Synthesis of a New Scaffold: the 7H,8H-Pyrimido[1,6-b]pyridazin-6,8-dione Nucleus

This paper describes a modified method of preparation of a number of alpha-aryl-alpha-(pyridazin-3-yl)-acetonitriles via the C-arylation reaction of the corresponding carbanionsof phenylacetonitriles using 3-chloropyridazine derivatives. KOH and DMSO were used inthe deprotonation process, which made the reaction very simple and safe to perform.Nitriles were obtained in the hydrolysis reaction to the corresponding alpha-aryl-alpha-(pyridazin-3-yl)-acetamide derivatives, which were next subjected to cyclization to afford the finalproducts. A number of new derivatives of 7H,8H-pyrimido[1,6-b]pyridazin-6,8-dione weresynthesized in the cyclocondensation reaction of respective alpha-aryl-alpha-(pyridazin-3-yl)-acetamides with diethyl carbonate in the presence of EtONa. The structure andcomposition of the new compounds were confirmed by IR, (1)H- and (13)C- NMR analysesand by elemental C, H and N analysis.


Introduction
The imide moiety, which is present in various heterocyclic systems, may have a significant effect on the biological activity of their derivatives.This functionality is a significant structural element of an important group of compounds belonging to the so-called long-chain arylpiperazines (LCAPs), ligands of 5-HT 1A receptors of high affinity (see Figure 1) [1][2][3][4][5].First Buspirone and then Tandospirone were introduced for medicinal purposes; both of them are prominent representatives of the LCAPs group.These drugs, as 5-HT 1A receptor agonists, revolutionized the treatment of anxiety disorder by the mechanism of serotonergic neurotransmission [2,4,[5][6][7][8][9][10].As shown by the investigations to date, the imide group in Buspirone is an element of the nonpharmacophore part and plays an important role in stabilization of the ligand -5-HT 1A receptor complex.Moreover, it also affects the lipophility of the ligand, which in turn has a significant effect on selectivity of the ligand to 5-HT 1A receptor, with respect to other receptors such as 5-HT 2A or α 1 [2,7,9].Our earlier investigations also dealt with the preparation of new heterocyclic systems, such as for instance the derivatives of pyrido [1,2-c]pyrimidine and pyrrole [1,2-a]pyrazine, which have the afore-mentioned grouping [11][12][13].We have now focused our studies on a little known pyrimido [1,6b]pyridazine system, the derivatives of which had already been described by Bemis et al.Our investigations were concerned with Kinase p38 inhibitors which possess antitumor activity [14].The aim of our investigations was to synthesize the new 7H,8H-pyrimido [1,6-b]pyridazin-6,8-dione derivatives 4a-f, 4f′ and 4g′, which contain the imide moiety in their structure.These derivatives should be important substrates for further synthesis of the potential ligands of 5-HT 1A receptors of higher selectivity in the LCAPs group.

