Translational Therapeutics

Inhibition of the deubiquitinase USP10 induces degradation of SYK

Abstract

Background

There is growing evidence that spleen tyrosine kinase (SYK) is critical for acute myeloid leukaemia (AML) transformation and maintenance of the leukemic clone in AML patients. It has also been found to be over-expressed in AML patients, with activating mutations in foetal liver tyrosine kinase 3 (FLT3), particularly those with internal tandem duplications (FLT3-ITD), where it transactivates FLT3-ITD and confers resistance to treatment with FLT3 tyrosine kinase inhibitors (TKIs).

Methods

We have previously described a pharmacological approach to treating FLT3-ITD-positive AML that relies on proteasome-mediated FLT3 degradation via inhibition of USP10, the deubiquitinating enzyme (DUB) responsible for cleaving ubiquitin from FLT3.

Results

Here, we show that USP10 is also a major DUB required for stabilisation of SYK. We further demonstrate that degradation of SYK can be induced by USP10-targeting inhibitors. USP10 inhibition leads to death of cells driven by active SYK or oncogenic FLT3 and potentiates the anti-leukemic effects of FLT3 inhibition in these cells.

Conclusions

We suggest that USP10 inhibition is a novel approach to inhibiting SYK and impeding its role in the pathology of AML, including oncogenic FLT3-positive AML. Also, given the significant transforming role SYK in other tumours, targeting USP10 may have broader applications in cancer.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1: Small molecule inhibition of USP10 leads to degradation of SYK and targeted killing of activated SYK-driven cells.
Fig. 2: HBX19818 analogues have differential effects on SYK protein levels in Ba/F3-FLT3-ITD cells.
Fig. 3: USP10 and SYK physically associate and genetic knockdown or knockout of USP10 leads to SYK degradation.
Fig. 4: USP10 KD or small molecule inhibition leads to ubiquitination and degradation of SYK and increased total cellular ubiquitination.
Fig. 5: Loss of USP10 shortens the half-life of SYK protein with no effect on SYK mRNA transcription.
Fig. 6: Midostaurin potentiates the effects of USP10 inhibitors against Ba/F3-SYK-TEL cells.

References

  1. 1.

    Carnevale, J., Ross, L., Puissant, A., Banerji, V., Stone, R. M., DeAngelo, D. J. et al. SYK regulates mTOR signaling in AML. Leukemia 27, 2118–2128 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  2. 2.

    Dennehy, K. M., Ferwerda, G., Faro-Trindade, I., Pyz, E., Willment, J. A., Taylor, P. R. et al. Syk kinase is required for collaborative cytokine production induced through Dectin-1 and Toll-like receptors. Eur. J. Immunol. 38, 500–506 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  3. 3.

    Rinaldi, A., Kwee, I., Taborelli, M., Largo, C., Uccella, S., Martin, V. et al. Genomic and expression profiling identifies the B-cell associated tyrosine kinase Syk as a possible therapeutic target in mantle cell lymphoma. Br. J. Haematol. 132, 303–316 (2006).

    CAS  PubMed  Google Scholar 

  4. 4.

    Chen, L., Monti, S., Juszczynski, P., Daley, J., Chen, W., Witzig, T. E. et al. SYK-dependent tonic B-cell receptor signaling is a rational treatment target in diffuse large B-cell lymphoma. Blood 111, 2230 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  5. 5.

    Buchner, M., Fuchs, S., Prinz, G., Pfeifer, D., Bartholomé, K., Burger, M. et al. Spleen tyrosine kinase is overexpressed and represents a potential therapeutic target in chronic lymphocytic leukemia. Cancer Res. 69, 5424 (2009).

    CAS  PubMed  Google Scholar 

  6. 6.

    Puissant, A., Fenouille, N., Alexe, G., Pikman, Y., Bassil, C. F., Mehta, S. et al. SYK is a critical regulator of FLT3 in acute myeloid leukemia. Cancer Cell 25, 226–242 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  7. 7.

    Weisberg, E. L., Puissant, A., Stone, R., Sattler, M., Buhrlage, S. J., Yang, J. et al. Characterization of midostaurin as a dual inhibitor of FLT3 and SYK and potentiation of FLT3 inhibition against FLT3-ITD-driven leukemia harboring activated SYK kinase. Oncotarget 8, 52026–52044 (2017).

    PubMed  PubMed Central  Google Scholar 

  8. 8.

