non-GCB Subtype Research Articles

Clinical and Biological Features of Primary Testicular B-Cell Lymphoma - a Single-Institution Study of 89 Cases

Context: Primary testicular lymphoma (PTL) is an aggressive form of extranodal lymphoma almost always of B-cell lineage. The data regarding prognostic indicators and appropriate treatment regimens in patients with PTL, especially in the era of rituximab-cyclophosphamide, hydroxydaunorubicin, vincristine and prednisone (R-CHOP) therapy are not well established.Objective: To evaluate the clinicopathologic features and their impact on outcome in patients with primary testicular lymphoma (PTL) of B-cell lineage.Patients and Design: We retrospectively analyzed the characteristics of 89 patients with PTL referred to our institution between September 1, 1998 and May 31, 2014. Follow up data was available for all patients. Detailed staging and therapy information were available for 69 and 63 patients, respectively. Cell of origin classification of DLBCL cases was determined by immunohistochemistry when possible.Results: The median age at diagnosis was 63.1 years (range, 2.9-96.2). Patients presented with stage I (26/66, 40%), stage II (14/66, 21%), stage III (4/66, 1%) and stage IV (25/66, 38%) disease, respectively. Histologic variants included: diffuse large B-cell lymphoma (DLBCL) (n=82), Burkitt lymphoma (BL) (n=2), B-cell lymphoma, unclassifiable, with features intermediate between DLBCL and BL (n=4) and blastoid mantle cell lymphoma (n=1). Twenty-six (40%) patients had involvement of spermatic cord, epididymis and paratesticular tissues and 16 (18%) tumors showed coagulative tumor necrosis. In 31 cases DLBCL was stratified into germinal center B-cell-like (n=16, 52%) and non-germinal center B-cell-like (n=15, 42%). Six of 36 (16%) tumors analyzed showed CD5 expression by immunohistochemistry and/or flow cytometry. The median Ki-67 proliferation index was 80% in 24 cases analyzed (range, 40-100%). Four of 16 cases (25%) analyzed were positive for MYC rearrangement by fluorescence in situ hybridization. Among patients with available data (n=63); all received systemic chemotherapy (41, R-CHOP, 9 CHOP and 13 other regimens) +/- prophylactic intrathecal chemotherapy (n=48) and/or consolidation involved field radiation therapy (n=37, 23 patients with stage I/II and 12 patients with stage III/IV disease). Response to therapy was documented in 61 patients; 85% achieved complete remission, 7% partial remission and 8% had progressive disease (PD). Recurrent and/or PD were seen in 28 of 65 (43%) patients with available data. The most frequent sites of recurrence included lymph nodes (23%), soft tissues (19%), leptomeninges/brain (19%) and contralateral testicle (13%). The median overall survival (OS) and progression free survival (PFS) were 84.7 and 55.7 months, respectively. Advanced Ann Arbor stage (p=0.05, PFS), involvement of ≥2 extranodal sites (p=0.05, OS), elevated serum LDH level (p=0.013 and 0.026, OS and PFS respectively), and International Prognostic Index ≥2 (p=0.04, OS) were significantly associated with poorer outcome. Administration of prophylactic intrathecal chemotherapy (p=0.0017 and 0.012, OS and PFS, respectively) and consolidation radiation therapy (Figure 1, p=0.01 and 0.033, OS and PFS, respectively) were significantly associated with better outcome by univariate analysis. The association of consolidation radiation therapy with outcome was diminished in patients with stage III and IV disease. There was no significant association between different chemotherapy regimens and outcome, although patients who received rituximab showed a trend for better OS and PFS. Microscopic involvement of spermatic cord, epididymis and paratesticular tissue was associated coagulative necrosis (p= 0.04) and poorer outcome (p=0.018 and 0.023, OS and PFS respectively). Non-DLBCL histology type was significantly associated with nodal involvement at presentation (p=0.05). We observed a trend for shorter PFS in patients whose tumors had a Ki-67 proliferation index of ≥90% (p=0.2). There was no significant association between GCB vs. non-GCB subtypes, CD5 expression or MYC gene rearrangement and outcome.Conclusions: Our results indicate that PTL of B-cell lineage represents a distinctive group of extranodal lymphomas with unique clinical, biological and prognostic features. The data further demonstrate the beneficial therapeutic impact of prophylactic intrathecal chemotherapy and consolidation radiation therapy in patients with PTL. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

The Absolute Lymphocyte to Monocyte Ratio at Diagnosis Further Stratifies Survival Based on Cell of Origin in Diffuse Large B-Cell Lymphoma

Background: Diffuse large B-cell Lymphoma (DLCBL) is a heterogeneous disease. Current prognostic models have limitations, but emerging research utilizing the cell of origin and host immune factors, such as the absolute monocyte count (AMC) and absolute lymphocyte count (ALC), may improve risk stratification in these patients. In the rituximab era, the majority of relapses are observed within the first 24 months following treatment. Therefore, event free survival at 24 months (EFS24) is a reliable surrogate for overall survival in DLBCL [Maurer et. al., J Clin Oncol, 2014 Apr 1; 32(10): 1066-73]. The primary objective of this study was to determine if the ALC/AMC ratio predicts EFS24 in DLBCL risk-stratified by the cell of origin (COO).Methods: Patients with de novo DLBCL who were evaluated at the University of Michigan Comprehensive Cancer Center between 1997 and 2011 and had archived diagnostic tissue available were eligible for participation in this retrospective study. We identified 243 eligible patients. The ALC and AMC were obtained from a complete blood count with differential at the time of diagnosis. Immunohistochemical staining for CD10, Bcl-6 and IRF4/MUM1 was performed and cases independently scored in a blinded fashion by up to three hematopathologists. A consensus opinion was obtained in order to determine the cell of origin (COO) using the Hans algorithm.Results: The median age at diagnosis was 61 years with a median follow-up of 3.1 years. Among the 218 patients for whom the revised IPI could be calculated, 46% were high risk (IPI 3-5). Most patients received rituximab based chemoimmunotherapy (88%), largely R-CHOP (81%). The median ALC/AMC ratio for the entire cohort was 2 (interquartile range: 1-3). The ALC/AMC ratio was analyzed as a continuous variable on univariate analysis and was associated with inferior EFS (hazard ratio (HR) =0.81, 95% confidence interval (CI) 0.68-0.93, p=0.003). A receiver operating characteristic curve (ROC) was generated utilizing EFS24 to establish the optimal cut-off point for the ALC/AMC ratio. An ALC/AMC ratio of 1.6 was best able to risk-stratify patients for EFS24. The 24-month EFS for patients with an ALC/AMC ≥1.6 was 73% and for ALC/AMC <1.6 was 47% (p<0.001), and the 5-year OS was 68% vs 51% (p=0.004), respectively. On multivariate analysis the ALC/AMC ratio and R-IPI were both independently significant for EFS (HR 1.5, 95% CI 1.0-2.4, p= 0.036 and HR 1.88, 95% CI 1.2-2.9, p=0.003, respectively).Tissue exhaustion precluded determination of the COO for many patients. Among the remaining 112 patients, 63% were GCB and 37% non-GCB. Mean ALC/AMC ratios for the GCB and non-GCB subtypes were not significantly different at 2.2 and 2.4, respectively. Among GCB DLBCL the EFS24 and 5-year OS for ALC/AMC ≥1.6 vs ALC/AMC <1.6 was 77% vs 46% (p = <0.0001) and 71% vs 50% (p = 0.0007), respectively. Among non-GCB DLBCL, the EFS24 was 70% for ALC/AMC ≥1.6 and 49% for ALC/AMC <1.6 (p=0.025). No significant difference in 5-year OS was observed when non-GCB DLBCL were stratified by the ALC/AMC ratio.Conclusions: The ALC/AMC ratio is an independent prognostic factor for EFS24 in both GCB and non-GCB DLBCL. DisclosuresNo relevant conflicts of interest to declare.

