477, 400-403.

5

[3H]DAGO (µ)

[3H]DTLET ( )

[3H]HACBO-Gly (NEP)

Photomicrographs illustrating carrageenin-evoked c-Fosprotein immunoreactivity in frontal sections (40 µm) of thedosal horm at the level of l4-L5 segments.

From Le Guen S., Noble F., Fournié-Zaluski, MC., Roques BP, Besson JM. & Buritova J..Eur. J. Pharmacol. (2002) 441, 141-150.

6

RB101(S), a dual inhibitor of enkephalinases does not induce antinociceptive tolerance,or cross-tolerance with morphine: a c-Fos study at the spinal level

7

Travaux effectués en collaboration entre nos deux unités

1 • concernant la distribution des récepteurs mu, delta, kappa, et inhibiteurs d’enzyme

1987. J.M. ZAJAC, P. DELAY-GOYET, M. PESCHANSKI, N. SALES, Y. CHARNAY, G. GUILBAUD, Y. AGID and B.P. ROQUES.Proceedings of Colloquim of Protides of Biological Fluids, H. Peeters, Ed., Pergamon Press Oxford, 1987, 35, pp. 157-160."Distribution in the brain spinal cord (rat and human of enkephalin targets (mu, delta opioid receptors and enekphalinase) in variousphysiophathological conditions ».

1988. J.M. ZAJAC, M. PESCHANSKI, J.M. BESSON and B.P. ROQUES.in Pain Res. Clinical Management, .R. Dubner, G.F. Gebhart and M.R. Bond Eds., Elsevier Science Publishers, Amsterdam, 1988, vol. 3, 436-441."High resolution autoradiography of opioid receptors and enkephalinase in the rat spinal cord".

1989. J.M. ZAJAC, M.C. LOMBARD, M. PESCHANSKI, J.M. BESSON and B.P. ROQUES.Brain Res., 1989, 477, 400-403."Autoradiographic study of mu and delta opioid binding sites and neutral endopeptidase 24-11 in rat after dorsal root rhizotomy".

1989. J.M. BESSON, M.C. LOMBARD, J.M. ZAJAC, D. BESSE, M. PECHANSKI and B.P. ROQUES.Proc. of Sensory Information in the Superficial Dorsal Horn of the Spinal Cord, F. Cervero, G.J. Bennett and P.M. Headlay Eds., Plenum PublishingCorporation, 1989, pp. 415-428."Opioid receptors in the dorsal horn of intact and deafferented rats : autoradiographic and electrophysiological studies".

1990. D. BESSE, M.C. LOMBARD, J.M. ZAJAC, B.P. ROQUES and J.M. BESSON.International Narcotic Research Conference 1989, Progress in Clinical and Biological Research, vol. 328, A.L. Liss Inc., New York, 1990, 179-182."The use of [3H]DAGO to label projections of this primary afferent fibres at the super ficial dorsal horn level of the rat spinal cord".

1990. D. BESSE, M.C. LOMBARD, J.M. ZAJAC, B.P. ROQUES and J.M. BESSON.International Narcotic Research Conference 1989, Progress in Clinical and Biological Research, vol. 328, A.L. Liss Inc., New York, 1990, 183-186."Pre- and post-synaptic location of mu, delta and kappa opioid receptors in the superficial layers of the dorsal horn of the rat spinal cord ».

8

2 • utilisation de cFos

1994. C. ABBADIE, P. HONORE, M.C. FOURNIE-ZALUSKI, B.P. ROQUES and J.M. BESSON.Eur. J. Pharmacol., 1994, 258, 215-227."Effects of opioids and non-opioids on c-Fos-like immunoreactivity induced in rat lumbar spinal cord neurons induced by noxious heat stimulation".

1997. P. HONORE, J. BURITOVA, M.C. FOURNIE-ZALUSKI, B.P. ROQUES and J.M. BESSON.J. Pharm. Exp. Ther., 1997, 281 (1), 208-217."Antinociceptive effects of RB 101, a complete inhibitor of enkephalin-catabolizing enzymes, are enhanced by a CCK-B receptor antagonist as revealedby noxiously-evoked spinal C-Fos expression in the rat".

