Epigenetic modulation by targeting bromodomain containing protein 9 (BRD9): Its therapeutic potential and selective inhibition.

The bromodomain-containing protein 9, a component of the SWI/SNF chromatin remodeling complex, functions as an 'epigenetic reader' selectively recognizing acetyl-lysine marks. It regulates chromatin structure and gene expression by recruitment of acetylated transcriptional regulators and by modulating the function of remodeling complexes. Recent data suggests that BRD9 plays an important role in regulating cellular growth and it has been suggested to drive progression of several malignant diseases such as cervical cancer, and acute myeloid leukemia.

Discovery of Potent and Exquisitely Selective Inhibitors of Kinase CK1 with Tunable Isoform Selectivity.

Casein kinases 1 (CK1) are key signaling molecules that have emerged recently as attractive therapeutic targets in particular for the treatment of hematological malignancies. Herein, we report the identification of a new class of potent and highly selective inhibitors of CK1α, δ and ϵ. Based on their optimal in vitro and in vivo profiles and their exclusive selectivity, MU1250, MU1500 and MU1742 were selected as quality chemical probes for those CK1 isoforms.

Comparative structural analyses of the NHL domains from the human E3 ligase TRIM-NHL family.

Tripartite motif (TRIM) proteins constitute one of the largest subfamilies of the RING-type E3 ubiquitin ligases that play a role in diverse processes from homeostasis and immune response to viral restriction. While TRIM proteins typically harbor an N-terminal RING finger, a B-box and a coiled-coil domain, a high degree of diversity lies in their C termini that contain diverse protein interaction modules, most of which, both structures and their roles in intermolecular interactions, remain unknown. Here, high-resolution crystal structures of the NHL domains of three of the four human TRIM-NHL proteins, namely TRIM2, TRIM3 and TRIM71, are presented.

Identification and Biochemical Characterization of Pyrrolidinediones as Novel Inhibitors of the Bacterial Enzyme MurA.

To develop novel antibiotics, targeting the early steps of cell wall peptidoglycan biosynthesis seems to be a promising strategy that is still underutilized. MurA, the first enzyme in this pathway, is targeted by the clinically used irreversible inhibitor fosfomycin. However, mutations in its binding site can cause bacterial resistance.

Development and Characterization of Type I, Type II, and Type III LIM-Kinase Chemical Probes.

LIMKs are important regulators of actin and microtubule dynamics, and they play essential roles in many cellular processes. Deregulation of LIMKs has been linked to the development of diverse diseases, including cancers and cognitive disabilities, but well-characterized inhibitors known as chemical probes are still lacking. Here, we report the characterization of three highly selective LIMK1/2 inhibitors covering all canonical binding modes (type I/II/III) and the structure-based design of the type II/III inhibitors.

PKC isoforms activate LRRK1 kinase by phosphorylating conserved residues (Ser1064, Ser1074 and Thr1075) within the CORB GTPase domain.

Leucine-rich-repeat-kinase 1 (LRRK1) and its homolog LRRK2 are multidomain kinases possessing a ROC-CORA-CORB containing GTPase domain and phosphorylate distinct Rab proteins. LRRK1 loss of function mutations cause the bone disorder osteosclerotic metaphyseal dysplasia, whereas LRRK2 missense mutations that enhance kinase activity cause Parkinson's disease. Previous work suggested that LRRK1 but not LRRK2, is activated via a Protein Kinase C (PKC)-dependent mechanism.

Pharmacokinetic Optimization of Small Molecule Janus Kinase 3 Inhibitors to Target Immune Cells.

Modulation of Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling is a promising method of treating autoimmune diseases, and the profound potency of clinical compounds makes this mode of action particularly attractive. Other questions that remain unanswered also include: What is the ideal selectivity between JAK1 and JAK3? Which cells are most relevant to JAK blockade? And what is the ideal tissue distribution pattern for addressing specific autoimmune conditions? We hypothesized that JAK3 selectivity is most relevant to low-dose clinical effects and interleukin-10 (IL-10) stimulation in particular, that immune cells are the most important compartment, and that distribution to inflamed tissue is the most important pharmacokinetic characteristic for disease modification. To test these hypotheses, we prepared modified derivatives of JAK3 specific inhibitors that target C909 near the ATP binding site based on FM-381, first reported in 2016; a compound class that was hitherto limited in uptake and exposure .

