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Several new benzodiazepines selectively interact with a benzodiazepine receptor subtype
Neurosc/emce Letters. 38 0983) 73-78 El~.-vier Scientific Publisher5 Ireland Ltd.
73
SEVERAL NEW B E N ~ I A Z E P I N E S SELECFlVEL¥ IN~ILACT W I T H A BENZODIAZEPINE ~ O R SUBTYPE
WERNER SIEGHART
Psyckiatrische Universit~tskliniko Department of Biochemical Psychiut~, Lazarengasse 14, 1097 Vienna (Austrial (Received ~,nuary 4th, 1983; Revised version received March 28th. 1983; Accepted April 7th0 1983)
Key words: benzodiazepine receptor subtype - [3H]flunitrazepam - quazepam - SCH 15-725 - SCH 23-324 The potency of several new bcnzodiazcpines as inhibitors o f IJH]flunitrazepam binding was investisated in membranes from rat cerebellum or hippocampus. It was found that quazepam and two of its metabolites have a hilgher affinity for benz~iazgpine receptors in cerebellum than for those in hippocampus, indicating a preferential interaction of these compounds with BZt-receptors. Other experimems indicated that these compounds have benzodiazgpine algonistic properties similar to diazepam or flunitrazepam. Thus, for the first time, benT~odiazepines have been identified which differentially interact with different benzndiazepine receptors.
A large number of different benz~iazepines have been synthesized in an effort to separate the anxiolytic, anticonvulsant, hypnotic and muscle relaxant effects of these drugs 11I]. However, all the benzodiazepines investigated more or less had the same spectrum of activity su88estin8 that all ~hese drubs interact with the same biologically active sites and that the diverse actions of benzodiazepines might invariably be linked with each other !11,121. This was seemingly confirmed later on when it was demonstrated that all benzodiazepines so far investigated had similar affinities for different benzodiazepine receptor binding sites !1,6] in brain membranes and that their affinity for these sites correlated highly with their clinical potency 112]. Since different receptors are possibly associated with different actions of benzodiazepines, compounds selectively interacting with a subtype of benzodiazepine receptors might have more selective clinical properties. In the present study, a new benzodiazepine, quazepam [4], together with its three main metabolites was investigated, and it was demonstrated that quazepam and two of its metabolites preferentially interact with a benzodiazepine receptor subtype. The potency of various compounds as inhibitors of [3H]flunitrazepam binding was investigated in membranes from rat cerebellum or hippocampus. As shown in Table !, quazepam and its metabolites SCH 15-725, SCH 23-324 and Ro 5-3367 (Fig. 1) are potent inhibitors of [3H]flunitrazepam binding to cerebellar membranes.
74
However, in comrast to Ro 5-3367, dla,flpam and f l u n ~ - which have a similar potency for inhibition of | ~ l ] f l u n ~ binding in ccrehellum and hip-
~ ~ ~ observed weviously forCI 2188/216] or the methyl-, ethyl- or propyl-esters of jt~-carboline-3-carboxylate (.8-CC'M, ~8-42CEor ~CCP) [1,2] (Table !). This l e d t o the hypothe~ that C'I 218 872 and the ~¢arbolinc-3-carboxylateesters selectively bind to a mbtyp¢ of benzodiazcpine recgptorg which is present predominantly in cerebellum ('cerebellar type', 'type-F- or BZt-benzodiazepine receptor) but not as abundantly in hippocampus [1,2,6,101. Thus, the present results suggest that quazcpam, ~ 15-725 and ~ 23-324 also seem to preferentially interact with BZt receptors. Furthermore these data indicate that the selectivity of benzodiazepines for BZt receptors is determined by the trifluoroethyl side-chain of the henzodiazepin¢ molecule since N-dcsalkyl-flurazepam (Ro 5.3367) does not ¢xTABLE I POTENCY OF VARIOUS BENZODIAZEPINE RECIEPT(~i *. ' ,GANDS AS INHIBITORS OF SPECIFIC [JHIFLUNITRAZEPAM BINDING TO MEM;.~,..,,.ES FROM CEREBELLUM OR HIPPOCAMPUS Membrane aliquots, derived from 2 ms of cerebellar or hippocampal tisstu~ [10J, were incubated for 90 rain at OoC with 2 nM [JHJflunitrat~am (84.8 Ci/mmol, New England Nuclear) in I ml of an incubation medium which contained 150 mM NaCI, 50 mM Tris~'itrate buffer, pH 7. I, and various concentrations of ben/odiazcpine receptor littandl to be t~ted. The ~tmpl~ were then filtered under vacuum through Whatman GF/B filters and immediately washed twice with 5 ml ice-cold buffer. Itm._dioactivityon the filters was measured by liquid icintillmion counting. Non-specific binding was determined in the pm~mce of .~uM unlabeled diazepmn and was subtrmeted from total binding to give specific binding. The ¢oucen. trations giving half maximal inhibition o f [JH]flmdtrazepam bindin$ (IC~o) are mean values ± S.E.M. from 3-5 independent experiments performed in duplicate In several preliminary ¢upcrimoms, the inhibition of [~H]flunitrazcpam binding by q ~ , . ~ H 15-725 or C1218 872 was investipted at 37°C. Although ICso values of these compounds were significantly larger at this higher temtperature, the differcnees in IC~ values between cerebellum and hippocampus were again demonstrated for all these compounds at this more physiological temperature.
