NCX-4040, a nitric oxide-releasing aspirin, sensitizes drug-resistant human ovarian xenograft tumors to cisplatin by depletion of cellular thiols

Background: Ovarian carcinoma is the leading cause of mortality among gynecological cancers in the world. The high mortality rate is associated with lack of early diagnosis and development of drug resistance. The antitumor efficacy and mechanism of NCX-4040, a nitric oxide-releasing aspirin derivative, against ovarian cancer is studied. Methods: NCX-4040, alone or in combination with cisplatin (cis-diamminedichloroplatinum, cDDP), was studied in cisplatin-sensitive (A2780 WT) and cisplatin-resistant (A2780 cDDP) cell lines as well as xenograft tumors grown in nude mice. Electron paramagnetic resonance (EPR) was used for measurements of nitric oxide and redox state. Immunoblotting analysis of A2780 cDDP tumor xenografts from mice was used for mechanistic studies. Results: Cells treated with NCX-4040 (25 μM) showed a significant reduction of cell viability (A2780 WT, 34.9 ± 8.7%; A2780 cDDP, 41.7 ± 7.6%; p < 0.05). Further, NCX-4040 significantly enhanced the sensitivity of A2780 cDDP cells (cisplatin alone, 80.6 ± 11.8% versus NCX-4040+cisplatin, 26.4 ± 7.6%; p < 0.01) and xenograft tumors (cisplatin alone, 74.0 ± 4.4% versus NCX-4040+cisplatin, 56.4 ± 7.8%; p < 0.05), to cisplatin treatment. EPR imaging of tissue redox and thiol measurements showed a 5.5-fold reduction (p < 0.01) of glutathione in NCX-4040- treated A2780 cDDP tumors when compared to untreated controls. Immunoblotting analysis of A2780 cDDP tumor xenografts from mice treated with NCX-4040 and cisplatin revealed significant downregulation of pEGFR (Tyr845 and Tyr992) and pSTAT3 (Tyr705 and Ser727) expression. Conclusion: The results suggested that NCX-4040 could resensitize drug-resistant ovarian cancer cells to cisplatin possibly by depletion of cellular thiols. Thus NCX-4040 appears to be a potential therapeutic agent for the treatment of human ovarian carcinoma and cisplatin-resistant malignancies

Studies of hypoxemic/reoxygenation injury: With aortic clamping XIII. Interaction between oxygen tension and cardioplegiccomposition in limiting nitric oxide production and oxidant damage

AbstractThis study tests the interaction between oxygen tension and cardioplegic composition on nitric oxide production and oxidant damage during reoxygenation of previously cyanotic hearts. Of 35 Duroc-Yorkshire piglets (2 to 3 weeks, 3 to 5 kg), six underwent 30 minutes of blood cardioplegic arrest with hyperoxemic (oxygen tension about 400 mm Hg), hypocalcemic, alkalotic, glutamate/aspartate blood cardioplegic solution during 1 hour of cardiopulmonary bypass without hypoxemia (control). Twenty-nine others were subjected to up to 120 minutes of ventilator hypoxemia (oxygen tension about 25 mm Hg) before reoxygenation on CPB. To simulate routine clinical management, nine piglets underwent uncontrolled cardiac reoxygenation , whereby cardiopulmonary bypass was started at oxygen tension of about 400 mm Hg followed by the aforementioned blood cardioplegic protocol 5 minutes later. All 20 other piglets underwent controlled cardiac reoxygenation , whereby cardiopulmonary bypass was started at the ambient oxygen tension (about 25 mm Hg), and reoxygenation was delayed until blood cardioplegia was given. The blood cardioplegia solution was kept normoxemic (oxygen tension about 100 mm Hg) in 10 piglets and made hyperoxemic (oxygen tension about 400 mm Hg) in 10 others. The cardioplegic composition was also varied so that the cardioplegic solution in each subgroup contained either KCl only (30 mEq/L) or components that theoretically inhibit nitric oxide synthase by including hypocalcemia, alkalosis, and glutamate/aspartate. Function (end-systolic elastance) and myocardial nitric oxide production, conjugated diene production, and antioxidant reserve capacity were measured. Blood cardioplegic arrest without hypoxemia did not cause myocardial nitric oxide or conjugated diene production, reduce antioxidant reserve capacity, or change left ventricular functional recovery. In contrast, uncontrolled cardiac reoxygenation raised nitric oxide and conjugated diene production 19- and 13-fold, respectively ( p < 0.05 vs control), reduced antioxidant reserve capacity 40%, and contractility recovered only 21% of control levels. After controlled cardiac reoxygenation at oxygen tension about 400 mm Hg with cardioplegic solution containing KCl only, nitric oxide and conjugated diene production rose 16- and 12-fold, respectively ( p < 0.05 vs control), and contractility recovered only 43% ± 5%. Normoxemic (oxygen tension of about 100 mm Hg) controlled cardiac reoxygenation with the same solution reduced nitric oxide and conjugated diene production 85% and 71%, and contractile recovery rose to 55% ± 7% ( p < 0.05 vs uncontrolled reoxygenation). In comparison, controlled cardiac reoxygenation with an oxygen tension of about 400 mm Hg hypocalcemic, alkalotic, glutamate/aspartate blood cardioplegic solution reduced nitric oxide and conjugated diene production 85% and 62%, respectively, and contractility recovered 63% ± 4% ( p < 0.05 vs KCl only). Normoxemic delivery of this solution resulted in negligible nitric oxide and conjugated diene production and 83% ± 8% recovery of contractility ( p < 0.05 vs all other groups). These data show correlation between nitric oxide production during initial reoxygenation and the extent of oxidant damage (i.e., conjugated diene production) and link functional recovery to suppression of excessive nitric oxide production and limitation of lipid peroxidation by the interaction of oxygen tension and cardioplegic composition during initial reoxygenation. (J THORAC CARDIOVASC SURG 1995; 110:1274-86

Long-term treatment with sulfhydryl angiotensin-converting enzyme inhibition reduces carotid intima-media thickening and improves the nitric oxide/oxidative stress pathways in newly diagnosed patients with mild to moderate primary hypertension.

