Pyrazine-based small molecule kinase inhibitors: clinical applications and patent review (2019-2023).

Protein kinases play a key role in cellular signaling pathways including proliferation, apoptosis, inflammation and immune regulation. Therefore, targeting kinases with small molecules has emerged as a therapeutic potential in cancers and other diseases including inflammatory and autoimmune disorders. The main chemical motifs of the available small molecule kinase inhibitors are heterocyclic, nitrogen-containing and six-membered rings including pyrazine. Several potent and selective pyrazine-based kinase inhibitors have been developed and progressed into clinical trials. The data of clinical application of kinase inhibitors demonstrate good clinical activity with manageable toxicity in several relapse-resistant malignancies and severe to moderate immunological disorders. All pyrazine-based kinase inhibitors are orally active. This paper reviews the most recent kinase literature (2019-2023) related to pyrazine-based small molecule inhibitors. This review includes the FDA (Food and Drug Administration)-approved and patent agents along with their targeted kinase, scaffold, potency, selectivity profile, assignee and biological results in clinical and preclinical studies.

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7-Amino-[1,2,4]triazolo[1,5-a][1,3,5]triazines as CK1δ inhibitors: Exploring substitutions at the 2 and 5-positions.

CK1δ is a serine-threonine kinase involved in several pathological conditions including neuroinflammation and neurodegenerative disorders like Alzheimer's disease, Parkinson's disease, and Amyotrophic Lateral Sclerosis. Specifically, it seems that an inhibition of CK1δ could have a neuroprotective effect in these conditions. Here, a series of [1,2,4]triazolo[1,5-a][1,3,5]triazines were developed as ATP-competitive CK1δ inhibitors. Both positions 2 and 5 have been explored leading to a total of ten compounds exhibiting IC50s comprised between 29.1 µM and 2.08 µM. Three of the four most potent compounds (IC50 < 3 µM) bear a thiophene ring at the 2 position. All compounds have been submitted to computational studies that identified the chain composed of at least 2 atoms (e.g., nitrogen and carbon atoms) at the 5 position as crucial to determine a key bidentate hydrogen bond with Leu85 of CK1δ. Most potent compounds have been tested in vitro, resulting passively permeable to the blood-brain barrier and, safe and slight neuroprotective on a neuronal cell model. These results encourage to further structural optimize the series to obtain more potent CK1δ inhibitors as possible neuroprotective agents to be tested on models of the above-mentioned neurodegenerative diseases.

Role of GSK-3ß Inhibitors: New Promises and Opportunities for Alzheimer's Disease.

Glycogen synthase kinase-3 (GSK-3) was discovered to be a multifunctional enzyme involved in a wide variety of biological processes, including early embryo formation, oncogenesis, as well cell death in neurodegenerative diseases. Several critical cellular processes in the brain are regulated by the GSK-3ß, serving as a central switch in the signaling pathways. Dysregulation of GSK-3ß kinase has been reported in diabetes, cancer, Alzheimer's disease, schizophrenia, bipolar disorder, inflammation, and Huntington's disease. Thus, GSK-3ß is widely regarded as a promising target for therapeutic use. The current review article focuses mainly on Alzheimer's disease, an age-related neurodegenerative brain disorder. GSK-3ß activation increases amyloid-beta (Aß) and the development of neurofibrillary tangles that are involved in the disruption of material transport between axons and dendrites. The drug-binding cavities of GSK-3ß are explored, and different existing classes of GSK-3ß inhibitors are explained in this review. Non-ATP competitive inhibitors, such as allosteric inhibitors, can reduce the side effects compared to ATP-competitive inhibitors. Whereas ATP-competitive inhibitors produce disarrangement of the cytoskeleton, neurofibrillary tangles formation, and lead to the death of neurons, etc. This could be because they are binding to a site separate from ATP. Owing to their interaction in particular and special binding sites, allosteric ligands interact with substrates more selectively, which will be beneficial in resolving drug-induced resistance and also helpful in reducing side effects. Hence, in this review, we focussed on the allosteric GSK-3ß inhibitors and discussed their futuristic opportunities as anti-Alzheimer's compounds.

