Philippe Gaulard

Molecular Diagnosis of Primary Mediastinal B Cell Lymphoma Identifies a Clinically Favorable Subgroup of Diffuse Large B Cell Lymphoma Related to Hodgkin Lymphoma

Using current diagnostic criteria, primary mediastinal B cell lymphoma (PMBL) cannot be distinguished from other types of diffuse large B cell lymphoma (DLBCL) reliably. We used gene expression profiling to develop a more precise molecular diagnosis of PMBL. PMBL patients were considerably younger than other DLBCL patients, and their lymphomas frequently involved other thoracic structures but not extrathoracic sites typical of other DLBCLs. PMBL patients had a relatively favorable clinical outcome, with a 5-yr survival rate of 64% compared with 46% for other DLBCL patients. Gene expression profiling strongly supported a relationship between PMBL and Hodgkin lymphoma: over one third of the genes that were more highly expressed in PMBL than in other DLBCLs were also characteristically expressed in Hodgkin lymphoma cells. PDL2, which encodes a regulator of T cell activation, was the gene that best discriminated PMBL from other DLBCLs and was also highly expressed in Hodgkin lymphoma cells. The genomic loci for PDL2 and several neighboring genes were amplified in over half of the PMBLs and in Hodgkin lymphoma cell lines. The molecular diagnosis of PMBL should significantly aid in the development of therapies tailored to this clinically and pathogenetically distinctive subgroup of DLBCL.

Survival Benefit of High-Dose Therapy in Poor-Risk Aggressive Non-Hodgkin’s Lymphoma: Final Analysis of the Prospective LNH87–2 Protocol—A Groupe d’Etude des Lymphomes de l’Adulte Study

PURPOSE To present the final analysis, with a median follow-up of 8 years, of the LNH87-2 randomized study, which compares consolidative sequential chemotherapy (ifosfamide plus etoposide, asparaginase, and cytarabine) with high-dose therapy (HDT) using cyclophosphamide, carmustine, and etoposide (CBV regimen) followed by stem-cell transplantation in patients with aggressive non-Hodgkins lymphoma in first complete remission after induction, focusing on high/intermediate- and high-risk patients identified by the age-adjusted international prognostic index. PATIENTS AND METHODS Among the 916 eligible patients, 451 presented with two (n = 318) or three (n = 133) risk factors. After reaching complete remission to induction therapy, 236 of these higher risk patients were assessable for the consolidation phase, with 125 patients in the HDT arm and 111 in the sequential chemotherapy arm. RESULTS Among these 451 higher risk patients, 277 (61%) achieved complete remission after induction treatment. In the population of 236 randomized patients, HDT was superior to sequential chemotherapy, with 8-year disease-free survival rates of 55% (95% confidence interval [CI], 46% to 64%) and 39% (95% CI, 30% to 48%), respectively (P =.02; relative risk, 1.56). The 8-year survival rate was significantly superior in the HDT arm (64%; 95% CI, 55% to 73%) compared with the sequential chemotherapy arm (49%; 95% CI, 39% to 59%) (P =.04; relative risk, 1.51). CONCLUSION On the basis of the final analysis of this prospectively treated series of patients, retrospectively analyzed on the basis of the International Prognostic Index, we hypothesize that HDT benefits patients at higher risk who achieve complete remission after induction treatment.

Immunohistochemical Prognostic Markers in Diffuse Large B-Cell Lymphoma: Validation of Tissue Microarray As a Prerequisite for Broad Clinical Applications—A Study From the Lunenburg Lymphoma Biomarker Consortium

