Tricyclic Azaergoline Analogues: Synthesis, Structural Modifications,and Pharmacological Studies

a) Institut fUr Phannazie und Lebensmittelchemie der Universitiit Miinchen, Sophienstra6e 10, SOOO Miinchen 2, Germany

b) Abteilung Biochemie, Boehringer Ingelheim KG, 6507 IngelheimlRhein, Germany

As an extension of previous investigations on synthesis and dopamineautoreceptor activity of bicyclic ergoline analogues the tricyclic azaergolineanalogues 9a and 9b were synthesized. Funhermore, the geometry of thearomatic ~-ethylamine moiety of 9a,b was modified by stereoselective con-struction of the cycloheptenyl fused pyrazolopyridine derivative 7 and theaminomethyl substituted tricycle 10. Binding affinity of these compounds atdopamine (OA) receptor sites was investigated employing rat striatum ho-mogenate: The compounds reveal modest to wealc, but selective binding to adopamine 0-2 receptor when it is labelled with the OA-autoreceptor agonist[3HI-SNO 919. In vivo studies with mice showed that 7, 9a,b, and 10 affecttheir CNS activity.

Tricyclische Azaergolin Analoge: Synthese, strukturelle Modifikationenund pharmakologische Studien

1m AnschluB an friihere Untersuchungen zur Synthese und Oopamin Autore-zeptor Aktivitiit bicyclischer Ergolin Analoger, wurden die tricyclischenAzaergolin Analogen 9a und 9b synthetisiert. AuBerdem sollte die Geome-trie der aromatisch substituierten ~-Ethylamin-Struktur vaniert werden. Eswurde deshalb das Cycloheptenylamin Derivat 7 und der aminomethyl-sub-stituierte Tricyclus 10 dargestellt. Die Affinitat dieser Verbindungen zu denOopamin (OA) Rezeptor Bindungsstellen wurde untersucht, wobei das Stria-tum der Ratte verwendet wurde. Die Verbindungen binden mittelstark bisschwach an den mit dem OA-Autorezeptor Agonisten eHI-SND 919 mar-kienen 0-2 Rezeptor. In vivo Untersuchungen mit Mausen zeigten, daB 7,9a,b und 10 die ZNS Aktivitiit der Tiere beeinflussen.

Since the fortuitous discovery of chlorpromazine!) a large number ofneuroleptics have been developed2). It is postulated that the antipsychoticactivity of the classical neuroleptics is due to a blocking effect at thepostsynaptic 0-2 receptors3). Since these 0-2 receptor antagonists induceextrapyramidal side effects (i.e., pseudo-parkinsonism), alternative ap-proaches for preventing the symptoms of schizophrenia attract considerableattention in recent years 4). One of these novel approaches is to employselective OA-autoreceptor agonists, which decrease synthesis and releaseof dopamine (OA)5,6). Thus the hyperactivity in the meso limbic dopaminer-gic system (A 10 system) of schizophrenia patients should be reduced.Since it is assumed that the binding sites of the postsynaptic 0-2 receptorand its prejunctional congener are very similar5), it seemed to be a valuablestrategy to modify the molecular structure of known postsynaptic 0-2agonists, in order to develop OA autoreceptor agonists.

In fact, we and others have shown that this is possible, byapplying the DA active bicyclic ergoline analogue 1 as amodel compound. Structural modifications at the aromaticregion resulted in selective autoreceptor activity of the deri-vatives 27), 3 (SND 919)8), and 4, 59).

As an extension of our work, we try to evaluate specificligands for the DA autoreceptor by structural changes ofthe tricyclic ergoline analogue 6, which is also known toexhibit DA agonistic activitylO). Thus, we are investigatingaza analogues with pyrazolo[1,5-a]pyridine (8)11) or tetra-hydropyrazolo[ I ,5-0 ]pyridine substructure (9). In thispaper, we report on syntheses and phannacological studiesof both diastereomers of the hexahydropyrazoloquinolinederivative 9. As a further modification, the geometry of thearomatic ethyl amine moiety of 9 was varied by construc-tion of the tetrahydropyrazolopyridyl fused cyclohepteny-lamine 7 and the aminomethyl substituted pyran derivative10.

c5:N-N

\H

NPr2 /'

~

~8

.::::::?

The synthesis of the target compounds 9a and 9b wasplanned by a stereospecific Clirtius rearrangement startingfrom the carboxylates 14a and 14b, which we obtained fromthe pyrazolopyridine carboxylate 11 via the ~-ketoester 12aand the iodide 13 as the key intermediatesl2). The hydrolysisof the esters 14a and 14b was performed by NaOH in diox-ane/H20 to yield the carboxylic acids ISa and ISb, respec-tively, both as pure diastereomers. Treatment of ISa,b withdiphenyl phosphorazidate (DPPA) in acetonitrile I3.14), fol-lowed by acidic hydrolysis of the intermediately formed iso-cyanates, afforded the primary amines 16a and 16b withcomplete retention of configuration, indicated by NMRspectroscopy. This reaction is best monitored by IR-spec-troscopy by observing the appearence/disappearence of thediagnostic bands at 2260 (N=C=O) and 1710 em'! (O=C-OH). FinaIly the primary amines 16a and 16b were trans-formed into the dipropylamines 9a and 9b by use of pro-pionic aldehyde and NaCNBH3. For both products the cy-clohexenyl ring adopts a half-chair conformation, when theamine substituent is positionated axiaIly for 9a and equator-ially for 9b, according to the NMR data.