1999. S. LE GUEN, P. HONORÉ, G. CATHELINE, M.C. FOURNIÉ-ZALUSKI, B.P. ROQUES, & J.M. BESSON.Brain Res. 1999, 834(1-2), 200-206."The effects of RB 101, a complete inhibitor of enkephalin-catabolizing enzymes, on carrageenin-induced spinal c-Fos expression are completelyblocked by ß-funaltrexamine, a selective µ-opioid receptor antagonist".

2002. S. LE GUEN, F. NOBLE, M.-C. FOURNIÉ-ZALUSKI, B.P. ROQUES, J.-M. BESSON, & J. BURITOVA.Eur. J. Pharmacol., 2002, 441(3), 141-150."RB 101(s), a dual inhibitor of enkephalinases does not induce antinociceptive tolerance, or cross-tolerance with morphine : a c-Fos study at the spinal level »

2003. S. LE GUEN, G. CATHELINE, M.-C. FOURNIÉ-ZALUSKI, B.P. ROQUES, J.-M. BESSON & J. BURITOVA.Brain Res. 2003, 967, 106-112.Further evidence for the interaction of m- and d-opioid receptors in the antinociceptive effects of the dual inhibitor of enkephalin catabolism, RB 101(S). A spinal c-Fos protein study in the rat under carrageenin inflammation.

2003. J. BURITOVA, S. LE GUEN, M.-C. FOURNIÉ-ZALUSKI, B.P. ROQUES & J.M. BESSON.Eur. J. Pain 2003, 7(3), 241-249.Antinociceptive effects of RB 101(S), a complete inhibitor of enkephalin-catabolizing enzymes, are enhanced by (+)-HA966, a functional NMDA receptor antagonist : A c-Fos study in the rat spinal cord.

Travaux effectués en collaboration entre nos deux unités

9

3 • études pharmacologiques des opioïdes sélectifs et des inhibiteurs d’enképhalinases sélectifs ou mixtes

1986. J.M. BENOIST, V. KAYSER, G. GACEL, J.M. ZAJAC, M. HAUTRON, B.P. ROQUES and G. GUILBAUD.Brain Res., 1986, 398, 49-56."Differential depressive action of two mu and delta opioids ligands on neuronal responses to noxious stimuli in the thalamic ventrobasal complex of rat ».

1986. A. NEIL, V. KAYSER, G. GACEL, J.M. BESSON and G. GUILBAUD.Eur. J. Pharmacol., 1986, 130, 203-208."Opioid receptor types and antinociceptive activity in chronic inflammation: both kappa and mu opiate agonistic effects are enhanced in arthritic rats".

1989. V. KAYSER, M.C. FOURNIE-ZALUSKI, G. GUILBAUD and B.P. ROQUES.Brain Res., 1989, 497, 94-101."Potent antinociceptive effects of kelatorphan (a highly efficient inhibitor of multiple enkephalin degrading enzymes) systemically administered in normal and arthritic rats".

1989. P. DELAY-GOYET, V. KAYSER, J.M. ZAJAC, G. GUILBAUD, J.M. BESSON and B.P. ROQUES.Neuropharmacol., 1989, 28(12), 1341-1348."Lack of significant changes in mu, delta opioid binding sites and neutral endopeptidase EC 3.4.24.11 in the brain and spinal cord of arthritic rats".

1993. J.A. DESMEULES, V. KAYSER, G. GACEL, G. GUILBAUD and B.P. ROQUES Brain Res., 1993, 611, 243-248."The highly selective delta agonist BUBU induces an analgesic effect in normal and arthritic rat and this action in not affected by repeated administration of low doses of morphine.

1993. S. PERROT, V. KAYSER, M.C. FOURNIE-ZALUSKI, B.P. ROQUES and G. GUILBAUD.Eur. J. Pharmacol., 1993, 241, 129-133."Antinociceptive effect of systemic PC 12, a prodrug mixed inhibitor of enkephalin-degrading enzymes, in normal and arthritic rats ».

Travaux effectués en collaboration entre nos deux unités

10

1991. B.P. ROQUES.in Towards a New Pharmacotherapy of Pain, DAHLEM Conference report, A.I. Basbaum and J.M. Besson, Eds, Chichester, J. Wiley & Sons Ltd., 1991, 257-277."What are the relevant features of the distribution, selective binding and metabolism of opioid peptides and how can these beapplied to drug design ?".

1992. J.M. BESSON et B.P. ROQUES.La Douleur : Au-delà des Maux, Editions des Archives Contemporaines, 1992, 39-48."Stratégie moderne dans le développement de nouveaux analgésiques : de l'opium aux endomorphines".

