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An allosteric MALT1 inhibitor is a molecular corrector rescuing function in an immunodeficient patient

Abstract

MALT1 paracaspase is central for lymphocyte antigen-dependent responses including NF-κB activation. We discovered nanomolar, selective allosteric inhibitors of MALT1 that bind by displacing the side chain of Trp580, locking the protease in an inactive conformation. Interestingly, we had previously identified a patient homozygous for a MALT1 Trp580-to-serine mutation who suffered from combined immunodeficiency. We show that the loss of tryptophan weakened interactions between the paracaspase and C-terminal immunoglobulin MALT1 domains resulting in protein instability, reduced protein levels and functions. Upon binding of allosteric inhibitors of increasing potency, we found proportionate increased stabilization of MALT1-W580S to reach that of wild-type MALT1. With restored levels of stable MALT1 protein, the most potent of the allosteric inhibitors rescued NF-κB and JNK signaling in patient lymphocytes. Following compound washout, MALT1 substrate cleavage was partly recovered. Thus, a molecular corrector rescues an enzyme deficiency by substituting for the mutated residue, inspiring new potential precision therapies to increase mutant enzyme activity in other deficiencies.

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Fig. 1: MLT-748 and MLT-747 inhibit MALT1 peptide cleavage.
Fig. 2: MLT-748 and MLT-747 mimics Trp580 in MALT1.
Fig. 3: MLT-748 binds to the same allosteric pocket as mepazine and displaces Trp580.
Fig. 4: Comparison of the allosteric inhibitors MLT-748, MLT-747 and mepazine in binding and stabilizing MALT1-W580S in vitro and in MALT1mut/mut B cells.
Fig. 5: MLT-748 blocks cleavage of MALT1 substrates in human lymphocytes.
Fig. 6: MLT-748 restores MALT1-W580S function in patient B and T cells.

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Data availability

Data are available from the authors upon reasonable request. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the data set identifier PXD008421.

References

  1. Turk, B. Targeting proteases: successes, failures and future prospects. Nat. Rev. Drug. Discov. 5, 785–799 (2006).

    Article  CAS  Google Scholar 

  2. Overall, C. M. & Kleifeld, O. Tumour microenvironment - opinion: validating matrix metalloproteinases as drug targets and anti-targets for cancer therapy. Nat. Rev. Cancer 6, 227–239 (2006).

    Article  CAS  Google Scholar 

  3. Klein, T., Eckhard, U., Dufour, A., Solis, N. & Overall, C. M. Proteolytic cleavage-mechanisms, function, and “omic” approaches for a near-ubiquitous posttranslational modification. Chem. Rev. 118, 1137–1168 (2018).

    Article  CAS  Google Scholar 

  4. Gomes, C. M. Protein misfolding in disease and small molecule therapies. Curr. Top. Med. Chem. 12, 2460–2469 (2012).

    Article  CAS  Google Scholar 

  5. Gámez, A. et al. Protein misfolding diseases: prospects of pharmacological treatment. Clin. Genet. 93, 450–458 (2018).

    Article  Google Scholar 

  6. Rudashevskaya, E. L., Stockner, T., Trauner, M., Freissmuth, M. & Chiba, P. Pharmacological correction of misfolding of ABC proteins. Drug Discov. Today. Technol. 12, e87–e94 (2014).

    Article  Google Scholar 

  7. Mohell, N. et al. APR-246 overcomes resistance to cisplatin and doxorubicin in ovarian cancer cells. Cell Death Dis. 6, e1794 (2015).

    Article  CAS  Google Scholar 

  8. Lieberman, R. L. et al. Structure of acid β-glucosidase with pharmacological chaperone provides insight into Gaucher disease. Nat. Chem. Biol. 3, 101–107 (2007).

    Article  CAS  Google Scholar 

  9. Jorge-Finnigan, A. et al. Pharmacological chaperones as a potential therapeutic option in methylmalonic aciduria cblB type. Hum. Mol. Genet. 22, 3680–3689 (2013).

    Article  CAS  Google Scholar 

  10. Denny, R. A., Gavrin, L. K. & Saiah, E. Recent developments in targeting protein misfolding diseases. Bioorg. Med. Chem. Lett. 23, 1935–1944 (2013).

