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Epilepsy is prevalent with about 50 million patients affected worldwide. There are many treatment gaps in the management of epilepsy in India. Within Anti-epileptic drugs (AED), Brivaracetam, which is a high-affinity, selective, and reversible ligand for synaptic vesicle 2A is approved by the Food and Drug Administration for monotherapy as well as adjunctive treatment of focal seizures. A series of meeting occurring during April 2020 inviting neurologist across India as panel members, reviewed the efficacy and safety of brivaracetam and discussed the use of brivaracetam in clinical settings and the drivers and barriers for the use of brivaracetam in the management of epilepsy. Brivaracetam has good efficacy and tolerability as adjunctive therapy in the treatment of focal (partial onset) seizures in patients 16 years of age and older. Brivaracetam is safe for prolonged use in patients with epilepsy and in children with epilepsy. The most common adverse events with brivaracetam are related to central nervous system and include fatigue, dizziness, and somnolence; these may improve or resolve during treatment. A consensus was sought for the use of brivaracetam in epilepsy management in routine neurology practices Brivaracetam is a safer AED with lesser behavioral AEs, lack of cognitive impairment, and no clinically relevant drug-drug interactions or dose adjustment for renal patients.

Ions of structure R2N[N(O)NO]- and their alkylation products have seen increasing use as nitric oxide (NO)-generating agents for biomedical research applications. Here we show that such diazeniumdiolate anions can readily displace halide from a variety of electrophilic aza- or nitroaromatic substrates to form O2-arylated derivatives of structure R2N−N(O)N−OAr. The site of arylation and the cis arrangement of the oxygens were confirmed by X-ray crystallography. Displacement by various nucleophiles showed R2N[N(O)NO]- to be a reasonably good leaving group, with rate constants for displacement by hydroxide, methoxide, and isopropylamine that were between those of chloride and fluoride in the SNAr reactions we surveyed. The Meisenheimer intermediate could be spectrally observed. These O2-aryl diazeniumdiolates proved capable of reacting with the nucleophilic sulfur of the HIV-1 p7 nucleocapsid protein's zinc finger assembly to eject the zinc, disrupting a structural motif critical to viral replication and sug...

Reaction of nitric oxide with N-benzylidene-4-methoxyaniline produced 4-methoxybenzenediazonium nitrate and benzaldehyde. This may represent an example of the electrophilic reaction of NO with a double bond.

3-Alkyl-1,2,3-oxadiazolinium ions 1 have been proposed as reactive intermediates in the activation of (2-hydroxyethyl)nitrosamines. The reaction of 3-methyl-1,2,3-oxadiazolinium tosylate (1a), 2-ethyl-1-methoxy-2-phenyldiazenium tetrafluoroborate (3), and 3-phenyl-1,2,3-oxadiazolinium triflate (1b) with thiols was investigated to determine the behavior of these compounds toward typical "cellular nucleophiles". Each of these substances oxidizes benzenethiol to diphenyl disulfide. The reaction in aqueous buffer at pH 7.4 is rapid. Reaction of 1b with benzenethiol gives, in addition to the disulfide, benzene, biphenyl, azobenzene, diphenyl sulfide, aniline, and glycolaldehyde. Similar products are obtained from 3. Phenyldiazene is postulated as an intermediate in this process, and its generation from phenyldiazoformate in the presence of benzenethiol gives similar products. Diazenes are presumed to arise by proton abstraction from the CH adjacent to N. The kinetics of reaction of 1a with N-acetylcysteine to give the corresponding disulfide show first order dependence on each reactant and base catalysis. The data from these model chemical experiments suggest that 1,2,3-oxadiazolinium ions could react with abundant thiols in cells to lead to either their detoxification or radical processes emanating from diazenes. The occurrence of thiol-oxadiazolinium ion redox transformations could modulate the alkylation chemistry of these substances as well.

