Impact of Protein Kinase C Activation and Monoclonal Antibodies on Immune Checkpoint Regulation and B Cell Function in Patients with Chronic Lymphocytic Leukemia

<b>Background</b>: Chronic lymphocytic leukemia (CLL) is characterized by the proliferation of dysfunctional B cells, resulting in significant immune dysregulation. Patients with CLL exhibit varied responses to B cell receptor (BCR) targeted therapies, emphasizing the need for tailored immunotherapy approaches. This study investigated B cell function in untreated patients with CLL, and we further explored the effects of ex vivo protein kinase C activation on immune checkpoint expression and B cell profiles. <b>Methods:</b> Peripheral blood samples were collected from 21 untreated patients with CLL at King Edward Hospital in South Africa, between 2019 and 2022. B cells were stimulated with phorbol myristate acetate (PMA) and ionomycin. Using flow cytometry, the study explored the levels of B cell subsets and immune checkpoint proteins programmed cell death 1 (PD-1), programmed cell death-ligand 1 (PD-L1), programmed cell death-ligand 2 (PD-L2) and cytotoxic T-lymphocyte associated protein 4 (CTLA-4) expression on various B cell subsets. <b>Results:</b> PMA and ionomycin B cell stimulation upregulated PD-1, CTLA-4 and PD-L2 expression on B cell subsets (<i>p</i> < 0.01). As expected, monoclonal antibodies targeting PD-1, PD-L1 and CTLA-4 significantly downregulated the CTLA-4 expression of B cell subsets (<i>p</i> < 0.05), while PD-L2 exhibited varied responses in different B cell subsets. Moreover, PD-1 and PD-L1 expression on total B cells significantly declined following their blockage (<i>p</i> < 0.01). In addition, these monoclonal antibodies increased the levels of CD19⁺CD27⁺ B cells (<i>p</i> < 0.0128) and activated CD19⁺CD27⁺ B cells (<i>p</i> < 0.01). <b>Conclusions:</b> Protein kinase C activation on B cells stimulates immune checkpoint expression. The use of monoclonal antibodies on B cells plays a critical role in the B cell function through the reduction in CD38 expressing activated B cells and upregulation of CD19⁺CD27⁺ B cells. Moreover, the monoclonal antibody targeting PD-1, PD-L1 and CTLA-4 are effective in reducing the expression of CTLA-4 on B cell subsets, while PD-1 and PD-L1 blockage may be effective in reducing the expression of these immune checkpoints on total B cells.