Ligand
Cerebellum IC~ (nM)
Hippocampus IC~o(nM) ICso ratio hippocampus/cerebellum
Quazepam SCH 15-725 SCH 23-324 Ro 5-3367 Diazepam Flunitrazepam CI 2188"/2 8-CCE 8-CCM
29.7± 3.4 12.8± 3.9 30.9± !.8 2.8± 0.7 12.8± 0.8 2.9± 0.4 190.3 ± 15.5 0.9+_ 0.1 0.8± 0.04
163.7± 18.4 101.4± 15.1 140.0± 5.6 3.1± 0.6 11.5± 0.3 2.7± 0.1 1296.7 ±60.6 4.5± 0.3 3.6± 0.5
5.5 7.9 4.5 I.i 0.9 0.9 6.8 5.0 4.5
75 bibit differential affmity for benzodiazepine receptors in cerebellum or hippocampus. Quazcpam, as well as SC'H 15-725 or SCH 23-324, boxylatc-mters [I], are competitive inhibitors of cerebellum and mixed type competitive inbihitors of [3H]flunitrazepam binding in hippocampns (experiments not shown). In addition, inhibition of [3H]flunitrazepam binding by these compounds resulted in Hill coefficients close to unity in cerebellum and I~ween 0.7 and 0.8 in hippocampus. These results support the hypothesis of the existence of different benz~iazepine receptors in cerebellum and hippocampns [1,6,10], and are in a~cement with the conclusion of a differential interaction of quazepam, SC'H 15-725 and $CH 23-324 with different benzodiazepine receptors. Recently, it was demonstrated that B-CCM has convulsant and 0-CCE proconvulsant properties in animals [2,8]. In addition, it was demonstrated that 0-CCE 12,8] and Ro 15-1788 [7], by binding to benzodiazepine receptors, antagonized the anxiolytic, anticonvulsant, muscle relaxant and sedative hypnotic actions of ben-
N
N
O
m,
CI'
CA
Qua~pem
SCH15 726
1 0
a
SCH 2 3 3 2 4
Ro 5 - 3 3 6 ?
