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Background and purpose: The ATP-gated P2X 7 receptor has been shown to play a role in several inflammatory processes, making it an attractive target for anti-inflammatory drug discovery. We have recently identified a novel set of cyclic imide compounds that inhibited P2X 7 receptor-mediated dye uptake in human macrophage THP-1 cells. In this study the actions and selectivity of one of these compounds, AZ11645373, were characterized. Experimental approach: We measured membrane currents, calcium influx, and YOPRO-1 uptake from HEK cells expressing individual P2X receptors, and YOPRO1 uptake and interleukin-1b release from THP-1 cells in response to ATP and the ATP analogue benzoylbenzoyl ATP (BzATP). Key results: AZ11645373 up to 10 mM, had no agonist or antagonist actions on membrane currents due to P2X receptor activation at human P2X 1 , rat P2X 2 , human P2X 3 , rat P2X 2/3 , human P2X 4 , or human P2X 5 receptors expressed in HEK cells. AZ11645373 inhibited human P2X 7 receptor responses in HEK cells in a non-surmountable manner with K B values ranging from 5 -20 nM, with mean values not significantly different between assays. K B values were not altered by removing extracellular calcium and magnesium. ATP-evoked IL-1b release from lipopolysaccharide-activated THP-1 cells was inhibited by AZ11645373, IC 50 ¼ 90 nM. AZ11645373 was 4 500-fold less effective at inhibiting rat P2X 7 receptor-mediated currents with less than 50% inhibition occurring at 10 mM. Conclusions and implications: AZ11645373 is a highly selective and potent antagonist at human but not rat P2X 7 receptors and will have much practical value in studies of human cells.

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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.

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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 ...

A novel series of antagonists of the human P2X7 receptor is described. Modification of substituents enabled identification of compounds selective for the rat P2X7 receptor and provides useful pharmacological tools for evaluation of the role of P2X7 in disease.

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