- Email: [email protected]
S-4-2 Therapeutic potential of drugs acting on excitatory amino acid receptors
169
S-4 Pharmacological treatment o/stroke" drugs, or ey adding directly the : l u c s to the p h o s p r o r y l a t i o n svstem vitro. The increase iii c~-CaMK h autophospr-orvla:~on could be accour'teO fo by: 1) increased p h o s p h a t e mcorporat;on nlto the k i ' l a s e 2) ncreased n e o s y n t h e s i s of the k=nase: 3) translocat=on of the klnase The level o ~ {~ C a M K li w a s analyzed in the s u b s y n a p t o s o m a l fractions from control a r a treated animals by q u a n t i t a t i v e m m u n o b l o t s h o w i n g that no s i g n i f c a n t c h a n g e of c~ C a M K II level o c c u r r e d in the presynaptic fractions of treated animals. The total e n z y m a t i c actw=ty of ~ C a M K I was also ~ e a s u r e d usmg a synthetic o e p t l d e s u b s t r a t e m the a n n a l s treated w=th f l u v o x a m m e and v e n l a f a x m e The activity w a s slgnlf,cantly mcreased in LP2 and LS2 ÷rac trons, but n o t in LP1 fraction, after t r e a t m e n t The increase was roughh/ e q u i v a l e n t in LS2 { - 7 0 - 8 2 % ' , with the t w o drugs, w h e r e a s m LP2 (synoptic vesicles) it w a s higher with v e n l a f a x m e (+ I£.5%) c o m p a r e d to fluvoxam ine(+60%) It =s a matter o ~ speculation w h e t h e r this difference is due to different specificity of the t w o drugs Long-lasting modifications m C a M K ii a u t o p h o s p h o r y l a t l o n and acbvltv have been showP in several different e v e n t s Pathological events n c l u d e i s c h e m i a g l u t a m a t e - i n d u c e d seurctoxlc~t'~ seizures ohvslologlcal ones =n clude long term p o t e n t arian and synaptrc olas~ city The results here re ported are in a g r e e m e n t witt ~ the ev d a n c e s u g g e s t ng a presynaptlc effec: for SSIRIs in one of the m a n s e r o t o n e r g l s pathways, ar'd c~sclose a n e w site in t h e r n e c h a q i s m o f a c t l o r " o ; a n t l c e p r e s s a n t drugs Also theyconfnr~" that protein kinase m o d i f i c a t o n s appear to be of s r i m a r y l m p o r t a n c e n t h E plastic c h a n g e s o c c u r n n g in CNS ;ollowiPg p~y51o ogical or oathological as well as p h a r m a c o l o g i c a l e v e n t s
Na ÷ Ca2+ Sperm=he
~ace,~
Spermlne
NRIA
Hlslamine
r
lenp oa
NR1A/ NR2A
NR1A/ NR2B
Sl0ermlne
NR1A/ NR2C
NR1A/ NR2D
Effect of spetmine 1 "Glycine-inaependent" stimulation
yes
no
ye~
no
no
2 "Glycine-dependent" slimulation
yes
yell
yell
nO
no
3 Vo~sge-dependerlt inhibrtion
yell
yes
yes
no
no
4 Decrease affinity 'cr agonists
ye$
rio
yli
ha
no
Potentiafion by histamine
yo=
no
ye=
no
no
6
Rolatr~e affinity 1or ifenprodil
high
low
high
low
low
7
Relat~'e aflFnily lor glycinB
high
~ow
high
high
v. high
high
high
high
low
low
(increase aflinily for glycine)
8 Ralahve affinity for Mg 2~
Ftgure 1 Subunit-dependetlt properties of recombinant N M D A receptors
References
Bliec R and de Mor-tign,r ( (1994) Currer, t e J'c t'er:qs r" t'~ trectrrler~t ' depression T. in Pharm Sc, ~5 220 225 Blier, P and Bouchard C (1994) Modulation of 5 HT re ease m the gumea p,g bran ;ol c,.*{ ing long-term administration of antidepressant drugs Br J Pharrracol 113. 485 49b Greengard. P Vatorta. F Czern~ AJ and Be~ferat~ ; 11993) Synoptic veslc!e Phos phoprotelns and regular~oP ol synoptic fupcr or/ Science 259, 780 785 Popoli, M (1993) Synaptotagmm ~s endogenous v phospbcrylated by {:a2+/calmOOU r protein kinase II m synapt~c yes c~es FEBS Lett 21,' 85 88
S-4 Pharmacological treatment of stroke Molecular pharmacology of NMDA receptors Ke~th Williams Depa~ment of ,Dhaffr~aco/og'y, Ur~,'ver~,,t), e/Penrls~'/vam~ ~, Schoo/ of MedJGrre, Ph/lade/ph/a, PA 79704 6(284. tJ'~A N M D A r e c e p t o r s are n v c l v e u n the inoui:tlc ~ : f ~a leo.~ rv[:i s of svr apt r plasticity in the central nervoL, s System and q m_~u'opathologles nslus 'ig ischemia induced neLronal ceh ~eath ] h e a e ,anal:tars ore~ent potentla targets for n e u r o p r o t e c t w e agents N M D A r e s e g l c ' s ale a e : v a t e d bY g : t a m a t e (or N M D A ) t o g e t h e r w t h glycme Ir~ acd t l o n t~e ~DreDe'tlea ai:~ acbvatidn of these receptors can be altere:~ 2~, a urge ',u'nb~r of e c n o g e nous and e x o g e n o u s m o d u l a t o r s and
i S-4-2 J Therapeutic potential of drugs acting on excitatory amino acid receptors
5 Scattor'. R Avenet. J Benavides. F Besnard. C Carter, D Duverger. i: C,~aham,~ l@onarden, R Santa Maria, S Z L a n g e r Synthe/abo
