Theodora A. M. Wormhoudt

Objective: Patients with Sjӧgren's syndrome (SS) have an increased risk of developing malignant B cell lymphomas, particularly mucosa‐associated lymphoid tissue (MALT)–type lymphomas. We have previously shown that a predominant proportion of patients with SS‐associated salivary gland MALT lymphoma express somatically hypermutated IgM with strong amino acid sequence homology with stereotypic rheumatoid factors (RFs). The present study was undertaken in a larger cohort of patients with SS‐associated MALT lymphoma to more firmly assess the frequency of RF reactivity and the significance of somatic IGV‐region mutations for RF reactivity.Methods: B cell antigen receptors (BCRs) of 16 patients with SS‐associated salivary gland MALT lymphoma were analyzed. Soluble recombinant IgM was produced of 12 MALT lymphoma samples, including 1 MALT lymphoma sample that expressed an IgM antibody fitting in a novel IGHV3‐30–encoded stereotypic IGHV subset. For 4 of the 12 IgM antibodies from MALT lymphoma samples, the somatically mutated IGHV and IGKV gene sequences were reverted to germline configurations. Their RF activity and binding affinity were determined by enzyme‐linked immunosorbent assay and surface plasmon resonance, respectively.Results: Nine (75%) of the 12 IgM antibodies identified in patients with SS‐associated salivary gland MALT lymphoma displayed strong monoreactive RF activity. Reversion of the IGHV and IGKV mutations to germline configuration resulted in RF affinities for IgG that were significantly lower for 3 of the 4 somatically mutated IgM antibodies. In stereotypic IGHV3‐7/IGKV3‐15–encoded RFs, a recurrent replacement mutation in the IGKV3‐15–third complementarity‐determining region was found to play a pivotal role in the affinity for IgG‐Fc.Conclusion: A majority of patients with SS‐associated salivary gland MALT lymphoma express somatically mutated BCRs that are selected for monoreactive, high‐affinity binding of IgG‐Fc. These data underscore the notion that soluble IgG, most likely in immune complexes in inflamed tissues, is the principal autoantigen in the pathogenesis of a variety of B cell lymphomas, particularly SS‐associated MALT lymphomas.

In the past years, the genomes of thousands of tumors have been elucidated. To date however, our knowledge on somatic gene alterations in normal cells is very limited. In this study, we demonstrate that tetanus-specific human memory B lymphocytes carry a substantial number of somatic mutations in the coding regions of the genome. Interestingly, we observed a statistically significant correlation between the number of exome mutations and those present in the immunoglobulin heavy variable regions. Our findings indicate that the majority of these genomic mutations arise in an antigen-dependent fashion, most likely during clonal expansion in germinal centers. The knowledge that normal B cells accumulate genomic alterations outside the immunoglobulin loci during development is relevant for our understanding of the process of lymphomagenesis.

Subsets of mucosa-associated lymphoid tissue (MALT) lymphoma, splenic marginal zone B-cell lymphoma (MZBCL) and chronic lymphocytic leukemia (CLL) have been identified that express near-identical B-cell receptors (BCRs), strongly suggesting selection by restricted antigenic epitopes. We here report a new IGV mutated stereotypic IGHV4-59/IGHJ5-encoded BCR subset, expressed in hepatitis C virus (HCV)-related B-cell lymphoma, MALT lymphoma and diffuse large B-cell lymphoma (DLBCL). We demonstrate that these mutated stereotypic IGHV4-59/IGHJ5 BCRs are high affinity monoreactive rheumatoid factors (RFs) underscoring the significance of IgG as a major auto-antigen in the pathogenesis of several types of B-cell lymphoma.

Objective: Among autoimmune diseases, Sjögren's syndrome (SS) is most strongly associated with the development of malignant B cell lymphoma, in particular mucosa‐associated lymphoid tissue (MALT)–type lymphoma. Previously, we have shown that in ∼40% of cases of salivary gland MALT lymphoma, high‐affinity stereotypic rheumatoid factor (RF) B cell receptors, specific for IgG‐Fc, are expressed. This study was undertaken to investigate whether in the inflamed salivary glands of patients with SS, a similar RF‐biased Ig repertoire is present.Methods: Extensive analyses of the B cell Ig V_H region repertoire were performed on microdissected tissue samples from the labial salivary glands of 4 patients with SS.Results: All SS labial salivary glands harbored expanded B cell clones, of which 1 or 2 were highly expanded and detected in >50% of the microdissected samples. However, among the identified 464 distinct Ig clonotypes, only 3 stereotypic RF–expressing clones were detected. In 2 patients with SS, an RF‐expressing clone was detected at low frequency in 1 of the microdissected samples, whereas 1 patient with SS harbored a highly expanded RF‐expressing clone that was detected in all microdissected samples and also detected in the peripheral blood. Two years after analysis of this sample, the latter patient developed a diffuse large B cell lymphoma originating from the same RF clone.Conclusion: Inflamed labial salivary glands in patients with SS generally harbor 1 or 2 highly expanded B cell clones. The repertoire strongly biased toward stereotypic RFs in salivary gland MALT lymphomas is not a reflection of a similar repertoire in the inflamed salivary glands of patients with SS; rather, in the latter, the repertoire is based on a strong selection advantage of incidental stereotypic RF–expressing B cells.

