Salivary Gland Mucosa-Associated Lymphoid Tissue-Type Lymphoma From Sjögren's Syndrome Patients in the Majority Express Rheumatoid Factors Affinity-Selected for IgG

Abstract

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.

Figure 1

VH–third complementarity‐determining region (CDR3) amino acid sequence homology of salivary gland mucosa‐associated lymphoid tissue (MALT)–type lymphomas of patients M86, M88, M93‐IB, M94, and M95 with 4 stereotypic rheumatoid factors (RFs) and with a MALT lymphoma (MALT‐L) from a hepatitis C virus (HCV)–infected patient. VH‐CDR3 homology is calculated as a percentage of identical (red) and similar (blue) amino acids.

Figure 2

MALT lymphoma IgM antibodies characterized as high‐affinity monoreactive RFs. In enzyme‐linked immunosorbent assays (ELISAs), IgM derived from MALT lymphoma salivary gland samples M86, M91, M93‐I, M94, and M96 show specific binding to coated human IgG (A), with different binding affinities (B). Values on the y‐axis are the absorbance measured at 450 nm. Sample M8 is an IgM from a patient with Sjӧgren’s syndrome–associated MALT lymphoma who was analyzed previously and does not show RF activity. Sample CLL57 is an IGV‐gene–unmutated IgM derived from a patient with B cell chronic lymphocytic leukemia (CLL) and shows strong polyreactivity. Black boxes indicate positive ELISA results at all IgM concentrations tested; the gray‐shaded box indicates positive ELISA results at the 2 highest IgM concentrations tested. The IgMs were tested in at least 4 concentrations, i.e., 125, 250, 500, and 1,000 ng/ml. LPS = lipopolysaccharide (see Figure 1 for other definitions).

Figure 3

Alignment of amino acid sequences of IGHV1‐69–, IGHV3‐7–, and IGKV3‐15– encoded RFs of salivary gland MALT lymphomas. Boxes highlight shared somatic replacement mutations, i.e., S36T in the VH‐CDR1 of samples M91 and M9, I59L in the VH‐CDR2 of samples M91 and M11, and A65P in the VH‐CDR2 of samples M88, M9, M11, and M22 (A), Q58P in the VH‐CDR2 of samples M93IB and M6 (B), and Q106H in the VK‐CDR3 of samples M89, M93, M94, M96, M6, and M5 (C). Somatic replacement mutations in the VH‐CDR3 are underlined. FR1 = framework region 1 (see Figure 1 for other definitions).

Figure 4

Binding affinities of MALT lymphoma RFs and germline‐reverted variants for human IgG, as measured by surface plasmon resonance assay. A, IGHV and IGKV configurations for the native lymphoma RFs and germline‐reverted variants of lymphoma RFs, are shown, along with their binding constants (K _D) to indicate affinity for human IgG. B, Histograms display the 1/K _D values of the native and germline‐reverted variants of lymphoma RFs. Each letter to the left of the bars for each sample represents an individual recombinantly produced IgM antibody. NB = no binding; 106‐H = histidine mutation on position 106 of VK‐CDR3; 106‐Q = germline glutamine on position 106 of VK‐CDR3 (see Figure 1 for other definitions).

Figure 5

Alignment of the IGHV1‐69–encoded RFs of YES8c and M91. A, Somatic replacement mutations, as compared with the IGHV1‐69 and IGKV3‐20 germline sequences, are indicated. B, Alignment of the VH‐CDR3 amino acid sequences in YES8c, M91, and the homologous T580B, which originated from a patient with a Sjögren's syndrome–related MALT lymphoma, are indicated. Somatic replacement mutations are underlined. Gray‐shaded boxes highlight the amino acids of RFs in YES8c that are the contact residues with IgG‐Fc. FR1 = framework region 1 (see Figure 1 for other definitions).