Generation and external validation of a histological transformation risk model for patients with follicular lymphoma.

Follicular lymphoma (FL) is the most frequent indolent lymphoma. 10-15% of patients suffer histological transformation (HT) to a more aggressive lymphoma, usually diffuse large B cell lymphoma (DLBCL). This study aimed to validate and improve a genetic risk model to predict HT at diagnosis. We collected mutational data from diagnosis biopsies of 64 FL patients. We combined them with the data from a previously published cohort (total n = 104, 62 from non-transformed, and 42 from patients who did transform to DLBCL). This combined cohort was used to develop a nomogram to estimate the risk of HT. Prognostic mutated genes and clinical variables were assessed using Cox regression analysis to generate a risk model. The model was internally validated by bootstrapping and externally validated in an independent cohort. Its performance was evaluated using a concordance index and a calibration curve. The clinicogenetic nomogram included the mutational status of three genes (HIST1HE1, KMT2D, and TNFSR14) and high-risk FLIPI and predicted HT with a concordance index of 0.746. Patients were classified as being at low or high risk of transformation. The probability HT function at 24 months was 0.90 in the low-risk group vs. 0.51 in the high-risk group and, at 60 months, 0.69 vs. 0.15, respectively. In the external validation cohort, the probability HT function in the low-risk group was 0.86 vs. 0.54 in the high-risk group at 24 months, and 0.71 vs. 0.32 at 60 months. The concordance index in the external cohort was 0.552. In conclusion, we propose a clinicogenetic risk model to predict FL HT to DLBLC, combining genetic alterations in HIST1H1E, KMT2D, and TNFRSF14 genes and clinical features (FLIPI) at diagnosis. This model could improve the management of FL patients and allow treatment strategies that would prevent or delay transformation.

Proposal and validation of a method to classify genetic subtypes of diffuse large B cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease whose prognosis is associated with clinical features, cell-of-origin and genetic aberrations. Recent integrative, multi-omic analyses had led to identifying overlapping genetic DLBCL subtypes. We used targeted massive sequencing to analyze 84 diagnostic samples from a multicenter cohort of patients with DLBCL treated with rituximab-containing therapies and a median follow-up of 6 years. The most frequently mutated genes were IGLL5 (43%), KMT2D (33.3%), CREBBP (28.6%), PIM1 (26.2%), and CARD11 (22.6%). Mutations in CD79B were associated with a higher risk of relapse after treatment, whereas patients with mutations in CD79B, ETS1, and CD58 had a significantly shorter survival. Based on the new genetic DLBCL classifications, we tested and validated a simplified method to classify samples in five genetic subtypes analyzing the mutational status of 26 genes and BCL2 and BCL6 translocations. We propose a two-step genetic DLBCL classifier (2-S), integrating the most significant features from previous algorithms, to classify the samples as N12-S, EZB2-S, MCD2-S, BN22-S, and ST22-S groups. We determined its sensitivity and specificity, compared with the other established algorithms, and evaluated its clinical impact. The results showed that ST22-S is the group with the best clinical outcome and N12-S, the more aggressive one. EZB2-S identified a subgroup with a worse prognosis among GCB-DLBLC cases.

Unraveling transformation of follicular lymphoma to diffuse large B-cell lymphoma.

Follicular lymphoma (FL) is an indolent but largely incurable disease. Some patients suffer histological transformation to a more aggressive subtype with poorer prognosis. This study aimed to improve our understanding of the genetics underlying FL histological transformation, and to identify genetic drivers or promoters of the transformation by elucidating the differences between FL samples from patients who did and did not transform. We conducted targeted massive parallel sequencing of 22 pre-transformed FL/transformed diffuse large B-cell lymphoma pairs and 20 diagnostic samples from non-transformed FL patients. Additionally, 22 matched samples from 11 transformed FL patients (pre-transformed FL and diffuse large B-cell lymphoma) and 9 non-transformed FLs were studied for copy number variation using SNP arrays. We identified recurrently mutated genes that were enriched at transformation, most notably LRP1B, GNA13 and POU2AF1, which have roles in B-cell differentiation, GC architecture and migration. Mutations in POU2AF1 might be associated with lower levels of expression, were more frequent in transformed FLs, and seemed to be specific to transformed- compared with de novo-diffuse large B-cell lymphomas. Pre-transformed FLs carried more mutations per sample and had greater subclonal heterogeneity than non-transformed FLs. Finally, we identified four mutated genes in FL samples that differed between patients who did and did not transform: NOTCH2, DTX1, UBE2A and HIST1H1E. The presence of mutations in these genes was associated with shorter time to transformation when mutated in the FL biopsies. This information might be useful for identifying patients at higher risk of transformation.

