RESUMEN
Mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinase 2 (ERK2) and p38α MAP kinase (p38α MAPK), regulate various cellular responses. ERK2 is a drug target for treating many diseases, such as cancer, whereas p38α has attracted much attention as a promising drug target for treating inflammatory disorders. ERK2 is a critical off-target for p38α MAPK and vice versa. In this study, an allosteric ERK2 inhibitor with a benzothiazole moiety (compound 1) displayed comparable inhibitory activity against p38α MAPK. Crystal structures of these MAPKs showed that compound 1 bound to the allosteric site of ERK2 and p38α MAPK in distinct manners. Compound 1 formed a covalent bond with Cys162 of p38α MAPK, whereas this covalent bond was absent in the ERK2 complex even though the corresponding cysteine is conserved in ERK2. Structural dissection combined with computational simulations indicated that an amino acid difference in the allosteric site is responsible for the distinct binding modes of compound 1 with ERK2 and p38α MAPK. These structural insights underline the feasibility of developing highly selective and potent ERK2 and p38α MAPK inhibitors.
Asunto(s)
Proteína Quinasa 14 Activada por Mitógenos , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Benzotiazoles/farmacologíaRESUMEN
BACKGROUND: Endothelial cell activation is tightly controlled by the balance between VEGF (vascular endothelial cell growth factor) and Notch signaling pathway. VEGF destabilizes blood vessels and promotes neovascularization, which are common features of sight-threatening ocular vascular disorders. Here, we show that BCL6B (B-cell CLL/lymphoma 6 member B protein), also known as BAZF, ZBTB28, and ZNF62, plays a pivotal role in the development of retinal edema and neovascularization. METHODS: The pathophysiological physiological role of BCL6B was investigated in cellular and animal models mimicking 2 pathological conditions: retinal vein occlusion and choroidal neovascularization. An in vitro experimental system was used in which human retinal microvascular endothelial cells were supplemented with VEGF. Choroidal neovascularization cynomolgus monkey model was generated to investigate the involvement of BCL6B in the pathogenesis. Mice lacking BCL6B or treated with BCL6B-targeting small-interfering ribose nucleic acid were examined for histological and molecular phenotypes. RESULTS: In retinal endothelial cells, the BCL6B expression level was increased by VEGF. BCL6B-deficient endothelial cells showed Notch signal activation and attenuated cord formation via blockage of the VEGF-VEGFR2 signaling pathway. Optical coherence tomography images showed that choroidal neovascularization lesions were decreased by BCL6B-targeting small-interfering ribose nucleic acid. Although BCL6B mRNA expression was significantly increased in the retina, BCL6B-targeting small-interfering ribose nucleic acid suppressed ocular edema in the neuroretina. The increase in proangiogenic cytokines and breakdown of the inner blood-retinal barrier were abrogated in BCL6B knockout (KO) mice via Notch transcriptional activation by CBF1 (C promotor-binding factor 1) and its activator, the NICD (notch intracellular domain). Immunostaining showed that Müller cell activation, a source of VEGF, was diminished in BCL6B-KO retinas. CONCLUSIONS: These data indicate that BCL6B may be a novel therapeutic target for ocular vascular diseases characterized by ocular neovascularization and edema.
