Evidence for Non-Random Immunoglobulin V_H Gene and Light Chain Isotype Usage in Follicular Non-Hodgkin’s Lymphoma.

Background: Antibody diversity is generated by recombination of individual immunoglobulin (Ig) gene segments and subsequent somatic diversification driven by antigen recognition. In the repertoire of expressed B cell receptors (BCR) among normal peripheral B cells, variable heavy (V_H) gene segments are not equally represented. The ratio of kappa to lambda light chain usage is also skewed; the normal κ/λ is 1.5. Investigation of the BCR repertoire may provide clues to the genesis of B cell malignancies, as suggested in chronic lymphocytic leukemia (CLL) and mantle cell lymphoma (MCL). BCR V gene identification during the production of recombinant antibodies used in our ongoing PhII and PhIII FavId® (idiotype/KLH) immunotherapy studies has enabled us to analyze V gene usage from 475 B cell follicular lymphoma (FL) tissue samples. This study reports the results of V_H gene and κ/λ gene expression in this FL sample collection.

Methods: Ig heavy chain (HC) and light chain (LC) isotypes from B cell FL samples were identified by flow cytometry. V_H and V_L regions were sequenced from gene specific cDNA libraries prepared from these samples. V_H gene usage and κ/λ ratios were compared to frequencies determined for normal peripheral B cells isolated from six healthy volunteers as well as published reports for normal peripheral B cells and other B cell malignancies.

Results: Compared to V_H gene family usage determined for normal B cells, VH3 usage is higher (68% vs. 42%), VH1 usage is lower (7.8% vs. 22%) and VH4 usage is equivalent (22% vs. 26%) in our cohort of FL patients while VH2, 5, 6 and 7 are infrequently used in both populations. Usage of the VH3 genes within FL derived sequences also depends upon isotype, in that this gene family is preferentially associated with the IgM HC isotype relative to IgG (76% and 57% respectively). Additionally, the combined usage of the specific genes VH3-23 and VH3-48 in our patient collection accounts for over 29% of all VH genes - compared to 9% among normal B cells. These V_H gene usages also differ from reports of V_H gene expression among CLL and MCL patients. With respect to LC usage, VH3 isolates are associated with a normal κ/λ ratio of 1.6 while VH4 gene isolates are preferentially associated with λ light chains with a κ/λ ratio of 0.9. Finally, FL B cells expressing the IgM HC isotype preferentially co-express κ light chains (κ/λ ratio of 2.4) while IgG expressing cells preferentially utilize λ chains (κ/λ ratio of 0.6).

Conclusions: Non-random V gene and LC expression among patients with FL is noted. These distortions in Ig gene expression suggest that lymphomagenesis in FL may be associated with B cell stimulation by common antigens. A program to investigate the epitopes recognized by FL derived BCRs via binding of recombinant FL derived antibodies to protein arrays containing common auto-antigens is currently underway.