Isotype switch variants reveal clonally related subpopulations in diffuse large B-cell lymphoma

Primary diffuse large B-cell lymphomas (DLBCLs) are aggressive tumors accounting for approximately 40% of B-cell malignancies. The immunoglobulin (Ig) variable region genes have undergone rearrangement and are commonly somatically mutated. The majority show intraclonal variation which indicates that somatic mutation has continued after transformation. Typically, cells of DLBCLs express Ig of a single isotype, but there may be accompanying cells that express alternative isotypes. To probe the status of the isotype switch process in DLBCL, 4 cases of tumor-derived constant region transcripts of all isotypes were investigated. Following the identification of the VDJ sequences, the presence of the major isotype expected from immunohistochemical analysis was confirmed at the RNA level. Another 3-4 alternative isotypes were revealed in all cases, some of which could also be detected by immunohistochemistry. All cases were somatically mutated with an intraclonal variation. In 2 cases there were clearly distinct patterns of somatic mutation between isotypes, which was consistent with independent evolution of the tumor subpopulations. There was apparent clustering of mutational patterns into either an IgMD/IgG3/IgA set or an IgG1/IgA set, indicating that the switch to IgA can occur by different routes. Alternative isotype expression is evident in DLBCL at both the RNA and protein levels. The pattern of mutation indicates that switching is occurring in subpopulations of the tumor after malignant transformation. The findings support the concept that isotype switch events may be a feature of DLBCL.