Single-Cell RNA-seq Analysis Reveals Distinct Tumor and Immunosuppressive T-Cell Phenotypes in Patients with CLL Treated with Ibrutinib

PURPOSE: The development of Bruton tyrosine kinase inhibitors (BTKi) and their introduction into clinical practice represents a major advance in the treatment of chronic lymphocytic leukemia (CLL). However, monotherapy with ibrutinib or other BTKis does not induce complete remissions or undetectable minimal residual disease even with extended therapy. Therefore, there is a need to understand the differences between ibrutinib-sensitive and -resistant CLL cells along with immune microenvironment to identify therapeutic approaches for controlling residual disease during BTKi treatment. EXPERIMENTAL DESIGN: Here, we investigated the cellular heterogeneity of peripheral blood mononuclear cells from patients with CLL treated with ibrutinib using single-cell RNA sequencing. RESULTS: We identified unique transcriptional heterogeneity within the B-cell cluster in the ibrutinib-sensitive and -resistant patients. Ibrutinib-sensitive cells showed enrichment of B-cell populations with upregulation of MHC I molecules and TNF family members. Additionally, we observed that inflammatory response and metabolism-related pathways were decreased, whereas cellular response to stress and DNA repair programs were increased in the ibrutinib-resistant samples. T cells in ibrutinib-resistant patients showed expansion of regulatory T cells and an exhausted CD8 effector T-cell compartment. Furthermore, CD14+ and CD16+ monocytes from ibrutinib-resistant patients preferentially expressed a gene expression program of antiviral immunity. CONCLUSIONS: At single-cell level, our findings demonstrate a picture of transcriptional heterogeneity in the tumor compartment and immune milieu. Overall, these findings highlight transcriptional changes in circulating immune cells associated with ibrutinib resistance, suggesting that T-cell exhaustion and monocyte polarization accompany and may contribute to resistance during long-term BTKi therapy.