Non-Stereotypic Follicular Lymphoma B-Cell Receptors Recognize Vimentin as a Shared Autoantigen

Abstract 139

The chromosomal translocation t(14;18) is present in most human follicular lymphomas (FL), however, it is not sufficient for lymphomagenesis. A number of studies have suggested a potential role for antigen activation of the B-cell receptor (BCR) in the pathogenesis of FL and other B-cell malignancies. However, the nature of the antigen(s) recognized by FL BCRs has not been well studied. We hypothesized that the antigen(s) recognized by FL BCRs is either a self or non-self antigen. The objective of this study was to identify self antigen(s) recognized by FL BCRs. We generated clonal tumor immunoglobulins (Igs) from 217 patients with FL using either hybridoma or recombinant DNA technologies. Purified tumor Igs were tested for autoreactivity against whole cell lysate prepared from a human epithelial cell line, HEp-2, in an enzyme-linked immunosorbent assay (ELISA). We observed that FL tumor Igs were frequently self-reactive. To determine the nature of the antigen recognized by the tumor Igs, we performed polyacrylamide gel electrophoresis of HEp-2 cell lysate. The single band recognized by the tumor Igs was analyzed by mass spectrometry and was determined to be vimentin. Using recombinant vimentin protein, we demonstrated that the tumor Igs specifically bound to vimentin when tested by ELISA and Western Blot. Furthermore, pre-incubation of the tumor Igs with recombinant vimentin protein significantly inhibited their reactivity against HEp-2 cell lysate in a dose-dependent manner. Using indirect immunofluorescence staining, we showed that the vimentin-reactive tumor Igs co-localized with a mouse anti-vimentin mAb in HEp-2 cells. Using a series of truncated recombinant vimentin protein fragments, we determined that the epitope recognized by the tumor Igs is localized to the N-terminal region of vimentin. Overall, 42/217 (19.35%) FL tumor Igs recognized N-terminal region of vimentin. Vimentin-reactivity was more commonly observed with IgG than IgM FL tumor Igs (30.4% vs. 10%; P=0.0022). However, comprehensive analysis of the sequences of the variable regions of heavy and light chains of the FL tumor Igs showed that vimentin-reactivity did not correlate with VH, JH, Vκ, Jκ, Vλ, and Jλ gene usage; CDR3 length; number of positively charged amino acid residues in CDR3; rate and pattern of mutations; or rate and pattern of N-glycosylation. In addition, we did not observe recurrent IgV_H- or IgV_L-CDR3 motifs amongst the vimentin-reactive FL tumor Igs. By immunohistochemistry, we showed that vimentin was expressed in the T-cell rich regions of FL suggesting its availability for binding with BCR within the tumor microenvironment. Interestingly, vimentin was also recognized by 9/18 (50%) mantle cell lymphoma (MCL) tumor Igs and 12/31 (38.7%) multiple myeloma (MM) tumor Igs. In conclusion, our results demonstrate that vimentin is a shared autoantigen recognized by non-stereotypic FL BCRs as well as tumor Igs from MCL and MM and suggest that it may play a role in the pathogenesis of multiple B-cell malignancies. The identification of a shared autoantigen recognized by tumor BCRs could lead to development of in vitro and in vivo models to better understand the pathogenesis and natural history of FL and other B-cell malignancies.