Identification and evaluation of small-molecule inhibitors against the dNTPase SAMHD1 via a comprehensive screening funnel.

SAMHD1 is a dNTP triphosphohydrolase governing nucleotide pool homeostasis and can detoxify chemotherapy metabolites controlling their clinical responses. To understand SAMHD1 biology and investigate the potential of targeting SAMHD1 as neoadjuvant to current chemotherapies, we set out to discover selective small-molecule inhibitors. Here, we report a discovery pipeline encompassing a biochemical screening campaign and a set of complementary biochemical, biophysical, and cell-based readouts for rigorous characterization of the screen output. The identified small molecules, TH6342 and analogs, accompanied by inactive control TH7126, demonstrated specific, low µM potency against both physiological and oncology-drug-derived substrates. By coupling kinetic studies with thermal shift assays, we reveal the inhibitory mechanism of TH6342 and analogs, which engage pre-tetrameric SAMHD1 and deter oligomerization and allosteric activation without occupying nucleotide-binding pockets. Altogether, our study diversifies inhibitory modes against SAMHD1, and the discovery pipeline reported herein represents a thorough framework for future SAMHD1 inhibitor development.

Author Correction: Targeted NUDT5 inhibitors block hormone signaling in breast cancer cells.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Targeted NUDT5 inhibitors block hormone signaling in breast cancer cells.

With a diverse network of substrates, NUDIX hydrolases have emerged as a key family of nucleotide-metabolizing enzymes. NUDT5 (also called NUDIX5) has been implicated in ADP-ribose and 8-oxo-guanine metabolism and was recently identified as a rheostat of hormone-dependent gene regulation and proliferation in breast cancer cells. Here, we further elucidate the physiological relevance of known NUDT5 substrates and underscore the biological requirement for NUDT5 in gene regulation and proliferation of breast cancer cells. We confirm the involvement of NUDT5 in ADP-ribose metabolism and dissociate a relationship to oxidized nucleotide sanitation. Furthermore, we identify potent NUDT5 inhibitors, which are optimized to promote maximal NUDT5 cellular target engagement by CETSA. Lead compound, TH5427, blocks progestin-dependent, PAR-derived nuclear ATP synthesis and subsequent chromatin remodeling, gene regulation and proliferation in breast cancer cells. We herein present TH5427 as a promising, targeted inhibitor that can be used to further study NUDT5 activity and ADP-ribose metabolism.

Novel spirocyclic systems via multicomponent aza-Diels-Alder reaction.

Here we present a two-step diastereoselective methodology building on a multicomponent aza-Diels-Alder reaction. Using previously unexplored cyclic ketones, heterocyclic amines and cyclopentadiene derivatives, we obtained novel spiro-heterocyclic frameworks at the interphase between "drug-like" molecules and natural products.

Tetrahydrobenzothiophene carboxamides: Beyond the kinase domain and into the fatty acid realm.

A series of tetrahydrobenzothiophene carboxamides, inspired by structural features present in kinase and SCD1 inhibitors, are presented here. Prototype compound 8 (MMDD13) modulates fatty acid elongase and desaturase indexes, lipid accumulation, while preserving kinase inhibitory activity. This chemotype represents a stepping stone towards chemical probes to study the consequences of lipid metabolism modulation through non-redundant pathways.

Inhibitory Kappa B Kinase α (IKKα) Inhibitors That Recapitulate Their Selectivity in Cells against Isoform-Related Biomarkers.

IKKß plays a central role in the canonical NF-kB pathway, which has been extensively characterized. The role of IKKα in the noncanonical NF-kB pathway, and indeed in the canonical pathway as a complex with IKKß, is less well understood. One major reason for this is the absence of chemical tools designed as selective inhibitors for IKKα over IKKß. Herein, we report for the first time a series of novel, potent, and selective inhibitors of IKKα. We demonstrate effective target engagement and selectivity with IKKα in U2OS cells through inhibition of IKKα-driven p100 phosphorylation in the noncanonical NF-kB pathway without affecting IKKß-dependent IKappa-Bα loss in the canonical pathway. These compounds represent the first chemical tools that can be used to further characterize the role of IKKα in cellular signaling, to dissect this from IKKß and to validate it in its own right as a target in inflammatory diseases.

Diverse heterocyclic scaffolds as dCTP pyrophosphatase 1 inhibitors. Part 1: Triazoles, triazolopyrimidines, triazinoindoles, quinoline hydrazones and arylpiperazines.

