Clinical Advances in Hematology & Oncology
January 2016, Volume 14, Issue 1
Stefan Koehrer, MD, and Jan A. Burger, MD
The authors are affiliated with the Department of Leukemia at the University of Texas MD Anderson Cancer Center in Houston, Texas.
Jan A. Burger, MD, PhD
Department of Leukemia, Unit 428
The University of Texas
MD Anderson Cancer Center
PO Box 301402
Houston, TX 77230-1402
Tel: (713) 563-1487 or (713) 792-1865
Fax: (713) 794-4297
Abstract: B-cell receptor (BCR) signaling has emerged as a key pathway for the expansion of neoplastic B-cell clones in several B-cell malignancies. The mechanisms that activate BCR signaling differ substantially among subtypes of B-cell lymphoma and leukemia. These include BCR stimulation by foreign or self-antigens, or the acquisition of mutations in components of the BCR pathway that result in autonomous or enhanced antigen-induced BCR signaling. Targeting BCR signaling with selective inhibitors of the BCR-associated kinases Bruton’s tyrosine kinase, spleen tyrosine kinase, and phosphoinositide 3-kinase δ induces high response rates in patients with chronic lymphocytic leukemia, mantle cell lymphoma, Waldenström macroglobulinemia, and diffuse large B-cell lymphoma of the activated B-cell–like subtype and is currently transforming the therapeutic landscape in these diseases. Here we review the mechanisms of BCR activation that govern growth and survival of malignant B cells. We also summarize recent clinical trials of BCR inhibitors, with a focus on the most clinically advanced agents.
B-Cell Receptor Expression and Signaling in Normal B Cells
The B-cell receptor (BCR) is a unique, defining feature of B lymphocytes. The purpose of B-cell development in the bone marrow and secondary lymphatic organs is to generate a diverse set of mature B cells, each equipped with a unique BCR. These unique BCRs allow B cells to recognize foreign antigens and mount specific antibody responses while sparing host (auto) antigens.1,2 B cells lacking a functional BCR rapidly undergo apoptosis.3 The BCR consists of 2 identical immunoglobulin heavy (IgH) chains and 2 identical immunoglobulin light (IgL) chains. The variable domains of IgH and IgL chains are products of gene rearrangements at the pro-B (IgH) and pre-B (IgL) cell stage and define the antigen specificity of the BCR.4 The transmembrane domain anchors the BCR to the cell membrane, where each BCR molecule associates noncovalently with a heterodimer of Igα (CD79a) and Igβ (CD79b).5 The Igα/Igβ heterodimer constitutes the signaling subunit of the BCR complex.6 Within their cytoplasmic tails, Igα and Igβ harbor 2 conserved tyrosine residues as part of a 26 amino acid–long sequence, also referred to as an immunoreceptor tyrosine–based activation motif (ITAM).7 Phosphorylation of these ITAM tyrosine residues through SRC kinases, such as Lck/Yes-related novel protein tyrosine kinase (LYN), FYN, B-lymphoid kinase (BLK), or spleen tyrosine kinase (SYK), marks the first step in signal transduction from the BCR to the nucleus.8,9
Two distinct means of BCR activation have been described: antigen-induced and antigen-independent autonomous (tonic) BCR signaling.10 Engagement of the BCR by antigen induces membrane movement and aggregation of BCR components, ITAM phosphorylation, and consequently the recruitment of SYK to the ITAM residues of Igα and Igβ (Figure 1).11,12 In proximity of the BCR, SYK gets activated through ITAM binding, and SYK phosphorylation occurs through SRC kinases and autophosphorylation.13 Active SYK, together with the SRC kinase LYN, phosphorylate the adaptor proteins CD19, B-cell adaptor for phosphoinositide 3-kinase (BCAP), and B-cell linker protein (BLNK), which are detrimental to the proper activation of BCR downstream signaling. CD19 and BCAP each recruit phosphoinosi-tide 3-kinase (PI3K) to the plasma membrane, where PI3K acts in concert with BLNK in order to activate Bruton’s tyrosine kinase (BTK) and its crucial downstream target phospholipase Cγ2 (PLCγ2).12 PLCγ2 ties BCR engagement to the activation of several signaling cascades. These include calcium mobilization, mitogen-associated protein (MAP) kinase and RAS activation, and activation of protein kinase C β and CARD11, causing recruitment of BCL10 and MALT1 into a multiprotein CBM complex that activates IκB kinase, thereby initiating nuclear factor κB (NFκB) signaling.14,15 Collectively, these signaling events promote B-cell survival and proliferation.15
In contrast to the transient nature of antigen-induced BCR signaling, tonic BCR activation is characterized by low-level signals continuously emerging from the BCR complex independent of antigenic stimulation.16 The presence of tonic BCR signaling was first noted when inducible ablation of BCR expression in mature B cells resulted in the induction of apoptosis and their disappearance from the periphery.3,17 Subsequently, an elegant study by Srinivasan and colleagues revealed that BCR-deficient mature B cells are rescued by constitutive PI3K signaling but not by NFκB or ERK activation, thus suggesting a crucial role for PI3K and protein kinase B (AKT) in tonic BCR signaling and the BCR-dependent survival of mature B cells.18 However, the mechanisms leading to tonic BCR activation remain controversial, potentially involving the self-aggregation of BCR molecules, an altered balance of positive and negative regulators of BCR signaling, or the hijacking of the BCR complex molecules by other receptors, such as the B-cell activating factor of the tumor necrosis factor (TNF) family (BAFF) receptor.19-21
In light of the importance of the BCR in normal B cells, it is not surprising that BCR signaling is also involved in the pathogenesis of a variety of B-cell malignancies. Indeed, the majority of B-cell lymphomas retain BCR expression despite their genomic instability,22,23 and the presence of active BCR signaling (antigen-induced, tonic, or both) has been confirmed for several B-cell lymphomas/leukemias, including chronic lymphocytic leukemia (CLL),24 diffuse large B-cell lymphoma (DLBCL),14 mantle cell lymphoma (MCL),25 hairy cell leukemia (HCL),26 and Burkitt leukemia/lymphoma (BL).27
Chronic Lymphocytic Leukemia
Several lines of evidence suggest the importance of BCR signaling in the pathophysiology of CLL. Markers associated with active BCR signaling, such as zeta chain–associated protein kinase 70kDa (ZAP-70) expression and increased expression of the T cell–attracting chemokines CCL3 and CCL4, are strong predictors of CLL progression and time to treatment.28-32 Moreover, the structure of the BCR itself strongly influences progression of the disease. Based on the degree of somatic hypermutation within the BCR antigen–binding site, CLL patients can be classified as unmutated if they have 98% or more homology with the germline sequence, or mutated if they have less than 98% sequence homology.33 Mutated CLL is typically associated with slower disease progression and better overall survival (OS), whereas unmutated CLL progresses faster, resulting in a shorter time to treatment and shorter survival.34,35
The nature of BCR engagement in CLL B cells remains controversial, with evidence for the presence of both tonic- and antigen-induced BCR activation. The importance of tonic BCR stimulation is supported by constitutive phosphorylation of BCR pathway components in primary CLL B cells, such as LYN,36 SYK,37 ERK,38 and subunits of NFκB.39 Furthermore, overexpression of MYC in murine B cells leads to a CLL-like disease in the absence of antigenic stimulation.40 A highly restricted IGHV gene repertoire,33 culminating in the presence of CLL BCRs with virtually identical (stereotyped) complementarity-determining region 3 (CDR3) sequences, argues for the necessity of antigenic stimulation during CLL pathogenesis.41,42 Consistently, BCRs from unmutated CLL exhibit polyreactivity against a variety of ubiquitous autoantigens, such as nonmuscle myosin heavy chain 2A and vimentin, as well as the Fc tail of immunoglobulin G, single-stranded DNA, double-stranded DNA, lipopolysaccharides, apoptotic cells, insulin, and oxidized low-density lipoproteins.43-48 Foreign antigens also have been implicated in the selection and expansion of CLL cells with certain BCR specificities. For instance, CLL BCRs harboring IGHV1-69 or IGHV3-21 interact with the cytomegalovirus-derived superantigen pUL32.49 Likewise, cases of mutated CLL with IGHV3-07 and short heavy-chain complementarity-determining region 3 (HCDR3) sequences (V3-7Sh) were shown to bind with high affinity to β(1,6)-glucan, a major antigenic determinant of yeast and filamentous fungi.50 A recent study demonstrated antigen-independent cell-autonomous activation of CLL BCRs, resulting in constitutive Ca++ signaling.51 Thus, CLL cells appear to depend on continuous and intermittent BCR signaling that drives cell survival and expansion.
