Exploring the effects of topoisomerase II inhibitor XK469 on anthracycline cardiotoxicity and DNA damage.

Anthracyclines, such as doxorubicin (adriamycin), daunorubicin, or epirubicin, rank among the most effective agents in classical anticancer chemotherapy. However, cardiotoxicity remains the main limitation of their clinical use. Topoisomerase IIß has recently been identified as a plausible target of anthracyclines in cardiomyocytes. We examined the putative topoisomerase IIß selective agent XK469 as a potential cardioprotective and designed several new analogs. In our experiments, XK469 inhibited both topoisomerase isoforms (α and ß) and did not induce topoisomerase II covalent complexes in isolated cardiomyocytes and HL-60, but induced proteasomal degradation of topoisomerase II in these cell types. The cardioprotective potential of XK469 was studied on rat neonatal cardiomyocytes, where dexrazoxane (ICRF-187), the only clinically approved cardioprotective, was effective. Initially, XK469 prevented daunorubicin-induced toxicity and p53 phosphorylation in cardiomyocytes. However, it only partially prevented the phosphorylation of H2AX and did not affect DNA damage measured by Comet Assay. It also did not compromise the daunorubicin antiproliferative effect in HL-60 leukemic cells. When administered to rabbits to evaluate its cardioprotective potential in vivo, XK469 failed to prevent the daunorubicin-induced cardiac toxicity in either acute or chronic settings. In the following in vitro analysis, we found that prolonged and continuous exposure of rat neonatal cardiomyocytes to XK469 led to significant toxicity. In conclusion, this study provides important evidence on the effects of XK469 and its combination with daunorubicin in clinically relevant doses in cardiomyocytes. Despite its promising characteristics, long-term treatments and in vivo experiments have not confirmed its cardioprotective potential.

Examination of diverse iron-chelating agents for the protection of differentiated PC12 cells against oxidative injury induced by 6-hydroxydopamine and dopamine.

Labile redox-active iron ions have been implicated in various neurodegenerative disorders, including the Parkinson's disease (PD). Iron chelation has been successfully used in clinical practice to manage iron overload in diseases such as thalassemia major; however, the use of conventional iron chelators in pathological states without systemic iron overload remains at the preclinical investigative level and is complicated by the risk of adverse outcomes due to systemic iron depletion. In this study, we examined three clinically-used chelators, namely, desferrioxamine, deferiprone and deferasirox and compared them with experimental agent salicylaldehyde isonicotinoyl hydrazone (SIH) and its boronate-masked prochelator BSIH for protection of differentiated PC12 cells against the toxicity of catecholamines 6-hydroxydopamine and dopamine and their oxidation products. All the assayed chelating agents were able to significantly reduce the catecholamine toxicity in a dose-dependent manner. Whereas hydrophilic chelator desferrioxamine exerted protection only at high and clinically unachievable concentrations, deferiprone and deferasirox significantly reduced the catecholamine neurotoxicity at concentrations that are within their plasma levels following standard dosage. SIH was the most effective iron chelator to protect the cells with the lowest own toxicity of all the assayed conventional chelators. This favorable feature was even more pronounced in prochelator BSIH that does not chelate iron unless its protective group is cleaved in disease-specific oxidative stress conditions. Hence, this study demonstrated that while iron chelation may have general neuroprotective potential against catecholamine auto-oxidation and toxicity, SIH and BSIH represent promising lead molecules and warrant further studies in more complex animal models.

Screening of Synthetic Heterocyclic Compounds as Antiplatelet Drugs.