Virtual screening-based discovery of AI-2 quorum sensing inhibitors that interact with an allosteric hydrophobic site of LsrK and their functional evaluation.

Introduction: Quorum sensing (QS) is a bacterial intracellular and intercellular communication system that regulates virulence factor production, biofilm formation, and antibiotic sensitivity. Quorum-sensing inhibitors (QSIs) are a novel class of antibiotics that can effectively combat antibiotic resistance. Autoinducer-2 (AI-2) is a universal signaling molecule that mediates inter- and intraspecies QS systems among different bacteria. Furthermore, LsrK plays an important role in regulating the activity and stability of the intracellular AI-2 signaling pathway. Thus, LsrK is considered an important target for the development of QSIs. Methods: We designed a workflow integrating molecular dynamic (MD) simulations, virtual screening, LsrK inhibition assays, cell-based AI-2-mediated QS interference assays, and surface plasmon resonance (SPR)-based protein affinity assays to screen for potential LsrK kinase inhibitors. Results: MD simulation results of the LsrK/ATP complex revealed hydrogen bonds and salt bridge formation among four key residues, namely, Lys 431, Tyr 341, Arg 319, and Arg 322, which are critical for the binding of ATP to LsrK. Furthermore, MD simulation results indicated that the ATP-binding site has an allosteric pocket that can become larger and be occupied by small molecule compounds. Based on these MD simulation results, a constraint of forming at least one hydrogen bond with Arg 319, Arg 322, Lys 431, or Tyr 341 residues was introduced when performing virtual screening using Glide's virtual screening workflow (VSW). In the meantime, compounds with hydrophobic group likely to interact with the allosteric hydrophobic pocket are preferred when performing visual inspection. Seventy-four compounds were selected for the wet laboratory assays based on virtual screening and the absorption, distribution, metabolism, and excretion (ADME) properties of these compounds. LsrK inhibition assays revealed 12 compounds inhibiting LsrK by more than 60% at a 200 µM concentration; four of these (Y205-6768, D135-0149, 3284-1358, and N025-0038) had IC50 values below 50 µM and were confirmed as ATP-competitive inhibitors. Six of these 12 LsrK inhibitors exhibited high AI-2 QS inhibition, of which, Y205-6768 had the highest activity with IC50 = 11.28 ± 0.70 µM. The SPR assay verified that compounds Y205-6768 and N025-0038 specifically bound to LsrK. MD simulation analysis of the docking complexes of the four active compounds with LsrK further confirmed the importance of forming hydrogen bonds and salt bridges with key basic amino acid residues including Lys 431, Tyr 341, Arg 319, and Arg 322 and filling the allosteric hydrophobic pocket next to the purine-binding site of LsrK. Discussion: Our study clarified for the first time that there is an allosteric site near the ATP-binding site of Lsrk and that it enriches the structure-activity relationship information of Lsrk inhibitors. The four identified compounds showed novel structures, low molecular weights, high activities, and novel LsrK binding modes, rendering them suitable for further optimization for effective AI-2 QSIs. Our work provides a valuable reference for the discovery of QSIs that do not inhibit bacterial growth, thereby avoiding the emergence of drug resistance.

ATP-competitive and allosteric inhibitors induce differential conformational changes at the autoinhibitory interface of Akt1.