Diffuse Large B-Cell Lymphoma
Similar to CLL, the IGHV gene repertoire in DLBCL is highly biased, with 3 IGHV gene segments (IGHV4-34, IGHV3-23, and IGHV4-39) accounting for one-third of DLBCL BCRs.52-55
Gene expression profiling studies revealed the existence of 2 subtypes of DLBCL, activated B-cell–like (ABC) and germinal center B-cell–like (GCB) DLBCL, based on their resemblance to either activated or germinal center B cells.56,57 It was initially noted that ABC-type DLBCL cells are dependent on the BCR downstream target NFκB.58 Retroviral suppression of NFκB signaling, as well as short hairpin RNA-mediated knockdown of the NFκB pathway components IKBKB, CARD11, MALT1, and BCL10, induce cell death selectively in ABC-type DLBCL.59 In 10% of ABC DLBCL cases, constitutive NFκB activation is caused by mutations in the coiled-coil domain of CARD11.60 In the remaining cases, NFκB activation results from chronic active BCR signaling, which is characterized by surface BCR clustering and resembles antigen-induced BCR activation.14 Consistently, ABC DLBCL cells harboring wild-type CARD11 were sensitive to knockdown of the BCR pathway components CD79A/B, IgM, Igκ, SYK, and BTK.14
In GCB DLBCL, direct evidence for the involvement of BCR signaling in disease pathogenesis is missing. However, Chen and colleagues recently reported the dependency of certain GCB DLBCL cell lines on SYK, suggesting a role for the BCR in subgroups of GCB DLBCL.61,62 In contrast to ABC DLBCL, GCB DLBCL cells do not depend on constitutive NFκB signaling, indicating that BCR signaling in GCB DLBCL mimics tonic rather than antigen-induced BCR activation. Consistently, expression of a constitutively active form of AKT successfully rescues SYK-deficient GCB DLBCL cells.62
Recurrent somatic mutations in essential components of the BCR signaling cascade are another sign of the important role of BCR signaling in DLBCL. In addition to carrying mutations in the CARD11 gene, about 21% of ABC DLBCL patients carry mutations in either CD79A (2.9%) or CD79B (18%).14,63 The majority of CD79A mutations cause deletions of large parts of the ITAM region, including the second ITAM tyrosine residue.14 CD79B mutations are almost exclusively missense mutations resulting in the replacement of the first ITAM tyrosine by a variety of other amino acids.14 Functionally, CD79A/B mutations were shown to increase BCR surface expression levels and to attenuate the autoinhibitory function of LYN, all likely leading to enhanced BCR pathway activation.14
Supporting the presence of tonic BCR signaling in GCB DLBCL, Pfeifer and colleagues recently reported the loss of tumor suppressor phosphatase and tensin homolog (PTEN) expression in 55% of GCB DLBCL cases.64 This was in part owing to genomic alterations, including deletions and mutations in the PTEN gene. PTEN-deficient GCB DLBCL cells are dependent on PI3K signaling and MYC, and are selectively sensitive to the PI3K inhibitor LY294002.64
Mantle Cell Lymphoma
In keeping with the requirement for antigenic stimulation during MCL pathogenesis, MCL BCRs are characterized by the use of a biased IGHV gene repertoire. Stereotyped HCDR3 regions are present in 10.4% of MCL cases.65 Based on somatic hypermutation status, MCL can be further subdivided into mutated and unmutated MCL, with 1 study reporting a more favorable outcome for mutated MCL cases (5-year OS: 59% for mutated MCL vs 40% for unmutated MCL).66 Further suggesting a crucial role for the BCR in subgroups of MCL, gene expression profiling and phosphoproteomic analysis revealed the presence of active BCR signaling in several MCL cell lines.67,68
Despite the strong preclinical evidence for the involvement of antigen and BCR signaling in MCL pathogenesis, the significance of the BCR pathway as a therapeutic target was fully appreciated after clinical trials with the BCR inhibitor ibrutinib (Imbruvica, Pharmacyclics/Janssen), resulting in clinical responses in a majority of MCL patients.69 Subsequently, Rahal and colleagues unraveled the importance of canonical NFκB signaling for the BTK-dependent survival of MCL cell lines, thus providing a biologic rationale for the clinical success of BCR pathway inhibition in MCL.25 It remains to be determined whether this is a consequence of chronic active BCR signaling, as seen in ABC-type DLBCL, or whether MCL cells utilize other means of NFκB activation.
Hairy Cell Leukemia
The most solid evidence for the involvement of the BCR and BCR signaling in HCL pathogenesis arises from structural analyses of HCL BCR molecules. These reveal a biased IGH and IGL variable gene segment repertoire, as well as the existence of mutated and unmutated HCL cases.70,71 Unmutated HCL and the usage of the IGHV4-34 gene segment are considered poor prognostic markers.72,73 Suggesting an involvement of BCR signaling in HCL pathogenesis, Weston-Bell and colleagues reported that BCRs of HCL cells respond to antibody-mediated cross-linking with an increase in cellular calcium levels, ERK phosphorylation, and apoptosis.74 On the contrary, we recently reported on the ability of BCR cross-linking to protect primary HCL cells from undergoing spontaneous apoptosis in vitro.26 Importantly, pretreatment with the BTK inhibitor ibrutinib completely abrogated these effects, suggesting therapeutic relevance of the BCR pathway in HCL.
