Anticancer property of Bryophyllum pinnata (Lam.) Oken. leaf on human cervical cancer cells.

BACKGROUND: Bryophyllum pinnata (B. pinnata) is a common medicinal plant used in traditional medicine of India and of other countries for curing various infections, bowel diseases, healing wounds and other ailments. However, its anticancer properties are poorly defined. In view of broad spectrum therapeutic potential of B. pinnata we designed a study to examine anti-cancer and anti-Human Papillomavirus (HPV) activities in its leaf extracts and tried to isolate its active principle. METHODS: A chloroform extract derived from a bulk of botanically well-characterized pulverized B. pinnata leaves was separated using column chromatography with step- gradient of petroleum ether and ethyl acetate. Fractions were characterized for phyto-chemical compounds by TLC, HPTLC and NMR and Biological activity of the fractions were examined by MTT-based cell viability assay, Electrophoretic Mobility Shift Assay, Northern blotting and assay of apoptosis related proteins by immunoblotting in human cervical cancer cells. RESULTS: Results showed presence of growth inhibitory activity in the crude leaf extracts with IC50 at 552 µg/ml which resolved to fraction F4 (Petroleum Ether: Ethyl Acetate:: 50:50) and showed IC50 at 91 µg/ml. Investigations of anti-viral activity of the extract and its fraction revealed a specific anti-HPV activity on cervical cancer cells as evidenced by downregulation of constitutively active AP1 specific DNA binding activity and suppression of oncogenic c-Fos and c-Jun expression which was accompanied by inhibition of HPV18 transcription. In addition to inhibiting growth, fraction F4 strongly induced apoptosis as evidenced by an increased expression of the pro-apoptotic protein Bax, suppression of the anti-apoptotic molecules Bcl-2, and activation of caspase-3 and cleavage of PARP-1. Phytochemical analysis of fraction F4 by HPTLC and NMR indicated presence of activity that resembled Bryophyllin A. CONCLUSIONS: Our study therefore demonstrates presence of anticancer and anti-HPV an activity in B. pinnata leaves that can be further exploited as a potential anticancer, anti-HPV therapeutic for treatment of HPV infection and cervical cancer.

Antibiotic resistance: mono- and dinuclear zinc complexes as metallo-beta-lactamase mimics.

Biomimetic systems containing one or two zinc(II) ions supported by phenolate ligands were developed as functional mimics of metallo-beta-lactamase. These complexes were shown to catalytically hydrolyze beta-lactam substrates, such as oxacillin and penicillin G. The dinuclear zinc complex 1, which has a coordinated water molecule, exhibits high beta-lactamase activity, whereas the dinuclear zinc complex 2, which has no water molecules, but labile chloride ligands, shows a much lower activity. The high beta-lactamase activity of complex 1 can be ascribed to the presence of a zinc-bound water molecule that is activated by being hydrogen bonded to acetate substituents. The kinetics of the hydrolysis of oxacillin by complex 1 and the effect of pH on the reaction rates are reported in detail. In addition, the kinetic parameters obtained for the synthetic analogues are compared with those of the natural metallo-beta-lactamase from Bacillus cereus (BcII). To understand the role of the second metal ion in hydrolysis, the syntheses and catalytic activities of two mononuclear complexes (3 and 4) that include coordinated water molecules are described. Interestingly, the mononuclear zinc complexes 3 and 4 also exhibit high activity, supporting the assumption that the second zinc ion is not crucial for the beta-lactamase activity.

Horseradish peroxidase inhibition and antioxidant activity of ebselen and related organoselenium compounds.

Horseradish peroxidase (HRP) inhibition and glutathione peroxidase (GPx) activities of ebselen and some related derivatives are described. These studies show that ebselen and ebselen ditelluride (EbTe(2)) with significant antioxidant activity, inhibit the HRP-catalyzed oxidation reactions. In addition, inhibition of lipid peroxidation and singlet oxygen quenching studies were carried out. Although the inhibition of HRP by ebselen is comparable with that of EbTe(2), the inhibitory effect on gamma-radiation induced lipid peroxidation and the GPx activity of ebselen is found to be much higher than that of EbTe(2).

Interaction of anti-thyroid drugs with iodine: the isolation of two unusual ionic compounds derived from Se-methimazole.

The inhibition of lactoperoxidase (LPO)-catalyzed iodination of l-tyrosine by the anti-thyroid drug methimazole (MMI) and its selenium analogue (MSeI) is described. MSeI inhibits LPO with an IC(50) value of 12.4 microM, and this inhibition could be completely reversed by increasing the peroxide concentration. In addition to the inhibition, MSeI reacts with molecular iodine to produce novel ionic diselenides, and the nature of the species formed in this reaction appear to be solvent-dependent. The formation of ionic species in the reaction is confirmed by single-crystal X-ray studies, FT-IR and FT-Raman spectroscopic investigations. This study provides the first experimental evidence that MSeI not only effectively inhibits the LPO-catalyzed iodination of tyrosine, but also reacts with I(2) to produce novel ionic diselenides. These results also suggest that MSeI reacts with iodine, even in its oxidized form, to form ionic diselenides containing iodide or polyiodide anions, which might be effective intermediates in the inhibition of thyroid hormones.