CC-122 Has Potent Anti-Lymphoma Activity through Destruction of the Aiolos and Ikaros Transcription Factors and Induction of Interferon Response Pathways

Background: IMiDs® Immunomodulatory agent lenalidomide (Len) is active in a range of hematologic cancers including diffuse large B-cell lymphoma (DLBCL). The molecular target of Len is cereblon (CRBN), a substrate receptor of the cullin 4 RING E3 ubiquitin ligase complex. In a CRBN-dependent manner, Len promotes ubiquitination of lymphoid transcription factors Ikaros and Aiolos leading to their subsequent degradation. This was shown to contribute to Len’s anti-Multiple Myeloma and T-cell activation properties. CC-122 is a non-phthalimide analog of thalidomide and has potent anti-proliferative, immunomodulatory and anti-angiogenic activities giving it a potentially broader range of activity than Len. CC-122, a first-in-class pleiotropic pathway modifier (PPM®), is currently in a Phase I clinical trial for DLBCL, multiple myeloma, and solid tumors (ClinicalTrials.gov NCT01421524) and showing encouraging clinical activity. The present study investigated the mechanism of action of CC-122 in pre-clinical models of DLBCL and in clinical samples from the ongoing Phase I study.Results: CC-122 inhibited proliferation (IC50=0.01-1.5μΜ) and induced apoptosis (6.5-12 fold) in 4 ABC and 3 GCB DLBCL cell lines in a dose-dependent manner. CC-122 induced rapid degradation of Aiolos and Ikaros in a CRBN-dependent manner; an effect blocked by MLN4924, a small molecule inhibitor of Cullin-RING E3 ligase complexes. In vivo anti-tumor activity of CC-122 was demonstrated in 2 xenograft models of human DLBCL, and was associated with significant reduction in Aiolos and Ikaros expression (94% and 69%, respectively, p<0.001) in tumor tissue within 6 hrs of administration. To identify proteins and signaling pathways affected by CC-122, tandem-mass tag proteomics was performed on DLBCL cells treated with vehicle or CC-122. While Aiolos and Ikaros levels decreased in a time- and dose-dependent manner, Interferon Stimulated Genes (ISGs) including IRF7, DDX58, and IFIT3 were notably increased (3, 5 and 12.5 fold, respectively). ISG induction with 24 hr CC-122 treatment was further confirmed by Western analysis. Surprisingly, the induction of ISGs occurred in the absence of increased interferon α, β, or γ levels, which was confirmed by qPCR and ELISA. Furthermore, shRNA targeting Aiolos led to inhibition of Aiolos protein expression (2-3 fold) and an increase in the mRNA and protein levels of IRF7, DDX58 and IFIT3 (10, 14 and 22 fold in mRNA expression, respectively). These data indicate that the increase in ISGs is a direct consequence of Aiolos degradation by CC-122 and bypasses the need for interferon production. Exploration of additional downstream functional gene candidates of CC-122 and shAiolos activity revealed a decrease in protein levels of Myc (up to 3 fold decrease with shAiolos), a key driver of DLBCL proliferation and survival associated with poor survival. Importantly, reduction of Aiolos expression and the consequent changes in the molecular pathways described led to inhibition of proliferation (2-4 fold versus control shRNA) and increased apoptosis (3-10 fold vs control shRNA), which paralleled CC-122 effects in DLBCL cells lines. Of note, CC-122 induction of ISGs was comparable in both ABC and GCB lines, whereas Len did not induce ISGs in GCB lines.The relationship between Aiolos and Ikaros protein expression and DLBCL cell of origin (COO) was further explored in primary DLBCL tissues. A tissue microarray containing 90 DLBCL patient cases immunohistochemically stained for Aiolos, Ikaros and COO markers (Hans) demonstrated Aiolos and Ikaros expression across GCB and non-GCB subtypes, suggesting that CC-122 may have broad-spectrum activity independent of COO. In fact, Aiolos and Ikaros were degraded in lymph node biopsies from both GCB and non-GCB DLBCL patients administered CC-122. The predictive value of Aiolos and Ikaros protein levels for clinical response will be assessed in the Phase I study.Conclusion: Aiolos degradation-dependent upregulation of ISGs represents a novel mechanism for the activity of CC-122. This mechanism bypasses the interferon-signaling pathway and occurs across cell of origin subtypes, suggesting this is a common mechanism accounting for the broad-spectrum activity of CC-122. Understanding of this mechanism has potential to greatly enhance the clinical development of CC-122 in DLBCL by guiding patient selection strategies and rational combination therapies. DisclosuresHagner:Celgene Corp: Employment, Equity Ownership. Wang:Celgene Corp: Employment, Equity Ownership. Couto:Celgene Corporation: Employment, Equity Ownership. Breider:Celgene Corp: Employment. Fontanillo:Celgene Corp: Employment, Equity Ownership. Trotter:Celgene Corporation: Employment, Equity Ownership. Bjorklund:Celgene Corp: Employment, Equity Ownership. Havens:Celgene Corporation: Employment, Equity Ownership, Patents & Royalties. Raymon:Celgene Corporation: Employment, Equity Ownership. Narla:Celgene Corporation: Employment, Equity Ownership. Barnes:Celgene Corporation: Employment, Equity Ownership. Waldman:Celgene Corporation: Employment, Equity Ownership. Daniel:Celgene Corp: Employment, Equity Ownership. Klippel:Celgene Corp: Employment. Pourdehnad:Celgene: Employment, Equity Ownership. Thakurta:Celgene Corp: Employment, Equity Ownership. Chopra:Celgene Corporation: Employment, Equity Ownership. Gandhi:Celgene Corp: Employment, Equity Ownership.