    Article  CAS  Google Scholar 

  11. Hayden, M. S. & Ghosh, S. NF-κB in immunobiology. Cell Res. 21, 223–244 (2011).

    Article  CAS  Google Scholar 

  12. Rosebeck, S., Rehman, A. O., Lucas, P. C. & Mcallister-lucas, L. M. From MALT lymphoma to the CBM signalosome: three decades of discovery. Cell Cycle 10, 2485–2496 (2011).

    Article  CAS  Google Scholar 

  13. Sun, L., Deng, L., Ea, C. K., Xia, Z. P. & Chen, Z. J. The TRAF6 ubiquitin ligase and TAK1 kinase mediate IKK activation by BCL10 and MALT1 in T lymphocytes. Mol. Cell 14, 289–301 (2004).

    Article  CAS  Google Scholar 

  14. Rebeaud, F. et al. The proteolytic activity of the paracaspase MALT1 is key in T cell activation. Nat. Immunol. 9, 272–281 (2008).

    Article  CAS  Google Scholar 

  15. Düwel, M. et al. A20 negatively regulates T cell receptor signaling to NF-kappaB by cleaving Malt1 ubiquitin chains. J. Immunol. 182, 7718–7728 (2009).

    Article  Google Scholar 

  16. Hailfinger, S. et al. Malt1-dependent RelB cleavage promotes canonical NF-kappaB activation in lymphocytes and lymphoma cell lines. Proc. Natl Acad. Sci. USA 108, 14596–14601 (2011).

    Article  CAS  Google Scholar 

  17. Klein, T. et al. The paracaspase MALT1 cleaves HOIL1 reducing linear ubiquitination by LUBAC to dampen lymphocyte NF-κB signalling. Nat. Commun. 6, 8777 (2015).

    Article  CAS  Google Scholar 

  18. Bornancin, F. et al. Deficiency of MALT1 paracaspase activity results in unbalanced regulatory and effector T and B cell responses leading to multiorgan inflammation. J. Immunol. 194, 3723–3734 (2015).

    Article  CAS  Google Scholar 

  19. Gewies, A. et al. Uncoupling Malt1 threshold function from paracaspase activity results in destructive autoimmune inflammation. Cell Rep. 9, 1292–1305 (2014).

    Article  CAS  Google Scholar 

  20. Jaworski, M. et al. Malt1 protease inactivation efficiently dampens immune responses but causes spontaneous autoimmunity. EMBO J. 33, 2765–2781 (2014).

    Article  CAS  Google Scholar 

  21. Yu, J. W. et al. MALT1 protease activity is required for innate and adaptive immune responses. PLoS One 10, e0127083 (2015).

    Article  Google Scholar 

  22. Qiao, Q. et al. Structural architecture of the CARMA1/Bcl10/MALT1 signalosome: nucleation-induced filamentous assembly. Mol. Cell 51, 766–779 (2013).

    Article  CAS  Google Scholar 

  23. Staal, J. et al. T-cell receptor-induced JNK activation requires proteolytic inactivation of CYLD by MALT1. EMBO J. 30, 1742–1752 (2011).

    Article  CAS  Google Scholar 

  24. Coornaert, B. et al. T cell antigen receptor stimulation induces MALT1 paracaspase-mediated cleavage of the NF-kappaB inhibitor A20. Nat. Immunol. 9, 263–271 (2008).

    Article  CAS  Google Scholar 

  25. Elton, L. et al. MALT1 cleaves the E3 ubiquitin ligase HOIL-1 in activated T cells, generating a dominant negative inhibitor of LUBAC-induced NF-κB signaling. FEBS. J. 283, 403–412 (2016).

    Article  CAS  Google Scholar 

  26. Douanne, T., Gavard, J. & Bidère, N. The paracaspase MALT1 cleaves the LUBAC subunit HOIL1 during antigen receptor signaling. J. Cell. Sci. 129, 1775–1780 (2016).

    Article  CAS  Google Scholar 

  27. Ginster, S. et al. Two Antagonistic MALT1 auto-cleavage mechanisms reveal a role for TRAF6 to unleash MALT1 activation. PLoS One 12, e0169026 (2017).

    Article  Google Scholar 

  28. Hailfinger, S. et al. Essential role of MALT1 protease activity in activated B cell-like diffuse large B-cell lymphoma. Proc. Natl Acad. Sci. USA 106, 19946–19951 (2009).