Ions of structure R2N[N(O)NO]- and their alkylation products have seen increasing use as nitric oxide (NO)-generating agents for biomedical research applications. Here we show that such diazeniumdiolate anions can readily displace halide from a variety of electrophilic aza- or nitroaromatic substrates to form O2-arylated derivatives of structure R2N−N(O)N−OAr. The site of arylation and the cis arrangement of the oxygens were confirmed by X-ray crystallography. Displacement by various nucleophiles showed R2N[N(O)NO]- to be a reasonably good leaving group, with rate constants for displacement by hydroxide, methoxide, and isopropylamine that were between those of chloride and fluoride in the SNAr reactions we surveyed. The Meisenheimer intermediate could be spectrally observed. These O2-aryl diazeniumdiolates proved capable of reacting with the nucleophilic sulfur of the HIV-1 p7 nucleocapsid protein's zinc finger assembly to eject the zinc, disrupting a structural motif critical to viral replication and sug...

Xinhua Ji1, Ajai Pal2, Ravi Kalathur1, Xun Hu2, Yijun Gu1,2, Joseph E Saavedra3, Gregory S Buzard3, Aloka Srinivasan4, Larry K Keefer4, Shivendra V Singh21Macromolecular Crystallography Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA; 2Department of Pharmacology and University of Pittsburgh Cancer Institute, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213, USA; 3Basic Research Program, SAIC-Frederick Inc., Frederick, MD 21702, USA; 4Laboratory of Comparative Carcinogenesis, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USAAbstract: Glutathione S-transferase (GST) is a superfamily of detoxification enzymes, represented by GSTα, GSTµ, GSTπ, etc. GSTα is the predominant isoform of GST in human liver, playing important roles for our well being. GSTπ is overexpressed in many forms of cancer, thus presenting an opportunity for selective targeting of cancer cells. Our structure-based design of prodrugs intended to release cytotoxic levels of nitric oxide in GSTπ-overexpressing cancer cells yielded PABA/NO, which exhibited anticancer activity both in vitro and in vivo with a potency similar to that of cisplatin. Here, we present the details on structural modification, molecular modeling, and enzymatic characterization for the design of PABA/NO. The design was efficient because it was on the basis of the reaction mechanism and the structures of related GST isozymes at both the ground state and the transition state. The ground-state structures outlined the shape and property of the substrate-binding site in different isozymes, and the structural information at the transition-state indicated distinct conformations of the Meisenheimer complex of prodrugs in the active site of different isozymes, providing guidance for the modifications of the molecular structure of the prodrug molecules. Two key alterations of a GSTα-selective compound led to the GSTπ-selective PABA/NO.Keywords: structure-based, drug design, anticancer, prodrug, PABA/NO

Endogenously generated nitric oxide (NO) mediates a host of important physiological functions, playing roles in the vascular, immunological, and neurological systems. As a result, exogenous agents that release NO have become important therapeutic interventions and research tools. O 2 -Vinyl 1-(pyrrolidin-1-yl)diazen-1-ium-1,2-diolate (V-PYRRO/NO) is a prodrug designed with the hypothesis that it might release nitric oxide via epoxidation of the vinyl group by cytochrome P450, followed by enzymatic and/or spontaneous epoxide hydration to release the ultimate NO-donating moiety, 1-(pyrrolidin-1-yl)diazen-1-ium-1,2-diolate (PYRRO/NO) ion. In this study, we investigated this hypothetical activation mechanism quantitatively for V-PYRRO/NO using cDNA-expressed human cytochrome P450 (CYP)2E1. Incubation with CYP2E1 and an NADPH-regenerating system resulted in a time-dependent decomposition of V-PYRRO/NO, with a turnover rate of 2.0 nmol/min/pmol CYP2E1. Nitrate and nitrite were detected in high yield as metabolites of NO. The predicted organic metabolites pyrrolidine and glycolaldehyde were also detected in near-quantitative yields. The enzymatic decomposition of V-PYRRO/NO was also catalyzed, albeit at lower rates, by CYP2A6 and CYP2B6. We conclude that the initial step in the metabolism of V-PYRRO/NO to NO in the liver is catalyzed efficiently but not exclusively by the alcohol-inducible form of cytochrome P450 (CYP2E1). The results confirm the proposed activation mechanism involving enzymatic oxidation of the vinyl group in V-PYRRO/NO followed by epoxide hydration and hydrolytic decomposition of the resulting PYRRO/NO ion to generate nitric oxide.