BACKGROUND: Antiplatelet drugs represent the keystone in the treatment and prevention of diseases of ischemic origin, including coronary artery disease. The current palette of drugs represents efficient modalities in most cases, but their effect can be limited in certain situations or associated with specific side effects. In this study, representatives of compounds selected from series having scaffolds with known or potential antiplatelet activity were tested. These compounds were previously synthetized by us, but their biological effects have not yet been reported. OBJECTIVE: The aim of this study was to examine the antiplatelet and anticoagulation properties of selected compounds and determine their mechanism of action. METHODS: Antiplatelet activity of compounds and their mechanisms of action were evaluated using human blood by impedance aggregometry and various aggregation inducers and inhibitors and compared to appropriate standards. Cytotoxicity was tested using breast adenocarcinoma cell cultures and potential anticoagulation activity was also determined. RESULTS: In total, four of 34 compounds tested were equally or more active than the standard antiplatelet drug Acetylsalicylic Acid (ASA). In contrast to ASA, all 4 active compounds decreased platelet aggregation triggered not only by collagen, but also partly by ADP. The major mechanism of action is based on antagonism at thromboxane receptors. In higher concentrations, inhibition of thromboxane synthase was also noted. In contrast to ASA, the tested compounds did not block cyclooxygenase- 1. CONCLUSION: The most active compound, 2-amino-4-(1H-indol-3-yl)-6-nitro-4H-chromene-3- carbonitrile (2-N), which is 4-5x times more potent than ASA, is a promising compound for the development of novel antiplatelet drugs.

The Effect of Silymarin Flavonolignans and Their Sulfated Conjugates on Platelet Aggregation and Blood Vessels Ex Vivo.

Silymarin is a traditional drug and food supplement employed for numerous liver disorders. The available studies indicate that its activities may be broader, in particular due to claimed benefits in some cardiovascular diseases, but the contributions of individual silymarin components are unclear. Therefore, we tested silymarin flavonolignans as pure diastereomers as well as their sulfated metabolites for potential vasorelaxant and antiplatelet effects in isolated rat aorta and in human blood, respectively. Eleven compounds from a panel of 17 tested exhibited a vasorelaxant effect, with half maximal effective concentrations (EC50) ranging from 20 to 100 µM, and some substances retained certain activity even in the range of hundreds of nM. Stereomers A were generally more potent as vasorelaxants than stereomers B. Interestingly, the most active compound was a metabolite-silychristin-19-O-sulfate. Although initial experiments showed that silybin, 2,3-dehydrosilybin, and 2,3-dehydrosilychristin were able to substantially block platelet aggregation, their effects were rapidly abolished with decreasing concentration, and were negligible at concentrations ≤100 µM. In conclusion, metabolites of silymarin flavonolignans seem to have biologically relevant vasodilatory properties, but the effect of silymarin components on platelets is low or negligible.

4-Methylcatechol, a Flavonoid Metabolite with Potent Antiplatelet Effects.

SCOPE: Intake of flavonoids from the diet can be substantial, and epidemiological studies suggest that these compounds can decrease the incidence of cardiovascular diseases by involvement with increased platelet aggregation. Although parent flavonoids possess antiplatelet effects, the clinical importance is disputable due to their very low bioavailability. Most of them are metabolized by human colon bacteria to smaller phenolic compounds, which reach higher plasma concentrations than the parent flavonoids. In this study, a series of 29 known flavonoid metabolites is tested for antiplatelet potential. METHODS AND RESULTS: Four compounds appear to have a biologically relevant antiplatelet effect using whole human blood. 4-Methylcatechol (4-MC) is clearly the most efficient being about 10× times more active than clinically used acetylsalicylic acid. This ex vivo effect is also confirmed using a potentially novel in-vivo-like ex ovo hen's egg model of thrombosis, where 4-MC significantly increases the survival of the eggs. The mechanism of action is studied and it seems that it is mainly based on the influence on intracellular calcium signaling. CONCLUSION: This study shows that some flavonoid metabolites formed by human microflora have a strong antiplatelet effect. This information can help to explain the antiplatelet potential of orally given flavonoids.

The influence of microbial isoflavonoid specific metabolites on platelets and transition metals iron and copper.

