New Insights into the Mechanism of Action of the Thienopyrimidine Antitubercular Prodrug TP053.

The thienopyrimidine TP053 is an antitubercular prodrug active against both replicating and nonreplicating Mycobacterium tuberculosis (M. tuberculosis) cells, which requires activation by the mycothiol-dependent nitroreductase Mrx2. The investigation of the mechanism of action of TP053 revealed that Mrx2 releases nitric oxide from this drug both in the enzyme assays with purified Mrx2 and in mycobacterial cultures, which can explain its activity against nonreplicating bacilli, similar to pretomanid activated by the nitroreductase Ddn. In addition, we identified a highly reactive metabolite, 2-(4-mercapto-6-(methylamino)-2-phenylpyrimidin-5-yl)ethan-1-ol, which can contribute to the antimycobacterial effects on replicating cells as well as on nonreplicating cells. In summary, we explain the mechanism of action of TP053 on both replicating and nonreplicating M. tuberculosis and report a novel activity for Mrx2, which in addition to Ddn, represents another example of nitroreductase releasing nitric oxide from its substrate. These findings are particularly relevant in the context of drugs targeting nonreplicating M. tuberculosis, which is shown to be killed by increased levels of nitric oxide.

Integrated serum proteins and fatty acids analysis for putative biomarker discovery in inflammatory bowel disease.

The diagnosis and management of Inflammatory bowel disease (IBD), including Crohn's disease (CD) and ulcerative colitis (UC), is still challenging. There is no definitive gold standard diagnostic test, which is made on patient history and with endoscopic and histological findings. This study analyzed serum proteins and fatty acids using mass spectrometry-based techniques. Quantitation of serum proteins was performed by depleting 14 high-abundance proteins, followed by tryptic digestion and LC-MS analysis, while fatty acids were analyzed using GC-MS. Bioinformatic tools were used to identify several new potential biomarkers for an early and non-invasive diagnosis of IBD, and to differentiate CD from UC. Moreover, the diagnostic power of the MS-identified biomarkers was also corroborated by Western Blot and ELISA assays. Hence, we identified the biological functions and pathways involved in the various subsets of IBD. Coagulation, fibrinolysis and acute phase response processes were found to be strongly involved in the condition. The involvement of several fatty acids, such as anti-inflammatory mediators, was also identified. Finally, proteomic and lipidomic data were integrated by using combinatorial and multivariate analyses to discover new combined biomarkers and to study the molecular pathways involved in IBD.

Trichostatin A alters cytoskeleton and energy metabolism of pancreatic adenocarcinoma cells: An in depth proteomic study.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal of all human cancers with a high mortality rate. Resistance to conventional treatments and chemotherapeutics is a typical feature of PDAC. To investigate the causes of drug resistance it is essential to deeply investigate the mechanism of action of chemotherapeutics. In this study, we performed an in depth shotgun proteomic approach using the label-free proteomic SWATH-MS analysis to investigate novel insights of the mechanism of action of the histone deacetylase (HDAC) inhibitor trichostatin A (TSA) in PDAC cells. This proteomic analysis in PaCa44 cells and data elaboration of TSA-regulated proteins by bioinformatics showed an overall up-regulation of cytokeratins and other proteins related to the cytoskeleton organization, keratinization, and apoptotic cell death. On the contrary, a large amount of the down-regulated proteins by TSA treatment belongs to the cellular energetic metabolism and to the machinery of protein synthesis, such as ribosomal proteins, determining synergistic cell growth inhibition by the combined treatment of TSA and the glycolytic inhibitor 2-deoxy-d-glucose in a panel of PDAC cell lines. Data are available via ProteomeXchange with identifier PXD007801.