Organotin Compound Derived from 3-Hydroxy-2-formylpyridine Semicarbazone: Synthesis, Crystal Structure, and Antiproliferative Activity

The novel diphenyltin(IV) compound [Ph2(HyFoSc)Sn] (2), where H2HyFoSc (1) is 3-hydroxy-2-formylpyridine semicarbazone, was prepared and characterized by vibrational and NMR (1H, 13C) spectroscopy. The structure of [Ph2(HyFoSc)Sn] was confirmed by single-crystal X-ray crystallography. The doubly deprotonated ligand is coordinated to the tin atom through the enolic-oxygen, the azomethine-nitrogen, and phenolic-oxygen, and so acts as an anionic tridentate ligand with the ONO donors. Two carbon atoms complete the fivefold coordination at the tin(IV) center. Intermolecular hydrogen bonding, C–H → π, and π → π interactions combine to stabilize the crystal structure. Compounds 1 and 2 have been evaluated for antiproliferative activity in vitro against the cells of three human tumor cell lines: MCF-7 (human breast cancer cell line), T24 (bladder cancer cell line), A549 (nonsmall cell lung carcinoma), and a mouse fibroblast L-929 cancer cell line.


Introduction
Organotin compounds are of interest in view of their considerable structural diversity [1]. Increasing interest in organotin(IV) chemistry has arisen in the last few decades and is attributed to their significantly important biological properties. Several di-and tri-organotin species have shown potential as antineoplastic and antituberculosis agents [2][3][4][5]. The binding ability of organotin compounds towards DNA depends on the coordination number and nature of groups bonded to the central tin atom. The phosphate group of DNA-sugar backbones usually acts as an anchoring site and DNA base-nitrogen binding is extremely effective and this often results in the stabilization of the octahedrally coordinated tin center. Recent studies have showed that low doses of organotins can exhibit antitumoral activity and have suggested a mode of action via a gene-mediated pathway in the cancer cells, opening a new research subarea on organotin compounds [6].
Thio-and semicarbazones (TSC) possess a wide range of bioactivities, and their chemistry and pharmacological applications have been extensively investigated. The more significant bioactivities of a variety of semicarbazones (antiprotozoa, anticonvulsant) and thiosemicarbazones (antibacterial, antifungal, antitumoral, antiviral) and their metal complexes have been reviewed together with proposed mechanisms of action and structure-activity relationships [7,8]. Casas et al. [9] have surveyed structural aspects of main group metal complexes of semicarbazones and thiosemicarbazones. The survey shows that heterocyclic and nonheterocyclic TSC's are very versatile coordination agents with these elements [9].
Following our interest in the chemistry and pharmacological properties of thiosemicarbazones [10][11][12][13][14][15][16][17] and towards organotins [18][19][20][21], herein, the preparation and spectroscopic characterization of a novel semicarbazone and a novel diphenyl organotin compound derived from the reaction of SnPh 2 O with 3-hydroxy-2-formylpyridine semicarbazone H 2 HyFoSc (1) are described with the final goal of developing new biologically active pharmaceuticals. The results of the cytotoxic activity of 1, SnPh 2 O, and of the organotin compound (2) against the cells of three human cancer cell lines: MCF-7 (human breast cancer cell line), T24 (bladder cancer cell line), A549 (non-small cell lung carcinoma), and a mouse fibroblast L-929 cancer cell line are also reported. To our knowledge, this is the first report of synthesis of 1 and 2.

X-Ray Crystallography.
Crystal data are given in Table 1, together with refinement details. All measurements were performed on a Kuma KM4CCD kappa-axis diffractometer with graphite-monochromated MoKα radiation (λ = 0.71073Å). The data were corrected for Lorentz and polarization effects. An analytical absorption correction was applied to the data using a multifaceted crystal model [24]. Data reduction and analysis were carried out with the Kuma Diffraction (Wroclaw) programs. The structure was solved by directmethods and refined by a full-matrix least-squares method on all F 2 data using the SHELXL97 [25]. Nonhydrogen atoms were refined with anisotropic displacement parameters; all hydrogen atoms were located from different Fourier maps. The C-bound H atoms were refined with the riding model approximation, while the N-bound H-atoms were freely refined isotropically. Molecular graphics were performed with PLATON 2004 [26].
Crystallographic data for 2 have been deposited with the Cambridge Crystallographic Data Centre, CCDC, 634269 for compound 3. Copies of this information may be obtained free of charge from The Director, CCDC, 12, Union Road, Cambridge CB2 1EZ [FAX +44(1223)336-033] or e-mail deposit@ccdc.cam.ac.uk or http://www.ccdc.cam.ac.uk.

