1314
J . Med. Chem. 1985, 28, 1314-1319
Benzodiazepine Receptor Binding and Anticonflict Activity in a Series of 3,6-Disubstituted Pyridazino[4,3-c]isoquinolines Devoid of Anticonvulsant Properties Emilio Toja,*' Giorgio Tarzia,' Domenico Barone,: Franco Luzzani,: and Licia Gallicd Departments of Chemistry and Pharmacology, Lepetit Research Center, 20158 Milano, Italy. Received November 6, 1984
A series of 3,6-disubstituted pyridazino[4,3-c]isoquinolineswere synthesized and tested for their ability to inhibit the binding of [3H]diazepamto rat brain receptors in vitro. Compounds bearing a phenyl, 4-methoxyphenyl, or methyl group at position 3 and a dialkylamino group at position 6 showed the highest affinity in the binding assay and were subsequently evaluated for their anticonflict and anticonvulsant effects. All of these compounds (5a-1 and 5q) were active in the Vogel rat conflict procedure, but none prevented convulsions in mice induced either by metrazol or bicuculline. 3-Phenyl-6-pyrrolidinylpyridazino[4,3-c]isoquinoline (5d)with a Ki = 11.4 nM in the binding assay exhibited the best potency in the anticonflict assay (MED 5 mg/kg ip) and did not produce neuromuscular impairment a t the highest dose tested (50 mg/kg ip).
The search for antianxiety agents that are devoid of the side effects associated with the benzodiazepines (BZs) has led to the discovery of various non-BZ classes of compounds with high in vitro activity in the BZ binding assay. Various members of these classes are being actively investigated as potential anxiolytics.' It is not apparent, however, how structurally unrelated chemical entities can interact with the same BZ receptor, nor is it understood why slight structural modifications can produce agonists and antagonists from the same parent compound.* The discovery of the high BZ binding site affinity of several 3-aryl-1,2,4-triazolo[3,4-a]phthalazines (I)3 and 2-arylpyrazolo [4,3-c]quinolin-3(5W-ones (11)2prompted us to synthesize a series of 3-arylpyridazino[4,3-c] isoquinolines (111) and evaluate their binding properties to this receptor (Chart I). On the basis of the structural similarity of the three classes of compounds it could reasonably be expected that they would interact with the BZ receptors in a similar way. Thus, a limited number of compounds belonging to class I11 were prepared, and their ability to displace [3H]diazepam (DZ) was assessed. Results of this study indicated that the enlargement of ring C from pyrazolo or triazolo to pyridazine did not greatly affect the binding. A series of derivatives of class I11 was then synthesized in order to clarify the influence of the substituents in positions 3 and 6 on the biological activity. Chemistry. We previously reported4 that the hydrazones obtained by condensation of N-aminophthalimidine with ethyl benzoylacetate or acetoacetate undergo a sodium ethoxide promoted rearrangement to give 4(1H)pyridazones la or Id (Scheme I) as the main products. This procedure was also employed in the conversion of the hydrazones prepared from 4-methoxy- and 4-chloro-substituted benzoylacetatesj to yield lb and IC, respectively. Lactonization of la-d was achieved either with an equimolar amount of dicyclohexylcarbodiimidein refluxing pyridine? or with acetic anhydride in toluene with the azeotropic removal of acetic acid. Treatment of 2a-d with an excess of dry ammonium acetate at ca. 200 "C in a steel cylinder quantitatively gave lactams 3a-d, which were then treated with phosphorus pentachloride and phosphorus oxychloride to yield 4a-d. The physical properties of intermediates 1-4 are shown in Table I. Substitution of the chlorine atom of 4a-d with various dialkylamines gave 64dialkylamino)pyridazino[4,3-c]isoquinolines 5 while 6-alkoxypyridazino[4,3-c]iso*Present address: Roussel Maestretti,Via Gran Sasso 18,20131 Milano, Italy. Department of Chemistry. ? Department of Pharmacology. 0022-2623/85/1828-1314$01.50/0
Chart I. 1,2,4-Triazolo[3,4-a]phthalazines (I), Pyrazolo[4,3-c]quinolin-3( 5H)-ones (11), and Pyridazino[4,3-c]isoquinolines(111)"
k
R
R 11.
Key: Ar = substituted phenyl; R = dialkylamino, alkoxy, hydrogen. a
Scheme I"
1 a-d
2 a-d
1
,efl \
/
PCI5-P0CI3
3bd
dn
CI
5 6
R R
7
R
i
=
dlalkylamino alkoxy
4
a-d
- H
a For compounds 1-4: a, R = C,H, ;b, R = 4OCH,C,H, ; C, R = 4-C1C,H4; d, R = CH,.
quinolines 6 were obtained by reaction of 4a-d with sodium alkoxides in the corresponding alcohol. Hydrogenation of 4a, in the presence of palladium on carbon with magnesium oxide as an acid acceptor, caused displacement of the chlorine atom and saturation of the 5,6 double bond. Oxidation of the dihydro derivative to 7 was achieved by treatment with an ethanolic solution of iodine and po(1) The subject has been reviewed by: Williams, M. J. Med. Chem. 1983, 26, 619. Martin, I. L. Trends Pharmacol. Sci. 1984, 5, 343. (2) The pyrazoloquinolines CGS 9896 and CGS 8216, which differ in a chlorine atom, are respectively an agonist and antagonist at the BZ receptor as reported by: Yokoyama, N.; Ritter, B.; Neubert, A. D. J. Med. Chem. 1982, 25, 337. (3) Occelli, E.; Barone, D.; Tarzia, G.; Giunta, A. Eur. Pat. A p p l . 85840, 1983. (4) Toja, E.; Omodei-Sale', A,; Nathansohn, G. Tetrahedron Lett. 1979, 2921. (5) Wallingford, V. H.; Homeyer, A. H.; Jones, D. M. J . Am. Chem. SOC.1941, 63, 2252.
