Attenuation of Antioxidant Capacity in Human Breast Cancer Cells by Carbon Monoxide through Inhibition of Cystathionine ß-Synthase Activity: Implications in Chemotherapeutic Drug Sensitivity.

Drug resistance is a major impediment to effective treatment of breast cancer. Compared to normal cells, cancer cells have an increased antioxidant potential due to an increased ratio of reduced to oxidized glutathione (GSH/GSSG). This is known to confer therapeutic resistance. Here, we have identified a mechanism, unique to breast cancer cells, whereby cystathionine ß-synthase (CBS) promotes elevated GSH/GSSG. Lentiviral silencing of CBS in human breast cancer cells attenuated GSH/GSSG, total GSH, nuclear factor erythroid 2-related factor 2 (Nrf2), and processes downstream of Nrf2 that promote GSH synthesis and regeneration of GSH from GSSG. Carbon monoxide (CO) reduced GSH/GSSG in three breast cancer cell lines by inhibiting CBS. Furthermore, CO sensitized breast cancer cells to doxorubicin. These results provide insight into mechanism(s) by which CBS increases the antioxidant potential and the ability for CO to inhibit CBS activity to alter redox homeostasis in breast cancer, increasing sensitivity to a chemotherapeutic.

Synthesis and structures of photoactive manganese-carbonyl complexes derived from 2-(pyridin-2-yl)-1,3-benzothiazole and 2-(quinolin-2-yl)-1,3-benzothiazole.

PhotoCORMs (photo-active CO-releasing molecules) have emerged as a class of CO donors where the CO release process can be triggered upon illumination with light of appropriate wavelength. We have recently reported an Mn-based photoCORM, namely [MnBr(pbt)(CO)3] [pbt is 2-(pyridin-2-yl)-1,3-benzothiazole], where the CO release event can be tracked within cellular milieu by virtue of the emergence of strong blue fluorescence. In pursuit of developing more such trackable photoCORMs, we report herein the syntheses and structural characterization of two MnI-carbonyl complexes, namely fac-tricarbonylchlorido[2-(pyridin-2-yl)-1,3-benzothiazole-κ2N,N']manganese(I), [MnCl(C12H8N2S)(CO)3], (1), and fac-tricarbonylchlorido[2-(quinolin-2-yl)-1,3-benzothiazole-κ2N,N']manganese(I), [MnCl(C16H10N2S)(CO)3], (2). In both complexes, the MnI center resides in a distorted octahedral coordination environment. Weak intermolecular C-H...Cl contacts in complex (1) and Cl...S contacts in complex (2) consolidate their extended structures. These complexes also exhibit CO release upon exposure to low-power broadband visible light. The apparent CO release rates for the two complexes have been measured to compare their CO donating capacity. The fluorogenic 2-(pyridin-2-yl)-1,3-benzothiazole and 2-(quinolin-2-yl)-1,3-benzothiazole ligands provide a convenient way to track the CO release event through the `turn-ON' fluorescence which results upon de-ligation of the ligands from their respective metal centers following CO photorelease.

Synthesis, Structures, and CO Release Capacity of a Family of Water-Soluble PhotoCORMs: Assessment of the Biocompatibility and Their Phototoxicity toward Human Breast Cancer Cells.

Two manganese(I) carbonyl complexes derived from 2-(pyridyl)benzothiazole (pbt) and 1,10-phenanthroline (phen) release carbon monoxide (CO) under low-power broad-band visible-light illumination. CO photorelease from [Mn(CO)3(pbt)(PTA)]CF3SO3 (1, where PTA = 1,3,5-triaza-7-phosphaadamantane) is accompanied by an emergence of a strong fluorescence around 400 nm from almost nonfluorescent preirradiated 1. However, [Mn(CO)3(phen)(PTA)]CF3SO3 (2) showed no such phenomenon upon prolonged illumination under similar experimental conditions. The two analogous rhenium(I) complexes, namely, [Re(CO)3(pbt)(PTA)]CF3SO3 (3) and [Re(CO)3(phen)(PTA)]CF3SO3 (4), have also been synthesized and characterized to compare their photo properties with the manganese congeners. Complexes 3 and 4 exhibit moderate CO release upon irradiation with low-power UV light. All four complexes are highly soluble in anaerobic/aerobic aqueous media and are also considerably more stable when kept under dark conditions. The inherently luminescent rhenium complex 3 was utilized to demonstrate cellular internalization of these types of compounds by MDA-MB-231 (human breast cancer) cells, while the two biocompatible manganese(I) complexes (1 and 2) have been applied to assess the cell viability of these malignant cells upon CO delivery.

Light-triggered CO delivery by a water-soluble and biocompatible manganese photoCORM.

