Alterations of the CD58 gene in classical Hodgkin lymphoma.

Immune evasion plays a central role in the pathophysiology of classical Hodgkin lymphoma (cHL). As mutations of the CD58 gene contribute to immune evasion of diffuse large B cell lymphoma tumor cells, we studied whether alterations of the CD58 gene also occur in Hodgkin and Reed/Sternberg (HRS) cells of cHL. Single nucleotide polymorphism chip analysis revealed homozygous deletions within the CD58 gene in two cHL cell lines (SUP-HD1 and U-HO1). Sequencing of the CD58 gene in seven cHL cell lines disclosed in addition a homozygous splice site mutation in cell line KM-H2. None of the three mutated lines expressed CD58 protein on their surface. Thus, three of seven cHL cell lines analyzed harbor destructive CD58 mutations. Molecular analysis of isolated HRS cells from 10 primary cases of cHL; however, did not reveal any case with a CD58 mutation. A FICTION study indicated heterozygous deletions of CD58 in 3 of 13 cHL analyzed. Overall, we report frequent inactivating mutations of CD58 in cHL cell lines, but their rare occurrence in primary HRS cells. As the three cHL cell lines with CD58 mutations were all established from HRS cells located in pleural effusions, i.e., outside the normal lymph node microenvironment, in end-stages of the disease, CD58 inactivation in cHL might be predominantly prevalent to such situations.

Transcriptional profiling of the nuclear factor-kappaB pathway identifies a subgroup of primary lymphoma of the central nervous system with low BCL10 expression.

Recent studies point to a role of nuclear factor (NF)-kappaB signaling in a subset of diffuse large B cell lymphomas. We have analyzed the expression of 21 genes encoding NF-kappaB family members, upstream modulators, and targets in 32 primary central nervous system lymphomas (PCNSLs) by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). Compared with nonmalignant germinal center centroblasts, expression of BCL10, REL, IAP1, and TRAF1 was significantly lower in PCNSLs, whereas that of BAX, BCLXL, BCL2, MALT1, CARD9, CARD10, CARD11, CARD14, CCND2, cFLIP, RELA, RELB, NFKB1, NFKB2, and IRF4 was higher. Hierarchical clustering of gene expression data revealed two distinct subgroups of PCNSLs, which were characterized by significantly different transcriptional levels, predominantly of BCL10, but also of REL and IAP1. Thus, these quantitative RT-PCR data with expression of genes of the NF-kappaB family as well as NF-kappaB-regulated genes together with immunohistochemical detection of nuclear RELA and REL indicate activation of the NF-kappaB pathway in PCNSLs, which may contribute to their high proliferative activity and the low level of apoptosis.

Chromosomal translocations fusing the BCL6 gene to different partner loci are recurrent in primary central nervous system lymphoma and may be associated with aberrant somatic hypermutation or defective class switch recombination.

Primary central nervous system lymphomas (PCNSLs) are diffuse large B cell lymphomas confined to the brain. Only minimal data exist on chromosomal aberrations underlying PCNSLs. We studied 41 PCNSLs by fluorescence in situ hybridization for breakpoints affecting the BCL6 locus in chromosomal band 3q27. Of 37 cases evaluable, 14 (38%) carried a breakpoint in the BCL6 locus. Two of these showed juxtaposition of BCL6 to the IGH locus. In 4 cases, the BCL6 breakpoints were cloned using long-distance inverse polymerase chain reaction. All breakpoints were located within the BCL6 major translocation cluster. The translocation partners were the IGH gene in 14q32.33, the IGL gene in 22q11.22, and the histone 1 H4I gene in 6p22.1. In the fourth case, a deletion in 3q leads to loss of an 837-kb fragment extending from the first intron of BCL6 to the third intron of the lipoma-preferred partner (LPP) gene. This deletion may bring the BCL6 gene under the control of regulatory elements of the LPP gene or the miRNA-28 gene located in intron 4 of LPP. DNA sequence analysis of the junctional sequences provided evidence that aberrant class switch recombination or somatic hypermutation may be involved in the generation of BCL6 translocations.

Molecular characterization of BCL6 breakpoints in primary diffuse large B-cell lymphomas of the central nervous system identifies GAPD as novel translocation partner.

Primary central nervous system lymphomas (PCNSL) constitute diffuse large B-cell lymphomas arising in and remaining confined to the brain. Little information is available on cytogenetic changes in PCNSL, and recurrent chromosomal translocations have not yet been identified. Fluorescence in situ hybridization (FISH) of a series of 13 PCNSL from immunocompetent patients revealed 3 cases with signal patterns of a BCL6-specific probe suggesting a breakpoint in this oncogene locus in chromosome band 3q27. Here, we describe cloning of the translocation breakpoints by long-distance inverse polymerase chain reaction (LDI-PCR) in 2 of these tumors. Both breakpoints affected the first intron of BCL6. In one PCNSL, the HSPCA (HSP90A) gene in 14q32.31 was identified as BCL6 partner. In the second lymphoma, the gene encoding glyceraldehyde-3-phosphate dehydrogenase (GAPD) on 12p13.31 was detected as a hitherto unknown partner of BCL6. Our results suggest translocation-mediated BCL6 oncogene activation as a so far unknown pathogenetically relevant mechanism in PCNSL.

Interphase cytogenetic analysis of lymphoma-associated chromosomal breakpoints in primary diffuse large B-cell lymphomas of the central nervous system.

Primary central nervous system lymphomas (PCNSLs) are germinal center-derived diffuse large B-cell lymphomas (DLBCLs) arising in and remaining confined to the brain, the pathogenesis of which is poorly understood. We investigated 13 PCNSLs from immunocompetent patients by means of interphase cytogenetics on cryopreserved cells derived from stereotactic biopsies. Interphase fluorescence in situ hybridization (FISH) was performed for the detection of structural alterations affecting the IGH (14q32), IGK (2p12), IGL (22q11), BCL6 (3q27), MYC (8q24), CCND1 (11q13), MLT, and BCL2 (both 18q21) loci. Signal constellations indicating breakpoints within the IGH and IGK locus were detected in 5 and 1 PCNSLs, respectively. There was no evidence for a t(8;14), t(11;14), or t(14;18) in this series of tumors. Breakpoints in the BCL6 locus were observed in 3 of the 13 cases, and nuclear Bcl-6 protein expression was detected in 6 of 9 PCNSLs, including those with genomic alterations of the encoding locus. Gains of 18q21 represented the most frequent imbalances present in more than one third of all cases. Interestingly, these gains included the MLT gene. Thus, this study provides the first evidence for recurrent chromosomal translocations in PCNSLs. While they share similarities with extracerebral DLBCL with respect to the presence of IGH translocations, they appear to differ in the usage of translocation partner genes, which remain to be identified.