The outcome of B-cell receptor signaling in chronic lymphocytic leukemia: proliferation or anergy

Abstract

Biologists and clinicians agree that the B-cell receptor influences the behavior of chronic lymphocytic leukemia, and promising new drugs are aimed at receptor-associated kinases. Engagement of surface immunoglobulin by antigen is a key driver of malignant cells with outcome influenced by the nature of the cell, the level of stimulation and the microenvironment. Analysis of surface immunoglobulin-mediated signaling in the two major disease subsets defined by IGHV mutational status reveals bifurcation of responses toward proliferation or anergy. Mutated chronic lymphocytic leukemia, generally of relatively good prognosis, is mainly, but not exclusively, driven towards anergy in vivo. In contrast, unmutated chronic lymphocytic leukemia shows less evidence for anergy in vivo retaining more responsiveness to surface immunoglobulin M-mediated signaling, possibly explaining increased tumor progression. Expression and function of surface immunoglobulin M in unmutated chronic lymphocytic leukemia appear rather homogeneous, but mutated chronic lymphocytic leukemia exhibits a highly heterogeneous profile that may relate to further variable clinical behavior within this subset. Anergy should increase susceptibility to apoptosis but, in leukemic cells, this may be countered by overexpression of the B-cell lymphoma-2 survival protein. Maintained anergy spreads to chemokines and adhesion molecules, restraining homing and migration. However, anergy is not necessarily completely benign, being able to reverse and regenerate surface immunoglobulin M-mediated responses. A two-pronged attack on proliferative and anti-apoptotic pathways may succeed. Increased understanding of how chronic lymphocytic leukemia cells are driven to anergy or proliferation should reveal predictive biomarkers of progression and of likely response to kinase inhibitors, which could assist therapeutic decisions.

Figure 1.

Differential BCR signaling responses and variable clinical outcome in CLL.

Figure 2.

A model for CLL recirculation and receptor modulation. Engagement of sIgM and CXCR4 by antigen and CXCL12 (from stromal cells), respectively, leads to downmodulation of these receptors and exit of cells into the blood where they are accessible for study. These cells therefore carry a temporary “imprint” of prior tissue-based stimulation. In the absence of stimulation there is recovery of sIgM and CXCR4 expression which primes these cells for re-entry into tissues and antigen restimulation and can be modelled by incubating cells in vitro. New inhibitors targeted towards BCR-associated kinases are likely to act at multiple points (indicated in red boxes), potentially blocking chemokine signals required for homing and migration into tissues, as well as signaling via sIg.

Figure 3.

BCR signaling in naive and anergic B cells. Antigen triggers signaling that differs between naive and chronically stimulated anergic cells. (A) In naive B cells, antigen engagement triggers activation of SRC-family kinases, including LYN, which catalyzes the dual phosphorylation of immunoreceptor tyrosine-based activatory motifs within CD79A and CD79B. This results in recruitment and activation of SYK and triggers activation of downstream effectors, including protein kinases (AKT, BTK) and adaptors (BLNK), as well as lipid metabolism via lipid kinases (PI3K) and lipases PLCγ2 leading to the production of lipid metabolites, PIP3, DAG and IP3. These pathways are naturally inhibited by LYN-dependent activation of both protein (SHP1) and lipid (SHIP1) phosphatases. (B) In anergic cells, chronic BCR engagement favors mono-phosphorylation of CD79A/B which results in weak SYK activation but efficient LYN activation. Activation of phosphatases, especially SHIP1, further suppress SYK-dependent signaling by preventing accumulation of PIP3, and suppresses activation of BCRs and distinct receptors such as CXCR4. Anergic cells are often characterized by raised basal levels of iCa²⁺ and ERK phosphorylation. Adapted from Yarkoni et al.

Figure 4.

Variable BCR signaling responses influence clinical outcome via differential effects on anergy and positive signaling. Antigen engagement appears to be ongoing in all CLL with anergy being the predominant outcome. However, low levels of growth promoting positive signaling may tip the balance towards progressive disease. A temporary imprint of these different tissue-based responses can be detected in circulating CLL cells. Thus, markers of anergy (sIgM downmodulation, raised basal ERK phosphorylation and NFAT) are more prominent in good prognosis subsets, whereas markers of positive signaling (MYC and MCL1) are associated with a poor prognosis. The balance between anergy and positive signaling is likely to be determined by both intrinsic and extrinsic factors.

Figure 5.

Potential mechanisms of suppression of anergy-associated apoptosis in CLL. In normal B cells, anergy is associated with increased expression of BIM, and reduced BAFF responsiveness (boxed). In CLL, overexpression of BCL2, commonly associated with loss of suppressive miRNAs on chromosome 13, appears to sequester BIM to suppress apoptosis. Autocrine or paracrine production of BAFF may also play an important role promoting additional survival signals. BH3 mimetics, such as ABT-199 and navitoclax, reverse sequestration of BIM by survival molecules such as BCL2, potentially revealing the pro-apoptotic potential of anergic CLL cells.