The discovery of salutary effects of low doses of carbon monoxide (CO) has spurred interest in designing exogenous molecules that can deliver CO to biological targets under controlled conditions. Herein we report a water-soluble photosensitive manganese carbonyl complex [MnBr(CO)3(pyTAm)] (2) (pyTAm = 2-(pyridyl)imino-triazaadamantane) that can be triggered to release CO upon exposure to visible light. Inclusion of a triazaadamantyl pharmacophore into the coligand of 2 improves its stability and solubility in water. Change in the coligand from 2-(pyridyl)imino-triazaadamantane to 2-(pyridyl)iminoadamantane (pyAm) or 2-(quinonyl)imino-triazaadamantane (qyTAm) dramatically alters these desired properties of the photoCORM. In addition to structures and CO-releasing properties of the three analogous complexes 1-3 from these three α-diimine ligands, theoretical calculations have been performed to determine the origin of Mn-CO bond labilization upon illumination. Rapid delivery of CO to myoglobin under physiological conditions attests the potential of 2 as a biocompatible photoCORM.

A Theranostic Two-Tone Luminescent PhotoCORM Derived from Re(I) and (2-Pyridyl)-benzothiazole: Trackable CO Delivery to Malignant Cells.

A Re(I) carbonyl complex derived from 2-(2-pyridyl)-benzothiazole (pbt), [Re(H2O)(CO)3(pbt)](CF3SO3) (1), rapidly releases CO under low-power UV illumination. CO photorelease from 1 is accompanied by a change in luminescence from orange to deep blue. These two distinct luminescence signals have been successfully employed to track (a) the entry of the pro-drug 1 into cancer cells and (b) the end of the CO (drug) delivery step within the target.

Exceptionally rapid CO release from a manganese(I) tricarbonyl complex derived from bis(4-chloro-phenylimino)acenaphthene upon exposure to visible light.

Two manganese(i) carbonyl complexes derived from α,α'-diimine ligands with extended conjugated framework namely [MnBr(CO)3(BIAN)] () and [MnBr(CO)3(MIAN)] (), have been synthesized and structurally characterized. Unlike the previously reported photoactive CO-releasing molecules (photoCORMs), these two complexes exhibit unusually high sensitivity toward low power (0.3-10 mW) visible light (λ ≥ 520 nm) even in the solid state and rapidly release carbon monoxide (CO) upon illumination. The role of the ligand frames in such activity has been examined with the help of theoretical calculations. Application of these photoCORMs in delivering high fluxes of CO to biological targets is anticipated.

Synthesis and Characterization of a "Turn-On" photoCORM for Trackable CO Delivery to Biological Targets.

A designed photoactive CO releasing molecule (photoCORM), namely, fac-[MnBr(CO)3(pbt)] (1, pbt = 2-(2-pyridyl)benzothiazole), promotes CO-induced death of MDA-MB-231 human breast cancer cells upon illumination with broadband visible light. The CO release from this photoCORM can be tracked by rise in fluorescence within the cellular matrix due to deligation of the pbt ligand. The results of this study suggest the potential of 1 in eradication of cancer cells through CO delivery.

Design strategies to improve the sensitivity of photoactive metal carbonyl complexes (photoCORMs) to visible light and their potential as CO-donors to biological targets.

The recent surprising discovery of the beneficial effects of carbon monoxide (CO) in mammalian physiology has drawn attention toward site-specific delivery of CO to biological targets. To avoid difficulties in handling of this noxious gas in hospital settings, researchers have focused their attention on metal carbonyl complexes as CO-releasing molecules (CORMs). Because further control of such CO delivery through light-triggering can be achieved with photoactive metal carbonyl complexes (photoCORMs), we and other groups have attempted to isolate such complexes in the past few years. Typical metal carbonyl complexes release CO when exposed to UV light, a fact that often deters their use in biological systems. From the very beginning, our effort therefore was directed toward identifying design principles that could lead to photoCORMs that release CO upon illumination with low-power (5-15 mW/cm(2)) visible and near-IR light. In our work, we have utilized Mn(I), Re(I), and Ru(II) centers (all d(6) ground state configuration) to ensure overall stability of the carbonyl complexes. We also hypothesized that transfer of electron density from the electron-rich metal centers to π* MOs of the ligand frame via strong metal-to-ligand charge transfer (MLCT) transitions in the visible/near-IR region would weaken metal-CO back-bonding and promote rapid CO photorelease. This expectation has been realized in a series of carbonyl complexes derived from a variety of designed ligands and smart choice of ligand/coligand combinations. Several principles have emerged from our systematic approach to the design of principal ligands and the choice of auxiliary ligands (in addition to the number of CO) in synthesizing these photoCORMs. In each case, density functional theory (DFT) and time-dependent DFT (TDDFT) study afforded insight into the dependence of the CO photorelease from a particular photoCORM on the wavelength of light. Results of these theoretical studies indicate that extended conjugation in the principal ligand frames as well as the nature of the donor groups lower the energy of the lowest unoccupied MOs (LUMOs) while auxiliary ligands like PPh3 and Br(-) modulate the energy of the occupied orbitals depending on their strong σ- or π-donating abilities. As a consequence, the ligand/coligand combination dictates the energy of the MLCT bands of the resulting carbonyl complexes. The rate of CO photorelease can be altered further by proper disposition of the coligands in the coordination sphere to initiate trans-effect or alter the extent of π back-bonding in the metal-CO bonds. Addition of more CO ligands blue shift the MLCT bands, while intersystem crossing impedes labilization of metal-CO bonds in several Re(I) and Ru(II) carbonyl complexes. We anticipate that our design principles will provide help in the future design of photoCORMs that could eventually find use in clinical studies.

