B细胞淋巴瘤中的免疫检查点抑制剂：当前证据、分子决定因素及未来方向

… the rationale for investigation of immune-checkpoint inhibition in the treatment of B-cell … for clinical trials of drugs targeting PD-1 in these subtypes of large-B-cell lymphomas. A phase I …

Primary mediastinal B-cell lymphoma (PMBL) is a rare, aggressive B-cell lymphoma that accounts for 2% to 4% of all non-Hodgkin lymphomas, and it typically affects young adults in the third or fourth decade with a slightly higher incidence among females. 1,2 First-line treatment with rituximab and a chemotherapeutic agent is curative for >80% of PMBL cases. In contrast, 10% to 20% are refractory to or relapse after the fi rst treatment approach, mostly within the fi rst 18 months. 3,4 Salvage treatment consists of intensive regimens, followed by autologous stem cell transplantation, but outcomes are poor with a 2-year overall survival (OS) rate of 15% to 29%. 5,6 Immune checkpoint inhibitors (ICIs) restore the T lymphocyte – mediated antitumor effect. In the fi nal analysis of the phase 2 KEYNOTE-170 study, pembrolizumab (a humanized monoclonal immunoglobulin G4 antibody directed against programmed cell death protein 1) in patients with relapsed or refractory (R/R) PMBL yielded an overall response rate (ORR)

Classical Hodgkin lymphoma (cHL) has been identified with universal genetic alterations of chromosome 9p24.1, which contains PD-L1/PD-L2 genes. The amplification of 9p24.1 is associated with the increased expression of PD-L1 and PD-L2 on RS cells, which promotes their immune evasion, and subsequently makes cHL sensitive to PD-1 blockade. Several PD-1 inhibitors have shown significant efficacies with overall response rate (ORR) of 70%−90% in relapse/refractory (r/r) cHL and have acquired the approvals for this indication. Recently, more and more studies are conducted to investigate PD-1 blockade in earlier disease course and in combination with neo-agents or chemotherapy. Unlike cHL, non-Hodgkin lymphoma (NHL) consists of numerous subtypes harboring highly biological heterogeneity. Only a few subtypes have been shown to have genetic alteration of9p24.1 including primary mediastinal B cell lymphoma (PMBL), gray zone lymphoma (GZL) with features intermediate between diffuse large B cell lymphoma (DLBCL) and cHL, primary central nervous system lymphoma (PCNSL) and primary testicular lymphoma (PTL). Epstein-Barr virus (EBV)-associated lymphomas have a virally mediated overexpression of PD-L1, also making them sensitive to PD-1 blockade. Therefore, PD-1 inhibitors are less effective in most r/r NHL than in r/r cHL. Further understanding of the biological features of NHL and immune checkpoint inhibitors (ICPi) combined therapy is the research focus in the future. In this review, we outlined the recent progress of ICPi in lymphoma originating from clinical studies.

Cancer cells escape immune recognition by exploiting the programmed cell‐death protein 1 (PD‐1)/programmed cell‐death 1 ligand 1 (PD‐L1) immune checkpoint axis. Immune checkpoint inhibitors that target PD‐1/PD‐L1 unleash the properties of effector T cells that are licensed to kill cancer cells. Immune checkpoint blockade has dramatically changed the treatment landscape of many cancers. Following the cancer paradigm, preliminary results of clinical trials in lymphoma have demonstrated that immune checkpoint inhibitors induce remarkable responses in specific subtypes, most notably classical Hodgkin lymphoma and primary mediastinal B‐cell lymphoma, while in other subtypes, the results vary considerably, from promising to disappointing. Lymphomas that respond to immune checkpoint inhibitors tend to exhibit tumor cells that reside in a T‐cell‐rich immune microenvironment and display constitutive transcriptional upregulation of genes that facilitate innate immune resistance, such as structural variations of the PD‐L1 locus, collectively referred to as T‐cell‐inflamed lymphomas, while those lacking such characteristics are referred to as noninflamed lymphomas. This distinction is not necessarily a sine qua non of response to immune checkpoint inhibitors, but rather a framework to move the field forward with a more rational approach. In this article, we provide insights on our current understanding of the biological mechanisms of immune checkpoint evasion in specific subtypes of B‐cell and T‐cell non‐Hodgkin lymphomas and summarize the clinical experience of using inhibitors that target immune checkpoints in these subtypes. We also discuss the phenomenon of hyperprogression in T‐cell lymphomas, related to the use of such inhibitors when T cells themselves are the target cells, and consider future approaches to refine clinical trials with immune checkpoint inhibitors in non‐Hodgkin lymphomas.

Classical Hodgkin lymphoma (cHL) is characterized by nearly universal genetic alterations in 9p24.1, resulting in constitutive expression of PD-1 ligands. This likely underlies the unique sensitivity of cHL to PD-1 blockade, with response rates of ∼70% in relapsed/refractory disease. There are now numerous clinical trials testing PD-1 inhibitors in earlier stages of treatment and in combination with many other therapies. In general, non-Hodgkin lymphomas (NHLs) do not display a high frequency of 9p24.1 alterations and do not share cHL's vulnerability to PD-1 blockade. However, a few entities have genetic or immunologic features that may predict sensitivity to immune checkpoint blockade. These include primary mediastinal B cell lymphoma, primary central nervous system lymphoma, and primary testicular lymphoma, which harbor frequent alterations in 9p24.1, as well as Epstein Barr virus (EBV)-infected lymphomas, where EBV infection leads to increased PD-L1 expression. Although these subtypes may be specifically vulnerable to PD-1 blockade, the majority of NHLs appear to be minimally sensitive to PD-1 blockade monotherapy. Current investigations in NHL are therefore focusing on targeting other checkpoints or studying PD-1-based combination therapy. Looking forward, additional insight into the most common mechanisms of resistance to immune checkpoint inhibitors will be important to guide rational clinical trial design. In this review, we describe the biological basis for checkpoint blockade in cHL and NHL and summarize the clinical data generated to date. Guided by our rapidly evolving understanding of the pathobiology of various lymphoma subtypes, we are hopeful that the role of checkpoint inhibitors in lymphoma treatment will continue to grow.

