Ibrutinib Therapy Increases T Cell Repertoire Diversity in Patients with Chronic Lymphocytic Leukemia

Abstract

The Bruton's tyrosine kinase inhibitor ibrutinib is a highly effective, new targeted therapy for chronic lymphocytic leukemia (CLL) that thwarts leukemia cell survival, growth, and tissue homing. The effects of ibrutinib treatment on the T cell compartment, which is clonally expanded and thought to support the growth of malignant B cells in CLL, are not fully characterized. Using next-generation sequencing technology, we characterized the diversity of TCRβ-chains in peripheral blood T cells from 15 CLL patients before and after 1 y of ibrutinib therapy. We noted elevated CD4⁺ and CD8⁺ T cell numbers and a restricted TCRβ repertoire in all pretreatment samples. After 1 y of ibrutinib therapy, elevated peripheral blood T cell numbers and T cell-related cytokine levels had normalized, and T cell repertoire diversity increased significantly. Dominant TCRβ clones in pretreatment samples declined or became undetectable, and the number of productive unique clones increased significantly during ibrutinib therapy, with the emergence of large numbers of low-frequency TCRβ clones. Importantly, broader TCR repertoire diversity was associated with clinical efficacy and lower rates of infections during ibrutinib therapy. These data demonstrate that ibrutinib therapy increases diversification of the T cell compartment in CLL patients, which contributes to cellular immune reconstitution.

Figure 1. T cell counts and the levels of related plasma cytokines significantly decreased during ibrutinib therapy

(a) T cell subsets decreased over time during ibrutinib therapy. (b) Levels of plasma Th1-, Th2-, and Th17-type cytokines and chemotactic factors significantly declined when compared to pretreatment levels. The bars represent mean values, and dotted lines indicate the normal ranges. “Pre”, “3 m”, “6 m”, and “12 m” refer to pretreatment, 3 month, 6 month, and 12 month on ibrutinib therapy, respectively. Ctrl. Indicates age-matched control group. (Asterisks represent statistical significance; *P < 0.05, ** P < 0.01, ***P < 0.005, **** P < 0.0001. ns, not significant.)

Figure 2. TCRβ repertoire diversity increased after one year of ibrutinib therapy

(a) There was no significant difference in numbers of productive total TCRβ sequences between samples taken before and after 1 year of ibrutinib therapy. In contrast, (b) the number of absolute productive unique sequences, representing the richness of the clones, and (c) relative productive unique sequences significantly increased after 1 year of ibrutinib therapy. “Pre” and “12 m” refer to pretreatment and 12 months on ibrutinib therapy, respectively. (*P < 0.05, ** P < 0.01, **** P < 0.0001; ns, not significant.)

Figure 3. High-frequency TCRβ clones diminished and low-frequency T cell clones increased during ibrutinib therapy

(a) Graphic representation of the model used for assignment of clones according to their frequency. (b and c) TCRβ repertoire diversity significantly increased after 1 year of ibrutinib therapy compared to pretreatment samples, as the fraction of high-frequency clones significantly declined and the fractions of “singleton” clones significantly increased. (*P < 0.05, ** P < 0.01, **** P < 0.0001; ns, not significant.)

Figure 4. Broader TCR repertoire diversification was associated with clinical response and lower infection rates during ibrutinib therapy

(a) Patients who achieved CR had greater TCR repertoire diversification than patients who achieved PR after one year of ibrutinib therapy. (b) Broader TCR repertoire diversity was associated with lower rates of infections during the first 6 months of ibrutinib therapy, and TCR repertoire diversity significantly increased after 1 year of ibrutinib therapy in patients with lower rates of infection. (*P < 0.05, ns, not significant.)

Figure 5. Longitudinal evaluation of shift of T cell clones from pretreatment to 1 year after ibrutinib therapy

The dominant clones present before treatment decreased in frequency or vanished, while a large numbers of new clones emerged 1 year after ibrutinib therapy.