Structure-based design and development of (benz)imidazole pyridones as JAK1-selective kinase inhibitors.

The mammalian Janus Kinases (JAK1, JAK2, JAK3 and TYK2) are intracellular, non-receptor tyrosine kinases whose activities have been associated in the literature and the clinic with a variety of hyperproliferative diseases and immunological disorders. At the onset of the program, it was hypothesized that a JAK1 selective compound over JAK2 could lead to an improved therapeutic index relative to marketed non-selective JAK inhibitors by avoiding the clinical AEs, such as anemia, presumably associated with JAK2 inhibition. During the course of the JAK1 program, a number of diverse chemical scaffolds were identified from both uHTS campaigns and de novo scaffold design. As part of this effort, a (benz)imidazole scaffold evolved via a scaffold-hopping exercise from a mature chemical series. Concurrent crystallography-driven exploration of the ribose pocket and the solvent front led to analogs with optimized kinome and JAK1 selectivities over the JAK2 isoform by targeting several residues unique to JAK1, such as Arg-879 and Glu-966.

Live cell screening to identify RNA-binding small molecule inhibitors of the pre-let-7-Lin28 RNA-protein interaction.

Dysregulation of the networking of RNA-binding proteins (RBPs) and RNAs drives many human diseases, including cancers, and the targeting of RNA-protein interactions (RPIs) has emerged as an exciting area of RNA-targeted drug discovery. Accordingly, methods that enable the discovery of cell-active small molecule modulators of RPIs are needed to propel this emerging field forward. Herein, we describe the application of live-cell assay technology, RNA interaction with protein-mediated complementation assay (RiPCA), for high-throughput screening to identify small molecule inhibitors of the pre-let-7d-Lin28A RPI. Utilizing a combination of RNA-biased small molecules and virtual screening hits, we discovered an RNA-binding small molecule that can disrupt the pre-let-7-Lin28 interaction demonstrating the potential of RiPCA for advancing RPI-targeted drug discovery.

Activation of Hepatocyte Growth Factor/MET Signaling as a Mechanism of Acquired Resistance to a Novel YAP1/TEAD Small Molecule Inhibitor.

Many tumor types harbor alterations in the Hippo pathway, including mesothelioma, where a high percentage of cases are considered YAP1/TEAD dependent. Identification of autopalmitoylation sites in the hydrophobic palmitate pocket of TEADs, which may be necessary for YAP1 protein interactions, has enabled modern drug discovery platforms to generate compounds that allosterically inhibit YAP1/TEAD complex formation and transcriptional activity. We report the discovery and characterization of a novel YAP1/TEAD inhibitor MRK-A from an aryl ether chemical series demonstrating potent and specific inhibition of YAP1/TEAD activity. In vivo, MRK-A showed a favorable tolerability profile in mice and demonstrated pharmacokinetics suitable for twice daily oral dosing in preclinical efficacy studies. Importantly, monotherapeutic targeting of YAP1/TEAD in preclinical models generated regressions in a mesothelioma CDX model; however, rapid resistance to therapy was observed. RNA-sequencing of resistant tumors revealed mRNA expression changes correlated with the resistance state and a marked increase of hepatocyte growth factor (HGF) expression. In vitro, exogenous HGF was able to fully rescue cytostasis induced by MRK-A in mesothelioma cell lines. In addition, co-administration of small molecule inhibitors of the MET receptor tyrosine kinase suppressed the resistance generating effect of HGF on MRK-A induced growth inhibition. In this work, we report the structure and characterization of MRK-A, demonstrating potent and specific inhibition of YAP1/TAZ-TEAD-mediated transcriptional responses, with potential implications for treating malignancies driven by altered Hippo signaling, including factors resulting in acquired drug resistance.

Building a Culture of Medicinal Chemistry Knowledge Sharing.

Increasing the efficiency of the drug discovery process is a challenge faced by drug hunters everywhere. One strategy medicinal chemists employ to meet this challenge is learning from knowledge sources within and beyond their organization. In this Perspective, we discuss the evolution of mechanisms for medicinal chemistry knowledge capture and sharing at Merck & Co. over the past 15 years. We describe our approach to knowledge management and report on the multiple enduring and complementary teams and initiatives we have created to capture and share knowledge within a geographically diverse medicinal chemistry community. In addition, this Perspective will share the benefits we have observed and also reflect on what has allowed our efforts to be both successful and sustainable.

Diminishing GSH-Adduct Formation of Tricyclic Diazepine-based Mutant IDH1 Inhibitors.