Background B-cell non-Hodgkin lymphomas (NHL) are hematologic malignancies that arise in the lymph node. Despite this, the malignant cells are not cleared by the immune cells present. The failure of anti-tumor immunity may be due to immune checkpoints such as the PD-1/PDL-1 axis, which can cause T-cell exhaustion. Unfortunately, unlike Hodgkin lymphoma, checkpoint blockade in NHL has shown limited efficacy. Methods We performed an extensive functional analysis of malignant and non-malignant lymph nodes using high dimensional flow cytometry. We compared follicular lymphoma (FL), diffuse large B-cell lymphoma (DLBCL), and lymph nodes harboring reactive hyperplasia (RH). Results We identified an expansion of CD8+PD1+ T-cells in the lymphomas relative to RH. Moreover, we demonstrate that these cells represent a mixture of activated and exhausted T-cells in FL. In contrast, these cells are nearly universally activated and functional in DLBCL. This is despite expression of counter-regulatory molecules such as PD-1, TIM-3, and CTLA-4, and the presence of regulatory T-cells. Conclusion These data may explain the failure of single-agent immune checkpoint inhibitors in the treatment of DLBCL. Accordingly, functional differences of CD8+ T-cells between FL and DLBCL may inform future therapeutic targeting strategies.

Abstract The multicohort phase 1b KEYNOTE-013 study (NCT01953692) evaluated the safety and efficacy of pembrolizumab in patients with relapsed or refractory NHL who were ineligible for or failed hematopoietic cell transplantation (HCT). Patients received pembrolizumab (cohort 4) or pembrolizumab plus lenalidomide (cohort 5). Primary end points were safety and objective response rate (ORR) per IWG 2007 criteria. Cohort 4 included 89 patients. ORR was 22% (19/86; 90% CI 15–31; 10 CR, nine PR); ORRs by disease type were 48% (10/21), 10% (2/20), 12% (5/41), and 50% (2/4), for PMBCL, FL, DLBCL, and ‘other’ NHL, respectively. Toxicity was as predicted. Cohort 5 included 19 patients. ORR was 39% (90% CI 20–61; four CR, three PR). Hematologic toxicities were the most common treatment-related AEs. In conclusion, pembrolizumab following HCT ineligibility/failure confirms prior experience in PMBCL but not with NHL subtypes in this study. Additional analyses in DLBCL may not be warranted.

Non-Hodgkin lymphomas (NHLs) encompass a diverse group of malignancies arising from B cells, T cells, and natural killer (NK) cells at various stages of differentiation. Conversely, classical Hodgkin lymphomas (cHLs) primarily feature Reed-Sternberg cells (RSCs) amid a background of reactive immune cells. Immunomodulatory pathways, notably the PD-1/PD-L1 axis, play pivotal roles in tumor immune evasion across both NHLs and cHLs. Elevated expression of PD-1 and PD-L1 is observed in a spectrum of lymphomas, influencing prognosis and treatment response. Therapeutically, immune checkpoint inhibitors (ICIs) targeting PD-1/PD-L1 have revolutionized lymphoma management, particularly in relapsed/refractory cases. Nivolumab and pembrolizumab, among others, have demonstrated efficacy in various B-cell lymphomas, with promising outcomes in cHL. Combination strategies incorporating ICIs with conventional chemotherapy or targeted agents show enhanced efficacy and are being explored extensively. In this review we discuss the most important features of the tumor microenvironment of NHLs and cHLs, address the therapeutic approaches with ICIs and try to outline future perspectives.

ABSTRACT Introduction: Pembrolizumab is a novel monoclonal antibody that targets the interaction between programmed cell death protein 1 (PD-1) and its ligand (PD-L1). Pembrolizumab has shown significant clinical efficacy in Hodgkin Lymphoma (HL), but results in non Hodgkin Lymphoma (NHL) are mixed. Some NHL subtypes, which share certain genetic features with HL, such as alterations in chromosome 9p24.1 and expression of PD-L1, have shown promising responses in early phase trials. Areas covered: In this review, we provide an overview of pembrolizumab as a compound, and present the available clinical efficacy and safety data in the treatment of diffuse large B cell lymphomas. Expert opinion: Current early phase data suggest that single agent pembrolizumab in NHL demonstrates both efficacy and a favorable safety profile. However, it is anticipated that future treatment strategies will be biomarker-driven and incorporate pembrolizumab into combination therapies with chemotherapy and/or immunotherapy agents.

Programmed cell death protein 1 (PD-1) blockade targeting the PD-1 immune checkpoint has demonstrated unprecedented clinical efficacy in the treatment of advanced cancers including hematologic malignancies. This article reviews the landscape of PD-1/programmed death-ligand 1 (PD-L1) expression and current PD-1 blockade immunotherapy trials in B-cell lymphomas. Most notably, in relapsed/refractory classical Hodgkin lymphoma, which frequently has increased PD-1+ tumor-infiltrating T cells, 9p24.1 genetic alteration, and high PD-L1 expression, anti-PD-1 monotherapy has demonstrated remarkable objective response rates (ORRs) of 65% to 87% and durable disease control in phase 1/2 clinical trials. The median duration of response was 16 months in a phase 2 trial. PD-1 blockade has also shown promise in a phase 1 trial of nivolumab in relapsed/refractory B-cell non-Hodgkin lymphomas, including follicular lymphoma, which often displays abundant PD-1 expression on intratumoral T cells, and diffuse large B-cell lymphoma, which variably expresses PD-1 and PD-L1. In primary mediastinal large B-cell lymphoma, which frequently has 9p24.1 alterations, the ORR was 35% in a phase 2 trial of pembrolizumab. In contrast, the ORR with pembrolizumab was 0% in relapsed chronic lymphocytic leukemia (CLL) and 44% in CLL with Richter transformation in a phase 2 trial. T cells from CLL patients have elevated PD-1 expression; CLL PD-1+ T cells can exhibit a pseudo-exhaustion or a replicative senescence phenotype. PD-1 expression was also found in marginal zone lymphoma but not in mantle cell lymphoma, although currently anti-PD-1 clinical trial data are not available. Mechanisms and predictive biomarkers for PD-1 blockade immunotherapy, treatment-related adverse events, hyperprogression, and combination therapies are discussed in the context of B-cell lymphomas.