BACKGROUND: Isoflavonoids seem to possess positive cardiovascular and other beneficial effects in humans. HYPOTHESIS: Their low bioavailability, however, indicates that small isoflavonoid metabolites formed by human microflora can significantly contribute to these activities. STUDY DESIGN: Testing antiplatelet activity ex vivo in human blood and interaction with transition metals in vitro. METHODS: The effect on platelet aggregation induced by different triggers (arachidonic acid, collagen, ADP, TRAP-6), and interactions with transition metals (iron and copper chelation/reduction) were evaluated against four isoflavonoid-specific metabolites: S-equol; O-desmethylangolensin; 2-(4-hydroxyphenyl) propionic acid (HPPA); and 4-ethylphenol. RESULTS: S-equol, 4-ethylphenol and O-desmethylangolensin blocked platelet aggregation induced by arachidonic acid and collagen. S-equol even matched the potency of acetylsalicylic acid in the case of collagen, which is the most physiological inducer of aggregation. Moreover, their effects in general seemed to be biologically relevant and attainable at achievable plasma concentrations, with the exception of HPPA which was ineffective. While only O-desmethylangolensin mildly chelated iron and copper, all four compounds markedly reduced cupric ions. Their direct free radical scavenging effects seem to have little clinical relevance. CONCLUSION: This study has shown that S-equol, O-desmethylangolensin and 4-ethylphenol, arising from isoflavonoid intake, can have biologically relevant effects on platelet aggregation.

Two flavonoid metabolites, 3,4-dihydroxyphenylacetic acid and 4-methylcatechol, relax arteries ex vivo and decrease blood pressure in vivo.

SCOPE: The flavonoid quercetin reduces arterial blood pressure in animals and humans but the mechanisms remains elusive. The aim of this study was to test the activity of flavonoid microbial metabolites, which can participate on the final vasorelaxant effect. METHODS AND RESULTS: Both ex vivo (isolated rat thoracic aorta and mesenteric artery) and in vivo (normotensive and spontaneously hypertensive rats) approaches were used in this study. 4-methylcatechol and 3,4-dihydroxyphenylacetic acid (DHPA) had greater vasorelaxant effects on mesenteric artery than 3-(3-hydroxyphenyl)propionic acid, the previously reported metabolite with vasorelaxant effect. In vivo testing confirmed their blood pressure decreasing effect given both as bolus and slow infusion. Their mechanism at molecular level was different. CONCLUSIONS: This study is the first to show that flavonoid metabolites DHPA and 4-methylcatechol decrease arterial blood pressure and hence a mixture of microbial metabolites formed in the gastrointestinal tract may be responsible for or contribute to the effect of orally ingested quercetin.

Correction to: Interaction of soy isoflavones and their main metabolites with hOATP2B1 transporter.

The published online version contains mistake in the caption of Figures 3, 4 and 5 for in front of the figure legends designations "a-g", "a-e", and "a-b" have been provided. Such data should be deleted.

Interaction of soy isoflavones and their main metabolites with hOATP2B1 transporter.

Membrane organic anion-transporting polypeptides (OATPs) are responsible for the drug transmembrane transport within the human body. The function of OATP2B1 transporter can be inhibited by various natural compounds. Despite increased research interest in soya as a part of human diet, the effect of its active components to interact with hOATP2B1 has not been elucidated in a complex extent. This in vitro study examined the inhibitory effect of main soy isoflavones (daidzin, daidzein, genistin, genistein, glycitin, glycitein, biochanin A, formononetin) and their metabolites formed in vivo (S-equol, O-desmethylangolensin) towards human OATP2B1 transporter. MDCKII cells overexpressing hOATP2B1 were employed to determine quantitative inhibitory parameters of the tested compounds and to analyze mechanism/s of the inhibitory interaction. The study showed that aglycones of soy isoflavones and the main biologically active metabolite S-equol were able to significantly inhibit hOATP2B1-mediated transport. The Ki values for most of aglycones range from 1 to 20 µM. In contrast, glucosides did not exhibit significant inhibitory effect. The kinetic analysis did not indicate a uniform type of inhibition towards the hOATP2B1 although predominant mechanism of inhibition seemed to be competitive. These findings may suggest that tested soy isoflavones and their metabolites might affect transport of xenobiotics including drugs across tissue barriers via hOATP2B1.

