Comparative expressed sequence hybridization studies of t(11;18)(q21;q21)-positive and -negative gastric MALT lymphomas reveal both unique and overlapping gene programs.

Among the genetic abnormalities reported to occur in MALT lymphomas, the translocation t(11;18)(q21;q21) is of particular interest because it is exclusively documented in MALT lymphomas, mainly with gastrointestinal location. It results in the creation of a fusion protein API2-MALT1 that activates the transcription factor NF-kappaB through enhanced IKK gamma polyubiquitination. Here, we apply the recently developed molecular technique termed comparative expressed sequence hybridization to identify differentially expressed chromosomal regions related to the pathogenesis of gastric MALT lymphomas. By comparing t(11;18)(q21;q21)-positive gastric MALT lymphomas to their t(11;18)(q21;q21)-negative counterparts, we found that the location of the MALT1 break point determines a difference in expression pattern within the t(11;18)(q21;q21)-positive group. Moreover, we could define a gastric MALT lymphoma signature, which most likely comprises the regions and genes with significance in the development of MALT lymphomas, by comparing both t(11;18)(q21;q21)-positive and -negative MALT lymphomas to normal lymphoid tissue. Finally, a significant imprint of the marginal zone signature, established by comparing microdissected, splenic B follicles with and without marginal zone, was evident in the expression profile of MALT lymphoma, further supporting a marginal zone origin for this type of B-cell non-Hodgkin's lymphoma.

Translocations targeting CCND2, CCND3, and MYCN do occur in t(11;14)-negative mantle cell lymphomas.

The genetics of t(11;14)(q13;q32)/cyclin D1-negative mantle cell lymphoma (MCL) is poorly understood. We report here 8 MCL cases lacking t(11;14) or variant CCND1 rearrangement that showed expression of cyclin D1 (2 cases), D2 (2 cases), and D3 (3 cases). One case was cyclin D negative. Cytogenetics and fluorescence in situ hybridization detected t(2;12)(p11;p13)/IGK-CCND2 in one of the cyclin D2-positive cases and t(6;14)(p21;q32)/IGH-CCND3 in one of the cyclin D3-positive cases. Moreover, we identified a novel cryptic t(2;14)(p24;q32) targeting MYCN in 2 blastoid MCLs: one negative for cyclin D and one expressing cyclin D3. Interestingly, both cases showed expression of cyclin E. Notably, all 3 blastoid MCLs showed a monoallelic deletion of RB1 associated with a lack of expression of RB1 protein and monoallelic loss of p16. In sum-mary, this study confirms frequent aberrant expression of cyclin D2 and D3 in t(11;14)-negative MCLs and shows a t(11;14)-independent expression of cy-clin D1 in 25% of present cases. Novel findings include cyclin E expression in 2 t(11;14)-negative MCLs characterized by a cryptic t(2;14)(p24;q32) and identification of MYCN as a new lymphoma oncogene associated with a blastoid MCL. Clinically important is a predisposition of t(11;14)-negative MCLs to the central nervous system involvement.

T-cell/histiocyte-rich large B-cell lymphoma shows transcriptional features suggestive of a tolerogenic host immune response.