Fig. !. Structural formulae of q~az_,~pam (7-chloro-1-(2,2,24rifluoroethyl)-1,3-dihydro-5-(2-fluoro-
phenyi)-2H-I.4-benzodiazepin-2-thione) and its metabolites SCH 15-725 (7-chloro-1-(2,2,2-trifluoroethyl)-l.3-dihydro-5-(2-fluorophenyl)-2H-1o4-benzodiazepin-2-one), SCH 23-324 (7-chlogo-1-(2,2,2-triflu•r•ethy•)-3•hydr•xy-542-flu•r•pheny•)••.3-dihydr••2H•••4-benz•diaz•pin•2••n•) and Ro 5-3367 (7-chloro-5-(2-flurophcnyl)-311-1.4-benzodiazcpin-2(IH)-one)o
76
zodiazepines in both experimental animal; and man. From this it was concluded that these compounds bind to b e n z o d i a ~ t e receptors in a way differen~ from ben-
bammazqgne ',n e u r o t r ' a n s m ~
sion [2,3,71. Several ~mple in vitro test s/stems were develot.~l which seem to distinguish between compounds eahibitin8 benmdiazepine alwni~e or antagonistic properties D,5,71. in the present study, one of these t a t sygems [71 ~as applied to investigate the properties of quazepam and its metabolites. In this test sygem, inhibition by several compounds o f [Jl-l]Ro 15-1788 binding to cerebellar membranes was investigated in the absence or presence of 100 FM GABA (~. As demonstrated previously [3,71, the inhibitory potency of benzodiazepine agonists is enhanced by GABA while that of antagonists like Ro 15-1788 remains unaltered or is even decreased by this amino acid in the case of ~ C C E or 0-CCM. These results were confirmed in the present study (Table 11), and it was demonstrated that the inhibitory potency of quazepam, SCH 15-725 and SCH 23-324, as well as that of diazepam, flunitrazepam and CI 218 872, is enhanced in the presence of 100/~M GABA indicating a similar type of interaction with benzodiazepine receptors of all these compounds. Thus, the present results indicate that quazepam and its metabolites SCH 15-725 and SCH 23-324 have a high affinity for benzodiazepine receptors and have ben-
TABLE II I'OTENt'Y OF VARIOUS BENZODIAZEPINE RECEPTOR LIGANDS AS INHIBITORS OF SPECIFIC {*HIRo I.~-1788 BINDING TO CEREBEiI+AR MEMBRANES IN THE ABSENCE OR PRESENCE OF GABA
Binding of 1,5 nM[~Hl~o 15-1788 (87,5 Ci/mmol. New England Nuclear) was determined in membrane~ from rat cerebellum in the absence or presence of 100 ~M GABA and of various concentrations of benzodiazepine receptor ligands to be lested as described in legend to Table I. lose values are means ± S.E.M. from 3~5 independent experiments performed in duplical¢. Similar GABA ratios were obtained when the experiments ~¢r¢ performed in rat hippocampus, indicating that all the compounds it~vestigated have similar agonisti¢ or amagoni,qic properties on different benzodiazepine receptors. IC~ (nM) Ligand
- GABA
+ GABA
GABA ratio
Quazepam SCH 15~725 SCH 23-324 Diazepam Flunitrazepam CI 2 1 8 872 Ro 15-1788 ~-CCE ~-CCM
42.1± 2.4 19.5± 2.3 49.8 ± !.8 29,8_+ 3.6 10.5 + 0.9 261.6 ± 18.8 2.24- 0.2 0.9-" O.I !.9_+ O.I
20.14- !.3 8.14- !.6 23.6 4- 4. I 13.2± !.1 4.3 ± 0.5 120.04- 10.4 , 2.0± 0.1 i.5_+ O.I 3.44- 0.2
2.1 2.4 2. I 2.3 2.4 2.2 !.1 0.6 0.5
77
zodiazepine agonistic properties similar to diazepam or flunitrazepam. In contrast to these latter substances, however, quazepam, SCH 15-725 and SCH 23-324 seem
structure of the benzodiazepine m o i r e new benzodiazepines might emerge with even higher selectivity for indi~dual receptors, Data from animal studies indicate that quazepam has a low toxicity and pharmacological properties which are similar to those of other benzodiazepines [9]. When quazepam was compared with flurazepam, assessing sedation and ataxia, quazepam appeared to be somewhat more potent than flurazepam, but caused less ataxia [9]. Preliminary data from clinical studies indicated that quazepam is a safe and effective hypnotic agent in short-term and intermediate-term treatment os subjects suffering from chronic insomnia [4]. However, further studies should be performed investigating more carefully the immediate actions of low concentrations of quazepam in animals and man. Since quazepam is rapidly metabolized to SCH 15-725 but only slowly further metabolized to Ro 5-3367 (Scherin8 Corp., data on file), the immediate actions of low concentrations of qnazepam might preferentially reflect interactions with BZt receptors. In any case, the present results should stimula~c research in identification of compounds selectively interacting with specific benzodiazepine receptor subtypes, and this possibly might lead to benzodiazepines with more selective actions. This work was supported by 'Fonds zur F6rderung der wissenschaftlichen Forschung in Osterreich'. ! thank A. Schuster and G. Drexler for excellent technical assistance and Dr. Mauracher, EssexChemie, Switzerland, for kindly providing me with quazepam, SCH 15-725 and $CH 23-324. I Braestrup, C. and Nielsen, M.J., [JHlPropyl-/3-carboline-3-carboxylate as a selective radioligan,', for the BZ, benzndiazepine receptor subclass, J. Neurochem., 37 (1981) 333-341. 