Recherche, CNS Research Dent, Bagneux 92220, France The s,/napt~c r e s o o n s e s elicited by e x c i t a t o r v amino acids (EAA) eg L :]iutamate and aspartate in the CNS are m e d i a t e d by d i s t i n c t classes of re:eplers which have been categorized into G o r a t e s c o u p l e d m e t a b o t r o p i c ~'ld i~gand-gated ionotrooic ( N M D A and AMPA-kainate s u b t y p e s } receptors. D e N M D A r e c e p t o r has attracted m u c h interest because its plays an import e r : role IF processes such as learning and m e m o r y and neuronal d e g e n e r ,inl h e ' e are a n J m b e r of distinct sites on the N M D A r e c e p t o r c o m p l e x g l u t a m a t e r e c o g n t l o n site, glycme and polyamine m o d u l a t o r y sites) w h i c h urov de potential targets f o r a p h a r m a c o l o g i c a l blockade of this r e c e p t o r a r d selective antagonists n o w exist for each of these sites (Scatton, 1993). :)ho, sDhonate analogs of carboxylic acid (eg CGS 19755, CGP 37849, LY 233053~ are c o m p e t i t i v e antagonists f o r the N M D A r e c o g n i t i o n site. Drugs u c b as phencychdine, dizocilgine [ M K - 8 0 I ) . m e m a n t i n e and d e x t r o r p h a n ~ct as cpen channel b o c k e - s of the N M D A r e c e p t o r A n t a g o n i s t s of the m o d u l a t o r y giyc ne site are derived f r o m kynurenic acid and indole acetic 1did and mclude 5.7 d i c h l o r o k y n u r e n a t e , L 689.560, M D L 100,748 and M D L :9951 F nally, p n e n y l e t h a n o i a m i n e s eg ifenprodil, eliprodil (SL 82.0715) i n d tlbalosme have been s u g g e s t e d to act as antagonists of the p o l y a m i n e ",oculatorv srte ~(:arter et al, 1991). V >lecular clonmg e x p e r i m e n t s have estabhshed that the N M D A r e c e p t o r , cor~oosed of t w o famdies of subunlts NR 1 (8 s p t c e variants) and NR 2 ,4 L~ionreric f o r m s A to D) with distinct b r a n regional distribution p a t t e r n s ; u ' c t i o n a l v d s t n c t N M D A r e c e p t o r s u b t y p e s are f o r m e d b y h e t e r o m e r i c s s e m b l v o; the NR 1 s u b u n l t with d # e r e n t rT'embers of the NR 2 s u b u n i t 'am i y ( N a k a n i s h , 1992). Certain N M D A ~eceptor antagonists appear to posa s s select~v ty for a gwen suotype. Thus, in X e n o p u s o o c y t e s t r a n s f e c t e d in :he i ' u c l e t w t h p a s m ~ d r n x t u r e s c o n t a m i n g t h e NR1 and NR 2 r e c o m b i n a n t :)NAs, eliprod,I and ifenprodd select,vely innibi: the N M D A - e v o k e d c u r r e n t ff h e t e r o m e n c NR~/NR28 receptors {respective IC50's 0 2 2 and 2 9 # M ) ' , u t / , a t of receptors c o m p r i s e d of the NR 1/NR2A subunits (IC50 > 30 # M ) , contrast, dizoc,tpine blocked t h e N M D A induced c u r r e n t w i t h an IC50 ~a,L.e of 30 nM in both NR1/NR2B and N R I / N R 2 B r e c e p t o r c o m b i n a t i o n s . ' h e selectivity of fenprodil f o r a s u b t y p e of N M D A r e c e p t o r is s u p p o r t e d :;y the distnbut~cn of p o l y a m i n e sensitive 3H-i÷enprodil binding sites w h i c h Iosevmatchestr]atof N M D A R 2 B r e c e p t o r m R N A Moreover, in r a t s t r i a t a l ,;l~:es, ~fenp~odd and eliprooil block the s t i m u a t e r y effects of N M D A on 14:: acetylcholme release (respective IC50's 5 and 3 /*M), b u t are w i t h o u t , f f e a t on N M D A e v o k e d s p e r m i d i n e release (IC50 > 100 / z M ) T h i s profile shared bv oolyamine spider toxins and the N M O A channel blockers d e x ~ a m e t h o r D h a n and m e m a n t i n e whde d~zocdpine, phencyclidine, C G P 3 7 8 4 9 . l ~ L 6 8 9 , 5 6 0 alack both responses w'Jth equal potency. Finally, in cerebellar
170
S-4 Pharmacological treatment of stroke
granule celia from the r eonatal rat denoroo:l and ehprodil partla ly anrago nlze (50%), whereas dizoc,lpine 2-APV and 7-chlorokynurenate rote ly L]tock NMDAqnduced increase in u]ternal Ca 2. concentrations. Overactwatlon of EAA receptors, particularly of the NMDA receptor s.