    Article  CAS  Google Scholar 

  29. Ferch, U. et al. Inhibition of MALT1 protease activity is selectively toxic for activated B cell-like diffuse large B cell lymphoma cells. J. Exp. Med. 206, 2313–2320 (2009).

    Article  CAS  Google Scholar 

  30. Nagel, D. et al. Pharmacologic inhibition of MALT1 protease by phenothiazines as a therapeutic approach for the treatment of aggressive ABC-DLBCL. Cancer Cell. 22, 825–837 (2012).

    Article  CAS  Google Scholar 

  31. Fontan, L. et al. MALT1 small molecule inhibitors specifically suppress ABC-DLBCL in vitro and in vivo. Cancer Cell. 22, 812–824 (2012).

    Article  CAS  Google Scholar 

  32. Turvey, S. E. et al. TheCARD11-BCL10-MALT1 (CBM) signalosome complex: stepping into the limelight of human primary immunodeficiency. J. Allergy Clin. Immunol. 134, 276–284 (2014).

    Article  CAS  Google Scholar 

  33. Pérez de Diego, R. et al. Genetic errors of the human caspase recruitment domain-B-cell lymphoma 10-mucosa-associated lymphoid tissue lymphoma-translocation gene 1 (CBM) complex: molecular, immunologic, and clinical heterogeneity. J. Allergy Clin. Immunol. 136, 1139–1149 (2015).

    Article  Google Scholar 

  34. McKinnon, M. L. et al. Combined immunodeficiency associated with homozygous MALT1 mutations. J. Allergy Clin. Immunol. 133, 1458–1462 (2014). 1462.e1–1462.e7.

    Article  CAS  Google Scholar 

  35. Punwani, D. et al. Combined immunodeficiency due to MALT1 mutations, treated by hematopoietic cell transplantation. J. Clin. Immunol. 35, 135–146 (2015).

    Article  CAS  Google Scholar 

  36. Charbit-Henrion, F. et al. Deficiency in mucosa-associated lymphoid tissue lymphoma translocation 1: a novel cause of IPEX-like syndrome. J. Pediatr. Gastroenterol. Nutr. 64, 378–384 (2017).

    Article  Google Scholar 

  37. Rozmus, J. et al. Successful clinical treatment and functional immunological normalization of human MALT1 deficiency following hematopoietic stem cell transplantation. Clin. Immunol. 168, 1–5 (2016).

    Article  CAS  Google Scholar 

  38. Wiesmann, C. et al. Structural determinants of MALT1 protease activity. J. Mol. Biol. 419, 4–21 (2012).

    Article  CAS  Google Scholar 

  39. Schlauderer, F. et al. Structural analysis of phenothiazine derivatives as allosteric inhibitors of the MALT1 paracaspase. Angew. Chem. Int. Ed. Engl. 52, 10384–10387 (2013).

    Article  CAS  Google Scholar 

  40. Kleifeld, O. et al. Isotopic labeling of terminal amines in complex samples identifies protein N-termini and protease cleavage products. Nat. Biotechnol. 28, 281–288 (2010).

    Article  CAS  Google Scholar 

  41. Bardet, M. et al. The T-cell fingerprint of MALT1 paracaspase revealed by selective inhibition. Immunol. Cell Biol. 96, 81–99 (2018).

    Article  CAS  Google Scholar 

  42. Yu, J. W., Jeffrey, P. D., Ha, J. Y., Yang, X. & Shi, Y. Crystal structure of the mucosa-associated lymphoid tissue lymphoma translocation 1 (MALT1) paracaspase region. Proc. Natl Acad. Sci. USA 108, 21004–21009 (2011).

    Article  CAS  Google Scholar 

  43. Demeyer, A., Staal, J. & Beyaert, R. Targeting MALT1 proteolytic activity in immunity, inflammation and disease: good or bad? Trends Mol. Med. 22, 135–150 (2016).

    Article  CAS  Google Scholar 

  44. Vonrhein, C. et al. Data processing and analysis with the autoPROC toolbox. Acta Crystallogr. D Biol. Crystallogr. 67, 293–302 (2011).

    Article  CAS  Google Scholar 

  45. Storoni, L. C., McCoy, A. J. & Read, R. J. Likelihood-enhanced fast rotation functions. Acta Crystallogr. D Biol. Crystallogr. 60, 432–438 (2004).

    Article  Google Scholar 

  46. Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D Biol. Crystallogr. 60, 2126–2132 (2004).