dOOC'H'~11

H~~)~:00

12a: n = 112b:n=2

/ ~

&;H'17: X =OH18: X=OMes19:X= I

~

biRR'H ~N~N20a: R = COZCH3.; R' = H20b: R = H; R' = c;02CH3

21: R = C02H; R' = H22: R = NH2; R' = H

7: R = NPr2; R' = H

14a: R = C02~H3.; R' = H14b. R = H, R = C02CH315a: R = C02H; R' = H15b: R = H: R' = C02H16a: R = NH2: R' = H16b: R = H; R' = NH29a: R = NPr2; R' = H9b: R = H; R' = NPr2

For the synthesis of the homologues cycloheptene fusedheterocycles the hydroxyethyl substituted ~-ketoester 12b,which can be efficiently derived from 11, according to pre-vious studiesI5), should serve as a suitable educt. Reductionof the aromatic ketone 12b was achieved by catalytic hy-drogenation (Pd/C) at 60 bar pressure and l30°C to give 17

in 92% yield. Subsequently the primary alcohol 17 was con-verted into the mesylate 18. Treatment of 18 with NaIyielded the electrophile 19, which could be cyclized by a7-(enolexo)-exo-tet ring closure using LDA as a base, at-78°e. Surprisingly, the ring closure proceeds stereoselec-tively to afford the trans diastereomer 20a exclusively, Bycontrast, the 6-(enolexo)-exo-tet cyclization of 13 affords14a and 14b in a I: I ratio12). We reason that the cyclohep-tene derivative 20a is formed selectively because - kineti-caIly controlled - the reaction proceeds through a chair typetransition state, leading to the trans configurated product(Figure I). On the other hand, formation of the 6-memberedring requires much higher temp. (O°C). Thus, under thestrongly basic conditions epimerization takes place and athermodynamicaIly controlled mixture of 14a and 14b isproduced. Such an equilibration can be also observed forthe 7-membered case, by treating 20a with LDA at O°C,when a I: I mixture of diastereomers (20a and 20b) wasisolated.

Our final product 7 was synthesized from 20a in 53%yield by hydrolysis, followed by Curtius rearrangement andreductive alkylation via the intermediates 21 and 22.

x) 0rtCH3

C)~,"12a: X = OH; R = H12c: X = I; R = H12d: X = OH; R = CH3

X=y ~OH(yrCH'~e'H[~~:H'

~ ~~ ..

23 24a; R = H" 24b: R = CH3

'" NOE /,.- H R' .¥

\OE( \ 0i+RH' ••~~A R

65a~, /)N.~25a: R = H; R' = COj!CH;)25b; R = CHJ: R' = c;02CH325c. R = H, R = C02H25d: R = H; R' = NH2

10: R = H; R' = NPr2

Previously, we have demonstrated that anionic cyclizationof the iodide 12c results in O-alkylation to give the enolether 23 eXclusively'S). We regarded this compound as auseful precursor for the synthesis of the amine 10, whichwas planned to be investigated as a potential DA equivalentwith a differently locked ethyl amine conformation. Ac-tually, catalytic hydrogenation (Pd/C) of the enol ether 23afforded the pseudo-equatorially substituted pyran deriva-tive 2Sa selectively. The nucleophilic approach from thepseudo-axial side might be explained by transition state sta-bilization due to interaction of the axially positioned oxygenlone-pair and the incipient low lying vacant CH-orbital0"*" 16). In case of an equatorial approach, torsional strainbetween the axial 0 lone-pair and the incipient 0"" CH-orbi-tal would destabilize the transition structure'?) (Fig. 2).

The structure of 25a was elucidated by IH-NMR spectroscopy observinga significant dipolar exchange of magnetism between H-5 •• and H-3, due to1,3-diaxial interference. The position of H-5ax was determined by the H,H-COSY technique, a diagnostic vicinal coupling constant (\ax'5a = 10.7Hz), and a significant NOE with H-6ax (1,3-diaxial interaction).

The conversion of the ester 2Sa into the target compound10 was performed by Curtius rearrangement through theintermediates 2Sc and 2Sd, as described for the syntheses of7 and 9.

Alternatively, 2Sa can be synthesized directly from theprimary alcohol 12at2) - a synthetic precursor of 12c and 23- by catalytic hydrogenation in acetic acid in 64% yield.This is an essential improvement of the synthesis since thepathway via 12c and 23 requires 3 additional steps and af-fords an overall yield of only 15%. We expect, that thereaction proceeds through the hemiacetal 24a. Then, 24acan either eliminate H20 to give 23 which is subsequentlyhydrogenated, or hydrogenation takes place on an inter-mediately formed stabilized carbenium ion. To investigatewhether the formation of an enolether by dehydration isessential for this one pot procedure the dimethyl substitutedanalogue 12d was heated using similar conditions (HOAc,140°C, 50 bar H2, Pd/C). In fact, the reaction works verysmoothly to give 2Sb as a single diastereomer, although -due to the quaternary C-atom - dehydration of the hemiace-tal 24b is impossible.

The educt 12d was prepared from the aromatic ester 2612)by ester condensation to give the p-keto ester 27. Afterdeprotonation by LDA 27 was reacted with benzyloxy-methyl chloride to afford 12e, which could be debenzylatedby hydrogenolysis to yield 12d.