Akt is a master regulator of pro-growth signaling in the cell. Akt is activated by phosphoinositides that disrupt the autoinhibitory interface between the kinase and pleckstrin homology (PH) domains and then is phosphorylated at T308 and S473. Akt hyperactivation is oncogenic, which has spurred development of potent and selective inhibitors as therapeutics. Using hydrogen deuterium exchange mass spectrometry (HDX-MS), we interrogated the conformational changes upon binding Akt ATP-competitive and allosteric inhibitors. We compared inhibitors against three different states of Akt1. The allosteric inhibitor caused substantive conformational changes and restricts membrane binding. ATP-competitive inhibitors caused extensive allosteric conformational changes, altering the autoinhibitory interface and leading to increased membrane binding, suggesting that the PH domain is more accessible for membrane binding. This work provides unique insight into the autoinhibitory conformation of the PH and kinase domain and conformational changes induced by Akt inhibitors and has important implications for the design of Akt targeted therapeutics.

Identification of potential ATP-competitive cyclin-dependent kinase 1 inhibitors: De novo drug generation, molecular docking, and molecular dynamics simulation.

Cyclin-dependent kinases 1 (CDK1) has been identified as a potential target for the search for new antitumor drugs. However, no clinically effective CDK1 inhibitors are now available for cancer treatment. Therefore, this study aimed to offer potential CDK1 inhibitors using de novo drug generation, molecular docking, and molecular dynamics (MD) simulation studies. We first utilized the BREED algorithm (a de novo drug generation approach) to produce a novel library of small molecules targeting CDK1. To initially obtain novel potential CDK1 inhibitors with favorable physicochemical properties and excellent druggability, we performed a virtual rule-based rational drug screening on our generated library and found ten initial hits. Then, the molecular interactions and dynamic stability of these ten initial hits and CDK1 complexes during their all-atom MD simulations (total 18 µs) and binding pose metadynamics simulations were investigated, resulting in five final hits. Furthermore, another MD simulation (total 2.1 µs) with different force fields demonstrated the binding ability of the five hits to CDK1. It was found that these five hits, CBMA001 (ΔG = -29.88 kcal/mol), CBMA002 (ΔG = -34.89 kcal/mol), CBMA004 (ΔG = -32.47 kcal/mol), CBMA007 (ΔG = -31.16 kcal/mol), and CBMA008 (ΔG = -34.78 kcal/mol) possessed much greater binding affinity to CDK1 than positive compound Flavopiridol (FLP, ΔG = -25.38 kcal/mol). Finally, CBMA002 and CBMA004 were identified as excellent selective CDK1 inhibitors in silico. Together, this study provides a workflow for rational drug design and two promising selective CDK1 inhibitors that deserve further investigation.

Investigation of morpholine isosters for the development of a potent, selective and metabolically stable mTOR kinase inhibitor.

Upregulation of mechanistic target of rapamycin (mTOR) signaling drives various types of cancers and neurological diseases. Rapamycin and its analogues (rapalogs) are first generation mTOR inhibitors, and selectively block mTOR complex 1 (TORC1) by an allosteric mechanism. In contrast, second generation ATP-binding site inhibitors of mTOR kinase (TORKi) target both TORC1 and TORC2. Here, we explore 3,6-dihydro-2H-pyran (DHP) and tetrahydro-2H-pyran (THP) as isosteres of the morpholine moiety to unlock a novel chemical space for TORKi generation. A library of DHP- and THP-substituted triazines was prepared, and molecular modelling provided a rational for a structure activity relationship study. Finally, compound 11b [5-(4-(3-oxa-8-azabicyclo[3.2.1]octan-8-yl)-6-(tetrahydro-2H-pyran-4-yl)-1,3,5-triazin-2-yl)-4-(difluoromethyl)pyridin-2-amine] was selected due its potency and selectivity for mTOR kinase over the structurally related class I phosphoinositide 3-kinases (PI3Ks) isoforms. 11b displayed high metabolic stability towards CYP1A1 degradation, which is of advantage in drug development. After oral administration to male Sprague Dawley rats, 11b reached high concentrations both in plasma and brain, revealing an excellent oral bioavailability. In a metabolic stability assay using human hepatocytes, 11b was more stable than PQR620, the first-in-class brain penetrant TORKi. Compound 11b also displayed dose-dependent anti-proliferative activity in splenic marginal zone lymphoma (SMZL) cell lines as single agent and when combined with BCL2 inhibition (venetoclax). Our results identify the THP-substituted triazine core as a novel scaffold for the development of metabolically stable TORKi for the treatment of chronic diseases and cancers driven by mTOR deregulation and requiring drug distribution also to the central nervous system.