Photodelivery of CO by designed PhotoCORMs: correlation between absorption in the visible region and metal-CO bond labilization in carbonyl complexes.

The therapeutic potential of photoactive CO-releasing molecules (photoCORMs) have called for close examination of the roles of the ligand(s) and the central metal atoms on the overall photochemical labilization of the metal-CO bonds. Along this line, we have synthesized four metal complexes, namely, [MnBr(azpy)(CO)3 ] (1), [Mn(azpy)(CO)3 (PPh3 )]ClO4 (2), [ReBr(azpy)(CO)3 ] (3), and [Re(azpy)(CO)3 (PPh3 )]ClO4 (4), derived from 2-phenylazopyridine. These complexes were characterized by spectroscopic and crystallographic studies. Although both 1 and 3 exhibit strong metal-to-ligand charge-transfer bands in the 500-600 nm region, only 1 photoreleases CO upon illumination with visible light. Results of theoretical studies were used to gain insight into this surprising difference. Strong spin-orbit coupling (prominent in heavy metals) appears to promote intersystem crossing to a triplet state in 3, a step that discourages CO release upon illumination with visible light. Slow release of CO from 2 and 4 also indicates that strong σ-donating ligands, such as Br(-) , accelerate the rate of CO photorelease relative to π-acid ligands, such as PPh3 .

Rapid CO release from a Mn(I) carbonyl complex derived from azopyridine upon exposure to visible light and its phototoxicity toward malignant cells.

Two Mn(I) carbonyl complexes namely [MnBr(azpy)(CO)3] and [Mn(azpy)(CO)3(PPh3)](ClO4) (azpy = 2-phenylazopyridine) have been synthesized and characterized. Both complexes exhibit rapid CO release upon exposure to low power visible light. [MnBr(azpy)(CO)3] shows significant phototoxicity toward two malignant cell lines HeLa and MDA-MB-231.

Photoactivity of mono- and dicarbonyl complexes of ruthenium(II) bearing an N,N,S-donor ligand: role of ancillary ligands on the capacity of CO photorelease.

One monocarbonyl and one dicarbonyl complex of ruthenium(II), namely, [Ru(Cl)(CO)(qmtpm)(PPh3)]BF4 (2) and [Ru(Cl)(CO)2(qmtpm)]ClO4 (3), derived from the tridentate ligand 2-quinoline-N-(2'-methylthiophenyl)methyleneimine (qmtpm) have been synthesized and structurally characterized. The qmtpm ligand binds in a meridional fashion in these carbonyl complexes, and in 3, the two carbon monoxide (CO) ligands are cis to each other. Solutions of 2 in ethanol, chloroform, or acetonitrile rapidly release CO upon illumination with low-power (3-15 mW) light in the 300-450 nm range. Loss of CO from 2 brings about a dramatic color change from yellow to magenta because of the formation of [Ru(Cl)(MeCN)(qmtpm)(PPh3)]BF4 (4). In acetonitrile, photorelease of CO from 3 under 360 nm light occurs in two steps, and the violet photoproduct [Ru(Cl)(MeCN)2(qmtpm)](+) upon reaction with Ag(+) and PPh3 affords red [Ru(MeCN)2(qmtpm)(PPh3)](ClO4)2 (5). The structure of 5 has also been determined by X-ray crystallography. Reduced myoglobin assay confirms that 2 and 3 act as photoactive CO-releasing molecules (photoCORMs) that deliver 1 and 2 equiv of CO, respectively. The results of density functional theory (DFT) and time-dependent DFT studies confirm that electronic transitions from molecular orbitals with predominantly Ru-CO character to ligand-based π* orbitals facilitate CO release from these two photoCORMs. Complexes 2-5 have provided an additional opportunity to analyze the roles of the ancillary ligands, namely, PPh3, Cl(-), and MeCN, in shifting the positions of the metal-to-ligand charge-transfer bands and the associated sensitivity of the two photoCORMs to different wavelengths of light. Collectively, the results provide helpful hints toward the future design of photoCORMs that release CO upon exposure to visible light.