2 BraeSlrup, C. and Nielsen, M.J., GABA reduces bi,~ding of 13H]melhyI-B-carboline-3-carboxylate to brain benzodiazepine receptors, Nature (Lond.), 294 (1981) 472-474. 3 Braestrup, C., Schmiechen, R., Neff, G., Nielsen, M.J. and Petersen, E.N., Interactions of convulsive ligands with benzcdiazepine receptors, Science, 216 0982) 124t-1243. 4 Kales, A., Scharf, M.B., Soldatos, C.R., Bixler, E.O., Bianchi, S.B. and Schweitzer, P.K.J., Quazepam, a new benzmliazepine hypnotic: intermediate-term sleep laboratory evaluation, J. clin. Pharmacolo, 20 (1980) 184-192. 5 Karobath, Mo and SupavJlai, P., Distinction of benzodiazepine agonists from antagonists by photoaffinity labeling of benzodiazepine receptors in vitro, Neurosci. Lett., 31 (1982) 65-69. 6 Klepner, C.A., Lippa, A.S., Benson, D.I., Sano, M.C. and Beer, B., Resolution of two biochemically and pbagmacolosicaHy distinct benzodiazepine receptors, Pharmacol. Biochem. Beh~v., Ii (1979) 457-462. 7 MOhler, H. and Richards, J.G., Agonist and antagonist benzodiazepine receptor interaction in vitro, Nature (Lond.), 294 (1981) 763-765.
78
$ C~tiq, N.R. and Jones, BA., The ~ cad~i~ad, o ~ L ~ , Euroo. J. i q m ~ o L ,
and ~ a s i n 8
~,ffecls of ethyl-~-
68 (1~0) 311t-~12.
14f~-!462.
(Load.). 2116(1~0) 211S-2S7, . . . . .... I! S~.rnlmch, L . H , The IN~zcli~lKme stm'y, in R.G, Prie~ t3. V ' ~ . IL Amrein andM, Skrcla ff.ds.), I 1 ~ , Today and Tomon'ow, MT'P Press, 1980, pp. 5--17. 12 Tallman. J.F., Paul. S.M.. Skoiaigk, P. and Gallal~. D.W., ~ for the age of ammiety: Isharmacomsy of the I m u ~ i m m ~ m . Sciaze. 207 (tee0) 274-2~.
73
SEVERAL NEW B E N ~ I A Z E P I N E S SELECFlVEL¥ IN~ILACT W I T H A BENZODIAZEPINE ~ O R SUBTYPE
WERNER SIEGHART
Psyckiatrische Universit~tskliniko Department of Biochemical Psychiut~, Lazarengasse 14, 1097 Vienna (Austrial (Received ~,nuary 4th, 1983; Revised version received March 28th. 1983; Accepted April 7th0 1983)
Key words: benzodiazepine receptor subtype - [3H]flunitrazepam - quazepam - SCH 15-725 - SCH 23-324 The potency of several new bcnzodiazcpines as inhibitors o f IJH]flunitrazepam binding was investisated in membranes from rat cerebellum or hippocampus. It was found that quazepam and two of its metabolites have a hilgher affinity for benz~iazgpine receptors in cerebellum than for those in hippocampus, indicating a preferential interaction of these compounds with BZt-receptors. Other experimems indicated that these compounds have benzodiazgpine algonistic properties similar to diazepam or flunitrazepam. Thus, for the first time, benT~odiazepines have been identified which differentially interact with different benzndiazepine receptors.
A large number of different benz~iazepines have been synthesized in an effort to separate the anxiolytic, anticonvulsant, hypnotic and muscle relaxant effects of these drugs 11I]. However, all the benzodiazepines investigated more or less had the same spectrum of activity su88estin8 that all ~hese drubs interact with the same biologically active sites and that the diverse actions of benzodiazepines might invariably be linked with each other !11,121. This was seemingly confirmed later on when it was demonstrated that all benzodiazepines so far investigated had similar affinities for different benzodiazepine receptor binding sites !1,6] in brain membranes and that their affinity for these sites correlated highly with their clinical potency 112]. Since different receptors are possibly associated with different actions of benzodiazepines, compounds selectively interacting with a subtype of benzodiazepine receptors might have more selective clinical properties. In the present study, a new benzodiazepine, quazepam [4], together with its three main metabolites was investigated, and it was demonstrated that quazepam and two of its metabolites preferentially interact with a benzodiazepine receptor subtype. The potency of various compounds as inhibitors of [3H]flunitrazepam binding was investigated in membranes from rat cerebellum or hippocampus. As shown in Table !, quazepam and its metabolites SCH 15-725, SCH 23-324 and Ro 5-3367 (Fig. 1) are potent inhibitors of [3H]flunitrazepam binding to cerebellar membranes.
