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Macrophages are abundant immune cells in the microenvironment of diffuse large B-cell lymphoma (DLBCL). Macrophage estimation by immunohistochemistry shows varying prognostic significance across studies in DLBCL, and does not provide a comprehensive analysis of macrophage subtypes. Here, using digital spatial profiling with whole transcriptome analysis of CD68+ cells, we characterize macrophages in distinct spatial niches of reactive lymphoid tissues (RLTs) and DLBCL. We reveal transcriptomic differences between macrophages within RLTs (light zone /dark zone, germinal center/ interfollicular), and between disease states (RLTs/ DLBCL), which we then use to generate six spatially-derived macrophage signatures (MacroSigs). We proceed to interrogate these MacroSigs in macrophage and DLBCL single-cell RNA-sequencing datasets, and in gene-expression data from multiple DLBCL cohorts. We show that specific MacroSigs are associated with cell-of-origin subtypes and overall survival in DLBCL. This study provides a spatially-resolved whole-transcriptome atlas of macrophages in reactive and malignant lymphoid tissues, showing biological and clinical significance., (© 2024. The Author(s).)

Cancers often overexpress multiple clinically relevant oncogenes, but it is not known if combinations of oncogenes in cellular subpopulations within a cancer influence clinical outcomes. Using quantitative multispectral imaging of the prognostically relevant oncogenes MYC, BCL2, and BCL6 in diffuse large B-cell lymphoma (DLBCL), we show that the percentage of cells with a unique combination MYC+BCL2+BCL6- (M+2+6-) consistently predicts survival across four independent cohorts (n = 449), an effect not observed with other combinations including M+2+6+. We show that the M+2+6- percentage can be mathematically derived from quantitative measurements of the individual oncogenes and correlates with survival in IHC (n = 316) and gene expression (n = 2,521) datasets. Comparative bulk/single-cell transcriptomic analyses of DLBCL samples and MYC/BCL2/BCL6-transformed primary B cells identify molecular features, including cyclin D2 and PI3K/AKT as candidate regulators of M+2+6- unfavorable biology. Similar analyses evaluating oncogenic combinations at single-cell resolution in other cancers may facilitate an understanding of cancer evolution and therapy resistance., Significance: Using single-cell-resolved multiplexed imaging, we show that selected subpopulations of cells expressing specific combinations of oncogenes influence clinical outcomes in lymphoma. We describe a probabilistic metric for the estimation of cellular oncogenic coexpression from IHC or bulk transcriptomes, with possible implications for prognostication and therapeutic target discovery in cancer. This article is highlighted in the In This Issue feature, p. 1027., (©2023 The Authors; Published by the American Association for Cancer Research.)

CNS prophylaxis is commonly used in Diffuse Large B-Cell Lymphoma (DLBCL) patients with risk features for CNS relapse. This systematic review and meta-analysis compares CNS relapse rates with and without CNS prophylaxis, for patients at intermediate to high CNS relapse risk. Studies reporting CNS relapse risk category and CNS outcomes with and without CNS prophylaxis for antiCD20-CHOP treated DLBCL patients were included. 10 studies with 3770 patients at intermediate to high CNS relapse risk were analyzed. No significant difference in the pooled Absolute Risk Difference (ARD 0.01, 95 % CI -0.01 to 0.02, P = 0.61) or Risk (RR 1.22, 95 % CI 0.81-1.83, P = 0.34) was noted in patients with and without CNS prophylaxis. There were also no differences within pre-specified subgroups of IV Methotrexate or IT chemotherapy. However, the quality of evidence supporting these observations was low. A meta-analysis of individual patient data will help evaluate the benefit of CNS prophylaxis strategies., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The role of central nervous system (CNS) prophylaxis with high-dose methotrexate (HDMTX) in DLBCL is controversial. In this retrospective study, we evaluated the efficacy of prophylactic HDMTX on isolated CNS relapse, concomitant CNS and systemic relapse, systemic relapse, and survival outcomes in 226 patients with newly diagnosed DLBCL and high-risk CNS International Prognostic Index (CNS-IPI) score treated with RCHOP. The three-year risk of isolated CNS relapse was significantly lower in patients who received HDMTX, at 3.1% compared to 14.6% (P = 0.032) in those who did not. However, neither concomitant CNS-systemic relapse rates, systemic relapse rates, nor three-year PFS and OS were significantly different between treatment groups in multivariable analysis. Among propensity score-matched patients (N = 102), HDMTX was also associated with significantly lower isolated CNS relapse rates (HR 0.06, 95% CI 0.004-0.946, P = 0.046). HDMTX was well tolerated with manageable toxicities when given at a dose of 3 g/m 2 by day 3 of RCHOP chemotherapy. Using propensity score matching and multivariable regression to yield treatment groups with well-balanced covariates, we showed that prophylactic HDMTX improved isolated CNS relapse rates but did not decrease concomitant CNS-systemic relapse rates, systemic relapse rates, or improve survival outcomes., (© 2021. The Author(s).)

