Perspectives for the use of structural information and chemical genetics to develop inhibitors of Janus kinases

Haan, Claude; Behrmann, Iris; Haan, Serge

doi:10.1111/j.1582-4934.2010.01018.x

Cited by 55 publications

(81 citation statements)

References 276 publications

(309 reference statements)

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“…disease-causing JH1 and JH2 mutations localize in or near the JH2-JH1 interface and are expected to destabilize the interaction (13,18). The structures of JAK1 and TYK2 JH2 are highly similar to JAK2 JH2 (5,19), and all three JH2s bind ATP (20).…”

Section: Significancementioning

confidence: 90%

“…The V617F mutation is found in ∼95% of patients with polycythemia vera (PV) (9-12) as well as in ∼60% of patients with essential thrombocythemia (ET) and primary myelofibrosis (PMF). After identification of the V617F mutation, a multitude of other mutations in JAK2, JAK1, and JAK3 have been found that are linked to myeloid and lymphoid malignancies and to immunological diseases as well as to some solid cancers (13,14). The mutations cluster mainly in exon 12 in the SH2-JH2 linker (numbering for human JAK2), exon 14 near Val617, and exon 16 (13).…”

mentioning

confidence: 99%

“…After identification of the V617F mutation, a multitude of other mutations in JAK2, JAK1, and JAK3 have been found that are linked to myeloid and lymphoid malignancies and to immunological diseases as well as to some solid cancers (13,14). The mutations cluster mainly in exon 12 in the SH2-JH2 linker (numbering for human JAK2), exon 14 near Val617, and exon 16 (13). Although most JAK JH2 mutations are gain-of-function, some JH2 mutations in JAK3 suppress JH1 activity, leading to severe combined immunodeficiency (2,15).…”

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confidence: 99%

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ATP binding to the pseudokinase domain of JAK2 is critical for pathogenic activation

Hammarén

Ungureanu

Grisouard

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

123

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Pseudokinases lack conserved motifs typically required for kinase activity. Nearly half of pseudokinases bind ATP, but only few retain phosphotransfer activity, leaving the functional role of nucleotide binding in most cases unknown. Janus kinases (JAKs) are nonreceptor tyrosine kinases with a tandem pseudokinase-kinase domain configuration, where the pseudokinase domain (JAK homology 2, JH2) has important regulatory functions and harbors mutations underlying hematological and immunological diseases. JH2 of JAK1, JAK2, and TYK2 all bind ATP, but the significance of this is unclear. We characterize the role of nucleotide binding in normal and pathogenic JAK signaling using comprehensive structure-based mutagenesis. Disruption of JH2 ATP binding in wildtype JAK2 has only minor effects, and in the presence of type I cytokine receptors, the mutations do not affect JAK2 activation. However, JH2 mutants devoid of ATP binding ameliorate the hyperactivation of JAK2 V617F. Disrupting ATP binding in JH2 also inhibits the hyperactivity of other pathogenic JAK2 mutants, as well as of JAK1 V658F, and prevents induction of erythrocytosis in a JAK2 V617F myeloproliferative neoplasm mouse model. Molecular dynamic simulations and thermal-shift analysis indicate that ATP binding stabilizes JH2, with a pronounced effect on the C helix region, which plays a critical role in pathogenic activation of JAK2. Taken together, our results suggest that ATP binding to JH2 serves a structural role in JAKs, which is required for aberrant activity of pathogenic JAK mutants. The inhibitory effect of abrogating JH2 ATP binding in pathogenic JAK mutants may warrant novel therapeutic approaches.T he Janus kinases (JAK1-3, TYK2) are a family of nonreceptor tyrosine kinases with essential functions in the regulation of hematopoiesis, the immune system, and cellular metabolism. JAKs interact specifically with various cytokine receptors and couple cytokine binding to cytoplasmic signaling cascades, including the signal transducers and activators of transcription (STAT) pathway. JAKs consist of an N-terminal FERM domain, an SH2-like (Src homology 2) domain, a pseudokinase domain (JAK homology 2, JH2), and the C-terminal tyrosine kinase domain (JH1). JH2 mediates critical regulatory functions in JAKs and primarily serves to inhibit basal JH1 activity. Experimental deletion of JH2 increases JH1 activity in full-length JAK in the absence of stimulation (1-3), and in recombinant systems addition of JH2 suppresses JH1 activity (4-6). JH2 is, however, also required for ligand-induced activation of full-length JAKs in cell (1-3, 6, 7). The regulatory functions of JH2 are corroborated by the multitude of human disease mutations identified in the domain. The most common JAK2 mutation, V617F, leads to cytokine-independent signaling through the exclusively JAK2-dependent homotypic receptors for erythropoietin (EPO), granulocyte colony stimulating factor (G-CSF), and thrombopoietin (8). The V617F mutation is found in ∼95% of patients with polycythemia vera (PV) (9...