Simple Summary The PD-1/PD-L1 axis is not only involved in anti-tumour immune evasion of germinal center (GC)-derived diffuse large B cell lymphomas (DLBCL), but also inherently in the fine-tuned regulation of normal GC reactions during humoral immune responses. This checkpoint axis modulates crosstalks between B and T cells that allow positive selection for survival and proliferation. Malignant DLBCL cells may deceive and take advantage of these mechanisms to establish an immunosuppressive microenvironment. This review delves into PD-1/PD-L1 role in the complex inter-cellular interactions from normal GC reactions to DLBCL progression, in order to highlight vulnerabilities that could be targeted by promising combination immunotherapies. Abstract Besides a recognized role of PD-1/PD-L1 checkpoint in anti-tumour immune evasion, there is accumulating evidence that PD-1/PD-L1 interactions between B and T cells also play an important role in normal germinal center (GC) reactions. Even when smaller in number, T follicular helper cells (TFH) and regulatory T (TFR) or B (Breg) cells are involved in positive selection of GC B cells and may result critical in the lymphoma microenvironment. Here, we discuss a role of PD-1/PD-L1 during tumour evolution in diffuse large B cell lymphoma (DLBCL), a paradigm of GC-derived lymphomagenesis. We depict a progression model, in two phases, where malignant B cells take advantage of positive selection signals derived from correct antigen-presentation and PD-1/PD-L1 inter-cellular crosstalks to survive and initiate tumour expansion. Later, a constant pressure for the accumulation of genetic/epigenetic alterations facilitates that DLBCL cells exhibit higher PD-L1 levels and capacity to secrete IL-10, resembling Breg-like features. As a result, a complex immunosuppressive microenvironment is established where DLBCL cells sustain proliferation and survival by impairing regulatory control of TFR cells and limiting IL-21-mediated anti-tumour functions of TFH cells and maximize the use of PD-1/PD-L1 signaling to escape from CD8+ cytotoxic activity. Integration of these molecular and cellular addictions into a framework may contribute to the better understanding of the lymphoma microenvironment and contribute to the rationale for novel PD-1/PD-L1-based combinational immunotherapies in DLBCL.

Simple Summary Immunotherapy using antibodies against programmed cell death protein 1 (PD-1) or its ligand PD-L1 can restore host antitumor immunity in many types of cancer. Hence, PD-1/PD-L1 blockade therapy has become the standard treatment for various cancer types in the last decade. However, apart from classic Hodgkin lymphoma and primary mediastinal B-cell lymphoma, PD-1/PD-L1 blockade therapy has shown limited efficacy in other lymphomas. To address this gap, several clinical trials of combination therapies with PD-1/PD-L1 inhibitors have been recently conducted or are underway in both frontline and relapsed/refractory settings. Here, we comprehensively review these clinical studies of combination therapies for lymphomas and discuss their outcomes in the hope of providing a perspective to develop novel therapeutic approaches for combination therapy. Abstract Immunotherapy with the programmed cell death protein 1 (PD-1)/PD-1 ligand (PD-L1) blockade has revolutionized the treatment of advanced solid cancers. However, these clinical benefits have been limited to cases of malignant lymphomas, showing promising results for only classic Hodgkin lymphoma (cHL) and primary mediastinal B-cell lymphoma (PMBCL). To bring clinical benefits to more patients with lymphoma, numerous combination therapies involving PD-1/PD-L1 blockade have been tested in clinical trials in both frontline and relapsed/refractory settings. This article reviews the current landscape of combination therapies with PD-1/PD-L1 blockade for lymphoma and discusses the potential therapeutic approaches. An interim analysis of a phase 3 study demonstrated increased progression-free survival with nivolumab combination therapy over the current frontline treatment in patients with advanced-stage cHL. The results of combination therapies for aggressive B-cell lymphomas, except for PMBCL, have been disappointing. Several clinical trials of combined PD-1/PD-L1 blockade and Bruton’s tyrosine kinase inhibitors are exploring its efficacy in patients with chronic lymphocytic leukemia (CLL) with Richter transformation. Several T-cell lymphoma subtypes respond to PD-1/PD-L1 blockade monotherapy. Further clinical trials are underway to investigate appropriate combination regimens with PD-1/PD-L1 blockade, especially for cHL, CLL with Richter transformation, and T-cell lymphoma, in both frontline and relapsed/refractory settings.

Key Points • The best response rates were observed with pembrolizumab before tisagenlecleucel; however, definitive conclusions cannot be made.• Pembrolizumab did not augment the cellular expansion of tisagenlecleucel but delayed peak expansion in the day 1 cohort.

… The beneficial antitumoral activity of programmed death-1 (PD-1) and programmed death-ligand … on the role of the PD-1/PD-L1 pathway in Diffuse Large B-cell lymphoma (DLBCL), and …

CD19-directed chimeric antigen receptor-modified T cells (CAR T cells) achieve durable remissions in about 30-40% of relapsed/refractory large B-cell lymphomas. T cell exhaustion and/or an immunosuppressive tumor-microenvironment may contribute to CAR T-cell failure. Pembrolizumab, an anti-PD1 immune checkpoint inhibitor, may reverse T-cell exhaustion following CAR T-cell therapy. We treated 12 patients with B-cell lymphomas who were either refractory to (N=9) or relapsed after (N=3) CD19-directed CAR T cell (4-1BB-costimulated) therapy with pembrolizumab 200mg IV every 3 weeks. Median time from CAR T-cell infusion to first pembrolizumab dose was 3.3 months (range: 0.4-42.8 months). Pembrolizumab was well-tolerated and the only ≥ grade 3 adverse events related to pembrolizumab were neutropenia (N=3; 25%). Best overall response rate after pembrolizumab was 3/12 (25%) [1 complete response; 2 partial responses]. One (8%) patient had stable disease, thus, 4/12 (33%) patients had clinical benefit. After pembrolizumab, 4 patients with clinical benefit had increase in percentage of CAR T cells by mass cytometry (CyTOF); 3 of 4 of these patients also had increases in CAR19 transgene levels by qPCR. Deep immune profiling using mass cytometry revealed increased CAR T cell activation and proliferation and less T-cell exhaustion in clinical responders. Together, PD1 blockade with pembrolizumab after CD19-directed CAR T-cell therapy appears safe and may achieve clinical responses in some patients with B-cell lymphomas refractory to or relapsed after CAR T-cell therapy.