The Impact of Concurrent MYC BCL2 Protein Expression on the Risk of Secondary Central Nervous System Relapse in Diffuse Large B-Cell Lymphoma (DLBCL)

Introduction: Recent studies have established that concurrent MYC and BCL2 protein expression by immunohistochemistry (IHC) identifies a subgroup of patients with diffuse large B-cell lymphoma (DLBCL) with a poor outcome. Classic dual translocation MYC/ BCL2, so called 'double hit’ disease, is associated with a high risk of central nervous system (CNS) relapse; however the impact of concurrent MYC and BCL2 protein expression on the risk of CNS relapse remains unknown. Further, robust biological markers that accurately predict the risk of CNS relapse in DLBCL would also be of value in clinical practice.Methods: Cases of pre-treatment formalin fixed paraffin embedded DLBCL in two tissue microarrays were independently scored by two expert hematopathologist (GWS and KLT or PF and AM) for expression of MYC (Epitomics Y69), BCL2 (Dako 124), CD10, BCL6 and MUM1 by IHC. MYC and BCL2 positivity were defined as ≥ 40% and ≥ 50% cells with staining, respectively, in accordance with previously established cutoffs (Johnson, JCO 2012; 30). Cases with discordant scores were reviewed by a third hematopathologist (RDG) to reach a consensus. Cell of origin (COO) was assigned according to the Hans IHC algorithm (Hans, Blood 103: 2004) as well as by the recently described gene expression profiling Lymph2Cx 20 gene assay based on NanoString technology (Scott, Blood 2014; 123) in the subset of patients with ≥ 40% tumor content. Patients treated with at least one cycle of R-CHOP chemotherapy with curative intent were included and those with established CNS disease at diagnosis were excluded.Results: 447 patients were identified with the following baseline clinical characteristics: Median age 65 y (16-92y); males n=280, 63%; performance status ≥ 2, n= 147, 33%; stage 3 or 4 disease n=242, 54%; elevated LDH n=219, 47%; EN > 1 n= 80, 17%. With a median follow-up of 6.75 years for living patients, the 3 year time to progression, progression-free and overall survival for all patients were 68%, 66%, and 73%, respectively. In total, 131 (29%) were MYC+BCL2+ and 316 (71%) were non-MYC+BCL2+. By COO assignment using the Hans algorithm (n=444), 192 were non-GCB (43%) and 252 were GCB (57%) and by the Lymph2Cx (n=308); 103 were ABC (33%), 172 were GCB (56%) and 33 (11%) were unclassifiable. The 2 year cumulative risk of CNS relapse for the whole cohort was 4.3%. The cumulative risk of CNS relapse was higher in cases that were MYC+BCL2+ (2 year risk 9.4% vs 2.4%, P=0.001) with similar results obtained if classic MYC+BCL2+ double hit cases are excluded. There were no cases of CNS relapse in cases MYC+ alone by IHC. By COO, patients with a non-GCB phenotype by the Hans algorithm had an increased risk of CNS relapse (2 year risk 6.9% vs 2.6%, P=0.03) and similarly, cases assigned as ABC DLBCL by the Lymph2Cx assay also identified a group with a higher risk of CNS relapse compared to GCB cases (9.5% vs 2.5%, P=0.03) (Figure 1). In Cox regression multivariate analysis including the COO (Hans), IPI group (0/1 vs 2/3 vs 4/5) and MYC/BCL2 IHC, only the IPI (HR 2.18, P=0.02) and MYC+BCL2+ IHC (HR=3.76, P=0.007) were associated with an increased risk of CNS relapse. Similar results were obtained using the Lymph2Cx COO designation. Within the IPI risk groups, MYC+BCL2+ status further stratified patients in the intermediate risk group (IPI 2 or 3, n=206) into a higher risk group (2 year CNS relapse 12.6%) and a low risk group (2 year CNS relapse 2.9%) (P=0.01). A similar trend was observed in the high IPI risk group (IPI 4 or 5, n=86, 2 year CNS relapse MYC+BCL2+ 17.2% vs 4.7%, P=.0.18) but it was not useful in the low IPI risk group (IP1 0 or 1 (n=155), 2 year CNS relapse 4% vs 1%, P=0.39) where the overall risk was low. Within the COO subgroups, MYC+BCL2+ status also defined a group at high cumulative risk of CNS relapse within the non-GCB subtype (12.9% vs 3%, P=0.001) and by the Lymph2Cx defined ABC subtype (16.9% vs 2.2%, P= 0.03) and a trend was observed for GCB defined by Lymph2Cx (6.6% vs 1.5%. P=.08) but not by Hans criteria (P=0.40).Conclusion: Concurrent expression of MYC and BCL2 protein in DLBCL defines a group of patients at high risk of CNS relapse, independent of the IPI and COO. MYC+BCL2+ status may help to further risk stratify patients in the intermediate and high IPI risk groups and within the ABC subtype to identify patients who should undergo additional diagnostic testing and in whom to explore the effectiveness of prophylactic CNS strategies. [Display omitted] DisclosuresSavage:F Hoffmann-La Roche: Other. Sehn:Roche: Research Funding. Connors:Seattle Genetics, Inc.: Research Funding; Roche: Research Funding. Gascoyne:Hoffman La-Roche: Research Funding.

The Impact of Immunophenotypic Subtypes and Treatment Regimens on Patient Outcomes in Monomorphic Post-Transplant Lymphoproliferative Disorders (Diffuse Large B-Cell Lymphoma)