Asunto(s)
Neovascularización Coroidal , Ácidos Nucleicos , Neovascularización Retiniana , Enfermedades Vasculares , Animales , Humanos , Ratones , Neovascularización Coroidal/genética , Neovascularización Coroidal/metabolismo , Células Endoteliales/metabolismo , Macaca fascicularis/metabolismo , Ácidos Nucleicos/metabolismo , Ácidos Nucleicos/uso terapéutico , Neovascularización Retiniana/genética , Neovascularización Retiniana/metabolismo , Ribosa/metabolismo , Ribosa/uso terapéutico , Enfermedades Vasculares/patología , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Mitogen-activated protein kinase kinases (MAP2Ks) 1, 4, and 7 are potential targets for treating various diseases. Here, we solved the crystal structures of MAP2K1 and MAP2K4 complexed with covalent inhibitor 5Z-7-oxozeaenol (5Z7O). The elucidated structures showed that 5Z7O was non-covalently bound to the ATP binding site of MAP2K4, while it covalently attached to cysteine at the DFG-1 position of the deep ATP site of MAP2K1. In contrast, we previously showed that 5Z7O covalently binds to MAP2K7 via another cysteine on the solvent-accessible edge of the ATP site. Structural analyses and molecular dynamics calculations indicated that the configuration and mobility of conserved gatekeeper methionine located at the central ATP site regulated the binding and access of 5Z7O to the ATP site of MAP2Ks. These structural features provide clues for developing highly potent and selective inhibitors against MAP2Ks. Abbreviations: ATP, adenosine triphosphate; FDA, Food and Drug Administration; MAP2Ks, mitogen-activated protein kinase kinases; MD, molecular dynamics; NSCLC, non-small cell lung cancer; 5Z7O, 5Z-7-oxozeaenol; PDB, protein data bank; RMSD, root-mean-square deviation.
RESUMEN
TGF-ß is an immunosuppressive cytokine and plays a key role in progression of cancer by inducing immunosuppression in tumor microenvironment. Therefore, inhibition of TGF-ß signaling pathway may provide a potential therapeutic intervention in treating cancers. Herein, we report the discovery of a series of novel thiazole derivatives as potent inhibitors of ALK5, a serine-threonine kinase which is responsible for TGF-ß signal transduction. Compound 29b was identified as a potent inhibitor of ALK5 with an IC50 value of 3.7 nM with an excellent kinase selectivity.
Asunto(s)
Diseño de Fármacos , Inhibidores de Proteínas Quinasas , Proteínas Serina-Treonina Quinasas , Receptor Tipo I de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta , Tiazoles , Tiazoles/química , Tiazoles/farmacología , Tiazoles/síntesis química , Receptor Tipo I de Factor de Crecimiento Transformador beta/antagonistas & inhibidores , Receptor Tipo I de Factor de Crecimiento Transformador beta/metabolismo , Humanos , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Relación Estructura-Actividad , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/metabolismo , Receptores de Factores de Crecimiento Transformadores beta/antagonistas & inhibidores , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Estructura Molecular , Relación Dosis-Respuesta a DrogaRESUMEN
Extracellular signal-regulated kinase 2 (ERK2), a mitogen-activated protein kinase (MAPK), plays an essential role in physiological cellular processes and is a drug target for treating cancers and type 2 diabetes. A previous in silico screening study focusing on an allosteric site that plays a crucial role in substrate anchoring conferred an ERK2 inhibitor (compound 1). In this report, compound 1 was found to show high selectivity toward ERK2 compared with the nearest off-target p38α MAPK, and the crystal structure revealed that compound 1 binds to the allosteric site of ERK2. Fragment molecular orbital calculations based upon this crystal structure provided the structural basis to improve potency of compound 1 derivatives. Further computational studies uncovered that the low entropic cost of binding conferred the high selectivity of compound 1 toward ERK2 over p38α MAPK. These findings demonstrate the feasibility of developing potent and selective ERK2 inhibitors.
Asunto(s)
Diabetes Mellitus Tipo 2 , Proteína Quinasa 1 Activada por Mitógenos , Humanos , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Sitio AlostéricoRESUMEN
Extracellular signal-regulated kinase 2 (ERK2) controls vital physiological processes involving proliferation and differentiation and is a drug target molecule for many diseases such as cancers. In silico screening focusing on an allosteric site that plays a crucial role in substrate anchoring conferred an ERK2 inhibitor (compound 1). However, a competitive binding assay indicated that compound 1 did not bind to the allosteric site. Here, the crystal structure of ERK2 in complex with compound 1 revealed a novel binding site. This finding demonstrates the feasibility of developing new types of ERK2 inhibitors.