~b type, by endogenous glutamate/aspartate is now accepted as a major mech anism of neuronal damage that occurs in a variety of neurologica condl tions including cerebral ischem a, CNS trauma and chronic neurodegeqer atwe diseases (see Scatton, 1994) NMDA receptor antagomsts targeted at the distinct recognition or modulatory sites have been shown to blunt glutamate induced inlury in cell cultures, to prevent hypoxia-induceo EPSP decrease ~n rat hippocampal shces and to salvage a large portion of bral~ tissue in anlma~ mooels of stroke and cerebral trauma The degree of neu roprotecton afforded by these d r u g s l s v e r y l a r g e For instance, oEzocilplne (! mg/kg ip) and eliprodl (1 '-'g/kg ivl administered following permanert occlusion of the m~adle cereora, artery ~n the rat diminish by --~50% the total infarction v o l u ~ e Sire!lady, "epeated administration of ehprodd {10 mg/kg ~p) to rats sub}ectec to lateral fluid percussion of the parietal cortex reduces by 60% the volume of the t~aumatic lesion. As a general r u e in models of focal iscqemla, charnel olockers and eliprodd are more potent and efficacious than NMDA recogmtloq or glycn'e site aqtagomsts probably because of their better brain per'etratlon Unfortunately, NMDA receptor antagomsm can be accompanied by a bat terv of undesirable side effects including, m va'y ng degree, phencyclidine like psychostlmulation, amnesia rntrinsic qeurotoxlclty on specific cortica! neuron populations, hypertenslor ~andtachycardla These effects arepartlc ularly intense with certain NMDA chaqne! blockers {although less so irq the case of dextrcrphan like comoounds), and can also be observed with coopetitwe NMDAantagonlsts Glycineslteantagon~stsappeartohavea more favourable p~ofile at neuroprotective doses, w6ile eliprodil is completely devoid of such side effects {G~roux et at 1994) Trle select:ve blockage by eliprodil (and to some extent non stimulant channel blockers} o{ a NMDA receptor sub~,ype may account for its favorable safety index as compared to conventional NMDA antagonist which block indtscriminately el' NMDA receptor subtypes In conclusion, subtype se:ect~ve NMDA receptor antagonists may rep resent a promtsing approach for the medical treatment of acute ischemlc stroke, CNS trauma and poss bly chrorlic neurodegenerative diseases Sev eral NMDA antagonists including CNS 1102, ehprodil, CGS 19755 and dex trorphan are now being evaluated in early treatment clinical trials m stroke o r bra~n trauma and useful theraptes may emerge with those drugs
References Carter, C. Benavides. ~ Dana, C Schoemaker. H Perrault, G r Sanger DJ and Scatto~ B (1991i Non comDetibve NMDA receptor antagonists acting on the poiyam~ne site In ExcltatoryAmlno Acid Antagonists, B S Meldrum Ed, Blackwell Scientific London pp 130 169 Glroux, C Rosen, P and Scatton, B 1994) :~reshn~calpharmacology and ci nical safew prof,le of eliprodil, an atypical NMDA receptor antagonist In Pharmacology of Cere bra Ischem~a J Krieglstein, H Oberplchler Schwenk (Edsi Wissenschaftlche Ve" lagsgesetlschaft mbH Stuttgart, pp 643 648 Nakamshi, S (1992} Molecular dive,s,ty of glutamate receptors and implicat~ors {o, bratP function Science, 258 597 603 Scatton, B (1993) The NMDA recepto ~complex Fundam Clin Pha,macol 7 389 400 Scatton, B (1994) Bxc;tatory amino ac*d recentor antagonists a novel treatment :or ~schemic cereb'ovascular d~seases L~le So, 55 2115 212/*
ogy lhe need for absolute certainty of anti-ischaemic benefit requires that a number of separate tssues are addressed in preclinlcal development. An Nllproved acute histological outcome with the drug must be demonstrated to demonstrate that drug efficacy is independent of physiological artefact. Chronic h~stological outcome must be shown to have been improved by drug ~ntervent~on to dispel doubt that the drug simply slows the maturation of tne ischaemic damage. Some investigations of anti-ischaemic efficacy must be performed in a gyrencephalic species Jn case the mechanism of interest contributes disproportionately to outcome in the lissencephalic brain {spreading depression being an obvious example). The question of therapeutic time w i n d o w must be addressed but it must be appreciated that the qmportant time w i n d o w is that in man and not that in young, healthy ammals with a carefully controlled single ischaemic event. Considerable attention needs to be directed to drug pharmacokinetics not just plasma revels, but CNS levels of the drug around the zone of ischaemia. Once unequivocal anti-ischaemic effect is demonstrated, the central issue becomes safety While preclinical investigations have focused on the adverse effects of the new neuroprotective drugs in the CNS, their effects on cardiovascular and respiratory function are of equal ior greater) importance in the clinical evaluation. NMDA Receptor Blockade. in animal models of focal cerebral ischaemia, unequivocal benefit has