ABSTRACT
N-Nitrosamines (also referred to as nitrosamines) are a class of substances, many of which are highly potent mutagenic agents which have been classified as probable human carcinogens. Nitrosamine impurities have been a concern within the pharmaceutical industry and by regulatory authorities worldwide since June 2018, when regulators were informed of the presence of N-nitrosodimethylamine (NDMA) in the angiotensin-II receptor blocker (ARB) medicine, valsartan. Since that time, regulatory authorities have collaborated to share information and knowledge on issues related to nitrosamines with a goal of promoting convergence on technical issues and reducing and mitigating patient exposure to harmful nitrosamine impurities in human drug products. This paper shares current scientific information from a quality perspective on risk factors and potential root causes for nitrosamine impurities, as well as recommendations for risk mitigation and control strategies.
Subject(s)
Nitrosamines , Humans , Angiotensin Receptor Antagonists , Angiotensin-Converting Enzyme Inhibitors , Risk Factors , Pharmaceutical PreparationsABSTRACT
BACKGROUND: Flow cytometry (FCM) is a powerful single-cell based measurement method to ascertain multidimensional optical properties of millions of cells. FCM is widely used in medical diagnostics and health research. There is also a broad range of applications in the analysis of complex microbial communities. The main concern in microbial community analyses is to track the dynamics of microbial subcommunities. So far, this can be achieved with the help of time-consuming manual clustering procedures that require extensive user-dependent input. In addition, several tools have recently been developed by using different approaches which, however, focus mainly on the clustering of medical FCM data or of microbial samples with a well-known background, while much less work has been done on high-throughput, online algorithms for two-channel FCM. RESULTS: We bridge this gap with flowEMMi, a model-based clustering tool based on multivariate Gaussian mixture models with subsampling and foreground/background separation. These extensions provide a fast and accurate identification of cell clusters in FCM data, in particular for microbial community FCM data that are often affected by irrelevant information like technical noise, beads or cell debris. flowEMMi outperforms other available tools with regard to running time and information content of the clustering results and provides near-online results and optional heuristics to reduce the running-time further. CONCLUSIONS: flowEMMi is a useful tool for the automated cluster analysis of microbial FCM data. It overcomes the user-dependent and time-consuming manual clustering procedure and provides consistent results with ancillary information and statistical proof.
Subject(s)
Algorithms , Flow Cytometry/methods , Microbiota , Models, Theoretical , Automation , Benchmarking , Cluster Analysis , Time FactorsABSTRACT
Inhibition of human histone deacetylases (HDACs) has emerged as a novel concept in the chemotherapeutic treatment of cancer. Two chemical entities, SAHA (ZOLINZA, Merck) and romidepsin (Istodax, Celgene) have been recently approved by the FDA as first-in-class drugs against cutaneous T-cell lymphoma. Clinical use of these drugs revealed several side effects including gastro-intestinal symptoms, fatigue, thrombocytopenia, thrombosis. Romidepsin is associated with an yet unresolved cardiotoxicity issue. A general hypothesis for the diminishment of unwanted adverse effects and an improved therapeutical window suggests the development of more isotype selective inhibitors. In this study the first time HDAC inhibitors with perfluorinated spacers between the zinc chelating moiety and the aromatic capping group were synthesized and tested against representatives of HDAC classes I, IIa and IIb. Competitive binding assays and a combined approach by using blind docking and molecular dynamics support binding of the perfluorinated analogs of SAHA to the active site of the HDAC-like amidohydrolase from Bordetella/Alcaligenes and presumably also to human HDACs. In contrast to the alkyl spacer of SAHA and derivatives, the perfluorinated alkyl spacer seems to contribute to or facilitate the induction of selectivity for class II, particularly class IIa, HDACs even though the overall potency of the perfluorinated SAHA analogs in this study against human HDACs remained still rather moderate in the micromolar range.
Subject(s)
Coordination Complexes/chemical synthesis , Diamide/chemistry , Diamide/pharmacology , Histone Deacetylase Inhibitors/chemical synthesis , Histone Deacetylase Inhibitors/pharmacology , Histone Deacetylases/chemistry , Binding Sites , Catalytic Domain , Cell Line , Coordination Complexes/chemistry , Coordination Complexes/pharmacology , Depsipeptides/pharmacology , Diamide/chemical synthesis , Enzyme Activation/drug effects , Histone Deacetylase Inhibitors/chemistry , Histone Deacetylases/genetics , Histone Deacetylases/metabolism , Humans , Hydroxamic Acids/pharmacology , Molecular Dynamics Simulation , Recombinant Proteins/antagonists & inhibitors , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Vorinostat , Zinc/chemistryABSTRACT
Cytosolic phospholipase A(2)alpha (cPLA(2)alpha) and fatty acid amide hydrolase (FAAH) are enzymes, which have emerged as attractive targets for the development of analgetic and anti-inflammatory drugs. We recently reported that 1-[3-(4-octylphenoxy)-2-oxopropyl]indole-5-carboxylic acid (10) and related compounds are inhibitors of cPLA(2)alpha. Since cPLA(2)alpha and FAAH possess several common structural features, we now screened this substance series together with some new derivatives for FAAH inhibition. Some of the assayed compounds proved to be selective cPLA(2)alpha inhibitors, while others showed high FAAH and moderate cPLA(2)alpha inhibitory potency. Furthermore, several derivatives were favorably active against both enzymes and, therefore, could represent agents, which have improved analgetic and anti-inflammatory qualities in comparison with selective cPLA(2)alpha and FAAH inhibitors.
Subject(s)
Amidohydrolases/antagonists & inhibitors , Enzyme Inhibitors/pharmacology , Group IV Phospholipases A2/antagonists & inhibitors , Heterocyclic Compounds/pharmacology , Indoles/pharmacology , Amidohydrolases/metabolism , Animals , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Group IV Phospholipases A2/isolation & purification , Group IV Phospholipases A2/metabolism , Heterocyclic Compounds/chemical synthesis , Heterocyclic Compounds/chemistry , Humans , Indoles/chemical synthesis , Indoles/chemistry , Molecular Conformation , Rats , Stereoisomerism , Structure-Activity RelationshipABSTRACT
The synthesis and structure-activity relationship study of a series of 1-indol-1-yl-3-phenoxypropan-2-one inhibitors of cytosolic phospholipase A(2)alpha (cPLA(2)alpha) are described. The compounds were evaluated in a vesicle assay with isolated cPLA(2)alpha and in cellular assays with intact human platelets. Systematic variation led to 3-methylhydrogen 1-[3-(4-decyloxyphenoxy)-2-oxopropyl]indole-3,5-dicarboxylate (57), which revealed the highest activity against the isolated enzyme. With an IC(50) value of 4.3 nM in this assay, it is one of the most potent in vitro cPLA(2)alpha inhibitors known today.
