Biochemical combinatorial techniques such as phage display, RNA display and oligonucleotide aptamers have proven to be reliable methods for generation of ligands to protein targets. Adapting these techniques to small synthetic molecules has been a long-sought goal. We report the synthesis and interrogation of an 800-million-member DNA-encoded library in which small molecules are covalently attached to an encoding oligonucleotide. The library was assembled by a combination of chemical and enzymatic synthesis, and interrogated by affinity selection. We describe methods for the selection and deconvolution of the chemical display library, and the discovery of inhibitors for two enzymes: Aurora A kinase and p38 MAP kinase.

The metalloprotease ADAMTS-5 is considered a potential target for the treatment of osteoarthritis. To identify selective inhibitors of ADAMTS-5, we employed encoded library technology (ELT), which enables affinity selection of small molecule binders from complex mixtures by DNA tagging. Selection of ADAMTS-5 against a four-billion member ELT library led to a novel inhibitor scaffold not containing a classical zinc-binding functionality. One exemplar, (R)-N-((1-(4-(but-3-en-1-ylamino)-6-(((2-(thiophen-2-yl)thiazol-4-yl)methyl)amino)-1,3,5-triazin-2-yl)pyrrolidin-2-yl)methyl)-4-propylbenzenesulfonamide (8), inhibited ADAMTS-5 with IC(50) = 30 nM, showing >50-fold selectivity against ADAMTS-4 and >1000-fold selectivity against ADAMTS-1, ADAMTS-13, MMP-13, and TACE. Extensive SAR studies showed that potency and physicochemical properties of the scaffold could be further improved. Furthermore, in a human osteoarthritis cartilage explant study, compounds 8 and 15f inhibited aggrecanase-mediated (374)ARGS neoepitope release from aggrecan and glycosaminoglycan in response to IL-1β/OSM stimulation. This study provides the first small molecule evidence for the critical role of ADAMTS-5 in human cartilage degradation.

Tuberculosis (TB) is one of the world's oldest and deadliest diseases, killing a person every 20 s. InhA, the enoyl-ACP reductase from Mycobacterium tuberculosis, is the target of the frontline antitubercular drug isoniazid (INH). Compounds that directly target InhA and do not require activation by mycobacterial catalase peroxidase KatG are promising candidates for treating infections caused by INH resistant strains. The application of the encoded library technology (ELT) to the discovery of direct InhA inhibitors yielded compound 7 endowed with good enzymatic potency but with low antitubercular potency. This work reports the hit identification, the selected strategy for potency optimization, the structure−activity relationships of a hundred analogues synthesized, and the results of the in vivo efficacy studies performed with the lead compound 65.

ABSTRACT:The aggrecan degrading metalloprotease ADAMTS-4 has been identified as a novel therapeutic target for osteoarthritis. Here, we use DNA-encoded Library Technology (ELT) to identify novel ADAMTS-4 inhibitors from a DNAencoded triazine library by affinity selection. Structure−activity relationship studies based on the selection information led to the identification of potent and highly selective inhibitors. For example, 4-(((4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)-6-(((4-methylpiperazin-1-yl)methyl)amino)-1,3,5-triazin-2-yl)amino)methyl)-N-ethyl-N-(m-tolyl)benzamide has IC 50 of 10 nM against ADAMTS-4, with >1000-fold selectivity over ADAMT-5, MMP-13, TACE, and ADAMTS-13. These inhibitors have no obvious zinc ligand functionality.

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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.

Undecaprenyl pyrophosphate synthase (UppS) is an essential enzyme in bacterial cell wall synthesis. Here we report the discovery of Staphylococcus aureus UppS inhibitors from an Encoded Library Technology screen and demonstrate binding to the hydrophobic substrate site through cocrystallography studies. The use of bacterial strains with regulated uppS expression and inhibitor resistant mutant studies confirmed that the whole cell activity was the result of UppS inhibition, validating UppS as a druggable antibacterial target.

In the version of this article initially published, the IC 50 values in the table in Figure 3c were listed as nM instead of µM. The error has been corrected in the HTML and PDF versions of the article. 772volume 5 number 10 october 2009 nature chemical biology e r r ata