B lymphocytes can directly participate in humoral immune responses as immune effector cells, and can also be used as antigen-presenting cells to capture antigens and present them to T lymphocytes. B cells mediate foreign antigen signals and complex biological effects, including B cell activation, proliferation, and differentiation, through their surface B cell receptors (BCR). This process involves a series of downstream signal transduction pathways that interweave to form a very complex B cell receptor signal transduction regulatory network. Many non-Hodgkin's lymphomas (NHL) originate from B lymphocytes. B cell receptor (BCR) complexes and their related protein kinases play important roles in the development, proliferation and survival of normal and malignant B cells. After the concept of antigen stimulation to induce lymphoma production, the BCR receptor signaling pathway has become an important pathway for lymphoma growth and survival. Therefore, various kinases in BCR and its signaling pathways can be new targets for lymphoma therapy.
BCR structure and function BCR is a heterologous oligomeric complex composed of membrane immunoglobulin (mIg) and Igα (CD79A)/Igβ (CD79B), which is one of the characteristic markers of B cells. mIg recognizes and binds antigens. The intracellular regions of Igα and Igβ have an immunoreceptor tyrosine-based activation motif (ITAM), which is involved in the transduction of antigenic stimulation signals after phosphorylation.
BCR signal transduction pathway
PLC-γ2 mediated signaling pathway BCR/Igα/Igβ complex enters the lipid raft on the B cell surface after BCR is stimulated by antigen. Activation of Lyn (SRC family kinases) phosphorylates ITAM in the cytoplasmic region of Igα/Igβ. Then Syk (spleen tyrosine kinase) is recruited to the lipid raft and then be phosphorylated. Activated Syk can phosphorylate B cell linker protein (BLNK) and CIN85 (Cbl-interacting protein of 85 ku), and then bind to phospholipase C-γ2 (phospholipase C-γ2, PLC-γ2) with Btk (Bruton tyrosine kinase). Activated PLC-γ2 hydrolyzes the membrane substrate PIP2 to generate a second messenger IP3 and DAG. Binding of IP3 to the IP3 receptor on the endoplasmic reticulum membrane opens the membrane Ca2++ channel and intracellular calcium stores, resulting in an increase in intracytoplasmic Ca2++ concentration. The calcineurin is then activated to dephosphorylate the transcription factor NFAT (phosphorylated factor NFAT) to activate and mediate the NFAT pathway. DAG can bind protein kinase Cβ (PKCβ) to the inner side of the cell membrane. Under the synergistic action of phosphatidylserine, PKCβ can be activated by DAG to activate Bcl-10/CARMA1/MALT1 complex, making IKK complex phosphorylated. Subsequently, the IκB serine residue is phosphorylated by the IKK complex and then degraded by ubiquitin labeling, releasing NF-κB from IκB. NF-κB translocation into the nuclear activation of the NF-κB pathway. DAG also activates the downstream MAPK pathway by activating Ras, which leads to the activation of the transcription factor AP1, and regulates gene expression.
Figure 1. The BCR signaling pathway
PI3K-mediated signaling pathway CD19 is a specific cell surface molecule expressed by B cells from pre-B cells to plasma cells. After BCR activation, CD19 and PI3K B cell adaptor molecule for PI3K (BCAP) can be activated by phosphorylation of Lyn, Syk, and Btk to activate PI3K. PI3K uses PIP2 as a substrate to generate PIP3, which allows Akt and Btk to be recruited to the membrane and activated. Activated Akt phosphorylates the IKK complex and ultimately activates signaling pathways such as NF-κB, MAPK, NFAT and RAS.
BCR mediates chronic lymphocytic leukemia (CLL). The BCR signaling pathway largely mediates the development and progression of chronic lymphocytic leukemia (CLL). Some patients have high expression of BCR-associated kinases such as ERK, Akt, and NFAT. In vitro experiments have shown that IgM on the surface of IgM monoclonal antibody cross-linked B cell membrane can cause intracellular transfer of calcium ions and activation of downstream signaling molecules, leading to anti-apoptotic reaction, and Bcl-2 family protein, Mcl-1 (the myeloid leukemia cell differentiation protein), and other pro-survival factors activated. The ability of BCR to respond to growth or survival factors depends on the level of expression of immunoglobulin on the membrane surface. Somatic mutations in the expression of immunoglobulin heavy chain variable region genes, somatic mutations in ZAP-70 (ζ-associated protein of 70 Ku), and intracellular tyrosine kinases can activate and recruit Syk. Therefore, the invasiveness of patients with positive ZAP expression also indirectly indicates the relevance of the BCR signaling pathway. This is presumed to be because the BCR signal has been fully activated in these malignant cells by antigen or autonomic signals. Another interesting explanation is that the BCR signal transduction tendency in CLL can be directly affected by microRNAs, thus affecting disease invasiveness.
