B lymphocyte stimulator: a new target for treating B cell malignancies (2008)

Blymphocyte stimulator (BLyS) is also known as B cell activating factor belonging to the tumor necrosis factor (TNF) family (BAFF), or TNF and apoptosis ligand-related leukocyte-expressed ligand 1 (TALL-1).1-3 As a member of the TNF family, BLyS plays an important role in B cell development, and has been implicated in autoimmune diseases and B cell malignancies.4-6In vitro experiments demonstrated that BLyS could bind B cells, stimulate their proliferation, and promote their survival.1,2,7 If the BLyS gene is deleted in mice, mature B cells are lost in peripheral lymphoid systems.8 On the other hand, over-expression of BLyS in transgenic mice results in a significant increase in peripheral B cells and leads to autoimmune disease-like symptoms.9 Recently, abundant evidence showed that BLyS could promote the survival of malignant B cells. In this review, we discussed the function of BLyS and its roles in B cell malignancies. FUNCTIONS OF BLYS BLyS is constitutively synthesized by myeloid cells like macrophages, monocytes and dendritic cells as a type II transmembrane protein consisting of 285 amino acids.1,2 After proteolytic cleavage of the peptide bound between R133 and A134, BLyS is released as a soluble form. Like other members of the TNF family, soluble BLyS can form a homotrimer as a functional unit. In addition, the production of BLyS can also be induced by type I and type II interferons, and IL-10, which may be involved in immune defenses against bacterial and viral infection.10,11 BLyS binds B cells and stimulates their proliferation and survival in vitro. Early B cell development from progenitor B cells to immature B cells occurs in bone marrow (BM). After leaving BM, immature B cells migrate to peripheral lymphoid tissues and develop through transitional stages 1 and 2 (T1 and T2). T2 B cells undergo one of two separate differentiation paths. The majority of the cells (80% of peripheral B cells) will become follicular (FO) B cells. The rest will mature to marginal zone (MZ) B cells.12,13 Deletion of the BLyS gene in mice results in loss of both FO and MZ B cells.8 Although the number of T1 B cells reduces slightly, there are almost no T2 cells in BLyS-/- mice. So BLyS is absolutely required for T1 to T2 cell transition. On the other hand, over-expression of BLyS leads to severe B cell hyperplasia and eventually results in autoimmune-like symptoms in BLyS transgenic mice.9,14 Thus BLyS is a critical factor that regulates B cell homeostasis in vivo. SIGNAL PATHWAYS AND RECEPTORS OF BLYS BLyS can bind to three receptors: B cell maturation antigen (BCMA), transmembrane activator and CAML-interactor (TACI) and BLyS receptor 3 (BR3).15,16 BCMA and BR3 are expressed on B cells, while TACI is expressed both on B cells and activated T lymphocytes. Upon binding of BLyS, BCMA and BR3 interact with TNF receptor associated factors (TRAFs), activate nuclear factor-κB (NF-κB) signal pathways, and upregulate the expression of anti-apoptosis genes including Bcl-2 and Bcl-XL.17-20 BR3 activates the alternative NF-κB pathway (NF-κB2),21,22 while BCMA activates the classical NF-κB pathway (NF-κB1).17,23 In addition, BLyS can block the nuclear translocation of protein kinase C-δ (PKCδ) in a c-myb dependent manner and protect B cells from apoptosis.24,25 If c-myb gene is deleted, BLyS no longer blocks the nuclear translocation of PKCδ and the cells are more susceptible to apoptosis.25 All these results demonstrate that BLyS provides survival signals for B cells (Figure).Figure.: Receptors and signal pathways of BLyS and APRIL. Upon binding of BLyS and APRIL, BCMA and TACI interact with TNF receptor associated factors (TRAFs), activate the classical NF-κB pathway (NF-κB1), and upregulate the expression of anti-apoptosis genes including Bcl-2 and Bcl-XL. BLyS can also interact with BR3 and activates the alternative NF-κB pathway (NF-κB2). In addition, BLyS can block the nuclear translocation of PKCδ in a c-myb dependent manner and protect B cells from apoptosis.Gene knockout experiments showed that mice without the BCMA gene had normal B cell development.8 However, there was a significant reduction in mature B cells in BR3-mutated A/WySnJ mice, similar to that found in BLyS-/- mice, suggesting that BR3 was the major signal transducer of BLyS activity.16 Further studies have demonstrated that BR3 is not involved in terminal differentiation of plasma cells (PCs). BCMA plays an indispensable role in the survival of PCs.26 In agreement with this phenomenon, there are differential expression patterns of BLyS receptors. BR3 is expressed by T2, immature, FO and MZ B cells, but is absent in PCs.27,28 BCMA is a predominant receptor of BLyS in PCs.29 It is not yet known how BCMA functions in PC differentiation. In addition, BCMA and TACI can also bind to a proliferation inducing ligand (APRIL).30,31 APRIL is also a member of the TNF family and shares 48% homology with BLyS.2 APRIL is expressed at low level in normal tissues but is highly expressed in human tumor tissues such as colon, thyroid and lymphoid cancers.10 It is not yet fully understood what role APRIL plays in B cell development. However, it is believed that APRIL functions at least partially by binding to BCMA and TACI and activating the classical NF-κB pathway. BLYS AND B CELL MALIGNANCIES B cell malignancies include non-Hodgkin's lymphoma (NHL), Hodgkin's lymphoma (HL) and multiple myeloma (MM). The tumor cells are transformed B cells that are clonally expanded from different differential stages. The mechanisms of B cell malignancies are complex.32-34 Gene translocations and over-expression of proto-oncogenes such as c-myc and ras are frequently observed in these tumors. In addition, excessive survival factors such as IL-6 in MM were also found to promote tumor growth. Recent studies demonstrated that both BLyS and APRIL