Nitric oxide (NO) is a potent and multifaceted bioregulatory agent. This project is aimed at finding ways to target NO to specific sites in the body or in a test tube for important research and/or therapeutic applications.? The essential starting point for all our research is the continuing physicochemical characterization of a versatile class of NO-releasing prodrugs, the diazeniumdiolates. As an example, during the past year, we have introduced O2-alkylated diazeniumdiolates of structure RHN-N(O)=NOR' to the literature via a description of their synthesis and solvolysis chemistry; one such agent for which R' is a beta-D-glucose residue was shown to be activated by glucosidase to generate NO (collaboration with S. Bohle and J. Ivanic). ? This fundamental chemical research program serves as a versatile platform for designing improved biomedical research tools as well as potential clinical applications.
Work aim ed at characterizing the mechanisms of NO versus HNO (nitroxyl, a newly identified bioeffector species) release in model diazeniumdiolates is being pursued in collaboration with K. Miranda and D. Wink.? Glycosylated diazeniumdiolates have been shown to be reasonably stable at neutral or acidic pH but to undergo ready cleavage in aqueous base. This finding has allowed us to develop a convenient protecting group strategy for acid-sensitive R2NN(O)=NO- anions (collaboration with M. Meyerhoff).? One such anion, PROLI/NO (where R2N is the L-prolyl residue) has shown particular promise for biomedical applications because of its favorable toxicological profile and the fact that its dissociation to NO is so rapid (half-life 2 seconds at pH 7.4 and 37oC) that the pharmacological effects can be effectively localized at the point of introduction into the body. But this sensitivity to decomposition has complicated various attempts to formulate it for biomedical use. Nevertheless, we have been able during the past year to devise a successful procedure for reliably formulating it for intravascular infusion (collaboration with G. Grimes).? Such R2NN(O)=NO- anions have proven ideal for studying the reactivity of NO in aerobic aqueous media. Molecular NO can itself be used for this purpose, but the fact that it rapidly oxidizes to produce reactive nitrous acid (HONO) can greatly complicate interpretation of the results--were the reactions observed due to aerobic NO or to acidified nitrite? Conducting the study in well buffered media can of course be employed, but one must still be wary of transient local excesses of nitrite ion at low pH when the reactants are mixed. Diazeniumdiolate anions allow the researcher to circumvent this problem, because a basic amine moiety is generated at each and every site in the medium where an NO is released, ensuring that the pH in that microcosm cannot be lowered to activate the otherwise unreactive nitrite ion. ? This advantage of the simple R2NN(O)=NO- ions has been exploited to show that aerobic NO can induce nitrosative deamidation of asparagine and glutamine residues in peptides and proteins. Current work is aimed at determining the potential biological significance of this possible post-translational modification, whose mechanism differs in very fundamental ways from that of the established hydrolytic deamidation pathway.? Work continues on other aspects of the chemistry and pharmacology of NO and the diazeniumdiolates, including those in which the -N(O)=NO- group is attached to carbon centers. Practical applications of diazeniumdiolate chemistry are being developed, with patents being obtained where warranted. Also published during the current reporting period was a review of the emerging commercial opportunities we believe this project to present.
Showing the most recent 10 out of 28 publications