Introduction: Post-Transplant Lymphoproliferative Disorders (PTLD) are a group of heterogeneous disorders that arise as a consequence of iatrogenic immunosuppression for solid organ or allogeneic bone marrow/stem cell transplantation. Though they all arise in a common clinical context, different types of PTLD differ with respect to their underlying biology, clinical presentation and treatment. The aims of this study were to: (1) Define the cell of origin (COO) of monomorphic Diffuse Large B-cell Lymphoma (DLBCL) PTLD and evaluate their impact on clinical presentation and survival and (2) assess the impact of different Rituximab containing treatment regimens on survival outcomes in monomorphic DLBCL PTLD patients.Methods: We conducted a retrospective review of our institutional databases to identify all the cases of monomorphic PTLD (DLBCL) diagnosed and treated at our medical center from 2000-2013. COO classification into germinal center B-cell like (GCB) and non-germinal center B-cell like (non-GCB) type was performed by immunohistochemistry using the Hans algorithm.Results: Cell of origin: 40 cases of monomorphic PTLD (DLBCL) were diagnosed during the study interval. Tissue material for COO subtyping was available for 25 patients. By immunohistochemistry 16/25 (64%) were non-GCB and 9/25 (36%) were GCB subtype, median age of presentation being 46 years (range 3-75) and 48 years (range 3-64), respectively. A trend towards EBV positivity (by in situ hybridization for EBV encoded RNA) was noted in the non-GCB group (75% vs. 33%) [p=0.09]. Non-GCB DLBCL PTLD presented earlier post-transplant at a median 1.5 years (range 0.2-15) vs. 3.9 years (range 0.7-17) for GCB cases. When comparing immunosuppressive therapy at the time of PTLD presentation, an association between Tacrolimus therapy and non-GCB phenotype was identified [p=0.03]. Non-GCB DLBCL PTLD demonstrated a trend towards higher rates of extra nodal involvement (88% vs. 44%) [p=0.06] and advanced stage disease (Stage III/IV 75% vs. 33%) [p=0.09]. No significant differences in organ transplanted, LDH, ECOG performance and IPI were observed. While acknowledging the heterogeneity of therapies administered, no significant differences in Progression Free Survival (PFS) (median PFS non-GCB = 17 months vs. GCB = 15 months [p=0.36]) and Overall Survival (OS) (median OS non-GCB = 33 months vs. GCB = 27 months [p=0.22]) were identified.Impact of Treatment: 35 adults (age≥18) were treated at our center. The four most common first line therapies administered were R-CHOP (14), R-EPOCH (7), Palliative Care (5) and Rituximab monotherapy (4). Five patients were given 4 other different therapies. In patients given Rituximab monotherapy, two patients presenting with stage I disease responded while two with stage IV disease progressed. When focusing on patients who commenced R-CHOP or R-EPOCH as their initial therapy, no significant differences in age, stage, LDH, extra nodal disease, ECOG performance status, IPI and immunosuppression therapy was identified between the groups. The complete response (CR) rate for R-CHOP was 50% vs. 71% for R-EPOCH. Primary refractory disease was present in 29% of patients receiving R-CHOP vs. 14% with R-EPOCH. Death during first line therapy occurred equally in both groups (14%). All four primary refractory disease patients in the R-CHOP arm died, while the one patient who was primary refractory to R-EPOCH is alive 3 years post autologous stem cell transplant. R-EPOCH demonstrated prolonged PFS (median PFS R-CHOP = 15 months vs. R-EPOCH not reached [p=0.049]) and prolonged OS (Figure 1) [p=0.036].Conclusions: (1) In monomorphic PTLD (DLBCL), the non-GCB subtype predominates and is associated with the use of Tacrolimus. It commonly presents with advanced stage disease and extra nodal involvement however, no difference in PFS and OS was noted when compared to GCB DLBCL PTLD. (2) R-EPOCH demonstrated prolonged PFS and OS when compared to R-CHOP. The survival differences reflect the higher rates of primary refractory disease in the R-CHOP group and the inability to salvage patients once they become relapse/refractory. Given the retrospective nature of our analysis, further studies in a larger cohort of patients are ongoing to validate these results. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Downregulated expression of Dicer1 predicts inferior survival in primary gastrointestinal diffuse large B-cell lymphoma treated with CHOP-like regimen and rituximab

The aim of this study was to detect the expression levels of Dicer1, Drosha, DGCR8, and Ago2 messenger ribonucleic acids (mRNAs) in patients with primary gastrointestinal diffuse large B-cell lymphoma (PGI-DLBCL) and determine their associations with clinical parameters and prognostic significance. The mRNA level expressions of Dicer1, Drosha, DGCR8, and Ago2 were detected by real-time quantitative polymerase chain reaction. Immunohistochemical staining of CD10, BCL6, and MUM1 was performed using EnVision™ system. The clinicopathologic features and follow-up data were analyzed using Kaplan-Meier estimator. The results show that the expression of Dicer1 (P=0.001), Drosha (P=0.01), DGCR8 (P=0.02), and Ago2 (P=0.002) mRNAs in cancer tissues of patients with PGI-DLBCL was significantly lower than those in normal tissues of healthy controls. Among the expression of CD10, BCL6, and MUM1, 27.4% (17/62) of the patients belonged to the germinal center B-cell (GCB) subtype and 72.6% (45/62) belonged to the non-GCB subtype. Dicer1 expression was significantly decreased in the non-GCB subgroup (P=0.02) and in the high International Prognostic Index (3-5 score) subgroup (P=0.03). Kaplan-Meier analysis showed that the low-Dicer1 subgroup had a shorter overall survival (P=0.02) and shorter progression-free survival (P=0.015) than the high-Dicer1 subgroup. Multivariate analysis identified Dicer1 as an independent prognostic factor in PGI-DLBCL. In Conclusion, Dicer1, Drosha, DGCR8, and Ago2 play key roles in the pathogenesis of PGI-DLBCL. Dicer1, an independent prognostic factor for predicting shortened survival of patients with PGI-DLBCL, can be used as a biomarker to guide the prognosis.

High Concordance of Gene Expression Profiling–correlated Immunohistochemistry Algorithms in Diffuse Large B-cell Lymphoma, Not Otherwise Specified

Diffuse large B-cell lymphoma (DLBCL) is classified into prognostically distinct germinal center B-cell (GCB) and activated B-cell subtypes by gene expression profiling (GEP). Recent reports suggest the role of GEP subtypes in targeted therapy. Immunohistochemistry (IHC) algorithms have been proposed as surrogates of GEP, but their utility remains controversial. Using microarray, we examined the concordance of 4 GEP-correlated and 2 non-GEP-correlated IHC algorithms in 381 DLBCLs, not otherwise specified. Subtypes and variants of DLBCL were excluded to minimize the possible confounding effect on prognosis and phenotype. Survival was analyzed in 138 cyclophosphamide, adriamycin, vincristine, and prednisone (CHOP)-treated and 147 rituximab plus CHOP (R-CHOP)-treated patients. Of the GEP-correlated algorithms, high concordance was observed among Hans, Choi, and Visco-Young algorithms (total concordance, 87.1%; κ score: 0.726 to 0.889), whereas Tally algorithm exhibited slightly lower concordance (total concordance 77.4%; κ score: 0.502 to 0.643). Two non-GEP-correlated algorithms (Muris and Nyman) exhibited poor concordance. Compared with the Western data, incidence of the non-GCB subtype was higher in all algorithms. Univariate analysis showed prognostic significance for Hans, Choi, and Visco-Young algorithms and BCL6, GCET1, LMO2, and BCL2 in CHOP-treated patients. On multivariate analysis, Hans algorithm retained its prognostic significance. By contrast, neither the algorithms nor individual antigens predicted survival in R-CHOP treatment. The high concordance among GEP-correlated algorithms suggests their usefulness as reliable discriminators of molecular subtype in DLBCL, not otherwise specified. Our study also indicates that prognostic significance of IHC algorithms may be limited in R-CHOP-treated Asian patients because of the predominance of the non-GCB type.

