Therapeutic applications of the gaseous mediators carbon monoxide and hydrogen sulfide

Background: Hydrogen sulfide (H₂S) and carbon monoxide (CO) are endogenously produced gaseous autacoids that regulate a number of physiological processes, including the inflammatory response, cell death and proliferation, neural transmission and smooth muscle tone. Objective/methods: The current review aims to provide a comprehensive overview of all recent patent applications that address the potential therapeutic applications of CO and H₂S. Results/conclusion: Beyond the direct administration of CO and H₂S, this review highlights the therapeutic applications of a variety of gas-releasing molecules that are being developed to deliver CO and H₂S to diseased tissues at therapeutic doses. The term autacoid, which, in addition to its pharmacological use to describe a locally-acting hormone, literally translates from Greek as ‘self-drug’, seems to particularly well describe the current approach to capture the potential therapeutic use of these two gasotransmitters. In summary, we can conclude that there is a markedly growing interest in harnessing the tissue-protective actions of CO and H₂S.     

Identification of novel nonsteroidal compounds as substrates or inhibitors of hASBT

A prodrug approach that employs the human apical sodium dependent bile acid transporter (hASBT) for absorption requires a recognition moiety for hASBT. Bile acids are natural ligands for hASBT, but are hormones with high molecular weight, such that a recognition moiety that is not a bile acid may be advantageous. The objective was to identify nonsteroidal small molecules that could potentially serve as promoieties in the design of prodrugs that target hASBT. Three searches for bile acid analogues were conducted and it involved molecular fingerprints as the computational tools for similarity searching, as well as traditional medicinal chemistry pattern recognition. Sixty-three compounds were tested using a hASBT-Madin-Darby canine kidney cell monolayer model. Twenty-three of these compounds were found to be hASBT inhibitors and represent novel hASBT inhibitors. Three were selected for hASBT uptake studies. Two were substrates, which represent the first reported nonsteroidal substrates of hASBT. Interestingly, each compound lacked a negative charge. These compounds promise to serve as leads to identify hASBT recognition moieties in a prodrug approach to target hASBT to increase drug absorption.     

Screening of organosulfur compounds as inhibitors of human CYP2A6.

The capacities to inhibit coumarin 7-hydroxylase activity of human cytochrome P450 2A6 (CYP2A6) by organosulfur compounds were evaluated. Five dialkyl sulfides and five dialkyl disulfides, with alkyl chains from methyl to amyl, were examined. In addition to these chemicals, diallyl sulfide, diallyl disulfide, allyl methyl sulfide, allyl n-propyl sulfide, allyl phenyl sulfide, diphenyl sulfide, diphenyl disulfide, difurfuryl disulfide, phenyl cyclopropyl sulfide, 2,2'-dipyridyl disulfide, 4,4'-dipyridyl sulfide, and 4,4'-dipyridyl disulfide were also examined for their capacity to inhibit CYP2A6. The membrane fraction of genetically engineered Escherichia coli cells expressing CYP2A6 together with NADPH-cytochrome P450 reductase was used as an enzyme source. Dialkyl disulfides inhibited CYP2A6 more strongly than did dialkyl sulfides. Among dialkyl disulfides examined, di-n-propyl disulfide, contained in onion oil, was the most potent competitive inhibitor of CYP2A6, with a K(i) value of 1.73 microM. Diallyl disulfide, present in garlic oil, inhibited CYP2A6 activity in a competitive/noncompetitive mixed manner, with the K(i) value of 2.13 microM. Among all of the organosulfur compounds tested, 4,4'-dipyridyl disulfide was the most potent inhibitor of CYP2A6, with a K(i) value of 60 nM, followed by 4,4'-dipyridyl sulfide, with a K(i) value of 72 nM. These chemicals inhibited CYP2A6 in a competitive manner. The preincubation time did not affect the inhibitory effects of di-n-propyl disulfide, diallyl disulfide, 4,4'-dipyridyl disulfide, and 4,4'-dipyridyl sulfide on CYP2A6, indicating that these chemicals were not mechanism-based inhibitors of CYP2A6. 4,4'-Dipyridyl disulfide also inhibited midazolam 1'-hydroxylase activity of CYP3A4. We discovered 4,4'-dipyridyl disulfide to be a potent and relatively selective inhibitor of CYP2A6.     

Synthesis and antioxidative properties of novel thiazolidinedione/imidazolidinedione compounds as retinoids

The general term "retinoids" refers to both naturally occurring as well as synthetic compounds which exhibit biological activity similar to vitamin A (retinol). Vitamin A and its two metabolites, retinaldehyde and retinoic acid, are fat-soluble unsaturated isoprenoids necessary for the growth, differentiation and maintenance of epithelial tissues. In this study, we have synthesized thiazolidinedione/imidazolidinedione compounds as retinoids. Their in vitro effects on rat liver microsomal NADPH-dependent lipid peroxidation (LP) levels and superoxide anion formation were determined.     

The combined toxicity of ethanol and hydrogen sulfide

An 8% solution of ethanol in saline was administered via intraperitoneal injection to male rats, at dose levels of 0.33 and 0.66 g/kg body weight. When exposed to an 800 ppm concentration of hydrogen sulfide, 30 min after injection, the alcohol-treated animals displayed a heightened sensitivity to the gas compared to saline-treated rats.     

