Nitrosothiols as no-donor drugs: Synthesis, mechanistic studies, chemical stability, pharmacological and physiological activity

S-Nitrosothiols (RSNO) are an important class of NO-donor drugs. They have been used clinically and occur naturally where they may have a role in several biological and physiological processes in the human body. The medical importance of S-nitrosothiols has been highlighted recently by several reports which describe the clinical use of GSNO (13) to inhibit platelet aggregation during coronary angioplasty and also to treat a form of preeclampsia, a high blood pressure condition suffered by some pregnant women. We set out to extend the range of compounds of this type by synthesising a novel series of biologically active S-nitrosothiols (1-13) and to look for a correlation between structure, chemical stability, and physiological activity. The situation has been complicated by the recent discovery that the main route for the release of NO from S-nitrosothiols is a copper- catalysed process. A detailed kinetic study of copper ions, and thiols on the stability of the compounds we synthesised has shown that the dominant pathway for the decomposition of S-nitrosothiols in most circumstances is one catalysed by Cu⁺ ions. We suggest that Cu⁺ ions are formed by the reaction of Cu²⁺ ions with thiol, present in the S-nitrosothiols as an impurity. The implications of this discovery for an understanding of the biological action of S-nitrosothiols is suggested. All the new S-nitrosated dipeptides (2-12) examined show less susceptibility to copper (I)- catalysed release of NO than SNAP (1) but are more reactive than GSNO (13). We found that S-nitrosated dipeptides are potent vasodilators and suitable inhibitors of platelet aggregation but are chemically very stable in the absence of copper ions. All thirteen compounds combine the favoured property of chemical stability with a high level of biological activity. We found that copper(I)-chelation induced reduction of the biological activity of S- nitrosothiols in smooth muscle relaxation. The results show that responses to both SNAP and GSNO are reversibly inhibited by neocuproine. We conclude that relaxation of vasodilator smooth muscle by SNAP and GSNO is caused in part by NO released into solution via a Cu⁺-dependent catalytic reaction, and provide evidence that endogenous Cu⁺ ions may also contribute to the maintenance of vasodilator 'store' in vivo by catalysing the decomposition of naturally-occurring S-nitrosothiols. A particularly interesting finding recently by Gordge et al. (1995) shows that the inhibition of platelet aggregation activity shown by GSNO is much reduced in the presence of neocuproine and the closely related bathocuproine, both specific Cu⁺-chelating agent. However, it was shown very recently (Schrammel et al, 1996) that copper ions inhibit basal and NO-stimulated recombinant soluble guanylate cyclase activity and that Cu⁺ is more effective than Cu²⁺ in this regard. L-Ascorbic acid (vitamin C) could play a role in the in vivo release of NO from naturally occurring and exogenous S-nitrosothiols and so play a part in smooth muscle relaxation and in inhibition of platelet aggregation. All thirteen compounds examined show the ability to release NO in vitro. The inhibitory effect of Hb, a recognised NO scavenger, was investigated. In smooth muscle, responses to intermediate doses of S-nitrosothiols were significantly inhibited by Hb, though not abolished entirely. In platelet, we found that the inhibitory activity of these S-nitrosothiols was reversed by haemoglobin, indicating the involvement of NO in the process. We found that the solution stability of the S-nitrosothiols did not correlate with relaxation of vascular smooth muscle or inhibition of platelet aggregation, again suggesting that the tissue specificity is a function of the R- group. We conclude that the biological activity of S-nitrosothiols depends upon the release of NO in a process catalysed by Cu(I), and that the decomposition may occur inside or outside the cell, depending upon the structure of RSNO

S-nitrosothiols as nitric oxide-donors: Chemistry, biology and possible future therapeutic applications

In recent years, the gaseous molecule nitric oxide (NO) has been shown to be involved in many important biological events. S-Nitrosothiols are biological substances derived endogenously from NO, and are found in a variety of tissues exhibiting NO-mimetic activity. Fundamental studies on the chemical aspects of S-nitrosothiols have become an integral part of NO research, with a view to further understanding the numerous biological functions both of NO and of S-nitrosothiols themselves. It has often been suggested that S-nitrosothiols represent a means either for the storage or transport of NO, although the evidence for this is sparse. Many S-nitrosothiols have now been synthesized chemically, and at present they show the most promise as clinically useful NO-donor drugs. In this review, we examine in detail the biological functions of S-nitrosothiols, as well as the chemical properties and biomedical applications of these compounds

Current status and future possibilities of nitric oxide-donor drugs: Focus on S-nitrosothiols

