Acoustic Mist Ionization-Mass Spectrometry: A Comparison to Conventional High-Throughput Screening and Compound Profiling Platforms.

Drug discovery usually begins with a high-throughput screen (HTS) of thousands to millions of molecules to identify starting points for medicinal chemistry. Conventional HTS platforms require expensive reagents and typically have complex assay formats. HTS platforms based on radioactivity are expensive, both in terms of reagent cost and disposal. Furthermore, nonspecific interferences common to these technologies result in an extensive attrition of hits during validation experiments. Mass spectrometry (MS) is a highly selective, label-free technology that can quantify multiple analytes in a single experiment. However, most commercial MS platforms typically involve a separation or cleanup prior to analysis and are too slow for large-scale screening campaigns. Recently, an MS platform (AMI-MS) was introduced that uses acoustically generated droplets to deliver analyte molecules directly from microtiter plates into the mass spectrometer at subsecond per well sampling rates. Here, we demonstrate the application of AMI-MS by developing an HTS-compatible assay that measures the inhibition of histone acetyltransferase activity. Real-time kinetic measurements from a single well were used to determine enzyme Km and Vmax values. We compare the AMI-MS readout with conventional platforms in single-shot screening and multipoint profiling modes. The AMI-MS assay identified 86% of hits previously identified, with a pIC50 ≥ 5.0, in a scintillation proximity assay (SPA) HTS at a lower hit rate and with a significantly reduced cost per well compared to the SPA-based readout. Furthermore, pIC50s, as measured by AMI-MS, showed a good correlation with values generated by RapidFire-MS. AMI-MS has the potential to provide significant improvements to high-throughput bioassays.

New IDH1 mutant inhibitors for treatment of acute myeloid leukemia.

Neomorphic mutations in isocitrate dehydrogenase 1 (IDH1) are driver mutations in acute myeloid leukemia (AML) and other cancers. We report the development of new allosteric inhibitors of mutant IDH1. Crystallographic and biochemical results demonstrated that compounds of this chemical series bind to an allosteric site and lock the enzyme in a catalytically inactive conformation, thereby enabling inhibition of different clinically relevant IDH1 mutants. Treatment of IDH1 mutant primary AML cells uniformly led to a decrease in intracellular 2-HG, abrogation of the myeloid differentiation block and induction of granulocytic differentiation at the level of leukemic blasts and more immature stem-like cells, in vitro and in vivo. Molecularly, treatment with the inhibitors led to a reversal of the DNA cytosine hypermethylation patterns caused by mutant IDH1 in the cells of individuals with AML. Our study provides proof of concept for the molecular and biological activity of novel allosteric inhibitors for targeting different mutant forms of IDH1 in leukemia.

Discovery and SAR Study of Boronic Acid-Based Selective PDE3B Inhibitors from a Novel DNA-Encoded Library.

Human genetic evidence shows that PDE3B is associated with metabolic and dyslipidemia phenotypes. A number of PDE3 family selective inhibitors have been approved by the FDA for various indications; however, given the undesirable proarrhythmic effects in the heart, selectivity for PDE3B inhibition over closely related family members (such as PDE3A; 48% identity) is a critical consideration for development of PDE3B therapeutics. Selectivity for PDE3B over PDE3A may be achieved in a variety of ways, including properties intrinsic to the compound or tissue-selective targeting. The high (>95%) active site homology between PDE3A and B represents a massive obstacle for obtaining selectivity at the active site; however, utilization of libraries with high molecular diversity in high throughput screens may uncover selective chemical matter. Herein, we employed a DNA-encoded library screen to identify PDE3B-selective inhibitors and identified potent and selective boronic acid compounds bound at the active site.

Thiol-based angiotensin-converting enzyme 2 inhibitors: P1 modifications for the exploration of the S1 subsite.

Screening of a metalloprotease library led to the identification of a thiol-based dual ACE/NEP inhibitor as a potent ACE2 inhibitor. Modifications of the P(1) benzyl moiety led to improvements in ACE2 potency as well as to increased selectivity versus ACE and NEP.

Author Correction: Prioritizing multiple therapeutic targets in parallel using automated DNA-encoded library screening.

This corrects the article DOI: 10.1038/ncomms16081.

Prioritizing multiple therapeutic targets in parallel using automated DNA-encoded library screening.

The identification and prioritization of chemically tractable therapeutic targets is a significant challenge in the discovery of new medicines. We have developed a novel method that rapidly screens multiple proteins in parallel using DNA-encoded library technology (ELT). Initial efforts were focused on the efficient discovery of antibacterial leads against 119 targets from Acinetobacter baumannii and Staphylococcus aureus. The success of this effort led to the hypothesis that the relative number of ELT binders alone could be used to assess the ligandability of large sets of proteins. This concept was further explored by screening 42 targets from Mycobacterium tuberculosis. Active chemical series for six targets from our initial effort as well as three chemotypes for DHFR from M. tuberculosis are reported. The findings demonstrate that parallel ELT selections can be used to assess ligandability and highlight opportunities for successful lead and tool discovery.