Green tea catechins as novel antitumor and antiangiogenic compounds

The concept of cancer prevention by use of naturally occuring substances that could be included in the diet is under investigation as a practical approach towards reducing cancer incidence, and therefore the mortality and morbidity associated with this disease. Tea, which is the most popularly consumed beverage aside from water, has been particularly associated with decreased risk of various proliferative diseases such as cancer and atherosclerosis in humans. Various studies have provided evidence that polyphenols are the strongest biologically active agents in green tea. Green tea polyphenols (GTPs) mainly consist of catechins (3-flavanols), of which (-)-epigallocatechin gallate is the most abundant and the most extensively studied. Recent observations have raised the possibility that green tea catechins, in addition to their antioxidative properties, also affect the molecular mechanisms involved in angiogenesis, extracellular matrix degradation, regulation of cell death and multidrug resistance. This article will review the effects and the biological activities of green tea catechins in relation to these mechanisms, each of which plays a crucial role in the development of cancer in humans. The extraction of polyphenols from green tea, as well as their bioavailability, are also discussed since these two important parameters affect blood and tissue levels of the GTPs and consequently their biological activities. In addition, general perspectives on the application of dietary GTPs as novel antiangiogenic and antitumor compounds are also presented.     

Garlic flavonoids and organosulfur compounds: impact on the hepatic pharmacokinetics of saquinavir and darunavir

The therapeutic efficacy of saquinavir and darunavir is affected by the presence of xenobiotics (such as garlic compounds) capable of modifying transporter-enzyme interplay. To ascertain the mechanism of interactions between antiretroviral drugs and garlic supplements and to identify the garlic constituents responsible, the hepatic pharmacokinetics of two antiretroviral drugs was investigated in the presence of garlic phytochemicals and aged garlic extract. For this purpose, rat liver slices and isolated rat hepatocytes were used. Aged garlic extract significantly inhibited saquinavir efflux from rat hepatocytes, while the efflux of darunavir significantly increased. Phytochemicals inducing distribution changes of saquinavir and darunavir were most probably flavonoids and lipophilic organosulfur compounds, respectively. All tested phytochemicals (except S-allyl L-cysteine) and aged garlic extract inhibited CYP3A4 metabolism of both drugs and modulated hepatic distribution of the corresponding saquinavir and darunavir metabolites. The competition between saquinavir and garlic constituent(s) for the same binding site on the efflux transporter and the positive cooperative effect between darunavir and garlic phytochemical(s), which bind to separate binding places on transporter, are the most probable mechanisms explaining the plasma profile changes, which could occur in vivo during concomitant consumption of antiretrovirals and garlic supplements.     

Therapeutic and diagnostic applications of nanoparticles

Nanoparticles are sphere-like biocompatible materials made of inert silica, metal or crystals of a few nanometers in size. They are emerging as a novel class of therapeutics for cancer treatment. Being more selective and specific toward their targets, nanoparticles have the ability to enhance the anticancer effects and to simultaneously reduce systemic toxicity compared with conventional therapeutics. Furthermore, they offer the potential to overcome drug resistance leading to higher intracellular drug accumulation. Nowadays, nanotechnologies are applied to molecular diagnostics and incorporated in cutting-edge molecular diagnostic methods, such as DNA and protein microarray biochips. Nanotechnologies enable diagnosis at the single-cell and single-molecule levels. Recent progress in cancer nanotechnology raises exciting opportunities for specific drug delivery. The purpose of this review is to give an overview about different types of nanoparticles and to summarize the latest results regarding their diagnostic and therapeutic applications in the clinic with more focus on cancer treatment. Furthermore, we discuss opportunities, limitations, and challenges faced by therapeutic nanoparticles.     

Effect of garlic-derived organosulfur compounds on mitochondrial function and integrity in isolated mouse liver mitochondria

The objectives of this work were to evaluate the direct effects of diallysulfide (DAS) and diallyldisulfide (DADS), two major organosulfur compounds of garlic oil, on mitochondrial function and integrity, by using isolated mouse liver mitochondria in a cell-free system. DADS produced concentration-dependent mitochondrial swelling over the range 125–1000 μM, while DAS was ineffective. Swelling experiments performed with de-energized or energized mitochondria showed similar maximal swelling amplitudes. Cyclosporin A (1 μM), or ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid (EGTA, 1 mM) were ineffective in inhibiting DADS-induced mitochondrial swelling. DADS produced a minor (12%) decrease in mitochondrial membrane protein thiols, but did not induce clustering of mitochondrial membrane proteins. Incubation of mitochondria with DADS (but not DAS) produced an increase in the oxidation rate of 2′,7′ dichlorofluorescein diacetate (DCFH-DA), together with depletion of reduced glutathione (GSH) and increased lipid peroxidation. DADS (but not DAS) produced a concentration-dependent dissipation of the mitochondrial membrane potential, but did not induce cytochrome c release. DADS-dependent effects, including mitochondrial swelling, DCFH-DA oxidation, lipid peroxidation and loss of mitochondrial membrane potential, were inhibited by antioxidants and iron chelators. These results suggest that DADS causes direct impairment of mitochondrial function as the result of oxidation of the membrane lipid phase initiated by the GSH- and iron-dependent generation of oxidants.     

