Diffuse large B-cell lymphomas in adults with aberrant coexpression of CD10, BCL6, and MUM1 are enriched in IRF4 rearrangements

Diffuse large B-cell lymphoma (DLBCL) with aberrant co-expression of CD10+BCL6+MUM1+ (DLBCL-AE), classified as germinal center B cell (GCB)-type by the Hans algorithm (HA), were genetically characterized. To capture the complexity of these DLBCL-AE, we used an integrated approach including gene expression profiling (GEP), fluorescence in-situ hybridization (FISH), targeted gene sequencing, and copy number (CN) arrays. According to GEP, 32/54 (59%) cases were classified as GCB-DLBCL, 16/54 (30%) as activated B-cell (ABC)-DLBCL and 6/54 (11%) as unclassifiable. The discrepancy between HA and GEP was 41%. Three genetic subgroups were identified. Group 1 included 13/50 (26%) cases without translocations and mainly showing and ABC/MCD molecular profile. Group 2 comprised 11/50 (22%) cases with IRF4 alterations (DLBCL-IRF4), frequent mutations in IRF4 (82%) and NF-?B pathway genes (MYD88, CARD11, and CD79B), and losses of 17p13.2. Five cases each were classified as GCB- or ABC-type. Group 3 included 26/50 (52%) cases with one or several translocations in BCL2/BCL6/MYC/IGH and GCB/EZB molecular profile predominated. Two cases in this latter group showed complex BCL2/BCL6/IRF4 translocations. DLBCL-IRF4 in adults showed a similar CN profile and share recurrent CARD11 and CD79B mutations when compared to LBCL-IRF4 in pediatric population. However, adult cases showed higher genetic complexity, higher mutational load with frequent MYD88 and KMT2D mutations, and more often ABC-GEP. IRF4 mutations were identified only in IRF4-rearranged cases indicating its potential utility in the diagnostic setting. In conclusion, DLBCL-AE are genetically heterogeneous and enriched in cases with IRF4 alterations. DLBCL-IRF4 in adults has many similarities to the pediatric counterpart.Copyright © 2021 American Society of Hematology

Assessment of 2-Year Storage Conditions on Protein, RNA, and DNA in Unstained Human Tissue Sections, Including a Novel Multiplex Digital Gene Expression Profiling Method with Implications for Biobanking.

Background: Formalin-fixed, paraffin-embedded (FFPE) tissues are a valuable resource for clinical and basic science research. Paraffin blocks and the resulting unstained sections (USS) are often stored for years before being used. Previous studies have evaluated the effects of time, temperature, humidity, and inert gases on preservation of USS; however, no study has examined all four variables together. Methods: In the current work, we prospectively and blindly assessed time points from 0 to 24 months, room versus refrigerated temperature, and presence of a desiccant and/or nitrogen atmosphere on a variety of benign and malignant tissues from North America and Africa. End points included immunohistochemistry (IHC), in situ hybridization (ISH), extracted RNA and DNA quantity and quality, and messenger RNA performance in a novel, multiplexed digital gene expression profiling assay of both housekeeping and tumor-specific genes. Results: We found that using current methods of antigen retrieval, staining, and extraction, the end points of IHC, ISH, RNA, and DNA were well preserved under the various conditions tested, with implications that pre-embedding factors contribute to variability in subsequent tissue integrity. We also document that spectrophotometric estimations of nucleic acid concentrations were in general estimated to be higher than with fluorimetric methods, which may be pertinent to end assay development. We further describe a new multiplex assay, the PlexSet digital gene expression assay, suitable for evaluating RNA quality in FFPE tissues. Conclusion: Altogether, these results may provide helpful guidance with regard to approaches for long-term storage conditions for USS

Transcriptional profiles define drug refractory disease in myeloma

Abstract Identifying biomarkers associated with disease progression and drug resistance are important for personalized care. We investigated the expression of 121 curated genes, related to immunomodulatory drugs (IMiDs) and proteasome inhibitors (PIs) responsiveness. We analyzed 28 human multiple myeloma (MM) cell lines with known drug sensitivities and 130 primary MM patient samples collected at different disease stages, including newly diagnosed (ND), on therapy (OT), and relapsed and refractory (RR, collected within 12 months before the patients’ death) timepoints. Our findings led to the identification of a subset of genes linked to clinical drug resistance, poor survival, and disease progression following combination treatment containing IMIDs and/or PIs. Finally, we built a seven‐gene model (MM‐IMiD and PI sensitivity‐7 genes [IP‐7]) using digital gene expression profiling data that significantly separates ND patients from IMiD‐ and PI‐refractory RR patients. Using this model, we retrospectively analyzed RNA sequcencing (RNAseq) data from the Mulltiple Myeloma Research Foundation (MMRF) CoMMpass (n = 578) and Mayo Clinic MM patient registry (n = 487) to divide patients into probabilities of responder and nonresponder, which subsequently correlated with overall survival, disease stage, and number of prior treatments. Our findings suggest that this model may be useful in predicting acquired resistance to treatments containing IMiDs and/or PIs

Bendamustine or high-dose cytarabine-based induction with rituximab in transplant-eligible mantle cell lymphoma.