BL joined the list of BCR-dependent B-cell malignancies recently, after a genomic screen of sporadic, endemic, and HIV-associated BL cases identified mutations in the transcription factor gene TCF3 and its negative regulator gene ID3 in more than 70% of cases.27 Extensive functional analysis revealed that both types of mutations entail the activation of TCF3 transcriptional activity, ultimately affecting the expression of several BCR pathway components, including upregulation of IgH and IgL and the suppression of SHP-1, a key negative regulator of BCR signaling.27 Consequently, mutations in TCF3 and ID3 were shown to enhance tonic BCR signaling, which, in cooperation with the deregulation of MYC, fuels proliferation and survival of BL cells and serves as a potential target for therapeutic intervention.
BCR Signaling Inhibitors in B-Cell Malignancies
The critical role of the BCR pathway in the pathogenesis of autoimmune disorders and several B-cell malignancies led to the development of kinase inhibitors that target BCR signaling. All currently available BCR inhibitors belong to the group of small-molecule tyrosine kinase inhibitors, and can be classified according to their target specificity in BTK, PI3K, and SYK inhibitors (Figure 2). In the following sections, we discuss their mechanism of action and their activity, with a focus on the clinically most advanced agents (see Table).
Bruton’s Tyrosine Kinase
BTK belongs to the TEC family of nonreceptor tyrosine kinases, and constitutes an essential part of the BCR signaling cascade. BTK is primarily expressed in hematopoietic cells, particularly in B cells and not in T cells or plasma cells.75 Upon BCR activation, increasing levels of the PI3K product phosphatidylinositol (3,4,5)-triphosphate recruit BTK to the plasma membrane via its pleckstrin homology domain.76 LYN and SYK subsequently activate BTK in concert through phosphorylation, ultimately resulting in activation of the transcription factor NFκB, B-cell proliferation, and B-cell survival.77 In addition to its involvement in BCR signaling, BTK has been shown to transduce signals from chemokine (CXCR4 and CXCR5) and integrin receptors, thus making it also a crucial component in the regulation of B-cell migration and tissue homing.78,79
Ibrutinib, formerly called PCI-32765, is the most clinically advanced BTK inhibitor. Ibrutinib blocks the enzymatic activity of BTK through covalent binding to a conserved cysteine residue (Cys-481) in the active site of BTK.80 In samples from patients with CLL, ibrutinib has been shown to block BCR-derived survival signals and downregulates the secretion of the BCR-dependent chemokines CCL3 and CCL4 in vitro, as well as in CLL patients receiving ibrutinib.81 Ibrutinib also antagonizes the effects of the prosurvival factors CD40 ligand, BAFF, and interleukin 6 on CLL cells.82 Apart from BCR signaling, ibrutinib has been shown to interfere with integrin-mediated CLL cell adhesion and with the migration of CLL cells towards the tissue homing factors CXCL12 and CXCL13.83 Alongside the compelling in vitro data, ibrutinib also effectively thwarted disease progression in the TCL1 mouse model of CLL.81
In DLBCL, ibrutinib blocks BTK-dependent NFκB activation, resulting in the selective killing of ABC-type DLBCL cells.14 These effects were augmented by the addition of lenalidomide (Revlimid, Celgene), resulting in enhanced cytotoxicity and superior effects in xenograft models of ABC-type DLBCL.84 In subsets of MCL, ibrutinib was shown to exert its effects in a similar fashion, depriving MCL cells from crucial NFκB survival signals and inducing apoptosis.25
In clinical trials, ibrutinib exhibited activity in CLL, MCL, Waldenström macroglobulinemia, and ABC DLBCL patients. In CLL, Byrd and colleagues reported that single-agent ibrutinib induced an overall response rate (ORR) of 71% in relapsed or refractory patients. An additional 15% to 20% of patients had a partial response with lymphocytosis.85 The response was independent of clinical and genomic risk factors present before treatment. At 26 months, the estimated progression-free survival (PFS) rate was 75% and the OS rate was 83%. O’Brien and colleagues assessed the safety and efficacy of single-agent ibrutinib in treatment-naive patients aged 65 years and older.86 Ibrutinib was well tolerated, with the most common side effects being mild to moderate diarrhea, nausea, fatigue, and hypertension. The objective response rate in this cohort of patients was 71%, with an additional 13% achieving a partial response with lymphocytosis. In a phase 3 clinical trial of ibrutinib vs the anti-CD20 antibody ofatumumab (Arzerra, GlaxoSmithKline) in previously treated CLL, the ORR was 42.6% in the ibrutinib group and 4.1% in the ofatumumab group.87 At a median follow-up of 9.4 months, ibrutinib also significantly improved PFS and OS. The results of these studies led to US Food and Drug Administration (FDA) approval of ibrutinib for the treatment of CLL patients who received at least 1 prior therapy and for all CLL patients with the 17p deletion. In a recent study, we investigated the activity and safety of the combination of ibrutinib with the anti-CD20 antibody rituximab (Rituxan, Genentech/Biogen Idec) in high-risk CLL. All participants had high-risk cytogenetic abnormalities (deletion 17p, deletion 11q, or TP53 mutations) or a PFS of less than 36 months after first-line chemoimmunotherapy. Despite these high-risk features, the 18-month PFS and OS of ibrutinib plus rituximab were 78% and 84%, respectively.88 Further evaluating the activity of ibrutinib in high-risk CLL, Farooqui and colleagues recently reported a 92% objective response rate (50% partial response rate; 42% rate of partial response with lymphocytosis) in previously untreated and relapsed/refractory patients with CLL who had TP53 aberrations.89 In a 3-year follow-up study, single-agent ibrutinib was well tolerated, and prolonged ibrutinib therapy was associated with durable remissions that improved in quality over time. Disease progression was rare and mainly occurred in subgroups of patients characterized by extensive prior therapy and high-risk cytogenetic features (deletion 17p and deletion 11q).
In addition to showing benefits in patients with CLL, the initial clinical evaluation of ibrutinib also suggested that MCL patients may benefit from ibrutinib therapy.69 In a subsequent phase 2 study, Wang and colleagues reported on single-agent efficacy of ibrutinib in patients with relapsed or refractory MCL, with a complete response rate of 21% and a partial response rate of 47%.90 The estimated median duration of response was 17.5 months, the estimated median PFS was 13.9 months, and the estimated OS rate was 58% at 18 months. The FDA consequently granted accelerated approval to ibrutinib for the treatment of MCL patients who have received at least 1 prior therapy.