Polo-like kinase-1 (PLK1) regulates the MYC-dependent kinome in aggressive B-cell lymphoma. However, the role of PLK1 and MYC toward proliferation in diffuse large B-cell lymphoma (DLBCL) is unknown. We use multiplexed fluorescent immunohistochemistry (fIHC) to evaluate the co-localization of MYC, PLK1 and Ki67 to study their association with proliferation in DLBCL. The majority (98%, 95% CI 95-100%) of MYC/PLK1-double positive tumor cells expressed Ki67, underscoring the key role of the MYC/PLK1 circuit in proliferation. However, only 38% (95% CI 23-40%) and 51% (95% CI 46-51%) of Ki67-positive cells expressed MYC and PLK1, respectively. Notably, 40% (95% CI 26-43%) of Ki67-positive cells are MYC- and PLK-negative. A stronger correlation exists between PLK1 and Ki67 expression ( R  = 0.74, p  < .001) than with MYC and Ki67 expression ( R  = 0.52, p  < .001). Overall, the results indicate that PLK1 has a higher association than MYC in DLBCL proliferation and there are mechanisms besides MYC and PLK1 influencing DLBCL proliferation.

Background : Previous studies in Western populations, using immunohistochemistry (IHC) methods to subtype diffuse large B-cell lymphoma (DLBCL), suggest that germinal center B-cell lymphomas (GCBs) have improved outcomes. However, data in Asians have been limited and conflicting. This study aims to evaluate the prognostic impact of cell-of-origin (COO) subtyping by IHC and Lymph2Cx in South-East Asian (SEA) DLBCL patients, and to summarize the existing literature. Methods : A single-center retrospective analysis of 384 DLBCL patients diagnosed 2013-2018 who received Rituximab-based chemotherapy was performed. Hans and Lymph2Cx were used to assign COO and correlated with outcomes. Results : International Prognostic Index (IPI) score was associated with overall survival (OS) and progression-free survival (PFS). The 5-yr-OS for non-GCB versus GCB for COO by Hans was 70% versus 71% p =0.39, while 5-yr-OS for ABC versus GCB for COO by Lymph2Cx was 74% versus 92% p =0.19. The 5-yr-PFS for non-GCB versus GCB for COO by Hans was 65% versus 70% p =0.26, while 5-yr-PFS for ABC versus GCB for COO by Lymph2Cx was 64% versus 86% p =0.07. Conclusions : IPI is reaffirmed to be relevant in the rituximab era. COO by Hans has no prognostic significance, while subtyping by Lymph2Cx trends toward GCBs having better PFS and OS.

Primary mediastinal large B-cell lymphoma (PMBCL) is a distinct clinico-pathological subtype of diffuse large B-cell lymphoma with unclear prognostic factors and limited clinical data. Optimal treatment and role for radiotherapy is not fully defined. We performed a multicenter retrospective review of 124 patients with newly diagnosed PMBCL between 2001 and 2016. Treatment regimens were R-CHOP (n = 41), R-CHOP + RT (n = 37), and DA-EPOCH-R (n = 46). 6% (n = 3) in the DA-EPOCH-R group received RT. With a median follow up of 45 months, the overall 5-year OS and PFS was 89.4% and 82.4%, respectively. The type of chemo-radiotherapy regimen, B symptoms and Ann-Arbor staging showed a significant association with OS on univariate analysis but only B symptoms remained prognostic (P = 0.012) after multivariate analysis. The chemo-radiotherapy regimen, Japanese IPI and Ann-Arbor stage was significantly associated with PFS in univariate analysis, but only chemo-radiotherapy regimen remained significant (P = 0.02) after multivariate analysis. Patients who received R-CHOP + RT or DA-EPOCH-R had better PFS than those receiving R-CHOP alone, with 5-year PFS of 90% vs 88.5% vs 56%, respectively (P = 0.02). In the subgroup analysis of patients with bulk (n = 71), R-CHOP alone (n = 21) had inferior 5-year PFS 56.6% compared to those who received R-CHOP + RT (n = 23) 91.3% or DA-EPOCH-R (n = 27) 92.6% (P = 0.007). In contrast, in patients without bulk (n = 42), there was no impact of treatment regimen on PFS (P = 0.25). In conclusion, R-CHOP + RT and DA-EPOCH-R provide excellent outcomes in patients with PMBCL. In patients with bulky disease, the use of DA-EPOCH-R may be preferable as it allows omission of RT without reduction in efficacy., (© 2019 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)