    Nijman, S. M., Luna-Vargas, M. P., Velds, A., Brummelkamp, T. R., Dirac, A. M., Sixma, T. K. et al. A genomic and functional inventory of deubiquitinating enzymes. Cell 123, 773–786 (2005).

    CAS  PubMed  Google Scholar 

  9. 9.

    Fraile, J. M., Quesada, V., Rodriguez, D., Freije, J. M. & Lopez-Otin, C. Deubiquitinases in cancer: new functions and therapeutic options. Oncogene 31, 2373–2388 (2012).

    CAS  PubMed  Google Scholar 

  10. 10.

    Jacq, X., Kemp, M., Martin, N. M. & Jackson, S. P. Deubiquitylating enzymes and DNA damage response pathways. Cell Biochem. Biophys. 67, 25–43 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  11. 11.

    Ristic, G., Tsou, W.-L. & Todi, S. V. An optimal ubiquitin-proteasome pathway in the nervous system: the role of deubiquitinating enzymes. Front. Mol. Neurosci. 7, 72–72 (2014).

    PubMed  PubMed Central  Google Scholar 

  12. 12.

    Ebner, P., Versteeg, G. A. & Ikeda, F. Ubiquitin enzymes in the regulation of immune responses. Crit. Rev. Biochem Mol. Biol. 52, 425–460 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  13. 13.

    Mofers, A., Pellegrini, P., Linder, S. & D’Arcy, P. Proteasome-associated deubiquitinases and cancer. Cancer Metastasis Rev. 36, 635–653 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. 14.

    Sacco, J. J., Coulson, J. M., Clague, M. J. & Urbe, S. Emerging roles of deubiquitinases in cancer-associated pathways. IUBMB Life 62, 140–157 (2010).

    CAS  PubMed  Google Scholar 

  15. 15.

    Katkere, B., Rosa, S. & Drake, J. R. The Syk-binding ubiquitin ligase c-Cbl mediates signaling-dependent B cell receptor ubiquitination and B cell receptor-mediated antigen processing and presentation. J. Biol. Chem. 287, 16636–16644 (2012). e-pub ahead of print 2012/03/28.

    CAS  PubMed  PubMed Central  Google Scholar 

  16. 16.

    Mohapatra, B., Ahmad, G., Nadeau, S., Zutshi, N., An, W., Scheffe, S. et al. Protein tyrosine kinase regulation by ubiquitination: critical roles of Cbl-family ubiquitin ligases. Biochim. Biophys. Acta 1833, 122–139 (2013).

    CAS  PubMed  Google Scholar 

  17. 17.

    Taylor, S. J., Thien, C. B., Dagger, S. A., Duyvestyn, J. M., Grove, C. S., Lee, B. H. et al. Loss of c-Cbl E3 ubiquitin ligase activity enhances the development of myeloid leukemia in FLT3-ITD mutant mice. Exp. Hematol. 43, 191–206 e191 (2015).

    CAS  PubMed  Google Scholar 

  18. 18.

    Weisberg, E. L., Schauer, N. J., Yang, J., Lamberto, I., Doherty, L., Bhatt, S. et al. Inhibition of USP10 induces degradation of oncogenic FLT3. Nat. Chem. Biol. 13, 1207–1215 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  19. 19.

    Cools, J., Mentens, N., Furet, P., Fabbro, D., Clark, J. J., Griffin, J. D. et al. Prediction of resistance to small molecule FLT3 inhibitors. Cancer Res. 64, 6385 (2004).

    CAS  PubMed  Google Scholar 

  20. 20.

    Matsuo, Y., MacLeod, R. A., Uphoff, C. C., Drexler, H. G., Nishizaki, C., Katayama, Y. et al. Two acute monocytic leukemia (AML-M5a) cell lines (MOLM-13 and MOLM-14) with interclonal phenotypic heterogeneity showing MLL-AF9 fusion resulting from an occult chromosome insertion, ins(11;9)(q23;p22p23). Leukemia 11, 1469–1477 (1997).

    CAS  PubMed  Google Scholar 

  21. 21.

    Weisberg, E., Boulton, C., Kelly, L. M., Manley, P., Fabbro, D., Meyer, T. et al. Inhibition of mutant FLT3 receptors in leukemia cells by the small molecule tyrosine kinase inhibitor PKC412. Cancer Cell 1, 433–443 (2002).

    CAS  PubMed  Google Scholar 

  22. 22.