Mutant isocitrate dehydrogenase 1 (IDH1) has been identified as an attractive oncology target for which >70% of grade II and III gliomas and ∼10% of acute myeloid leukemia (AML) harbor somatic IDH1 mutations. These mutations confer a neomorphic gain of function, leading to the production of the oncometabolite (R)-2-hydroxyglutarate (2-HG). We identified and developed a potent, selective, and orally bioavailable brain-penetrant tricyclic diazepine scaffold that inhibits mutant IDH1. During the course of in vitro metabolism studies, GSH-adduct metabolites were observed. The hypothesis for GSH-adduct formation was driven by the electron-rich nature of the tricyclic core. Herein, we describe our efforts to reduce the electron-rich nature of the core. Ultimately, a strategy focused on core modifications to block metabolic hot spots coupled with substitution pattern changes (C8 N → C linked) led to the identification of new tricyclic analogues with minimal GSH-adduct formation across species while maintaining an overall balanced profile.

Structure-Guided Discovery of Aminoquinazolines as Brain-Penetrant and Selective LRRK2 Inhibitors.

The leucine-rich repeat kinase 2 (LRRK2) protein has been genetically and functionally linked to Parkinson's disease (PD), a disabling and progressive neurodegenerative disorder whose current therapies are limited in scope and efficacy. In this report, we describe a rigorous hit-to-lead optimization campaign supported by structural enablement, which culminated in the discovery of brain-penetrant, candidate-quality molecules as represented by compounds 22 and 24. These compounds exhibit remarkable selectivity against the kinome and offer good oral bioavailability and low projected human doses. Furthermore, they showcase the implementation of stereochemical design elements that serve to enable a potency- and selectivity-enhancing increase in polarity and hydrogen bond donor (HBD) count while maintaining a central nervous system-friendly profile typified by low levels of transporter-mediated efflux and encouraging brain penetration in preclinical models.

Discovery and Optimization of Potent, Selective, and Brain-Penetrant 1-Heteroaryl-1H-Indazole LRRK2 Kinase Inhibitors for the Treatment of Parkinson's Disease.

Inhibition of leucine-rich repeat kinase 2 (LRRK2) kinase activity represents a genetically supported, chemically tractable, and potentially disease-modifying mechanism to treat Parkinson's disease. Herein, we describe the optimization of a novel series of potent, selective, central nervous system (CNS)-penetrant 1-heteroaryl-1H-indazole type I (ATP competitive) LRRK2 inhibitors. Type I ATP-competitive kinase physicochemical properties were integrated with CNS drug-like properties through a combination of structure-based drug design and parallel medicinal chemistry enabled by sp3-sp2 cross-coupling technologies. This resulted in the discovery of a unique sp3-rich spirocarbonitrile motif that imparted extraordinary potency, pharmacokinetics, and favorable CNS drug-like properties. The lead compound, 25, demonstrated exceptional on-target potency in human peripheral blood mononuclear cells, excellent off-target kinase selectivity, and good brain exposure in rat, culminating in a low projected human dose and a pre-clinical safety profile that warranted advancement toward pre-clinical candidate enabling studies.

Recent Therapeutic Approaches to Modulate the Hippo Pathway in Oncology and Regenerative Medicine.

The Hippo pathway is an evolutionary conserved signaling network that regulates essential processes such as organ size, cell proliferation, migration, stemness and apoptosis. Alterations in this pathway are commonly found in solid tumors and can lead to hyperproliferation, resistance to chemotherapy, compensation for mKRAS and tumor immune evasion. As the terminal effectors of the Hippo pathway, the transcriptional coactivators YAP1/TAZ and the transcription factors TEAD1-4 present exciting opportunities to pharmacologically modulate the Hippo biology in cancer settings, inflammation and regenerative medicine. This review will provide an overview of the progress and current strategies to directly and indirectly target the YAP1/TAZ protein-protein interaction (PPI) with TEAD1-4 across multiple modalities, with focus on recent small molecules able to selectively bind to TEAD, block its autopalmitoylation and inhibit YAP1/TAZ-TEAD-dependent transcription in cancer.

Discovery and characterization of novel peptide inhibitors of the NRF2/MAFG/DNA ternary complex for the treatment of cancer.

Pathway activating mutations of the transcription factor NRF2 and its negative regulator KEAP1 are strongly correlative with poor clinical outcome with pemetrexed/carbo(cis)platin/pembrolizumab (PCP) chemo-immunotherapy in lung cancer. Despite the strong genetic support and therapeutic potential for a NRF2 transcriptional inhibitor, currently there are no known direct inhibitors of the NRF2 protein or its complexes with MAF and/or DNA. Herein we describe the design of a novel and high-confidence homology model to guide a medicinal chemistry effort that resulted in the discovery of a series of peptides that demonstrate high affinity, selective binding to the Antioxidant Response Element (ARE) DNA and thereby displace NRF2-MAFG from its promoter, which is an inhibitory mechanism that to our knowledge has not been previously described. In addition to their activity in electrophoretic mobility shift (EMSA) and TR-FRET-based assays, we show significant dose-dependent ternary complex disruption of NRF2-MAFG binding to DNA by SPR, as well as cellular target engagement by thermal destabilization of HiBiT-tagged NRF2 in the NCI-H1944 NSCLC cell line upon digitonin permeabilization, and SAR studies leading to improved cellular stability. We report the characterization and unique profile of lead peptide 18, which we believe to be a useful in vitro tool to probe NRF2 biology in cancer cell lines and models, while also serving as an excellent starting point for additional in vivo optimization toward inhibition of NRF2-driven transcription to address a significant unmet medical need in non-small cell lung cancer (NSCLC).