A high-throughput screening campaign using a commercial compound library (ChemBridge DiverSET) revealed diverse chemotypes as inhibitors of the human dCTP pyrophosphatase 1 (dCTPase). Triazole, triazolopyrimidine, triazinoindole, quinoline hydrazone and arylpiperazine hits were clustered, confirmed by IC50 determinations, and their preliminary structure-activity-relationships (SAR) and ligand efficiency scores are discussed in this letter.

Diverse heterocyclic scaffolds as dCTP pyrophosphatase 1 inhibitors. Part 2: Pyridone- and pyrimidinone-derived systems.

Two screening campaigns using commercial (Chembridge DiverSET) and proprietary (Chemical Biology Consortium Sweden, CBCS) compound libraries, revealed a number of pyridone- and pyrimidinone-derived systems as inhibitors of the human dCTP pyrophosphatase 1 (dCTPase). In this letter, we present their preliminary structure-activity-relationships (SAR) and ligand efficiency scores (LE and LLE).

Piperazin-1-ylpyridazine Derivatives Are a Novel Class of Human dCTP Pyrophosphatase 1 Inhibitors.

The dCTP pyrophosphatase 1 (dCTPase) is a nucleotide pool "housekeeping" enzyme responsible for the catabolism of canonical and noncanonical nucleoside triphosphates (dNTPs) and has been associated with cancer progression and cancer cell stemness. We have identified a series of piperazin-1-ylpyridazines as a new class of potent dCTPase inhibitors. Lead compounds increase dCTPase thermal and protease stability, display outstanding selectivity over related enzymes and synergize with a cytidine analogue against leukemic cells. This new class of dCTPase inhibitors lays the first stone toward the development of drug-like probes for the dCTPase enzyme.

Identification of Triazolothiadiazoles as Potent Inhibitors of the dCTP Pyrophosphatase 1.

The dCTP pyrophosphatase 1 (dCTPase) is involved in the regulation of the cellular dNTP pool and has been linked to cancer progression. Here we report on the discovery of a series of 3,6-disubstituted triazolothiadiazoles as potent dCTPase inhibitors. Compounds 16 and 18 display good correlation between enzymatic inhibition and target engagement, together with efficacy in a cellular synergy model, deeming them as a promising starting point for hit-to-lead development.

Structure-metabolism-relationships in the microsomal clearance of piperazin-1-ylpyridazines.

In this study, we provide insight into the metabolic profile of a series of piperazin-1-ylpyridazines suffering from rapid in vitro intrinsic clearance in a metabolic stability assay using liver microsomes (e.g. compound 1 MLM/HLM t1/2 = 2/3 min). Aided by empirical metabolite identification and computational predictive models, we designed the structural modifications required to improve in vitro intrinsic clearance by more than 50-fold (e.g. compound 29 MLM/HLM t1/2 = 113/105 min).

Crystal Structure of the Emerging Cancer Target MTHFD2 in Complex with a Substrate-Based Inhibitor.

To sustain their proliferation, cancer cells become dependent on one-carbon metabolism to support purine and thymidylate synthesis. Indeed, one of the most highly upregulated enzymes during neoplastic transformation is MTHFD2, a mitochondrial methylenetetrahydrofolate dehydrogenase and cyclohydrolase involved in one-carbon metabolism. Because MTHFD2 is expressed normally only during embryonic development, it offers a disease-selective therapeutic target for eradicating cancer cells while sparing healthy cells. Here we report the synthesis and preclinical characterization of the first inhibitor of human MTHFD2. We also disclose the first crystal structure of MTHFD2 in complex with a substrate-based inhibitor and the enzyme cofactors NAD+ and inorganic phosphate. Our work provides a rationale for continued development of a structural framework for the generation of potent and selective MTHFD2 inhibitors for cancer treatment. Cancer Res; 77(4); 937-48. ©2017 AACR.

Discovery of the First Potent and Selective Inhibitors of Human dCTP Pyrophosphatase 1.

The dCTPase pyrophosphatase 1 (dCTPase) regulates the intracellular nucleotide pool through hydrolytic degradation of canonical and noncanonical nucleotide triphosphates (dNTPs). dCTPase is highly expressed in multiple carcinomas and is associated with cancer cell stemness. Here we report on the development of the first potent and selective dCTPase inhibitors that enhance the cytotoxic effect of cytidine analogues in leukemia cells. Boronate 30 displays a promising in vitro ADME profile, including plasma and mouse microsomal half-lives, aqueous solubility, cell permeability and CYP inhibition, deeming it a suitable compound for in vivo studies.

