Constitutive activation of distinct BCR-signaling pathways in a subset of CLL patients: a molecular signature of anergy

Abstract

Stimulation through the B-cell antigen receptor (BCR) is believed to be involved in the natural history of chronic lymphocytic leukemia (CLL). Some cases respond to the in vitro cross-linking of surface immunoglobulin (sIg) with effective activation. In contrast, the remaining cases do not respond to such stimulation, thereby resembling B cells anergized after antigen encounter in vivo. However the biochemical differences between the 2 groups are ill defined, and in humans the term B-cell anergy lacks a molecular definition. We examined the expression and activation of key molecules involved in signaling pathways originating from the BCR, and we report that a proportion of CLL patients ( IntroductionChronic lymphocytic leukemia (CLL) is a chronic lymphoid malignancy characterized by the accumulation of CD5 ϩ monoclonal B lymphocytes in primary and secondary lymphoid tissues. Several lines of evidence, including molecular and functional analysis of the monoclonal immunoglobulin (Ig), strongly support the hypothesis that a stimulation through the B-cell receptor (BCR) may be involved in the selection and expansion of the malignant clone. 1,2 At molecular level, the leukemic cells have a preferential IGHV gene usage; in at least half of the cases, they carry somatically mutated IGHV genes 3,4 ; in more than 20% of the cases, they express closely homologous if not identical ("stereotyped") complementarity-determining region 3 (CDR3) sequences on both heavy and light chains. 5-11 All these observations suggest that a limited set of antigenic elements may be specifically recognized by the leukemic Ig.At functional level, it is long known that CLL cells characteristically express low levels of surface Ig (sIg) a property shared by murine B cells anergized by the exposure to antigens (Ags). 1,2 This anergy-related feature does not necessarily indicate a functional impairment of BCR signaling, as at least half of the cases can actually be stimulated in vitro through their sIg, irrespective of their low level. 12 In contrast, the remaining CLL cases are unresponsive to BCR cross-linking, thereby resembling also at functional level B cells anergized in vivo after chronic stimulation by an antigen. 13 Still, and despite extensive studies, the molecular bases for this functional difference are ill defined.In normal B lymphocytes, BCR engagement triggers a cascade of signaling events resulting in enzymatic activation of various kinases and phosphorylation of the relevant substrates. 14 In particular, BCR-proximal Lyn and Syk kinases induce PLC␥2 phosphorylation and Ras activation. Ras binds to and activates Raf kinase that subsequently activates MEK1 and MEK2 that lay immediately upstream of extracellular signal-related kinase (ERK)1/2. In a parallel signaling cascade, PI3K phosphorylates and activates AKT kinase. BCR stimulation activates gene transcription also through activation of nuclear factor (NF)-kB and nuclear factor of activated T cells (NF-AT) transcription factors [14][15][16] (and see the Science STKE website fo...

Pro‐survival signal inhibition by CDK inhibitor dinaciclib in Chronic Lymphocytic Leukaemia

“…The importance of signalling pathways in cell responses to dinaciclib is likely to vary in different CLL patients. For instance, not all patients have a high level ERK activation (Muzio, et al 2008) and AKT Serine 473 phosphorylation, a marker of its activation, was very low in unstimulated CLL cells (Ringshausen, et al 2002). These data supports the view that the signalling networks in CLL cells are very diverse and complex and that drugs that can target more than one of these signals are likely to be effective at treating a greater number of patients.…”

“…Once engaged, it activates pro-survival signals such as the RAF/MEK/ERK, PI3K/AKT, p38, NF-κB and STAT3 pathways, which play a pathogenic role in CLL (Cuní, et al 2004, Hazan-Halevy, et al 2010, Kawauchi, et al 2002, Ogasawara, et al 2003, Pickering, et al 2007, Ringshausen, et al 2002, Rozovski, et al 2014, Sainz-Perez, et al 2006. Nearly half of CLL cases have high levels of constitutively phosphorylated ERK1/2 (Muzio, et al 2008) and in freshly isolated CLL cells, phosphorylated p38 is predominately activated (Sainz-Perez, et al 2006). PI3K is also frequently activated in CLL cells and its inhibition induces apoptosis in some models (Cuní, et al 2004).…”

The functional in vitro response to CD40 ligation reflects a different clinical outcome in patients with chronic lymphocytic leukemia

“…2 The stimulation through the B-cell receptor has been found to be a key factor, 3,4 although the response to B-cell receptor stimulation is heterogeneous and only in a subset of patients the malignant cells are responsive in vitro to the immunoglobulin (Ig) crosslinking that leads to an effective cell activation. 5,6 CLL cases unresponsive to Ig ligation show the functional and biochemical features of B cells anergized by an in vivo antigen (Ag) encounter and a have a better clinical outcome 5,7 as compared with CLL cases responsive to anti-IgM who have a poorer clinical outcome and tend to express other negative prognostic factors (for example, ZAP70 or CD38 positivity, absence of IGHV gene mutations). 6,[8][9][10] There is substantial evidence that CLL cells retain the capacity to respond in vitro also to the so-called 'second signal' delivered through CD40, 11 a member of the tumor necrosis factor receptor family.…”

Proliferation centers in chronic lymphocytic leukemia: the niche where NF-κB activation takes place

“…11,12 NFATc1 can also be activated in response to CD40 or BCR signaling and is active in those cases that are unresponsive to surface Ig ligation. 13 In our series, we observed activated nuclear NFATc1, which can activate BAFF at the transcriptional level, contributing with NF-kB to create a positive feedback loop that favors survival of malignant B cells 14 ( Figure 1). These data suggest that BAFF acts autocrinally, promoting survival of prolymphocytes in PCs, as has been shown for circulating CLL/SLL cells.…”