Employing rat striatal membranes, the novel tricyclicazaergoline analogues 7, 9a,b, and 10 were evaluated fortheir binding affinity to the dopamine D-l receptor labelledwith eHl-SCH 23390 and to the D-2 receptor sites labelledwith eH]-spiroperidol and eH]-SND 919, a compoundwhich in functional in vivo experiments pointed out to be anautoreceptor agonisfl). It turned out, that there exists a signifi-cant affinity to the DA receptor site labelled with eH]-SND919, which is selective since an affinity to the postsynapticD-l and to the D-2 receptor labelled with eH]-spiroperidolcould not be observed. However, the ICso values demonstratethe ability of 7, 9a,b, and 10 to displace eH]-SND 919 isonly modest (Table 1). The most active compound was theequatorially substituted cis isomer 9b, which gave an ICsovalue of 4.4 flM. (By contrast, our previously developed bi-cyclic ergoline analogue (S)-3 showed a 147-fold higher af-finity (ICso = 0.03 flM)9». The trans configuration of the cy-cloheptene fused homologue 7 resulted only in a slight loss ofaffinity (ICso = 6.3 flM), when compared to 9b. This is easyto understand because the dipropylamino group of 7 is alsoequatorially orientated. On the other hand, the cyclohexeny-lamine fused trans diastereomer 9a with an axially posi-tioned dipropylamino group had a 4.3 fold lower affinity than9b. The lowest ICso value (23 flM) was measured for thepyran derivative 10, when also a conformational change ofthe ethylamine moiety resulted in a decrease of the affinity tothe DA-receptorlabelled with eH]-SND 919.

ICso [11M]

compd 0-1' , D-2b DA-autoreceptor c

7 > 100 > 100 6.39a > 100 > 100 199b > 100 > 100 4.410 > 100 > 100 23

In accordance to the receptor binding data, 9b and 7turned out to be the most active compounds in vivo. Ourbehavioural-activity screening with mice showed that all thetest compounds (7, 9a,b, and 10) caused excitation includ-ing Straub tail, vocalization and convulsion, immediatelyafter injection. After this period of excitation both cyclo-

Table 2: Locomotor Activity Measurements (0-60 min)

compound counts (x ± S.E.M.) control counts C (it ± S.E.M.) t1d P7 (25mg) 3046±532 1992±275 +53%

was filtered (CeliteR AFA) and the filtrate was evaporated. The residue waspurified by flash chromatography (CH2Cl2 - CH30H 97:3) to give 130 mg(86%) ofl2d as a colorless oil.- CI4H2oN204 (280.3) Calcd. C 60.0 H 7.19 N10.0 Found C 60.0 H 7.16 N 10.0; mol.-mass 280 (ms).- IR (NaCI): 3390;2950; 1740; 1660 cm·l._ IH-NMR (CDeI3): () (ppm) = 1.48 (s, 3H, CCH3),1049 (s, 3H, CCH3), 1.78-2.20 (m, 4H, H2C-5, H2C-6), 2.60 (br-s, IH, OH),3.64-3.66 (m, 2H, CH20), 3.68 (s, 3H, OCH3), 3.85-3.86 (m, IH. HC-4),4.00 (m, I H, H•• C-7), 4.27 (m, I H, HeqC-7), 7.22 (s, 1H, HC-2).

Methyl (±}-3-(4-benzyloxymethyl-4,5 .6,7-tetrahydropyrazolo[1,5-a]pyridill-3-yl)2,2-dimethyl-3-oxo-propionate (Ue)

To a solution of 27 (250 mg, 1 mmol) in 10 ml of THF were addeddropwise 3 ml of freshly prepared LDA (0.32 molar in THF) at _78°C. Thereaction mixture was stirred at this temp. for 30 min, when it was addeddropwise to a precooled solution (-78°C) of benzyloxymethylchloride(BOM-CI; 234 mg, 1.5 mmol) in 10 ml ofTHF. After 30 min it was addedto a mixture of ether and satd. aqueous solution of NaHC03. The org. layerwas dried (MgS04) and evaporated and the residue was separated by flashchromatography (petroleum ether - EtOAc 1:1) to give 250 mg (68%) ofpure 12e as a colorless solid; m.p. 83°C.- C21H26N204 (370.5) Calcd. C68.1 H 7.07 N 7.6 Found C 67.9 H 7.10 N 7.7; mol.-mass 370 (ms).- IR(KBr): 3040; 2950; 2870; 1740; 1660 cm-I.-IH-NMR (CDCI3): 0 (ppm) =1.46 (s, 3H, CCH3), 1.47 (s, 3H, CCH3), 1.73-2.29 (m, 4H, H2C-5, H2C-6),3.55 (dd, J = 9.5, 8.8 Hz, 1H, PhCH20C.!h), 3.65 (s, 3H, OCH3), 3.74 (dd,J = 9.5,3.0 Hz, tH, PhCH20Cl:!.l), 3.83-3.85 (m, IH. HC-4). 3.96 (ddd. J =13.2, 11.7, 5.1 Hz, IH, H••C-7), 4.21 (br-dd, J = 13.2, 3.6 Hz, IH, HeqC-7),4046 (d, J = 11.7 Hz, IH, PhCH2), 4.66 (d, J = 11.7 Hz, 1H, PhCH2),7.25-7.36 (m, 5H, Ph), 7.69 (s, IH, HC-2).

(4RS ,5aRS}-4,5 ,Sa ,6 ,7 ,8-Hexahydro-3 H-pyrazolo[2,3 ,4-j,i] quinolin-4-yl-carboxylic acid (ISa)

A solution of 14a (82 mg, 0.37 mmol) in 2 ml of dioxane and 1.5 ml of NNaOH was stirred at room temp. for I h. Then the solution was extractedwith ether. The aqueous layer was acidified to pH = 3-4 with 2 N HCl andsubsequently extracted with CH2CI2 - CH30H 9:1. The org. layer gave 50mg (66%) of ISa as a colorless solid, m.p. 191°C, after drying (MgS04),evaporation, and flash chromatography (CH2CI2 - CH30H 9:1).-CIlH14N202 (206.3) Calcd. C 64.1 H 6.84 N 13.6 Found C 64.2 H 7.07 N13.2; mol.-mass 206 (ms).- IR (KBr): 3430; 2930; 2850; 1700 cm·I.- IH_NMR (CDCI3): () (ppm) = 1.20 (br-q, J = 12 Hz, IH, H••C-6), 1.45 (ddd, J= 12.5, 12.5,8 Hz, IH, H••C-5), 2.01-2.15 (m, 3H, HeqC-6, H2C-7), 2042(br-d, J = 13 Hz, IH, HeqC-5), 2.67 (dd, J = 16.2,6.6 Hz, IH, H••C-3), 2.78(m, IH, Ha.C-5a), 3.09 (m, IH, HeqC-4), 3.15 (d, J = 16.2 Hz, IH, HeqC-3),3.85-3.90 (m, 1H, Ha.C-8), 4.26-4.30 (m, IH, HeqC-8), 7.31 (s, 1H. HC-2).