CD30 Expression by B and T Cells: A Frequent Finding in Angioimmunoblastic T-Cell Lymphoma and Peripheral T-Cell Lymphoma-Not Otherwise Specified.

CD30 expression in peripheral T-cell lymphoma (PTCL) and angioimmunoblastic T-cell lymphoma (AITL) is currently of great interest because therapy targeting CD30 is of clinical benefit, but the clinical and therapeutic relevance of CD30 expression in these neoplasms still remains uncertain. The aim of this study was to better quantify CD30 expression in AITL and PTCL-not otherwise specified (NOS). The secondary objective was to determine whether CD30 cells exhibit a B-cell or a T-cell phenotype. Gene expression profiling was studied in a series of 37 PTCL cases demonstrating a continuous spectrum of TNFRSF8 expression. This prompted us to study CD30 immunohistochemical (IHC) expression and mRNA levels by reverse transcription polymerase chain reaction (RT-PCR) in a different series of 51 cases (43 AITLs and 8 PTCL-NOSs) in routine samples. Double stainings with PAX5/CD30, CD3/CD30, and LEF1/CD30 were performed to study the phenotype of CD30 cells. Most (90%) of the cases showed some level of CD30 expression by IHC (1% to 95%); these levels were high (>50% of tumoral cells) in 14% of cases. CD30 expression was not detected in 10% of the cases. Quantitative RT-PCR results largely confirmed these findings, demonstrating a moderately strong correlation between global CD30 IHC and mRNA levels (r=0.65, P=1.75e-7). Forty-four of the positive cases (98%) contained CD30-positive B cells (PAX5), whereas atypical CD30-positive T cells were detected in 42 cases (93%). In conclusion, our data show that most AITL and PTCL-NOS cases express CD30, exhibiting very variable levels of CD30 expression that may be measured by IHC or RT-PCR techniques.

EBV-positive diffuse large B-cell lymphoma of the elderly is an aggressive post-germinal center B-cell neoplasm characterized by prominent nuclear factor-kB activation.

Here, we report a retrospective series of 47 EBV-positive diffuse large B-cell lymphoma associated with advanced age. Histopathology allowed to the identification of different histological patterns: cases with polymorphic diffuse large B-cell lymphoma (29 cases), Hodgkin-like (8 cases) and polymorphic lymphoproliferative disorder-like (9 cases) patterns. One case was purely monomorphic diffuse large B-cell lymphoma. We show that this lymphoma type is a neoplasm with prominent classical and alternative nuclear factor-kB pathway activation in neoplastic cells (79% of the cases showed nuclear staining for p105/p50, 74% for p100/p52 and 63% for both proteins), with higher frequency than that observed in a control series of EBV-negative diffuse large B-cell lymphoma (χ(2) <0.001). Most cases showed an activated phenotype (95% non-germinal center (Hans algorithm); 78% activated B cell (Choi algorithm)). Clonality testing demonstrated IgH and/or K/Kde/L monoclonal rearrangements in 64% of cases and clonal T-cell populations in 24% of cases. C-MYC (1 case), BCL6 (2 cases) or IgH (3 cases) translocations were detected by FISH in 18% cases. These tumors had a poor overall survival and progression-free survival (the estimated 2-year overall survival was 40 ± 10% and the estimated 2-year progression-free survival was 36 ± 9%). Thus, alternative therapies, based on the tumor biology, need to be tested in patients with EBV-positive diffuse large B-cell lymphoma of the elderly.