PURPOSE The results of immunohistochemical class prediction and prognostic stratification of diffuse large B-cell lymphoma (DLBCL) have been remarkably various thus far. Apart from biologic variations, this may be caused by differences in laboratory techniques, scoring definitions, and inter- and intraobserver variations. In this study, an international collaboration of clinical lymphoma research groups from Europe, United States, and Canada concentrated on validation and standardization of immunohistochemistry of the currently potentially interesting prognostic markers in DLBCL. PATIENTS AND METHODS Sections of a tissue microarray from 36 patients with DLBCL were stained in eight laboratories with antibodies to CD20, CD5, bcl-2, bcl-6, CD10, HLA-DR, MUM1, and MIB-1 according to local methods. The study was performed in two rounds firstly focused on the evaluation of laboratory staining variation and secondly on the scoring variation. RESULTS Different laboratory staining techniques resulted in unexpectedly highly variable results and very poor reproducibility in scoring for almost all markers. No single laboratory stood out as uniformly poor or excellent. With elimination of variation due to staining, high agreement was found for CD20, HLA-DR, and CD10. Poor agreement was found for bcl-6 and Ki-67. Optimization of techniques and uniformly agreed on scoring criteria improved reproducibility. CONCLUSION This study shows that semiquantitative immunohistochemistry for subclassification of DLBCL is feasible and reproducible, but exhibits varying rates of concordance for different markers. These findings may explain the wide variation of biomarker prognostic impact reported in the literature. Harmonization of techniques and centralized consensus review appears mandatory when using immunohistochemical biomarkers for treatment stratification.

A neuronal receptor, neuropilin-1, is essential for the initiation of the primary immune response.

The initiation of a primary immune response requires contact between dendritic cells (DCs) and resting T cells. However, little is known about the proteins that mediate this initial contact. We show here that neuropilin-1, a receptor involved in axon guidance, was expressed by human DCs and resting T cells both in vitro and in vivo. The initial contact between DCs and resting T cells led to neuropilin-1 polarization on T cells. DCs and resting T cells specifically bound soluble neuropilin-1, and resting T cells formed clusters with neuropilin-1–transfected COS-7 cells in a neuropilin-1–dependent manner. Functionally, preincubation of DCs or resting T cells with blocking neuropilin-1 antibodies inhibited DC-induced proliferation of resting T cells. These data suggest that neuropilin-1 mediates interactions between DCs and T cells that are essential for initiation of the primary immune response and show parallels between the nervous and immune systems.

Efficacy of L-asparaginase with methotrexate and dexamethasone (AspaMetDex regimen) in patients with refractory or relapsing extranodal NK/T-cell lymphoma, a phase II study.

Extranodal NK/T-cell lymphoma, nasal type, is a rare and highly aggressive disease with a grim prognosis. No therapeutic strategy is currently identified in relapsing patients. We report the results of a French prospective phase II trial of an L-asparaginase-containing regimen in 19 patients with relapsed or refractory disease treated in 13 centers. Eleven patients were in relapse and 8 patients were refractory to their first line of treatment. L-Asparaginase-based treatment yielded objective responses in 14 of the 18 evaluable patients after 3 cycles. Eleven patients entered complete remission (61%), and only 4 of them relapsed. The median overall survival time was 1 year, with a median response duration of 12 months. The main adverse events were hepatitis, cytopenia, and allergy. The absence of antiasparaginase antibodies and the disappearance of Epstein-Barr virus serum DNA were significantly associated with a better outcome. These data confirm the excellent activity of L-asparaginase-containing regimens in extranodal NK/T-cell lymphoma. L-Asparaginase-based treatment should thus be considered for salvage therapy, especially in patients with disseminated disease. First-line L-asparaginase combination therapy for extranodal NK/T-cell lymphoma warrants evaluation in prospective trials. This trial is registered at www.clinicaltrials.gov as #NCT00283985.

Mediastinal Gray Zone Lymphoma: The Missing Link Between Classic Hodgkin's Lymphoma and Mediastinal Large B-cell Lymphoma