74
However, in comrast to Ro 5-3367, dla,flpam and f l u n ~ - which have a similar potency for inhibition of | ~ l ] f l u n ~ binding in ccrehellum and hip-
~ ~ ~ observed weviously forCI 2188/216] or the methyl-, ethyl- or propyl-esters of jt~-carboline-3-carboxylate (.8-CC'M, ~8-42CEor ~CCP) [1,2] (Table !). This l e d t o the hypothe~ that C'I 218 872 and the ~¢arbolinc-3-carboxylateesters selectively bind to a mbtyp¢ of benzodiazcpine recgptorg which is present predominantly in cerebellum ('cerebellar type', 'type-F- or BZt-benzodiazepine receptor) but not as abundantly in hippocampus [1,2,6,101. Thus, the present results suggest that quazcpam, ~ 15-725 and ~ 23-324 also seem to preferentially interact with BZt receptors. Furthermore these data indicate that the selectivity of benzodiazepines for BZt receptors is determined by the trifluoroethyl side-chain of the henzodiazepin¢ molecule since N-dcsalkyl-flurazepam (Ro 5.3367) does not ¢xTABLE I POTENCY OF VARIOUS BENZODIAZEPINE RECIEPT(~i *. ' ,GANDS AS INHIBITORS OF SPECIFIC [JHIFLUNITRAZEPAM BINDING TO MEM;.~,..,,.ES FROM CEREBELLUM OR HIPPOCAMPUS Membrane aliquots, derived from 2 ms of cerebellar or hippocampal tisstu~ [10J, were incubated for 90 rain at OoC with 2 nM [JHJflunitrat~am (84.8 Ci/mmol, New England Nuclear) in I ml of an incubation medium which contained 150 mM NaCI, 50 mM Tris~'itrate buffer, pH 7. I, and various concentrations of ben/odiazcpine receptor littandl to be t~ted. The ~tmpl~ were then filtered under vacuum through Whatman GF/B filters and immediately washed twice with 5 ml ice-cold buffer. Itm._dioactivityon the filters was measured by liquid icintillmion counting. Non-specific binding was determined in the pm~mce of .~uM unlabeled diazepmn and was subtrmeted from total binding to give specific binding. The ¢oucen. trations giving half maximal inhibition o f [JH]flmdtrazepam bindin$ (IC~o) are mean values ± S.E.M. from 3-5 independent experiments performed in duplicate In several preliminary ¢upcrimoms, the inhibition of [~H]flunitrazcpam binding by q ~ , . ~ H 15-725 or C1218 872 was investipted at 37°C. Although ICso values of these compounds were significantly larger at this higher temtperature, the differcnees in IC~ values between cerebellum and hippocampus were again demonstrated for all these compounds at this more physiological temperature.
Ligand
Cerebellum IC~ (nM)
Hippocampus IC~o(nM) ICso ratio hippocampus/cerebellum
Quazepam SCH 15-725 SCH 23-324 Ro 5-3367 Diazepam Flunitrazepam CI 2188"/2 8-CCE 8-CCM
29.7± 3.4 12.8± 3.9 30.9± !.8 2.8± 0.7 12.8± 0.8 2.9± 0.4 190.3 ± 15.5 0.9+_ 0.1 0.8± 0.04
163.7± 18.4 101.4± 15.1 140.0± 5.6 3.1± 0.6 11.5± 0.3 2.7± 0.1 1296.7 ±60.6 4.5± 0.3 3.6± 0.5
5.5 7.9 4.5 I.i 0.9 0.9 6.8 5.0 4.5
75 bibit differential affmity for benzodiazepine receptors in cerebellum or hippocampus. Quazcpam, as well as SC'H 15-725 or SCH 23-324, boxylatc-mters [I], are competitive inhibitors of cerebellum and mixed type competitive inbihitors of [3H]flunitrazepam binding in hippocampns (experiments not shown). In addition, inhibition of [3H]flunitrazepam binding by these compounds resulted in Hill coefficients close to unity in cerebellum and I~ween 0.7 and 0.8 in hippocampus. These results support the hypothesis of the existence of different benz~iazepine receptors in cerebellum and hippocampns [1,6,10], and are in a~cement with the conclusion of a differential interaction of quazepam, SC'H 15-725 and $CH 23-324 with different benzodiazepine receptors. Recently, it was demonstrated that B-CCM has convulsant and 0-CCE proconvulsant properties in animals [2,8]. In addition, it was demonstrated that 0-CCE 12,8] and Ro 15-1788 [7], by binding to benzodiazepine receptors, antagonized the anxiolytic, anticonvulsant, muscle relaxant and sedative hypnotic actions of ben-
N
N
O
m,
CI'
CA
Qua~pem
SCH15 726
1 0
a
SCH 2 3 3 2 4
Ro 5 - 3 3 6 ?