Improved demonstration of immunohistochemical prognostic markers for survival in follicular lymphoma cells.

Follicular lymphoma (FL) is one of the most common forms of the low-grade non-Hodgkin's lymphoma in adults, with a characteristic translocation, t(14;18)(q32;q21) that deregulates the expression of the BCL2 gene. The clinical course of FL patients is variable, whereby a subset of patients survive for long periods even without relapses, whereas the majority have frequent relapses with shorter survival. We have analyzed a series of 186 FLs, studying the correlation between clinical outcome and the tumor cell expression of a set of immunohistochemical markers, using an automated procedure for tissue microarrays to reduce the subjectivity of scoring. The results identified several markers associated with differences in overall survival (OS) in univariate analyses, such as Cyclin E, Mdm2, CD10, p21, IgD, Bcl-xL, CD30, and E2F6. Cases with a higher level of expression of Cyclin E, Mdm2, p21, IgD, Bcl-xL, CD30, and E2F6 were associated with a significantly shorter OS. On the other hand, strong CD10 expression was linked to a significantly better outcome. A Cox model was then constructed, integrating the Follicular Lymphoma International Prognostic Index (FLIPI) score and a restricted selection of three immunohistochemical markers: Cyclin E, Mdm2, and CD10 expression. A potentially useful finding is that the integrated FLIPI plus immunohistochemical model can be used to identify a subset of 26 patients (almost 20% of the total series), with a survival probability of 100% at 5 years. This not only confirms that a group of FL cases may have a very good clinical course, but also indicates that this group can be identified using this integrated clinical and immunohistochemical approach.

Functional signatures identified in B-cell non-Hodgkin lymphoma profiles.

Gene-expression profiling in B-cell lymphomas has provided crucial data on specific lymphoma types, which can contribute to the identification of essential lymphoma survival genes and pathways. In this study, the gene-expression profiling data of all major B-cell lymphoma types were analyzed by unsupervised clustering. The transcriptome classification so obtained, was explored using gene set enrichment analysis generating a heatmap for B-cell lymphoma that identifies common lymphoma survival mechanisms and potential therapeutic targets, recognizing sets of coregulated genes and functional pathways expressed in different lymphoma types. Some of the most relevant signatures (stroma, cell cycle, B-cell receptor (BCR)) are shared by multiple lymphoma types or subclasses. A specific attention was paid to the analysis of BCR and coregulated pathways, defining molecular heterogeneity within multiple B-cell lymphoma types.

TCL1A expression delineates biological and clinical variability in B-cell lymphoma.

The assembly of a collection of gene-expression signatures of the major types of B-cell non-Hodgkin's lymphoma has identified increased T-cell leukemia/lymphoma 1A (TCL1) expression in multiple lymphoma types and cases, and has enabled the investigation of the functional and clinical importance of TCL1 expression. Specifically, Burkitt's lymphoma cases show a homogeneously strong expression of TCL1, whereas diffuse large B-cell lymphoma, follicular lymphoma, mantle cell lymphoma, chronic lymphocytic leukemia, nodal marginal zone lymphoma, and splenic marginal zone lymphoma display a striking variability in the intensity of TCL1 staining. This was validated in two independent series. A Gene-Set Enrichment Analysis of the genes correlated with TCL1A expression found that variation in the level of expression of TCL1A was significantly associated with some of the most important gene signatures recognizing B-cell lymphoma pathogenesis and heterogeneity, such as germinal center, B-cell receptor, NF-kappaB (and its target genes), death, MAP kinases, TNFR1, TOLL, and IL1R. Additionally, TCL1 expression was correlated with shorter time to treatment in chronic lymphocytic leukemia cases and shorter lymphoma-specific survival in mantle cell lymphoma series, thus indicating the clinical and biological significance of TCL1 expression, and suggesting TCL1A as a potential therapeutic target.

Peripheral T-cell lymphoma with follicular T-cell markers.