(4RS ,5aSR)-4 ,5 ,5a.6 ,7,8-hexahydro- 3H-pyrazolo[2,3 ,4-j,i]quinolill-4-yl-carboxylic acid (ISb)

A solution of 14b (70 mg, 0.32 mmol) in 2 ml of dioxane and 1.5 ml of NNaOH was stirred at room temp. for I h and worked up as described for ISato give 45 mg (68%) of ISb as a colorless solid; m.p. 209°C.- CII H 14N202(206.3) Calcd. C 64.1 H 6.84 N 13.6 Found C 64.2 H 6.99 N 1304; mol.-mass206 (ms).- IR (KBr): 3430; 2950; 2850; 1710 cm-I.- IH-NMR (CDCI3): ()(ppm) = 1.20 (br-q, J = 13 Hz, IH, Ha.C-6), 1.46 (q, J = 12.5 Hz. IH. HaxC-5),2.01-2.22 (m. 3H, HeqC-6. H2C-7), 2.33 (ddd, J = 12.5,404,2.2 Hz. IH.HeqC-5), 2.68-2.78 (m, 2H, H••C-3. H••C-5a), 2.81-2.89 (m, IH. H••C-4),2.98 (dd, J = 14.7.5.8 Hz, IH, HeqC-3), 3.88 (ddd, J = 12.5, 12.5,5.5 Hz. IH,H••C-8), 4.30 (dd, J = 12.5,5.9 Hz, IH, HeqC-8), 7.33 (s, IH, HC-2).

(4RS ,SaRS )-4-Amino-4.5 'sa,6,7,8-hexahydro-3 H-pyrazolo[2,3,4-j.i]quinoline (16a)

A mixture of ISa (50 mg. 0.24 mmol). diphenyl phosphorazidate (67 mg,0.24 mmol) and triethylamine (24 mg, 0.24 mmol) in 15 ml of acetonitrile

was stirred at 60°C for 5 h. After the addition of 0.5 N HCI (3 ml) thereaction mixture was stirred at room temp. for 1.5 h. The solution wasconcentrated, then extracted with ether, basified with 2 N NaOH and sub-sequently extracted with CH2CI2. The org. layer was dried (MgS04) andevaporated to give 28 mg (65%) of 16a as a colorless oil.- ClOH15N3(177.3) mol.-mass 177 (ms).- IR (NaCl): 3340; 2920; 2850 cm·l._ IH-NMR(CDCI3): 0 (ppm) = 1.25 (dddd. J = 12.5, 12.5, 12.1, 3 Hz, IH. Hax C-6),1.37 (ddd, J = 12.5, 12.1,2.6 Hz, IH, H••C-5), 1.95-2.01 (m. 2H. HeqC-5,HeqC-6), 2.05-2.23 (m. 2H. H2C-7), 2047 (d. J = 16.1 Hz. IH. HeqC-3), 2.80(ddd, J = 16.1,5.2,1.8 Hz, IH. H••C-3), 2.91 (dddd. J = 12.1. 11.7.5.5.5.5Hz, IH, H.xC-5a), 3.60 (br-s, IH, HeqC-4), 3.88 (ddd. J = 12.5. 12.1,5.9Hz, IH, H••C-8), 4.28 (dd, J = 12.5, 6 Hz, IH, HeqC-8), 7.24 (s, IH,HC-2).- Elemental analysis was performed after conversion of 16a into itsdibenzoyl hydrogen tartrate using I equiv. of (L)-dibenzoyltartaric acid inether.- C2sH29N30S (535.6) Calcd. C 62.8 H 5046 N 7.8 Found C 62.9 H5047 N 7.8.

(4RS'saSR)-4-amino-4,5 ,Sa.6.7.8-hexahydro-3H -pyrazolo[2 ,3 ,4-j.i]quinoline (16b)

A mixture of ISb (43 mg. 0.21 mmol). diphenyl phosphorazidate (58mg, 0.21 mmol) and triethylamine (21 mg, 0.21 mmol) in 15 ml of ace-tonitrile was stirred at 60°C for 5 h. After the addition of 0.5 N HCl (3ml) the reaction mixture was stirred at room temp. for 2.5 h and workedup as described for 16a to give 24 mg (65%) of 16b as a colorless oil.-ClOH15N3 (177.3) mol.-mass 177 (ms).- IR (NaCl): 3340: 2920: 2850 em'1._ IH-NMR (CDCI3): () (ppm) = 1.23 (dddd, J = 13.0, 12.5, 12.5.2.8 Hz,IH, H••C-6), 1.27 (ddd, J = 11.8, 11.7, 11.6 Hz, IH, H••C-5). 1.97-2.04(m, 2H, HeqC-5. HeqC-6), 2.05-2.10 (m. IH. Ha.C-7), 2.14-2.20 (m. I H,HeqC-7). 2.19 (ddd, J = 15.0: 10: 2 Hz, IH. HaxC-3). 2.74 (dddd. J = 12.5.11.8, 5.5, 4 Hz, I H, Ha,C-5a), 2.92 (dd, J = 15.0, 5.9 Hz, IH, HeqC-3),3.26 (dddd, J = 11.6, 1004,5.8.2.9 Hz, IH, H••C-4), 3.85 (ddd. J = 12.5,12.5.5.9 Hz, 1H, H.,C-8), 4.26 (dd. J = 12.5.5.5 Hz. IH. HeqC-8), 7.27(s, IH, HC-2).

