Interaction of 5′-Guanosine Monophosphate with Organotin(IV) Moieties: Synthesis, Structural Characterization, and Anti-Inflammatory Activity

Reaction(s) of 5′-guanosine monophosphate (5′GMP) with di- and triorganotin(IV) chloride(s) led to formation of organotin(IV) derivatives of general formulae, [R2Sn(5′-GMP)·H2O]n and [(R′3Sn)2(5′-GMP)·H2O]n, where R = Me, n-Bu, and Ph; R′ = Me, i-Pr, n-Bu, and Ph; (5′-GMP)2− = 5′-guanosine monophosphate. An attempt has been made to prove the structures of the resulting derivatives on the basis of FT-IR, multinuclear 1H, 13C, and 119Sn NMR and 119Sn Mössbauer spectroscopic studies. These investigations suggest that both di- and triorganotin(IV)-5′-guanosine monophosphates are polymeric in which (5′-GMP)2− is bonded through phosphate group resulting in a distorted trigonal bipyramidal geometry around tin. The ribose conformation in all of the derivatives is C3′-endo, except diphenyltin(IV) and tri-i-propyltin(IV) derivatives where it is C2′-endo. All of the studied derivatives exhibited mild-to-moderate anti-inflammatory activity (~15.64–20.63% inhibition) at 40 mg kg−1 dose and LD50 values > 400 mg kg−1 in albino rats.


Introduction
The field of cancer chemotherapy has been developed enormously during the past fifty years. Prior to 1969, however, the arsenal of chemotherapeutic agents was devoid of compounds which are inorganic in nature because of generally accepted belief that most metals and their compounds were potentially carcinogenic [1]. In 1969, Rosenberg and his coworkers made the serendipitous discovery [2] that certain Pt compounds were potent antitumor agents against Sarcoma 180 tumors and L1210 leukemia in mice and must be considered to be an outstanding development in the field of metal compounds in medicine [3]. Cis-platin is the first drug from inorganic chemistry to have come under routine clinical use in medical oncology [3]; in 1986, it was the largest selling anticancer drug worldwide. Its success placed the coordination chemists on the front line in the fight against cancer and stimulated the search for other metal-containing compounds with potential anticancer activity. In last 20 years about more than 12000 complexes of 55 metals have been tested [4], many of them are now entering for clinical trials, and some may ultimately rival cis-platin [5][6][7]. Although the majority of these successes involved complexes containing transition metal ions such as Cr, Co, Cu, Pd, Rh, Ru, and Au [5][6][7][8], but some main group metals (i.e., Al, Ga, In, ad Tl; Ge, Sn, and Pb; Bi and Po) compounds [1], especially organotins, have also been discovered which show promise as future members of man's anticancer arsenal [9][10][11][12][13]. Further, several organotin(IV) derivatives have been reported to exhibit good anti-inflammatory activity [14][15][16][17][18][19][20][21][22].
The mechanism of mode of action of cis-platin is due to the formation of an intrastrand crosslink with DNA, involving the N7 of two guanine residues [23,24]. The mode of action of organotin compounds is not very well documented. In order to obtain a better insight about the interaction of organotins with DNA inside the biological systems, their studies with basic constituent units of DNA are indispensable. In view of this, some studies on organotinnucleotides both in solid-state and in solution have been carried out [25][26][27][28][29][30]. Stannylated ribonucleotides in the 2 ISRN Organic Chemistry presence of iodine as activating agent have been used in chemical synthesis of m 7 G 5 ppNu (Nu = A, G, C, and U) [31]. In continuation to our recent studies on the interaction of organotin(IV) moieties with guanine [21] and guanosine [22], in this paper, we wish to report the results of the interaction of 5 -guanosine monophosphate with tri-and diorganotin(IV) moieties.

Experimental
Solvents were dried and distilled before use. Dimethyltin(IV) dichloride di-n-butyltin(IV) dichloride, diphenyltin(IV) dichloride, trimethyltin(IV) chloride, tri-i-propyltin(IV) chloride, tri-n-butyltin(IV) chloride, triphenyltin(IV) chloride (E. Merck), di-n-octyltin(IV) oxide (Aldrich), and disodium salt of 5 -guanosine monophosphate (Na 2 (5 -GMP) (Sigma) were used as received. The elemental analysis, namely, melting points, carbon, hydrogen, nitrogen, and tin of the synthesized compounds was determined on the same instruments as reported earlier [21,22]. Infrared and far-infrared spectra were recorded on a Perkin-Elmer 1600 series FT IR spectrophotometer in the range 4000-400 cm −1 from KBr discs and 600-200 cm −1 from CsI discs. 1 H and 13 C spectra were recorded on a Bruker DRX 300 (300 MHz FT NMR) spectrometer at the Central Drug Research Institute, Lucknow, India, using CD 3 OD as solvent and TMS as the internal standard. 119 Sn NMR spectra were recorded on a Bruker DRX 500 (500 MHz FT NMR) spectrometer at the Institute Instrumentation Centre, IIT, Roorkee, India, using DMSO-d 6 /CD 3 OD as solvent and TMS as the internal standard [21,22]. 119 Sn Mössbauer spectra were recorded on Mössbauer spectrometer model MS-900 according to the procedure reported previously at the Department of Chemistry and Physics, University of The District of Columbia, Washington, DC, USA [21,22]. Toxicity (LD 50 : average lethal dose at 50% survival) and anti-inflammatory activity of the studied derivatives were determined according to the procedures reported earlier [21,22].