BACKGROUND: Gene expression profiling has successfully identified the prognostic significance of the host response in lymphomas. The aggressive T-cell/histiocyte-rich large B-cell lymphoma and the indolent nodular lymphocyte-predominant Hodgkin's lymphoma are both characterized by a paucity of tumor cells embedded in an overwhelming background. The tumor cells of both lymphomas share several characteristics, while the cellular composition of their microenvironment is clearly different. DESIGN AND METHODS: We collected 33 cases of T-cell/histiocyte-rich large B-cell lymphoma and 56 cases of nodular lymphocyte-predominant Hodgkin's lymphoma and performed microarray gene expression profiling on ten cases of each lymphoma, to obtain a better understanding of the lymphoma host response. By quantitative reverse transcriptase polymerase chain reaction we verified that these 20 selected cases were representative of the entire population of T-cell/histiocyte-rich large B-cell and nodular lymphocyte-predominant Hodgkin's lymphomas. RESULTS: We observed that the microenvironment in nodular lymphocyte-predominant Hodgkin's lymphoma is molecularly very similar to a lymph node characterized by follicular hyperplasia, while the microenvironment in T-cell/histiocyte-rich large B-cell lymphoma is clearly different. The T-cell/histiocyte-rich large B-cell lymphoma signature is hallmarked by up-regulation of CCL8, interferon-gamma, indoleamine 2,3 dioxygenase, VSIG4 and Toll-like receptors. These features may be responsible for the recruitment and activation of T cells, macrophages and dendritic cells, characterizing the stromal component of this lymphoma, and may point towards innate immunity and a tumor tolerogenic immune response in T-cell/histiocyte-rich large B-cell lymphoma. CONCLUSIONS: The gene expression profile of T-cell/histiocyte-rich large B-cell lymphoma, in comparison with that of nodular lymphocyte-predominant Hodgkin's lymphoma, shows features suggestive of a distinct tolerogenic host immune response that may play a key role in the aggressive behavior of this lymphoma, and that may serve as a potential target for future therapy.

Telomeric IGH losses detectable by fluorescence in situ hybridization in chronic lymphocytic leukemia reflect somatic VH recombination events.

Routine interphase fluorescence in situ hybridization (FISH) analysis of chronic lymphocytic leukemia (CLL) with LSI IGH/CCND1 assay, applied to differentiate CLL from leukemic mantle cell lymphoma, identified a subset of cases (42/174) with translocation-like IGH signal pattern. To unravel the underlying 14q32/IGH aberrations, 14 of these cases were subjected to cytogenetic, detailed FISH, and V(H) mutation analyses. FISH identified cryptic losses of various portions of the IGHV region in all 14 cases. Fine mapping of these V(H) deletions revealed a strict correlation between their distal border and localization of the used VH gene, suggesting that they are not oncogenic but reflect physiological events accompanying somatic V-D-J assembly. This hypothesis was further supported by FISH analysis of 20 CLL and hairy cell leukemia cases with the known V(H) usage showing a constant loss of sequences proximal to the used gene, identification of V(H) deletions in normal B cells, and their exclusive demonstration in B cell malignancies, but not of T cell and myeloid linage. Given that these cryptic physiological VH losses in B cells may seriously complicate analysis of B cell leukemia/lymphoma and lead to false conclusions, FISH users should take them into consideration when interpreting IGH aberrations in these malignancies.

Is there a difference in childhood T-cell acute lymphoblastic leukaemia and T-cell lymphoblastic lymphoma?

To distinguish the similarities or differences between T-cell acute lymphoblastic leukaemia (T-ALL) and T-cell lymphoblastic lymphoma (T-LBL), we retrospectively analyzed the clinical, immunophenotypic, cytogenetic, and molecular characteristics in 37 children diagnosed between December 1990 and December 2003. Comparative Expressed Sequence Hybridisation (CESH) was used to determine gene expressing profile in both diseases. Twenty two patients suffered from T-ALL and 15 patients were diagnosed as T-LBL. Immunophenotyping demonstrated a more immature phenotype in T-ALL and a more mature phenotype in T-LBL. Cytogenetic and molecular genetic aberrations were found in 82% of T-ALL compared with 73% of T-LBL. By CESH gene expression profiling, the investigated cases were segregated into two groups that largely corresponded with T-ALL and T-LBL. The clinical presentation and cytogenetic characteristics are largely similar for T-ALL and T-LBL supporting the concept that both represent a spectrum of one single disease. The differences that were found between both neoplasms, in particular in their phenotype and in their expression profile may suggest that most T-ALL derive from a T-cell progenitor of the bone marrow, while thymocytes represent the normal counterpart of T-LBL.

Large cleaved and immunoblastic lymphoma may represent two distinct clinicopathologic entities within the group of diffuse large B-cell lymphomas.

