Discovery of BI-9787, a potent zwitterionic ketohexokinase inhibitor with oral bioavailability.

Fructose metabolism by ketohexokinase (KHK) is implicated in a variety of metabolic disorders. KHK inhibition is a potential therapeutic strategy for the treatment of diseases including diabetes, non-alcoholic fatty liver disease, and non-alcoholic steatohepatitis. The first small-molecule KHK-inhibitors have entered clinical trials, but it remains unclear if systemic inhibition of KHK by small-molecules will eventually benefit patients. Here we report the discovery of BI-9787, a potent, zwitterionic KHK inhibitor characterized by high permeability and favorable oral rat pharmacokinetics. BI-9787 was identified by optimizing chemical starting points generated via a ligand-based virtual screening of Boehringer's virtual library of synthetically accessible compounds (BICLAIM). It serves as a high-quality in vitro and in vivo tool compound for investigating the role of fructose metabolism in disease.

Design, synthesis and evaluation of MCH receptor 1 antagonists--Part I: Optimization of HTS hits towards an in vivo efficacious tool compound BI 414.

Despite recent approvals of anti-obesity drugs there is still a high therapeutic need for alternative options with higher efficacy in humans. As part of our MCH-R1 antagonist program for the treatment of obesity, a series of biphenylacetamide HTS hits was evaluated. Several issues of the initial lead structures had to be resolved, such as potency, selectivity over related GPCRs and P-gp efflux limiting brain exposure in this series. We could demonstrate that all parameters can be significantly improved by structural modifications resulting in BI 414 as a potent and orally available MCH-R1 antagonist tool compound with acceptable in vivo efficacy in an animal model of obesity.

Design, synthesis and evaluation of MCH receptor 1 antagonists--Part III: Discovery of pre-clinical development candidate BI 186908.

Although overweight and obesity are highly prevalent conditions, options to treat them are still very limited. As part of our search for safe and effective MCH-R1 antagonists for the treatment of obesity, two series of pyridones and pyridazinones were evaluated. Optimization was aimed at improving DMPK properties by increasing metabolic stability and improving the safety profile by reducing inhibition of the hERG channel and reducing the potential to induce phospholipidosis. Steric shielding of a labile keto moiety with an ortho-methyl group and fine-tuning of the polarity in several parts of the molecule resulted in BI 186908 (11 g), a potent and selective MCH-R1 antagonist with favorable DMPK and CMC properties. Chronic administration of BI 186908 resulted in significant body weight reduction comparable to sibutramine in a 4 week diet-induced obesity model in rats. Based on its favorable safety profile, BI 186908 was advanced to pre-clinical development.

Design, synthesis and evaluation of MCH receptor 1 antagonists--Part II: Optimization of pyridazines toward reduced phospholipidosis and hERG inhibition.

Despite recent success there remains a high therapeutic need for the development of drugs targeting diseases associated with the metabolic syndrome. As part of our search for safe and effective MCH-R1 antagonists for the treatment of obesity, a series of 3,6-disubstituted pyridazines was evaluated. During optimization several issues of the initial lead structures had to be resolved, such as selectivity over related GPCRs, inhibition of the hERG channel as well as the potential to induce phospholipidosis. Utilizing property-based design, we could demonstrate that all parameters can significantly be improved by consequently increasing the polarity of the compounds. By this strategy, we succeeded in identifying potent and orally available MCH-R1 antagonists with good selectivity over M1 and 5-HT2A and an improved safety profile with respect to hERG inhibition and phospholipidosis.

Discovery and development of BI 1265162, an ENaC inhibitor for the treatment of cystic fibrosis.

Lung selective inhibition of the endothelial sodium channel (ENaC) is a potential mutation agnostic treatment of Cystic Fibrosis (CF). We describe the discovery and development of BI 1265162, the first ENaC inhibitor devoid of the amiloride structural motif that entered clinical trials. The design of BI 1265162 focused on its suitability for inhalation via the Respimat® Soft Mist™ Inhaler and a long duration of action. A convergent and scalable route for the synthesis of BI 1265162 as dihydrogen phosphate salt is presented, that was applied to support clinical trials. A phase 2 study with BI 1265162 did not provide a clear sign of clinical benefit. Whether ENaC inhibition will be able to hold its promise for CF patients remains an open question.

Crystal structures of human and mouse ketohexokinase provide a structural basis for species- and isoform-selective inhibitor design.

A molecular understanding of the proteins involved in fructose metabolism is essential for controlling the current spread of fructose-related obesity, diabetes and related adverse metabolic states in Western populations. Fructose catabolism starts with the phosphorylation of D-fructose to fructose 1-phosphate by ketohexokinase (KHK). KHK exists in two alternatively spliced isoforms: the hepatic and intestinal isoform KHK-C and the peripheral isoform KHK-A. Here, the structure of apo murine KHK (mKHK), which differs from structures of human KHK in overall conformation, is reported. An isoform-selective ligand, which offers a 50-fold higher potency on mKHK and human KHK-A compared with KHK-C, is further characterized. In mKHK, large-scale conformational changes are observed upon ligand binding. The structures suggest a combined strategy for the design of species- and isoform-selective KHK inhibitors.