In recent years, overlap in biologic and morphologic features has been identified between classic Hodgkin lymphoma (cHL) and B-cell non-Hodgkin lymphoma. Nevertheless, the therapeutic approaches for these diseases remain different. We undertook a study of “mediastinal gray zone lymphomas” (MGZL), with features transitional between cHL nodular sclerosis (NS) and primary mediastinal large B-cell lymphoma (MLBCL) to better understand the morphologic and immunophenotypic spectrum of such cases. Twenty-one MGZL cases were identified over a 20-year period. We also studied 6 cases of composite or synchronous lymphoma with two distinct components at the same time (cHL-NS and MLBCL) and 9 sequential cases with MLBCL and cHL-NS at different times. All patients had a large mediastinal mass. Immunohistochemical studies focused on markers known to discriminate between cHL and MLBCL, including B-cell transcription factors. VJ-PCR was performed in 8 cases to look at clonality of the immunoglobulin heavy chain gene (IgH). Of the gray zone cases, 11 had morphology reminiscent of cHL-NS, but with unusual features, including a large number of mononuclear variants, diminished inflammatory background, absence of classic Hodgkin phenotype, and strong CD20 expression (11 of 11). Ten cases had morphology of MLBCL, but with admixed Hodgkin/Reed-Sternberg and lacunar cells, absent (3 of 10) or weak (7 of 10) CD20 expression, and positivity for CD15 in 7 cases. B-cell transcription factor expression in the gray zone cases more closely resembled MLBCL than cHL with expression of Pax5, Oct2, and BOB.1 in all but 1 case studied (14 of 15). MAL staining was found in 7 of 10 MGZL, and in at least one component of 6 of 7 evaluable composite or sequential MLBCL/cHL cases. Two cases of sequential lymphoma showed rearrangements of the IgH gene of identical size: one in which MLBCL was the first diagnosis and one in which MLBCL was diagnosed at relapse, indicating clonal identity for the two components of cHL and MLBCL. There is accumulating evidence that MLBCL and cHL are related entities. Further support for a relationship between MLBCL and cHL-NS is provided by composite and metachronous lymphomas in the same patient, as well as the existence of MGZL with transitional morphology and phenotype.

The Germinal Center/Activated B-Cell Subclassification Has a Prognostic Impact for Response to Salvage Therapy in Relapsed/Refractory Diffuse Large B-Cell Lymphoma: A Bio-CORAL Study

PURPOSE To evaluate the prognostic value of the cell of origin (COO) in patients with relapsed/refractory diffuse large B-cell lymphoma (DLBLC), prospectively treated by rituximab, dexamethasone, high-dose cytarabine, and cisplatin (R-DHAP) versus rituximab, ifosfamide, carboplatin, and etoposide and followed by intensive therapy plus autologous stem-cell transplantation on the Collaborative Trial in Relapsed Aggressive Lymphoma (CORAL) trial. PATIENTS AND METHODS Among the 396 patients included on the trial, histologic material was available for a total of 249 patients at diagnosis (n = 189 patients) and/or at relapse (n = 147 patients), which included 87 matched pairs. The patient data were analyzed by immunochemistry for CD10, BCL6, MUM1, FOXP1, and BCL2 expression and by fluorescent in situ hybridization for BCL2, BCL6 and c-MYC breakpoints. The correlation with survival data was performed by using the log-rank test and the Cox model. RESULTS Characteristics of immunophenotype and chromosomal abnormalities were statistically highly concordant in the matched biopsies. In univariate analysis, the presence of c-MYC gene rearrangement was the only parameter to be significantly correlated with a worse progression-free survival (PFS; P = .02) and a worse overall survival (P = .04). When treatment interaction was tested, the germinal center B (GCB) -like DLBCL that was based on the algorithm by Hans was significantly associated with a better PFS in the R-DHAP arm. In multivariate analysis, independent prognostic relevance was found for the GCB/non-GCB the Hans phenotype interaction treatment (P = .04), prior rituximab exposure (P = .0052), secondary age-adjusted International Prognostic Index (P = .039), and FoxP1 expression (P = .047). Confirmation was obtained by gene expression profiling in a subset of 39 patients. CONCLUSION COO remains a major and independent factor in relapsed/refractory DLBCL, with a better response to R-DHAP in GCB-like DLBCL. This needs confirmation by a prospective study.

Expression of CXCL13 by neoplastic cells in angioimmunoblastic T-cell lymphoma (AITL): a new diagnostic marker providing evidence that AITL derives from follicular helper T cells.