Asunto(s)
Adenosina Trifosfato/metabolismo , Proteína Quinasa 1 Activada por Mitógenos/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Sitio Alostérico , Sitios de Unión , Unión Competitiva , Cristalografía por Rayos X , Humanos , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Fosforilación , Conformación ProteicaRESUMEN
Casein kinase 2 (CK2) is a vital protein kinase that consists of two catalytic subunits (CK2α1 and/or CK2α2) and two regulatory subunits (CK2ß). CK2α1 is a drug target for nephritis and cancers, while CK2α2 is a serious off-target because its inhibition causes testicular toxicity. High similarity between the isozymes CK2α1 and CK2α2 make it difficult to design CK2α1-specific inhibitors. Herein, the crystal structures of CK2α1 and CK2α2 complexed with a 3-amino-pyrazole inhibitor revealed the remarkable differences in the protein-inhibitor interaction modes. This inhibitor bound to the ATP binding sites of both isozymes in apparently distinct orientations. In addition, another molecule of this inhibitor bound to CK2α1, but not to CK2α2, at the CK2ß protein-protein interface. Binding energy calculations and biochemical experiments suggested that this inhibitor possesses the conventional ATP-competitive characteristics with moderate allosteric function in a molecular glue mechanism. These results will assist the potential design of potent and selective CK2α1 inhibitors.
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Quinasa de la Caseína II , Isoenzimas , Pirazoles/farmacología , Adenosina Trifosfato , Quinasa de la Caseína II/metabolismo , Isoenzimas/metabolismo , Inhibidores de Proteínas Quinasas/farmacologíaRESUMEN
BACKGROUND: Metastasis is the primary cause of death in cancer patients, and its management is still a major challenge. Epithelial to mesenchymal transition (EMT) has been implicated in the process of cancer metastasis, and its pharmacological interference holds therapeutic promise. METHODS: Traf2- and Nck-interacting kinase (TNIK) functions as a transcriptional coregulator of Wnt target genes. Given the convergence of Wnt and transforming growth factor-ß (TGFß) signalling, we examined the effects of a small-molecule TNIK inhibitor (named NCB-0846) on the TGFß1-induced EMT of lung cancer cells. RESULTS: NCB-0846 inhibited the TGFß1-induced EMT of A549 cells. This inhibition was associated with inhibition of Sma- and Mad-Related Protein-2/3 (SMAD2/3) phosphorylation and nuclear translocation. NCB-0846 abolished the lung metastasis of TGFß1-treated A549 cells injected into the tail veins of immunodeficient mice. The inhibition of EMT was mediated by suppression of the TGFß receptor type-I (TGFBR1) gene, at least partly through the induction of microRNAs targeting the TGFBR1 transcript [miR-320 (a, b and d) and miR-186]. CONCLUSIONS: NCB-0846 pharmacologically blocks the TGFß/SMAD signalling and EMT induction of lung cancer cells by transcriptionally downregulating TGFBRI expression, representing a potentially promising approach for prevention of metastasis in lung cancer patients.
Asunto(s)
Transición Epitelial-Mesenquimal/efectos de los fármacos , Neoplasias Pulmonares , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Factor de Crecimiento Transformador beta1/metabolismo , Células A549 , Animales , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Ratones , Transducción de Señal/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
p38α mitogen activated protein kinase (MAPK) plays important roles in multiple cellular functions by phosphorylating a wide variety of substrates, and therefore, p38α MAPK has been considered as an important drug target. In this study, we designed peptide-based inhibitors for p38α MAPK, which can only inhibit the Smad3 phosphorylation specifically, by targeting the KIM binding site of p38α MAPK. Peptide 6 showed a significant inhibitory potency for the Smad3 phosphorylation by p38α MAPK. Peptide 6 showed no ATP dependency, and did not inhibit the phosphorylation of other substrates by p38α MAPK. The discovery of peptide 6 by targeting the KIM binding site likely provide an opportunity for the discovery of a novel class of allosteric and substrate-specific p38α MAPK inhibitors.