Dipyridamole interacts with the N-terminal domain of HSP90 and antagonizes the function of the chaperone in multiple cancer cell lines.

Molecular chaperone HSP90 has been considered as a promising target for anti-cancer drug development for years. However, due to the heat shock response induced by the ATP competitive inhibitors against HSP90, the therapeutic efficacies of the compounds are compromised, which consequently restricts the clinical use of HSP90-targeted inhibitors. Therefore, there is a need to discover novel HSP90-targeted modulators which exhibit acceptable inhibition activity against the chaperone and do not induce significant heat shock response in the meantime. Here in this study, we firstly developed a tip-based affinity selection-mass spectrometry platform with optimized experimental conditions/parameters for HSP90-targeted active compound screening, and then applied it to fish out inhibitors against HSP90 from a collection of 2,395 compounds composed of FDA-approved drugs and drug candidates. Dipyridamole, which acts as an anti-thrombotic agent by modulating multiple targets and has a long history of safe use, was identified to interact with HSP90's N-terminal domain. The following conducted biophysical and biochemical experiments demonstrated that Dipyridamole could bind to HSP90's ATP binding pocket and function as an ATP competitive inhibitor of the chaperone. Finally, cellular-based assays including CESTA, cell viability assessment and proteomic analysis etc. were performed to evaluate whether the interaction between HSP90 and Dipyridamole contributes to the anti-tumor effects of the compound. We then found that Dipyridamole inhibits the growth and proliferation of human cancer cells by downregulating cell cycle regulators and upregulating apoptotic cell signaling, which are potentially mediated by the binding of Dipyridamole to HSP90 and to PDEs (phosphodiesterases), respectively.

New Insights on the Nuclear Functions and Targeting of FAK in Cancer.

Focal adhesion kinase (FAK) is a non-receptor tyrosine kinase over-expressed and activated in both adult and pediatric cancers, where it plays important roles in the regulation of pathogenesis and progression of the malignant phenotype. FAK exerts its functions in cancer by two different ways: a kinase activity in the cytoplasm, mainly dependent on the integrin signaling, and a scaffolding activity into the nucleus by networking with different gene expression regulators. For this reason, FAK has to be considered a target with high therapeutic values. Indeed, evidence suggests that FAK targeting could be effective, either alone or in combination, with other already available treatments. Here, we propose an overview of the novel insights about FAK's structure and nuclear functions, with a special focus on the recent findings concerning the roles of this protein in cancer. Additionally, we provide a recent update on FAK inhibitors that are currently in clinical trials for patients with cancer, and discuss the challenge and future directions of drug-based anti-FAK targeted therapies.

Marine-Derived Natural Products as ATP-Competitive mTOR Kinase Inhibitors for Cancer Therapeutics.

The mammalian target of rapamycin (mTOR) is a serine/threonine kinase portraying a quintessential role in cellular proliferation and survival. Aberrations in the mTOR signaling pathway have been reported in numerous cancers including thyroid, lung, gastric and ovarian cancer, thus making it a therapeutic target. To attain this objective, an in silico investigation was designed, employing a pharmacophore modeling approach. A structure-based pharmacophore (SBP) model exploiting the key features of a selective mTOR inhibitor, Torkinib directed at the ATP-binding pocket was generated. A Marine Natural Products (MNP) library was screened using SBP model as a query. The retrieved compounds after consequent drug-likeness filtration were subjected to molecular docking with mTOR, thus revealing four MNPs with better scores than Torkinib. Successive refinement via molecular dynamics simulations demonstrated that the hits formed crucial interactions with key residues of the pocket. Furthermore, the four identified hits exhibited good binding free energy scores through MM-PBSA calculations and the subsequent in silico toxicity assessments displayed three hits deemed essentially non-carcinogenic and non-mutagenic. The hits presented in this investigation could act as potent ATP-competitive mTOR inhibitors, representing a platform for the future discovery of drugs from marine natural origin.