INTRODUCTION: Peripheral T-cell lymphomas (PTCLs) in western countries are uncommon tumors with unfavorable prognosis. They may be subclassified as anaplastic large-cell lymphomas (ALCLs), angioimmunoblastic-T-cell lymphomas (AITLs), or unspecified peripheral T-cell lymphomas (PTCLs-U). It has recently been demonstrated that AITLs originate from germinal center follicular helper T cells (TFH), whereas the normal counterparts of other PTCLs remain essentially unknown. The aim of this study was to establish whether other PTCL subgroups also express TFH cell markers. MATERIALS AND METHODS: One hundred forty-six PTCLs were analyzed for programmed death-1 (PD-1) expression in tissue microarrays using a new monoclonal antibody called NAT-105. PD-1-positive cases, which did not fulfill all the criteria for AITL, were further evaluated in whole-tissue sections for another 12 immunohistochemical markers, including the TFH cell markers CXCL13, CD10, and BCL6. Clonal Ig and T-cell receptor rearrangements and Epstein-Barr virus-encoded RNA expression were also evaluated. Morphologic, clinical, and follow-up data were reviewed. RESULTS: Twenty-five out of 87 non-AITL cases (28.75%) showed PD-1 immunostaining. CXCL13, BCL6, and CD10 expression was found in 24/25 (96%), 16/25 (64%), and 6/25 (24%) cases, respectively. All cases expressed at least 2 TFH cell markers. Moreover, 5 cases were positive for all 4 markers. Most cases (17/25, 68%) displayed some AITL-like features. Of the remainder, 1 was considered to be early AITL, 1 was diagnosed as ALCL-anaplastic lymphoma kinase-negative, and 4 of the other 6 PTCLs-U had morphology consistent with lymphoepithelioid (Lennert's) lymphoma. Three AITL-like cases showed IgH clonal rearrangement, 2 of which were associated with Epstein-Barr virus expression. Our series of patients did not differ significantly in their clinical presentation from most reported PTCL cases in the literature: 55% of them were alive and 35% were in complete remission after a median follow-up of 15 months after cyclophosphamide, dexorubicin, vincristine, and prednisone-based chemotherapy. CONCLUSIONS: TFH cell markers, especially PD-1, were expressed in a subset of PTCLs not classified as AITL, although most of them shared some morphologic features with AITL. This suggests that the spectrum of AITL may be wider than previously thought, possibly including cases of lymphoepithelioid (Lennert's) lymphoma. Additionally, the results suggest that a subgroup of PTCLs-U, distinct from AITL and including some cases denominated as ALCL, may also be derived from TFH cells, although they develop along a distinct pathogenic pathway.

Somatic hypermutation signature in B-cell low-grade lymphomas.

BACKGROUND: Immunoglobulin gene somatic hypermutation is a biologically relevant and clinically useful prognostic factor in different types of low-grade B-cell lymphomas, including chronic lymphocytic leukemia, mantle cell lymphoma and splenic marginal zone lymphoma. DESIGN AND METHODS: With the aim of identifying surrogate markers of somatic hypermutation, a combined investigation of IgV(H) mutational status and expression profiles of 93 samples from patients with small B-cell lymphoma was performed. RESULTS: The analysis identified an somatic hypermutation signature of genes involved in the regulation of gene transcription, DNA repair and replication, and chromosome maintenance. Eight of these genes were subjected to protein analysis using tissue microarrays, for a set of 118 cases. We found a clear link between RAD51C and CDK7 protein expression and somatic hypermutation status, in that positive expression of either marker was significantly associated with a mutated status (p<0.003). We also found that positive expression of TFDP1 and POLA was significantly associated with ongoing somatic hypermutation (p<0.001). To assess the potential clinical applicability of these somatic hypermutation markers, we studied a series of cases of mantle cell lymphoma included in a tissue microarray. The expression of RCC1 and CDK7, separately and together, was found to be significantly associated with longer overall survival. CONCLUSIONS: An somatic hypermutation signature has been identified for different types of small B-cell lymphoma. This has a potential mechanistic and diagnostic value.