Purpose Pembrolizumab, a programmed cell death protein 1 (PD1) inhibitor inhibits the interplay between PD1 of T-cell and programmed cell death ligand 1 (PDL1) on tumor cells. Although pembrolizumab has been tried to various subtypes of non-Hodgkin lymphoma (NHL), real-world data about the efficacy of pembrolizumab in NHL patients are limited. Materials and methods We analyzed the outcome of 30 relapsed or refractory NHL patients treated with pembrolizumab, and compared the outcome between Epstein-Barr virus (EBV)‒positive and negative subtypes because EBV infection of tumor cells can upregulate PDL1 expression. Results Seven patients with EBV-positive NHL showed a response including NK/T-cell lymphoma (6/14, 44%) and primary mediastinal B-cell lymphoma (1/4, 25%) whereas EBV-negative subtypes did not respond such as diffuse large B-cell lymphoma and T-lymphoblastic lymphoma. We also evaluated PDL1 expression using tumor tissue of 76 patients. High PDL1 expression (positive staining of > 50% of tumor cells) was more frequent in NK/T-cell lymphoma and primary mediastinal B-cell lymphoma than other subtypes. Thus, PDL1 expression was significantly higher in EBV-positive (18/32, 56%) than EBV-negative NHL (4/38, 11%, p < 0.001). Furthermore, NK/T-cell lymphoma patients with high PDL1 expression showed a higher response (4/6, 67%) than those with low PDL1 expression (1/5, 20%). Conclusion Pembrolizumab could be useful as a salvage treatment for relapsed or refractory EBV-positive NHL, especially NK/T-cell lymphoma. However, its efficacy in EBV-negative NHL with low or absent PDL1 expression is still not clear although pembrolizumab could be a potential treatment option for relapsed or refractory NHL.

Key Points Treatment with pembrolizumab plus dinaciclib resulted in antitumor activity with acceptable safety for patients with rrCLL and rrDLBCL. A careful and comprehensive approach to explore PD-1 inhibition and CDK9 inhibition in combination with other agents is needed.

Primary mediastinal large B-cell lymphoma (PMBCL) is a subtype of diffuse large B-cell lymphoma (DLBCL). PMBCL comprises approximately 10% of DLBCLs, thus making it a rare variant of DLBCL. Cure rates for PMBCL with upfront regimens like DA-REPOCH exceed 90%. However, if there is a poor response to this first-line therapy, relapsed/refractory PMBCL (rrPMBCL) has limited treatment options. The historic trend is to treat rrPMBCL with salvage regimens commonly used for DLBCL followed by high-dose therapy and autologous stem cell transplant (HDT-ASCT); however, response rates to salvage therapy remain low and few patients are able to proceed to transplant. An interesting feature of PMBCL is that even though it is classified as a subtype of DLBCL, PMBCL actually shares many clinical, pathologic, and genetic features with classical Hodgkin lymphoma (cHL). For example, both frequently express program death ligand 1 and 2 (PD-L1/2), which is not seen in other mature B-cell lymphomas. The expression of PD-L1/2 in PMBCL makes PDL1 inhibitors, such as pembrolizumab, an attractive therapeutic target. Pembrolizumab is an effective and well-tolerated therapy now approved for a number of cancer types from advanced melanoma to relapsed/refractory cHL. There are now multi-institutional trials underway assessing the role of pembrolizumab in the treatment of rrPMBCL.

Refractory or relapsed diffuse large B-cell lymphoma (DLBCL) often associates with the activated B-cell-like (ABC) subtype and genetic alterations that drive constitutive NF-κB activation and impair B-cell terminal differentiation. Here, we show that DNA damage response by p53 is a central mechanism suppressing the pathogenic cooperation of IKK2ca-enforced canonical NF-κB and impaired differentiation resulting from Blimp1 loss in ABC-DLBCL lymphomagenesis. We provide evidences that the interplay between these genetic alterations and the tumor microenvironment select for additional molecular addictions that promote lymphoma progression, including aberrant coexpression of FOXP1 and the B-cell mutagenic enzyme activation-induced deaminase, and immune evasion through major histocompatibility complex class II downregulation, PD-L1 upregulation, and T-cell exhaustion. Consistently, PD-1 blockade cooperated with anti-CD20-mediated B-cell cytotoxicity, promoting extended T-cell reactivation and antitumor specificity that improved long-term overall survival in mice. Our data support a pathogenic cooperation among NF-κB-driven prosurvival, genetic instability, and immune evasion mechanisms in DLBCL and provide preclinical proof of concept for including PD-1/PD-L1 blockade in combinatorial immunotherapy for ABC-DLBCL.

Aim To evaluate the therapeutic value and find out potential combination agents of programmed death 1/programmed death-ligand 1 (PD-1/PD-L1) monoclonal antibody (mAb) in relapsed/refractory (r/r) diffuse large B-cell lymphoma (DLBCL). Methods We conducted a meta-analysis to assess the efficacy and safety of PD-1/PD-L1 mAb in r/r DLBCL, potential qualified studies were searched in PubMed, Embase, Web of Science, and ClinicalTrials.gov. This meta-analysis had been registered on the PROSPERO platform (CRD42023340031). Results After systematic screening, a total of 32 records involving 29 studies were included, pooled survival curves indicated better progression-free survival (PFS) (p < 0.0001; HR = 0.51, 95% CI [0.42–0.62]) and overall survival (OS) (p = 0.013; HR = 0.71, 95% CI [0.57–0.88]) for combination therapy compared with monotherapy. Combination therapy group also achieved a better pooled complete response rate (CRR) (14.6% vs. 3.0%; p < 0.001) and overall response rate (ORR) (30.5% vs. 10.3%; p < 0.001). Analysis of the incidence of adverse events (AEs) did not demonstrate additional toxicities of combination therapy. The limitation was the predominance of single-arm trials, precluding the direct comparison of combination versus partner agents alone. Conclusions These findings support further exploration of PD-1/PD-L1-mAb-based combination therapy to identify long-term survival benefits, while application of monotherapy in unselected DLBCL patients is not recommended.

… antitumour immunity is PD-L1. Responders expressed higher levels of PD-L1 on peripheral … lymphoma” and “rituximab” and “PD-1”). We did not identify any other clinical trials targeting …

Nivolumab, an anti-programmed death-1 (PD-1) monoclonal antibody, showed promising activity in relapsed or refractory (R/R) follicular lymphoma (FL) in a phase 1 study. We conducted a phase 2 trial to evaluate further its efficacy and safety in patients with R/R FL and to explore biomarkers of response. Patients with R/R FL and at least two prior lines of therapy, each containing a CD20 antibody or an alkylating agent, were treated with nivolumab 3 mg/kg every 2 weeks. The primary endpoint was objective response rate (ORR) assessed by independent radiologic review committee. Biomarker analyses included gene expression profiling and multiplex immunofluorescence studies of pretreatment tumor samples. A total of 92 patients were treated. After a minimum follow-up of 12 months, ORR was 4% (4/92). Median PFS was 2.2 months (95% confidence interval [CI], 1.9-3.6). Median DOR was 11 months (95% CI, 8-14). Exploratory analyses suggested that responders had significantly higher proportion of CD3+ T cells in the tumor microenvironment than non-responders, but no significant differences in PD-1 or PD-L1 expression were observed. High expression of a set of tumor-associated macrophage genes was associated with reduced PFS (hazard ratio, 3.28; 95% CI, 1.76-6.11; P = .001). The safety profile was consistent with previous reports of nivolumab. In conclusion, nivolumab monotherapy was associated with very limited activity in patients with R/R FL. Better understanding of the immune biology of this disease may facilitate the development of effective checkpoint-based strategies. This trial was registered at www.clinicaltrials.gov as #NCT02038946.