Optimization of brain-penetrant picolinamide derived leucine-rich repeat kinase 2 (LRRK2) inhibitors.

The discovery of potent, kinome selective, brain penetrant LRRK2 inhibitors is the focus of extensive research seeking new, disease-modifying treatments for Parkinson's disease (PD). Herein, we describe the discovery and evolution of a picolinamide-derived lead series. Our initial optimization efforts aimed at improving the potency and CLK2 off-target selectivity of compound 1 by modifying the heteroaryl C-H hinge and linker regions. This resulted in compound 12 which advanced deep into our research operating plan (ROP) before heteroaryl aniline metabolite 14 was characterized as Ames mutagenic, halting its progression. Strategic modifications to our ROP were made to enable early de-risking of putative aniline metabolites or hydrolysis products for mutagenicity in Ames. This led to the discovery of 3,5-diaminopyridine 15 and 4,6-diaminopyrimidine 16 as low risk for mutagenicity (defined by a 3-strain Ames negative result). Analysis of key matched molecular pairs 17 and 18 led to the prioritization of the 3,5-diaminopyridine sub-series for further optimization due to enhanced rodent brain penetration. These efforts culminated in the discovery of ethyl trifluoromethyl pyrazole 23 with excellent LRRK2 potency and expanded selectivity versus off-target CLK2.

Discovery of N-(Indazol-3-yl)piperidine-4-carboxylic Acids as RORγt Allosteric Inhibitors for Autoimmune Diseases.

The clinical success of anti-IL-17 monoclonal antibodies (i.e., Cosentyx and Taltz) has validated Th17 pathway modulation for the treatment of autoimmune diseases. The nuclear hormone receptor RORγt is a master regulator of Th17 cells and affects the production of a host of cytokines, including IL-17A, IL-17F, IL-22, IL-26, and GM-CSF. Substantial interest has been spurred across both academia and industry to seek small molecules suitable for RORγt inhibition. A variety of RORγt inhibitors have been reported in the past few years, the majority of which are orthosteric binders. Here we disclose the discovery and optimization of a class of inhibitors, which bind differently to an allosteric binding pocket. Starting from a weakly active hit 1, a tool compound 14 was quickly identified that demonstrated superior potency, selectivity, and off-target profile. Further optimization focused on improving metabolic stability. Replacing the benzoic acid moiety with piperidinyl carboxylate, modifying the 4-aza-indazole core in 14 to 4-F-indazole, and incorporating a key hydroxyl group led to the discovery of 25, which possesses exquisite potency and selectivity, as well as an improved pharmacokinetic profile suitable for oral dosing.

Total synthesis of the marine natural product (-)-clavosolide A. A showcase for the Petasis-Ferrier union/rearrangement tactic.

[Structure: see text] The total synthesis of the marine diolide (-)-clavosolide A has been achieved in 17 steps (longest linear sequence) from commercially available crotonaldehyde exploiting the Petasis-Ferrier union/rearrangement tactic to construct the requisite aglycon monomer. A one-pot esterification/lactonization employing the Yamaguchi protocol, followed by bis-glycosidation, furnished (-)-clavosolide A.

An Efficient Protocol for the Oxidative Hydrolysis of Ketone SAMP Hydrazones Employing SeO(2) and H(2)O(2) under Buffered (pH 7) Conditions.

An effective oxidative protocol for the liberation of ketones from SAMP hydrazones employing peroxyselenous acid under aqueous buffered conditions (pH 7) has been developed. The procedure proceeds without epimerization of adjacent stereocenters or dehydration, respectively, in representative SAMP alkylation and aldol reaction adducts.

Interaction of halide and carboxylate ions with 4,5-diacetamidoacridine-9(10H)-one: thermodynamics of association and deprotonation events.

4,5-Diacetamidoacridine-9(10H)-one was prepared, and its interactions with halide and benzoate anions were studied using a combination of NMR, fluorescence, and isothermal titration calorimetry experiments. Whereas chloride and bromide exhibited simple association, both fluoride and benzoate exhibited initial entropy-driven association followed by an enthalpically favorable deprotonation of the receptor by a second equivalent of the anion.