Computational Insights into the Potential of Withaferin-A, Withanone and Caffeic Acid Phenethyl Ester for Treatment of Aberrant-EGFR Driven Lung Cancers.

The anticancer activities of Withaferin-A (Wi-A) and Withanone (Wi-N) from Ashwagandha and Caffeic Acid Phenethyl Ester (CAPE) from honeybee propolis have been well documented. Here, we examined the binding potential of these natural compounds to inhibit the constitutive phosphorylation of epidermal growth factor receptors (EGFRs). Exon 20 insertion mutants of EGFR, which show resistance to various FDA approved drugs and are linked to poor prognosis of lung cancer patients, were the primary focus of this study. Apart from exon 20 insertion mutants, the potential of natural compounds to serve as ATP competitive inhibitors of wildtype protein and other common mutants of EGFR, namely L858R and exon19del, were also examined. The potential of natural compounds was compared to the positive controls such as erlotinib, TAS6417 and poziotinib. Similar to known inhibitors, Wi-A and Wi-N could displace and binds at the ATP orthosteric site of exon19del, L858R and exon20, while CAPE was limited to wildtype EGFR and exon 20 insertion mutants only. Moreover, the binding free energy of the natural drugs against EGFRs was also comparable to the positive controls. This computational study suggests that Wi-A and Wi-N have potential against multiple mutated EGFRs, warranting further in vitro and in vivo experiments.

Data set of competitive and allosteric protein kinase inhibitors confirmed by X-ray crystallography.

A data set was generated comprising currently available competitive and allosteric human protein kinase inhibitors confirmed by X-ray crystallography. This data set has been used to systematically explore structural relationships between these types of inhibitors with different mechanisms of action. A major finding of this study has been that these different inhibitor types frequently displayed structural relationships and essentially represented a structural continuum [1]. Use of the data set is not limited to the inhibitor-centric exploration of structural relationships. The collection of kinase inhibitors with structurally confirmed distinct mechanisms of action can also be used, for example, to aid in structure-based drug design or the search for new allosteric kinase inhibitors.

Systematic comparison of competitive and allosteric kinase inhibitors reveals common structural characteristics.

Allosteric and ATP-competitive kinase inhibitors act by distinct mechanisms and are expected to have high and low kinase selectivity, respectively. This also raises the question whether or not these different types of inhibitors might be structurally distinct. To address this question, we have assembled data sets of currently available competitive and allosteric kinase inhibitors confirmed by X-ray crystallography and systematically compared these compounds on the basis of different structural criteria. Many competitive and allosteric inhibitors were found to contain the same or similar substructures and a subset of allosteric inhibitors was found to share core structures with ATP site-directed inhibitors. In some instances, small chemical modifications of common cores were found to yield either allosteric or competitive inhibitors. Hence, these different categories of inhibitors with distinct mechanisms of action were often structurally related and represented much more of a structural continuum than discrete states. Additional target annotations were frequently identified for competitive inhibitors, but were rare for allosteric inhibitors. As a part of this study, our collection of kinase inhibitors and the associated information are made freely available to enable further assessment of chemical modifications that distinguish similar kinase inhibitors with distinct mechanisms of action.

Potential of Withaferin-A, Withanone and Caffeic Acid Phenethyl ester as ATP-competitive inhibitors of BRAF: A bioinformatics study.