PURPOSE: The reliability of immunohistochemistry for subdividing diffuse large B-cell lymphomas (DLBCL) into germinal center B-cell-like (GCB) and non-GCB prognostic subgroups is debated. In this study we evaluated the prognostic significance of such subgrouping on a series of 153 DLBCL patients. Furthermore, we investigated whether both subgroups could comprise clinicopathologic entities recognized by their morphology and characterized by a distinct phenotype, specific genetic abnormalities, and clinical characteristics. PATIENTS AND METHODS: All samples from patients were reviewed and morphologically subdivided into large cleaved, immunoblastic, and not otherwise specified DLBCL. GCB and non-GCB immunohistochemical profiles were established. The presence of chromosomal translocations involving BCL2, BCL6, and MYC and/or rearrangements of these genes was investigated. RESULTS: Subdividing DLBCL with either a GCB or non-GCB immunophenotypic profile was not of prognostic significance. Nevertheless, CD10 expression was a predictor of favorable outcome, whereas high bcl-2 expression and BCL6 rearrangement were adverse predictors of disease-free survival. Interestingly, large cleaved DLBCL was clearly associated with a GCB immunophenotypic profile, CD10 expression, BCL2 rearrangement, age younger than 60 years, and low to low/intermediate International Prognostic Index risk, but was not of prognostic significance. In contrast, immunoblastic morphology was associated with a non-GCB profile and was a significant predictor of unfavorable DFS. CONCLUSION: Subdividing DLBCL into subgroups based on their immunohistochemical profile was not of prognostic significance. Nevertheless, it allowed the additional characterization of two lymphoma subgroups previously recognized in the Working Formulation. Both correspond to two distinct clinicopathologic entities within the DLBCL.

T(11;18)(q21;q21)-positive gastrointestinal MALT lymphomas are heterogeneous with respect to the V(H) gene mutation status.

AIM: To investigate how t(11;18)(q21;q21)-positive gastrointestinal MALT lymphomas relate to other marginal zone lymphomas with respect to the somatic mutation pattern of the V(H) genes and the expression of the marker CD27. METHODS: The V(H) gene of 7 t(11;18)(q21;q21)-positive gastrointestinal MALT lymphomas was amplified by PCR using family specific V(H) primers and a consensus J(H) primer. PCR products were sequenced and mutation analysis of the CDR and the FR regions was performed. All cases were immunostained for CD27. RESULTS: One case showed unmutated V(H) genes while the others showed mutated V(H) genes with mutation frequencies ranging from 1.3 to 14.7% and with evidence of antigen selection in 2 cases. These data suggest that the translocation t(11;18)(q21;q21) can target either B-cells at different stages of differentiation or naive B-cells that retain the capacity to differentiate upon antigen stimulation. All cases but one displayed weak to strong CD27 expression which did not correlate with the V(H) gene mutation status. CONCLUSION: t(11;18)(q21;q21)-positive gastrointestinal MALT lymphomas are heterogeneous with respect to the V(H) mutation status and CD27 is not a marker of somatically mutated B-cells.

Comparative expressed sequence hybridization reveals differential gene expression in morphological breast cancer subtypes.

In this study, comparative expressed sequence hybridization (CESH) has been used to compare gene expression patterns in three morphologically different breast cancer subtypes: classic-type invasive lobular carcinoma (ILC), poorly differentiated ERBB2-negative invasive ductal carcinoma-not otherwise specified (IDC-NOS), and poorly differentiated ERBB2-positive IDC-NOS. CESH allows global detection of chromosomal regions with differential gene expression in a way similar to that of comparative genomic hybridization (CGH). Eight cases of each breast cancer subtype were included in the study. For each subtype, two pools of four cases each were constructed. CESH was used to compare both pools within the same morphological subtype, followed by a comparison of pools belonging to different subtypes. This revealed three chromosomal regions that were differentially expressed in ductal and lobular carcinomas, including relative overexpression at 8q13-q23 and 16q22, and relative underexpression at 8p21-p22. In addition, an expression signature characterized by relative overexpression at 3q24-q26.3, 14q23-31, 17q12, and 20q12-13 was identified for ERBB2-positive IDC-NOS. In summary, CESH analysis highlights chromosomal regions of differential gene expression that are associated with morphologically defined breast cancer subtypes and suggests that regions on chromosome 8 are of interest in the discrimination between ductal and lobular carcinomas. In addition, using CESH, it was possible to identify an ERBB2 expression signature, comprising four chromosomal regions with potential significance in the aggressive behaviour of ERBB2-positive IDC-NOS.