Biomimetic studies on selenoenzymes: modeling the role of proximal histidines in thioredoxin reductases.

The roles of built-in thiol cofactors and the basic histidine (His) residues in the active site of mammalian thioredoxin reductases (TrxRs) are described with the help of experimental and density functional theory calculations on small-molecule model compounds. The reduction of selenenyl sulfides by thiols in selenoenzymes such as glutathione peroxidase (GPx) and TrxR is crucial for the regeneration of the active site. Experimental as well as theoretical studies were carried out with model selenenyl sulfides to probe their reactivity toward incoming thiols. We have shown that the nucleophilic attack of thiols takes place at the selenium center in the selenenyl sulfides. These thiol exchange reactions would hamper the regeneration of the active species selenol. Therefore, the basic His residues are expected to play crucial roles in the selenenyl sulfide state of TrxR. Our model study with internal amino groups in the selenenyl sulfide state reveals that the basic His residues may play important roles by deprotonating the thiol moiety in the selenenic acid state and by interacting with the sulfur atom in the selenenyl sulfide state to facilitate the nucleophilic attack of thiol at sulfur rather than at selenium, thereby generating the catalytically active species selenol. This model study also suggests that the enzyme may use the internal cysteines as cofactors to overcome the thiol exchange reactions.

Bioinorganic chemistry in thyroid gland: effect of antithyroid drugs on peroxidase-catalyzed oxidation and iodination reactions.

Propylthiouracil (PTU) and methimazole (MMI) are the most commonly used antithyroid drugs. The available data suggest that these drugs may block the thyroid hormone synthesis by inhibiting the thyroid peroxidase (TPO) or diverting oxidized iodides away from thyroglobulin. It is also known that PTU inhibits the selenocysteine-containing enzyme ID-1 by reacting with the selenenyl iodide intermediate (E-SeI). In view of the current interest in antithyroid drugs, we have recently carried out biomimetic studies to understand the mechanism by which the antithyroid drugs inhibit the thyroid hormone synthesis and found that the replacement of sulfur with selenium in MMI leads to an interesting compound that may reversibly block the thyroid hormone synthesis. Our recent results on the inhibition of lactoperoxidase (LPO)-catalyzed oxidation and iodination reactions by antithyroid drugs are described.

Anti-thyroid drugs and thyroid hormone synthesis: effect of methimazole derivatives on peroxidase-catalyzed reactions.

Syntheses and characterization of the selenium analogue (MSeI) of anti-thyroid drug methimazole and a series of organoselenium compounds bearing N-methylimidazole pharmacophore are described. In contrast to the sulfur compound that exists predominantly in its thione form, the selenium analogue exists in a selenol form, which spontaneously oxidizes in air to produce the corresponding diselenide. The reduction of the diselenide by GSH or NaBH(4) affords the biologically active selenol, which effectively inhibits the lactoperoxidase (LPO) activity in vitro. The monoselenides having N-methylimidazole moiety are found to be much less active than the selenol, suggesting that the presence of a selenol moiety is important for the LPO inhibition. The kinetic and mechanistic studies reveal that MSeI inhibits the LPO activity by reducing the H(2)O(2), providing a novel method to reversibly inhibit the enzyme. Although MSeI strongly inhibits LPO, the enzyme's activity can be completely recovered by increasing the H(2)O(2) concentration. On the other hand, the inhibition by methimazole (MMI), the sulfur analogue, cannot be reversed by increasing the H(2)O(2) concentration, leading to a complete inactivation of the enzyme. The reversible inhibition of LPO by some of the selenium derivatives is correlated with their glutathione peroxidase (GPx) activity, and the high GPx activity of the selenium compounds as compared with their sulfur analogues suggests that the selenium derivatives may protect the thyroid gland from oxidative damage.

Glutathione peroxidase (GPx)-like antioxidant activity of the organoselenium drug ebselen: unexpected complications with thiol exchange reactions.

The factors that are responsible for the relatively low glutathione peroxidase (GPx)-like antioxidant activity of organoselenium compounds such as ebselen (1, 2-phenyl-1,2-benzisoselenazol-3(2H)-one) in the reduction of hydroperoxides with aromatic thiols such as benzenethiol and 4-methylbenzenethiol as cosubstrates are described. Experimental and theoretical investigations reveal that the relatively poor GPx-like catalytic activity of organoselenium compounds is due to the undesired thiol exchange reactions that take place at the selenium center in the selenenyl sulfide intermediate. This study suggests that any substituent that is capable of enhancing the nucleophilic attack of thiol at sulfur in the selenenyl sulfide state would enhance the antioxidant potency of organoselenium compounds such as ebselen. It is proved that the use of thiol having an intramolecularly coordinating group would enhance the biological activity of ebselen and other organoselenium compounds. The presence of strong S...N or S...O interactions in the selenenyl sulfide state can modulate the attack of an incoming nucleophile (thiol) at the sulfur atom of the -Se-S- bridge and enhance the GPx activity by reducing the barrier for the formation of the active species selenol.