1992. J.M. BESSON and B.P. ROQUES.The Puzzle of PAIN, Gordon and Breach Art International , 1992, 38-48."Modern strategies in the development of new analgesics : from opium to endogenous opioids".

Travaux effectués en collaboration entre nos deux unités

11

From D.L. Donnelly-Roberts, Abbott Laboratories.

La suppression de la douleur : un objectif loin d’être atteint …

Depuis des décennies aucun analgésique innovant aussi puissant que la morphine mais

dépourvu de ses effets indésirables majeurs n’a été mis sur le marché.

Le besoin de nouveaux analgésiques est urgent aussi bien pour traiter les douleurs aigües permanentes

(cancers, accidents graves, etc) que les douleurs neuropathiques (allodynie, hyperalgésie) dues à des

lésions ou affections les produisant au niveau des terminaisons et fibres périphériques avec projection

centrale.

Pas de traitement réellement efficace, nombreux effets secondaires pour les anticonvulsivants et les

antidépresseurs.

La découverte des récepteurs opioïdes et de leurs agonistes endogènes,

les enképhalines, avait laissé espérer le développement d’une « endomorphine idéale » … !

Les raisons de l’échec, les moyens de le surmonter.

EOS

ECS

EOS

ECS

EOS, ECS

Attacking pain at

its source

Relieving or reducing pain at its source

More than 50% of MO effects are attributable toperipheral neurons (nociceptors)

(From Roques, B.P., Fournié-Zaluski, M.C. and Wurm, M., Nature Reviews Drug Discovery, 2012))

Three levels of pain control by endogenous opioid system (EOS)and endogenous cannabinoids system (ECS)

13

NEP

Contrôle de la douleur par exaltation del’action des morphines endogènes

YGGFM(L) YFGGFM(L)

Y

+ GGPM(L)

YGG

+ FM(L)

APN

PENK

ENKs

MOR, DOR

Zn Zn

ENKs

SV

Plasma membrane

cytosol

DENKI DENKI

Extended synaptic area

Opioid signaling

(Exogènes)

Récepteurs opioïdes

Morphine

DORMOR

Hyperstimulation ubiquitaire de tous les récepteurs opioïdes (ORs)

même ceux qui ne sont pas impliqués dans le contrôle de la douleur.

Effets indésirables

Tyr – Gly – Gly – Phe – Met (Leu)Enképhalines(Endogènes)

APN NEP

Stimulation restreinte aux sites où les ENKs sont

secrétées en quantité suffisante sous l’effet d’un

stimulus (ex. douleur). Idée : développer des

inhibiteurs mixtes (DENKIs) bloquant NEP et APN qui agiront sélectivement sur un nombre limité de ORs.

Absence d’effet indésirable

Roques, B.P., Fournié-Zaluski, M.C. and Wurm, M., Nature Reviews Drug Discovery, 2012, 11, 292-311.

The two leading families of DENKIs

8x10-9M

2x10-9M

3x10-9M4x10-9M

APN NEP

Enkephalins

Tyr -Gly -Gly -Phe -Met(Leu)

Cascade ester (*)

(*) Increases bio -availability

PL 37PL800 PL804

PL804PL800

NEPAPN

(*) Increases bio -availability

Cascade carbamate (*)

PL 265

PL 254

PL

Enkephalins

Tyr -Gly -Gly -Phe -Met(Leu)

254

Ki = 8 nMKi = 2 nM

Ki = 4 nMKi = 3 nM

PL37 : R1 = H ; R2 = -CH(CH3)OCOOEt

PL265 : R1 = CH3 ; R2 = CH3 ; NH3+ = -NH-COO-CH(CH3)-OCOIPr

PL254 : R1 = CH3 ; R2 = CH3 ; NH3+

Effect of DENKIs on the extracellular efflux of Met-enkephalinin the nucleus accumbens, periaqueductal grey (PAG) and cerebrospinalfluid (CSF) Microdialysis or superfusion in awake and freely moving rats

Bourgoin S. et al., JPET, 1986.

Sub-arachnoidal

PAGNucleus accumbens

artificial CSF

17

Potentiation de l’action de la Met-enkephaline (ME) sur les neurones du locus coeruleus par le kélatorphan.Absence d’effet propre du kélatorphan.