Asunto(s)
Péptidos/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Proteína smad3/antagonistas & inhibidores , Proteínas Quinasas p38 Activadas por Mitógenos/antagonistas & inhibidores , Regulación Alostérica/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Humanos , Modelos Moleculares , Estructura Molecular , Péptidos/síntesis química , Péptidos/química , Fosforilación/efectos de los fármacos , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Proteína smad3/metabolismo , Relación Estructura-Actividad , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Mitogen-activated protein kinase kinase 4 (MAP2K4) plays a critical role in regulating the stress-activated protein kinase signaling cascade. A small angle X-ray scattering experiment, a powerful technique for analyzing a solution structure cleared from the structural artifacts due to crystal packing, provided the ensemble structures of human non-phosphorylated MAP2K4 in three states involving the apo form, the binary complex with an ATP analogue, and the ternary complex with the ATP analogue and substrate peptide. These ensemble structures provided more detailed mechanisms for regulating MAP2K4 in addition to those delineated only by the crystal structures in three states.
Asunto(s)
MAP Quinasa Quinasa 4/análisis , MAP Quinasa Quinasa 4/metabolismo , Adenosina Trifosfato/química , Adenosina Trifosfato/metabolismo , Humanos , Conformación Proteica , Dispersión del Ángulo Pequeño , Difracción de Rayos XRESUMEN
The production of TrkA-selective inhibitors is considerably difficult because the kinase domains of TrkA and its isoforms TrkB/C have highly homologous amino acid sequences. Here we describe the structural basis for the acquisition of selectivity for a isoform-selective TrkA inhibitor, namely compound V1. The X-ray structure revealed that V1 acts as a molecular glue to stabilize the symmetrical dimer of the TrkA kinase domains. V1 binds to the ATP-binding site and simultaneously engages in the dimeric interface of TrkA. The region of the dimeric interface in TrkA is not conserved in TrkB/C; thus, dimer formation may be a novel mechanism for the production of selective TrkA inhibitors. The biochemical and biophysical assay results confirmed that V1 selectively inhibited TrkA and induced the dimer formation of TrkA, but not TrkB. The binding pocket at the TrkA dimer interface can be used for the production of new isoform-selective TrkA inhibitors.
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Isoformas de Proteínas/metabolismo , Receptor trkA/metabolismo , Secuencia de Aminoácidos , Humanos , Modelos MolecularesRESUMEN
BTK is a promising target for the treatment of multiple diseases such as B cell malignances, asthma, and rheumatoid arthritis. Here, we report the discovery of a series of novel pyrimidine analogs as potent, highly selective, non-covalent inhibitors of BTK. Compound 25d demonstrated higher affinity to an unactivated conformation of BTK that resulted in an excellent kinase selectivity. Compound 25d showed a good oral bioavailability in mice, and significantly inhibits the PCA reaction in mice.
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Diseño de Fármacos , Anafilaxis Cutánea Pasiva/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Pirimidinas/farmacología , Administración Oral , Animales , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Canales de Potasio Éter-A-Go-Go/antagonistas & inhibidores , Canales de Potasio Éter-A-Go-Go/metabolismo , Humanos , Ratones , Estructura Molecular , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/química , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/metabolismo , Pirimidinas/administración & dosificación , Pirimidinas/química , Relación Estructura-ActividadRESUMEN
The majority of kinase inhibitors have been developed as ATP competitors to interact with a hinge region in ATP binding sites of kinases. 7-Azaindole has been found as an excellent hinge binding motif by making two hydrogen bonds with the kinase hinge region. Vemurafenib, a B-RAF kinase (serine-threonine kinase [STK]) inhibitor approved by the U.S. Food and Drug Administration (FDA) for the treatment of melanoma, was created from this simple 7-azaindole fragment by successful use of structure-based drug design techniques. The huge potential of 7-azaindole as a hinge-binding motif has encouraged many researchers to employ it as a kinase privileged fragment. This paper will review recent examples of 7-azaindole-based kinase inhibitors, and discusses their binding interactions with the kinase hinge regions.