We have found O(2)-substituted diazeniumdiolates, compounds of structure R(2)N-N(O)=NOR' that are under development for various possible pharmaceutical uses, to be rather photosensitive. With R = ethyl and R' = methyl, benzyl, or 2-nitrobenzyl, the observed product distributions suggest that two primary pathways are operative. A minor pathway involves the extrusion of nitrous oxide (N(2)O) with simultaneous generation of R(2)N(*) and R'O(*), which may then form amines, aldehydes, and alcohols. The major reaction pathway is an interesting photochemical cleavage of the N=N bond to form a nitrosamine (R(2)NN=O) and an oxygen-substituted nitrene (R'ON). The intermediacy of the O-nitrene was inferred from the production of abundant oxime, via rearrangement of the O-nitrene to a C-nitroso compound (R'ON --O=NR'), and subsequent tautomerization to the more stable oxime. Involvement of the O-nitrene was confirmed by trapping with 2,3-dimethyl-2-butene to form the aziridine and with oxygen to generate the nitrate ester. 2-Nitro substitution on the benzyl derivative had surprisingly little effect on the reaction course. For each compound examined, minor amounts of nitric oxide (NO), presumably produced by secondary photolysis of the nitrosamine, were observed. Time-resolved infrared experiments provided additional support for the above reaction pathways and confirmed that the nitrosamine is a primary photoproduct. We have also found that the relative contributions of the reaction pathways can be altered in certain derivatives. For example, when R' = 2,4-dinitrophenyl, the contribution of the nitrosamine/O-nitrene-forming pathway was diminished. Pharmacological implications of these results are discussed.

The involvement of nitric oxide in numerous biological functions has led to the intense study of nitric oxide (NO) generation by the nitric oxide synthases (NOS) responsible. In addition to NO, nitric oxide synthases produceNG-hydroxy- l -arginine, superoxide anion and, indirectly, NOx species such as peroxynitrite and, possibly, nitrous oxide (N2O). Consequently, the interactions ofNG-hydroxy- l -arginine with NO and other oxides of nitrogen (NOx) are of considerable interest.NG-Hydroxy- l -arginine and other monosubstituted hydroxyguanidines react with aqueous aerobic NO, peroxynitrite, and various NOx and nitrosating agents to form compounds that subsequently release NO and N2O. Spectrometric data indicate that the nitrosation product ofNG-hydroxy- l -arginine is of the sameN-nitroso-N-hydroxy/diazeniumdiolate (formerly “NONOate”) structure as previously found for the nitrosation products of other model hydroxyguanidines. These decompose in aqueous solution in a pH-dependent manner to yield mainly NO and ureas at low pH, N2O and cyanamides at basic pH, and what appear to be primary nitrosamines/nitrosoimines. Studies on purified iNOS using a mass spectrometer with a gas-permeable membrane inlet identified both NO and N2O (or15NO and15N15NO with15N-labeled l -arginine as substrate) as products of NOS activity. These experiments suggest that much more NO than N2O is produced under the conditions studied and that N2O formation can be rationalized via the reaction of NOx species withNG-hydroxy- l -arginine.

N-Hydroxyguanidines can be nitrosatively converted to zwitterionic diazeniumdiolates of crystallographically-confirmed structure H2N+C[NHR][N(O)NO]–, whose hydrolytic dissociation at physiological pH leads to both NO and N2O; the results appear to account for the formation of the ‘potential intercellular nitric oxide carrier’ produced on exposing NG- hydroxy-L-arginine (a metabolic intermediate in mammalian NO biosynthesis) to aerobic NO.

We show here that a azoxyalkene is produced by the electrocyclic ring opening of a novel dihydrodiazete N-oxide produced from the isomerization of a N-nitrosoaziridinium ion, and we discuss our experimental and theoretical probing of this process