The objective of this study was to explore differences in outcomes between first-line R-B and R-CHOP/R-DHAP in transplant-eligible patients with MCL. A population-based cohort of 97 patients aged 18-65 years with stage II-IV MCL, consecutively treated with R-B was retrospectively identified at BC Cancer. Baseline characteristics, response rates, and outcomes were compared to the cohort of 232 MCL patients randomized to the R-CHOP/R-DHAP arm of the MCL Younger trial. The primary aim was to estimate the hazard ratio (HR) of the progression-free survival (PFS) comparison between both groups, adjusted for MCL International Prognostic Index (MIPI), Ki67 index, and blastoid/pleomorphic morphology. Secondary endpoints included response rate, event free survival, overall survival, and time to next treatment. Ann Arbor stage, lactate dehydrogenase, MIPI, blastoid morphology, and MCL35 assignments were similar between both groups. The overall response rate (ORR) to R-B was 90% (54% complete response [CR]); 77% of patients proceeded to autologous stem cell transplantation (ASCT) and 78% received maintenance rituximab (MR). The ORR to R-CHOP/R-DHAP was 94% (54% CR); 78% proceeded to ASCT and 2% received MR. There were no differences in PFS in unadjusted (HR 0.87 [95% CI 0.53-1.41], p=0.56) or adjusted (HR 0.79 [95% CI 0.45-1.37], p=0.40) comparisons. There were no clear differences in secondary endpoints in unadjusted or adjusted analyses. This retrospective adjusted comparison of two independent cohorts of younger patients with MCL suggests that R-B with ASCT and maintenance rituximab is a feasible and effective first-line treatment, with outcomes comparable to R-CHOP/R-DHAP with ASCT

Evaluation of clinical parameters and biomarkers in older, untreated mantle cell lymphoma patients receiving bendamustine-rituximab

Mantle cell lymphoma (MCL) is clinically and biologically heterogeneous. While various prognostic features have been proposed, none currently impact therapy selection, particularly in older patients, for whom treatment is primarily dictated by age and comorbidities. Herein, we undertook a comprehensive comparison of clinicopathological features in a cohort of patients 60 years and older, uniformly treated with bendamustine and rituximab, with a median survival of >8 years. The strongest prognostic indicators in this cohort were a high-risk call by a simplified MCL international prognostic index (s-MIPI) (HR: 3.32, 95% CI: 1.65-6.68 compared to low risk), a high-risk call by MCL35 (HR: 10.34, 95% CI: 2.37-45.20 compared to low risk) and blastoid cytology (HR: 4.21, 95% CR: 1.92-9.22 compared to classic). Patients called high risk by both the s-MIPI and MCL35 had the most dismal prognosis (HR: 11.58, 95% CI: 4.10-32.72), while those with high risk by either had a moderate but clinically relevant prognosis (HR: 2.95, 95% CI: 1.49-5.82). A robust assay to assess proliferation, such as MCL35, along with stringent guidelines for cytological evaluation of MCL, in combination with MIPI, may be a strong path to risk-stratify older MCL patients in future clinical trials

Distinct molecular profile of IRF4-rearranged large B-cell lymphoma.

Pediatric large B-cell lymphomas (LBCLs) share morphological and phenotypic features with adult types but have better prognosis. The higher frequency of some subtypes such as LBCL with IRF4 rearrangement (LBCL-IRF4) in children suggests that some age-related biological differences may exist. To characterize the genetic and molecular heterogeneity of these tumors, we studied 31 diffuse LBCLs (DLBCLs), not otherwise specified (NOS); 20 LBCL-IRF4 cases; and 12 cases of high-grade B-cell lymphoma (HGBCL), NOS in patients ≤25 years using an integrated approach, including targeted gene sequencing, copy-number arrays, and gene expression profiling. Each subgroup displayed different molecular profiles. LBCL-IRF4 had frequent mutations in IRF4 and NF-κB pathway genes (CARD11, CD79B, and MYD88), losses of 17p13 and gains of chromosome 7, 11q12.3-q25, whereas DLBCL, NOS was predominantly of germinal center B-cell (GCB) subtype and carried gene mutations similar to the adult counterpart (eg, SOCS1 and KMT2D), gains of 2p16/REL, and losses of 19p13/CD70. A subset of HGBCL, NOS displayed recurrent alterations of Burkitt lymphoma-related genes such as MYC, ID3, and DDX3X and homozygous deletions of 9p21/CDKN2A, whereas other cases were genetically closer to GCB DLBCL. Factors related to unfavorable outcome were age >18 years; activated B-cell (ABC) DLBCL profile, HGBCL, NOS, high genetic complexity, 1q21-q44 gains, 2p16/REL gains/amplifications, 19p13/CD70 homozygous deletions, and TP53 and MYC mutations. In conclusion, these findings further unravel the molecular heterogeneity of pediatric and young adult LBCL, improve the classification of this group of tumors, and provide new parameters for risk stratification