Methyl(±)-3 -[4.5.6.7 ·tetrahydro-4-(2 -hydroxyethyl)-pyrazolo[1,5-a]pyrid-3-yl]propiollate (17)

A mixture ofl2b (1900 mg. 7.14 mmol) and I g of Pd/C (10%) in 60 mlof methanol was stirred at 1300C for 3 h under H2 of 60 bar. Then it wasfiltered (CeliteR AFA) and the filtrate was evaporated to give 1650 mg(92%) of 17. Flash chromatography (CH2CI2-CH30H 97:3) gave an analyti-cally pure sample as a colorless oil.- C13H20N203 (252.3) Calcd. C 61.9 H7.99 N 11.1 Found C 61.7 H 8.18 N 11.1: mol.-mass 252 (ms).-IR (NaCI):3340; 2950: 2860; 1740 cm-I.- IH-NMR (CDCI3): () (ppm) = 1.71-2.13 (m,6H, H2C-5, H2C-6, CH2CH20). 2.57-2.60. 2.74-2.79 (2m. 4H.CH2CH2CO), 3.18-3.23 (m. IH, HC-4), 3.68 (s. 3H. OCH3), 3.72-3.77 (m,2H, CH20), 3.98 (ddd, J = 12.5,9.5,5.1 Hz, IH, H••C-7), 4.19 (ddd, J =12.5,5.1,404 Hz, IH, HeqC-7), 7.28 (s, IH, HC-2).

Methyl(±}-3 -[4,5.6.7 ·tetrahydro-4-(2 -mesyloxyethyl }-pyrazolo[l,S-a]pyrid-3-yl]propionate (18)

To a solution of 17 (1500 mg. 5.95 mmol) in 100 ml of THF was addedtriethylamine (0.55 ml, 7 mmol) and methanesulfonyl chloride (0.97 ml. 7mmol). After stirring at room temp. for 2 h the mixture was evaporated andthe residue purified by flash chromatography (CH2ClrCH30H 94:6) toafford 1890 mg (96%) of 18 as a colorless oil.- CI4H22N20SS (33004)Calcd. C 50.9 H 6.71 N 8.5 Found C 50.6 H 6.63 N 804: mol.-mass 330(ms).- IR (NaCI): 2950: 1740: 1350: 1170 cm·'.- IH-NMR (CDCI.,): ()(ppm) = 1.82-2.16 (m, 6H. H2C-5. H2C-6. Cl:bCH20), 2.55-2.61. 2.70-2.75(2m, 4H, CH2CH2CO), 3.06 (s. 3H. OS02CH.,), 3.17-3.21 (m. IH. HC-4),3.68 (s. 3H. OCH3). 3.96-4.04 (m. IH. H".C-7l. 4.17-4.22 (m. IH. HeqC-7).

4.30-4.35 (m, 2H, CHzO), 7.30 (s, IH, HC-2).

Methyl (±)-3-f45,6,7 -tetrahydro-4-(2 -iodoethyl )-pyrazolo[J ,5-a]pyrid-3-yl] propionate (19)

A mixture of 18 (1700 mg, 5.15 mmol) and NaI (7700 mg, 51.3 mmol)was stirred in 100 ml of boiling acetone for 4 h. After cooling to roomtemp. the solvent was removed and the residue extracted with ether. Theextract was evaporated and the residue purified by flash chromatography(petroleum ether - EtOAc 4:6) to give 1230 mg (66%) of 19 as a colorlessoil.- Ct3Ht9INzOz (362.2) Calcd. C 43.1 H 5.29 N 7.7 Found C 43.0 H 5.45N 7.7; mol.-mass 362 (ms).- IR (NaCI): 2950; 2850; 1740 cm-t._ tH-NMR(CDCI3): /) (ppm) = 1.75-2.24 (m, 6H, HzC-5, HzC-6, CHzCHzI), 2.57-2.61, 2.74-2.78 (2m, 4H, CHzCHzCO), 3.12-3.20 (m, 2H, HC-4, CHzI),3.32 (ddd, J = 12.5,7.3,4.4 Hz, IH, CHzI), 3.69 (s, 3H, OCH3), 4.00 (ddd,J = 12.5,9.6,5.1 Hz, IH, H••C-7), 4.18 (ddd, J = 12.5,5.2,5.1 Hz, IH,HeqC-7), 7.29 (s, I H, HC-2).

Methyl(5aRS ,8RS)-3,4.5.5a.6 ,7.8 .9-0ctahydl'o-2 .2a-diazabenzo[ c .d]azulene-8-cal'bo:tylate (20a)

Methyl(5aRS ,8SR )-3,4.5 ,5a.6 ,7,8 .9-0ctahydro-2 ,2a-diazabenzo[ c ,d]aut/ene-8-cal'bmylate (20b)