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Section: Significancementioning

confidence: 90%

mentioning

confidence: 99%

mentioning

confidence: 99%

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ATP binding to the pseudokinase domain of JAK2 is critical for pathogenic activation

Hammarén

Ungureanu

Grisouard

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

123

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“…As in other protein kinases, ATP binding may prime for critical conformational changes (40,41). Indeed, structural studies showed that the isolated JH1 domains of the four Jaks complexed with ATP-competitive inhibitors adopt an active conformation with an exposed loop (42)(43)(44). With regard to intact Jaks, specific ATP competitive inhibitors that block catalytic function also promote activation loop phosphorylation in cells (45).…”

Section: Discussionmentioning

confidence: 99%

Two Rare Disease-Associated Tyk2 Variants Are Catalytically Impaired but Signaling Competent

Gakovic

Ragimbeau

et al. 2013

The Journal of Immunology

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Tyk2 belongs to the Janus protein tyrosine kinase family and is involved in signaling of immunoregulatory cytokines (type I and III IFNs, IL-6, IL-10, and IL-12 families) via its interaction with shared receptor subunits. Depending on the receptor complex, Tyk2 is coactivated with either Jak1 or Jak2, but a detailed molecular characterization of the interplay between the two enzymes is missing. In human populations, the Tyk2 gene presents high levels of genetic diversity with >100 nonsynonymous variants being detected. In this study, we characterized two rare Tyk2 variants, I684S and P1104A, which have been associated with susceptibility to autoimmune disease. Specifically, we measured their in vitro catalytic activity and their ability to mediate Stat activation in fibroblasts and genotyped B cell lines. Both variants were found to be catalytically impaired but rescued signaling in response to IFN-α/β, IL-6, and IL-10. These data, coupled with functional study of an engineered Jak1 P1084A, support a model of nonhierarchical activation of Janus kinases in which one catalytically competent Jak is sufficient for signaling provided that its partner behaves as proper scaffold, even if inactive. Through the analysis of IFN-α and IFN-γ signaling in cells with different Jak1 P1084A levels, we also illustrate a context in which a hypomorphic Jak can hamper signaling in a cytokine-specific manner. Given the multitude of Tyk2-activating cytokines, the cell context–dependent requirement for Tyk2 and the catalytic defect of the two disease-associated variants studied in this paper, we predict that these alleles are functionally significant in complex immune disorders.

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“…8 Currently several JAK2 inhibitors are under development or/and undergoing clinical trials for MPNs. 27 Identification of JAK inhibitor-resistant mutations in JAK2 is, therefore, of great importance. We, therefore, introduced the Y931C, a homologous mutation of JAK1 F958C, into WT or V617F JAK2 protein and selected these cells for autonomous proliferation.…”

Section: Jak2 Y931c the Homologous Mutation Of Jak1 F958c Confers Rmentioning

confidence: 99%

Oncogenic JAK1 and JAK2-activating mutations resistant to ATP-competitive inhibitors

Hornakova¹,

Springuel²,

Devreux³

et al. 2011

Haematologica

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BackgroundActivating mutations in JAK1 and JAK2 have been described in patients with various hematologic malignancies including acute lymphoblastic leukemia and myeloproliferative neoplasms, leading to clinical trials with JAK inhibitors. While there has been a tremendous effort towards the development of specific JAK inhibitors, mutations conferring resistance to such drugs have not yet been observed. Design and MethodsTaking advantage of a model of spontaneous cellular transformation, we sequenced JAK1 in selected tumorigenic BaF3 clones and identified 25 de novo JAK1 activating mutations, including 5 mutations already described in human leukemias. We further used this library of JAK1 mutation-positive cell lines to assess their sensitivity to ATP-competitive inhibitors. ResultsWhile most JAK1 mutants were sensitive to ATP-competitive JAK inhibitors, mutations targeting Phe958 and Pro960 in the hinge region of the kinase domain rendered JAK1 constitutively active but also resistant to all tested JAK inhibitors. Furthermore, mutation of the homologous Tyr931 in JAK2 wild-type or JAK2 V617F mutant found in patients with myeloproliferative neoplasms also conferred resistance to JAK inhibitors, such as INCB018424, which is currently in clinical use. ConclusionsOur data indicate that some activating mutations not only promote autonomous cell proliferation but also confer resistance to ATP-competitive inhibitors. In vivo, such a mutation can potentially occur as primary JAK-activating mutations but also as secondary mutations combining oncogenicity with drug resistance.Key words: JAK, tyrosine kinase, mutations, inhibitors, STAT.Citation: Hornakova T, Springuel L, Devreux J, Dusa A, Constantinescu SN, Knoops L, and Renauld J-C. Oncogenic JAK1 and JAK2-activating mutations resistant to ATP-competitive inhibitors. Haematologica 2011;96(6):845-853. doi:10.3324/haematol.2010 This is an open-access paper. Oncogenic JAK1 and JAK2-activating mutations resistant to ATP-competitive inhibitors

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Perspectives for the use of structural information and chemical genetics to develop inhibitors of Janus kinases

Cited by 55 publications

References 276 publications

ATP binding to the pseudokinase domain of JAK2 is critical for pathogenic activation

ATP binding to the pseudokinase domain of JAK2 is critical for pathogenic activation

Two Rare Disease-Associated Tyk2 Variants Are Catalytically Impaired but Signaling Competent

Oncogenic JAK1 and JAK2-activating mutations resistant to ATP-competitive inhibitors

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