Transfer mechanism of cell-free synthesized membrane proteins into mammalian cells.

Nanodiscs are emerging to serve as transfer vectors for the insertion of recombinant membrane proteins into membranes of living cells. In combination with cell-free expression technologies, this novel process opens new perspectives to analyze the effects of even problematic targets such as toxic, hard-to-express, or artificially modified membrane proteins in complex cellular environments of different cell lines. Furthermore, transferred cells must not be genetically engineered and primary cell lines or cancer cells could be implemented as well.

A Toolbox for the Generation of Chemical Probes for Baculovirus IAP Repeat Containing Proteins.

E3 ligases constitute a large and diverse family of proteins that play a central role in regulating protein homeostasis by recruiting substrate proteins recruitment domains to the proteasomal degradation machinery. Small molecules can either inhibit, modulate or hijack E3 function. The latter class of small molecules led to the development of selective protein degraders, such as PROTACs (PROteolysis TArgeting Chimeras), that recruit protein targets to the ubiquitin system leading to a new class of pharmacologically active drugs and to new therapeutic options.

Enabling pseudokinases as potential drug targets.

Pseudokinases play significant roles in disease development. Similar to active kinases, their cellular functions can be targeted pharmacologically. But notably, instead of inhibiting an enzymatic activity, drug-like molecules act by stabilizing distinct pseudokinase conformations, by interfering with protein interactions, or by inducing proteasomal degradation.

TLR8 is activated by 5'-methylthioinosine, a Plasmodium falciparum-derived intermediate of the purine salvage pathway.

The innate immune recognition of the malaria-causing pathogen Plasmodium falciparum (P. falciparum) is not fully explored. Here, we identify the nucleoside 5'-methylthioinosine (MTI), a Plasmodium-specific intermediate of the purine salvage pathway, as a pathogen-derived Toll-like receptor 8 (TLR8) agonist.

LRRK2 dynamics analysis identifies allosteric control of the crosstalk between its catalytic domains.

The 2 major molecular switches in biology, kinases and GTPases, are both contained in the Parkinson disease-related leucine-rich repeat kinase 2 (LRRK2). Using hydrogen-deuterium exchange mass spectrometry (HDX-MS) and molecular dynamics (MD) simulations, we generated a comprehensive dynamic allosteric portrait of the C-terminal domains of LRRK2 (LRRK2RCKW). We identified 2 helices that shield the kinase domain and regulate LRRK2 conformation and function.

Structural Aspects of LIMK Regulation and Pharmacology.

Malfunction of the actin cytoskeleton is linked to numerous human diseases including neurological disorders and cancer. LIMK1 (LIM domain kinase 1) and its paralogue LIMK2 are two closely related kinases that control actin cytoskeleton dynamics. Consequently, they are potential therapeutic targets for the treatment of such diseases.

TDP-43 Modulation by Tau-Tubulin Kinase 1 Inhibitors: A New Avenue for Future Amyotrophic Lateral Sclerosis Therapy.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease without any effective treatment. Protein TDP-43 is a pathological hallmark of ALS in both sporadic and familiar patients. Post-translational modifications of TDP-43 promote its aggregation in the cytoplasm.

Inhibitors of the Hippo Pathway Kinases STK3/MST2 and STK4/MST1 Have Utility for the Treatment of Acute Myeloid Leukemia.

Serine/threonine-protein kinases 3 and 4 (STK3 and STK4, respectively) are key components of the Hippo signaling pathway, which regulates cell proliferation and death and provides a potential therapeutic target for acute myeloid leukemia (AML). Herein, we report the structure-based design of a series of pyrrolopyrimidine derivatives as STK3 and STK4 inhibitors. In an initial screen, the compounds exhibited low nanomolar potency against both STK3 and STK4.

Discovery of a Potent and Highly Isoform-Selective Inhibitor of the Neglected Ribosomal Protein S6 Kinase Beta 2 (S6K2).