To a solution of 19 (362 mg, I mmol) in 30 ml of THF were addeddropwise 3.4 ml of freshly prepared LDA (0.32 molar in THF) at _78°e.The reaction mixture was stirred at this temp. for 15min, when it was addedto a mixture of ether and satd. aqueous solution of NaHC03. The org. layerwas dried (MgS04) and evaporated and the residue was separated by flashchromatography (petroleum ether - EtOAc 1:1) to give 140 mg (60%) ofpure 20a as a colorless solid; m.p. noc.- Ct3HtsNzOz (234.3) Calcd. C66.6 H 7.74 N 12.0 Found C 66.6 H 7.98 N 11.8; mol.-mass 234 (ms).- IR(KBr): 2930; 2840; 1735 cm-t._ IH-NMR (CDCI3): /) (ppm) = 1.25-1.45(m, 2H, H.,C-5, Ha,C-6), 1.75 (br-q, J = 13 Hz, IH, Ha,C-7), 1.87-1.99 (m,2H, H.,C-4, H.,C-6), 2.04-2.10 (m, 2H, HeqC-4, HeqC-5), 2.36-2.43 (m,2H, HeqC-7, HeqC-8), 2.62 (dd, J 0 14.7, 11.8 Hz, IH, Ha,C-9), 2.73-2.78(m, IH, He-5a). 2.97 (ddd, J = 14.7,2.2,2.2 Hz, IH, HeqC-9), 3.70 (s, 3H,OCH3), 3.94 (ddd, 12.5, 12.5,3.7 Hz, IH, H.,C-3), 4.22 (dd, J = 12.5,5.1Hz, IH, HeqC-3), 7.23 (s, IH, HC-I).- 13C_NMR (CDCI3): /) (ppm) = 23.1(HzC-4), 28.8 (H2C-9), 28.9 (HzC-5), 33.8 (HzC-7), 34.3 (HzC-6), 36.5(HC-5a), 45.5 (HC-8), 47.5 (H2C-3), 51.7 (CH3), 115.5 (C-9a), 138.1 (HC-I), 141.0 (C-2b), 176.3 (C02),

Treatment of 20a (23.4 mg, 0.1 mmol) with freshly prepared LDA (0.3ml, 0.32 molar in THF) in THF (5 ml) at O°C for 30 min gave a I: I mixtureof 20a and 20b after usual work up.

20b: IH-NMR (CDCI3): /) (ppm) = 1.33-1.43 (m, IH, Ha,C-5), 1.62(br-q, J = 13 Hz, IH, Ha,C-6), 1.74-1.83 (m, IH, HeqC-7), 1.90-2.10 (m,5H, HzC-4, HeqC-5, HeqC-6, HeqC-7), 2.70 (dd, J = 14.7,3 Hz, IH, Ha,C-9),2.70-2.78 (m, IH, Ha,C-5), 2.89 (dddd, J = 6; 6; 3; 3 Hz, IH, HeqC-8),3.05 (dd, J = 14.7,5.9 Hz, IH. HeqC-9), 3.62 (s, 3H. CH3), 3.89-3.97 (m,IH, Ha,C-3), 4.18-4.24 (m, IH, HeqC-3), 7.24 (s, IH, HC-I).

(5aRS .8RS )-3,4,5 .5a.6 ,7 ,8 ,9-0ctahydro-2 ,2a-diazabenzo[ c.d]azu/ene-8-carboxylic acid (21)

A solution of20a (160 mg, 0.68 mmol) in 10 ml of dioxane and 10 ml ofN NaOH was stirred at room temp. for 2.5 h. Then the solution was ex-tracted with ether. The aqueous layer was concentrated and then acidifiedto pH 3-4 with citric acid (10% in H20). After standing at room temp. for 2h. filtration and drying of the residue gave 125 mg (83%) of 21 as acolorless solid; m.p. 253°C.- C12HI6NZ02 (220.3) Calcd. C 65.4 H 7.32 N12.7 Found C 65.8 H 7.23 N 12.5; mol.-mass 220 (ms).- IR (KBr): 3430;2930; 2850; 1710 cm-I._ IH-NMR (CDCl3): /) (ppm) = 1.34 (br-g. J = 13Hz. IH. Ha,C-6), 1.41 (br-g. J = 12 Hz. IH. Ha,C-5), 1.75 (br-g, J = 13 Hz,IH, Ha,C-7), 1.88-2.02 (m, 2H, Ha,C-4, HeqC-6), 2.07-2.13 (m. 2H. HeqC-

4, HeqC-5), 2.30 (It, J = 11.7,2.2 Hz, IH, HaxC-8), 2.39 (ddd. J = 13; 6.6; 2Hz, IH, HeqC-7), 2.57 (dd, J = 15; 12 Hz, IH, HaxC-9), 2.75-2.81 (m, IH,HC-5a), 2.97 (ddd, J = 14.7,2.2,2.2 Hz, IH, HeqC-9), 3.90 (ddd, J = 12.5,11.7,3.7 Hz, IH. HaxC-3), 4.13 (dd. J = 12.5.5.0 Hz. IH, HcqC-3), 7.20 (s,IH, HC-I).- 13C-NMR (CDC!.,): /) (ppm) = 22.6 (H2C-4), 28.3 (H1C-5,H1C-9), 33.4 (H1C-7), 33.9 (H1C-6), 35.8 (HC-5a), 45.1 (HC-8), 47.1(H1C-3), 115.0 (C-9a), 137.3 (HC-I), 140.5 (C-2b), 176.6 (COz).

(5aRS .8RS)-8-Amino-3.4.5.5a .6.7.8 .9-octah\'dI'0-2 .2a-diazabenzo[c.d]azu/ene (22) .

A mixture of 21 (300 mg. 1.36 mmol). diphenyl phosphorazidate (375mg, 1.36 mmol) and triethylamine (138 mg, 1.36 mmol) in 25 ml of ace-tonitrile was stirred at 60°C for 4 h. After addition of 0.5 N HCI (20 ml) thereaction mixture was stirred at room temp. for 2.5 h and worked up asdescribed for 16a to give 234 mg (87%) of 22 as a colorless solid; m.p.85°C.- C11H17N3 (191.3) mol.-mass 191 (ms).- IR (KBr): 3360; 2910; 2830cm-I.- IH-NMR (CDCI3): /) (ppm) = 1.37-1.46 (m. 2H, Ha,C-5, Ha,C-6),1.58-1.66 (m, IH, HaxC-7), 1.84-1.96 (m, 2H. Ha,C-4, HeqC-6). 2.03-2.11

(m. 3H, HeqC-4, HeqC-5. HeqC-7), 2.41 (dd. J = 13.9. 11.0 Hz. IH, HaxC-9),2.70-2.86 (m, 3H, HaxC-5a. Ha,C-8, HeqC-9), 3.93 (ddd, J = 12.5. 12.5,4Hz, IH, HaxC-3), 4.21 (dd, J = 12.5,5.1 Hz, IH, HeqC-3), 7.21 (s, IH,HC-I).