Comparative genome profiling across subtypes of low-grade B-cell lymphoma identifies type-specific and common aberrations that target genes with a role in B-cell neoplasia.

BACKGROUND: Low-grade B-cell lymphomas are a very heterogeneous group of tumors, whose differential diagnosis is frequently compromised by the lack of specific cytogenetic or molecular features. Our objective was to search for genomic features that allow a better molecular identification of the different types of lymphoma studied. DESIGN AND METHODS: We selected a panel of 87 low-grade B-cell lymphoma tumor samples that were unambiguously diagnosed (clinically and cytogenetically) as: follicular, splenic marginal zone, nodal marginal zone, lymphoplasmacytic, mantle cell, extranodal marginal zone MALT-type lymphoma or B-cell chronic lymphocytic leukemia. All samples were subjected to the same high-resolution genomic DNA analysis (array-based comparative genomic hybridization): a whole genome platform that contained 44000 probes distributed across the genome. Genomic imbalances were recorded, compiled and analyzed. RESULTS: Eighty percent of analyzed cases showed genomic imbalances (deletions and gain/amplifications) but the frequency of these imbalances ranged from 100% in mantle cell lymphomas to 33% in MALT lymphomas. A total of 95 new genomic imbalances affecting all lymphoma subtypes, were defined. We evaluated the extension of the genomic instability, detecting distinct patterns of genomic instability within subtypes. Specific pathways, such as nuclear factor kB (gains of REL and BCL11A, and losses of COMMD3, BIRC1, IKK1 and NFKB2), Polycomb group proteins (gain of BMI1 and deletion of PCGF6), DNA repair checkpoint pathways (deletion of 16q24 involving CDT1), or miRNA with a role in B-cell lymphoma pathogenesis (MIRN15A, MIRN16-1), were targeted by this genomic instability. CONCLUSIONS: Although all subtypes of lymphomas showed gains and losses of DNA, the analysis of their genomic profiles indicated that there are specific aberrations in almost every subtype as well as frequent aberrations that are common to a large number of lymphoma types. These common aberrations target genes that are important in B-cell lymphomagenesis.

Identification of survival-related genes of the phosphatidylinositol 3'-kinase signaling pathway in glioblastoma multiforme.

BACKGROUND: Knowledge of the molecular mechanisms involved in the biology of glioblastoma multiforme (GBM) is essential for the identification of candidate prognostic markers, new putative therapeutic targets, and early detection strategies predictive of survival. METHODS: The authors performed expression-profiling analyses in a series of primary GBMs by using complementary DNA microarrays. Validation of putative targets was performed in large series of GBMs by immunohistochemistry on tissue microarrays, real-time quantitative reverse transcription-polymerase chain reaction analysis, and Western blot analysis. RESULTS: The expression signature consisted of 159 up-regulated genes and 186 down-regulated genes. Most of these genes were involved in cell adhesion, signal transduction, cell cycle, apoptosis, and angiogenesis. Among the genes from the molecular signature, annexin 1 (ANXA1) and ubiquitin-specific protease 7 (USP7) were evaluated in wider series of GBMs. ANXA1 analysis carried out in different types of gliomas revealed exclusive overexpression in astrocytomas. Furthermore, survival analysis by using functional clusters of genes related with cancer and glioma biology revealed 7 genes involved in the PI3K-signaling pathway that presented a significant association with clinical outcome. Among these genes, positive expression of BCL2-associated X protein (BAX) was associated significantly with better survival in a larger series of tumors. In addition, activation of the PI3K/Akt pathway was demonstrated in this set of GBMs. CONCLUSIONS: The authors concluded that there is a significant role for PI3K pathway survival-related genes in patients with GBM, and putative prognostic markers associated with glioma tumorigenesis were identified. The detailed study of these candidate genes and the molecular pathways regulating PI3K activation reveal that they are promising targets for the clinical management of patients with glioma.

Gcet1 (centerin), a highly restricted marker for a subset of germinal center-derived lymphomas.