General Method for Synthesis of Triorganotin(IV)
Derivatives of (5 -GMP) 2− . The procedure for the syntheses of triorganotin(IV) derivatives of (5 -GMP) 2− was same as discussed in the previous paragraph using the stoichiometric ratio of Na 2 (5 -GMP) and triorganotin(IV) chloride equal to 2 : 1. 4'
All of the synthesized compounds are obtained as white or cream solid in 65-81% yield and stable towards air and moisture. They are insoluble in common organic solvents but sparingly soluble in DMSO. They decomposed at high temperature instead of melting, which indicates their polymeric nature. The analytical data of all of the newly synthesized derivatives of (5 -GMP) 2− suggest that the resulting complexes are crystallized with 1 : 1 stoichiometry in case of diorganotin(IV) derivatives of (5 -GMP) 2− , whereas 2 : 1 (Sn: 5 -GMP 2− ) stoichiometry is observed for triorganotin(IV) derivatives of (5 -GMP) 2− . In the entire studied derivatives one molecule of water is also involved.
In the infrared spectra of di-and triorganotin(IV) derivatives of (5 -GMP) 2− , three bands due to the ν(NH 2 ) and ν(OH) are observed in the 3117-3362 cm −1 region as compared to a single broadband observed at 3314 cm −1 in Na 2 (5 -GMP). Further, NH 2 deformation vibration undergoes some shifts (∼±10 cm −1 ) in these organotin(IV) derivatives as compared to Na 2 (5 -GMP) (1638 cm −1 ). These shifts may be due to the different extent of hydrogen bonding in organotin(IV) derivatives in the solid-state. An additional band observed beyond 3400 cm   associated with the C3 -endo and at ∼>820 cm −1 associated with the C2 -endo, the two most commonly found ribose puckers in nucleotides and nucleic acids. Ph 2 Sn(IV) and iso-Pr 3 Sn(IV) derivatives have ribose pucker band at 860 and 822 cm −1 , respectively, whereas all other complexes show this band at 805 ± 5 cm −1 , which indicate the C2 -endo conformation in the former and C3 -endo in the latter complexes.
The symmetric stretching vibration of the phosphate group (PO 3 ) 2− of Na 2 (5 -GMP) gets shifted towards higher wave number except in [Ph 2 Sn(5 -GMP)·H 2 O] n upon complexation, whereas the smaller shifts are also observed for the asymmetric stretching vibrations in all of the studied complexes, which indicate the bonding of the phosphate group with the organotin moiety. The appearance of new bands of medium intensity in the region 452-512 cm −1 in the studied complexes, which may be assigned to ν(Sn-O), further confirms the coordination of the (PO 3 ) 2− , group of (5 -GMP) 2− to tin through covalent bonding [29]. Therefore, coordination of (5 -GMP) 2− through NH 2 and C=O groups of nucleobase is unlikely. The ν(Sn-C 2 ) bands observed at around 594 ± 17 cm −1 and 521 ± 9 cm −1 can be identified as ν as (Sn-C) and ν s (Sn-C), respectively, which is consistent with the cis-disposition of alkyl groups, whereas for the phenyl derivatives, the corresponding ν(Sn-C 2 ) stretching bands are observed in the far-IR region of 222-280 cm −1 [21,22].
The 119 Sn Mössbauer spectral data of the studied compounds are presented in Table 1. The structures of R 2 Sn(IV) and R 3 Sn(IV) derivatives of (5 -GMP) 2− are considerably more complex than those of guanosine [22]. The 119 Sn Mössbauer spectra of di-and trialkyltin(IV) derivatives of (5 -GMP) 2− exhibit a doublet centered (IS) at 1.14 ± 1 and 1.40 mm s −1 , respectively, and quadrupole splitting in the   13 C, and 19 Sn NMR spectral data of di-and triorganotin(IV) derivatives of (5 -GMP) 2 (Figure 1). Therefore, on the basis of the QS values, the geometry adopted by all of the triorganotin(IV) derivatives would be similar to that as shown in Figure 1(b). The slightly low value of QS (2.52 mm s −1 ) for triphenyltin(IV) derivatives is in accordance with the reported the fact that QS and IS values decrease when an alkyl group is replaced by a phenyl group. Therefore, polymeric structures involving a bidentate phosphate group in axial position and three organic groups in equatorial position leading to either 2-or 3-dimensional associated lattice have been proposed for triorganotin(IV) derivatives of (5 -GMP) 2− as shown in Figure 2. A monomeric structure involving a four coordinate R 3 Sn(IV) moiety bonded individually to (PO 3 ) 2− and 3 -O has been ruled out on the basis of the presence of only one tin species in 119 Sn Mössbauer spectra with ρ value greater than four.