been demonstrated after NMDA receptor blockade ,n every species examined {baboon, cat, rabbit, rat, mouse). The maximum reduction (typically 50%) in the volume of infarcted tissue is broadly similar ~rrespective of whether the drug acts at the agonist recognition site of the NMDA receptor complex (e.g. D-CPPene, CGS 19755)the ion channel site {eg. MK-801, CNS 1102, or remacemide hydrochlobde) or polyamine site {eg. eliprodil) (Chen et el., 1991; McCulloch, 1992: Bannan et el., 1994). Although the evidence of anti ischaemic efficacy of the NMDA receptor antagon,sts is overwhelming, the clinical evaluation of these drugs was delayed oy concern over their effects on CNS function such as vacuole formation, immediate early gene or heat shock protein expression in the cingulate cortex, hypermetabolism visualised with deoxyglucose autoradiography e t c (McCulloch, 1992) AMPA Receptor Blockade. There is growing evidence that AMPA recep tor blockers (NBQX, YM 900, LY-293558, GYKI 42566} also reduce damage a~ter experimental focal ischaemia but the volumes of tissue saved are typically smaller than for NMDA antagonists {Bullock et el, 1994), The effects of AMPA receptor antagonists on CNS function bear little resemblance to t~ose of NMDA antagonists. The critical utility of AMPA receptor antagonists ~s constrained by their potent respiratory depressant actions. {Browne and McCulloch, 1994). Glutamate Release lnhlT~itors. Attenuation of the elevations in extracelI~.lar glutamate concentrations which occur in cerebral ischaemia has the advantage of reducing the resultant activation of both NMDA and AMPA receptors There is growing evidence that inhibition of glutamate release wa diverse strategies (Na + and Ca ++ channel blockers, kappa opioid ago n~sts etc) can reduce ischaemic damage in experimental focal ischaemia (Mackay et el, 1993). The effects on CNS function of glutamate release inhibit on are less pronounced than NMDA or AMPA receptor blockade. In summary, it is worth emphasising that for the first time, a class of orugs is entenng clinical trials in human ischaemia based on a certainty of anti-ischaemic benefit in animal models and the data emerging from these chnical trials is most encouraging.
References Glutamate antagonists in animal models of cerebral ischaemia We//come Surgical Institute ~ Hugh Fraser Neuroscience Labs, Un/versity o/Glasgow, Garscube Estate, Bearsder-, Road, Glasgow G6t tOH, Scotland, United Kingdom James McCulloch
Animal models of cerebral ischaerma provide the c~ucla hnk between tqe hypothesis that excttotoxic mecr~anlsms contnbute to iscrlaemlc injury to neurones and the development of new therapeutic strategies for man Models of focal cerebral iscnaem~a (typically occlusion of a middle cerebral artery} have been the most powerful tools in characterising neurop'otechve efficacy of glutamate receptor antagonists and data in these models underpin the current chnical use o~ NMDA antagomsta and glutamate release innibitors In contrast, models of gtobai cerebral ischaemia have not been particularly valuable in antl-lscnaemlc orug development because of con cerP of the pertinence of the selectwe neuronal necrosis of global ischaemia to the processes of infarctton iq stroke (McCulloch. 1992} Anti-ischaem/c Drug Development. A Strategy Although many expert mental endpoints can provide mechanistic insight, the deflmtwe endpo nt for a putahve neuroprotective dr,~g must always be quantltattve blstopat~'oi
Barman, BE, Graham, DI, Lees, K R and McCulloch. J {1994). Neuroprotective effect of remacemide hydroch!oride in focal cerebral ischaemia in the cat Brain Research 664 271-275 Browne, S E 8 McCulloch, J (1994) AMPA receptor antagonists end local cerebral glucose utilization in the rat Brain Research 641, 10 20 Bullock, R Graham, DI, Swanson, S fi McCulloch, J {1994) Neuroprotective effect of the AMPA receptor antagonist LY-293558 in focal cerebral ischemia in the cat. Journal of Cerebrai Blood Flow and Metabolism 14, 46£:~471 McCulloch, J (1992) Excitatory amino acid antagonists and their potential for the treatment of ischaemic brain damage in man. British Journal of Clinical Pharmacology 34, 106.114 Mackav, K B , Kusumoto, K, Graham, D I and McCulloch, J (1993) Focal cerebral ischemia in the cat: pretreatment with a kappa-t opioid receptor agonist, CI-977 Brain Research 618, 213 219 Chen, M Bullock R , Graham, D I , Frey, B, Lowe, D and McCulloch,J (1991). Evaluation of a competitive NMDA antagonist (D-CPPenelin feline focal cerebral ischemia Annals of Neurology 30, 62 70