Exogenous ME (or MO) applied by pressure pulse () on slice containing the LC, leads to hyperpolarization. The response is amplified by kelatorphan (J.T. Williams et al., J. Pharmacol. Exp. Ther., 243(1), 397-401, 1987)

De ce fait les DENKIs n’ont pas d’effet indésirable sur la respiration tant chez l’animal (E. Boudinot et al., Pain 90, 7-13 (2001) que chez l’homme (étudeclinique du PL37)

Le locus coeruleus est une structure cérébrale impliquée dans le contrôle de la

respiration et du rythme cardiaque ainsi que dans les phénomènes addictifs.

Le Kelatorphan seul est inactif indiquant l’existence d’une libération basaletrès faible des ENKs dans cette structure

18

Puissance analgésique similaire de la morphine et des enképhalines par voiei.v. dans les modèles des douleurs aiguës.

i.c.v. in miceED50 Met-E+kelatorphan (50 µg) : 1.2 nmol/mouse (Fournié-Zaluski, EJP 1984)ED50 MO : 0.63 nmol/mouse; ED50 Met-E : 330 nmol/mouse (Interussi, PNAS 1980)

vehicle (EtOH/cremophor/H2O) improving BBB crossing

Comparison between i.v. morphine and i.v. PL37 in the Hot Plate Test (HPT)

% analgesia = 100 x

(test latency – control

latency) / (cut-off time

– control latency)

** p

19

Induction of peripheral

nociceptors hyperexcitability

Schwann cell

Blood capillary

Oligodendrocyte

EOS and neuropathic pain

NORMAL STATE

NEUROPATHIC STATE

Spontaneous discharges by experexpression of Na+ channels in injured nerves

Slow weakly myelinated (∆ds) and unmyelinated (C) fibers

Fast myelinated fibers Aa, Aß

non noxiousstimuli

noxious

stimuli(allodynia)

noxious

stimuli(hyperalgesia)

to spinalcord

( ) Hyperexpression of ORsenrichment in ENKs

Sites ofDENKIs action

Induction of peripheral

nociceptors hyperexcitability

(Alteration of Blood Nerve Barrier with transfert ofpronociceptive compounds; TNF, interleukins, cytokines, …)

(intact Blood Nerve Barrier)

to spinalcord

to spinalcord

non noxiousstimuli

to spinalcord

Pain modulation by descending systems

central

sensitization

Hyperactivation of Glu system.

Reduction of GABA system.

Final State

Ex. tactile

20

PL37 reduction of neuropathic pain (allodynia and thermal hyperalgesia) induced by partial ligation of sciatic nerve in mice

**p

PL37 is active per os at the peripheral level in a model of inflammatory pain.

Administration of the peripherally selective opioid

antagonist naloxone methiodide (i.p.) antagonized

antinociception elicited by oral administration of PL37

(100 mg/kg).

PL37 activates peripheral opioid receptors

Antinociceptive effects of PL37 (100 mg/kg/po) on vocalization threshold to paw pressure in rats with

carrageenan-induced hindpaw inflammation

BaselinevehiclePL37100

200

300

400

inflamed paw

# # #

Vo

cali

zati

on

th

resh

old

(g

)

0 20 40 600

100

200

300

400

500vehiclePL37/NaClPL37/Nlxe-MET

Time, mn

22

First demonstration of synergistic interaction between gabapentin and DENKI-protected endogenous opioids in neuropathic pain alleviation (thermal hyperalgesia).

Collaboration with Ana Baamonde, Universidad de Oviedo, Spain

Experimental ED50 (with synergy) : 7.03

Computed ED50without synergy : 12.8

PL265 and gabapentin show the same synergistic effects in this model

Other synergistic effects are observed between PL37 or PL265 and P2X3R, CCKBR antagonists and CB2R agonists

Synergistic effects of PL37 combinedwith gabapentin, both at subactive

doses.

PL37 antihyperalgesic effect is reversed by naloxone

methiodide. Isobolographic analysis

Peripheral analgesia.

PL37

NAL-MET

25 25- -

2 2- - 2

- 25 25-

- - 2

Menendez et al. Eur. J. Pharmacol., 2008

Synergistic antihyperalgesic effects of oral PL37 and gabapentin in murine tibial osteosarcoma

From Rice A.S.C. et al. The Lancet (2014) 383, 1637-1647.

23