Diffuse large B-cell lymphoma with histone H3 trimethylation at lysine 27: another poor prognostic phenotype independent of c-Myc/Bcl2 coexpression.

Deregulation of histone H3 trimethylation at lysine 27 (H3K27me3) via aberration of the histone methyltransferase, enhancer of zeste homologue 2 (EZH2), is suggested to play a critical role in cancers including hematologic malignancies. In the present study, implications of H3K27me3 were investigated in diffuse large B-cell lymphoma (DLBCL) with respect to clinicopathological factors, especially in association with c-Myc/Bcl2 coexpression and germinal center B-like (GCB) or non-GCB subtype. By immunohistochemistry, a high level of H3K27me3 was observed in approximately one-third (35.3%, 79/224) of DLBCL cases, and this subset of cases was related to poor performance status (Eastern Cooperative Oncology Group scores ≥ 2) (P = .013), elevated lactate dehydrogenase level (P = .001), and a higher international prognostic index risk group (scores ≥3) (P = .005). H3K27me3 level was significantly correlated with EZH2 expression (P = .004) and c-Myc protein expression (P = .003) but not correlated with c-Myc/Bcl2 coexpression or with GCB or non-GCB subtype. A high level of H3K27me3 was related to an inferior overall survival (P = .006) and was shown to be an independent prognostic factor for overall survival along with the higher international prognostic index risk group and c-Myc/Bcl2 coexpression. In conclusion, H3K27me3 was related to EZH2 and c-Myc expression, suggesting formation of a MYC-EZH2-H3K27me3 loop in a subgroup of DLBCL cases. H3K27me3 was associated with poor patient outcome and revealed as an independent predictor for overall survival of DLBCL patients. H3K27me3 in DLBCL may be another high-risk phenotype independent of the phenotype of c-Myc/Bcl2 coexpression or other known poor prognostic subgroups.

PROGNOSTIC FACTORS FOR PRIMARY CENTRAL NERVOUS SYSTEM LYMPHOMAS TREATED WITH HIGH-DOSE METHOTREXATE-BASED CHEMO-RADIOTHERAPY

BACKGROUND: Chemotherapy with high-dose methotrexate (HD-MTX) followed by whole brain radiotherapy (WBRT) is a conventional approach to treat primary central nervous system lymphomas (PCNSL), but some tumors relapse early leading to unfavorable outcome. Several biomarkers have been identified as prognostic factors in PCNSL, however, the correlation of both clinical factors including those related to MTX metabolism and B-cell differentiation and oncogenic biomarkers with response to and outcome by therapy is yet unclear. METHODS: We investigated 32 immunocompetent patients (19 males, 13 females) with PCNSL (all diffuse large B-cell type) treated with HD-MTX based therapy with or without WBRT since 2000 in our institution. Paraffin-embedded formalin-fixed tumor tissue sections were stained immunohistochemically with antibodies against following factors: B-cell differentiation markers (CD10, Bcl-6, Mum-1, CD138); MTX metabolism-related (MRP family, LRP, DHFR); cell cycle-related (p27KIP1, MIB-1); drug resistance-related (MGMT, MLH1, MSH2, MSH6, PMS2); and oncogenes (Myc, Bcl-2). Correlation between positivity of these factors and clinical outcomes were evaluated using logrank test and cox regression analysis. RESULTS: Among these factors, complete response to HD-MTX was significantly associated with longer progression-free survival (PFS)(P = 0.0012), while Bcl-6 expression as well as histological subtype (non-germinal center B-cell, non-GCB) was closely correlated with shorter PFS. Age (>60) (P = 0.006) and MSH2 expression (P = 0.017) were found to be better predictor for overall survival (OS), but in multivariate analysis, they were no longer significant. Other factors involved in MTX metabolism, DNA repair enzymes, and oncogenes did not affect outcome. CONCLUSIONS: Non-GCB subtype and Bcl-6 expression may be associated with worse outcome in patients with PCNSL treated with HD-MTX, while MTX-metabolism related factors did not influence prognosis. Further investigation is needed to assess Bcl-6 as a potential prognostic factor in PCNSL. SECONDARY CATEGORY: Clinical Neuro-Oncology.

Immunohistochemical and molecular characteristics with prognostic significance in diffuse large B-cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is an aggressive non-Hodgkin lymphoma with marked biologic heterogeneity. We analyzed 100 cases of DLBCL to evaluate the prognostic value of immunohistochemical markers derived from the gene expression profiling-defined cell origin signature, including MYC, BCL2, BCL6, and FOXP1 protein expression. We also investigated genetic alterations in BCL2, BCL6, MYC and FOXP1 using fluorescence in situ hybridization and assessed their prognostic significance. BCL6 rearrangements were detected in 29% of cases, and BCL6 gene alteration (rearrangement and/or amplification) was associated with the non-germinal center B subtype (non-GCB). BCL2 translocation was associated with the GCB phenotype, and BCL2 protein expression was associated with the translocation and/or amplification of 18q21. MYC rearrangements were detected in 15% of cases, and MYC protein expression was observed in 29% of cases. FOXP1 expression, mainly of the non-GCB subtype, was demonstrated in 37% of cases. Co-expression of the MYC and BCL2 proteins, with non-GCB subtype predominance, was observed in 21% of cases. We detected an association between high FOXP1 expression and a high proliferation rate as well as a significant positive correlation between MYC overexpression and FOXP1 overexpression. MYC, BCL2 and FOXP1 expression were significant predictors of overall survival. The co-expression of MYC and BCL2 confers a poorer clinical outcome than MYC or BCL2 expression alone, whereas cases negative for both markers had the best outcomes. Our study confirms that DLBCL, characterized by the co-expression of MYC and BCL2 proteins, has a poor prognosis and establishes a significant positive correlation with MYC and FOXP1 over-expression in this entity.

Effect of lenalidomide combined with R-CHOP (R2CHOP) on negative prognostic impact of nongerminal center (non-GCB) phenotype in newly diagnosed diffuse large B-cell lymphoma: A phase 2 study.

8520 Background: The non-germinal center B-cell like (non-GCB) subtype of diffuse large cell lymphoma (DLBCL) is associated with a worse outcome when treated with RCHOP chemotherapy. Lenalidomide h...

