Synthesis of Novel Aryl(heteroaryl)sulfonyl Ureas of Possible Biological Interest

The course of reaction of aryl and heteroaryl sulfonamides with diphenylcarbonate (DPC) and 4-dimethylaminopyridine (DMAP) was found to depend on the pKa of the sulfonamide used. Aryl sulfonamides with pKa ~ 10 gave 4-dimethylamino-pyridinium arylsulfonyl-carbamoylides, while the more acidic heteroaryl sulfonamides (pKa ~ 8) furnished 4-dimethylaminopyridinium heteroarylsulfonyl carbamates. Both the carbamoylides and carbamate salts reacted with aliphatic and aromatic amines with the formation of appropriate aryl(heteroaryl)sulfonyl ureas, and therefore, can be regarded as safe and stable substitutes of the hazardous and difficult to handle aryl(heteroaryl)sulfonyl isocyanates.

Recently, we have described a facile method for the preparation of arylsylfonyl ureas of general formula C (Scheme 1) using 4-dimethylaminopyridinium arylsulfonyl-carbamoylides B, which constitute non-hazardous substitutes of arylsulfonyl isocyanates [12,13]. As shown in Scheme 1, the method consists in the reaction of aromatic sulfonamides A with diphenyl carbonate (DPC) in the presence of 4-dimethylaminopyridine (DMAP), followed by the reaction with primary or secondary aliphatic or aromatic amine. The above procedure worked well with phenylsulfonamide and its para-substituted congeners, such as p-alkyl, p-methoxy-and p-chloro-phenylsulfonamide as the substrates. The unique structure of the carbamoylides obtained were confirmed by IR and NMR spectra as well as single crystal X-ray structure analysis [13]. Carbamoylides B compose of an appropriate arylsulfonyl isocyanate and a DMAP molecule. The stability of these highly polarizable adducts is mainly due to the delocalization of the positive charge on the pyridine ring and the negative charge on the arylsulfonylcarbamoyl moiety. In order to explore the scope of this procedure, an analogous previously not attempted reaction sequence starting from variously substituted phenylsulfonamides, naphthylsulfonamide as well as heteroarylsulfonamides, such as 2-thienyl-and benzothiazol-2-yl-sulfonamide has now been attempted.

Results and Discussion
The results of the investigations are presented in Scheme 2. First, it was found that the reaction of variously substituted phenylsulfonamides 1a-c (including p-nitrophenyl-, o-chlorophenyl-and 2-naphthylsulfonamide) with DPC in the presence of DMAP proceeded smoothly at room temperature to give appropriate carbamoylides 3a-c in good yields. The reactions of these stable arylsulfonyl isocyanates substitutes carried out in CH 3 CN with both aliphatic and aromatic amines at elevated temperature afforded the desired arylsulfonyl ureas, which could be easily separated from the reaction mixtures upon treatment with 1% aqueous HCl.
However, when 2-thienyl and benzothiazol-2-yl-sulphonamides 1d-f were treated with DPC in the presence of DMAP, the pyridinium carbamates 4a-c were obtained as the sole products. Neither prolonged reaction times nor elevated temperatures changed the reaction course.
The structures of pyridinium carbamoylides 3 and pyridinium carbamates 4 were confirmed by IR and NMR spectroscopic data as well as X-ray single crystal structure analysis of 4b and 4c ( Figure 1). The compounds 4b and 4c are organic salts with the proton transferred from the sulfonylcarbamate group to 4-dimethylaminopyridine. In the crystal the anions and the pyridinium cations form ionic pairs via N-H + ···Nhydrogen bonds. The anions assume similar conformation with one of the SO 2 group O atoms approximately in the plane of benzothiazole moiety and the bond lengths and angles are as expected. The crystal packing is, to a large extent, governed by electrostatic interactions, with a pair of 4-dimethylaminopyridinium cations stacked in anti-parallel manner and completely surrounded by the anions.
The major difference between the two classes of sulfonamide derivatives studied lies in their relative acidity, hence, it was reasoned that the more acidic heteroarylsulfonyl sulfonamides 1d-f (pK a ~ 8) [14] formed the more acidic carbamates 2d-f which, in turn, suffered proton abstraction by DMAP to give pyridinium salts of carbamates 4a-c, while the less acidic arylsulfonamides 1a-c (pK a ~ 10) [15] gave rise to the formation of less acidic carbamates 2a-c which underwent nucleophilic substitution reaction with DMAP to give the desired carbamoylides 3a-c.
In order to confirm the above hypothesis the reaction of DPC/DMAP couple with 4-chloropyridin-3-yl-sulfonamide (1g), characterized by a pK a value of 8.9 [16], was performed. As shown in Scheme 3, the above reaction carried out at ambient temperature afforded two products: the carbamate pyridinium salt 4d and the carbamoylide 3d which could be separated from the reaction mixture by fractional crystallization in 48% and 39% yield, respectively. Interestingly enough, while the desired aryl(heteroaryl)sulfonyl ureas 5 and 6 were obtained in good yields from the reactions of either 3a-c or 4a-c with aliphatic, aromatic and heteroaromatic amines, upon treatment of both the carbamate 4d and carbamoylide 3d with an excess of a secondary amine, 3-(indolin-1-yl)pyrido [3,4- 3]oxathiazine 1,1-dioxide (8) was obtained. This might be formed as a result of an intramolecular nucleophilic substitution reaction in the transiently formed arylsulfonyl ureidate 7 (Scheme 3).
All the newly prepared compounds 5, 6 and 8, including benzothiazol-2-yl analogues of sulofenur, were screened in vitro for their potential cytotoxic activity using human urinary bladder cancer 5637, small cell lung cancer A-427 and large cell lung cancer LCLC-103H cell lines. None of these compounds exhibited cytotoxic activity at concentrations below 20 μM.