Serine/threonine-protein kinase B-raf (BRAF) plays a significant role in regulating cell division and proliferation through MAPK/ERK pathway. The constitutive expression of wild-type BRAF (BRAFWT) and its mutant forms, especially V600E (BRAFV600E), has been linked to multiple cancers. Various synthetic drugs have been approved and are in clinical trials, but most of them are reported to become ineffective within a short duration. Therefore, combinational therapy involving multiple drugs are often recruited for cancer treatment. However, they lead to toxicity and adverse side effects. In this computational study, we have investigated three natural compounds, namely Withaferin-A (Wi-A), Withanone (Wi-N) and Caffeic Acid Phenethyl ester (CAPE) for anti-BRAFWT and anti-BRAFV600E activity. We found that these compounds could bind stably at ATP-binding site in both BRAFWT and BRAFV600E proteins. In-depth analysis revealed that these compounds maintained the active conformation of wild-type BRAF protein by inducing αC-helix-In, DFG-In, extended activation segment and well-aligned R-spine residues similar to already known drugs Vemurafenib (VEM), BGB283 and Ponatinib. In terms of binding energy, among the natural compounds, CAPE showed better affinity towards both wild-type and V600E mutant proteins than the other two compounds. These data suggested that CAPE, Wi-A and Wi-N have potential to block constitutive autophosphorylation of BRAF and hence warrant in vitro and in vivo experimental validation.

Recent advances in Bcr-Abl tyrosine kinase inhibitors for overriding T315I mutation.

BCR-ABL is a gene produced by the fusion of the bcr gene and the c-abl proto-oncogene and is considered to be the main cause of chronic myelogenous leukemia (CML) production. Therefore, the development of selective Bcr-Abl kinase inhibitors is an attractive strategy for the treatment of CML. However, in the treatment of CML with a Bcr-Abl kinase inhibitor, the T315I gatekeeper mutant disrupts the important contact interaction between the inhibitor and the enzyme, resistant to the first- and second-generation drugs currently approved, such as imatinib, bosutinib, nilotinib, and dasatinib. In order to overcome this special resistance, several different strategies have been explored, and many molecules have been studied to effectively inhibit Bcr-Abl T315I. Some of these molecules are still under development, and some are being studied preclinically, and still others are in clinical research. Herein, this review reports some of the major examples of third-generation Bcr-Abl inhibitors against the T315I mutation.

Synthesis, biological properties and structural study of new halogenated azolo[4,5-b]pyridines as inhibitors of CK2 kinase.

The new halogenated 1H-triazolo[4,5-b]pyridines and 1H-imidazo[4,5-b]pyridines were synthesised as analogues of known CK2 inhibitors: 4,5,6,7-tetrabromo-1H-benzotriazole (TBBt) and 4,5,6,7-tetrabromo-1H-benzimidazole (TBBi). Their influence on the activity of recombinant human CK2α, CK2α' and PIM1 kinases was determined. The most active inhibitors were di- and trihalogenated 1H-triazolo[4,5-b]pyridines (4a, 5a and 10a) with IC50 values 2.56, 3.82 and 3.26 µM respectively for CK2α. Furthermore, effect on viability of cancer cell lines MCF-7 (human breast adenocarcinoma) and CCRF-CEM (T lymphoblast leukemia) of all final compounds was evaluated. Finally, three crystal structures of complexes of CK2α1-335 with inhibitors 4a, 5a and 10a were obtained. In addition, new protocol was used to obtain high-resolution crystal structures of CK2α'Cys336Ser in complex with four inhibitors (4a, 5a, 5b, 10a).

Dynamically Shaping Chaperones. Allosteric Modulators of HSP90 Family as Regulatory Tools of Cell Metabolism in Neoplastic Progression.