Of note, in CLL and MCL patients, ibrutinib induces the rapid egress of tumor B cells from lymphatic tissues into the peripheral blood. Clinically, this is associated with a rapid reduction in lymph node size and peripheral blood lymphocytosis. The extent of lymphocytosis is variable among patients, is asymptomatic, and usually resolves during the first months of therapy. In a recent study by Woyach and colleagues, prolonged lymphocytosis (>12 months) in CLL patients receiving ibrutinib did not predict an inferior outcome.91 Similar effects on lymph node size and peripheral blood lymphocyte counts have been observed with PI3K and SYK inhibitors, suggesting that this is a BCR inhibitor–specific phenomenon, most likely due to their effects on cell migration and tissue homing.
Wilson and colleagues recently reported the results of a phase 1/2 clinical trial of single-agent ibrutinib in relapsed/refractory DLBCL patients. Ibrutinib therapy resulted in complete or partial responses in 37% of ABC-type but only in 5% of GCB-type DLBCL, corroborating the preclinical data of ibrutinib in DLBCL.92 Patients responded frequently in ABC-type cases with BCR mutations (5/9), as did patients with combined BCR and MYD88 mutations (4/5). CARD11 mutations were associated with ibrutinib resistance.
Phosphoinositide 3-Kinase δ
PI3Ks are key regulators of proliferation and survival in various cell types. Based on sequence homology and substrate specificity, PI3Ks can be divided into 3 classes: I, II, and III. Class I PI3Ks, which are particularly involved in cancer,93 comprise PI3Kα, PI3Kβ, PI3Kγ, and PI3Kδ. The PI3Kα and PI3Kβ isoforms are ubiquitously expressed, whereas the PI3Kγ and PI3Kδ isoforms are limited to hematopoietic cells. PI3Kγ has a role in T-cell activation, and PI3Kδ plays a critical role in B-cell homeostasis and function.94 Mice lacking a functional PI3Kδ isoform are characterized by a highly dysfunctional B-cell compartment resulting in reduced numbers of B1 and marginal zone B cells, low levels of immunoglobulins, poor responses to immunization, and defective BCR and CD40 signaling. In B-cell malignancies, an array of cell surface receptors have been shown to activate PI3Ks, including the BCR and CXCR4.95,96
Idelalisib (Zydelig, Gilead), previously called CAL-101 or GS-1101, is a highly selective PI3Kδ inhibitor with preclinical activity in a variety of B-cell malignancies.97 Idelalisib was shown to effectively block constitutive and BCR-induced PI3K activation in several B-cell lymphomas, including CLL and MCL.98 In addition to its effect on the BCR signaling cascade, idelalisib was also shown to counteract the effects of BAFF, TNF-α, and fibronectin stimulation on primary CLL cells.99
Based on the encouraging preclinical data, Gopal and colleagues conducted a single-group, open-label phase 2 study of idelalisib monotherapy in relapsed/refractory indolent non-Hodgkin lymphomas.100 The study included patients with follicular lymphoma (72 patients), small lymphocytic lymphoma (28 patients), marginal-zone lymphoma (15 patients), and lymphoplasmacytic lymphoma with or without Waldenström macroglobulinemia (10 patients). The response rate was 57%, with similar response rates across all subtypes. The most common grade 3 or higher adverse events were neutropenia (27%), elevations in aminotransferase levels (13%), diarrhea (13%), and pneumonia (7%). The results of this study led to the accelerated approval of idelalisib by the FDA for the treatment of patients with relapsed follicular B-cell non-Hodgkin lymphoma or relapsed small lymphocytic lymphoma. Furman and colleagues evaluated the efficacy of idelalisib in combination with rituximab vs rituximab plus placebo in relapsed CLL.101 This multicenter phase 3 trial enrolled 220 patients who were randomly assigned to receive rituximab and either 150 mg of idelalisib or placebo twice daily. Owing to the superior efficacy of idelalisib, the study was terminated early at the first interim analysis. ORRs for patients receiving idelalisib vs placebo were 81% and 13%, respectively. OS at 12 months was 92% for rituximab plus idelalisib and 81% for rituximab plus placebo. Serious adverse events occurred in 40% of the patients receiving idelalisib and rituximab and in 35% of those receiving placebo and rituximab. The FDA consequently granted approval of idelalisib for the treatment of patients with relapsed CLL.
Spleen Tyrosine Kinase
SYK is a nonreceptor tyrosine kinase that belongs to the family of SYK/ZAP-70 protein tyrosine kinases. SYK is predominantly expressed in hematopoietic cells and is crucial for the function of an array of cell surface receptors, including the BCR, integrin receptors, Fc receptors, and pattern recognition receptors of the innate immune system.102 Homozygous inactivation of SYK in mice is consistently associated with perinatal lethality, and selective deletion of SYK in the hematopoietic system severely impairs B-cell differentiation, with a block at the pro-B cell to pre-B cell transition.103,104 Moreover, in vivo studies recently demonstrated that SYK is critical for survival and maintenance of mature normal and malignant B cells.19,105
The SYK inhibitor fostamatinib disodium (R788, FosD) was the first BCR inhibitor under clinical evaluation for the treatment of BCR-dependent malignancies. The initial phase 1/2 study of fostamatinib in recurrent non-Hodgkin lymphoma patients reported an objective response rate of 22% for all cases, and 55% for CLL patients in particular.106 In vitro, fostamatinib kills DLBCL and CLL cells. Fostamatinib also blocks migration towards the tissue-homing chemokines CXCL12 and CXCL13 in CLL cells.61,107 Despite these encouraging results, further development of this drug has focused on rheumatoid arthritis, and most recently has focused on idiopathic thrombocytopenic purpura.108 Alternative SYK-specific inhibitors are under development and have demonstrated promising preclinical and clinical activity.109 Sharman and colleagues recently reported results of a phase 2 clinical trial of the SYK-specific inhibitor entospletinib (GS-9973) in relapsed/refractory CLL and non-Hodgkin lymphoma.110 The reported objective response rate in CLL patients was 61%, including 3 patients (7.3%) who achieved nodal response with persistent lymphocytosis. Cerdulatinib (PRT062070), a combined SYK/JAK inhibitor with promising in vitro activity in DLBCL and Burkitt lymphoma models,111 is currently being evaluated in a phase 1 dose-escalation study in CLL and non-Hodgkin lymphoma patients.
BCR inhibitors are currently transforming the therapeutic landscape in several B-cell malignancies. The induction of durable remissions, irrespective of established risk factors, alongside the favorable toxicity profile—especially a lack of myelotoxicity—has led to a wide use of BCR inhibitors in patients with CLL, MCL, and follicular lymphoma. Response rates and durability are particularly high in patients with CLL, whereas response rates and/or duration appear to be lower in other B-cell malignancies. The clinical success of these novel agents has fueled a series of translational studies to study the consequences of BCR inhibition in humans. As with any major discovery, these new concepts are challenging us with new questions. For instance, the available data on the efficacy of BCR inhibitors in B-cell malignancies are mostly derived from single-agent studies in high-risk and heavily pretreated patient populations, and do not take into account potential synergisms between BCR inhibitors and established therapeutic agents. However, with the FDA approval of ibrutinib and idelalisib, follow-up clinical trials are now testing the benefit of combination regimens to better define the optimal use of BCR signaling inhibitors in patients with B-cell malignancies.