Results and Discussion
The compounds described in this paper were obtained according to Scheme 1. Scheme 1. Synthesis of the title compounds and substituents.The starting α-aryl-α-(pyridazin-3-yl)-acetonitriles 2a-h, which were necessary for the synthesis, were obtained in a C-arylation reaction of stabilized carbanions of appropriate arylacetonitriles using 3-chloropyridazine derivatives.A modified method of deprotonation of the above-mentioned arylacetonitriles was applied, using KOH in DMSO.This method was elaborated by us earlier and was used for the synthesis of a number of derivatives of α-aryl-α-(2-pyridyl)-acetonitriles in a C-arylation reaction using 2-bromopyridine [11,12].The aim of the present studies was first of all to simplify the process of C-arylation to obtain a method which gives reproducible yields and is safe to perform.
The modified method used in the present work allowed us to obtain a number of new nitriles 2b-e, 2g, 2h, 2g′ in good yields.During the next stage of our studies the nitriles 2g and 2f, possessing chlorine in position 6 of the pyridazine moiety, were subjected to dehalogenation using ammonium formate and Pd/C in MeOH, instead of the hydrogenolysis process which had been used by Abbotto et al. during preparation of the derivative 2f′ [16].
A number of new derivatives of α-aryl-α-(pyridazin-3-yl)-acetamide 3a-f, 3h, 3f′ and 3g′, were obtained by hydrolyzing the above-obtained nitriles 2a-f, 2h, 2g′, 2f′ in acidic medium.The hydrolysis process was performed under different conditions.The nitriles 2a-f, 2h, 2f′ and 2g′ were hydrolyzed with sulphuric acid at the temperature of 50 °C.Hydrolysis of the nitriles 2c and 2d was performed in the sulphuric acid -acetic acid mixture at the temperature of 100 °C for 1 h for compound 3c, whereas the derivative 3d was obtained at 50 °C and the process was continued for 10 h.
The final compounds, the derivatives of 7H,8H-pyrimido [1,6-b]pyridazin-6,8-dione 4a-f, 4f′ and 4g′ were obtained in good yields in an intermolecular cyclocondensation reaction of the respective amides 3a-f, 3h, 3f′ and 3g′ with diethyl carbonate in the presence of EtONa.During the cyclization process of the derivatives 3f and 3h the formation of one cyclization product 4f was observed as a result of simultaneous chlorine substitution in position 2 of the cyclic compound on the -OEt group in amide 3f, whereas in the case of amide 3h the -OMe group was exchanged into -OEt (see Scheme 2).Scheme 2.
The structures and composition of the new intermediate and final compounds were confirmed by analysis of their IR, 1 H-and 13 C-NMR spectra, and also by elemental C, H and N analyses.The NMR spectra are mutually correlated and were in agreemeent with the literature data for similar systems, as well as with the theoretical spectra calculated according to ACD/NMR Predictor v. 8.09 program.In the amide proton spectra (compounds of series 3) we observed two NH proton signals from the NH 2 group, which could indicate that there is nonequivalence of magnetic surroundings of these protons due to inhibited rotation about the C-N bond.

General
Melting points of the substances were determined on a Mel-Temp® 3.0 (Barnsted/Thermolyne, USA) apparatus and are uncorrected.Elemental analysis was performed on a Perkin-Elmer CHN model 2400 analyzer.The IR spectra (KBr tablets) were performed on a Shimadzu FT IR-8300 apparatus. 1H-and 13 C-NMR spectra were recorded on the following spectrometers: a Bruker Avance DMX WB of basic frequency for the protons: 400.133MHz, for nuclei 13 C: 100.623MHz and a Varian type Unity Plus of basic frequency for the protons: 500.605MHz, for nuclei 13 C: 125.877MHz at room temperature.CDCl 3 was used as solvent and tetramethylsilane as internal standard.The chemical shifts of resonance signals are given in ppm, and the coupling constants are in Hz.Thin-layer chromatography (TLC) was performed on Merck DC-Platten Kiesegel 60 F 254 plates, developed in the dioxane, toluene, ethanol systems, 25% NH 4 OH (6.0:3.2:0.5:0.2, v/v) or chloroform, methanol, diethyl ether, 25% NH 4 OH (6.0:2.0:1.8:0.2, v/v) and visualized on a UV lamp.Analytical samples of the compounds were obtained by purification on the chromatographic column using the flash technique and Merck Kieselgel 60 (230-400 mesh) filling.They were eluted with methylene chloride/methanol mixtures (99:1; 97:3; 95:5, v/v).The starting phenylacetonitrile derivatives, 3,6-dichloropyridazine and 3-chloro-6-methoxypyridazine were commercial products purchased from Aldrich and used without further purification.The other starting reagent 3-chloro-6-phenylpyridazine was prepared by the reported procedure [18,19].

General procedure for the preparation of α-aryl-α-(3-pyridazin-3-yl)acetonitriles 2a-h
To DMSO (32.5 mL) was added KOH (16.5 g, 0.29 mol) and the mixture was stirred for 0.5 h at room temperature.Next an appropriate phenylacetonitrile derivative (0.11 mol) in DMSO (10 mL) was added dropwise and stirring was continued for another 0.5 h at room temperature.Next the appropriate 3-chloropyridazine derivative (0.07 mol) was added portionwise to the mixture, which was stirred at a temperature of 50 °C for 12 h.Next the post-reaction mixture was poured into ice water (1000 mL).The separated precipitates were filtered off.The obtained crude products 2c, 2d, 2e, 2f, 2f′ were purified by flash chromatography using CH 2 Cl 2 /MeOH (97:3 v/v), and then CH 2 Cl 2 /MeOH (99:1 v/v) as eluents, and then compounds 2c crystallized from absolute EtOH, 2d and 2f′ from EtOH.