Development of a Series of Kynurenine 3-Monooxygenase Inhibitors Leading to a Clinical Candidate for the Treatment of Acute Pancreatitis

Recently, we reported a novel role for KMO in the pathogenesis of acute pancreatitis (AP). A number of inhibitors of kynurenine 3-monooxygenase (KMO) have previously been described as potential treatments for neurodegenerative conditions and particularly for Huntington’s disease. However, the inhibitors reported to date have insufficient aqueous solubility relative to their cellular potency to be compatible with the intravenous (iv) dosing route required in AP. We have identified and optimized a novel series of high affinity KMO inhibitors with favorable physicochemical properties. The leading example is exquisitely selective, has low clearance in two species, prevents lung and kidney damage in a rat model of acute pancreatitis, and is progressing into preclinical development

Carbenoxolone, 20nM inhibitor of 11β-HSD1.

<p>Carbenoxolone, 20nM inhibitor of 11β-HSD1.</p

The UFSRAT Scoring function.

<p>UFSRAT descriptor values have been calculated for the molecules shown and their values input to the scoring function shown. Sqc is the similarity score between the two molecules q and c (query and candidate), M being a vector representing the 48 geometric distribution descriptors.</p

Small molecules bound to FKBP12 in ESI-MS.

<p>P represents FKBP12 apoprotein and P:L protein in complex with a ligand.</p

The UFSRAT workflow.

<p>The UFSRAT workflow.</p

Binding data for 11β-HSD1.

<p>Molecules 5, 6, 7 and 8 were retrieved using carbenoxolone as a query molecule supplied to the UFSRAT technique and screened against the EDULISS database. Binding of the top hits was tested using a scintillation proximity assay (SPA) and fluorescence assay (Fluoresc).</p><p>* HEK-293 cells transfected with human 11β-HSD1</p><p># Recombinant 11β-HSD1</p><p>Binding data for 11β-HSD1.</p

Binding data for FKBP12.

<p>Molecules 1, 2 and 3 were retrieved using query molecules Q1, Q2 and Q3 and the UFSRAT technique, screened against the EDULISS database. Binding of the top hits was tested using thermal denaturation (TDI) and mass spectrometry (ESI-MS).</p><p>* Recombinant human FKBP12</p><p># K_d 15 μM, 27°C [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0116570#pone.0116570.ref031" target="_blank">31</a>]</p><p>Binding data for FKBP12.</p

Generation of UFSRAT all atom descriptors for the P1 distribution.

<p>The atoms in a molecule are all considered as an array of points (black open circles). P1 represents the geometric centre of the atoms (black dot). Euclidean distances are calculated to all atoms from P1 (grey lines). The 3 descriptors for the P1 distribution are the mean, variance and skew of the distance distribution.</p

The four UFSRAT distributions from typed atoms.

<p>A molecule is broken down into four distributions, consisting of all atoms, hydrophobic, hydrogen bond acceptor and hydrogen bond donor.</p

Mass Spectrometry Imaging for Dissecting Steroid Intracrinology within Target Tissues

Steroid concentrations within tissues are modulated by intracellular enzymes. Such “steroid intracrinology” influences hormone-dependent cancers and obesity and provides targets for pharmacological inhibition. However, no high resolution methods exist to quantify steroids within target tissues. We developed mass spectrometry imaging (MSI), combining matrix assisted laser desorption ionization with on-tissue derivatization with Girard T and Fourier transform ion cyclotron resonance mass spectrometry, to quantify substrate and product (11-dehydrocorticosterone and corticosterone) of the glucocorticoid-amplifying enzyme 11β-HSD1. Regional steroid distribution was imaged at 150–200 μm resolution in rat adrenal gland and mouse brain sections and confirmed with collision induced dissociation/liquid extraction surface analysis. In brains of mice with 11β-HSD1 deficiency or inhibition, MSI quantified changes in subregional corticosterone/11-dehydrocorticosterone ratio, distribution of inhibitor, and accumulation of the alternative 11β-HSD1 substrate, 7-ketocholesterol. MSI data correlated well with LC-MS/MS in whole brain homogenates. MSI with derivatization is a powerful new tool to investigate steroid biology within tissues