    Chou, T. C. & Talalay, P. Quantitative analysis of dose-effect relationships: the combined effects of multiple drugs or enzyme inhibitors. Adv. Enzym. Regul. 22, 27–55 (1984).

    CAS  Google Scholar 

  23. 23.

    Reverdy, C., Conrath, S., Lopez, R., Planquette, C., Atmanene, C., Collura, V. et al. Discovery of specific inhibitors of human USP7/HAUSP deubiquitinating enzyme. Chem. Biol. 19, 467–477 (2012).

    CAS  PubMed  Google Scholar 

  24. 24.

    Chauhan, D., Tian, Z., Nicholson, B., Kumar, K. G. S., Zhou, B., Carrasco, R. et al. A small molecule inhibitor of ubiquitin-specific protease-7 induces apoptosis in multiple myeloma cells and overcomes bortezomib resistance. Cancer Cell 22, 345–358 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  25. 25.

    Paolini, R., Molfetta, R., Piccoli, M., Frati, L. & Santoni, A. Ubiquitination and degradation of Syk and ZAP-70 protein tyrosine kinases in human NK cells upon CD16 engagement. Proc. Natl Acad. Sci. USA 98, 9611–9616 (2001).

    CAS  PubMed  Google Scholar 

  26. 26.

    Young, R. M., Hardy, I. R., Clarke, R. L., Lundy, N., Pine, P., Turner, B. C. et al. Mouse models of non-Hodgkin lymphoma reveal Syk as an important therapeutic target. Blood 113, 2508–2516 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  27. 27.

    Baudot, A. D., Jeandel, P. Y., Mouska, X., Maurer, U., Tartare-Deckert, S., Raynaud, S. D. et al. The tyrosine kinase Syk regulates the survival of chronic lymphocytic leukemia B cells through PKCδ and proteasome-dependent regulation of Mcl-1 expression. Oncogene 28, 3261 (2009).

  28. 28.

    Perova, T., Grandal, I., Nutter, L. M. J., Papp, E., Matei, I. R., Beyene, J. et al. Therapeutic potential of spleen tyrosine kinase inhibition for treating high-risk precursor B cell acute lymphoblastic leukemia. Sci. Transl. Med. 6, 236ra262 (2014).

    Google Scholar 

  29. 29.

    Uckun, F. M., Qazi, S., Cely, I., Sahin, K., Shahidzadeh, A., Ozercan, I. et al. Nanoscale liposomal formulation of a SYK P-site inhibitor against B-precursor leukemia. Blood 121, 4348–4354 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. 30.

    Hahn, C. K., Berchuck, J. E., Ross, K. N., Kakoza, R. M., Clauser, K., Schinzel, A. C. et al. Proteomic and genetic approaches identify Syk as an AML target. Cancer Cell 16, 281–294 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  31. 31.

    Gao, J., Aksoy, B. A., Dogrusoz, U., Dresdner, G., Gross, B., Sumer, S. O. et al. Integrative analysis of complex cancer genomics and clinical profiles using the cBioPortal. Sci. Signal. 6, pl1–pl1 (2013).

    PubMed  PubMed Central  Google Scholar 

  32. 32.

    Kuno, Y., Abe, A., Emi, N., Iida, M., Yokozawa, T., Towatari, M. et al. Constitutive kinase activation of the TEL-Syk fusion gene in myelodysplastic syndrome with t(9;12)(q22;p12). Blood 97, 1050 (2001).

    CAS  PubMed  Google Scholar 

  33. 33.

    Dierks, C., Adrian, F., Fisch, P., Ma, H., Maurer, H., Herchenbach, D. et al. The ITK-SYK fusion oncogene induces a T-cell lymphoproliferative disease in mice mimicking human disease. Cancer Res. 70, 6193 (2010).

    CAS  PubMed  Google Scholar 

  34. 34.

    Sargin, B., Choudhary, C., Crosetto, N., Schmidt, M. H. H., Grundler, R., Rensinghoff, M. et al. Flt3-dependent transformation by inactivating c-Cbl mutations in AML. Blood 110, 1004 (2007).

    CAS  PubMed  Google Scholar 

  35. 35.

    Fernandes, M. S., Reddy, M. M., Croteau, N. J., Walz, C., Weisbach, H., Podar, K. et al. Novel oncogenic mutations of CBL in human acute myeloid leukemia that activate growth and survival pathways depend on increased metabolism. J. Biol. Chem. 285, 32596–32605 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  36. 36.