Clinicopathological characteristics and prognostic analysis of 92 cases with primary gastrointestinal diffuse large B-cell lymphoma

To analyze the clinical and pathological features, molecular biological markers and prognosis of primary gastrointestinal diffuse large B-cell lymphoma (DLBCL). A retrospective study was conducted in 92 cases of primary gastrointestinal DLBCL. The data of clinical characteristics, pathological and immunohistochemical features were analyzed. The relationship between different factors at diagnosis and prognosis were studied. Of the 92 patients, the male-female ratio was 1.56:1 with a median age of 59(7-89) years. The most frequently involved site was stomach (50.0%), followed by small intestine (15.2%), colon (12.0%), ileocecum (10.9%) and multiple gastrointestinal tract involvements (9.8%). The ratio of non-germinal center B cell (non-GCB) subtype to germinal center B cell (GCB) was 2:1. Among the cases, Ki-67 had a high level expression with a median positive rate >80%. Positive expression of Bcl-2 was detected in 52.1% cases (25/48). Evidence of Helicobacter pylori (Hp) infections was detected in 34.0% cases with stomach involvement. Of the patients, 75 were included in a follow-up study with a median time of 48 months (3-130 months). The complete remission(CR)rate was 66.7%(50/75),the partial remission (PR) rate was 22.7% (17/75) and the overall response rate was 89.4% (67/75). The 1-, 3- and 5- year survival rates of the patients were 85.3%, 66.2% and 60.5%, respectively. The incidence of secondary malignancy after chemotherapy was 2.7%. The multivariate analysis indicated that primary colic DLBCL or multiple involvements of gastrointestinal tract and low serum albumin level (<35 g/L) at diagnosis were independently associated with poor outcome. Patients who received chemotherapy combined with rituximab (43 cases) had a higher CR rate (79.1% vs 46.9%, P=0.008) and 5-year overall survival (77.5% vs 53.1%, P=0.0045) than those without rituximab (32 case). Primary gastrointestinal DLBCL was a highly invasive and heterogeneous malignancy. In our data, the proportion of non-GCB subtype was higher. Primary colic DLBCL and those with multiple involvements of gastrointestinal tract had poor survival. Low serum albumin at diagnosis indicated poor outcome. Rituximab treatment in addition to chemotherapy might help to improve the clinical outcome. Further prospective trails were needed to confirm our results.

The biology of diffuse large b cell lymphoma (DLBCL)

Diffuse large B cell lymphoma represents the most common non-Hodgkin lymphoma, accounting for 30-40% of all lymphomas world-wide. Patients with DLBCL show marked morphologic, phenotypic, genetic and clinical heterogeneity. The current standard of care, R-CHOP, produces cures in ~65-70% of patients. A subset of the 30-35% of patients whose primary therapy fails are defined as primary refractory disease; a finding that is associated with particularly dismal outcome. Gene expression profiling (GEP) studies have defined 3 major cell-of-origin (COO) sub-groups of DLBCL including a germinal centre B cell (GCB) type an activated B cell (ABC) type and primary mediastinal B cell lymphoma (PMBCL). Within DLBCL, binary immunohistochemical algorithms have been used to translate these GEP data into GCB and non-GCB subtypes with variable success. Mutational studies using next-generation sequencing have added texture to these data, with some mutations recognised as COO specific. Important oncogenic pathways in DLBCL have been described and include alteration of normal apoptotic signalling, constitutive NF-kB activation, altered histone modification, abnormal B cell receptor signalling, oncogenic events conferring a proliferative advantage, blocks to terminal differentiation and immune escape. All of these mechanisms will be discussed with a focus on the molecular events that underlie treatment failure.

Primary Malignant Lymphoma of Prostate with Silent &amp;lt;i&amp;gt;MYD&amp;lt;/i&amp;gt;88 Mutation

We present a case of primary malignant lymphoma of the prostate in a 77-year-old Japanese man. Immunohistochemical examinations revealed the presence of a non-GCB subtype of DLBCL (CD10 (-), Bcl-6 (-), MUM1 (+)), and genetic analysis disclosed a lack of typical codon 206 or 265 missense mutation in MYD88, suggesting that the case was of type3 (non-GCB andnon-ABC), a subtype of DLBCL. Three courses of chemotherapy (rituximab in combination with cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP)) were effective, and the patient has been free of the disease with no local or systemic recurrence for three years. Although the significance of silent mutation at hot spot of the highly oncogenic MYD88 gene in this case is unclear, a minute increase in translational efficiency by silent mutation may have contributed to the activation of MYD88.

Concurrent Expression Of MYC/BCL2 Protein In Newly Diagnosed DLBCL Is Not Associated With An Inferior Survival Following EPOCH-R Therapy

▪ BackgroundWhile several studies have now demonstrated that patients with MYC-rearranged DLBCL have a worse outcome following treatment with CHOP -with or without rituximab - recent reports suggest that concurrent expression of MYC and BCL2 protein by immunohistochemistry (IHC), independent of a MYC rearrangement, is associated with an inferior clinical outcome following R-CHOP (Johnson et al. JCO 2012 Oct 1; 30 (28): 3452-9). We previously reported that DLBCL patients with a MYC rearrangement have a similar outcome to MYC negative cases (4 year EFS of 83% versus 76% respectively: p=0.46) following dose-adjusted (DA)-EPOCH-R (Dunleavy et al. Ann Oncol 2011 (22) Suppl 4 # 71). MethodsTo assess the prognostic role of MYC and BCL2 protein expression, we performed immunohistochemistry for MYC and BCL2 in patients with newly diagnosed DLBCL who received DA-EPOCH-R or short-course (SC)-EPOCH-RR (if HIV infected). Primary mediastinal B-cell lymphoma (PMBL) cases were excluded as they have a different biology. Tumors were scored positive for MYC if ≥ 40% of cells stained positive. For BCL2, the staining intensity was compared with that in control T cells present in the tumor samples – tumor cells were considered positive if they stained the same or more intensely than T cells. We used the Hans algorithm, as previously described, to predict cell of origin as germinal center B-cell like (GCB) or non-GCB type and correlated this with outcome. ResultsOf 66 patients enrolled on study, characteristics were median (range) age: 48 (18-76) years; male sex 47 (71%); IPI score low versus intermediate/high 18 (27%) versus 48 (73%); HIV positive 22 (33%). IHC was positive for MYC in 28/48 (58%) cases and positive for BCL2 in 24/51 (47%). 36/51 (71%) and 15/51 (29%) of cases were of GCB and non-GCB origin respectively. At a median follow-up time of greater than 10 years, progression-free survival (PFS) and overall survival (OS) were 67.5% and 75% for all patients. We compared survival in the 4 groups: MYC+/BCL2+, MYC+/BCL2-, MYC-/BCL2+ and MYC-/BCL2-. PFS and OS were not significantly inferior in any group (global p value=0.5 (PFS) and 0.8 (OS)). Cell of origin did however predict outcome and at 10 years follow-up, PFS was 78% (GCB) versus 43% (non-GCB) (p=0.016) and OS 80% (GCB) versus 65% (non-GCB) (p=0.24). There was a significant association between MYC+/BCL2+ cases and non-GCB subtype (p=0.01) but as 42% were of GCB origin, MYC+/BCL2+ status alone did not predict a poor outcome. ConclusionsIn DLBCL patients treated with DA-EPOCH-R and SC-EPOCH-RR, concurrent expression of MYC and BCL2 protein did not correspond with a worse clinical outcome. However, cell of origin (GCB versus non-GCB) was predictive of outcome. MYC+/BCL2+ cases segregate with the non-GCB subtype. The overall survival of PMBL cases was 97% at 5 years follow-up. We are currently prospectively studying the DA-EPOCH-R regimen in MYC-rearranged DLBCL in a multicenter study (NCT01092182). Disclosures:No relevant conflicts of interest to declare.