GCET1 (germinal center B cell-expressed transcript-1) gene codes for a serpin expressed in germinal center (GC) B cells. Following the observation that follicular lymphoma cases exhibit an increased level of Gcet1 expression, compared with follicular hyperplasia, we have characterized Gcet1 protein expression in human tissues, cell lines, and a large series of lymphomas. To this end, we have performed immunohistochemical and Western blot analyses using a newly generated monoclonal antibody that is reactive in paraffin-embedded tissues. Our results demonstrate that Gcet1 is expressed exclusively by neoplasms hypothetically to be arrested at the GC stage of differentiation, including follicular lymphoma, nodular lymphocyte predominant Hodgkin lymphoma, and a subset of diffuse large B-cell lymphoma, T-cell/histiocyte rich B-cell lymphoma, and Burkitt lymphoma. Within these tumors, Gcet-1 protein expression is restricted to a subset of GC B cells, establishing the existence of a distinct heterogeneity among normal and neoplastic GC B cells. None of the other B-cell lymphomas, that is, chronic lymphocytic leukemia, splenic marginal zone lymphoma, and mantle cell lymphoma, was Gcet1(+), which underlines the potential utility of Gcet1 expression in lymphoma diagnosis. The results of RNA and protein expression should prompt further investigation into the role of Gcet1 in regulating B-cell survival.

The presence of STAT1-positive tumor-associated macrophages and their relation to outcome in patients with follicular lymphoma.

BACKGROUND AND OBJECTIVES: The presence of tumor-associated macrophages (TAM) is a prognostic factor for survival in follicular lymphoma (FL). Overexpression and/or activation of the signal transducer and activator of transcription 1 (STAT1) in these TAM have also been observed. The aim of this study was to determine the extent to which macrophages are present in FL and to investigate the expression of STAT1 in these cells. DESIGN AND METHODS: We retrospectively analyzed 211 patients with distinct stages and grades of FL. Expression of the CD68 proteins, chosen as a marker for macrophages, and STAT1 was quantified by immunohistochemistry and double immunofluorescence. RESULTS: Automated determinations revealed the presence of CD68-positive macrophages in all FL tissues studied (mean 57.6+/-45.1 cells/field), while STAT1 protein was expressed in 29.94% of cases. Double-fluorescence staining confirmed that STAT1 protein co-localized exclusively with CD68, indicating the presence of a subset of STAT1-expressing TAM localized principally in the vicinity of tumor cells. Multivariate analysis showed that, besides the Follicular Lymphoma International Prognostic Index (FLIPI) classification, expression of STAT1 was an important independent prognostic factor for shorter overall survival in FL. INTERPRETATION AND CONCLUSIONS: These results demonstrate the presence of STAT1-expressing TAM in FL and their association with an adverse outcome, thus emphasizing the relevance of non-tumor cells in the control of the growth and survival of lymphoma cells.

Implication of polycomb members Bmi-1, Mel-18, and Hpc-2 in the regulation of p16INK4a, p14ARF, h-TERT, and c-Myc expression in primary breast carcinomas.

PURPOSE: Deregulation of mammalian Polycomb group (PcG) members may contribute to human carcinogenesis. p16INK4a and p14ARF tumor suppressors, human telomerase reverse transcriptase (h-TERT), and oncoprotein c-Myc have been implicated in the regulation of the cell cycle and proliferation mediated by PcG proteins, mainly Bmi-1, in mice and in cell culture experiments. Here, we examine whether these in vitro findings can be extrapolated to the in vivo situation. EXPERIMENTAL DESIGN: We measure the expression of PcG members Bmi-1, Mel-18, and Hpc-2 and their potential targets by reverse transcription-PCR, immunostaining, and Western blotting in a series of 134 breast carcinomas and correlate the data with several clinical-pathologic variables of the tumors. RESULTS: Expression of PcG genes was variably detected, but overexpression of Bmi-1 was the most frequent PcG alteration observed. In addition, statistical direct correlation in expression level of the three PcG members was detected. A correlation between c-Myc and Bmi-1 expression levels was observed; however, there was no correlation between expression of Bmi-1 and p16INK4a, p14ARF, or h-TERT. However, expression of the other PcG members Mel-18 and Hpc-2 correlated with the cell cycle regulators. Moreover, PcG mRNA-altered expression correlated significantly with certain clinical-pathologic variables associated with poor prognosis. CONCLUSIONS: Our data suggest that the oncogenic role of Bmi-1 in human primary breast carcinomas is not determined by its capacity to inhibit INK4a/ARF proteins or to induce telomerase activity.

