Noncatalytic Bruton's tyrosine kinase activates PLCγ2 variants mediating ibrutinib resistance in human chronic lymphocytic leukemia cells

Martin Wist, Laura Meier, Orit Gutman, Jennifer Haas, Sascha Endres, Yuan Zhou, Reinhild Rösler, Sebastian Wiese, Stephan Stilgenbauer, Elias Hobeika, Yoav I. Henis, Peter Gierschik, Claudia Walliser*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

21 Scopus citations

Abstract

Treatment of patients with chronic lymphocytic leukemia (CLL) with inhibitors of Bruton's tyrosine kinase (BTK), such as ibrutinib, is limited by primary or secondary resistance to this drug. Examinations of CLL patients with late relapses while on ibrutinib, which inhibits BTK's catalytic activity, revealed several mutations in BTK, most frequently resulting in the C481S substitution, and disclosed many mutations inPLCG2, encoding phospholipase C-γ2 (PLCγ2). The PLCγ2 variants typically do not exhibit constitutive activity in cell-free systems, leading to the suggestion that in intact cells they are hypersensitive to Rac family small GTPases or to the upstream kinases spleen-associated tyrosine kinase (SYK) and Lck/Yes-related novel tyrosine kinase (LYN). The sensitivity of the PLCγ2 variants to BTK itself has remained unknown. Here, using genetically-modified DT40 Blymphocytes, along with various biochemical assays, including analysis of PLCγ2-mediated inositol phosphate formation, inositol phospholipid assessments, fluorescence recovery after photobleaching (FRAP) static laser microscopy, and determination of intracellular calcium ([Ca2+]i), we show that various CLL-specific PLCγ2 variants such as PLCγ2S707Y are hyperresponsive to activated BTK, even in the absence of BTK's catalytic activity and independently of enhanced PLCγ2 phospholipid substrate supply. At high levels of B-cell receptor (BCR) activation, which may occur in individual CLL patients, catalytically- inactive BTK restored the ability of the BCR to mediate increases in [Ca2+]i. Because catalytically-inactiveBTKis insensitive to active-siteBTKinhibitors, the mechanism involving the noncatalytic BTK uncovered here may contribute to preexisting reduced sensitivity or even primary resistance of CLL to these drugs.

Original languageEnglish
Pages (from-to)5717-5736
Number of pages20
JournalJournal of Biological Chemistry
Volume295
Issue number17
DOIs
StatePublished - 24 Apr 2020

Funding

FundersFunder number
Deutsche ForschungsgemeinschaftSFB 1074/TP A8
Hessisches Ministerium für Wissenschaft und Kunst
International Graduate School in Molecular Medicine UlmGSC 279

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