Aim. Sulfhydryl angiotensin-converting enzyme (ACE) inhibitors exert antiatherosclerotic effects in preclinical models and antioxidant effects in patients. However, whether ACE inhibitors have any clinically significant antiatherogenic effects remains still debated. Objectives. In mildly hypertensive patients, we evaluated the effect of the sulfhydryl ACE inhibitor zofenopril in comparison with the carboxylic ACE inhibitor enalapril on carotid atherosclerosis (intima-media thickness [IMT] and vascular lumen diameter) and systemic oxidative stress (nitrite/nitrate, asymmetrical dimethyl-L-arginine, and isoprostanes). Material and methods. In 2001, we started a small prospective randomized clinical trial on 48 newly diagnosed mildly hypertensive patients with no additional risk factors for atherosclerosis (eg, hyperlipidemia, smoke habit, familiar history of atherosclerosisrelated diseases or diabetes). Patients were randomly assigned either to the enalapril (20 mg/d, n = 24) or the zofenopril group (30 mg/d, n = 24); the planned duration of the trial was 5 years. Carotid IMT and vascular lumen diameter were determined by ultrasonography for all patients at baseline and at 1, 3, and 5 years. Furthermore, nitrite/nitrate, asymmetrical dimethyl-L-arginine, and isoprostane levels were measured. Results. In our conditions, IMT of the right and left common carotid arteries was similar at baseline in both groups (P = NS). Intima-media thickness measurements until 5 years revealed a significant reduction in the zofenopril group but not in the enalapril group (P b .05 vs enalapril-treated group). This effect was coupled with a favorable nitric oxide/oxidative stress profile in the zofenopril group. Conclusion. Long-term treatment with the sulfhydryl ACE inhibitor zofenopril besides its blood pressure–lowering effects may slow the progression of IMT of the carotid artery in newly diagnosed mildly hypertensive patients. (Am Heart J 2008;156:1154.e1-1154.e8.

Biological hydropersulfides and related polysulfides – a new concept and perspective in redox biology

The chemical biology of thiols (RSH, e.g., cysteine and cysteine‐containing proteins/peptides) has been a topic of extreme interest for many decades due to their reported roles in protein structure/folding, redox signaling, metal ligation, cellular protection, and enzymology. While many of the studies on thiol/sulfur biochemistry have focused on thiols, relatively ignored have been hydropersulfides (RSSH) and higher order polysulfur species (RSSnH, RSSnR, n > 1). Recent and provocative work has alluded to the prevalence and likely physiological importance of RSSH and related RSSnH. RSSH of cysteine (Cys‐SSH) has been found to be prevalent in mammalian systems along with Cys‐SSH‐containing proteins. The RSSH functionality has not been examined to the extent of other biologically relevant sulfur derivatives (e.g., sulfenic acids, disulfides, etc.), whose roles in cell signaling are strongly indicated. The recent finding of Cys‐SSH biosynthesis and translational incorporation into proteins is an unequivocal indication of its fundamental importance and necessitates a more profound look into the physiology of RSSH. In this Review, we discuss the currently reported chemical biology of RSSH (and related species) as a prelude to discussing their possible physiological roles

the novel role of epigenetics in primary prevention of cardiovascular diseases

A great deal of evidences indicate that impaired fetal growth and in utero exposure to risk factors, especially maternal hypercholesterolemia, may be relevant for human pathophysiological signs of atherosclerosis and subsequent development of cardiovascular disease (CVD) during different life stages. Despite the underlying mechanisms of fetal programming are still unknown, epigenetics has been suggested as one of the possible explanations for the associations between intrauterine risk factors and CVD development. Indeed, a lot of translational studies support the hypothesis that epigenetic changes are related to increased CVD risk although it is still not possible to establish a direct causality in humans. Notably, epigenetic modifications can be reversible through therapeutic approaches employing histone deacetylase inhibitors, histone acetyltransferase inhibitors and commonly used drugs like statins. Thus, the whole comprehension of these mechanisms will provide in the next future the rationale for the development of novel tools to be used in the primary prevention and therapy of CVD

Comment on “Evidence that the ProPerDP method is inadequate for protein persulfidation detection due to lack of specificity”

The recent report by Fan et al. alleged that the ProPerDP method is inadequate for the detection of protein persulfidation. Upon careful evaluation of their work, we conclude that the claim made by Fan et al. is not supported by their data, rather founded in methodological shortcomings. It is understood that the ProPerDP method generates a mixture of cysteine-containing and non–cysteine-containing peptides. Instead, Fan et al. suggested that the detection of non–cysteine-containing peptides indicates nonspecific alkylation at noncysteine residues. However, if true, then such peptides would not be released by reduction and therefore not appear as products in the reported workflow. Moreover, the authors’ biological assessment of ProPerDP using Escherichia coli mutants was based on assumptions that have not been confirmed by other methods. We conclude that Fan et al. did not rigorously assess the method and that ProPerDP remains a reliable approach for analyses of protein per/polysulfidation