    Abram, C. L. & Lowell, C. A. The expanding role for ITAM-based signaling pathways in immune cells. Sci. STKE 2007, re2 (2007).

    PubMed  Google Scholar 

  37. 37.

    Lu, J., Lin, W. H., Chen, S. Y., Longnecker, R., Tsai, S. C., Chen, C. L. et al. Syk tyrosine kinase mediates Epstein-Barr virus latent membrane protein 2A-induced cell migration in epithelial cells. J. Biol. Chem. 281, 8806–8814 (2006).

    CAS  PubMed  Google Scholar 

  38. 38.

    Katz, E., Dubois-Marshall, S., Sims, A. H., Faratian, D., Li, J., Smith, E. S. et al. A gene on the HER2 amplicon, C35, is an oncogene in breast cancer whose actions are prevented by inhibition of Syk. Br. J. Cancer 103, 401–410 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Singh, A., Greninger, P., Rhodes, D., Koopman, L., Violette, S., Bardeesy, N. et al. A gene expression signature associated with “K-Ras addiction” reveals regulators of EMT and tumor cell survival. Cancer Cell 15, 489–500 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Zhang, J., Benavente, C. A., McEvoy, J., Flores-Otero, J., Ding, L., Chen, X. et al. A novel retinoblastoma therapy from genomic and epigenetic analyses. Nature 481, 329–334 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  41. 41.

    Udyavar, A. R., Hoeksema, M. D., Clark, J. E., Zou, Y., Tang, Z., Li, Z. et al. Co-expression network analysis identifies Spleen Tyrosine Kinase (SYK) as a candidate oncogenic driver in a subset of small-cell lung cancer. BMC Syst. Biol. 7(Suppl 5), S1–S1 (2013).

    PubMed  PubMed Central  Google Scholar 

  42. 42.

    Luangdilok, S., Box, C., Patterson, L., Court, W., Harrington, K., Pitkin, L. et al. Syk tyrosine kinase is linked to cell motility and progression in squamous cell carcinomas of the head and neck. Cancer Res. 67, 7907 (2007).

    CAS  PubMed  Google Scholar 

  43. 43.

    Shin, G., Kang, T.-W., Yang, S., Baek, S.-J., Jeong, Y.-S. & Kim, S.-Y. GENT: gene expression database of normal and tumor tissues. Cancer Inform. 10, 149–157 (2011).

Download references

Acknowledgements

We thank Dr Nathanael Gray and Dr Richard Stone at Dana-Farber Cancer Institute—Harvard Medical School for valuable guidance on literature searches and critical revision of the manuscript.

Author information

Affiliations

Authors

Contributions

J.Y., C.M. and E.W. conceptualised, designed and performed the studies. J.Y. and E.W. carried out data analyses and wrote the manuscript. A.C. assisted with the writing of the manuscript and preparation of figures. I.L. assisted with shRNA KD studies. R.M. helped design and provided important suggestions for His pulldown assay. S.A. provided important immunoblotting reagents. J.W. and N.G. provided FLT3 inhibitors. S.L. provided primary cells. R.S. provided valuable scientific feedback and guidance. M.S. provided valuable scientific feedback and guidance. S.B. conceptualised and designed the studies, carried out data analyses and wrote the manuscript. J.G. conceptualised and designed the studies.

Corresponding authors

Correspondence to Sara Buhrlage or James D. Griffin.

Ethics declarations

Ethics approval and consent to participate

Prior to acquisition of normal PBMC samples, the subjects provided their informed consent to participate. Our studies were performed in accordance with the Declaration of Helsinki. IRB protocol number is 13-351 (consents 01-206 and 11-104 (which is now known as 17-000).

Consent to publish

N/A

Data availability

The authors declare that the main data supporting the findings of this study are available within the article and its supplementary information files.

Competing interests

The authors declare no competing interests.

Funding information

This study is supported by the National Institutes of Health research project grant (R01) CA211681.

Additional information

Note This work is published under the standard license to publish agreement. After 12 months the work will become freely available and the license terms will switch to a Creative Commons Attribution 4.0 International (CC BY 4.0).

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Yang, J., Meng, C., Weisberg, E. et al. Inhibition of the deubiquitinase USP10 induces degradation of SYK. Br J Cancer 122, 1175–1184 (2020). https://doi.org/10.1038/s41416-020-0731-z

Download citation

Further reading

Search