B cell chronic lymphocytic leukemia (CLL), the most common leukemia in adults, is a clonal expansion of CD5⁺CD19⁺ B lymphocytes. Two types of CLLs are being distinguished as carrying either unmutated or somatically mutated immunoglobulins (Igs), which are associated with unfavorable and favorable prognoses, respectively. More than 30% of CLLs can be grouped based on their expression of stereotypic B cell receptors (BCRs), strongly suggesting that distinctive antigens are involved in the development of CLL. Unmutated CLLs, carrying Ig heavy chain variable (IGHV) genes in germline configuration, express low-affinity, poly-, and self-reactive BCRs. However, the antigenic specificity of CLLs with mutated IGHV-genes (M-CLL) remained elusive. In this study, we describe a new subset of M-CLL, expressing stereotypic BCRs highly specific for β-(1,6)-glucan, a major antigenic determinant of yeasts and filamentous fungi. β-(1,6)-glucan binding depended on both the stereotypic Ig heavy and light chains, as well as on a distinct amino acid in the IGHV-CDR3. Reversion of IGHV mutations to germline configuration reduced the affinity for β-(1,6)-glucan, indicating that these BCRs are indeed affinity-selected for their cognate antigen. Moreover, CLL cells expressing these stereotypic receptors proliferate in response to β-(1,6)-glucan. This study establishes a class of common pathogens as functional ligands for a subset of somatically mutated human B cell lymphomas.

Ocular adnexal marginal zone B-cell lymphomas (OAMZLs) arise in the connective tissues of the orbit or in the mucosa-associated lymphoid tissue of the conjunctiva. Here, we present the immunological and genetic analyses of 20 primary Chlamydia psittaci (Cp)-negative OAMZLs. Analysis of the immunoglobulin variable heavy chain (IgV_H) gene usage demonstrated a significant preference for V_H4-34. A combined analysis across all previously published OAMZLs confirmed that this is a general feature of OAMZL, in particular of the Cp-negative group. Our series of OAMZLs did not express the characteristic rheumatoid factor V_HDJ_H rearrangements that were previously found in salivary gland- and gastric-marginal zone B-cell lymphomas (MZBCLs). We did not detect the MZBCL-specific chromosomal translocations, t(11;18) API2-MALT1 (mucosa-associated lymphoid tissue1) and t(14;18) IgH/MALT1. Two cases contained a premature stop codon in the A20 gene (TNFAIP3) and one case harbored the activating MYD88 hotspot mutation L265P. Variable nuclear expression of BCL10, NFκB1 (p50) and NFκB2 (p52) suggests that other additional genetic abnormalities affecting the NFκB pathway exist within this group of lymphomas. OAMZL showed variable expression of the chemokine receptor CXCR3 and integrin α4β7 by the tumor B cells, and low interferon-γ and interlukin-4 mRNA levels in the tissue, indicative of an inflammatory environment with features in between those previously found in cutaneous and other extranodal MZBCL. The strongly biased usage of V_H4-34 in Cp-negative OAMZLs suggests involvement of a particular stimulatory (auto-) antigen in their development.

Extranodal marginal zone B-cell lymphomas (MZBCLs) arise on a background of chronic inflammation resulting from organ-specific autoimmunity, infection, or by unknown causes. Well-known examples are salivary gland MZBCL in Sjögren9s sialadenitis and gastric MZBCL in Helicobacter pylori gastritis. MZBCLs express CXCR3, a receptor for interferon-γ–induced chemokines highly expressed in the chronic inflammatory environment. The immunoglobulin (Ig) variable heavy/light chain (IgV_H/IgV_L) gene repertoire of salivary gland and gastric MZBCL appears restricted and frequently encodes B-cell receptors with rheumatoid factor reactivity. Primary cutaneous marginal zone B-cell lymphomas (PCMZLs) are regarded as the skin-involving counterparts of extranodal MZBCLs. Although PCMZLs have been associated with Borrelia burgdorferi dermatitis, PCMZLs generally arise because of unknown causes. We studied an extensive panel of PCMZLs and show that PCMZLs do not conform to the general profile of extranodal MZBCL. Whereas most noncutaneous MZBCLs express IgM, PCMZLs in majority express IgG, IgA, and IgE and do not show an obvious immunoglobulin repertoire bias. Furthermore, the isotype-switched PCMZLs lack CXCR3 and seem to arise in a different inflammatory environment, compared with other extranodal MZBCLs.