This work was supported in part by a Leukemia & Lymphoma Society Scholar in Clinical Research Award to Dr. Burger, by the MD Anderson Cancer Center Moon Shots Program, and by a Cancer Center Support Grant (CA016672) from the National Cancer Institute.
Dr Burger has received research funding from Pharmacyclics and Gilead. Dr Koehrer has declared no competing financial interests.
1. Reth M, Hombach J, Wienands J, et al. The B-cell antigen receptor complex. Immunol Today. 1991;12(6):196-201.
2. Macallan DC, Wallace DL, Zhang Y, et al. B-cell kinetics in humans: rapid turnover of peripheral blood memory cells. Blood. 2005;105(9):3633-3640.
3. Lam K-P, Kühn R, Rajewsky K. In vivo ablation of surface immunoglobulin on mature B cells by inducible gene targeting results in rapid cell death. Cell. 1997;90(6):1073-1083.
4. LeBien TW. Fates of human B-cell precursors. Blood. 2000;96(1):9-23.
5. Schamel WW, Reth M. Monomeric and oligomeric complexes of the B cell antigen receptor. Immunity. 2000;13(1):5-14.
6. Papavasiliou F, Jankovic M, Suh H, Nussenzweig MC. The cytoplasmic domains of immunoglobulin (Ig) alpha and Ig beta can independently induce the precursor B cell transition and allelic exclusion. J Exp Med. 1995;182(5):1389-1394.
7. Reth M. Antigen receptor tail clue. Nature. 1989;338(6214):383-384.
8. Johnson SA, Pleiman CM, Pao L, Schneringer J, Hippen K, Cambier JC. Phosphorylated immunoreceptor signaling motifs (ITAMs) exhibit unique abilities to bind and activate Lyn and Syk tyrosine kinases. J Immunol. 1995;155(10):4596-4603.
9. Rolli V, Gallwitz M, Wossning T, et al. Amplification of B cell antigen receptor signaling by a Syk/ITAM positive feedback loop. Mol Cell. 2002;10(5):1057-1069.
10. LeBien TW, Tedder TF. B lymphocytes: how they develop and function. Blood. 2008;112(5):1570-1580.
11. Treanor B, Depoil D, Bruckbauer A, Batista FD. Dynamic cortical actin remodeling by ERM proteins controls BCR microcluster organization and integrity. J Exp Med. 2011;208(5):1055-1068.
12. Dal Porto JM, Gauld SB, Merrell KT, Mills D, Pugh-Bernard AE, Cambier J. B cell antigen receptor signaling 101. Mol Immunol. 2004;41(6-7):599-613.
13. Rowley RB, Burkhardt AL, Chao HG, Matsueda GR, Bolen JB. Syk protein-tyrosine kinase is regulated by tyrosine-phosphorylated Ig alpha/Ig beta immunoreceptor tyrosine activation motif binding and autophosphorylation. J Biol Chem. 1995;270(19):11590-11594.
14. Davis RE, Ngo VN, Lenz G, et al. Chronic active B-cell-receptor signalling in diffuse large B-cell lymphoma. Nature. 2010;463(7277):88-92.
15. Shaffer AL III, Young RM, Staudt LM. Pathogenesis of human B cell lymphomas. Annu Rev Immunol. 2012;30:565-610.
16. Monroe JG. ITAM-mediated tonic signalling through pre-BCR and BCR complexes. Nat Rev Immunol. 2006;6(4):283-294.
17. Kraus M, Alimzhanov MB, Rajewsky N, Rajewsky K. Survival of resting mature B lymphocytes depends on BCR signaling via the Igalpha/beta heterodimer. Cell. 2004;117(6):787-800.
18. Srinivasan L, Sasaki Y, Calado DP, et al. PI3 kinase signals BCR-dependent mature B cell survival. Cell. 2009;139(3):573-586.
19. Schweighoffer E, Vanes L, Nys J, et al. The BAFF receptor transduces survival signals by co-opting the B cell receptor signaling pathway. Immunity. 2013;38(3):475-488.
20. Yang J, Reth M. Oligomeric organization of the B-cell antigen receptor on resting cells. Nature. 2010;467:465-469.
21. Pierce SK, Liu W. The tipping points in the initiation of B cell signalling: how small changes make big differences. Nat Rev Immunol. 2010;10(11):767-777.
22. Young RM, Staudt LM. Targeting pathological B cell receptor signalling in lymphoid malignancies. Nat Rev Drug Discov. 2013;12(3):229-243.
23. Küppers R. Mechanisms of B-cell lymphoma pathogenesis. Nat Rev Cancer. 2005;5(4):251-262.
24. Herishanu Y, Pérez-Galán P, Liu D, et al. The lymph node microenvironment promotes B-cell receptor signaling, NF-kappaB activation, and tumor proliferation in chronic lymphocytic leukemia. Blood. 2011;117(2):563-574.
25. Rahal R, Frick M, Romero R, et al. Pharmacological and genomic profiling identifies NF-κB-targeted treatment strategies for mantle cell lymphoma. Nat Med. 2014;20(1):87-92.
26. Sivina M, Kreitman RJ, Arons E, Ravandi F, Burger JA. The bruton tyrosine kinase inhibitor ibrutinib (PCI-32765) blocks hairy cell leukaemia survival, proliferation and B cell receptor signalling: a new therapeutic approach. Br J Haematol. 2014;166(2):177-188.
27. Schmitz R, Young RM, Ceribelli M, et al. Burkitt lymphoma pathogenesis and therapeutic targets from structural and functional genomics. Nature. 2012;490(7418):116-120.
28. Gobessi S, Laurenti L, Longo PG, Sica S, Leone G, Efremov DG. ZAP-70 enhances B-cell-receptor signaling despite absent or inefficient tyrosine kinase activation in chronic lymphocytic leukemia and lymphoma B cells. Blood. 2007;109(5):2032-2039.
29. Nolz JC, Tschumper RC, Pittner BT, Darce JR, Kay NE, Jelinek DF. ZAP-70 is expressed by a subset of normal human B-lymphocytes displaying an activated phenotype. Leukemia. 2005;19(6):1018-1024.
30. Cutrona G, Colombo M, Matis S, et al. B lymphocytes in humans express ZAP-70 when activated in vivo. Eur J Immunol. 2006;36(3):558-569.
31. Sivina M, Hartmann E, Kipps TJ, et al. CCL3 (MIP-1α) plasma levels and the risk for disease progression in chronic lymphocytic leukemia. Blood. 2011;117(5):1662-1669.