The azulene scaffold from a medicinal chemist's perspective: Physicochemical and in vitro parameters relevant for drug discovery.

Azulene is a bicyclic scaffold rarely applied in medicinal chemistry. Here we report physicochemical and in vitro parameters relevant for drug discovery for a series of diversely substituted azulenes. We synthesized and characterized several scaffold hopping series of analogously substituted azulenes, indoles and naphthalenes. This enabled a comparison of azulene with the more common scaffolds indole and naphthalene. Our data indicates that undesirably low photostability of azulenes is restricted to certain substitution patterns. Generally, we conclude that azulene is an underused lipophilic bicycle and should be considered as a valuable complement to the collection of medicinal chemistry scaffolds.

Discovery of BI 99179, a potent and selective inhibitor of type I fatty acid synthase with central exposure.

Based on a high-throughput screen, cyclopentanecarboxanilides were identified as a new chemotype of non-covalent inhibitors of type I fatty acid synthase (FAS). Starting from initial hits we aimed at generating a tool compound suitable for the in vivo validation of FAS as a therapeutic target. Optimisation yielded BI 99179 which is characterised by high potency, remarkably high selectivity and significant exposure (both peripheral and central) upon oral administration in rats.

Chemical Derivatization Enables MALDI-TOF-Based High-Throughput Screening for Microbial Trimethylamine (TMA)-Lyase Inhibitors.

Microbial-dependent trimethylamine (TMA) generation from dietary precursors such as choline was recently linked to cardiovascular diseases (CVDs) as well as chronic kidney disease (CKD). Inhibition of TMA-generating enzymes in gut bacteria would be an innovative approach to treat these diseases. The potential to accurately quantify secreted TMA levels highlights the capacity of mass spectrometry (MS) for tracking microbial TMA-lyase activity. However, high-throughput screening (HTS) by conventional MS instrumentation is hampered by limited sample throughput. Recent advancement in liquid handling and instrumentation of matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS provides an HTS-compatible MS technology. The deciphering of enzymatic reactions using this label-free readout has been successfully applied but has thus far been limited to peptide/protein-centric activity assays. Here, we demonstrate the versatile applicability of MALDI-TOF by tracking a small molecule within a highly complex sample background. The key to success for this concept was chemical derivatization of the target molecule enabling quantitative assessment of microbial TMA formation. Further, its potential was demonstrated in a side-by-side comparison to RapidFire-MS in a primary screen and subsequent dose-response experiments. Overall, the established assay enables the screening for microbial TMA-lyase inhibitors and serves as a proof of concept for the applicability of MALDI-TOF for demanding assay concepts per se.

Cytotoxic efficacy and influence on cellular phospholipid metabolism of 2-hydroxy- and 2-O-acetyl-octadecylphosphocholines.

The cytotoxic efficacy and influence on phospholipid composition of the new alkylphosphocholines (APC) 2-hydroxy and 2-O-acetyl-octadecylphosphocholines both synthesised in R- and S-configuration (R/S-OH and R/S-O-acetyl) were examined in vitro using HL-60 and MDA-MB-468 cells. IC50- and LC50-values were measured by MTT- and cell count assay. All tested APC showed higher or similar cytotoxic efficacy compared to the well known APC hexadecylphosphocholine (HePC). However, while S-configured APC (IC50) revealed considerably higher cytotoxic activities, only R- (natural)-configured APC caused significant changes in the phospholipid composition of tumour cells. Further investigations revealed an increase in R-O-acyl and loss of PC up to 70% in the membrane of both cell lines. Similar to PC, R-O-acyl bears two long non-polar hydrocarbon chains and stabilises cell membranes structurally, thus, possibly explaining less cytotoxicity and lack of apoptosis induction by R-configured APC. Nevertheless, an enrichment of R-O-acyl up to 70% at the expense of PC in cell membrane is tremendous and may inhibit tumour development by influencing the intracellular lipid signalling. In conclusion, our findings reveal the antitumoral efficacy of all tested new APC and offer new perspectives in drug development targeting phospholipid metabolism.

Design, synthesis, and evaluation of indolinones as triple angiokinase inhibitors and the discovery of a highly specific 6-methoxycarbonyl-substituted indolinone (BIBF 1120).

Inhibition of tumor angiogenesis through blockade of the vascular endothelial growth factor (VEGF) signaling pathway is a new treatment modality in oncology. Preclinical findings suggest that blockade of additional pro-angiogenic kinases, such as fibroblast and platelet-derived growth factor receptors (FGFR and PDGFR), may improve the efficacy of pharmacological cancer treatment. Indolinones substituted in position 6 were identified as selective inhibitors of VEGF-, PDGF-, and FGF-receptor kinases. In particular, 6-methoxycarbonyl-substituted indolinones showed a highly favorable selectivity profile. Optimization identified potent inhibitors of VEGF-related endothelial cell proliferation with additional efficacy on pericyctes and smooth muscle cells. In contrast, no direct inhibition of tumor cell proliferation was observed. Compounds 2 (BIBF 1000) and 3 (BIBF 1120) are orally available and display encouraging efficacy in in vivo tumor models while being well tolerated. The triple angiokinase inhibitor 3 is currently in phase III clinical trials for the treatment of nonsmall cell lung cancer.