BCR mediates diffuse large B-cell lymphoma
The chronic activating BCR signaling pathway is involved in the pathogenesis of activated B cell like diffuse large B cell lymphoma (ABC DLBCL). ABC DLBCL tumor cells rely on the anti-apoptotic NF-κB pathway for survival. Gene expression profiling showed that the expression of the NF-κB target gene in ABC DLBCL was significantly higher than that of germinal center type, and the NF-κB pathway has become a potential therapeutic target. The CARD11 mutation showed that the NF-κB pathway involved in its disease. Mutations in tumor somatic cells affect the coiled-curled structure of CADR11, thereby spontaneously recruiting signal molecules downstream of the NF-κB pathway. RNA interference showed that the CBM (CARD11-BCL-10- MALT1) complex played a part in the activation of the ABC DLBCL NF-κB pathway. In normal B cells, the CBM complex binds BCR and IKK and is a key regulator of the NF-κB pathway. Approximately 10% of the subtype mutations in CARD11 in ABC DLBCL can cause activation of the CBM complex. RNA interference also showed that Btk is a key factor in NF-κB pathway activation and cell survival. Approximately 20% of ABC DLBCL develops CD79A and CD79B mutations in the ITAM motif. Knockout of BCR components (IgH, Igκ, CD79A or CD79B) or BCR signaling pathway effectors (Syk, BLNK, PLCγ2, PI3Kδ or PKCβ) can kill ABC DLBCL tumor cells. Overexpression of Bcl6 reduces the inhibitory tyrosine phosphatase and increases Syk activity to amplify the BCR signaling pathway.
Inhibitor against BCR signaling pathway
The BCR signaling pathway mediates the development and progression of malignant lymphoma, suggesting that drug inhibitors can be used to block the further development of the disease. At present, a variety of inhibitors have been developed to exert targeted therapeutic effects by inhibiting BCR signaling pathway enzymes. It is important to emphasize that these small molecules are usually capable of inhibiting multiple enzymes, and even inhibitors against a specific enzyme cannot underestimate the potential off-target effects of the drug. In fact, each inhibitor has an off-target effect, so pay attention to drug toxicity and the biological activity and clinical effects of the drug is of great importance.
Figure 2. BCR signaling inhibitors
Btk is a member of the Tec family of non-receptor kinases. Btk's passivation mutations can slow down the development of B cells and severely reduce antibody secretion, leading to X-chain gamma globulin deficiency. Ibrutinib (PCI-32765) is a highly selective and potent Btk inhibitor that can covalently modify enzymes. It is also an irreversible inhibitor. Ibrutinib reduces the initial cellular DNA synthesis and viability in CLL patients. In a mouse model of CLL-like cells transfected with Eμ-TCL1, Ibrutinib was found to prevent progression of the disease. Ibrutinib reduces the expression of chemokines CLL3 and CLL4 in plasma from early CLL patients. Ibrutinib inhibits Btk and inhibits the binding of chemokines in the lymph node microenvironment to CXC chemokine receptor type 4 (CXCR4) and CXCR5, thereby blocking the homing of CLL cells. Ibrutinib or IDelaliSb alone has a strong monotherapy activity, but it does not cure patients. Related studies in patients with CLL have shown that, similar to Irtreuib, IDelaliSb impairs BCR signaling, cellular interactions, and chemokine production.
It is unclear whether clinically used BCR inhibitors completely block BCR signaling in vivo. B cells from patients treated with ibrutinib can still be activated by BCR and TLR stimulation, although the response is weaker than the stimulation of CD40L by BTK130. Treatment with ibrutinib also causes upregulation of AKT, which may compensate for inhibition of BCR signaling and support cell survival. The results show that in mice, complete depletion of BCR signaling in mature B cells results in relatively rapid apoptosis, whereas the use of the BCR inhibitor ibrutinib in CLL patients allows for high B cells for several months.
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