Purpose Follicular lymphoma (FL) is an indolent, yet generally incurable neoplasia with a median survival exceeding 10 years. However, a subset of FL patients experiences histological transformation (HT) to a more aggressive lymphoma, in the majority of cases to diffuse large B-cell lymphoma (DLBCL). This affects both the clinical course and the prognostic outcome, resulting in a markedly reduced survival after transformation. Thus, early risk stratification and prediction of patients at risk of HT would be highly valuable in the clinical setting. Here, we investigated the potential of the immune inhibitory programmed death 1 (PD-1) receptor as a biomarker predictive of HT. Patients and Methods Immunohistochemical staining and quantification by digital image analysis of PD-1 was performed on diagnostic tumor-tissue samples from FL patients with and without subsequent transformation (n=34 and n=46, respectively), and on paired samples from the transformed lymphoma (n=34). Results At the time of initial FL diagnosis, samples from patients with subsequent HT had significantly higher tumor-tissue expression of PD-1 compared with diagnostic FL samples from patients without subsequent HT (p=0.010). At the time of transformation, PD-1 expression was significantly reduced (p<0.001). No difference was observed in intra-follicular PD-1 expression at FL diagnosis between samples from patients with or without HT; however, high intra-follicular levels of PD-1 were associated with significantly shorter transformation-free survival times (p<0.043). Conclusion Our data suggest that pre-treatment tumor-tissue PD-1 expression already predicts the risk of subsequent transformation to DLBCL, as early as the time of FL diagnosis.

ABSTRACT Follicular lymphoma (FL) accounts for approximately 35% of all non-Hodgkin lymphomas and can progress to diffuse large B cell lymphoma (DLBCL) at a rate of 2% per year. Here, we present a 56-year-old female patient who was diagnosed with grade 3a FL. Further pathological investigation revealed that the lymphoma had transformed into DLBCL following six courses of R-CHOP regimen, and further disease progression was observed after two courses of R2-GemOx. We ultimately failed to collect hematopoietic stem cells after two courses of R2-ICE. CD-22 CAR-T cell therapy salvaged the patient; however, a new soft tissue mass of 4.8 × 4.1 cm rapidly emerged in the patient’s right lung. Following the observation of strong tissue staining of PD-L1 (90%), the patient was administered PD-1 inhibitor and 26 Gy of radiotherapy and has maintained progression-free survival at more than 15 months of follow-up. Transformed FL with strong PD-L1 expression showed a poor response to standard immunochemotherapy. Our findings support the potential benefit of PD-1 inhibitor and combination therapies in this type of transformed FL.

Key Points Combination therapy with pembrolizumab and rituximab was well tolerated in patients with relapsed/refractory follicular lymphoma. In this single-arm, phase 2 study, the overall response rate was 67%, with 50% complete response and median PFS of 12.6 months.

Previously considered a subtype of diffuse large B-cell lymphoma (DLBCL), primary mediastinal B-cell lymphoma (PMBCL) is now recognized by the World Health Organization as an independent entity. PMBCL has clinicopathologic features that are separate from systemic DLBCL and harbors some biologic characteristics which overlap with nodular sclerosing classic Hodgkin’s lymphoma (cHL). Similar to cHL, copy number alterations of 9p24.1 are frequently seen in PMBCL, which leads to increased expression of key genes in the region, including programmed death-ligand 1( PD-L1), PD-L2, and JAK2. In addition, PMBCL cells express CD30 in a mostly patchy fashion. In the upfront setting, dose-adjusted etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin, and rituximab (i.e., DA-EPOCH-R) is the only regimen that has been shown in a prospective setting to result in outstanding outcomes without consolidative radiation to the mediastinum, with a 5-year event-free survival rate of 93% and overall survival rate of 97%. Thus, in recent years, DA-EPOCH-R has been recognized as the preferred frontline regimen. Despite the encouraging results in the frontline setting, the outcomes in the relapsed/refractory setting remain poor. The current approach of salvage chemotherapy followed by autologous stem cell transplantation, as used in patients with DLBCL, does not result in high rates of cure in patients with rrPMBCL. In recent years, the characteristic molecular features identified in PMBCL have provided more treatment opportunities for this patient population. In the relapsed setting, single-agent PD-1 inhibitor pembrolizumab have demonstrated high and durable remission rates. Despite the expression of CD30, the CD30 antibody drug-conjugate brentuximab vedotin (BV) as a single agent has been deemed inactive in this disease. On the contrary, the combinations of BV and PD-1 inhibitor have shown higher response rates than PD-1 inhibitor alone. Moreover, anti-CD19 chimeric antigen receptor T-cell (CAR T-cell) therapy has been positioned as another successful strategy for patients with rrPMBCL. Axicabtagene ciloleucel and lisocabtagene maraleucel are two products used in rrPMBCL.

… Treatment and response assessment of patient with a refractory primary mediastinal B-cell lymphoma treated with PD-1 and JAK1/2 inhibition. a Treatment schedule and response …

Primary mediastinal large B cell lymphoma (PMBCL) is an aggressive large B cell lymphoma originating in the mediastinum, that mainly expresses B cell surface molecules, such as CD19, CD20, CD22, andCD79a. Clinically, they are characterized by rapidly increasing anterior mediastinal masses, which can cause compression of the surrounding tissues. The diagnosis of PMBCL mainly depends on the pathological features, imaging examination and clinical features. Currently, the most commonly used therapeutic regimens are R‐CHOP and R‐EPOCH. Radiotherapy is beneficial in some patients, but it can also lead to long‐term toxicity. The research and development of novel therapies are ongoing, and some studies have achieved encouraging results, including those conducted on chimeric antigen receptor‐modified T (CAR‐T) cell therapy and anti‐PD‐1 drugs. However, randomized controlled trials with larger sample sizes are still needed. Positron emission tomography‐computed tomography (PET‐CT) is mainly used to assess the curative effect after treatment and to guide the subsequent treatment strategy.