Molecular chaperones have recently emerged as fundamental regulators of salient biological routines, including metabolic adaptations to environmental changes. Yet, many of the molecular mechanisms at the basis of their functions are still unknown or at least uncertain. This is in part due to the lack of chemical tools that can interact with the chaperones to induce measurable functional perturbations. In this context, the use of small molecules as modulators of protein functions has proven relevant for the investigation of a number of biomolecular systems. Herein, we focus on the functions, interactions and signaling pathways of the HSP90 family of molecular chaperones as possible targets for the discovery of new molecular entities aimed at tuning their activity and interactions. HSP90 and its mitochondrial paralog, TRAP1, regulate the activity of crucial metabolic circuitries, making cells capable of efficiently using available energy sources, with relevant implications both in healthy conditions and in a variety of disease states and especially cancer. The design of small-molecules targeting the chaperone cycle of HSP90 and able to inhibit or stimulate the activity of the protein can provide opportunities to finely dissect their biochemical activities and to obtain lead compounds to develop novel, mechanism-based drugs.

Theoretical Studies on the Selectivity Mechanisms of Glycogen Synthase Kinase 3ß (GSK3ß) with Pyrazine ATP-competitive Inhibitors by 3DQSAR, Molecular Docking, Molecular Dynamics Simulation and Free Energy Calculations.

BACKGROUND: Glycogen synthase kinase-3 (GSK3) is associated with various key biological processes and has been considered as an important therapeutic target for the treatment of many diseases. Great efforts have been made on the development of GSK3 inhibitors, especially ATP-competitive GSK3ß inhibitor, but it is still a great challenge to develop selective GSK3ß inhibitors because of the high sequence homology with other kinases. OBJECTIVE: In order to reveal the selectivity mechanisms of GSK3ß inhibition at the molecular level, a series of ATP-competitive GSK3ß inhibitor was analyzed by a systematic computational method, combining 3DQSAR, molecular docking, molecular dynamic simulations and free energy calculations. METHODS: Firstly, 3D-QSAR with CoMFA was built to explore the general structure activity relationships. Secondly, CDOCKER and Flexible docking were employed to predicted the reasonable docking poses of all studied inhibitors. And then, both GSK3ß and CDK2 complexes were selected to conduct molecular dynamics simulations. Finally, the free energy calculations were employed to find the key selective-residues. RESULTS: CoMFA model suggested the steric, hydrophobic fields play key roles in the bioactivities of inhibitors, and the binding mechanisms were well analyzed through molecular docking. The binding free energies predicted are in good agreement with the experimental bioactivities and the free energy calculations showed that the binding of GSK3ß/inhibitors was mainly contributed from hydrogen bonding and hydrophobic interaction. CONCLUSION: Some key residues for selective binding were highlighted, which may afford important guidance for the rational design of novel ATP-competitive GSK3ß inhibitors.

ATP-competitive DNA gyrase and topoisomerase IV inhibitors as antibacterial agents.

INTRODUCTION: The bacterial topoisomerases DNA gyrase and topoisomerase IV are validated targets for development of novel antibacterial agents. Fluoroquinolones inhibit the catalytic GyrA and/or ParC(GrlA) subunit and have been commonly used, although these have toxicity liabilities that restrict their use. The ATPase GyrB and ParE(GrlB) subunits have been much less explored and after withdrawal of novobiocin, there are no further marketed inhibitors . ATP-competitive inhibitors of GyrB and/or ParE(GrlB) are of special interest, as this target has been validated, and it is expected that many of the problems associated with fluoroquinolones can be avoided. AREAS COVERED: This review summarises the development of ATP-competitive inhibitors of GyrB and/or ParE(GrlB) as novel antibacterial agents over the last 10 years. Structural features of the new inhibitors and their optimisation strategies are highlighted. EXPERT OPINION: The development of novel ATP-competitive inhibitors of GyrB and/or ParE(GrlB) is ongoing in industrial and academical research. Development of resistance is one of the most problematic issues, but GyrB/ParE(GrlB) inhibitors do not show cross-resistance with fluoroquinolones. Other common issues, such as low solubility, high protein binding, development of off-target resistance, are related to the structures of the inhibitors themselves, which is thus a main focus of design strategies. With some now in early clinical development, there is reasonable expectation that novel ATP-competitive inhibitors of GyrB/ParE(GrlB) will reach the market in the near future.