S-4 Pharmacological treatment o/stroke" drugs, or ey adding directly the : l u c s to the p h o s p r o r y l a t i o n svstem vitro. The increase iii c~-CaMK h autophospr-orvla:~on could be accour'teO fo by: 1) increased p h o s p h a t e mcorporat;on nlto the k i ' l a s e 2) ncreased n e o s y n t h e s i s of the k=nase: 3) translocat=on of the klnase The level o ~ {~ C a M K li w a s analyzed in the s u b s y n a p t o s o m a l fractions from control a r a treated animals by q u a n t i t a t i v e m m u n o b l o t s h o w i n g that no s i g n i f c a n t c h a n g e of c~ C a M K II level o c c u r r e d in the presynaptic fractions of treated animals. The total e n z y m a t i c actw=ty of ~ C a M K I was also ~ e a s u r e d usmg a synthetic o e p t l d e s u b s t r a t e m the a n n a l s treated w=th f l u v o x a m m e and v e n l a f a x m e The activity w a s slgnlf,cantly mcreased in LP2 and LS2 ÷rac trons, but n o t in LP1 fraction, after t r e a t m e n t The increase was roughh/ e q u i v a l e n t in LS2 { - 7 0 - 8 2 % ' , with the t w o drugs, w h e r e a s m LP2 (synoptic vesicles) it w a s higher with v e n l a f a x m e (+ I£.5%) c o m p a r e d to fluvoxam ine(+60%) It =s a matter o ~ speculation w h e t h e r this difference is due to different specificity of the t w o drugs Long-lasting modifications m C a M K ii a u t o p h o s p h o r y l a t l o n and acbvltv have been showP in several different e v e n t s Pathological events n c l u d e i s c h e m i a g l u t a m a t e - i n d u c e d seurctoxlc~t'~ seizures ohvslologlcal ones =n clude long term p o t e n t arian and synaptrc olas~ city The results here re ported are in a g r e e m e n t witt ~ the ev d a n c e s u g g e s t ng a presynaptlc effec: for SSIRIs in one of the m a n s e r o t o n e r g l s pathways, ar'd c~sclose a n e w site in t h e r n e c h a q i s m o f a c t l o r " o ; a n t l c e p r e s s a n t drugs Also theyconfnr~" that protein kinase m o d i f i c a t o n s appear to be of s r i m a r y l m p o r t a n c e n t h E plastic c h a n g e s o c c u r n n g in CNS ;ollowiPg p~y51o ogical or oathological as well as p h a r m a c o l o g i c a l e v e n t s
Na ÷ Ca2+ Sperm=he
~ace,~
Spermlne
NRIA
Hlslamine
r
lenp oa
NR1A/ NR2A
NR1A/ NR2B
Sl0ermlne
NR1A/ NR2C
NR1A/ NR2D
Effect of spetmine 1 "Glycine-inaependent" stimulation
yes
no
ye~
no
no
2 "Glycine-dependent" slimulation
yes
yell
yell
nO
no
3 Vo~sge-dependerlt inhibrtion
yell
yes
yes
no
no
4 Decrease affinity 'cr agonists
ye$
rio
yli
ha
no
Potentiafion by histamine
yo=
no
ye=
no
no
6
Rolatr~e affinity 1or ifenprodil
high
low
high
low
low
7
Relat~'e aflFnily lor glycinB
high
~ow
high
high
v. high
high
high
high
low
low
(increase aflinily for glycine)
8 Ralahve affinity for Mg 2~
Ftgure 1 Subunit-dependetlt properties of recombinant N M D A receptors
References
Bliec R and de Mor-tign,r ( (1994) Currer, t e J'c t'er:qs r" t'~ trectrrler~t ' depression T. in Pharm Sc, ~5 220 225 Blier, P and Bouchard C (1994) Modulation of 5 HT re ease m the gumea p,g bran ;ol c,.*{ ing long-term administration of antidepressant drugs Br J Pharrracol 113. 485 49b Greengard. P Vatorta. F Czern~ AJ and Be~ferat~ ; 11993) Synoptic veslc!e Phos phoprotelns and regular~oP ol synoptic fupcr or/ Science 259, 780 785 Popoli, M (1993) Synaptotagmm ~s endogenous v phospbcrylated by {:a2+/calmOOU r protein kinase II m synapt~c yes c~es FEBS Lett 21,' 85 88
S-4 Pharmacological treatment of stroke Molecular pharmacology of NMDA receptors Ke~th Williams Depa~ment of ,Dhaffr~aco/og'y, Ur~,'ver~,,t), e/Penrls~'/vam~ ~, Schoo/ of MedJGrre, Ph/lade/ph/a, PA 79704 6(284. tJ'~A N M D A r e c e p t o r s are n v c l v e u n the inoui:tlc ~ : f ~a leo.~ rv[:i s of svr apt r plasticity in the central nervoL, s System and q m_~u'opathologles nslus 'ig ischemia induced neLronal ceh ~eath ] h e a e ,anal:tars ore~ent potentla targets for n e u r o p r o t e c t w e agents N M D A r e s e g l c ' s ale a e : v a t e d bY g : t a m a t e (or N M D A ) t o g e t h e r w t h glycme Ir~ acd t l o n t~e ~DreDe'tlea ai:~ acbvatidn of these receptors can be altere:~ 2~, a urge ',u'nb~r of e c n o g e nous and e x o g e n o u s m o d u l a t o r s and
i S-4-2 J Therapeutic potential of drugs acting on excitatory amino acid receptors
5 Scattor'. R Avenet. J Benavides. F Besnard. C Carter, D Duverger. i: C,~aham,~ l@onarden, R Santa Maria, S Z L a n g e r Synthe/abo
