Structure-based design and synthesis of potent benzothiazole inhibitors of interleukin-2 inducible T cell kinase (ITK)

MacKinnon, Colin H.; Lau, Kevin; Burch, Jason D.; Chen, Yuan; Dines, Jonathon A.; Ding, Xiao; Heifetz, Alexander; Jaochico, Allan; Johnson, Adam R.; Kraemer, Joachim; Kruger, Susanne; Krülle, Thomas M.; Liimatta, Marya; Ly, Justin Q.; Maghames, Rosemary; Montalbetti, Christian; Ortwine, Daniel F.; Perez‐Fuertes, Yolanda; Shia, S.; Stein, Daniel; Trani, Giancarlo; Vaidya, Darshan G.; Wang, Xiaolu; Bromidge, Steven M.; Wu, Lawren C.; Pei, Zhonghua

doi:10.1016/j.bmcl.2013.09.069

Cited by 15 publications

(20 citation statements)

References 21 publications

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“…The ITK kinase dimers, found in five different crystal forms in the BRAF ProtCID cluster, are a reasonable hypothesis for the mechanism of homodimerization of ITK at the membrane. These ITK dimers are not discussed or annotated by the authors of these structures [67][68][69][70][71][72][73][74] . crystal forms.…”

Section: Extending Dimers With Established Biological Activity To Othmentioning

confidence: 99%

ProtCID: A data resource for structural information on protein interactions

Dunbrack

2019

Preprint

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Structural information on the interactions of proteins with other molecules is plentiful, and for some proteins and protein families, there may be 100s of available structures. It can be very difficult for a scientist who is not trained in structural bioinformatics to access this information comprehensively. Previously, we developed the Protein Common Interface Database (ProtCID), which provided clusters of the interfaces of full-length protein chains as a means of identifying biological assemblies. Because proteins consist of domains that act as modular functional units, we have extended the analysis in ProtCID to the individual domain level. This has greatly increased the number of large protein-protein clusters in ProtCID, enabling the generation of hypotheses on the structures of biological assemblies of many systems. The analysis of domain families allows us to extend ProtCID to the interactions of domains with peptides, nucleic acids, and ligands. ProtCID provides complete annotations and coordinate sets for every cluster.

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Section: Extending Dimers With Established Biological Activity To Othmentioning

confidence: 99%

ProtCID: A data resource for structural information on protein interactions

Dunbrack

2019

Preprint

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“…Benzothiazole (BZT) ITK Inhibitors. As previously reported 29 the benzothiazole HTS hit, compound 2d (Table 2), had a moderate ITK K i of 460 nM. FMO analysis of the ITK−2d complex (Figure 6A) revealed six key interactions.…”

Section: T H Imentioning

confidence: 99%

“…Compounds 2p and 2q demonstrated 112-and 657-fold increases in ITK potency, respectively, over the original HTS hit (2d). 29 The existence of 10 interactions in 2q has been validated by the crystal structure of the ITK−2q complex (PDB entry 4MF1 29 ). FMO methodology was useful for exploring the interactions of 2d with ITK and identifying potential interactions that this compound lacked.…”

Section: T H Imentioning

confidence: 99%

Fragment Molecular Orbital Method Applied to Lead Optimization of Novel Interleukin-2 Inducible T-Cell Kinase (ITK) Inhibitors

et al. 2016

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Inhibition of inducible T-cell kinase (ITK), a nonreceptor tyrosine kinase, may represent a novel treatment for allergic asthma. In our previous reports, we described the discovery of sulfonylpyridine (SAP), benzothiazole (BZT), indazole (IND), and tetrahydroindazole (THI) series as novel ITK inhibitors and how computational tools such as dihedral scans and docking were used to support this process. X-ray crystallography and modeling were applied to provide essential insight into ITK-ligand interactions. However, "visual inspection" traditionally used for the rationalization of protein-ligand affinity cannot always explain the full complexity of the molecular interactions. The fragment molecular orbital (FMO) quantum-mechanical (QM) method provides a complete list of the interactions formed between the ligand and protein that are often omitted from traditional structure-based descriptions. FMO methodology was successfully used as part of a rational structure-based drug design effort to improve the ITK potency of high-throughput screening hits, ultimately delivering ligands with potency in the subnanomolar range.

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“…ITK-KD Mutant A was also used previously in our SHG assay. In the SHG assay, BMS-509744, 29 Pfizer compound 9, 21 and Pfizer compound 40 30 showed low signal intensities, whereas staurosporine, GSK compound 13, 31 Genentech compound 19, 32 Sanofi US compound, 34 and Genentech GNE-9822 33 showed high signal intensities. These results suggest that these active form inhibiters induced αC-helix-in conformation and oriented the SHG dye on αC-helix parallel to the z axis.…”

Section: Discussionmentioning

confidence: 99%

“…To our knowledge, no type II inhibitors that target Asp-Phe-Gly-out (DFG-out) conformation of ITK are known. 28 A comparison of the reported ITK crystal structures of inactive conformation (Protein Data Bank ID codes 3MJ2, 29 4M14, 21 and 4HCU 30 ) with those of active conformation (Protein Data Bank ID codes 4L7S, 31 4MF1, 32 and 4PQN 33 ) revealed that an αC-helix of inactive ITK rotates away from the catalytic center. We focused on movements of the αC-helix as an important determinant of inhibitor selectivity.…”

Section: Discussionmentioning

confidence: 99%

Identification of a New Inhibitor That Stabilizes Interleukin-2-Inducible T-Cell Kinase in Its Inactive Conformation

et al. 2019

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Interleukin-2-inducible T-cell kinase (ITK) plays an important role in T-cell signaling and is considered a promising drug target. As the ATP binding sites of protein kinases are highly conserved, the design of selective kinase inhibitors remains a challenge. Targeting inactive kinase conformations can address the issue of kinase inhibitor selectivity. It is important for selectivity considerations to identify compounds that stabilize inactive conformations from the primary screen hits. Here we screened a library of 390,000 compounds with an ADP-Glo assay using dephosphorylated ITK. After a surface plasmon resonance (SPR) assay was used to filter out promiscuous inhibitors, 105 hits were confirmed. Next, we used a fluorescent biosensor to enable the detection of conformational changes to identify inactive conformation inhibitors. A single-cysteine-substituted ITK mutant was labeled with acrylodan, and fluorescence emission was monitored. Using a fluorescent biosensor assay, we identified 34 inactive conformation inhibitors from SPR hits. Among them, one compound was bound to a site other than the ATP pocket and exhibited excellent selectivity against a kinase panel. Overall, (1) biochemical screening using dephosphorylated kinase, (2) hit confirmation by SPR assay, and (3) fluorescent biosensor assay that can distinguish inactive compounds provide a useful platform and offer opportunities to identify selective kinase inhibitors.

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Structure-based design and synthesis of potent benzothiazole inhibitors of interleukin-2 inducible T cell kinase (ITK)

Cited by 15 publications

References 21 publications

ProtCID: A data resource for structural information on protein interactions

ProtCID: A data resource for structural information on protein interactions

Fragment Molecular Orbital Method Applied to Lead Optimization of Novel Interleukin-2 Inducible T-Cell Kinase (ITK) Inhibitors

Identification of a New Inhibitor That Stabilizes Interleukin-2-Inducible T-Cell Kinase in Its Inactive Conformation

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