Multi-omics exploration of microenvironmental changes during BTK covalent inhibition in chronic lymphocytic leukemia

Continuous treatment with ibrutinib not only exerts tumor control but also enhances T cell function in patients with chronic lymphocytic leukemia (CLL). We conducted longitudinal multiomics analyses in samples from CLL patients receiving ibrutinib upfront to identify potential adaptive mechanisms to Bruton Tyrosine Kinase (BTK) inhibition during the first 12 months of continuous therapy. Wefound that in T cells, ibrutinib reduced the expression of exhaustion markers, the proportion of Tregs and Tfh cells, as well as expression of genes related to activation, proliferation, differentiation, and metabolism. In CLL cells, we observed a downregulation of immunosuppression, adhesion, and migration mechanisms. Adaptation at molecular level, characterized by an increase in cancer cell fraction of CLL cells with mutated driver genes, was observed in around half of the patients. Interestingly, BTK C481S mutations were detected as early as after 6 months of treatment, particularly enriched in subsets of malignant cells retaining migrative capacity. These CLL cells with potential migrative capacity under ibrutinib also exhibited a distinct transcriptomic profile including upregulation of mTOR-AKT and Myc pathways. We identified high expression of TMBIM6 as a potential novel independent poor prognostic factor. Of note, BIA, a TMBIM6 antagonist, induced CLL cell apoptosis and synergized with ibrutinib. In summary, our comprehensive multi-omics analysis of CLL patients undergoing ibrutinib therapy has unveiled early immunomodulatory effects on T cells and adaptative mechanisms in CLL cells. These findings can contribute to the identification of resistance mechanisms and the discovery of novel therapeutic targets. Overall design: To assess the transcriptomic changes in T cells from chronic lymphocytic leukemia (CLL) after 6 months of ibrutinib treatment, we isolated total RNA from T cells (CD3+ purification, negative selection) from peripheral blood mononuclear cells samples from CLL patients at before treatment and after 6 months of ibrutinib. To assess the transcriptomic changes in migrated B cells from chronic lymphocytic leukemia (CLL) towards CXCL12 before and after 6 months of ibrutinib treatment. We previously isolated B cells (CD19+ purification, negative selection) from peripheral blood mononuclear cells samples from CLL patients at before treatment and after 6 months of ibrutinib.