Recherche, CNS Research Dent, Bagneux 92220, France The s,/napt~c r e s o o n s e s elicited by e x c i t a t o r v amino acids (EAA) eg L :]iutamate and aspartate in the CNS are m e d i a t e d by d i s t i n c t classes of re:eplers which have been categorized into G o r a t e s c o u p l e d m e t a b o t r o p i c ~'ld i~gand-gated ionotrooic ( N M D A and AMPA-kainate s u b t y p e s } receptors. D e N M D A r e c e p t o r has attracted m u c h interest because its plays an import e r : role IF processes such as learning and m e m o r y and neuronal d e g e n e r ,inl h e ' e are a n J m b e r of distinct sites on the N M D A r e c e p t o r c o m p l e x g l u t a m a t e r e c o g n t l o n site, glycme and polyamine m o d u l a t o r y sites) w h i c h urov de potential targets f o r a p h a r m a c o l o g i c a l blockade of this r e c e p t o r a r d selective antagonists n o w exist for each of these sites (Scatton, 1993). :)ho, sDhonate analogs of carboxylic acid (eg CGS 19755, CGP 37849, LY 233053~ are c o m p e t i t i v e antagonists f o r the N M D A r e c o g n i t i o n site. Drugs u c b as phencychdine, dizocilgine [ M K - 8 0 I ) . m e m a n t i n e and d e x t r o r p h a n ~ct as cpen channel b o c k e - s of the N M D A r e c e p t o r A n t a g o n i s t s of the m o d u l a t o r y giyc ne site are derived f r o m kynurenic acid and indole acetic 1did and mclude 5.7 d i c h l o r o k y n u r e n a t e , L 689.560, M D L 100,748 and M D L :9951 F nally, p n e n y l e t h a n o i a m i n e s eg ifenprodil, eliprodil (SL 82.0715) i n d tlbalosme have been s u g g e s t e d to act as antagonists of the p o l y a m i n e ",oculatorv srte ~(:arter et al, 1991). V >lecular clonmg e x p e r i m e n t s have estabhshed that the N M D A r e c e p t o r , cor~oosed of t w o famdies of subunlts NR 1 (8 s p t c e variants) and NR 2 ,4 L~ionreric f o r m s A to D) with distinct b r a n regional distribution p a t t e r n s ; u ' c t i o n a l v d s t n c t N M D A r e c e p t o r s u b t y p e s are f o r m e d b y h e t e r o m e r i c s s e m b l v o; the NR 1 s u b u n l t with d # e r e n t rT'embers of the NR 2 s u b u n i t 'am i y ( N a k a n i s h , 1992). Certain N M D A ~eceptor antagonists appear to posa s s select~v ty for a gwen suotype. Thus, in X e n o p u s o o c y t e s t r a n s f e c t e d in :he i ' u c l e t w t h p a s m ~ d r n x t u r e s c o n t a m i n g t h e NR1 and NR 2 r e c o m b i n a n t :)NAs, eliprod,I and ifenprodd select,vely innibi: the N M D A - e v o k e d c u r r e n t ff h e t e r o m e n c NR~/NR28 receptors {respective IC50's 0 2 2 and 2 9 # M ) ' , u t / , a t of receptors c o m p r i s e d of the NR 1/NR2A subunits (IC50 > 30 # M ) , contrast, dizoc,tpine blocked t h e N M D A induced c u r r e n t w i t h an IC50 ~a,L.e of 30 nM in both NR1/NR2B and N R I / N R 2 B r e c e p t o r c o m b i n a t i o n s . ' h e selectivity of fenprodil f o r a s u b t y p e of N M D A r e c e p t o r is s u p p o r t e d :;y the distnbut~cn of p o l y a m i n e sensitive 3H-i÷enprodil binding sites w h i c h Iosevmatchestr]atof N M D A R 2 B r e c e p t o r m R N A Moreover, in r a t s t r i a t a l ,;l~:es, ~fenp~odd and eliprooil block the s t i m u a t e r y effects of N M D A on 14:: acetylcholme release (respective IC50's 5 and 3 /*M), b u t are w i t h o u t , f f e a t on N M D A e v o k e d s p e r m i d i n e release (IC50 > 100 / z M ) T h i s profile shared bv oolyamine spider toxins and the N M O A channel blockers d e x ~ a m e t h o r D h a n and m e m a n t i n e whde d~zocdpine, phencyclidine, C G P 3 7 8 4 9 . l ~ L 6 8 9 , 5 6 0 alack both responses w'Jth equal potency. Finally, in cerebellar
170
S-4 Pharmacological treatment of stroke
granule celia from the r eonatal rat denoroo:l and ehprodil partla ly anrago nlze (50%), whereas dizoc,lpine 2-APV and 7-chlorokynurenate rote ly L]tock NMDAqnduced increase in u]ternal Ca 2. concentrations. Overactwatlon of EAA receptors, particularly of the NMDA receptor s.