32. Yan XJ, Dozmorov I, Li W, et al. Identification of outcome-correlated cytokine clusters in chronic lymphocytic leukemia. Blood. 2011;118(19):5201-5210.
33. Fais F, Ghiotto F, Hashimoto S, et al. Chronic lymphocytic leukemia B cells express restricted sets of mutated and unmutated antigen receptors. J Clin Invest. 1998;102(8):1515-1525.
34. Damle RN, Wasil T, Fais F, et al. Ig V gene mutation status and CD38 expression as novel prognostic indicators in chronic lymphocytic leukemia. Blood. 1999;94(6):1840-1847.
35. Hamblin TJ, Davis Z, Gardiner A, Oscier DG, Stevenson FK. Unmutated Ig V(H) genes are associated with a more aggressive form of chronic lymphocytic leukemia. Blood. 1999;94(6):1848-1854.
36. Contri A, Brunati AM, Trentin L, et al. Chronic lymphocytic leukemia B cells contain anomalous Lyn tyrosine kinase, a putative contribution to defective apoptosis. J Clin Invest. 2005;115(2):369-378.
37. Gobessi S, Laurenti L, Longo PG, et al. Inhibition of constitutive and BCR-induced Syk activation downregulates Mcl-1 and induces apoptosis in chronic lymphocytic leukemia B cells. Leukemia. 2009;23(4):686-697.
38. Muzio M, Apollonio B, Scielzo C, et al. Constitutive activation of distinct BCR-signaling pathways in a subset of CLL patients: a molecular signature of anergy. Blood. 2008;112(1):188-195.
39. Hewamana S, Alghazal S, Lin TT, et al. The NF-kappaB subunit Rel A is associated with in vitro survival and clinical disease progression in chronic lymphocytic leukemia and represents a promising therapeutic target. Blood. 2008;111(9):4681-4689.
40. Refaeli Y, Young RM, Turner BC, Duda J, Field KA, Bishop JM. The B cell antigen receptor and overexpression of MYC can cooperate in the genesis of B cell lymphomas. PLoS Biol. 2008;6(6):e152.
41. Messmer BT, Albesiano E, Efremov DG, et al. Multiple distinct sets of stereotyped antigen receptors indicate a role for antigen in promoting chronic lymphocytic leukemia. J Exp Med. 2004;200(4):519-525.
42. Stamatopoulos K, Belessi C, Moreno C, et al. Over 20% of patients with chronic lymphocytic leukemia carry stereotyped receptors: pathogenetic implications and clinical correlations. Blood. 2007;109(1):259-270.
43. Bröker BM, Klajman A, Youinou P, et al. Chronic lymphocytic leukemic (CLL) cells secrete multispecific autoantibodies. J Autoimmun. 1988;1(5):469-481.
44. Sthoeger ZM, Wakai M, Tse DB, et al. Production of autoantibodies by CD5-expressing B lymphocytes from patients with chronic lymphocytic leukemia. J Exp Med. 1989;169(1):255-268.
45. Kostareli E, Gounari M, Janus A, et al. Antigen receptor stereotypy across B-cell lymphoproliferations: the case of IGHV4-59/IGKV3-20 receptors with rheumatoid factor activity. Leukemia. 2012;26(5):1127-1131.
46. Hoogeboom R, Wormhoudt TA, Schipperus MR, et al. A novel chronic lymphocytic leukemia subset expressing mutated IGHV3-7-encoded rheumatoid factor B-cell receptors that are functionally proficient. Leukemia. 2013;27(3):738-740.
47. Hervé M, Xu K, Ng YS, et al. Unmutated and mutated chronic lymphocytic leukemias derive from self-reactive B cell precursors despite expressing different antibody reactivity. J Clin Invest. 2005;115(6):1636-1643.
48. Lanemo Myhrinder A, Hellqvist E, Sidorova E, et al. A new perspective: molecular motifs on oxidized LDL, apoptotic cells, and bacteria are targets for chronic lymphocytic leukemia antibodies. Blood. 2008;111(7):3838-3848.
49. Steininger C, Widhopf GF II, Ghia EM, et al. Recombinant antibodies encoded by IGHV1-69 react with pUL32, a phosphoprotein of cytomegalovirus and B-cell superantigen. Blood. 2012;119(10):2293-2301.
50. Hoogeboom R, van Kessel KP, Hochstenbach F, et al. A mutated B cell chronic lymphocytic leukemia subset that recognizes and responds to fungi. J Exp Med. 2013;210(1):59-70.
51. Dühren-von Minden M, Übelhart R, Schneider D, et al. Chronic lymphocytic leukaemia is driven by antigen-independent cell-autonomous signalling. Nature. 2012;489(7415):309-312.
52. Küppers R, Rajewsky K, Hansmann ML. Diffuse large cell lymphomas are derived from mature B cells carrying V region genes with a high load of somatic mutation and evidence of selection for antibody expression. Eur J Immunol. 1997;27(6):1398-1405.
53. Lossos IS, Okada CY, Tibshirani R, et al. Molecular analysis of immunoglobulin genes in diffuse large B-cell lymphomas. Blood. 2000;95(5):1797-1803.
54. Nakamura N, Kuze T, Hashimoto Y, et al. Analysis of the immunoglobulin heavy chain gene variable region of CD5-positive and -negative diffuse large B cell lymphoma. Leukemia. 2001;15(3):452-457.
55. Sebastián E, Alcoceba M, Balanzategui A, et al. Molecular characterization of immunoglobulin gene rearrangements in diffuse large B-cell lymphoma: antigen-driven origin and IGHV4-34 as a particular subgroup of the non-GCB subtype. Am J Pathol. 2012;181(5):1879-1888.
56. Alizadeh AA, Eisen MB, Davis RE, et al. Distinct types of diffuse large B-cell lymphoma identified by gene expression profiling. Nature. 2000;403(6769):503-511.
57. Rosenwald A, Wright G, Chan WC, et al; Lymphoma/Leukemia Molecular Profiling Project. The use of molecular profiling to predict survival after chemotherapy for diffuse large-B-cell lymphoma. N Engl J Med. 2002;346(25):1937-1947.
58. Davis RE, Brown KD, Siebenlist U, Staudt LM. Constitutive nuclear factor kappaB activity is required for survival of activated B cell-like diffuse large B cell lymphoma cells. J Exp Med. 2001;194(12):1861-1874.
59. Ngo VN, Davis RE, Lamy L, et al. A loss-of-function RNA interference screen for molecular targets in cancer. Nature. 2006;441(7089):106-110.
60. Lenz G, Davis RE, Ngo VN, et al. Oncogenic CARD11 mutations in human diffuse large B cell lymphoma. Science. 2008;319(5870):1676-1679.
61. Chen L, Monti S, Juszczynski P, et al. SYK-dependent tonic B-cell receptor signaling is a rational treatment target in diffuse large B-cell lymphoma. Blood. 2008;111(4):2230-2237.