Methy/( 3RS .5aSR)-(5a.6 .7.8.-retrahydI'0-3H,5H -4-o.l'a-/.8a-diaza-acenaphthy/en-3-y/)-acctate (25a)

A mixture of 12a (50 mg, 0.2 mmol) and 100 mg of Pd/C (10%) in 10 mlof acetic acid was stirred at 120°C for 3 h under H, of 70 bar. Then it wasfiltered (CeliteR AFA) and the filtrate was evap;rated. A satd. aqueoussolution of NaHC03 was added and the mixture was extracted with EtOAc.The org. layer was dried (MgS04) and evaporated and the residue wasseparated by flash chromatography (petroleum ether - EtOAc I: I) to give30 mg (64%) of 25a as a colorless solid; m.p. 76°C.- ClzHI6N20, (236.3)Calcd. C 61.0 H 6.83 N 11.9 Found C 61.3 H 6.84 N 11.9: mol.-mass 236(ms).- IR (KBr): 2930; 2850; 1740 cm-I._ IH-NMR (CDCI3): /) (ppm) =1.10 (dddd, J = 13; 12.8; 12.5; 3.0 Hz, IH, HaxC-6), 1.91-2.25 (m. 3H,HeqC-6, HzC-7), 2.74 (d, J = 6.8 Hz, 2H, CH2CO), 2.97-3.02 (m, IH,HC-5a), 3.18 (t, J = 10.7 Hz, IH, HaxC-5), 3.74 (s. 3H, CH3), 3.89 (ddd, J =12.5, 12.5,5.5 Hz, IH, HaxC-8), 4.13 (dd, J = 10.7,5.1 Hz, IH, HeqC-5),4.32 (dd, J = 12.5,6.0 Hz, IH, HeqC-8), 5.15 (ddd, J = 6.8, 6.4, 1.7 Hz, I H,Ha,C-3), 7.27 (s, IH, HC-2).

Methy/(3RS .5aRS )-(5a,6.7,8-tetrahydro-3H 5H -4-o.l'a-/ ,8a-diaza-acenaphthy/en- 3 -yl}-dimethylacetate (25b)

A mixture of 12d (60 mg, 0.21 mmol) and 100 mg of Pd/C (10%) in 15ml of acetic acid was stirred at 140°C for 5 h under H2 of 60 bar. Then itwas filtered (CeliteR AFA) and the filtrate was evaporated. The residue wasseparated by flash chromatography (petroleum ether - EtOAc 6:4) to give22 mg (40%) of 25b as a colorless oil.- C14H20N203 (264.3) Calcd. C 63.7H 7.63 N 10.6 Found C 63.5 H 7.69 N 10.7; mol.-mass 264 (ms).- IR(NaCI): 2950; 2870; 1730 cm-I._ IH_NMR (CDCI,): /) (ppm) = 1.07 (dddd,J = 12.5, 12.5. 12.5, 2.9 Hz, I H. Ha,C-6), 1.13 (s. 3H, CCH3), 1.22 (s, 3H,CCH3), 1.90-2.23 (m, 3H, HeqC-6, H2C-7), 2.90-2.96 (m, IH, HC-5a). 3.08(t, J = 10.3 Hz, IH. Ha,C-5), 3.75 (s, 3H. OCH3), 3.89 (ddd. J = 12.5. 12.5,5.9 Hz, IH, Ha,C-8), 4.08 (dd, J = 10.3,5.1 Hz. IH. HeqC-5), 4.31 (dd, J =12.5. 5.8 Hz, I H. HeqC-8), 5.01 (d, J = 1.5 Hz. IH. HC-3). 7.21 (s, I H.HC-2).

(3RS .5aSR )-(5a.6 .7,8- Tetrahydro-3H.5 H-4-o.l'a-/ ,8a-dia za-ace III/phthylcn-3-yl)aceric acid (2Se)

A solution of 2Sa (500 mg. 2.12 mmol) in 20 ml of dioxane and 20 ml ofN NaOH was stirred at room temp. for 4 h. Then the solution was extracted

with ether and the aqueous layer was concentrated. then acidified to pH 3-4with citric acid (10% in H20) and extracted with CH2Cl2• The org. layerwas dried (MgS04) and evaporated to give 330 mg (70%) of 25e as acolorless solid; m.p. 192°C.- CIIHI4N203 (222.2) Calcd. C 59.5 H 6.35 N12.6 Found C 59.6 H 5.76 N 12.6; moL-mass 222 (ms).- IR (KBr): 2950;2870; 2750; 2490; 1710 em-I.- IH_NMR (CD30D): l) (ppm) = 1.12 (dddd. J= 13.2, 12.5, 12.5,2.5 Hz. IH, HaxC-6), 1.90-2.25 (m. 3H, lIeqC-6, H2C-7).2.80 (br-d, J = 5.9 Hz. 2H. CH2CO), 2.98-3.05 (m. I H. HC-5a). 3.22 (t. J =10.3 Hz. IH. HaxC-5). 3.91 (ddd, J = 12.5, 12.5,5.1 Hz, IH. HaxC-8), 4.17(dd, J = 10.3.5.1 Hz. IH, HeqC-5), 4.34 (dd, J = 12.5.5.9 Hz. IH. HeqC-8),5.15 (br-t, J = 5.9 Hz, IH. HC-3), 7.35 (s, IH, HC-2).