Fig. !. Structural formulae of q~az_,~pam (7-chloro-1-(2,2,24rifluoroethyl)-1,3-dihydro-5-(2-fluoro-
phenyi)-2H-I.4-benzodiazepin-2-thione) and its metabolites SCH 15-725 (7-chloro-1-(2,2,2-trifluoroethyl)-l.3-dihydro-5-(2-fluorophenyl)-2H-1o4-benzodiazepin-2-one), SCH 23-324 (7-chlogo-1-(2,2,2-triflu•r•ethy•)-3•hydr•xy-542-flu•r•pheny•)••.3-dihydr••2H•••4-benz•diaz•pin•2••n•) and Ro 5-3367 (7-chloro-5-(2-flurophcnyl)-311-1.4-benzodiazcpin-2(IH)-one)o
76
zodiazepines in both experimental animal; and man. From this it was concluded that these compounds bind to b e n z o d i a ~ t e receptors in a way differen~ from ben-
bammazqgne ',n e u r o t r ' a n s m ~
sion [2,3,71. Several ~mple in vitro test s/stems were develot.~l which seem to distinguish between compounds eahibitin8 benmdiazepine alwni~e or antagonistic properties D,5,71. in the present study, one of these t a t sygems [71 ~as applied to investigate the properties of quazepam and its metabolites. In this test sygem, inhibition by several compounds o f [Jl-l]Ro 15-1788 binding to cerebellar membranes was investigated in the absence or presence of 100 FM GABA (~. As demonstrated previously [3,71, the inhibitory potency of benzodiazepine agonists is enhanced by GABA while that of antagonists like Ro 15-1788 remains unaltered or is even decreased by this amino acid in the case of ~ C C E or 0-CCM. These results were confirmed in the present study (Table 11), and it was demonstrated that the inhibitory potency of quazepam, SCH 15-725 and SCH 23-324, as well as that of diazepam, flunitrazepam and CI 218 872, is enhanced in the presence of 100/~M GABA indicating a similar type of interaction with benzodiazepine receptors of all these compounds. Thus, the present results indicate that quazepam and its metabolites SCH 15-725 and SCH 23-324 have a high affinity for benzodiazepine receptors and have ben-
TABLE II I'OTENt'Y OF VARIOUS BENZODIAZEPINE RECEPTOR LIGANDS AS INHIBITORS OF SPECIFIC {*HIRo I.~-1788 BINDING TO CEREBEiI+AR MEMBRANES IN THE ABSENCE OR PRESENCE OF GABA
Binding of 1,5 nM[~Hl~o 15-1788 (87,5 Ci/mmol. New England Nuclear) was determined in membrane~ from rat cerebellum in the absence or presence of 100 ~M GABA and of various concentrations of benzodiazepine receptor ligands to be lested as described in legend to Table I. lose values are means ± S.E.M. from 3~5 independent experiments performed in duplical¢. Similar GABA ratios were obtained when the experiments ~¢r¢ performed in rat hippocampus, indicating that all the compounds it~vestigated have similar agonisti¢ or amagoni,qic properties on different benzodiazepine receptors. IC~ (nM) Ligand
- GABA
+ GABA
GABA ratio
Quazepam SCH 15~725 SCH 23-324 Diazepam Flunitrazepam CI 2 1 8 872 Ro 15-1788 ~-CCE ~-CCM
42.1± 2.4 19.5± 2.3 49.8 ± !.8 29,8_+ 3.6 10.5 + 0.9 261.6 ± 18.8 2.24- 0.2 0.9-" O.I !.9_+ O.I
20.14- !.3 8.14- !.6 23.6 4- 4. I 13.2± !.1 4.3 ± 0.5 120.04- 10.4 , 2.0± 0.1 i.5_+ O.I 3.44- 0.2
2.1 2.4 2. I 2.3 2.4 2.2 !.1 0.6 0.5
77
zodiazepine agonistic properties similar to diazepam or flunitrazepam. In contrast to these latter substances, however, quazepam, SCH 15-725 and SCH 23-324 seem