Clinical Outcome Of R-CHOP Or CHOP Regimen For Elderly Patients With Diffuse Large B-Cell Lymphoma In China：A Single Center's Experience

BackgroundDiffuse large-B-cell lymphoma (DLBCL) is the most frequent type of lymphoma which accounts for about 40 percent of new cases of lymphoma and more than 80 percent of aggressive lymphomas. It is reported to represent approximately 60 percent of all B-cell lymphomas in Eastern Asia and more than half of patients with DLBCL were found over 60 years old. The current first-line therapy is cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) combined with Rituximab. However, positive effects of Rituximab use in the elderly were not certain before the year 2002 when clinical trials proved that it was more effective with Rituximab. However, the addition of Rituximab has brought more toxicities, thus a better medical care to prevent toxicities and weighing over toxicities and benefits should be carefully done before initiation of the RCHOP regimen in the elderly. It has been recognized now that DLBCL is a heterogeneous disease with different molecular characteristics, such as the germinal center B-cell (GCB) and non-GCB subtypes. Though both commonly seen in both Asia and the West, DLBCL has been reported to differ in some molecular characteristics. Studies revealed differences in the proportion of GCB and non-GCB DLBCL between Asian and Western patients. Whether this difference contributes to clinical treatment and response to the current RCHOP regimen still remains controversial. An overall similar outcome was observed in DLBCL elderly patients from both East Asia and West treated with RCHOP. However, more data is needed to determine the GC proportion, the benefits and toxicity files of RCHOP in Asian population. Materials and MethodsA total of 84 elderly DLBCL patients who are 60 years old or older admitted to our center between 2009 and 2013 were included. Among them, 48 received CHOP with Rituximab and 36 received CHOP only chemotherapy due to economic reasons. No significant differences in gender, age distribution were found between the two groups. We stopped follow-up of these patients till May 2013. Overall survival time (OS), progression free survival time (PFS), GCB subtypes and toxicity files after chemotherapy were recorded. To better assess the stratification of IPI, we grouped these patients based on IPI score in two: IPI 0-1; 2; 3; 4-5 and IPI 0-2; 3-5. Cox-regression was performed for GCB subtypes, chemotherapy, different IPI groups, extranodal involvement with OS or PFS using SPSS 16.0 software. Kaplan-Meier survival curves also plotted between different factors with PFS or OS. P value less than 0.05 was considered significant. ResultsIn the four-group IPI scoring stratification system, the Cox regression model demonstrated a longer OS and PFS in Chinese elderly DLBCL patients treated with RCHOP than those receiving CHOP alone (p=0.014 and p<0.001). OS did not differ with GCB subtypes, extranodal involvement and the four IPI groups. PFS did not show difference with GCB subtypes, extranodal involvement, however, demonstrated an advantage in IPI group 1 (IPI=0 or 1) and group 2 (IPI=2). In the two-group IPI scoring stratification system, results showed a longer OS and PFS in patients treated with RCHOP (p=0.007 and p<0.001) and in patients with an IPI score less than 3 (p=0.002 and p=0.008). OS and PFS still did not differ with GCB subtypes, extranodal involvement. The estimated OS in these Chinese elderly DLBCL patients were 34.2 and 47.2 months in CHOP only group and RCHOP group while the PFS were 17.1 and 33.8 months respectively. 56% of the patients receiving RCHOP relapsed, but only 33% remission rate in CHOP only patients (p=0.048). Toxicities are more commonly seen in patients receiving RCHOP (p=0.013) including febrile neutropenia, infections, interstitial pneumonia and cardiomyopathy. Two patients receiving RCHOP even died with severe infection. ConclusionIn the Chinese elderly patients diagnosed with DLBCL, IPI score can predict the OS and PFS when it is stratified into two group (0-2; 3-5). RCHOP use achieved longer OS and PFS than CHOP only regimen. No significant differences were observed in whether the patients have extranodal involvement or GCB subtype. However, Rituximab brought an increased risk of toxicities. In conclusion, Chinese elderly DLBCL patients achieved a longer OS and PFS with RCHOP than CHOP only regimen. However, clinicians should be aware of the toxicities which came along. Disclosures:No relevant conflicts of interest to declare.

Distinct CD30 Expression Patterns In Germinal Center B-Cell (GCB) and Non-GCB Diffuse Large B-Cell Lymphoma (DLBCL)

BackgroundDLBCL is the most common type of non-Hodgkin lymphoma, which demonstrates morphologic, immunophenotypic, molecular, and clinical heterogeneity. Gene expression profiling studies define two molecular subtypes of DLBCL, namely germinal center B-cell-like (GCB) and activated B-cell-like (ABC) DLBCL. Hans' algorithm was developed to provide an immunohistochemical correlation to the molecular subtypes of DLBCL. In the pre and post-rituximab era, ABC subtype of DLBCL is known to demonstrate poor overall survival when compared to GCB subtype. In addition, relapsed or primary refractory DLBCL responds poorly to current therapeutic strategies. These findings underscore the necessity to identify novel therapeutic targets in DLBCL to achieve better clinical outcomes. Recent data suggests that CD30, a member of the tumor necrosis factor receptor family, is a potential therapeutic target in DLBCL. CD30 is widely expressed in classical Hodgkin lymphoma (cHL) and anaplastic large cell lymphoma (ALCL) thus serving as an attractive target of immunotherapy for cHL and ALCL. In this study we conducted a retrospective evaluation of CD30 expression in GCB and non-GCB DLBCL, treated at our Institution with standard front-line chemo-immunotherapy (i.e. R+CHOP or R+ DA-EPOCH). Materials and MethodsWe identified 60 patients with confirmed DLBCL for which archived formalin fixed paraffin embedded tissue was available in the form of a tissue micro-array (39 Cases) or diagnostic biopsy material (21 Cases). Immunohistochemical detection of CD30 was performed using routine methods (Biocare #PM031, ready to use aliquots). Demographic, clinical and pharmacological parameters were obtained for each patient. DLBCL cases were subtyped as GCB or non-GCB using immunohistochemistry-based Hans' algorithm. Membranous and Golgi pattern of CD30 expression in the tumor cells was designated as positive. Results21 patients (48.8%) were classified as GCB, 22 patients (51.2%) non-GCB. CD30 expression was detected in 13.3% of all DLBCL patients. Differences in CD30 expression were noted between GCB- and non-GCB DLBCL. CD30 expression was detected in 9.5% and 23% of the GCB- and non-GCB DLBCL subtypes respectively. Demographics and clinical characteristics were equally distributed between GCB- and non-GCB and between CD30 positive or negative DLBCL. Patients received either R+CHOP (82%) or R+DA-EPOCH (18%). The complete response (CR) rate of the entire cohort was 67% and no differences were observed between GCB- or non-GCB DLBCL. CD30 (+) DLBCL had a higher CR (complete response) rate than CD30 (-) DLBCL (88% vs 63.4%). However, the numbers were too small to reach statistical significance. No significant differences were observed between CD30 (+) or (-) DLBCL in terms of progression free survival (PFS) (CD30[-]37m vs. CD30[+]16.5m, P =0.785) or overall survival (OS) (CD30[-]86m vs. CD30[+]57.4m, P =0.99). In contrast to previously reported by other investigators, there was no difference in the clinical outcome between GCB vs. non-GCB DLBCL treated with R+CHOP or R+DA-EPOCH. ConclusionsOur data suggests that CD30 expression is more prevalent in non-GCB DLBCL patients based on our small cohort. While CD30 expression may not confer a prognostic value in newly diagnosed DLBCL (Table 1), routine testing for it may identify a group of patients that may benefit from CD30-targeted therapeutic strategies (i.e. antibody-drug conjugates) in the relapsed/refractory setting. (Research, in part, supported by The Eugene and Connie Corasanti Lymphoma Research Fund) Disclosures:Czuczman:Genetech, Onyx, Celgene, Astellas, Millennium, Mundipharma: Advisory Committees Other. Hernandez-Ilizaliturri:Seattle Genetics: Consultancy, Honoraria.