62. Chen L, Monti S, Juszczynski P, et al. SYK inhibition modulates distinct PI3K/AKT- dependent survival pathways and cholesterol biosynthesis in diffuse large B cell lymphomas. Cancer Cell. 2013;23(6):826-838.
63. Bohers E, Mareschal S, Bouzelfen A, et al. Targetable activating mutations are very frequent in GCB and ABC diffuse large B-cell lymphoma. Genes Chromosomes Cancer. 2014;53(2):144-153.
64. Pfeifer M, Grau M, Lenze D, et al. PTEN loss defines a PI3K/AKT pathway-dependent germinal center subtype of diffuse large B-cell lymphoma. Proc Natl Acad Sci U S A. 2013;110(30):12420-12425.
65. Hadzidimitriou A, Agathangelidis A, Darzentas N, et al. Is there a role for antigen selection in mantle cell lymphoma? Immunogenetic support from a series of 807 cases. Blood. 2011;118(11):3088-3095.
66. Navarro A, Clot G, Royo C, et al. Molecular subsets of mantle cell lymphoma defined by the IGHV mutational status and SOX11 expression have distinct biologic and clinical features. Cancer Res. 2012;72(20):5307-5316.
67. Pighi C, Gu T-L, Dalai I, et al. Phospho-proteomic analysis of mantle cell lymphoma cells suggests a pro-survival role of B-cell receptor signaling. Cell Oncol (Dordr). 2011;34(2):141-153.
68. Rinaldi A, Kwee I, Taborelli M, et al. Genomic and expression profiling identifies the B-cell associated tyrosine kinase Syk as a possible therapeutic target in mantle cell lymphoma. Br J Haematol. 2006;132(3):303-316.
69. Advani RH, Buggy JJ, Sharman JP, et al. Bruton tyrosine kinase inhibitor ibrutinib (PCI-32765) has significant activity in patients with relapsed/refractory B-cell malignancies. J Clin Oncol. 2013;31(1):88-94.
70. Forconi F, Sahota SS, Raspadori D, et al. Hairy cell leukemia: at the crossroad of somatic mutation and isotype switch. Blood. 2004;104(10):3312-3317.
71. Arons E, Sunshine J, Suntum T, Kreitman RJ. Somatic hypermutation and VH gene usage in hairy cell leukaemia. Br J Haematol. 2006;133(5):504-512.
72. Forconi F, Sozzi E, Cencini E, et al. Hairy cell leukemias with unmutated IGHV genes define the minor subset refractory to single-agent cladribine and with more aggressive behavior. Blood. 2009;114(21):4696-4702.
73. Arons E, Suntum T, Stetler-Stevenson M, Kreitman RJ. VH4-34+ hairy cell leukemia, a new variant with poor prognosis despite standard therapy. Blood. 2009;114(21):4687-4695.
74. Weston-Bell NJ, Forconi F, Kluin-Nelemans HC, Sahota SS, Variant B. Variant B cell receptor isotype functions differ in hairy cell leukemia with mutated BRAF and IGHV genes. PLoS One. 2014;9(1):e86556.
75. Genevier HC, Hinshelwood S, Gaspar HB, et al. Expression of Bruton’s tyrosine kinase protein within the B cell lineage. Eur J Immunol. 1994;24(12):3100-3105.
76. Mohamed AJ, Yu L, Bäckesjö CM, et al. Bruton’s tyrosine kinase (Btk): function, regulation, and transformation with special emphasis on the PH domain. Immunol Rev. 2009;228(1):58-73.
77. Petro JB, Rahman SM, Ballard DW, Khan WN. Bruton’s tyrosine kinase is required for activation of IkappaB kinase and nuclear factor kappaB in response to B cell receptor engagement. J Exp Med. 2000;191(10):1745-1754.
78. de Gorter DJ, Beuling EA, Kersseboom R, et al. Bruton’s tyrosine kinase and phospholipase Cgamma2 mediate chemokine-controlled B cell migration and homing. Immunity. 2007;26(1):93-104.
79. Spaargaren M, Beuling EA, Rurup ML, et al. The B cell antigen receptor controls integrin activity through Btk and PLCgamma2. J Exp Med. 2003;198(10):1539-1550.
80. Honigberg LA, Smith AM, Sirisawad M, et al. The Bruton tyrosine kinase inhibitor PCI-32765 blocks B-cell activation and is efficacious in models of autoimmune disease and B-cell malignancy. Proc Natl Acad Sci U S A. 2010;107(29):13075-13080.
81. Ponader S, Chen S-S, Buggy JJ, et al. The Bruton tyrosine kinase inhibitor PCI-32765 thwarts chronic lymphocytic leukemia cell survival and tissue homing in vitro and in vivo. Blood. 2012;119(5):1182-1189.
82. Herman SEM, Gordon AL, Hertlein E, et al. Bruton tyrosine kinase represents a promising therapeutic target for treatment of chronic lymphocytic leukemia and is effectively targeted by PCI-32765. Blood. 2011;117(23):6287-6296.
83. de Rooij MF, Kuil A, Geest CR, et al. The clinically active BTK inhibitor PCI-32765 targets B-cell receptor- and chemokine-controlled adhesion and migration in chronic lymphocytic leukemia. Blood. 2012;119(11):2590-2594.
84. Yang Y, Shaffer AL III, Emre NC, et al. Exploiting synthetic lethality for the therapy of ABC diffuse large B cell lymphoma. Cancer Cell. 2012;21(6):723-737.
85. Byrd JC, Furman RR, Coutre SE, et al. Targeting BTK with ibrutinib in relapsed chronic lymphocytic leukemia. N Engl J Med. 2013;369(1):32-42.
86. O’Brien S, Furman RR, Coutre SE, et al. Ibrutinib as initial therapy for elderly patients with chronic lymphocytic leukaemia or small lymphocytic lymphoma: an open-label, multicentre, phase 1b/2 trial. Lancet Oncol. 2014;15(1):48-58.
87. Byrd JC, Brown JR, O’Brien S, et al; RESONATE Investigators. Ibrutinib versus ofatumumab in previously treated chronic lymphoid leukemia. N Engl J Med. 2014;371(3):213-223.
88. Burger JA, Keating MJ, Wierda WG, et al. Safety and activity of ibrutinib plus rituximab for patients with high-risk chronic lymphocytic leukaemia: a single-arm, phase 2 study. Lancet Oncol. 2014;15(10):1090-1099.
89. Farooqui MZH, Valdez J, Martyr S, et al. Ibrutinib for previously untreated and relapsed or refractory chronic lymphocytic leukaemia with TP53 aberrations: a phase 2, single-arm trial. Lancet Oncol. 2015;16(2):169-176.
90. Wang ML, Rule S, Martin P, et al. Targeting BTK with ibrutinib in relapsed or refractory mantle-cell lymphoma. N Engl J Med. 2013;369(6):507-516.