Syntheses, structures, and properties of new manganese carbonyls as photoactive CO-releasing molecules: design strategies that lead to CO photolability in the visible region.

The unusual role of CO as a signaling molecule in several physiological pathways has spurred research in the area of synthesizing new CO-releasing molecules (CORMs) as exogenous CO donors. Auxiliary control on CO delivery can be achieved if CO can be released under the control of light. To synthesize such photoactive CORMs (photoCORMs) with the aid of smart design, a series of manganese carbonyls have been synthesized with ligands that contain extended conjugation and electron-rich donors on their frames. Five such photoCORMs, namely, [Mn(pimq)(CO)(3)(MeCN)]ClO(4) (1, where pimq = (2-phenyliminomethyl)quinoline), [Mn(qmtpm)(CO)(3)(MeCN)]ClO(4) (2, where qmtpm = 2-quinoline-N-(2'-methylthiophenyl) methyleneimine), [Mn(qmtpm)(CO)(3)Br] (3) [Mn(pmtpm)(CO)(3)(MeCN)]ClO(4) (4, where pmtpm = 2-pyridyl-N-(2'-methylthiophenyl)methyleneimine), and [Mn(pmtpm)(CO)(3)Br] (5), have been synthesized and structurally characterized. These designed carbonyls readily release CO upon exposure to light (400-550 nm). The apparent CO release rates and quantum yield values at 509 nm (φ(509)) of the photoCORMs increase steadily with rise in conjugation in the ligand frame and inclusion of a -SMe group. Addition of Br(-) as an ancillary ligand also improves the CO-donating capacity. Results of density functional theory (DFT) and time dependent DFT (TDDFT) studies indicate that low energy metal-to-ligand charge transfer (MLCT) transitions from Mn-CO bonding into ligand-π orbitals lead to reduction of M-CO(π*) back-bonding and loss of CO from these photoCORMs. Inclusion of -SMe and Br(-) in the coordination sphere attenuates the energies of the HOMO and LUMO levels and causes further enhancement of CO photorelease. Collectively, the results of this work demonstrate that new photoCORMs with excellent sensitivity to visible light can be synthesized on the basis of smart design principles.

Small molecule signaling agents: the integrated chemistry and biochemistry of nitrogen oxides, oxides of carbon, dioxygen, hydrogen sulfide, and their derived species.

Several small molecule species formally known primarily as toxic gases have, over the past 20 years, been shown to be endogenously generated signaling molecules. The biological signaling associated with the small molecules NO, CO, H2S (and the nonendogenously generated O2), and their derived species have become a topic of extreme interest. It has become increasingly clear that these small molecule signaling agents form an integrated signaling web that affects/regulates numerous physiological processes. The chemical interactions between these species and each other or biological targets is an important factor in their roles as signaling agents. Thus, a fundamental understanding of the chemistry of these molecules is essential to understanding their biological/physiological utility. This review focuses on this chemistry and attempts to establish the chemical basis for their signaling functions.

The reaction of H(2)S with oxidized thiols: generation of persulfides and implications to H(2)S biology.

Hydrogen sulfide is an endogenously generated molecule with many reported physiological functions. Although several biological targets have been proposed, the biochemical mechanisms by which it elicits activity are not established. Thus, in an effort to begin to delineate the fundamental biological chemistry of H(2)S, we have examined the reaction of H(2)S with oxidized thiols and thiol proteins in order to determine whether persulfide formation occurs, is stable and how this may affect protein function. We have found that persulfides are easily generated, relatively stable and can alter enzyme activity. Moreover, we have begun to develop methodology for in situ generation of persulfides to facilitate further study of this potentially important species.

HNO signaling mechanisms.

Due to recent discoveries of important and novel biological activity, nitroxyl (HNO) has become a molecule of significant interest. Although it has been used in the past as a treatment for alcoholism, it is currently being touted as a treatment for heart failure. It is becoming increasingly clear that many of the biological actions of HNO can be attributed to its ability to react with specific thiol- and, possibly, heme-proteins. Herein is discussed the chemistry of HNO with likely biological targets. A particular focus is given to targets associated with the pharmacological utility of HNO as a cardiovascular agent and for the treatment of alcoholism.