General
Melting points were measured on a Boetius 545 apparatus and are not corrected. The IR spectra were obtained on a Nicolet 380 FTIR spectrometer using potassium bromide pellets and the frequencies were quoted in cm -1 . The 1 H-NMR spectra were recorded on a Varian Gemini spectrometer at 200 MHz or a Varian Unity Plus apparatus at 500 MHz. The chemical shifts (δ) are expressed in ppm in relation to tetramethylsilane as a standard and the coupling constants (J) are given in Hz. The starting substrates were commercial reagents.

General procedure for the preparation of 4-dimethylaminopyridinium arylsulfonyl carbamoylides 3a-c and 4-dimethylaminopyridinium heteroarylsulfonyl carbamates 4a-c
A solution of appropriate arylsulfonamide 1a-c (33 mmol) in acetonitrile (40 mL) was treated with 4-dimethylaminopyridine (DMAP, 4.1 g, 66 mmol) and the reaction mixture was stirred at room temperature until sufonamide 1 had dissolved. Then diphenyl carbonate (DPC, 2.3 g, 37 mmol) was added and the reaction mixture was left overnight at room temperature. The solid that precipitated was separated by suction, washed with dry acetonitrile and dried to give pure carbamoylides 3a-d or carbamates 4a-d.
According to the above procedure the following compounds were obtained: 3.3. General procedure for preparation of arylsulfonyl ureas 5 and heteroarylsulfonyl ureas 6. Reaction of carbamoylides 3a-c and carbamates 4a-c with aliphatic and aromatic amines A mixture of carbamoylide 3 or carbamate 4 (2.8 mmol) and appropriate aliphatic or aromatic amine (3 mmol) in acetonitrile (10 mL) was heated at reflux for 10 min (in case of aliphatic) or 1 h (in case of aromatic) amine. The reaction mixture was cooled to room temperature and the solvent was evaporated under reduced pressure to dryness. The oily residue was suspended in methanol and treated with 10% aqueous hydrochloric acid. The sulphonylureas 5 or 6 that precipitated were separated by suction, washed with methanol and water and re-crystallized from suitable solvent.

X-ray structure analysis
The diffraction data were collected with a KumaCCD diffractometer using graphite monochromated Mo K α radiation. The intensity data were collected and processed using Oxford Diffraction CrysAlis Software [17]. The crystal structures were solved by direct methods with the program SHELXS-97 [18] and refined by full-matrix least-squares method on F 2 with SHELXL-97 [18].
Crystal data for C 14  Crystal data for C 16  Crystallographic data for compounds 4b and 4c have been deposited with Cambridge Crystallographic Data Centre (CCDC deposition numbers CCDC 742624-742625). Copies of the data can be obtained upon request from CCDC, 12 Union Road, Cambridge CB2 1EZ, UK, quoting the deposition numbers.

In vitro cytotoxicity assay
The following primary screening of the new compounds was done to indicate whether a substance possesses enough activity at the concentration of 20 μM to inhibit human tumor cell growth by 50% (GI 50 < 20 μM).
The in vitro cytotoxic activity of all arylsulfonylureas 5a-j and heteroarylsulfonylureas 6a-i were evaluated [19,20] using human urinary bladder cancer 5637, small cell lung cancer A-427 and large cell lung cancer LCLC-103H cell lines. The assay was carried out in 96-well microtiter plates. When the cells were putted into the plates after 24 h cells were treated with appropriate drug solutions. The cytotoxic effects of the compounds were measured after a 96 h continuous exposure to the substances. The cell growth inhibition values were estimated by staining the adherent cells with crystal violet. Only viable cells remained attached to the plastic surface of the wells and bind the dye. The unbound dye was washed out with water and stain remaining in the wells was redissolved with 70% ethanol. Finally, the optical density (OD) was measured with a microplate reader set at λ = 570 nm.

Conclusions
The current work has addressed the use of environmentally non hazardous aryl(heteroaryl)sulfonamides and diphenyl carbonate (DPC) in the synthesis of 4-dimethylaminopyridinium N-[aryl(heteroaryl)sulfonyl] carbamoylides of type 3, the stable and easy-to-handle substitutes of aryl(heteroaryl)sulfonyl isocyanates. The comparison of the existing literature methods for the preparation of arylsulfonyl/heteroaryl carbamates and ureas from arylsulfonamides suggests that the DPC/DMAP approach is superior. Some of the advantages include mild reaction conditions, the ease of preparation and product separation and the extended shelf-life of the parent ylides. Moreover, a very high reactivity of pyridinium carbamoylides renders them suitable for the syntheses of arylsulfonyl ureas. It should be emphasized, however, that in the above procedure diphenyl carbonate (DPC) could not be replaced by the less reactive diethyl or dimethyl carbonates.
It has been also found that the course of the reaction between sulphonamide and DPC/DMAP couple depends on pK a of the substrate. Thus, arylsulphonamides with pK a in the range of 9-10 give the desired carbamoylides 3, while the more acidic heteroarylsulfonamides (pK a ~ 8) react preferentially with the formation of pyridinium salts of the intermediary formed carbamates 4.