0 1985 American Chemical Society
Journal of Medicinal Chemistry, 1985, Vol. 28, No. 9 1315
Pyridazino[4,3-c]isoquinolines Table I. Intermediates 1-4 of Scheme I
'H NMR spectral datac 1, HE, br, 1, NH 13.2-15.0 C17H12N203 6.62 C18H14N204 6.60 13.47 (9) IC C17H13C1N204d 6.72 13.8- 14.2 Id Ci~HitA"03 6.22 12.4-13.6 'H NMR spectral data' no. vield." % mtx "C cryst solvent formulab 9, 1, H, dd, 1,Hin s, 1, NH 2a 92 205-206 AcOEt C17H10N202 7.65* 8.94 ( J = 8.5, 1.5) 2b 84 230-231 AcOH C18H12N203 8.32 8.88 ( J = 9, 1.5) 2C 83 273-275 AcOH C17H9C1N202 8.49 9.03 ( J = 8.5, 1.5) 2d 97 218-219 AcOEt ClZH8N202 7.35* 8.95 ( J = 8, 1.5) 3a 97 340-342 EtOH C17HllN30 7.65 8.87 (J = 8.5, 2) 11.87 3b 94 314-3 15 AcOH C18H13N302 7.80 9.07 ( J = 9, 1.5) 12.20 >350 3c 71 C17H10C1N30 7.82 9.04 ( J = 9, 1.5) 12.27 AcOH >350 3d 96 ClZH9N30 7.88 8.76 ( J = 8, 1.5) 13.18 DMF 177-178 4a 94 C17H10C1N3e 8.37* 9.53 ( J = 9, 1.5) MezCO 224-227 MeC6H5 C18H12ClN30f 8.83 9.51 ( J = 8.5, 1.5) 4b 98 228-230 C17H9C12N3 8.94 9.53 ( J = 8.5, 1.5) 4c 98 MeC6H5 7.86* 9.50 ( J = 9, 1.5) 4d 89 168-169 MezCO C12H8C1N3 The a The yield is based on recrystallized compounds for 1 and 4 and on crude reaction products for 2 and 3. See Experimental Section. compounds were analyzed for C, H, and N; analytical results were within &0.4% of theoretical values except for 4a and 4b. "Chemical shifts in 6 for the indicated protons in MezSO-d6 (* in CDCl,); coupling constants, J ortho and meta, in Hz. dObtained as monohydrate; desolvation at 130-150 "C and decomposition at 272 "C as shown by DSC. 'C: calcd, 69.99; found, 69.44. f C : calcd, 67.19; found, 67.70. no. la lb
yield," % 77 74 79 66
mp, "C 214-216 dec 224-227 dec 270-272 dec 209-210 dec
cryst solvent EtOH MeOH MeOH EtOH
formulab
8,
~
tassium acetate. The physical properties of pyridazino[4,3-c]isoquinolines 5-7 are listed in Table 11. Biological Results and Discussion. The in vitro activity in the BZ binding assay of 3,6-disubstituted pyridazin0[4,3-~]isoquinolines is shown in Table 11. By keeping the phenyl ring constant in position 3 we determined the effect of the 6-substituent of this series in inhibiting the specific [3H]DZ binding. In contrast to the unsubstituted compound 7, which has a very low affinity for this receptor, it appeared that compounds possessing dialkylamino groups (5a-e) show strongly enhanced affinity with Ki values lower than those obtained for Medazepam and C1218872. Members of this series bearing an alkoxy group (6a-b) show an affinity approaching lactam 3a, whereas the presence of a chlorine atom (4a) diminishes affinity as compared to 3a. Therefore, compounds bearing 6-dialkylamino groups and selected substituents on the phenyl ring were investigated. The p methoxy and -chloro substituents were considered in order to compare the affinities of these compounds with those reported for classes I3 and 112 (Chart I). Compounds having the 4-OCH3 group (5f-1) showed a lower affinity than the corresponding compounds with the unsubstituted phenyl, whereas the presence of the chlorine group practically abolished binding affinity in these molecules (5mp). Thus, the substituent in the 3-position also influences the interaction of these compounds with the BZ receptors. The unexpected good affinity of 5q, which possesses a methyl group a t the 3-position, precludes any correlation of structure and activity on the basis of steric or electronic effects. The influence of the various dialkylamino groups on BZ receptor binding can be ranked in decreasing order: 1pyrrolidinyl > dimethylamino = azetidinyl > N(CH3)CH,CHOHCH, > 4-morpholinyl > 1-piperidinyl > N(CH2CH20CHJ2 in the series of 3-aryl-substituted compounds. To ascertain whether members of this series with Ki's
