Arylidene pyruvic acids motif in the synthesis of new thiopyrano[2,3-d]thiazoles as potential biologically active compounds

Chemistry

The synthesis of starting 5-arylidene-4-thioxo-2-thiazolidinones (5-arylideneisorhodanines) 1a–d was accomplished by the reaction of 4-thioxo-2-thiazolidinone with appropriate aldehydes in glacial acetic acid in the presence of a catalytic amount of fused sodium acetate [8, 10]. Same results were obtained for the reaction conducted in ethanol in the presence of ethylenediaminediacetate (EDDA). The APAs were synthesized by the reaction of a corresponding aromatic aldehyde with a pyruvic acid in an aqueous methanol solution of KOH [16]. The hetero-Diels-Alder reaction of 2a–f with 5-arylidene-4-thioxo-2-thiazolidinones 1a–d regioselectively and diastereoselectively yielded a series of novel rel-(5R,6S,7S)-2-oxo-5,7-diaryl-3,5,6,7-tetrahydro-2H-thiopyrano[2,3-d]thiazol-6-yl-oxo-acetic acids (Scheme 2).

Scheme 2

Synthesis of 2-oxo-5,7-diaryl-3,5,6,7-tetrahydro-2H-thiopyrano[2,3-d]thiazol-6-yl-oxo-acetic acids.

The structure of the newly synthesized compounds was confirmed by elemental analysis and spectroscopic data (¹H NMR and ¹³C NMR). The stereochemical features of the above hetero-Diels-Alder reaction can be predicted. In particular, we have observed that the use of APAs in the [4+2]-cyclocondensation with 5-arylideneisorhodanines leads to a pair of rel-(5R,6S,7S)-diastereomers. This claim is based on the coupling constant values within the range of 10.4–10.8 Hz and the observed spectral patterns of the thiopyran fragment (triplet and two doublets at 4.18–4.85 ppm), which show an axial-axial interaction of the 5-H, 6-H and the 6-H, 7-H proton pairs. Importantly, a similar pattern was observed earlier for cinnamic acids and their amides as dienophiles in the hetero-Diels-Alder reactions [9–11].

Evaluation of anticancer activity in vitro

Synthesized rel-(5R,6S,7S)-2-oxo-5,7-diaryl-3,5,6,7-tetrahydro-2H-thiopyrano[2,3-d]thiazol-6-yl-oxo-acetic acids 3e and 3c were selected by the National Cancer Institute (NCI) Developmental Therapeutic Program (www.dtp.nci.nih.gov) and evaluated for antitumor activity at 10-μm concentration toward a panel of approximately 60 cancer cell lines. The human tumor cell lines were representing leukemia, melanoma, lung, colon, central nervous system, ovarian, renal, prostate, and breast cancers. Anticancer assays were performed according to the NCI protocol, which is described elsewhere [17–19]. The compounds were added at a single concentration, and the cell cultures were incubated for 48 h. The results for each compound are reported as the percentage of growth (GP%) of treated cells when compared with untreated control cells. The screening results are shown in Table 1.

The tested compounds 3e and 3c did not show significant activity in almost cancer cell lines. Nevertheless, slight in vitro cytostatic effect was observed in the growth of SK-MEL-2 (melanoma), TK-10 and UO-31 (renal cancer), and T-47-D (breast cancer). Moreover, 3e possessed moderate effect on the colon cancer HT-29 cell line (GP% = 53.15%).

Antiexudative activity

For antiexudative tests, adult male and female rats weighing 140–150 g were used. The carrageenan-induced hind paw edema in rats was produced using the method of Winter et al. [20]. Diclofenac sodium (10 mg/kg) and ketorolac tromethamine (10 mg/kg) were used as reference compounds. Carrageenan solution (1.0% in sterile 0.9% NaCl solution) was injected subcutaneously into the subplanar region of the hind paw (in volume of 0.1 mL to each paw) 1 h after administration of the test compound. The synthesized compounds were intraperitoneally injected (1 h before carrageenan) in a dose 100 mg/kg. Control rats only received a solution of 0.5% carboxymethylcellulose with one drop of Tween-80. The hind paw volume was measured with an electronic onkograph immediately before and 4 h after carrageenan injection. The effect of the test compounds on decreasing paw edema was compared with control rats. The antiexudative activity was expressed as a decrease in rat paw edema and is given in percentage (Table 2).

Among tested compounds, 3b was found to be the most active. It decreased carrageenan-induced edema on 42.1%. This value is comparable with the effect of reference compounds. For other derivatives, 3a, 3d, and 3g, moderate antiexudative activity was observed.

Chemistry

All materials were purchased from commercial sources and used without purification. Melting points were measured in open capillary tubes and are uncorrected. Elemental analyses were performed using Perkin-Elmer 2400 CHN analyzer. The ¹H NMR (400 MHz) and ¹³C NMR (100 MHz) spectra were recorded on Varian Gemini 400 or Bruker 125 in DMSO-d₆ using tetramethylsilane as internal standard. The purity of all obtained compounds was checked by thin-layer chromatography.

The starting compounds, 2,4-thiazolidinedione [21] and 4-thioxo-2-thiazolidinone [22], were obtained as described previously. 5-Arylidene-4-thioxo-2-thiazolidinones 1a–d were prepared by Knoevenagel condensation using method A or B.

General procedure for hetero-Diels-Alder reaction affording 3a–g

A mixture of appropriate 5-arylidene-4-thioxo-2-thiazolidinone (5 mmol), APA (5.5 mmol), and a catalytic amount of hydroquinone (2–3 mg) in 15 mL of glacial acetic acid was heated under reflux for 4–7 h and then left overnight at room temperature. The resultant solid product was collected by filtration, washed with water, methanol (5–10 mL), diethyl ether, and crystallized from acetic acid or ethanol.

Cytotoxic activity against malignant human tumor cells

Anticancer in vitro assay was performed on the human tumor cell lines panel derived from nine neoplastic diseases, in accordance with the protocol of the Drug Evaluation Branch, National Cancer Institute, Bethesda, MD, USA [17–19]. Tested compounds were added to the culture at a single concentration (10^-5m), and the cultures were incubated for 48 h. End point determinations were made with a protein binding dye, sulforhodamine B. Results for each tested compound were reported as the GP% of the treated cells when compared with the untreated control cells. GP% was evaluated spectrophotometrically vs. controls not treated with test agents.