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Chemical Derivatization Enables MALDI-TOF-Based High-Throughput Screening for Microbial Trimethylamine (TMA)-Lyase Inhibitors.
Winter, Martin; Bretschneider, Tom; Thamm, Sven; Kleiner, Carola; Grabowski, Daniel; Chandler, Sarah; Ries, Robert; Kley, Jörg T; Fowler, Danielle; Bartlett, Christina; Binetti, Ralph; Broadwater, John; Luippold, Andreas H; Bischoff, Daniel; Büttner, Frank H.
Afiliación
  • Winter M; 1 Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
  • Bretschneider T; 1 Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
  • Thamm S; 1 Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
  • Kleiner C; 1 Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
  • Grabowski D; 1 Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
  • Chandler S; 1 Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
  • Ries R; 1 Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
  • Kley JT; 2 Medicinal Chemistry, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
  • Fowler D; 3 Cardiometabolic Diseases Research, Boehringer Ingelheim Pharmaceuticals, Ridgefield, CT, USA.
  • Bartlett C; 3 Cardiometabolic Diseases Research, Boehringer Ingelheim Pharmaceuticals, Ridgefield, CT, USA.
  • Binetti R; 4 Cancer Immunology & Immune Modulation, Boehringer Ingelheim Pharmaceuticals, Ridgefield, CT, USA.
  • Broadwater J; 3 Cardiometabolic Diseases Research, Boehringer Ingelheim Pharmaceuticals, Ridgefield, CT, USA.
  • Luippold AH; 1 Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
  • Bischoff D; 1 Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
  • Büttner FH; 1 Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riß, Germany.
SLAS Discov ; 24(7): 766-777, 2019 08.
Article en En | MEDLINE | ID: mdl-31059309
Microbial-dependent trimethylamine (TMA) generation from dietary precursors such as choline was recently linked to cardiovascular diseases (CVDs) as well as chronic kidney disease (CKD). Inhibition of TMA-generating enzymes in gut bacteria would be an innovative approach to treat these diseases. The potential to accurately quantify secreted TMA levels highlights the capacity of mass spectrometry (MS) for tracking microbial TMA-lyase activity. However, high-throughput screening (HTS) by conventional MS instrumentation is hampered by limited sample throughput. Recent advancement in liquid handling and instrumentation of matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS provides an HTS-compatible MS technology. The deciphering of enzymatic reactions using this label-free readout has been successfully applied but has thus far been limited to peptide/protein-centric activity assays. Here, we demonstrate the versatile applicability of MALDI-TOF by tracking a small molecule within a highly complex sample background. The key to success for this concept was chemical derivatization of the target molecule enabling quantitative assessment of microbial TMA formation. Further, its potential was demonstrated in a side-by-side comparison to RapidFire-MS in a primary screen and subsequent dose-response experiments. Overall, the established assay enables the screening for microbial TMA-lyase inhibitors and serves as a proof of concept for the applicability of MALDI-TOF for demanding assay concepts per se.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción / Inhibidores Enzimáticos / Descubrimiento de Drogas / Ensayos Analíticos de Alto Rendimiento / Liasas / Metilaminas Tipo de estudio: Diagnostic_studies / Screening_studies Límite: Humans Idioma: En Revista: SLAS Discov Año: 2019 Tipo del documento: Article País de afiliación: Alemania Pais de publicación: Estados Unidos
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción / Inhibidores Enzimáticos / Descubrimiento de Drogas / Ensayos Analíticos de Alto Rendimiento / Liasas / Metilaminas Tipo de estudio: Diagnostic_studies / Screening_studies Límite: Humans Idioma: En Revista: SLAS Discov Año: 2019 Tipo del documento: Article País de afiliación: Alemania Pais de publicación: Estados Unidos