A considerable number of possible structures (Figure 3) may be proposed for diorganotin(IV) derivatives of (5 -GMP) 2− , which correspond to a distorted trigonalbipyramidal geometry involving one water molecule with either two axial or axial-equatorial disposition of both organic groups and a bidentate phosphate group (Figure 3(a) and Figure 3(b)), and a distorted cis-octahedral geometry (Figure 3(c)). The structure as shown in Figure 3(c) may be ruled out on the basis of 119 Sn NMR chemical shift (discussed later) corresponding to five-coordinated tin ( Table 2).
The characteristic resonances in the 1 H, 13 C, and 119 Sn NMR spectral data of the studied di-and triorganotin(IV) derivatives of (5 -GMP) 2− , recorded in dimethyl-sulfoxided 6 , are presented in Table 2  Na 2 (5 -GMP) are also included in Table 2 for comparison. In di-and triorganotin(IV) derivatives of (5 -GMP) 2− , all the resonances of (5 -GMP) 2− are observed at the expected position as compared to Na 2 (5 -GMP). The H-5 resonances are considerably shifted indicating the involvement of (PO 3 ) 2− group in bonding with organotin(IV) moiety. The resonances observed due to the tin-alkyl/phenyl protons in the studied organotin(IV) derivatives of (5 -GMP) 2− are observed in the expected regions. The downfield shifts in N(1)-H and NH 2 resonances may be due to the different extent of hydrogen bonding in the studied derivatives. The 13 C NMR spectra of [n-Bu 2 Sn(5 -GMP)·H 2 O] n and [(Me 3 Sn) 2 (5 -GMP)·H 2 O] n could not be recorded because of their extremely low solubility in DMSO-d 6 / CDCl 3 /CD 3 OD. The chemical shifts of various magnetically nonequivalent carbons of (5 -GMP) 2− have been assigned in the studied derivatives. The C-5 resonances in organotin(IV) derivatives of (5 -GMP) 2− are shifted towards downfield upon complexation as compared with that of ligand, which indicate the involvement of phosphate group (PO 3 ) 2− in bonding with tin. While all other carbon shifts remains almost unchanged, the 13 C chemical shifts of alkyl and phenyl groups attached to tin are also observed in the expected regions which are consistent with previously reported values [13,29]. The characteristic resonances in the 119 Sn NMR spectra of some of the studied derivatives, recorded in dimethylsulfoxide-d 6 , are also presented in Table 2. The satisfactory 119 Sn NMR spectra of 1, 2, 4, and 6 could not be recorded due to their poor solubility even in DMSO-d 6 . The 119 Sn chemical shifts in iso-Pr 3 Sn(IV), Ph 2 Sn(IV), and Ph 3 Sn(IV) derivatives of (5 -GMP) 2− are observed at δ −256, −225, and −226 ppm, which are characteristic of the five-coordinated organotin(IV) derivatives [13,17,21,22,32].
The anti-inflammatory activity (% inhibition) and toxicity data of di-and triorganotin(IV) derivatives of (5 -GMP) 2− are presented in Table 3. The activity of the studied derivatives is influenced by the nature of the ligand and the organic groups attached to tin. Organotin(IV) derivatives of (5 -GMP) 2− show better activity as compared to those of guanosine (∼7.51-9.21% inhibition at 40 mg kg −1 dose) [22], whereas di-and triorganotin(IV) derivatives of (5 -GMP) 2− displayed mild-to-moderate anti-inflammatory activity (∼15.64-20.63% inhibition at 40 mg kg −1 dose) which is significantly lower than that of phenylbutazone (34.56% inhibition). It has been observed that the activity decreases with the increases in size of the alkyl group, that is, Me 2 Sn(IV) derivative is better than n-Bu 2 Sn(IV), and iso-Pr 3 Sn(IV) derivative is better than n-Bu 3 Sn(IV) derivative. Further, phenyltin(IV) derivatives show better activity as compared to their alkyl analogues. Furthermore, triorganotin(IV) derivatives of guanosine and (5 -GMP) 2− show slightly higher activity than the corresponding diorganotin(IV) derivatives. [(Ph 3 Sn) 2 (5 -GMP)·H 2 O] n exhibited the highest anti-inflammatory activity (20.63% inhibition) among the studied derivatives. The higher activity of diphenyltin(IV) and triphenyltin(IV) derivatives of (5 -GMP) 2− among the studied derivatives may be due to the formation and frequent transportation of Ph 2 Sn(IV) 2+ /Ph 3 Sn(IV) + moiety across the cellular membrane as part of the mechanism for inhibition.
The observed LD 50 values (Table 3) indicate that di-and triorganotin(IV) derivatives of (5 -GMP) 2− are less toxic (LD 50 > 400 mg kg −1 ) than the corresponding derivatives of guanosine (LD 50 > 200 mg kg −1 ) [22]. Further, it has been observed that the LD 50 values of the studied derivatives are comparable (>400 mg kg −1 ) with those of other compounds reported earlier [33] and much higher than those of the diorganotin(IV) derivatives of the simple α-amino acids 8 ISRN Organic Chemistry (<50 mg/kg) [34], indicating that the bigger biomolecules lower the toxicities.