硫化氢对离体大鼠心脏缺血/再灌注损伤的影响及机制初探

目的观察内源性CSE/H2S通路的改变以及给予H2S供体对缺血/再灌注心脏的影响,探讨该通路与心脏缺血/再灌注损伤的关系及作用机制。方法采用Langendorff离体灌流装置、通过停灌30min/复灌30min方式造成Wistar大鼠心肌缺血/再灌注损伤模型;采用外源性NaHS(40μmol.L-1)分别在停灌30min前(SIR)与停灌30min后处理(IRS)对缺血/再灌注心脏的影响。记录心脏收缩期左心室内压上升的最大变化速率(+dp/dtmax)、舒张期左心室内压下降的最大变化速率(-dp/dtmax)及左室内压差(LVP=左室收缩压-左室舒张压)。采用比色法检测灌流液中乳酸脱氢酶(LDH)、心肌MDA及SOD;采用比色法检测心肌胱硫醚-γ-裂解酶(CSE)活性;采用RT-PCR方法测定心肌组织CSEmRNA表达。结果与缺血/再灌注组(I/R)30min相比,SIR组及IRS组±dp/dtmax、LVP均增高,LDH降低;I/R组MDA水平高于对照组(CON)、SIR组及IRS组(P<0.05,P<0.01);IR组SOD活性低于SIR组及IRS组(P<0.05),但与CON组差别无显著性;I/R组大鼠心肌CSE活性低于CON组(P<0.05);而大鼠心肌CSEmRNA的表达与CON组差异无显著性。结论在缺血前后给予外源性NaHS均可改善因再灌注损伤引起的心肌收缩及舒张功能障碍;其作用机制可能是通过提高心肌SOD活性,增加氧自由基清除而拮抗缺血/再灌注引起的心功能及细胞膜损伤;心肌缺血/再灌注时内源性CSE活性抑制可能与心功能障碍及细胞损伤有关。     

The ozonation of unsaturated fatty acids: aldehydes and hydrogen peroxide as products and possible mediators of ozone toxicity

The products of the reactions of ozone with aqueous emulsions of unsaturated fatty acids and with liposomes made from phosphatidylcholine esters were characterized. Ozonolysis of emulsions of methyl oleate yields approximately 1 mol of hydrogen peroxide and 2 mol of aldehydes per mole of ozone used and fatty acid reacted. That is, the net equation that occurs is RCH = CHR' + O3 + H2O----RCHO + R'CHO + H2O2 . Ozonolysis of emulsions of oleic, linoleic, linolenic, and arachidonic acids gives 1 mol of hydrogen peroxide per mole of ozone used. Only very low yields (less than 5%) of reducible materials other than hydrogen peroxide are observed, suggesting that the yields of organic peroxidic materials, including Criegee ozonides and lipid hydroperoxides, are small. Ozonolysis of rat erythrocyte ghost membranes and rat bronchoalveolar lavage also gives significant yields (about 50%) of hydrogen peroxide based on the moles of ozone consumed. Reactions of ozone with bovine serum albumin, glutathione, and glucose do not produce hydrogen peroxide, implying that the hydrogen peroxide formed during the ozonation of biological materials arises almost exclusively from ozone/olefin reactions. Hydrogen peroxide and aldehydes are suggested to be important mediators of the modifications observed in both the lung and extrapulmonary tissues when ozone is inhaled.     

Selective inhibitors of cardiac ADPR cyclase as novel anti-arrhythmic compounds

ADP-ribosyl cyclases (ADPRCs) catalyse the conversion of nicotinamide adenine dinucleotide to cyclic adenosine diphosphoribose (cADPR) which is a second messenger involved in Ca(2+) mobilisation from intracellular stores. Via its interaction with the ryanodine receptor Ca(2+) channel in the heart, cADPR may exert arrhythmogenic activity. To test this hypothesis, we have studied the effect of novel cardiac ADPRC inhibitors in vitro and in vivo in models of ventricular arrhythmias. Using a high-throughput screening approach on cardiac sarcoplasmic reticulum membranes isolated from pig and rat and nicotinamide hypoxanthine dinuleotide as a surrogate substrate, we have identified potent and selective inhibitors of an intracellular, membrane-bound cardiac ADPRC that are different from the two known mammalian ADPRCs, CD38 and CD157/Bst1. We show that two structurally distinct cardiac ADPRC inhibitors, SAN2589 and SAN4825, prevent the formation of spontaneous action potentials in guinea pig papillary muscle in vitro and that compound SAN4825 is active in vivo in delaying ventricular fibrillation and cardiac arrest in a guinea pig model of Ca(2+) overload-induced arrhythmia. Inhibition of cardiac ADPRC prevents Ca(2+) overload-induced spontaneous depolarizations and ventricular fibrillation and may thus provide a novel therapeutic principle for the treatment of cardiac arrhythmias.