The ribosomal protein S6 kinase beta 2 (S6K2) is thought to play an important role in malignant cell proliferation, but is understudied compared to its closely related homolog S6 kinase beta 1 (S6K1). To better understand the biological function of S6K2, chemical probes are needed, but the high similarity between S6K2 and S6K1 makes it challenging to selectively address S6K2 with small molecules. We were able to design the first potent and highly isoform-specific S6K2 inhibitor from a known S6K1-selective inhibitor, which was merged with a covalent inhibitor engaging a cysteine located in the hinge region in the fibroblast growth factor receptor kinase (FGFR) 4 via a nucleophilic aromatic substitution (SAr) reaction.

Discovery of a Potent Dual SLK/STK10 Inhibitor Based on a Maleimide Scaffold.

SLK (STE20-like kinase) and STK10 (serine/threonine kinase 10) are closely related kinases whose enzymatic activity is linked to the regulation of ezrin, radixin, and moesin function and to the regulation of lymphocyte migration and the cell cycle. We identified a series of 3-anilino-4-arylmaleimides as dual inhibitors of SLK and STK10 with good kinome-wide selectivity. Optimization of this series led to multiple SLK/STK10 inhibitors with nanomolar potency.

Identification of Pyrimidine-Based Lead Compounds for Understudied Kinases Implicated in Driving Neurodegeneration.

The pyrimidine core has been utilized extensively to construct kinase inhibitors, including eight FDA-approved drugs. Because the pyrimidine hinge-binding motif is accommodated by many human kinases, kinome-wide selectivity of resultant molecules can be poor. This liability was seen as an advantage since it is well tolerated by many understudied kinases.

Addressing a Trapped High-Energy Water: Design and Synthesis of Highly Potent Pyrimidoindole-Based Glycogen Synthase Kinase-3β Inhibitors.

In small molecule binding, water is not a passive bystander but rather takes an active role in the binding site, which may be decisive for the potency of the inhibitor. Here, by addressing a high-energy water, we improved the IC value of our co-crystallized glycogen synthase kinase-3β (GSK-3β) inhibitor by nearly two orders of magnitude. Surprisingly, our results demonstrate that this high-energy water was not displaced by our potent inhibitor ()-3-(3-((7-ethynyl-9-pyrimido[4,5-]indol-4-yl)(methyl)amino)piperidin-1-yl)propanenitrile (-, IC value of 6 nM).

Conformation and dynamics of the kinase domain drive subcellular location and activation of LRRK2.

To explore how pathogenic mutations of the multidomain leucine-rich repeat kinase 2 (LRRK2) hijack its finely tuned activation process and drive Parkinson's disease (PD), we used a multitiered approach. Most mutations mimic Rab-mediated activation by "unleashing" kinase activity, and many, like the kinase inhibitor MLi-2, trap LRRK2 onto microtubules. Here we mimic activation by simply deleting the inhibitory N-terminal domains and then characterize conformational changes induced by MLi-2 and PD mutations.

Controlling the Covalent Reactivity of a Kinase Inhibitor with Light.

Covalent kinase inhibitors account for some of the most successful drugs that have recently entered the clinic and many others are in preclinical development. A common strategy is to target cysteines in the vicinity of the ATP binding site using an acrylamide electrophile. To increase the tissue selectivity of kinase inhibitors, it could be advantageous to control the reactivity of these electrophiles with light.

Synthetic Opportunities and Challenges for Macrocyclic Kinase Inhibitors.

Macrocycles are typically cyclic variants of inhibitors derived from uncyclized canonical molecules or from natural products. For medicinal chemistry, drug-like macrocycles have received increasing interest over the past few years, since it has been demonstrated that macrocyclization can favorably alter the biological and physiochemical properties as well as selectivity in comparison to the acyclic analogue. Recent drug approvals such as Lorlatinib, glecaprevir, or voxilaprevir underline the clinical relevance of drug-like macrocycles.

Development of a potent and selective chemical probe for the pleiotropic kinase CK2.

Building on the pyrazolopyrimidine CK2 (casein kinase 2) inhibitor scaffold, we designed a small targeted library. Through comprehensive evaluation of inhibitor selectivity, we identified inhibitor 24 (SGC-CK2-1) as a highly potent and cell-active CK2 chemical probe with exclusive selectivity for both human CK2 isoforms. Remarkably, despite years of research pointing to CK2 as a key driver in cancer, our chemical probe did not elicit a broad antiproliferative phenotype in >90% of >140 cell lines when tested in dose-response.