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Indoles/química , Indoles/farmacología , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Animales , Humanos , Inhibidores de Proteínas Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismoRESUMEN
Mitogen-activated protein kinase kinase 7 (MAP2K7) regulates stress and inflammatory responses, and is an attractive drug discovery target for several diseases including arthritis and cardiac hypertrophy. Intracellular proteins such as MAP2K7 are prone to aggregation due to cysteine-driven oxidation in in vitro experiments. MAP2K7 instability due to the four free cysteine residues on the molecular surface abrogated the crystal growth and led to a low-resolution structure with large residual errors. To acquire a higher resolution structure for promoting rational drug discovery, we explored stable mutants of MAP2K7 by replacing the surface cysteine residues, Cys147, Cys218, Cys276 and Cys296. Single-site mutations, except for Cys147, maintained the specific activity and increased the protein yield, while all the multi-site mutations massively reduced the activity. The C218S mutation drastically augmented the protein production and crystallographic resolution. Furthermore, the C218S crystals grown under microgravity in a space environment yielded a 1.3 Å resolution structure, providing novel insights for drug discovery: the precisely assigned water molecules in the active site, the double conformations in the flexible region and the C-terminal extension bound to the N-terminal region of the adjacent molecules. The latter insight is likely to promote the production of allosteric MAP2K7 inhibitors.
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MAP Quinasa Quinasa 7/química , MAP Quinasa Quinasa 7/ultraestructura , Regulación Alostérica , Sitios de Unión , Simulación por Computador , Activación Enzimática , Modelos Químicos , Modelos Moleculares , Unión Proteica , Conformación Proteica , Relación Estructura-ActividadRESUMEN
Mitogen-activated protein kinase kinase 7 (MAP2K7) is an indispensable kinase of the c-Jun N-terminal kinase signal cascade and is rigorously regulated via phosphorylation. To investigate the regulatory mechanism of the inactive non-phosphorylated state of MAP2K7, the crystal structures of the wild-type and C218S mutant were solved. The wild-type apo-structure revealed an unprecedented auto-inhibition form that occluded the ATP site. This closed form was configured by the n-σ* interaction of Cys218, a non-conserved residue among the MAP2K family kinases, with Gly145 in the glycine-rich loop. The interaction was unaltered in the presence of an ATP analog, whereas the C218S mutation precluded the closed configuration. These structural insights are potentially valuable for drug discovery of highly selective MAP2K7 inhibitors.
Asunto(s)
MAP Quinasa Quinasa 7/química , MAP Quinasa Quinasa 7/metabolismo , Adenosina Trifosfato/metabolismo , Cristalografía por Rayos X , Humanos , MAP Quinasa Quinasa 7/genética , Modelos Moleculares , Mutación Puntual , Conformación ProteicaRESUMEN
Bruton's Tyrosine Kinase (BTK) is one of the crucial kinases for the B cell maturation and mast cell activation, and specific inhibitors of BTK are considered to be attractive targets in drug discovery research. In this Letter, we have designed and synthesized a new fluorescent probe for TR-FRET-based high-throughput screening, to identify compounds that preferentially bind to an inactive conformation of BTK which has a unique structural feature. A set of kinase-focused compound library was screened using this assay method, and compound 31 was successfully identified as a potent inhibitor which preferentially bind to the inactive conformation of BTK. These results suggest that this screening method has a great potential for the discovery of novel selective BTK inhibitors.
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Bioensayo/métodos , Colorantes Fluorescentes/química , Inhibidores de Proteínas Quinasas/aislamiento & purificación , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Agammaglobulinemia Tirosina Quinasa , Unión Competitiva , Sistemas de Liberación de Medicamentos , Diseño de Fármacos , Activación Enzimática/efectos de los fármacos , Concentración 50 Inhibidora , Modelos Biológicos , Unión Proteica , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Estructura Terciaria de ProteínaRESUMEN
BTK has emerged as a promising target for treating B-cell malignancies and autoimmune diseases, and there has been a growing demand to identify selective BTK inhibitors efficiently. In this Letter, we have designed and synthesized a new fluorescent probe to screen compounds that preferentially bind to an unactivated state of BTK (BTK [U]). The fluorescence of the probe was turned on in the presence of BTK [U], and quenched by the addition of compounds which preferentially bind to BTK [U]. This unique fluorescent probe was successfully applied to the screening of a kinase focused compound library. The results suggest that this new method is a simple and easy-to-perform assay to screen inhibitors of BTK [U].