Internally stabilized selenocysteine derivatives: syntheses, 77Se NMR and biomimetic studies.

Selenocystine ([Sec]2) and aryl-substituted selenocysteine (Sec) derivatives are synthesized, starting from commercially available amino acid l-serine. These compounds are characterized by a number of analytical techniques such as NMR (1H, 13C and 77Se) and TOF mass spectroscopy. This study reveals that the introduction of amino/imino substituents capable of interacting with selenium may stabilize the Sec derivatives. This study further suggests that the oxidation-elimination reactions in Sec derivatives could be used for the generation of biologically active selenols having internally stabilizing substituents.

Biomimetic studies on anti-thyroid drugs and thyroid hormone synthesis.

The selenium analogues of anti-thyroid drugs exhibit their anti-thyroid action by a mechanism different from that of MMI. The selenium analogue of MMI and related selenium compounds exhibit high GPx activity, providing a novel method for the reversible inhibition of thyroid hormone biosynthesis.

Glutathione peroxidase-like antioxidant activity of diaryl diselenides: a mechanistic study.

The synthesis, structure, and thiol peroxidase-like antioxidant activities of several diaryl diselenides having intramolecularly coordinating amino groups are described. The diselenides derived from enantiomerically pure R-(+)- and S-(-)-N,N-dimethyl(1-ferrocenylethyl)amine show excellent peroxidase activity. To investigate the mechanistic role of various organoselenium intermediates, a detailed in situ characterization of the intermediates has been carried out by (77)Se NMR spectroscopy. While most of the diselenides exert their peroxidase activity via selenol, selenenic acid, and selenenyl sulfide intermediates, the differences in the relative activities of the diselenides are due to the varying degree of intramolecular Se.N interaction. The diselenides having strong Se.N interactions are found to be inactive due to the ability of their selenenyl sulfide derivatives to enhance the reverse GPx cycle (RSeSR + H(2)O(2) = RSeOH). In these cases, the nucleophilic attack of thiol takes place preferentially at selenium rather than sulfur and this reduces the formation of selenol by terminating the forward reaction. On the other hand, the diselenides having weak Se.N interactions are found to be more active due to the fast reaction of the selenenyl sulfide derivatives with thiol to produce diphenyl disulfide and the expected selenol (RSeSR + PhSH = PhSSPh + RSeH). The unsubstituted diaryl diselenides are found to be less active due to the slow reactions of these diselenides with thiol and hydrogen peroxide and also due to the instability of the intermediates. The catalytic cycles of 18 and 19 strongly resemble the mechanism by which the natural enzyme, glutathione peroxidase, catalyzes the reduction of hydroperoxides.

Structure-activity correlation between natural glutathione peroxidase (GPx) and mimics: a biomimetic concept for the design and synthesis of more efficient GPx mimics.

Among the organoselenium compounds that mimic the action of the natural enzyme glutathione peroxidase (GPx), there are certain basic differences in the activity, substrate specificity and mechanism. These differences arise mainly from the nature of the substituents near the reaction center, and stability and reactivity of the intermediates. As an attempt to draw some general concepts for the development of new mimics, a structure - activity correlation between natural GPx and some existing mimics is described.

Synthesis and Structural Characterization of Monomeric Selenolato Complexes of Zinc, Cadmium, and Mercury.

The synthesis and characterization of homoleptic zinc(II), cadmium(II), and mercury(II) selenolates incorporating the intramolecularly chelating oxazoline ligand are described. The derivatives, M[Se(Ox)](2) [M = Zn (1), Cd (2), Hg (3); Ox = 2-(4,4-dimethyl-2-oxazolinyl)benzene], were prepared in good yield via the metathesis reactions of MCl(2) with a lithium areneselenolate, OxSe(-)Li(+). The mercury complex 3 was also synthesized by treating the corresponding diselenide with elemental mercury. The complexes are quite stable and highly soluble in common nonpolar organic solvents. X-ray diffraction results show that the complexes are monomeric in the solid state. The geometry around the metal ion in all complexes is found to be distorted tetrahedral. The crystal structure of Zn[Se(Ox)](2) (1) shows that the complex is "helically" chiral and enantiomerically pure. The spontaneous splitting of the racemates indicates the solubility differences between the racemates and pure enantiomers. (1)H, (13)C, and (77)Se NMR measurements indicate that complex 1 retains its "helical" structure in solution. Crystal data (Mo Kalpha; 293(2) K) are as follows: 1, monoclinic space group P2(1), a = 9.3900(12) Å, b = 11.618(2) Å, c = 10.8822(14) Å, beta = 98.245(8) degrees, Z = 2; 2, orthorhombic space group Pbca, a = 12.777(6) Å, b = 17.841(10) Å, c = 21.010(8) Å, Z = 8; 3, monoclinic space group P2(1)/c, a = 9.087(2) Å, b = 11.889(2) Å, c = 22.456(4) Å, beta = 98.780(13) degrees, Z = 4.