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Abstract Late‐stage functionalization (LSF) of drug molecules is an approach to generate modified molecules that retain functional groups present in the active drugs. Here, we report a study that seeks to characterize the potential value of high‐throughput desorption electrospray ionization mass spectrometry (HT DESI‐MS) for small‐scale rapid LSF. In conventional route screening, HT‐based DESI‐MS provides contactless, rapid analysis, reliable and reproducible data, minimal sample requirement, and exceptional tolerance to high salt concentrations. Ezetimibe (E), an established hypertension drug, is targeted for modification by LSF. C−H alkenylation and azo‐click reactions are utilized to explore this approach to synthesis and analytical characterization. The effect of choice of reactant, stoichiometry, catalyst, and solvent are studied for both reactions using high throughput DESI‐MS experiments. Optimum conditions for the formation of LSF products are established with identification by tandem mass spectrometry (MS/MS).
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Desorption Electrospray Ionization Mass Spectrometry Assay for Label‐Free Characterization of SULT2B1b Enzyme Kinetics
Abstract The sulfotransferase (SULT) 2B1b, which catalyzes the sulfonation of 3β‐hydroxysteroids, has been identified as a potential target for prostate cancer treatment. However, a major limitation for SULT2B1b‐targeted drug discovery is the lack of robust assays compatible with high‐throughput screening and inconsistency in reported kinetic data. For this reason, we developed a novel label‐free assay based on high‐throughput (>1 Hz) desorption electrospray ionization mass spectrometry (DESI‐MS) for the direct quantitation of the sulfoconjugated product (CV<10 %; <1 ng analyte). The performance of this DESI‐based assay was compared against a new fluorometric coupled‐enzyme method that we also developed. Both methodologies provided consistent kinetic data for the reaction of SULT2B1b with its major substrates, indicating the affinity trend pregnenolone>DHEA>cholesterol, for both the phospho‐mimetic and wild‐type SULT2B1b forms. The novel DESI‐MS assay developed here is likely generalizable to other drug discovery efforts and is particularly promising for identification of SULT2B1b inhibitors with potential as prostate cancer therapeutics.
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- Award ID(s):
- 1905087
- PAR ID:
- 10446438
- Publisher / Repository:
- Wiley Blackwell (John Wiley & Sons)
- Date Published:
- Journal Name:
- ChemMedChem
- Volume:
- 17
- Issue:
- 9
- ISSN:
- 1860-7179
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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