Antiproliferative Assay In Vitro
Compounds. Test solutions of the tested compounds (1 mg/mL) were prepared by dissolving the substance in 100 μL of DMSO completed with 900 μL of tissue culture medium. Afterwards, the tested compounds were diluted in culture medium to reach the final concentrations of 100, 50, 10, 1, and 0.1 ng/μL. The solvent (DMSO) in the highest concentration used in the test did not reveal any cytotoxic activity.
Cells. The cell lines are maintained in the Cell Culture Collection of the University of Ioannina. Twenty-four hours before addition of the tested agents, the cells were plated in 96-well plates at a density of 10 4 cells per well. The  [27], while T24 cells by the MTT assay [28].The in vitro tests were performed as described previously [29].

Results and Discussion
3.1. Synthesis. Compound 1 was synthesized by means of the Heinert-Martell reaction (Scheme 1) [22]. The corresponding diorganotin compound 2 was prepared by reacting diorganotin(IV) oxide with the semicarbazone in benzene solution in a 1 : 1 molar ratio.

Crystal Structure of 2.
A perspective view of 2, together with the atom-labelling scheme, is given in Figure 1 and selected bond lengths and angles are given in Table 2.
The doubly deprotonated ligand is coordinated to the tin atom through the enolic-oxygen, azomethinenitrogen, and phenolic-oxygen atoms. Two carbon atoms complete the fivefold coordination at the diorganotin(IV) fragment. Analysis of the shape determining angles using the approach of Addison et al. [30] yields τ = 0.54 (τ = 0.0 and 1.0 for SPY and TBPY geometries, resp.). The metal coordination geometry is therefore described as distorted trigonal bipyramidal with the O(2) and O(1) atoms occupying the apical positions around the tin atom.
The dianionic, tridentate ONO ligand has a ZEZ configuration, Figure 1. The coordinated part of the ligand is made of three rings, two chelates Sn (1) Table 2, are similar to other organotin complexes presented in the literature [18][19][20][21].
The polar hydrogen atoms on N(3) participate in two intermolecular hydrogen bonds. The monomers of 2 are connected into dimers by a pair of cooperative hydrogen bonds [see Table 3 for geometric parameters describing these interactions]. Further, adjacent dimers are connected by C-H· · · N contacts, as illustrated in Figure 2. The presence of additional C-H → π and π → π contacts further stabilizes the crystal structure.

Spectroscopic Studies.
In the IR spectrum of H 2 HyFoSc (1), the strong bands at 3147 and 3088 cm −1 are assigned to the asymmetric and symmetric modes of terminal NH 2 , respectively. The ν(NH) band appears at 2920 cm −1 . The strong broad band at ca. 2650 cm −1 is assigned to the ν(NH· · · O) and ν(OH· · · O) mode due to strong intraor intermolecular hydrogen bonding. The absence of the ν(NH) stretching motion at 2 is indicative of deprotonation of the amide proton. The strong broad band at ca. 2650 cm −1 at 2 is probably due to the ν(NH· · · N) intermolecular hydrogen bonds as confirmed by X-ray crystallography. The coordination of the azomethine-N atom to the tin center was suggested in the IR spectrum by a shift of the ν(C=N) band to a lower frequency, along with the occurrence of a ν(N-N) band to higher frequency [12,13,16]. An IR band at 1322 cm −1 for 1 was assigned to ν(C-O). This band was found to be shifted to 1283-1289 cm −1 , in the spectrum of 2, which indicates the coordination of this O atom. The low energy of the ν(C=O) vibration in the spectra of 1, that is, 1664 cm −1 , is indicative that the carbonyl O-atom is involved in hydrogen bonding. The replacement of the hydrogen by the metal atom does not shift this band to lower frequency. Coordination of the imine nitrogen is also consistent with the presence of a band at 446 cm −1 , assignable to ν (Sn-N).    Table 3: Geometric parameters for hydrogen bonds and for C-H-π and π · · · π interactions in 2.
The IC 50 values for H 2 HyFoSc (1)  against the L-929 and T-24 cell lines IC 50 are >555 μM. Thus, 1 is considered as nonactive against these tested cell lines. Compound 2 is also considered nonactive against T24 cell line. The IC 50 values for 2 against the L-929 and MCF-7 cell lines are 1.19 and 8.65 μM, respectively. These values are in the same range as observed for cisplatin and Ph 2 SnO, indicating that the observed cytotoxicity is probably due to the cytotoxicity of Ph 2 SnO. Compound 2 may then be considered as a vehicle for activation of the Ph 2 SnO as the cytotoxic agent. The IC 50 value for 2 against A-549 is 0.086 μM and therefore 2 is significantly more active compared to Ph 2 SnO (47.1 μM) and cisplatin (1.53 μM), respectively. Compound 2 is thus 547.7 and 17.8 times more cytotoxic than the Ph 2 SnO and cisplatin, respectively, against this cell line. Thus, 2 exhibits selectivity and is considered as an agent with potential antitumor activity against A-549 tumor cell line and can therefore be a candidate for further stages of screening in vitro and/or in vivo.