Primary central nervous system lymphomas are derived from germinal-center B cells and show a preferential usage of the V4-34 gene segment

Abstract

Primary central nervous system lymphomas (PCNSLs) have recently received considerable clinical attention due to their increasing incidence. To clarify the histogenetic origin of these intriguing neoplasms, PCNSLs from 10 HIV-negative patients were analyzed for immunoglobulin (Ig) gene rearrangements. All tumors exhibited clonal IgH gene rearrangements. Of the 10 cases, 5 used the V4-34 gene segment, and all of these lymphomas shared an amino acid exchange from glycine to aspartate due to a mutation in the first codon of the complementarity-determining region 1. No preferential usage of D(H), J(H), V(kappa), J(kappa), V(lambda), or J(lambda) gene segments was observed. All potentially functional rearrangements exhibited somatic mutations. The pattern of somatic mutations indicated selection of the tumor cells (or their precursors) for expression of a functional antibody. Mean mutation frequencies of 13. 2% and 8.3% were detected for the heavy and light chains, respectively, thereby exceeding other lymphoma entities. Cloning experiments of three tumors showed ongoing mutation in at least one case. These data suggest that PCNSLs are derived from highly mutated germinal-center B cells. The frequent usage of the V4-34 gene and the presence of a shared replacement mutation may indicate that the tumor precursors recognized a shared (super) antigen.

Figure 1.

Histopathological characteristics of PCNSL. a: Lymphoma cells strongly express the CD20 antigen (CD20 immunostaining, original magnification, ×280). b: Characteristic growth pattern of a PCNSL with splitting of the blood vessel walls and a diffuse infiltration of the brain parenchyma. Note the marked edema in the affected brain tissue (H&E staining; original magnification, ×220). c: More than 50% of the tumor cells are in a proliferating state as demonstrated by nuclear Ki-67 immunoreactivity (MIB-1 immunostaining; original magnification, ×280). d: The tumor cells are embedded in an infiltrate of small, mature T cells, which reside in a perivascular location and are also scattered throughout the brain (CD3 immunostaining; original magnification, ×280).

Figure 2.

PCR analysis of rearranged IgH genes. The PCR product of 550 bp indicates the presence of a monoclonal B cell population that has rearranged a gene of the VH4 family. PCR analysis of VH families 3–5 discloses additional faint bands indicating admixed polyclonal, presumably nonmalignant B cells.

Figure 3.

Biased usage of the V4–34 gene segment in PCNSL. a: The V_H sequences (codons 1–94) of cases 1, 5, 7, 8, and 10 were aligned to the most homologous germline V_H gene segment (V4–34*02). Dashes indicate sequence identity. Point mutations are shown either as small letters for silent mutations or as capital letters for replacement mutations. Mutations in codons 93 to 94 of cases 1, 5, and 10 were not taken into account for calculation of the R/S values or the mutation frequency. The arrow points to an insertion of 3 bp between codons 27 and 28 in case 1. b: Translation of the V_H sequences from a and alignment of the amino acid sequences. Dashes indicate sequence identity and a point indicates a gap of one amino acid for better alignment. c: Complementarity-determining region 3 and beginning of FRIV of the IgH sequences (codons 95–103) of five PCNSLs, using the V4–34 gene segment. A sequence comparison demonstrates unique sequences for each case.

Figure 4.

Intraclonal diversity in a subset of PCNSL. PCR products of rearranged IgH genes were cloned in cases 4 (a), 8 (b), and 9 (c). Eight separate clones were analyzed in each case. Only those codons are shown in which sequence heterogeneity was observed. Dashes indicate sequence identity. Point mutations are labeled either as small letters for silent mutations or as capital letters for replacement mutations. gl, germline sequence.