Cardiovasc Drug Rev. 2006 Summer;24(2):148-68.
Pharmacologic profile and therapeutic potential of NCX 4016, a nitric oxide-releasing aspirin, for cardiovascular disorders.
Gresele P, Momi S. Department of Internal Medicine, Division of Internal and Cardiovascular Medicine, University of Perugia, Perugia, Italy.
NCX 4016, 2-(acetyloxy)benzoic acid 3-[(nitrooxy)methyl]phenyl ester, is a new molecule in which a nitric oxide (NO)-releasing moiety is covalently linked to aspirin. After enzymatic metabolism, NCX 4016 releases both components. In vitro and in some animal models, these components exert their pharmacologic effects simultaneously. Nitric oxide (NO) is a small gaseous molecule that exerts several activities which may prevent atherothrombotic disorders. Moreover, it displays a protective activity on the gastric mucosa. NCX 4016 has been shown to inhibit platelet activation in vitro more effectively than aspirin, to inhibit smooth muscle cell proliferation, to exert an endothelial cell protective activity and to suppress the function of several inflammatory cells potentially involved in atherothrombosis. In animal models, NCX 4016 protected from platelet thromboembolism, prevented restenosis in atherosclerosis-prone animals, protected the heart from ischemia/reperfusion injury, and induced neoangiogenesis in critically ischemic limbs. Moreover, it displayed little or no gastric toxicity and appeared to protect stomach from noxious stimuli, including aspirin. NCX 4016 has been evaluated in healthy volunteers and found to inhibit platelet cyclo-oxygenase-1 (COX-1) similarly to or slightly less than aspirin, to raise the circulating levels of NO-degradation products, and to have little or no gastric toxicity in short term studies. In particular, in phase II studies, NCX 4016 had favorable effects on effort-induced endothelial dysfunction in intermittent claudication and on platelet-activation parameters elicited by short-term hyperglycemia in type II diabetics. In patients with type II diabetes the effects of NCX 4016 on microalbuminuria and on some hemodynamic parameters were promising. The pharmacokinetics of in vivo aspirin- and NO- released by NCX 4016, as well as the bioavailability of the two molecules, were not yet adequately studied. Also, the long-term tolerability of NCX 4016, as well as its possible effectiveness in preventing ischemic cardiovascular events and progression of atherosclerosis, should be explored.
Cell Cycle. 2006 Aug;5(15):1669-74. Epub 2006 Aug 1.
Nitrogen oxide-releasing aspirin induces histone H2AX phosphorylation, ATM activation and apoptosis preferentially in S-phase cells: involvement of reactive oxygen species.
Tanaka T, Kurose A, Halicka HD, Huang X, Traganos F, Darzynkiewicz Z. Brander Cancer Research Institute and Department of Pathology, New York Medical College, Valhalla, New York, USA.
Nitric oxide-releasing acetylsalicylic acid (NO-ASA; NO-aspirin) developed as an anti-inflammatory agent that was expected to avoid some of the adverse effects of aspirin (ASA), was recently shown to be cytotoxic to cells of different tumor lines. The cytotoxic properties and potency of NO-ASA are different than those of ASA which implies that the intracellular targets for NO-ASA and ASA, and their mechanism of action, are different. The aim of the present study was to reveal whether the cytotoxicity induced by NO-ASA is mediated by damage to DNA. We observed that even brief (1 h) treatment of human B-lymphoblastoid TK6 cells with >or=5 microM NO-ASA led to DNA damage revealed by the alkaline and neutral comet assays, histone H2AX phosphorylation on Ser 139, and ATM phosphorylation on Ser 1981, a marker of activation of this kinase. The induction of H2AX phosphorylation was preferential to S-phase cells. Exposure to >or=5 microM NO-ASA for over 3 h led to apoptosis, also preferentially of S-phase cells. Apoptosis was atypical; while chromatin was highly condensed there was no evidence of nuclear fragmentation nor were the cells positive in the TUNEL assay though they did express activated caspase-3. The induction of phosphorylation of H2AX on Ser 139 by NO-ASA was markedly attenuated in the presence of N-acetyl-L-cysteine, a scavenger of reactive oxygen species (ROS). The data imply that the NO-ASA induces DNA damage through oxidative stress; the oxidation-generated lesions provide a signal for induction of H2AX phosphorylation during DNA replication, perhaps when the progressing replication forks collide with the primary lesions converting them to DNA double-strand breaks. Because neither induction of H2AX phosphorylation nor apoptosis were observed at equimolar concentrations of ASA, the NO moiety attached to ASA appeared to mediate these effects.
Apoptosis. 2006 Aug;11(8):1321-30.
Molecular characterization of cytotoxic and resistance mechanisms induced by NCX 4040, a novel NO-NSAID, in pancreatic cancer cell lines.
Rosetti M, Tesei A, Ulivi P, Fabbri F, Vannini I, Brigliadori G, Amadori D, Bolla M, Zoli W. Istituto Oncologico Romagnolo, Meldola, Italy.
Although non steroidal antiinflammatory drugs (NSAIDs) have been shown to be effective as chemopreventive agents, important side-effects limit their clinical use. A promising novel class of drugs, nitric oxide-donating NSAIDs (NO-NSAIDs), has been found to be more active than classical NSAIDs. This study explored the effect of the NO-donating aspirin derivative, NCX 4040, on three human pancreatic adenocarcinoma cell lines (Capan-2, MIA PaCa-2 and T3M4). NCX 4040 activity was compared with that of NCX 4016 (an NO(2)-positional isomer of NCX 4040), SNAP (a standard NO-releasing molecule), NCX 4042 (denitrated analog of NCX 4040), and aspirin. NCX 4040 showed a striking cytocidal activity in all cell lines, already inducing significant percentages of apoptotic cells at 10 muM in Capan-2 cell lines. This study focused on the biological mechanisms of sensitivity and resistance to NCX 4040, highlighting that the cytotoxic action of this drug may be due to the hyperexpression of Bax, its translocation to the mitochondria, the release of Cytochrome C, and the activation of caspases-9 and -3, overall in a p53-independent manner. Moreover, the use of a specific COX-2 inhibitor (NS 398) in the experimental models showed that COX-2 hyperexpression could partially explain the resistance mechanisms to NCX 4040.
J Submicrosc Cytol Pathol. 2006 Jun-Sep;38(2-3):149-54.
Ultracytochemical demonstration of soluble guanylate cyclase activation in rat aorta by NCX4016, a NO-releasing aspirin derivative.
Rambotti MG, Mariucci G, Tantucci M, Ambrosini MV.
Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Perugia, Italy.
Biochemical studies demonstrate that the NO-releasing-aspirin derivative (NCX4016) stimulates soluble guanylate cyclase (sGC) activity and increases cyclic GMP (cGMP) in human platelet and monocytes by releasing NO. In the present study, an ultracytochemical technique for electron microscopy was used to investigate the effects of NCX4016 (2 mM) on sGC activity in rat thoracic aorta, using sodium nitroprusside (0.01 mM) as reference NO-donor. Guanylyl-imidodiphosphate sodium salt [Gpp(NH)p], a synthetic non-hydrolyzable analogue of GTP, was used as sGC substrate. NO-activated sGC released imidodiphosphate ions which were precipitated with lead ions, giving rise to deposits of electron-dense granules (reaction product). Ultracytochemistry allowed us to demonstrate that NCX4016 stimulated sGC activity in smooth muscle cells, and particularly in vascular endothelial cells, as sodium nitroprusside did. This result could explain the protective effects of chronic treatment with NCX4016 on aortic endothelium of diabetic rats demonstrated by scanning and transmission electron microscopy.