(3RS ,5aSR)-3 -Aminomethy/-5a .6.7.8-tetrahydro-3H ,5H -4-oxa-1.8a-diaza-acenaphthy/ene (25d)

A mixture of 25e (220 mg, I mmol), diphenyl phosphorazidate (275 mg,I mmol) and triethylamine (101 mg, I mmol) in 10 ml of acetonitrile wasstirred at 60°C for 20 h. After the addition of 0.5 N HCI (20 ml) thereaction mixture was stirred at room temp. for 5 h and worked up asdecribed for 16a to give 145 mg (75%) of 25d as a colorless oil.-ClOHI5N30 (193.3).- IR (NaC1): 3420; 2930; 2860 cm·'.- 'H-NMR(CDCI3): l) (ppm) = 1.01 (dddd, J = 12.5, 12.5, 12.5,2.2 Hz, IH, HaxC-6),1.93-2.24 (m, 3H, HeqC-6, H2C-7), 2.78 (br-s, 4H, CH2NH2), 2.97-3.04 (m,IH, HC-5a), 3.18 (I. J = 10.3 Hz, IH, HaxC-5), 3.88 (ddd, J = 12.5, 12.5,5.9 Hz, IH, HaxC-8), 4.15 (dd, J = 10.3,5.1 Hz, IH, lIeqC-5), 4.31 (dd, J =12.5.5.9 Hz, IH, HeqC-8), 4.82 (br-s, IH, HC-3), 7.31 (s, IH, HC-2).

M ethy/2 ,2-dimethy/-3 -oxo-3 -(4,5.6.7 -tetrahydropyraz%[J ,5-aj-pyridin-3-y/jpropionate (27)

A solution of methyl 2-methylpropionate (357 mg, 3.5 mmol) in 5 ml ofTHF was added dropwise to 10.75 ml of freshly prepared LDA (0.32 molarin THF) at _?S0c. The reaction mixture was stirred at this temp. for 15 minand then warmed up to O°C. Subsequently a solution of 26 (180 mg, Immol) in 5 ml of THF was added dropwise and then stirred at this temp.for I h. 10 ml of a satd. aqueous solution of NaHC03 were added and themixture was extracted with ether. The org. layer was dried (MgS04). evap-orated, and the residue was purified by flash chromatography (petroleumether - EtOAc 1:1) to give 170 mg (70%) of 27 as a colorless solid; m.p.99°C.- C13H18N203 (250.3) Calcd. C 62.4 H 7.25 N 11.2 Found C 62.3 H7.27 N 11.1.- IR (KBr): 2980; 2950; 2930; 2870; 1730; 1650 cm-I.- IH_NMR (CDCI3): l) (ppm) = 1.48 (s, 6H, 2 CCH3), 1.86-1.91 (m, 2H, H2C-5),2,01-2.08 (m, 2H, H2C-6), 3.11 (t, J = 6 Hz, 2H, H2C-4), 3.68 (s, 3H,OCH3), 4.14 (t. J = 6 Hz, 2H, H2C-7), 7.69 (s, I H, HC-2).

Receptor Binding Assay

DA receptor binding was performed as described using eH]-SCH2339018) and eH]-spiroperidoI19) as radioligands in concentrations of 0.3nM and 0.5 nM, respectively. In the receptor binding assay for the charac-terization of the DA autoreceptor, eH]-SND 919 (51 Ci/mmol specificactivity) was used in a concentration of 0.5 nM. The experimental proce-dure was performed in analogy to the binding assay with [3H]-spiroperidolas radioligand. For all receptor binding tests rat brain striatum was used.

Modified and Extended Behavioura/-Activity Screening. based on RefL 20,21

All drugs were tested in a minimum of 3 doses (100/50/25 mg/kg) em-ploying 3 mice (male NMRI mice, 20-33 g) for each dosis. For the first 30min after the test compound was injected i.p. (dose volume = 10 m1lkg, in0.9 aqueous NaCI solution or 0.5 tragant suspension) only fundamentalobservations (e.g .. convulsion, Straub tail, respiratory depression) were

noted. Then the animals were investigated, including the following parame-ters: irregular/dyspnoic respiration, vocalization, restlessness, piloerection,retracted sides, abdominal cramps, exophthalmus, twitches, convulsion,pinna response, corneal response, palpebral closure, motor activity, spatialorientation, !ail position, gait, hyperactivity/vocalization after stimulation,aggressivity, tremors, body position, righting reflex, hypothermia, hyper-thermia, lacrimation, dacryorrhea, salivation, sialhorrhea, diarrhea, skincolor, pupil size, light-pupil response, body tone, motor coordination, cata-lepsy, hot plate response, stereotype movements, death. Observations dif-fering from saline treated control animals were observed again after 4 and24 h. Each animal was used for one experiment only.

Three mice (male NMRI mice, 20-33 g) in a macrolon cage (type III)with free access to food and water were placed into a scanner box (RBM 3,MSE/INTRON, Miinchen/Miinsing, Germany) using electromagneticalfield for the measurement at 3.45 pm. After 30 min aqueous 0.9% NaCIsolution was injected i.p. Then the motor activity was recorded by a separ-ate printer for 2 h. Following the same schedule at the following day theprocedure was repeated after injection of the test compound. Totally, 8groups of 3 mice were investigated using each animal only once. Data wereexamined using the Hest for paris of observations according to Studellt andWi/coxon's matched pair signed rank statistic22•23).

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57 (1992).10 N.J. Bach, E.C. Kornfeld, N.D. Jones, M.O. Chaney, D.E. Dorman,

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