Primary mediastinal B‐cell lymphoma (PMBCL) is a distinct disease closely related to classical nodular sclerosing Hodgkin lymphoma. Conventional diagnostic paradigms utilising clinical, morphological and immunophenotypical features can be challenging due to overlapping features with other B‐cell lymphomas. Reliable diagnostic and prognostic biomarkers that are applicable to the conventional diagnostic laboratory are largely lacking. Nuclear factor kappa B (NF‐κB) and Janus kinase/signal transducers and activators of transcription (JAK‐STAT) signalling pathways are characteristically dysregulated in PMBCL and implicated in several aspects of disease pathogenesis, and the latter pathway in host immune evasion. The tumour microenvironment is manipulated by PMBCL tumours to avoid T‐cell mediated destruction via strategies that include loss of tumour cell antigenicity, T‐cell exhaustion and activation of suppressive T‐regulatory cells. R‐CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisolone) and DA‐EPOCH‐R (dose‐adjusted etoposide, prednisolone, vincristine, cyclophosphamide, doxorubicin, rituximab) are the most common first‐line immunochemotherapy regimens. End of treatment positron emission tomography scans are the recommended imaging modality and are being evaluated to stratify patients for radiotherapy. Relapsed/refractory disease has a relatively poor outcome despite salvage immunochemotherapy and subsequent autologous stem cell transplantation. Novel therapies are therefore being developed for treatment‐resistant disease, targeting aberrant cellular signalling and immune evasion.

Key Points Treatment options for relapsed/refractory PMBCL are limited, and prognosis is generally poor. Pembrolizumab had a manageable safety profile and promising antitumor activity in heavily pretreated rrPMBCL patients.

Primary mediastinal large B-cell lymphoma (PMBCL) is an aggressive B-cell lymphoma arising from thymic B-cells having clinicopathologic features distinct from systemic diffuse large B-cell lymphoma (DLBCL). PMBCL comprises 2% to 4% of all non-Hodgkin lymphomas (NHL), 7% of DLBCL and seen predominantly in young females with a median age of 35 years at diagnosis. The annual incidence of PMBCL is 0.4 per million with a 5-year survival rate exceeding 70% with improving supportive care and genetic characterization of the disease. Pathogenesis involves dysregulation of Janus kinase-signal transducer and activator of transcription (JAK-STAT), nuclear factor-kB (NF-kB) pathways and amplification of the 9p24.1 region of chromosome 9. PMBCL patients have a prolonged life expectancy necessitating the need for treatment approaches that are based on maximizing cure with minimal long-term toxicity. Due to rarity and its recognition as a distinct entity, therapeutic decisions are guided by clinical presentation, clinician and center experience, and analysis of patients with PMBCL within DLBCL registries. Historically R-CHOP has been the usual first line treatment for PMBCL followed by involved site radiotherapy (ISRT), however clinical practice varies across centers with emerging consensus to avoid upfront RT by utilizing dose intense regimens (DA-EPOCH-R) in younger and fit patients. Prognosis of relapsed refractory PMBCL not responding to salvage chemotherapy is dismal, however there are many emerging options including Brentuximab Vedotin, immune check point inhibitors and chimeric antigen receptor T-cell therapy. In this article, we focus on the pathogenesis, current and evolving treatments, and provide recommendations for optimal management of patients with PMBCL.

Key Points • Nivolumab + BV showed durable safety and efficacy as salvage in patients with R/R PMBL after 3-year follow-up, with no new safety signals.• High 2-year complete response rates (80%-100%) after subsequent HCT indicate potential for nivo + BV as a bridging therapy to transplant.

… two immune checkpoint inhibitors, resulting in the inhibition of T-… /kappa monoclonal antibody targeting PD-1. The common … by lymphoma cells support the use of anti-PD-1 blockers in …

ABSTRACT Introduction: Primary mediastinal large B-cell lymphoma (PMBL) is a rare subtype of lymphoma, clinically and biologically distinct from diffuse large B-cell lymphoma (DLBCL) that shows overlapping features with classical Hodgkin lymphoma (cHL). If first-line strategies lead to 80–85% of curability, relapse occurs early with a chemo-refractory disease and a poor outcome. The presence of 9p24.1 rearrangement, conducting to the overexpression of the immune checkpoint molecules PDL1 and 2, has paved the way for immune checkpoint blockers development in these entities. Pembrolizumab, an anti PD-1 checkpoint antibody, was initially approved in solid cancer and later on in the lymphoma field in cHL. Areas covered: We summarize the biology and clinical need in PMBL, leading to the rationale for checkpoint inhibitors development, as well as pembrolizumab clinical studies in this entity. To do so, we performed a PubMed search using the terms: ‘PMBCL,’ ‘lymphoma,’ ‘Immune checkpoint,’ and ‘Pembrolizumab.’ Expert opinion: Pembrolizumab showed tolerable safety profile and efficacy data in patients with PMBL who have relapsed after, or are ineligible for autologous stem cell transplant (ASCT). Some combination strategies have shown promising preliminary results, while others are currently being conducted.

Primary mediastinal large B-cell lymphoma (PMBCL) is a distinct clinicopathologic disease from other types of diffuse large B-cell lymphoma (DLBCL) with unique prognostic features and limited availability of clinical data. The current standard treatment for newly diagnosed PMBCL has long been dependent on a dose-intensive, dose-adjusted multi-agent chemotherapy regimen of rituximab plus etoposide, prednisone, vincristine, cyclophosphamide, and doxorubicin (DA-R-EPOCH). Recent randomized trials have provided evidence that R-CHOP followed by consolidation radiotherapy (RT) is a valuable alternative option to first-line treatment. For recurrent/refractory PMBCL (rrPMBCL), new drugs such as pembrolizumab and CAR-T cell therapy have proven to be effective in a few studies. Positron emission tomography-computed tomography (PET-CT) is the preferred imaging modality of choice for the initial phase of lymphoma treatment and to assess response to treatment. In the future, baseline quantitative PET-CT can be used to predict prognosis in PMBCL. This review focuses on the pathology of PMBCL, underlying molecular basis, treatment options, radiotherapy, targeted therapies, and the potential role of PET-CT to guide treatment choices in this disease.

