Phospho-proteomic analysis of mantle cell lymphoma cells suggests a pro-survival role of B-cell receptor signaling.

BACKGROUND: Mantle cell lymphoma (MCL) is currently an incurable entity, and new therapeutic approaches are needed. We have applied a high-throughput phospho-proteomic technique to MCL cell lines to identify activated pathways and we have then validated our data in both cell lines and tumor tissues. METHODS: PhosphoScan analysis was performed on MCL cell lines. Results were validated by flow cytometry and western blotting. Functional validation was performed by blocking the most active pathway in MCL cell lines. RESULTS: PhosphoScan identified more than 300 tyrosine-phosporylated proteins, among which many protein kinases. The most abundant peptides belonged to proteins connected with B-cell receptor (BCR) signaling. Active BCR signaling was demonstrated by flow cytometry in MCL cells and by western blotting in MCL tumor tissues. Blocking BCR signaling by Syk inhibitor piceatannol induced dose/time-dependent apoptosis in MCL cell lines, as well as several modifications in the phosphorylation status of BCR pathway members and a collapse of cyclin D1 protein levels. CONCLUSION: Our data support a pro-survival role of BCR signaling in MCL and suggest that this pathway might be a candidate for therapy. Our findings also suggest that Syk activation patterns might be different in MCL compared to other lymphoma subtypes

Identification of a three-gene model as a powerful diagnostic tool for the recognition of ALK negative ALCL

Anaplastic Large Cell Lymphomas (ALCL) are a clinically and biologically heterogeneous disease including the ALK+ and ALK- systemic forms. While ALK+ ALCL are molecularly characterized and can be readily diagnosed, specific immunophenotypic or genetic features to define ALK- ALCL are missing, and their distinction from other T-Cell Non- Hodgkin Lymphomas (T-NHL) remains controversial. Here, we undertook a transcriptional profiling meta-analysis of 309 cases, including ALCL and other primary T-NHL samples. Pathway discovery and prediction analyses defined a minimum set of genes capable to recognize ALK- ALCL. Application of RT-qPCR in independent data sets from cryopreserved and formalin-fixed paraffin embedded (FFPE) samples validated a three-gene model (TNFRSF8, BATF3, TMOD1) able to successfully separate ALK- ALCL from PTCLNOS, with overall accuracy near 97%. In conclusion, our data justify the possibility to translate RT-qPCR protocols to routine clinical settings as a new approach to objectively dissect T-NHL and to select more appropriate therapeutic protocols

Identification of a 3-gene model as a powerful diagnostic tool for the recognition of ALK-negative anaplastic large-cell lymphoma

Anaplastic large-cell lymphomas (ALCLs) are a group of clinically and biologically heterogeneous diseases including the ALK(+) and ALK(-) systemic forms. Whereas ALK(+) ALCLs are molecularly characterized and can be readily diagnosed, specific immunophenotypic or genetic features to define ALK(-) ALCL are missing, and their distinction from other T-cell non-Hodgkin lymphomas (T-NHLs) remains controversial. In the present study, we undertook a transcriptional profiling meta-analysis of 309 cases, including ALCL and other primary T-NHL samples. Pathway discovery and prediction analyses defined a minimum set of genes capable of recognizing ALK(-) ALCL. Application of quantitative RT-PCR in independent datasets from cryopreserved and formalin-fixed paraffin-embedded samples validated a 3-gene model (TNFRSF8, BATF3, and TMOD1) able to successfully separate ALK(-) ALCL from peripheral T-cell lymphoma not otherwise specified, with overall accuracy near 97%. In conclusion, our data justify the possibility of translating quantitative RT-PCR protocols to routine clinical settings as a new approach to objectively dissect T-NHL and to select more appropriate therapeutic protocols

GeneChip analyses point to novel pathogenetic mechanisms in mantle cell lymphoma

The translocation t(11;14)(q13;q32) is the genetic hallmark of mantle cell lymphoma (MCL) but is not sufficient for inducing lymphomagenesis. Here we performed genome-wide 100K GeneChip Mapping in 26 t(11;14)-positive MCL and six MCL cell lines. Partial uniparental disomy (pUPD) was shown to be a recurrent chromosomal event not only in MCL cell lines but also in primary MCL. Remarkably, pUPD affected recurrent targets of deletion like 11q, 13q and 17p. Moreover, we identified 12 novel regions of recurrent gain and loss as well as 12 high-level amplifications and eight homozygously deleted regions hitherto undescribed in MCL. Interestingly, GeneChip analyses identified different genes, encoding proteins involved in microtubule dynamics, such as MAP2, MAP6 and TP53, as targets for chromosomal aberration in MCL. Further investigation, including mutation analyses, fluorescence in situ hybridisation as well as epigenetic and expression studies, revealed additional aberrations frequently affecting these genes. In total, 19 of 20 MCL cases, which were subjected to genetic and epigenetic analyses, and five of six MCL cell lines harboured at least one aberration in MAP2, MAP6 or TP53. These findings provide evidence that alterations of microtubule dynamics might be one of the critical events in MCL lymphomagenesis contributing to chromosomal instability

CD30 expression by bone marrow mast cells from different diagnostic variants of systemic mastocytosis

AimsCD30 expression by bone marrow (BM) mast cells (MC) has been reported recently in systemic mastocytosis (SM) patients. The aim of this study was to investigate the potential diagnostic and prognostic value of CD30 expression in SM as assessed by multiparameter flow cytometry. Methods and resultsA total of 163 consecutive BM samples corresponding to 142 SM patients and 21 non-mastocytosis cases were studied. CD30 was positive in most SM patients (80\%), but in only one non-mastocytosis case (4.8\%). When combined with CD25, CD30 contributed to an improved accuracy over that of CD25 alone (98\% versus 93\%) mainly because most (eight of nine) of the well-differentiated SM (WDSM), who lacked CD25, were CD30(+). Similar levels of expression of CD30 were observed among all different subgroups of SM except mast cell leukaemia; among indolent SM (ISM) patients, no significant association was observed between the levels of CD30 expression and other clinical and biological features of the disease. ConclusionsThe increased expression of CD30 associated with absence of CD25 contributes to the diagnosis of WDSM and its distinction from other subtypes of SM. By contrast, CD30 expression did not contribute either to prognostic stratification of ISM or to the differential diagnosis between ISM and aggressive SM cases