Recent literature has claimed that inhibition of the enzyme MTH1 can eradicate cancer. MTH1 is one of the "housekeeping" enzymes that are responsible for hydrolyzing damaged nucleotides in cells and thus prevent them from being incorporated into DNA. We have developed orthogonal and chemically distinct tool compounds to those published in the literature to allow us to test the hypothesis that inhibition of MTH1 has wide applicability in the treatment of cancer. Here we present the work that led to the discovery of three structurally different series of MTH1 inhibitors with excellent potency, selectivity, and proven target engagement in cells. None of these compounds elicited the reported cellular phenotype, and additional siRNA and CRISPR experiments further support these observations. Critically, the difference between the responses of our highly selective inhibitors and published tool compounds suggests that the effect reported for the latter may be due to off-target cytotoxic effects. As a result, we conclude that the role of MTH1 in carcinogenesis and utility of its inhibition is yet to be established.

The bile acid-sensing transcription factor farnesoid X receptor (FXR) regulates multiple metabolic processes. Modulation of FXR is desired to overcome several metabolic pathologies but pharmacological administration of full FXR agonists has been plagued by mechanism-based side effects. We have developed a modulator that partially activates FXR in vitro and in mice. Here we report the elucidation of the molecular mechanism that drives partial FXR activation by crystallography- and NMR-based structural biology. Natural and synthetic FXR agonists stabilize formation of an extended helix α11 and the α11-α12 loop upon binding. This strengthens a network of hydrogen bonds, repositions helix α12 and enables co-activator recruitment. Partial agonism in contrast is conferred by a kink in helix α11 that destabilizes the α11-α12 loop, a critical determinant for helix α12 orientation. Thereby, the synthetic partial agonist induces conformational states, capable of recruiting both co-repressors and co-activators leading to an equilibrium of co-activator and co-repressor binding.

Steroid receptor drugs have been available for more than half a century, but details of the ligand binding mechanism have remained elusive. We solved X-ray structures of the glucocorticoid and mineralocorticoid receptors to identify a conserved plasticity at the helix 6-7 region that extends the ligand binding pocket toward the receptor surface. Since none of the endogenous ligands exploit this region, we hypothesized that it constitutes an integral part of the binding event. Extensive all-atom unbiased ligand exit and entrance simulations corroborate a ligand binding pathway that gives the observed structural plasticity a key functional role. Kinetic measurements reveal that the receptor residence time correlates with structural rearrangements observed in both structures and simulations. Ultimately, our findings reveal why nature has conserved the capacity to open up this region, and highlight how differences in the details of the ligand entry process result in differential evolutionary constraints across the steroid receptors.

Synthetic glucocorticoids (GC) are essential for the treatment of a broad range of inflammatory diseases. However, their use is limited by target related adverse effects on, e.g., glucose homeostasis and bone metabolism. Starting from a nonsteroidal GR ligand (4) that is a full agonist in reporter gene assays, we exploited key functional triggers within the receptor, generating a range of structurally diverse partial agonists. Of these, only a narrow subset exhibited full anti-inflammatory efficacy and a significantly reduced impact on adverse effect markers in human cell assays compared to prednisolone. This led to the discovery of AZD9567 (15) with excellent in vivo efficacy when dosed orally in a rat model of joint inflammation. Compound 15 is currently being evaluated in clinical trials comparing the efficacy and side effect markers with those of prednisolone.

Lead generation for difficult-to-drug targets that have large, featureless, and highly lipophilic or highly polar and/or flexible binding sites is highly challenging. Here, we describe how cores of macrocyclic natural products can serve as a high-quality in silico screening library that provides leads for difficult-to-drug targets. Two iterative rounds of docking of a carefully selected set of natural-product-derived cores led to the discovery of an uncharged macrocyclic inhibitor of the Keap1-Nrf2 protein–protein interaction, a particularly challenging target due to its highly polar binding site. The inhibitor displays cellular efficacy and is well-positioned for further optimization based on the structure of its complex with Keap1 and synthetic access. We believe that our work will spur interest in using macrocyclic cores for in silico -based lead generation and also inspire the design of future macrocycle screening collections.

A lead generation and optimization program delivered the highly selective and potent CatC inhibitor 10 as an in vivo tool compound and potential development candidate. Structural studies were undertaken to generate SAR understanding.

A class of potent, nonsteroidal, selective indazole ether-based glucocorticoid receptor modulators (SGRMs) was developed for the inhaled treatment of respiratory diseases. Starting from an orally available compound with demonstrated anti-inflammatory activity in rat, a soft-drug strategy was implemented to ensure rapid elimination of drug candidates to minimize systemic GR activation. The first clinical candidate 1b (AZD5423) displayed a potent inhibition of lung edema in a rat model of allergic airway inflammation following dry powder inhalation combined with a moderate systemic GR-effect, assessed as thymic involution. Further optimization of inhaled drug properties provided a second, equally potent, candidate, 15m (AZD7594), that demonstrated an improved therapeutic ratio over the benchmark inhaled corticosteroid 3 (fluticasone propionate) and prolonged the inhibition of lung edema, indicating potential for once-daily treatment.

Matrilysin or matrix metalloproteinase 7 (MMP7) is a member of a class of zinc-dependent endopeptidases (MMPs) capable of degrading extracellular matrix proteins and thought to play an important role in tissue remodeling associated with various physiological and pathological processes. It has broad specificity and cleaves a number of matrix substances, including proteoglycans and collagen III/IV/V/IX/X/XI, [1] which underlies a potential role for MMP7 inhibitors in the treatment of disease associated with tissue degradation/remodeling. In addition, MMP7 has been reported to have a potential role in tumor metastasis and inflammatory processes.[2] It has also been reported to be expressed in osteoarthritic cartilage, where it is colocalized with the tetraspanin, CD151, which has been implicated in its pericellular activation. [3] As a class, MMPs have been the subject of intense study within the pharmaceutical industry over the last decade, and inhibitors have been described for many of the individual MMPs. [4][5][6] However, MMP selectivity remains a significant hurdle for most MMP inhibitors due to a reliance on zinc chelation to provide a significant component of the binding energy. Despite high therapeutic value, there has been a consistent lack of success with such inhibitors in the clinic, and this can largely be attributed to several factors: poor selectivity, poor target validation for the targeted therapy, and poorly defined predictive preclinical animal models for safety and efficacy. The selectivity problem is particularly true with compounds bearing groups that chelate strongly to zinc (e.g., hydroxamate, reverse hydroxamate) because a large component of the binding energy is derived from a feature common to all MMPs.With this in mind, we initiated a program to identify selective MMP7 inhibitors employing a high-throughput screening approach, with MMP selectivity assessment very early in the screening process. To provide an initial indication of selectivity, actives were screened against MMP1 and MMP14. Compounds showing good selectivity against these two MMPs were then screened against MMP2, MMP12 and MMP13 to give an indication of wider MMP selectivity. As anticipated, potent hydroxamate and reverse hydroxamate inhibitors were identified but none of them showed selectivity over other MMPs. In fact, without exception, hydroxamate, reverse hydroxamate and also hydantoin inhibitors proved more active against many of the other MMPs than against MMP7. One active hit that emerged from these initial selectivity screens was the carboxylic acid compound I.[7] The compound has low potency against MMP7, but there was no indication of activity against any of the other MMPs tested. The low potency limited a true assessment of selectivity but, nonetheless, this initial lead was pursued further, driven largely by two factors: ease of synthesis and crystallization. Firstly, we were able to successfully crystallize this compound within the MMP7 protein and determine the complex structure to enable an understanding of ...