Forkhead box protein P1 expression in mucosa-associated lymphoid tissue lymphomas predicts poor prognosis and transformation to diffuse large B-cell lymphoma.

PURPOSE: Gene expression profiling studies have reported upregulated mRNA expression of forkhead box protein P1 (FOXP1) in response to normal B-cell activation and high expression in a poor prognosis subtype of diffuse large B-cell lymphoma (DLBCL). Recently, it was also found that FOXP1 rearrangements and expression of its protein occur in mucosa-associated lymphoid tissue (MALT) lymphomas. In this study, we investigated FOXP1 expression in its relationship to morphology, genetic features, and prognosis in a series of 70 MALT lymphomas. PATIENTS AND METHODS: All samples were morphologically reviewed and stained for FOXP1. Presence of structural and/or numeric aberrations of the FOXP1, BCL10, and MALT1 genes was investigated. For all patients, a complete clinical data set was collected. RESULTS: We detected nuclear expression of FOXP1 in 20 of the 70 MALT lymphomas (nine of them featuring structural or numeric aberrations of the FOXP1 locus). FOXP1 positivity was confined to MALT lymphomas with poor clinical outcome (with impact of FOXP1 expression on relapse rate and disease-free survival). It was also found that MALT lymphomas with strong FOXP1 expression are at risk of transforming into an aggressive DLBCL of nongerminal center phenotype if they feature, in addition, a polymorphic histology and the presence of trisomy 3 and 18. CONCLUSION: The data presented show that FOXP1 expression is an independent prognostic factor in MALT lymphomas. The data also support the hypothesis that a subgroup of nongerminal center DLBCLs (those marked by FOXP1 expression and trisomy 3 and 18) might represent a large-cell variant of MALT lymphomas.

Comparative expressed sequence hybridization studies of hairy cell leukemia show uniform expression profile and imprint of spleen signature.

Comparative expressed sequence hybridization (CESH) to chromosomes is a recently introduced technique that identifies chromosomal regions corresponding to a differential gene expression. This technique is analogous to comparative genomic hybridization (CGH) that detects genomic imbalances. We applied CESH for the study of hairy cell leukemia (HCL), a disorder with a largely unknown expression profile. Twelve HCL cases with spleen involvement were investigated by CESH and CGH. While the latter analysis identified only a few nonrecurrent genomic imbalances, CESH showed a consistent expression profile in all HCL cases. In addition, pairing normal spleen with normal lymph node, a "spleen signature" was established by CESH. This signature most likely reflects the expression profile of spleen-specific components, such as the sinusoidal lining cells from the red pulp and the marginal zone B cells from the white pulp. Imprint of the spleen signature was found in the HCL expression profile, suggesting that HCL may originate from a particular B-cell subset present in these splenic components. Besides pairing HCL with normal lymph node and spleen, we identified an "HCL signature" comprising several chromosome regions with altered expression. The most significantly underexpressed regions include 3p24, 3p21, 3q13.3-q22, 4p16, 11q23, 14q22-q24, 15q21-q22, 15q24-q25, and 17q22-q24; and 13q31 and Xq13.3-q21 were the most significantly overexpressed. These regions possibly harbor genes related to the biology and the pathogenesis of HCL. Their identification warrants further molecular investigations.