Identification of novel candidate target genes in amplicons of Glioblastoma multiforme tumors detected by expression and CGH microarray profiling.

BACKGROUND: Conventional cytogenetic and comparative genomic hybridization (CGH) studies in brain malignancies have shown that glioblastoma multiforme (GBM) is characterized by complex structural and numerical alterations. However, the limited resolution of these techniques has precluded the precise identification of detailed specific gene copy number alterations. RESULTS: We performed a genome-wide survey of gene copy number changes in 20 primary GBMs by CGH on cDNA microarrays. A novel amplicon at 4p15, and previously uncharacterized amplicons at 13q32-34 and 1q32 were detected and are analyzed here. These amplicons contained amplified genes not previously reported. Other amplified regions containing well-known oncogenes in GBMs were also detected at 7p12 (EGFR), 7q21 (CDK6), 4q12 (PDGFRA), and 12q13-15 (MDM2 and CDK4). In order to identify the putative target genes of the amplifications, and to determine the changes in gene expression levels associated with copy number change events, we carried out parallel gene expression profiling analyses using the same cDNA microarrays. We detected overexpression of the novel amplified genes SLA/LP and STIM2 (4p15), and TNFSF13B and COL4A2 (13q32-34). Some of the candidate target genes of amplification (EGFR, CDK6, MDM2, CDK4, and TNFSF13B) were tested in an independent set of 111 primary GBMs by using FISH and immunohistological assays. The novel candidate 13q-amplification target TNFSF13B was amplified in 8% of the tumors, and showed protein expression in 20% of the GBMs. CONCLUSION: This high-resolution analysis allowed us to propose novel candidate target genes such as STIM2 at 4p15, and TNFSF13B or COL4A2 at 13q32-34 that could potentially contribute to the pathogenesis of these tumors and which would require futher investigations. We showed that overexpression of the amplified genes could be attributable to gene dosage and speculate that deregulation of those genes could be important in the development and progression of GBM. Our findings highlight the important influence in GBM of signaling pathways such as the PI3K/AKT, consistent with the invasive features of this tumor.

Nuclear bcl10 expression characterizes a group of ocular adnexa MALT lymphomas with shorter failure-free survival.

Ocular adnexa B-cell lymphomas are a relatively rare group of extranodal lymphomas, marginal-zone B-cell lymphomas of mucosa-associated lymphoid tissue (MALT lymphomas) being the most frequent type at this location. As with other nongastrointestinal MALT lymphomas, ocular adnexa MALT lymphomas have distinct characteristics from those of the gastric MALT model, implying specific pathogenic events, which could be of interest in the prediction of clinical behavior and the choice between therapeutic options. In a series of 39 cases of ocular adnexa MALT lymphomas, studied using a tissue microarray, we observed that the most frequent alteration was related to apoptosis regulation. Thus, caspase 3 activity was completely abolished, and phosphorylated IkappaBalpha, a marker of NF-kappaB activation, showed increased expression, while cases with an increased number of large cells displayed increased expression of survivin and other cell-cycle-related proteins, such as cyclin A, cyclin E and Ki67, and p16 expression was reduced. There were no occurrences of t(11;18)(q21,q21), while 5/37 cases exhibited t(14;18)(q32;q21). Aberrant nuclear expression of bcl10 was observed in 11 cases, independently of the presence of translocations, and was significantly associated with phosphorylated IkappaBalpha expression and a reduced TdT-mediated biotin-dUTP nicked-end labeling apoptotic index. Moreover, patients with tumoral bcl10 nuclear expression showed shorter failure-free survival.