91. Woyach JA, Smucker K, Smith LL, et al. Prolonged lymphocytosis during ibrutinib therapy is associated with distinct molecular characteristics and does not indicate a suboptimal response to therapy. Blood. 2014;123(12):1810-1817.
92. Wilson WH, Young RM, Schmitz R, et al. Targeting B cell receptor signaling with ibrutinib in diffuse large B cell lymphoma. Nat Med. 2015;21(8):922-926; advance online publication.
93. Vivanco I, Sawyers CL. The phosphatidylinositol 3-Kinase AKT pathway in human cancer. Nat Rev Cancer. 2002;2(7):489-501.
94. Jou ST, Carpino N, Takahashi Y, et al. Essential, nonredundant role for the phosphoinositide 3-kinase p110delta in signaling by the B-cell receptor complex. Mol Cell Biol. 2002;22(24):8580-8591.
95. Longo PG, Laurenti L, Gobessi S, Sica S, Leone G, Efremov DG. The Akt/Mcl-1 pathway plays a prominent role in mediating antiapoptotic signals downstream of the B-cell receptor in chronic lymphocytic leukemia B cells. Blood. 2008;111(2):846-855.
96. Burger JA, Burger M, Kipps TJ. Chronic lymphocytic leukemia B cells express functional CXCR4 chemokine receptors that mediate spontaneous migration beneath bone marrow stromal cells. Blood. 1999;94(11):3658-3667.
97. Lannutti BJ, Meadows SA, Herman SEM, et al. CAL-101, a p110δ selective phosphatidylinositol-3-kinase inhibitor for the treatment of B-cell malignancies, inhibits PI3K signaling and cellular viability. Blood. 2011;117(2):591-594.
98. Hoellenriegel J, Meadows SA, Sivina M, et al. The phosphoinositide 3′-kinase delta inhibitor, CAL-101, inhibits B-cell receptor signaling and chemokine networks in chronic lymphocytic leukemia. Blood. 2011;118(13):3603-3612.
99. Herman SEM, Gordon AL, Wagner AJ, et al. Phosphatidylinositol 3-kinase-δ inhibitor CAL-101 shows promising preclinical activity in chronic lymphocytic leukemia by antagonizing intrinsic and extrinsic cellular survival signals. Blood. 2010;116(12):2078-2088.
100. Gopal AK, Kahl BS, de Vos S, et al. PI3Kδ inhibition by idelalisib in patients with relapsed indolent lymphoma. N Engl J Med. 2014;370(11):1008-1018.
101. Furman RR, Sharman JP, Coutre SE, et al. Idelalisib and rituximab in relapsed chronic lymphocytic leukemia. N Engl J Med. 2014;370(11):997-1007.
102. Mócsai A, Ruland J, Tybulewicz VLJ. The SYK tyrosine kinase: a crucial player in diverse biological functions. Nat Rev Immunol. 2010;10(6):387-402.
103. Turner M, Mee PJ, Costello PS, et al. Perinatal lethality and blocked B-cell development in mice lacking the tyrosine kinase Syk. Nature. 1995;378(6554):298-302.
104. Cheng AM, Rowley B, Pao W, Hayday A, Bolen JB, Pawson T. Syk tyrosine kinase required for mouse viability and B-cell development. Nature. 1995;378(6554):303-306.
105. Young RM, Hardy IR, Clarke RL, et al. Mouse models of non-Hodgkin lymphoma reveal Syk as an important therapeutic target. Blood. 2009;113(11):2508-2516.
106. Friedberg JW, Sharman J, Sweetenham J, et al. Inhibition of Syk with fostamatinib disodium has significant clinical activity in non-Hodgkin lymphoma and chronic lymphocytic leukemia. Blood. 2010;115(13):2578-2585.
107. Quiroga MP, Balakrishnan K, Kurtova AV, et al. B-cell antigen receptor signaling enhances chronic lymphocytic leukemia cell migration and survival: specific targeting with a novel spleen tyrosine kinase inhibitor, R406. Blood. 2009;114(5):1029-1037.
108. Weinblatt ME, Kavanaugh A, Genovese MC, Musser TK, Grossbard EB, Magilavy DB. An oral spleen tyrosine kinase (Syk) inhibitor for rheumatoid arthritis. N Engl J Med. 2010;363(14):1303-1312.
109. Hoellenriegel J, Coffey GP, Sinha U, et al. Selective, novel spleen tyrosine kinase (Syk) inhibitors suppress chronic lymphocytic leukemia B-cell activation and migration. Leukemia. 2012;26(7):1576-1583.
110. Sharman J, Hawkins M, Kolibaba K, et al. An open-label phase 2 trial of entospletinib (GS-9973), a selective spleen tyrosine kinase inhibitor, in chronic lymphocytic leukemia. Blood. 2015;125(15):2336-2343.
111. Coffey G, Betz A, DeGuzman F, et al. The novel kinase inhibitor PRT062070 (Cerdulatinib) demonstrates efficacy in models of autoimmunity and B-cell cancer. J Pharmacol Exp Ther. 2014;351(3):538-548.
112. Younes A, Thieblemont C, Morschhauser F, et al. Combination of ibrutinib with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) for treatment-naive patients with CD20-positive B-cell non-Hodgkin lymphoma: a non-randomised, phase 1b study. Lancet Oncol. 2014;15(9):1019-1026.
113. Maddocks K, Christian B, Jaglowski S, et al. A phase 1/1b study of rituximab, bendamustine, and ibrutinib in patients with untreated and relapsed/refractory non-Hodgkin lymphoma. Blood. 2015;125(2):242-248.
114. Brown JR, Barrientos JC, Barr PM, et al. The Bruton tyrosine kinase inhibitor ibrutinib with chemoimmunotherapy in patients with chronic lymphocytic leukemia. Blood. 2015;125(19):2915-2922.
115. Flinn IW, Kahl BS, Leonard JP, et al. Idelalisib, a selective inhibitor of phosphatidylinositol 3-kinase-δ, as therapy for previously treated indolent non-Hodgkin lymphoma. Blood. 2014;123(22):3406-3413.
116. Brown JR, Byrd JC, Coutre SE, et al. Idelalisib, an inhibitor of phosphatidylinositol 3-kinase p110δ, for relapsed/refractory chronic lymphocytic leukemia. Blood. 2014;123(22):3390-3397.
117. Kahl BS, Spurgeon SE, Furman RR, et al. A phase 1 study of the PI3Kδ inhibitor idelalisib in patients with relapsed/refractory mantle cell lymphoma (MCL). Blood. 2014;123(22):3398-3405.
118. Burger JA, Tedeschi A, Barr PM, et al; RESONATE-2 Investigators. Ibrutinib as initial therapy for patients with chronic lymphocytic leukemia [published online December 6, 2015]. N Engl J Med. doi:10.1056/NEJMoa1509388.