Angioimmunoblastic T-cell lymphoma (AITL) represents a distinct entity among peripheral T-cell lymphomas (PTCLs). The cellular origin of AITL remains unknown, although a possible derivation from follicular helper T cells (TFH) has been suggested based on the CD4/Bcl-6 phenotype. It has been recently shown that expression of CXCL13, a chemokine critically involved in B-cell migration into germinal centers, is characteristic of TFH cells, as compared with other T helper subsets. We compared CXCL13 expression in 29 AITLs, 20 PTCLs, unspecified, 10 anaplastic large cell lymphomas (ALCL), and 4 other PTCLs. We showed that CXCL13 is expressed by AITL (29 of 29, 100%) and a subset of PTCL, unspecified (6 of 20, 30%), which all showed borderline features with AITL, but in only 1 of 10 (10%) ALCLs, and 0 of 4 other PTCLs. Two-color immunostainings further showed that CXCL13 was found in the cytoplasm of atypical CD5-positive T cells that expressed CD10. We conclude that CXCL13 expression is a common characteristic of AITL, which can help to delineate the morphologic spectrum of the disease, and further supports its derivation from TFH cells. CXCL13 expression may also provide an additional useful tool for the diagnosis of AITL.

CD4+ CD56+ cutaneous neoplasms : A distinct hematological entity ?

We report seven cases of particular cutaneous tumors selected from the register of the French Study Group on Cutaneous Lymphomas. The patients (three men, four women) were aged 37-86 years. They initially presented with cutaneous nodules or papules. Three cases presented with regional lymph nodes. Stagings were negative, except for one patient with bone marrow involvement. Histological features were relevant with pleomorphic medium T-cell lymphoma, but these cells exhibited a distinguishing phenotype. They were positive for CD4, CD56, and also CD45, CD43, and HLA-DR. All other T-cell and B-cell markers were negative. The myelomonocytic markers (CD13, CD14, CD15, CD33, CD117, myeloperoxidase, and lysozyme) were negative excepted CD68, which was clearly positive in four cases and weakly in two cases. Others natural killer cell markers (CD16, CD57, TiA1, granzyme B), TdT, and CD34 were negative. Polymerase chain reaction studies did not detect any B or T clonal rearrangement. The cytogenetic studies, performed in five cases, showed a del(5q) in two cases. All patients were treated successfully by polychemotherapy, but relapsed quickly in the skin, between 4 and 28 months. Five patients developed bone marrow involvement, with leukemia in three cases, and they died in 5-27 months. One patient died at 17 months with skin progression. The seventh patient is alive at 33 months, with cutaneous progression. The origin of these cells is unclear. Despite expression of CD4 or CD56, we failed to demonstrate a T-cell, natural killer cell origin. However, CD4 and CD56 are not specific for T or natural killer lineages. Although these two markers are also known to be expressed by monocytic cells, classic myeloid antigens were negative. These seven cases, together with other rare similar cases already reported, seem to represent a distinct entity likely developed from hematological precursor cells.

Prognostic significance of immunohistochemical biomarkers in diffuse large B-cell lymphoma: a study from the Lunenburg Lymphoma Biomarker Consortium

The Lunenburg Lymphoma Biomarker Consortium (LLBC) evaluated the prognostic value of IHC biomarkers in a large series of patients with diffuse large B-cell lymphoma (DLBCL). Clinical data and tumor samples were retrieved from 12 studies from Europe and North America, with patients treated before or after the rituximab era. Using tissue microarrays from 1514 patients, IHC for BCL2, BCL6, CD5, CD10, MUM1, Ki67, and HLA-DR was performed and scored according to previously validated protocols. Optimal cut points predicting overall survival of patients treated in the rituximab era could only be determined for CD5 (P = .003) and Ki67 (P = .02), whereas such cut points for BCL2, BCL6, HLA-DR, and MUM1 could only be defined in patients not receiving rituximab. A prognostic model for patients treated in the rituximab era identified 4 risk groups using BCL2, Ki67, and International Prognostic Index (IPI) with improved discrimination of low-risk patients. Newly recognized correlations between specific biomarkers and IPI highlight the importance of carefully controlling for clinical and biologic factors in prognostic models. These data demonstrate that the IPI remains the best available index in patients with DLBCL treated with rituximab and chemotherapy.