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Colorantes Fluorescentes/química , Proteínas Tirosina Quinasas/análisis , Agammaglobulinemia Tirosina Quinasa , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Tirosina Quinasas/antagonistas & inhibidoresRESUMEN
5Z-7-Oxozeaenol (5Z7O) is a covalent bonding inhibitor against the several protein kinases (e.g., ERK2 and TAK1) that possess a free cysteine at the gatekeeper-2 position. In addition to this cysteine, MAP2K7 has three other cysteine residues that are candidate for covalent bonding by the inhibitor 5Z7O. The crystal structure of the MAP2K7/5Z7O complex revealed that the inhibitor binds to MAP2K7 at a cysteine residue located at the end of the hinge region and not at the gatekeeper-2 residue. The structural insights into the interaction of 5Z7O with MAP2K7 should aid the development of 5Z7O derivatives with improved potency and selectivity.
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Cisteína/química , MAP Quinasa Quinasa 7/química , Zearalenona/análogos & derivados , Adenosina Trifosfato/química , Adenosina Trifosfato/metabolismo , Sitios de Unión , MAP Quinasa Quinasa 7/metabolismo , Proteína Quinasa 1 Activada por Mitógenos/química , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Simulación de Dinámica Molecular , Estructura Terciaria de Proteína , Zearalenona/químicaRESUMEN
Stimulator of interferon genes (STING) is critical for the type I interferon response to pathogen- or self-derived DNA in the cytosol. STING may function as a scaffold to activate TANK-binding kinase 1 (TBK1), but direct cellular evidence remains lacking. Here we show, using single-molecule imaging of STING with enhanced time resolutions down to 5 ms, that STING becomes clustered at the trans-Golgi network (about 20 STING molecules per cluster). The clustering requires STING palmitoylation and the Golgi lipid order defined by cholesterol. Single-molecule imaging of TBK1 reveals that STING clustering enhances the association with TBK1. We thus provide quantitative proof-of-principle for the signaling STING scaffold, reveal the mechanistic role of STING palmitoylation in the STING activation, and resolve the long-standing question of the requirement of STING translocation for triggering the innate immune signaling.
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Lipoilación , Red trans-Golgi , Red trans-Golgi/metabolismo , Microscopía , Imagen Individual de Molécula , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Colesterol , Análisis por Conglomerados , Inmunidad InnataRESUMEN
INTRODUCTION: CDC7 is a serine/threonine kinase which plays an important role in DNA replication. Inhibition of CDC7 in cancer cells causes lethal S phase or M phase progression, whereas inhibition of CDC7 in normal cells does not cause cell death and only leads to cell cycle arrest at the DNA replication checkpoint. Therefore, CDC7 has been recognized as a potential target for novel therapeutic interventions in cancers. AREAS COVERED: Patent literature claiming novel small molecule compounds inhibiting CDC7 disclosed from 2017 to 2022. EXPERT OPINION: Despite the indisputable positive impact of CDC7 as a drug target, there have been reported only a handful of chemical scaffolds as CDC7 inhibitors. Several CDC7 inhibitors have been progressed into clinical trials for cancer treatments, but they did not result in satisfactory efficacies in those trials. One possible reason for the failure might be due to the dose-limiting toxicities, and some of the observed toxicities were thought to be not related to CDC7 inhibition, suggesting it should be important to identify novel chemical scaffolds to eliminate unwanted toxicities. Another important factor is the patient stratification that would enable greater response, and the identification of such predictive biomarkers should be the key to success for the development of CDC7 inhibitors.