Vinylic MIDA Boronates: New Building Blocks for the Synthesis of Aza-Heterocycles.

A two-step synthesis of structurally diverse pyrrole-containing bicyclic systems is reported. ortho-Nitro-haloarenes coupled with vinylic N-methyliminodiacetic acid (MIDA) boronates generate ortho-vinyl-nitroarenes, which undergo a "metal-free" nitrene insertion, resulting in a new pyrrole ring. This novel synthetic approach has a wide substrate tolerance and it is applicable in the preparation of more complex "drug-like" molecules. Interestingly, an ortho-nitro-allylarene derivative furnished a cyclic ß-aminophosphonate motif.

Lysophosphatidic acid receptor (LPAR) modulators: The current pharmacological toolbox.

Lysophosphatidic acids (LPA) are key lipid-signalling molecules that regulate a remarkably diverse set of cellular events, such as motility, chemotaxis, cell cycle progression, viability, and wound healing. The physiological and pathophysiological consequences of LPA signalling are evident and misregulation of LPA signalling can lead to pathologies like cancer, atherosclerosis, ischaemia, and fibrosis. LPA exerts its biological actions mainly through several types of G protein-coupled receptors, some of which display opposing or redundant effects. For this reason, selective LPA receptor small-molecule ligands can shine light on LPA biology and present an exciting opportunity for drug discovery endeavours. This review provides insights into the detailed chemical nature and pharmacological profile of the small-molecules thus far developed as LPA receptor modulators, as well as information on the preparation of key pharmaceuticals. This summary will facilitate future research efforts and nurture collaboration between chemists and biologists working in this emerging field.

Stereoselective synthesis of carbocyclic analogues of the nucleoside Q precursor (PreQ0).

A convergent and stereoselective synthesis of chiral cyclopentyl- and cyclohexylamine derivatives of nucleoside Q precursor (PreQ0) has been accomplished. This synthetic route allows for an efficient preparation of 4-substituted analogues with interesting three-dimensional character, including chiral cyclopentane-1,2-diol and -1,2,3-triol derivatives. This unusual substitution pattern provides a useful starting point for the discovery of novel bioactive molecules.

Chemical strategies for development of ATR inhibitors.

ATR protein kinase is one of the key players in maintaining genome integrity and coordinating of the DNA damage response and repair signalling pathways. Inhibition of ATR prevents signalling from stalled replication forks and enhances the formation of DNA damage, particularly under conditions of replication stress present in cancers. For this reason ATR/CHK1 checkpoint inhibitors can potentiate the effect of DNA cross-linking agents, as evidenced by ATR inhibitors recently entering human clinical trials. This review aims to compile the existing literature on small molecule inhibitors of ATR, both from academia and the pharmaceutical industry, and will provide the reader with a comprehensive summary of this promising oncology target.

Small-molecule inhibitors of IκB kinase (IKK) and IKK-related kinases.

The transcription factors NF-κB and IFN control important signaling cascades and mediate the expression of a number of important pro-inflammatory cytokines, adhesion molecules, growth factors and anti-apoptotic survival proteins. IκB kinase (IKK) and IKK-related kinases (IKKε and TBK1) are key regulators of these biological pathways and, as such, modulators of these enzymes may be useful in the treatment of inflammatory diseases and cancer. We have reviewed the most recent IKK patent literature (2008-2012), added publications of interest overlooked in previous patent reviews and identified all the players involved in small-molecule inhibitors of the IKKs. This will provide the reader with a decisive summary of the IKK arena, a field that has reached maturity over a decade of research.

Manganese dioxide mediated one-pot synthesis of methyl 9H-pyrido[3,4-b]indole-1-carboxylate: Concise synthesis of alangiobussinine.

The carboline ring system is an important pharmacophore found in a number of biologically important targets. Development of synthetic routes for the preparation of these compounds is important in order to prepare a range of analogues containing the carboline heterocyclic moiety. A manganese dioxide mediated one-pot method starting with an activated alcohol and consisting of alcohol oxidation, Pictet-Spengler cyclisation, and oxidative aromatisation, offers a convenient process that allows access to ß-carbolines. This one-pot process for the preparation of methyl 9H-pyrido[3,4-b]indole-1-carboxylate has subsequently been used as the key step in the synthesis of alangiobussinine and a closely related analogue.