Comprehensive review of cardiovascular toxicity of drugs and related agents.

Cardiovascular diseases are a leading cause of morbidity and mortality in most developed countries of the world. Pharmaceuticals, illicit drugs, and toxins can significantly contribute to the overall cardiovascular burden and thus deserve attention. The present article is a systematic overview of drugs that may induce distinct cardiovascular toxicity. The compounds are classified into agents that have significant effects on the heart, blood vessels, or both. The mechanism(s) of toxic action are discussed and treatment modalities are briefly mentioned in relevant cases. Due to the large number of clinically relevant compounds discussed, this article could be of interest to a broad audience including pharmacologists and toxicologists, pharmacists, physicians, and medicinal chemists. Particular emphasis is given to clinically relevant topics including the cardiovascular toxicity of illicit sympathomimetic drugs (e.g., cocaine, amphetamines, cathinones), drugs that prolong the QT interval, antidysrhythmic drugs, digoxin and other cardioactive steroids, beta-blockers, calcium channel blockers, female hormones, nonsteroidal anti-inflammatory, and anticancer compounds encompassing anthracyclines and novel targeted therapy interfering with the HER2 or the vascular endothelial growth factor pathway.

9-(4'-dimethylaminophenyl)-2,6,7-trihydroxy-xanthene-3-one is a Potentially Novel Antiplatelet Drug which Antagonizes the Effect of Thromboxane A2.

BACKGROUND: Currently, used oral antiplatelet drugs are both limited and associated with the risk of treatment failure/resistance. Research in this area is hence highly desired. A series of xanthene-3-ones derivatives, we had synthesized, showed us that these derivatives had antiplatelet activity. As far as we know, no research on the effects of xanthen-3-ones in this area has been done. OBJECTIVE: The aim was to study the antiplatelet potential of a series of synthesised 9-phenylxanthene- 3-ones and to find the ideal structural feature(s) for antiplatelet potential and determine the mechanism of action. METHODS: The compounds were synthesized from 1,2,4-triacetoxybenzene and various benzaldehydes. The reaction proceeded smoothly under acidic alcoholic conditions, furnishing the desired products in good yields. The compounds were first screened in whole human blood where platelet aggregation was induced by arachidonic acid. Further analysis was targeted at search of the mechanism of action. RESULTS: Initial screening showed that a majority of the synthesized derivatives had substantial antiplatelet potential. None of the compounds were able to block cyclooxygenase 1 or thromboxane synthase. The mechanism appeared to be based on antagonism of thromboxane effects. The most potent compound 9-(4'-dimethylaminophenyl)-2,6,7-trihydroxy-xanthene-3-one had better potential to block collagen induced platelet aggregation than clinically used acetylsalicylic acid. CONCLUSION: The last mentioned derivative is promising for further in vivo testing.

The isoflavonoid tectorigenin has better antiplatelet potential than acetylsalicylic acid.

BACKGROUND: One reason for the lower incidence of cardiovascular diseases in Asian countries may be the high intake of isoflavonoids and their antiplatelet effects may be an important factor. To date, there is limited comparison of a range of isoflavonoids and knowledge of their effects at different levels of platelet aggregation. PURPOSE: To screen the antiplatelet effects of a number of isoflavonoids on the arachidonic acid based aggregation pathway and investigate how the antiplatelet activity might occur. METHODS: The antiplatelet effects were first screened in whole human blood where platelet aggregation was induced by arachidonic acid. Further analysis was targeted at search of the mechanism of action. RESULTS: Thirteen of the eighteen tested isoflavonoids had significant inhibitory effect on platelet aggregation in whole human blood. Genistein had the same potency as clinically used acetylsalicylic acid (ASA) while tectorigenin was clearly stronger than ASA. Further analyses showed that the effect of tectorigenin was not based on inhibition of cyclooxygenase-1 in contrast to ASA or thromboxane synthase but by competitive antagonism at thromboxane receptors. CONCLUSION: Tectorigenin is a more potent antiplatelet compound than ASA and thus an interesting substance for further testing.