Introduction: Currently available therapy options for primary mediastinal large B-cell lymphoma (PMBCL) generally yield poor treatment outcomes. Like classical Hodgkin lymphoma (cHL), PMBCL frequently exhibits 9p24.1/PD-L1/PD-L2 copy number alterations and rearrangements and associated PD-L1 and/or PD-L2 overexpression, which may facilitate immune evasion. The genetics of PMBCL could thus make it susceptible to PD-1 blockade. KEYNOTE-013 (NCT01953692) is an ongoing multicenter, multicohort Phase 1b trial evaluating safety, tolerability, and antitumor activity of pembrolizumab, a humanized anti-PD-1 monoclonal antibody, in patients with hematologic malignancies. Here we report results from the first 19 patients enrolled in the PMBCL cohort of KEYNOTE-013, with a follow-up of up to 2 years. Methods: This independent cohort of KEYNOTE-013 is enrolling patients with relapsed/refractory (R/R) PMBCL who have relapsed after or are ineligible for autologous stem cell transplant (SCT). Patients received pembrolizumab IV 10 mg/kg every 2 weeks (Q2W), which was later changed by protocol amendment to a fixed dose of 200 mg every 3 weeks (Q3W), following PK/PD studies demonstrating both regimens to be equivalent. Treatment continues for up to 2 years or until unacceptable toxicity or confirmed disease progression. Treatment response is evaluated using IHP 2007 criteria by positron emission tomography and computed tomography at weeks 6 and 12, and every 9 weeks thereafter. Primary end points are safety and objective response rate (ORR). Secondary end points include complete remission (CR) rate and duration of response (DOR). The safety population consists of all patients who receive ≥1 dose of study drug and the efficacy population of all patients who progress prior to or reach the first efficacy evaluation. Whole blood was collected at predefined time points before and during pembrolizumab treatment for RNA and DNA extraction and biomarker analysis using a number of genomic-based assays, including NanoString and RNA sequencing. Results: As of the analysis cutoff date (May 27, 2016), 19 patients were enrolled in the PMBCL cohort, 18 were treated, and 16 had ≥1 post-baseline efficacy evaluation. The first 11 patients were to receive pembrolizumab IV 10 mg/kg Q2W (1 was not treated due to early progressive disease); all subsequent patients received 200 mg Q3W. Median age was 30.5 years (range, 22-62). Most patients (72%) were female. 61% of patients had ≥3 prior lines of therapy, 33% had prior autologous SCT, and 61% prior radiation. In the efficacy population, 16 patients were evaluable for response: one discontinued treatment based on clinical progression before the first response assessment (this patient was considered a nonresponder), the other had not reached the first assessment.The ORR was 41% (7/17), with 2 patients achieving a CR and 5 patients a partial response; 35% (6/17) had stable disease as best response. Overall, 81% (13/16) of evaluable patients had target lesion reductions (Figure). With a median follow-up duration of 11.3 months (range, 3.4 to 27.4 months), median DOR was not reached, and there were 6 ongoing responses at time of current data cutoff. DOR ranged from 2.4+ to 22.5+ months; DOR in the 2 patients with CR was 2.4+ and 20.5+ months. Two patients received an allogeneic SCT: one had SD, the other PD on pembrolizumab. Ten patients discontinued treatment: 5 due to progressive disease based on imaging, 4 for clinical progression, and 1 due to physician decision. Two patients reached the maximum 2 years of treatment and remain in remission. Six patients experienced serious AEs, and none discontinued due to AEs. Eleven patients (61%) experienced treatment-related adverse events (TRAEs), mostly grade 1-2. One patient experienced a TRAE of grade 3 neutropenia and another a TRAE of grade 4 venoocclusive liver disease (VOD; after allogeneic SCT during the follow-up period after pembrolizumab was discontinued), the only serious TRAE. The patient recovered from the VOD. There were no treatment-related deaths. Conclusions: These results from an on-going study in heavily pretreated R/R PMBCL patients demonstrate that PD-1 blockade with pembrolizumab has a manageable safety profile and promising antitumor activity. Due to these results, a pivotal global multi-center Phase 2 trial, KEYNOTE-170, is further evaluating single agent pembrolizumab in patients with R/R PMBCL. Figure. Figure. Disclosures Zinzani: MorphoSys: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees. Ribrag:Esai: Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Infinity: Membership on an entity's Board of Directors or advisory committees; Pharmamar: Membership on an entity's Board of Directors or advisory committees; ArgenX: Research Funding; Incyte: Membership on an entity's Board of Directors or advisory committees; NanoString: Membership on an entity's Board of Directors or advisory committees. Moskowitz:Seattle Genetics: Consultancy, Research Funding; Pharmacyclics: Research Funding; Merck &amp; Co., Inc.: Consultancy, Research Funding; Celgene Corporation: Consultancy; Genentech BioOncology: Consultancy. Michot:Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees. Kuruvilla:Merck &amp; Co., Inc.: Consultancy, Honoraria. Balakumaran:Merck &amp; Co.: Employment, Other: stock, stock options. Zhang:Merck &amp; Co., Inc.: Employment, Other: stock, stock options. Marinello:Merck &amp; Co., Inc.: Employment, Other: stock, stock options. Chlosta:Merck &amp; Co., Inc.: Employment, Other: stock, stock options. Gustafson:Merck &amp; Co., Inc.: Employment, Other: stock, stock options. Shipp:Merck &amp; Co., Inc.: Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees, Research Funding. Armand:Otsuka: Research Funding; Merck &amp; Co., Inc.: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Infinity: Consultancy; Roche: Research Funding; Sequenta: Research Funding; Tensha: Research Funding; Sigma Tau: Research Funding.