Immunophenotyping and molecular genetic analysis of diffuse large B-cell lymphoma

To provide genetic diagnosis and counseling for patients from two families affected with X-linked hypohidrotic ectodermal dysplasia. Potential mutation of the ED1 gene was screened by DNA sequencing. For family 1, multiplex ligation-dependent probe amplification (MLPA) analysis and haplotyping of ED1 gene were also carried out for prenatal diagnosis. For the patient from family 1, deletion of the exon 1 of the ED1 gene and 2 short tandem repeat(STR) sites (DXS8269 and DXS1422) were detected. His daughter was carrier of the deletion. Upon prenatal diagnosis, the fetus was confirmed to be a normal male, for whom the haplotype of ED1 gene has differed from that of the proband. In family 2, a c.463C>T mutation in exon 3 of the ED1 gene was detected in the proband, whose mother was heterozygous for the same mutation. The deletion (exon 1) and missense (R155C) mutation in ED1 gene have probably underlied the disease in the two families. During prenatal diagnosis, it may be necessary to obtain precise results through combining mutation detection and haplotype analysis of the ED1 gene.

Certification and role of local pathologists for diffuse large B-cell lymphoma (DLBCL) subtyping and eligibility determination in the phase II PYRAMID study.

8559 Background: The role of local pathologists in promoting patient (pt) accrual and evaluating eligibility criteria involving a complicated immunohistochemical (IHC) algorithm has rarely been investigated. The phase II PYRAMID trial (NCT00931918) is assessing R-CHOP ± bortezomib for newly diagnosed pts with non-germinal-center B-cell (non-GCB) subtype DLBCL. In this trial, local pathologists were encouraged to perform DLBCL subtyping at the point of biopsy, identify suitable pts for the trial, and facilitate accrual. Methods: Determination of GCB vs non-GCB subtype is per the Hans method, an algorithm based on IHC for CD10, BCL-6, and IRF4/MUM1. In stage 1, pathologists demonstrated IHC subtyping proficiency by evaluating a tissue microarray (TMA) of 12 DLBCL cases; those with ≥80% of samples in agreement with central lab results were certified for determining trial eligibility. In stage 2, to broaden participation, pathologist certification occurred via teleconference outlining trial eligibility criteria, tissue subtyping requirements, and determining pathologists’ experience with the Hans method. Results: 182 pathologists have been certified for local subtyping, 50 via TMA and 132 by teleconference. 66/88 active study sites have ≥1 certified pathologist. Only 1 of the 10 top enrolling sites lacks a certified pathologist. 52% (84/162) of pts have been enrolled based on local pathologist subtyping prior to central lab confirmation. Discordance with central lab results occurred in 9/84 cases (11%). Enrollment rates pre- and post-local pathologist certification were 0.053 and 0.096 pts/site/month; an improvement of 81%. Trial accrual correlates with the presence of a certified local pathologist (p=0.0026). The rate of ineligible GCB cases sent for central lab testing was lower from sites with a certified pathologist (23% [69/299 cases] vs 38% [40/106 cases] for sites without). Conclusions: Engagement of local pathologists in trials requiring pathology selection can significantly improve accrual. This study demonstrates the effectiveness of various training modalities in improving selection by local pathologists using a complex IHC algorithm. Clinical trial information: NCT00931918.

Germinal-center type B-cell classification and clinical characteristics of Chinese pediatric diffuse large B-cell lymphoma: a report of 76 cases

Pediatric diffuse large B-cell lymphoma (DLBCL) is a highly aggressive disease with unique clinical characteristics. This study analyzed the germinal-center type B-cell (GCB) classification and clinical characteristics of Chinese pediatric DLBCL. A total of 76 patients with DLBCL newly diagnosed in Sun Yat-sen University Cancer Center between February 2000 and May 2011, with an age younger than 18 years, were included in the analysis. The male/female ratio was 3.47:1. The median age was 12 years (range, 2 to 18 years), and 47 (61.8%) patients were at least 10 years old. Of the 76 patients, 48 (63.2%) had stage III/IV disease, 9 (11.8%) had bone marrow involvement, 1 (1.3%) had central nervous system (CNS) involvement, and 5 (6.6%) had bone involvement. The GCB classification was assessed in 45 patients: 26 (57.8%) were classified as GCB subtype, and 19 (42.2%) were classified as non-GCB subtype. The modified B-NHL-BFM-90/95 regimen was administered to 50 patients, and the 4-year event-free survival (EFS) rate was 85.8%. Among these 50 patients, 31 were assessed for the GCB classification: 17 (54.8%) were classified as GCB subtype, with a 4-year EFS rate of 88.2%; 14 (45.2%) were classified as non-GCB subtype, with a 4-year EFS rate of 92.9%. Our data indicate that bone marrow involvement and stage III/IV disease are common in Chinese pediatric DLBCL patients, whereas the percentage of patients with the GCB subtype is similar to that of patients with the non-GCB subtype. The modified B-NHL-BFM-90/95 protocol is an active and effective treatment protocol for Chinese pediatric patients with DLBCL.