structure of the benzodiazepine m o i r e new benzodiazepines might emerge with even higher selectivity for indi~dual receptors, Data from animal studies indicate that quazepam has a low toxicity and pharmacological properties which are similar to those of other benzodiazepines [9]. When quazepam was compared with flurazepam, assessing sedation and ataxia, quazepam appeared to be somewhat more potent than flurazepam, but caused less ataxia [9]. Preliminary data from clinical studies indicated that quazepam is a safe and effective hypnotic agent in short-term and intermediate-term treatment os subjects suffering from chronic insomnia [4]. However, further studies should be performed investigating more carefully the immediate actions of low concentrations of quazepam in animals and man. Since quazepam is rapidly metabolized to SCH 15-725 but only slowly further metabolized to Ro 5-3367 (Scherin8 Corp., data on file), the immediate actions of low concentrations of qnazepam might preferentially reflect interactions with BZt receptors. In any case, the present results should stimula~c research in identification of compounds selectively interacting with specific benzodiazepine receptor subtypes, and this possibly might lead to benzodiazepines with more selective actions. This work was supported by 'Fonds zur F6rderung der wissenschaftlichen Forschung in Osterreich'. ! thank A. Schuster and G. Drexler for excellent technical assistance and Dr. Mauracher, EssexChemie, Switzerland, for kindly providing me with quazepam, SCH 15-725 and $CH 23-324. I Braestrup, C. and Nielsen, M.J., [JHlPropyl-/3-carboline-3-carboxylate as a selective radioligan,', for the BZ, benzndiazepine receptor subclass, J. Neurochem., 37 (1981) 333-341. 2 BraeSlrup, C. and Nielsen, M.J., GABA reduces bi,~ding of 13H]melhyI-B-carboline-3-carboxylate to brain benzodiazepine receptors, Nature (Lond.), 294 (1981) 472-474. 3 Braestrup, C., Schmiechen, R., Neff, G., Nielsen, M.J. and Petersen, E.N., Interactions of convulsive ligands with benzcdiazepine receptors, Science, 216 0982) 124t-1243. 4 Kales, A., Scharf, M.B., Soldatos, C.R., Bixler, E.O., Bianchi, S.B. and Schweitzer, P.K.J., Quazepam, a new benzmliazepine hypnotic: intermediate-term sleep laboratory evaluation, J. clin. Pharmacolo, 20 (1980) 184-192. 5 Karobath, Mo and SupavJlai, P., Distinction of benzodiazepine agonists from antagonists by photoaffinity labeling of benzodiazepine receptors in vitro, Neurosci. Lett., 31 (1982) 65-69. 6 Klepner, C.A., Lippa, A.S., Benson, D.I., Sano, M.C. and Beer, B., Resolution of two biochemically and pbagmacolosicaHy distinct benzodiazepine receptors, Pharmacol. Biochem. Beh~v., Ii (1979) 457-462. 7 MOhler, H. and Richards, J.G., Agonist and antagonist benzodiazepine receptor interaction in vitro, Nature (Lond.), 294 (1981) 763-765.
78
$ C~tiq, N.R. and Jones, BA., The ~ cad~i~ad, o ~ L ~ , Euroo. J. i q m ~ o L ,
and ~ a s i n 8
~,ffecls of ethyl-~-
68 (1~0) 311t-~12.
14f~-!462.
(Load.). 2116(1~0) 211S-2S7, . . . . .... I! S~.rnlmch, L . H , The IN~zcli~lKme stm'y, in R.G, Prie~ t3. V ' ~ . IL Amrein andM, Skrcla ff.ds.), I 1 ~ , Today and Tomon'ow, MT'P Press, 1980, pp. 5--17. 12 Tallman. J.F., Paul. S.M.. Skoiaigk, P. and Gallal~. D.W., ~ for the age of ammiety: Isharmacomsy of the I m u ~ i m m ~ m . Sciaze. 207 (tee0) 274-2~.