Follicular lymphoma-associated BTK mutations are inactivating resulting in augmented AKT activation.

Purpose Based on the recent discovery of mutations in Bruton's tyrosine kinase (BTK) in Follicular Lymphoma (FL), we studied their functional properties. Experimental design We identified novel somatic BTK mutations in 7% of a combined total of 139 FL and 11 transformed FL cases, none of which had received prior treatment with B cell receptor (BCR) targeted drugs. We reconstituted WT and mutant BTK into various engineered lymphoma cell lines. We measured BCR-induced signal transduction events in engineered cell lines and primary human FL B cells. Results We uncovered that all BTK mutants destabilized the BTK protein and some created BTK kinase-dead mutants. The PLCγ2 is a substrate of BTK but the BTK mutants did not alter PLCγ2 phosphorylation. Instead, we discovered that BTK mutants induced an exaggerated AKT phosphorylation phenotype in anti-immunoglobulin (IG) treated recombinant lymphoma cell lines. The shRNA-mediated knock-down of BTK expression in primary human non-malignant lymph node-derived B cells resulted in strong anti-IG-induced AKT activation, as did the degradation of BTK protein in cells lines using ibrutinib-based proteolysis targeting chimera (PROTAC). Finally, through analyses of primary human FL B cells carrying WT or mutant BTK, we detected elevated AKT phosphorylation following surface IG crosslinking in all FL B cells, including all BTK mutant FL. The augmented AKT phosphorylation following BCR crosslinking could be abrogated by pre-treatment with a PI3Kδ inhibitor. Conclusions Altogether, our data uncover novel unexpected properties of FL-associated BTK mutations with direct implications for targeted therapy development in FL.