Drugs which release nitric oxide (NO) have great therapeutic potential. The organic nitrates and sodium nitroprusside have been used in cardiovascular therapeutics for many years, but several drawbacks limit their usefulness. In this review, we consider novel and potential future developments in NO-donor drugs, and the possible clinical usefulness of such compound

Neocuproine, a selective Cu (I) chelator, and the relaxation of rat vascular smooth muscle by S-nitrosothiols

1 A study has been made of the effect of neocuproine, a specific Cu(I) chelator, on vasodilator responses of rat isolated perfused tail artery to two nitrosothiols: S-nitroso-N-acetyl-D,L-penicillamine (SNAP) and S-nitroso-glutathione (GSNO).2 Bolus injections (10 mu l) of SNAP or GSNO (10(-7)-10(-3) M) Were delivered into the lumen of perfused vessels pre-contracted with sufficient phenylephrine (1-7 mu M) to develop pressures of 100-120 mmHg. Two kinds of experiment were made: SNAP and GSNO were either (a) pre-mixed with neocuproine (10(-4) M) and then injected into arteries; or (b) vessels were continuously perfused with neocuproine (10(-5) M) and then injected with either pure SNAP or GSNO.3 In each case, neocuproine significantly attenuated vasodilator responses to both nitrosothiols, although the nature of the inhibitory effect differed in the two types of experiment. We conclude that the ability of exogenous nitrosothiols to relax vascular smooth muscle in our ex vivo model is dependent upon a Cu(I) catalyzed process. Evidence is presented which suggests that a similar Cu(I)-dependent mechanism is responsible for the release of NO from endogenous nitrosothiols and that this process may assist in maintaining vasodilator tone in vivo.</p

A novel family of S-nitrosothiols: Chemical synthesis and biological actions

S-Nitrosothiols are a class of chemical compounds that decompose to release nitric oxide and show promise in the treatment of a variety of cardiovascular diseases. Some of these are present in vivo and others have been synthesized in vitro. However, those discovered or synthesized to date have very little tissue selectivity or specificity. We synthesized a number of novel S-nitrosated dipeptides of high purity and examined their effects on vasorelaxation using rat mesenteric arteries and on inhibition of platelet aggregation using platelets from healthy human subjects. For comparison, we also tested the effects of S-nitroso-L-glutathione (GSNO, an S-nitrosothiol present in vivo) and S-nitroso-N-acetyl-D-beta,beta -dimethylcystein (SNAP(D), the D-isomer of SNAP, a commonly used S-nitrosothiol previously synthesized in vitro) in these biological systems. Satisfactory elemental analyses were obtained for all compounds synthesized (less than +/- 0.3%), and all accurate mass measurements were within 1-5 ppm of the expected mass. The novel S-nitrosated dipeptides all elicited vasorelaxation with significantly higher potency, of the order of one log molar unit, than either GSNO or SNAP(D). However, all compounds inhibited U46619-induced platelet aggregation with similar potency to GSNO and SNAP(D). These findings indicate a degree of tissue selectivity which may prove to be of therapeutic usefulness. (C) 2000 Academic Press

Neocuproine, a selective Cu(I) chelator, and the relaxation of rat vascular smooth muscle by S-nitrosothiols

1. A study has been made of the effect of neocuproine, a specific Cu(I) chelator, on vasodilator responses of rat isolated perfused tail artery to two nitrosothiols: S-nitroso-N-acetyl-D,L-penicillamine (SNAP) and S-nitroso-glutathione (GSNO). 2. Bolus injections (10 μl) of SNAP or GSNO (10(−7)–10(−3) M) were delivered into the lumen of perfused vessels pre-contracted with sufficient phenylephrine (1–7 μM) to develop pressures of 100–120 mmHg. Two kinds of experiment were made: SNAP and GSNO were either (a) pre-mixed with neocuproine (10(−4) M) and then injected into arteries; or (b) vessels were continuously perfused with neocuproine (10(−5) M) and then injected with either pure SNAP or GSNO. 3. In each case, neocuproine significantly attenuated vasodilator responses to both nitrosothiols, although the nature of the inhibitory effect differed in the two types of experiment. We conclude that the ability of exogenous nitrosothiols to relax vascular smooth muscle in our ex vivo model is dependent upon a Cu(I) catalyzed process. Evidence is presented which suggests that a similar Cu(I)-dependent mechanism is responsible for the release of NO from endogenous nitrosothiols and that this process may assist in maintaining vasodilator tone in vivo