~b type, by endogenous glutamate/aspartate is now accepted as a major mech anism of neuronal damage that occurs in a variety of neurologica condl tions including cerebral ischem a, CNS trauma and chronic neurodegeqer atwe diseases (see Scatton, 1994) NMDA receptor antagomsts targeted at the distinct recognition or modulatory sites have been shown to blunt glutamate induced inlury in cell cultures, to prevent hypoxia-induceo EPSP decrease ~n rat hippocampal shces and to salvage a large portion of bral~ tissue in anlma~ mooels of stroke and cerebral trauma The degree of neu roprotecton afforded by these d r u g s l s v e r y l a r g e For instance, oEzocilplne (! mg/kg ip) and eliprodl (1 '-'g/kg ivl administered following permanert occlusion of the m~adle cereora, artery ~n the rat diminish by --~50% the total infarction v o l u ~ e Sire!lady, "epeated administration of ehprodd {10 mg/kg ~p) to rats sub}ectec to lateral fluid percussion of the parietal cortex reduces by 60% the volume of the t~aumatic lesion. As a general r u e in models of focal iscqemla, charnel olockers and eliprodd are more potent and efficacious than NMDA recogmtloq or glycn'e site aqtagomsts probably because of their better brain per'etratlon Unfortunately, NMDA receptor antagomsm can be accompanied by a bat terv of undesirable side effects including, m va'y ng degree, phencyclidine like psychostlmulation, amnesia rntrinsic qeurotoxlclty on specific cortica! neuron populations, hypertenslor ~andtachycardla These effects arepartlc ularly intense with certain NMDA chaqne! blockers {although less so irq the case of dextrcrphan like comoounds), and can also be observed with coopetitwe NMDAantagonlsts Glycineslteantagon~stsappeartohavea more favourable p~ofile at neuroprotective doses, w6ile eliprodil is completely devoid of such side effects {G~roux et at 1994) Trle select:ve blockage by eliprodil (and to some extent non stimulant channel blockers} o{ a NMDA receptor sub~,ype may account for its favorable safety index as compared to conventional NMDA antagonist which block indtscriminately el' NMDA receptor subtypes In conclusion, subtype se:ect~ve NMDA receptor antagonists may rep resent a promtsing approach for the medical treatment of acute ischemlc stroke, CNS trauma and poss bly chrorlic neurodegenerative diseases Sev eral NMDA antagonists including CNS 1102, ehprodil, CGS 19755 and dex trorphan are now being evaluated in early treatment clinical trials m stroke o r bra~n trauma and useful theraptes may emerge with those drugs
References Carter, C. Benavides. ~ Dana, C Schoemaker. H Perrault, G r Sanger DJ and Scatto~ B (1991i Non comDetibve NMDA receptor antagonists acting on the poiyam~ne site In ExcltatoryAmlno Acid Antagonists, B S Meldrum Ed, Blackwell Scientific London pp 130 169 Glroux, C Rosen, P and Scatton, B 1994) :~reshn~calpharmacology and ci nical safew prof,le of eliprodil, an atypical NMDA receptor antagonist In Pharmacology of Cere bra Ischem~a J Krieglstein, H Oberplchler Schwenk (Edsi Wissenschaftlche Ve" lagsgesetlschaft mbH Stuttgart, pp 643 648 Nakamshi, S (1992} Molecular dive,s,ty of glutamate receptors and implicat~ors {o, bratP function Science, 258 597 603 Scatton, B (1993) The NMDA recepto ~complex Fundam Clin Pha,macol 7 389 400 Scatton, B (1994) Bxc;tatory amino ac*d recentor antagonists a novel treatment :or ~schemic cereb'ovascular d~seases L~le So, 55 2115 212/*
ogy lhe need for absolute certainty of anti-ischaemic benefit requires that a number of separate tssues are addressed in preclinlcal development. An Nllproved acute histological outcome with the drug must be demonstrated to demonstrate that drug efficacy is independent of physiological artefact. Chronic h~stological outcome must be shown to have been improved by drug ~ntervent~on to dispel doubt that the drug simply slows the maturation of tne ischaemic damage. Some investigations of anti-ischaemic efficacy must be performed in a gyrencephalic species Jn case the mechanism of interest contributes disproportionately to outcome in the lissencephalic brain {spreading depression being an obvious example). The question of therapeutic time w i n d o w must be addressed but it must be appreciated that the qmportant time w i n d o w is that in man and not that in young, healthy ammals with a carefully controlled single ischaemic event. Considerable attention needs to be directed to drug pharmacokinetics not just plasma revels, but CNS levels of the drug around the zone of ischaemia. Once unequivocal anti-ischaemic effect is demonstrated, the central issue becomes safety While preclinical investigations have focused on the adverse effects of the new neuroprotective drugs in the CNS, their effects on cardiovascular and respiratory function are of equal ior greater) importance in the clinical evaluation. NMDA Receptor Blockade. in animal models of focal cerebral ischaemia, unequivocal benefit has been demonstrated after NMDA receptor blockade ,n every