    Article  Google Scholar 

  47. Chung, B. K. et al. Innate immune control of EBV-infected B cells by invariant natural killer T cells. Blood 122, 2600–2608 (2013).

    Article  CAS  Google Scholar 

  48. Miller, G. & Lipman, M. Release of infectious Epstein-Barr virus by transformed marmoset leukocytes. Proc. Natl Acad. Sci. USA 70, 190–194 (1973).

    Article  CAS  Google Scholar 

  49. Davis, R. E. et al. Chronic active B-cell-receptor signalling in diffuse large B-cell lymphoma. Nature 463, 88–92 (2010).

    Article  CAS  Google Scholar 

  50. Schlapbach, A. et al. N-aryl-piperidine-4-carboxamides as a novel class of potent inhibitors of MALT1 proteolytic activity. Bioorg. Med. Chem. Lett. 28, 2153–2158 (2018).

    Article  CAS  Google Scholar 

  51. Klein, T., Viner, R. I. & Overall, C. M. Quantitative proteomics and terminomics to elucidate the role of ubiquitination and proteolysis in adaptive immunity. Philos. Trans. A Math. Phys. Eng. Sci. 374, 2079 (2016).

    Article  Google Scholar 

  52. Evenou, J.-P. et al. The potent protein kinase C-selective inhibitor AEB071 (sotrastaurin) represents a new class of immunosuppressive agents affecting early T-cell activation. J. Pharmacol. Exp. Ther. 330, 792–801 (2009).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank C.H. Régnier for helpful discussions. We gratefully acknowledge the contributions of C. Malinverni and J. Heng in the early part of this work. We acknowledge the Paul Scherrer Institut, Villigen, Switzerland for provision of synchrotron radiation beamtime at beamline PXII (X10/SA) of the SLS. We also thank the beamline staff, as well as J. Diez and his staff at Expose GmbH for their excellent help with data collection. We thank F. Sirokin for providing a model of the MALT1 complex with mepazine. We thank M. Shipp (Harvard University MA, USA) for providing the OCI-Ly3 B cell line. C.M.O. holds a Canada Research Chair in Protease Proteomics and Systems Biology (number: 950-20-3877). This work was supported by Canadian Institutes of Health Research grants (MOP-133691 to S.E.T. and FDN: 148408, MOP-37937 to C.M.O.), Natural Sciences and Engineering Research Council of Canada (RGPIN 435829-201 to S.E.T), the Michael Smith Foundation for Health Research to establish the British Columbia Proteomics Network (IN-NPG-00105-156 to C.M.O.) and the Canada Foundation for Innovation (31059 to C.M.O.).

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Contributions

J.Q., A.S. and R.H. oversaw the discovery and synthesis of the compounds. T.K. and S.-Y.F. performed MALT1 stabilization, cleavage and signaling and functional experiments in B and T cells. T.K., S.Y.F., J.J.P. and S.K., performed experiments on expanded primary T cells. T.K., M.A.B. and R.I.V. performed kinetic experiments by TAILS, LC–MS/MS analysis and data analysis. J.K. synthesized the aldehyde polymer used for N-terminomic TAILS analyses, and T.K. analyzed proteomic data and made figures. M.R. conceived the crystallographic study, collected and analyzed crystallographic data, solved crystal structures and made figures. F.V. performed crystallization experiments. N.H. and P.E. designed and executed the SPR and DSF experiments. L.I. designed and executed the Trp580 quenching and enzymatic activity experiments. J.B. designed and oversaw the screening campaign and follow-up compound triage. J.Q., J.E., S.E.T., and C.M.O. conceived the research idea. J.Q., T.K., S.Y.F., S.E.T., F.B. and C.M.O. wrote the paper with contributions from all authors.

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Correspondence to Jean Quancard, Frédéric Bornancin or Christopher M. Overall.

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J.Q., M.R., N.H., L.I., J.B., A.S., P.E., F.V., R.H., J.E., and F.B. are employees of the Novartis Institute of Biomedical Research. R.I.V. is and M.B. was an employee of Thermo Fisher Scientific, developer and distributor of the Orbitrap Fusion Lumos mass spectrometer.

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Quancard, J., Klein, T., Fung, SY. et al. An allosteric MALT1 inhibitor is a molecular corrector rescuing function in an immunodeficient patient. Nat Chem Biol 15, 304–313 (2019). https://doi.org/10.1038/s41589-018-0222-1

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