protected malignant B cells from apoptosis and promoted their survival.35-39 The serum level of BLyS in NHL and MM is elevated, suggesting that excessive production of BLyS occurs in these patients.36,39,40 In MM, BM stromal cells and osteoclast cells can secrete BLyS and provide favorable microenvironments for the survival of tumor cells.41,42 More significantly, MM and B lymphoma tumor cells can express both BLyS and APRIL, which function locally on tumor cells.35,36,38,39 In HL, Hodgkin and Reed-Sternberg (HRS) cells also express membrane-bound BLyS. Inhibition of endogenous BLyS expression by interfering RNA (RNAi) induces apoptosis of HRS cells.43 Thus it is likely that BLyS synthesized by malignant B cells forms an autocrine loop to attenuate apoptosis of tumor cells and plays an important role in the development and progression of tumors. We analyzed expression of BLyS on two Burkitt's lymphoma cell lines (Daudi and Raji) and two MM cell lines (SKO-007 and XG-7) and found that BLyS was expressed on SKO-007 and XG-7 cells, but not on Daudi and Raji cells.44 Briones et al45 also reported that there was no cell-surface expression of BLyS by malignant B cells from 6 NHL patients, including follicular lymphoma, diffuse large-cell lymphoma, and mantle-cell lymphoma, whereas others studies indicated that NHL B cells did express membrane-bound BLyS.35,37,40 To understand the expression of BLyS in different types of NHL, more NHL patients need to be studied. In vitro studies demonstrated that BLyS activated the signal pathways of NF-κB and mitogen activated protein kinase (MAPK), and upregulated the expression of the anti-apoptosis genes Bcl-2 and Mcl-1 in MM cell lines as well as NHL.35,39 Exogenous BLyS also activates NF-κB and upregulates the expression of the anti-apoptosis genes bcl-2, bcl-XL, and c-myc in cultured HRS cells. However, the receptors that mediated BLyS function in malignant B cells are not fully understood. In normal B cells, activation of the alternative NF-κB pathway by BLyS through BR3 is required for the survival of B cells. But in B chronic lymphocytic leukemia (CLL), activation of the alternative NF-κB pathway is not required for survival of CLL cells. On the contrary, activation of the canonical NF-κB pathway through BCMA and TACI is required for protecting CLL cells from apoptosis.46 If expression of TACI and BCMA in HRS cells is silenced by RNAi or BLyS activity is blocked by a BLyS decoy receptor, TACI-Fc, these cells undergo apoptosis.43 The expression patterns of BLyS receptors in B malignancies are different from those of normal B cells. BCMA is highly expressed in MM cells.37,41 BR3 and TACI are expressed to various levels in NHL. In HL, HRS cells express BCMA and TACI, but not BR3.43 These results indicate that transmission of the BLyS signal through its receptors in normal and malignant B cells may be different, resulting in distinct responses to BLyS. In addition, low level expression of TACI on malignant B cells and high level expression of APRIL in B cell malignancies are associated with poor prognosis.47 BLYS-TARGETING THERAPY The incidence rate of B cell malignances has gradually increased in recent years. However, therapies for these diseases are not always effective and satisfactory. For example, traditional chemotherapy for MM results in a median survival duration of no more than three years, a 25% five-year survival rate, and a ten-year survival rate of less than 5%.48 After recent evidence convincingly demonstrated that BLyS and its receptors were associated with B cell malignancies, BLyS-targeting therapy is an attractive therapeutic approach for MM and lymphoma treatment. The inhibitors of BLyS may take the form of decoyed receptors or monoclonal antibodies. The decoyed receptor, TACI-Fc, has been tested for treating autoimmune diseases such as systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) in animals and has generated optimistic results.49-51 A clinical trial using a monoclonal antibody, lymphostat-B (Human Genome Sciences, USA) against human BLyS protein is underway for treating SLE. As a B cell binding cytokine, BLyS conjugated with radioisotope or toxin may deliver therapeutic reagents to malignant B cells and kill the tumor cells. For example, 125I labeled BLyS has been tested to treat B cell tumors.52 BLyS fused to toxin Gelonin can bind CLL cells and induce apoptosis of the cells in vitro.53 Several companies are currently conducting clinical studies to evaluate the safety and efficacy of BLyS inhibitors in treatment of B cell malignancies. A news release from ZymoGenetics showed that TACI-Ig, Atacicept, was more effective than BR3-Ig in inhibiting MM growth in animals. In a phase 1b clinical trial, refractory or relapsed MM responded well to Atacicept. Fifteen patients with relapsed or refractory NHL have shown decreasing serum Ig concentration after 5 weeks treatment with Atacicept, but tumors responded poorly to Atacicept (http://www.zymogenetics.com/ir/newsItem.php?id=941352). 131I labeled BLyS, LymphoRad131 developed by Human Genomic Sciences Inc. is also being tested in a phase 1 clinical trial to treat MM and NHL. In addition to decoyed receptors and monoclonal antibodies as described above, small molecules such as peptides may be an alternative approach to inhibiting BLyS activity. Peptides used as antagonists to block the biological activity of cytokines have been investigated intensively in vitro.54,55 We previously designed a BLyS antagonist peptide based on the complex structure of BCMA and BLyS.56 The peptide could inhibit the binding of BCMA with BLyS and partially block the biological activity of BLyS on mouse splenocytes. With the crystal structures of BLyS and its receptors available, more potent BLyS inhibitors can be designed by theoretically analyzing the interaction of BLyS with its receptors, and these inhibitors can be evaluated by biological experiments.