species examined {baboon, cat, rabbit, rat, mouse). The maximum reduction (typically 50%) in the volume of infarcted tissue is broadly similar ~rrespective of whether the drug acts at the agonist recognition site of the NMDA receptor complex (e.g. D-CPPene, CGS 19755)the ion channel site {eg. MK-801, CNS 1102, or remacemide hydrochlobde) or polyamine site {eg. eliprodil) (Chen et el., 1991; McCulloch, 1992: Bannan et el., 1994). Although the evidence of anti ischaemic efficacy of the NMDA receptor antagon,sts is overwhelming, the clinical evaluation of these drugs was delayed oy concern over their effects on CNS function such as vacuole formation, immediate early gene or heat shock protein expression in the cingulate cortex, hypermetabolism visualised with deoxyglucose autoradiography e t c (McCulloch, 1992) AMPA Receptor Blockade. There is growing evidence that AMPA recep tor blockers (NBQX, YM 900, LY-293558, GYKI 42566} also reduce damage a~ter experimental focal ischaemia but the volumes of tissue saved are typically smaller than for NMDA antagonists {Bullock et el, 1994), The effects of AMPA receptor antagonists on CNS function bear little resemblance to t~ose of NMDA antagonists. The critical utility of AMPA receptor antagonists ~s constrained by their potent respiratory depressant actions. {Browne and McCulloch, 1994). Glutamate Release lnhlT~itors. Attenuation of the elevations in extracelI~.lar glutamate concentrations which occur in cerebral ischaemia has the advantage of reducing the resultant activation of both NMDA and AMPA receptors There is growing evidence that inhibition of glutamate release wa diverse strategies (Na + and Ca ++ channel blockers, kappa opioid ago n~sts etc) can reduce ischaemic damage in experimental focal ischaemia (Mackay et el, 1993). The effects on CNS function of glutamate release inhibit on are less pronounced than NMDA or AMPA receptor blockade. In summary, it is worth emphasising that for the first time, a class of orugs is entenng clinical trials in human ischaemia based on a certainty of anti-ischaemic benefit in animal models and the data emerging from these chnical trials is most encouraging.
References Glutamate antagonists in animal models of cerebral ischaemia We//come Surgical Institute ~ Hugh Fraser Neuroscience Labs, Un/versity o/Glasgow, Garscube Estate, Bearsder-, Road, Glasgow G6t tOH, Scotland, United Kingdom James McCulloch
Animal models of cerebral ischaerma provide the c~ucla hnk between tqe hypothesis that excttotoxic mecr~anlsms contnbute to iscrlaemlc injury to neurones and the development of new therapeutic strategies for man Models of focal cerebral iscnaem~a (typically occlusion of a middle cerebral artery} have been the most powerful tools in characterising neurop'otechve efficacy of glutamate receptor antagonists and data in these models underpin the current chnical use o~ NMDA antagomsta and glutamate release innibitors In contrast, models of gtobai cerebral ischaemia have not been particularly valuable in antl-lscnaemlc orug development because of con cerP of the pertinence of the selectwe neuronal necrosis of global ischaemia to the processes of infarctton iq stroke (McCulloch. 1992} Anti-ischaem/c Drug Development. A Strategy Although many expert mental endpoints can provide mechanistic insight, the deflmtwe endpo nt for a putahve neuroprotective dr,~g must always be quantltattve blstopat~'oi
Barman, BE, Graham, DI, Lees, K R and McCulloch. J {1994). Neuroprotective effect of remacemide hydroch!oride in focal cerebral ischaemia in the cat Brain Research 664 271-275 Browne, S E 8 McCulloch, J (1994) AMPA receptor antagonists end local cerebral glucose utilization in the rat Brain Research 641, 10 20 Bullock, R Graham, DI, Swanson, S fi McCulloch, J {1994) Neuroprotective effect of the AMPA receptor antagonist LY-293558 in focal cerebral ischemia in the cat. Journal of Cerebrai Blood Flow and Metabolism 14, 46£:~471 McCulloch, J (1992) Excitatory amino acid antagonists and their potential for the treatment of ischaemic brain damage in man. British Journal of Clinical Pharmacology 34, 106.114 Mackav, K B , Kusumoto, K, Graham, D I and McCulloch, J (1993) Focal cerebral ischemia in the cat: pretreatment with a kappa-t opioid receptor agonist, CI-977 Brain Research 618, 213 219 Chen, M Bullock R , Graham, D I , Frey, B, Lowe, D and McCulloch,J (1991). Evaluation of a competitive NMDA antagonist (D-CPPenelin feline focal cerebral ischemia Annals of Neurology 30, 62 70