Lgr5-mediated restraint of β-catenin is essential for B-lymphopoiesis and leukemia-initiation

Abstract

Upon productive immunoglobulin gene rearrangement, expression of a functional pre-B cell receptor (pre-BCR) initiates positive selection of pre-B cells, clonal expansion and self-renewal1-2. Studying mechanisms driving this first wave of B-lymphopoiesis, we identified the G-protein coupled receptor Lgr5 as an essential initiator of positive selection. Lgr5 was extensively studied as determinant of stem cell populations in multiple tissues3-6, but not in B-cells. While undetectable throughout the hematopoietic system, positively selected pre-B cells were marked with a sharp peak of Lgr5 expression. Conditional deletion of Lgr5 preceding the pre-BCR checkpoint induced negative selection and complete abortion of B-cell development. Proteomic studies of Lgr5-ablation revealed massive (>250-fold) accumulation of {beta}-catenin and suppression of MYC. Lgr5-deficient pre-B cells fully recovered by concurrent {beta}-catenin-deletion, demonstrating a central role of Lgr5-mediated restraint of {beta}-catenin at the pre-BCR checkpoint. In other cell types, {beta}-catenin/TCF4 complexes drive transcriptional activation of MYC7-9. Instead of TCF4, proximity-based interactome studies in pre-B cells identified the B-lymphoid transcription factors IKZF1 and IKZF310-11 as {beta}-catenin-binding partners, which had the opposite effect and caused transcriptional repression of MYC. On positively selected pre-B cells, Lgr5 prevented accumulation of {beta}-catenin and formation of complexes with IKZF1 and IKZF3, which relieved transcriptional repression of MYC. Activating {beta}-catenin-mutations are common throughout all main types of cancer7-8, but were conspicuously absent in pre-B leukemia (B-ALL). Like pre-B cells, B-ALL cells were uniquely sensitive to genetic and pharmacological {beta}-catenin hyperactivation, which recapitulated the effects of Lgr5-deletion and compromised colony formation and leukemia-initiation. A new LGR5 antibody-drug conjugate targeted leukemia-initiating cells in patient-derived B-ALL and achieved long-term disease-control. Likewise, small molecule hyperactivation of {beta}-catenin selectively killed B-ALL but not other cell types. Hence, Lgr5-mediated restraint of {beta}-catenin activation is essential for B-lymphopoiesis and revealed an unexpected vulnerability that can be leveraged for the treatment of drug-resistant B-ALL.