Gene expression profiling has revealed that diffuse large B-cell lymphoma (DLBCL) is composed of at least 3 different sub-groups, each having distinct oncogenic mechanisms that respond to therapies in different ways. Germinal Center B-Cell like (GCB) DLBCLs appear to arise from normal germinal center B cells, while Activated B-cell like (ABC) DLBCLs are thought to arise from postgerminal center B cells that are arrested during plasmacytic differentiation. The differences in gene expression between GCB DLBCL and ABC DLBCL are as vast as the differences between distinct types of leukemia, but these conditions have historically been grouped together and treated as the same disease.
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Distinguishing features
A gene translocation between chromosome 14 (containing the antibody heavy chain locus) and chromosome 18 (containing the BCL-2 locus) is present in 45% of GCB DLBCLs but has never been found in ABC DLBCLs. This T(14,18) translocation places the BCL-2 gene close to the heavy chain gene enhancer and results in the overexpression of the Bcl-2 protein. Bcl-2 proteins prevent the activation of the caspases that lead to programmed cell death (apoptosis).
Activation of the nuclear factor kappa-light-chain-enhancer of activated B-cells (NF-κB) pathway is found only in ABC DLBCLs and not GCB DLBCLs.
GCB DLBCL shows amplification of the oncogenic mir-17–92 microRNA cluster and deletion of the tumor suppressor PTEN but these events have not been found in ABC DLBCL
Normal B-cell maturation process
B-cells form in the bone marrow and undergo gene rearrangement in order to develop B-cell receptors (BCRs) that bind to a specific antigen. Once activated by an antigen, B-cells proliferate and further differentiate into plasma cells and memory B-cells. B-cells that have not encountered an antigen are called naive B cells. When naïve B-cells encounter an antigen, one of the pathways that they can follow is through the germinal center environment. B-cells within the germinal center proliferate and undergo immunoglobulin somatic hypermutation (SHM) of IgV region genes to revise their antigen receptors. The rearranging of genes makes the cells capable of generating antibodies with a higher or lower affinity to the specified antigen. Follicular dendritic cells and T cells help to select the B-cells that have a high affinity to the antigen for further differentiation into plasma cells and memory cells. A large fraction of germinal center B-cells acquire somatic mutations that prohibit antigen binding and these undergo apoptosis.
Mechanisms of malignant transformation
Two oncogenic mechanisms that appear to be active in GCB DLBCL are the prevention of apoptosis and the blocking of terminal differentiation.
Preventing apoptosis
Normal germinal center B-cells appear to be poised for apoptosis unless they are selected to progress to the next stage of differentiation. Most normal germinal center B-cells express low levels of anti-apoptotic proteins such as Bcl-2. In GBC DLBCLs, the T(14,18) translocation can result in an increase of the Bcl-2 protein, which may reduce the number of cells that undergo apoptosis.
Blocking differentiation
Blocking the differentiation of germinal center B cells is dangerous because the cells are programmed to divide rapidly at this stage. The SHM that occurs in the germinal center can also target non-immunoglobulin loci and may be responsible for translocation of the BCL-6 gene. BCL-6 genes are involved in several cell processes that can affect the ability of the B-cell to differentiate and proliferate. BCL-6 genes produce BCL-6 proteins. These proteins work with other transcription factors (BLIMP1, PAX5, XBP1) to form a regulatory circuit that controls the progression of germinal center B cells to plasma cells. BCL-6 proteins repress genes involved in terminal differentiation and promote proliferation by blocking expression of a cell-cycle inhibitor (p27KIP1). BCL-6 is also an inhibitor of cellular senescence. Cellular senescence is a programmed response that prevents a cell from dividing after some number of cell divisions. The expression of key transcription factors in B cell development is frequently regulated by non-coding RNAs termed microRNAs. MicroRNAs (miRNAs) participate in pathways fundamental to B cell development like B cell receptor (BCR) signalling, B cell migration/adhesion, cell-cell interactions in immune niches, and the production and class-switching of immunoglobulins.
Treatment
DLBCL patients are at higher risk when they relapse early after R-CHOP chemotherapy and have a poor response to second-line rituximab-containing treatments even when these regimens involve high-dose therapy and autologous stem cell transplant. Approximately half of DLBCL patients develop CHOP-resistant cells. A study of DLBCL cell lines indicated that 14-3-3ζ proteins may play a role in mediating resistance of DLBCL cells to CHOP. 14-3-3 proteins exert anti-apoptotic activity by interfering with the function of BH3-only proteins and has been validated as a potential molecular target for anticancer therapeutic development in other types of cancers.
Monoclonal antibodies
Monoclonal antibodies are made by injecting human cancer cells into mice so that their immune systems create antibodies against foreign antigens. Monoclonal antibodies target specific antigens on cancer cells and may enhance the patient's immune response. They can be administered alone or be linked (conjugated) to anticancer drugs, radioisotopes, or other biologic response modifiers. There are several therapeutic mechanisms for monoclonal antibodies:
- Directly initiates apoptosis in the targeted cells
- Antibody-dependent cell-mediated cytotoxicity (ADCC) -- Recruits monocytes, macrophages, and natural killer cells to destroy the targeted cells
- Complement-dependent cytotoxicity (CDC)-- Initiates the complement system which activates the membrane attack complex causing cell lysis and death.
- Delivers chemotherapy or radiation in a targeted manner which allows higher concentrations to be administered
Monoclonal antibodies for treatment of B-cell malignancies
Bcl-2 inhibitors
Apoptosis is one of the major mechanisms of cell death targeted by cancer therapies. Reduced susceptibility to apoptosis increases the resistance of cancer cells to radiation and cytotoxic agents. B-cell lymphoma-2 (Bcl-2) family members create a balance between pro and anti-apoptotic proteins. Pro-apoptotic proteins include Bax and Bak. Anti-apoptotic proteins include Bcl-2, Bcl-XL, Bcl-w, Mcl-1. When anti-apoptotic family members are overexpressed, apoptotic cell death becomes less likely.
mTOR (mammalian target of rapamycin) inhibitors
mTOR inhibitors :
mTOR is a kinase enzyme inside the cell that regulates cell growth, proliferation, and survival. mTOR inhibitors lead to cell cycle arrest in the G1 phase and also inhibits tumor angiogenesis by reducing synthesis of VEGF.
A Phase II trial of Evorolimus on relapsed DLBCL patients showed a 30% Overall Response Rate (ORR).
Syk (Spleen Tyrosine Kinase) inhibitors
Syk inhibitors include :
Chronic signaling through the B-cell receptor appears to contribute to the survival of DLBCL. These survival signals can be blocked by Syk inhibitors. However, since the BCR signaling pathway is not as important to the GCB DLBCL as it is to the ABC subtype, Syk inhibitors may not be effective against GCB DLBCL
Proteasome inhibitors
Proteasome inhibitors inhibit the NF-κB pathway. Since this pathway is not a significant factor in GCB DLBCL, proteasome inhibitors have not been found to be effective against GCB DLBCL. A clinical trial of bortezomib showed that bortezomib alone had no activity in DLBCL, but when combined with chemotherapy, it demonstrated an ORR of 83% in ABC DLBCL and 13% in GCB DLBCL, suggesting that bortezomib enhances the activity of chemotherapy for ABC but not GCB DLBCL when combined with conventional chemotherapy.