PURPOSE Patients with relapsed or refractory primary mediastinal large B-cell lymphoma (rrPMBCL) have a poor prognosis, and their treatment represents an urgent and unmet need. Because PMBCL is associated with genetic aberrations at 9p24 and overexpression of programmed cell death-1 (PD-1) ligands (PD-L1), it is hypothesized to be susceptible to PD-1 blockade. METHODS In the phase IB KEYNOTE-013 (ClinicalTrials.gov identifier: NCT01953692) and phase II KEYNOTE-170 (ClinicalTrials.gov identifier: NCT02576990) studies, adults with rrPMBCL received pembrolizumab for up to 2 years or until disease progression or unacceptable toxicity. The primary end points were safety and objective response rate in KEYNOTE-013 and objective response rate in KEYNOTE-170. Secondary end points included duration of response, progression-free survival, overall survival, and safety. Exploratory end points included association between biomarkers and pembrolizumab activity. RESULTS The objective response rate was 48% (7 complete responses; 33%) among 21 patients in KEYNOTE-013 and 45% (7 complete responses; 13%) among 53 patients in KEYNOTE-170. After a median follow-up time of 29.1 months in KEYNOTE-013 and 12.5 months in KEYNOTE-170, the median duration of response was not reached in either study. No patient with complete response experienced progression, including 2 patients with complete response for at least 1 year off therapy. Treatment-related adverse events occurred in 24% of patients in KEYNOTE-013 and 23% of patients in KEYNOTE-170. There were no treatment-related deaths. Among 42 evaluable patients, the magnitude of the 9p24 gene abnormality was associated with PD-L1 expression, which was itself significantly associated with progression-free survival. CONCLUSION Pembrolizumab is associated with high response rate, durable activity, and a manageable safety profile in patients with rrPMBCL.

Primary central nervous system large B-cell lymphoma (PCNS-LBCL) is a rare, aggressive lymphoma that affects immune-privileged sites. Immune checkpoint molecules have been implicated in its aggressive biology, and promising results have emerged from immune checkpoint inhibitor therapy in relapsed/refractory cases. This study evaluates the tumor microenvironment (TME), immune checkpoint molecule expression, and the relationship with 9p24.1 gene region alterations in a large cohort of PCNS-LBCL, with detailed quantitative analyses. Tissue microarrays were constructed with 57 PCNS-LBCL cases and 45 systemic non-germinal center diffuse large B-cell lymphoma (DLBCL) controls. Immunostaining for CD3, CD8, CD68, CD163, PD-1, PD-L1, PD-L2, EBER in situ hybridization (ISH), and FISH for PD-L1/PD-L2 copy number alterations and translocations were performed. Digital image analysis was used for quantitative evaluations, which were compared with clinical and pathologic parameters. PCNS-LBCL showed significantly lower T-cell and histiocyte presence in the TME compared with nodal DLBCL (P<0.001), independent of preoperative steroid therapy. Cytotoxic T-cell ratio was higher in PCNS-LBCL (P<0.001). PD-1, PD-L1, and PD-L2 expressions in the TME of PCNS-LBCL were 89%, 96%, and 90%, respectively, and were positively correlated with TME density. Tumor cell expressions of PD-L1 and PD-L2 were 31% and 34%, respectively. FISH alterations in the 9p24.1 region were infrequent and did not consistently correlate with protein expression in either PCNS-LBCL or DLBCL. Higher CD8+ T-cell and CD68+ histiocyte counts were associated with better survival in PCNS-LBCL. Lower TME density and high expression of PD-1/PD-L1/PD-L2 in PCNS-LBCL reflect the unique CNS microanatomy and may contribute to poorer prognosis. These findings support the potential benefit of immune checkpoint inhibitors in treating PCNS-LBCL, aligning with ongoing clinical trials and current literature.

Copy number alterations (CNAs) of 9p24.1 occur frequently in Hodgkin lymphoma, primary mediastinal large B-cell lymphoma (PMBCL), primary central nervous system lymphoma, and primary testicular lymphoma, resulting in overexpression of PD-L1 and sensitivity to PD-1 blockade-based immunotherapy. While 9p24.1 CNA was also reported in diffuse large B-cell lymphoma (DLBCL), little is known about its molecular or clinical significance. In this study, we analyzed the prevalence of 9p24.1 CNA in newly diagnosed DLBCL and examined its association with PD-L1, PD-L2, and JAK2 expression, clinical characteristics, and outcome. We found that 10% of DLBCL cases had CNA of 9p24.1, with 6.5% gains, and 3.5% amplifications. Only the cases with a 9p24.1 amplification had high levels of PD-L1, PD-L2, and JAK2 expression. Gains or amplifications of 9p24.1 were associated with a younger age and the ABC/non-GCB subtype. Compared with DLBCL cases without 9p24.1 CNA, the cases with a 9p24.1 amplification had a trend of better event-free survival. Furthermore, the amplification cases had a gene expression and mutation profile similar to those of PMBCL. Our data suggest that amplification of 9p24.1 identifies a unique subset of DLBCL with clinical and molecular features resembling PMBCL that may be amenable to PD-1 blockade-based immunotherapy.

Epstein-Barr virus (EBV) associated diffuse large B-cell lymphoma (DLBCL) represents a rare aggressive B-cell lymphoma subtype characterized by an adverse clinical outcome. EBV infection of lymphoma cells has been associated with different lymphoma subtypes while the precise role of EBV in lymphomagenesis and specific molecular characteristics of these lymphomas remain elusive. To further unravel the biology of EBV associated DLBCL, we present a comprehensive molecular analysis of overall 60 primary EBV positive (EBV+) DLBCLs using targeted sequencing of cancer candidate genes (CCGs) and genome-wide determination of recurrent somatic copy number alterations (SCNAs) in 46 cases, respectively. Applying the LymphGen classifier 2.0, we found that less than 20% of primary EBV + DLBCLs correspond to one of the established molecular DLBCL subtypes underscoring the unique biology of this entity. We have identified recurrent mutations activating the oncogenic JAK-STAT and NOTCH pathways as well as frequent amplifications of 9p24.1 contributing to immune escape by PD-L1 overexpression. Our findings enable further functional preclinical and clinical studies exploring the therapeutic potential of targeting these aberrations in patients with EBV + DLBCL to improve outcome.

Epstein–Barr virus (EBV)‐positive diffuse large B‐cell lymphoma (EBV‐posDLBCL) is an aggressive B‐cell lymphoma that often presents similar morphological and immune phenotype features to that of EBV‐negative DLBCL (EBV‐negDLBCL).

Epstein-Barr virus-positive (EBV+) diffuse large B-cell lymphoma (DLBCL) and EBV+ classic Hodgkin lymphoma (CHL) are major B-cell lymphomas with EBV infection in elderly patients. Although they are regarded as distinct clinicopathologic entities, distinguishing EBV+ CHL from EBV+ DLBCL is often challenging because of their overlapping histological and immunophenotypic features. We characterized the spectrum of EBV+ large B-cell lymphoma in 57 patients aged ≥50 years, including 35 EBV+ DLBCL (12 polymorphic DLBCL [pDLBCL] and 23 monomorphic DLBCL [mDLBCL]) and 22 EBV+ CHL. Gene expression profiling revealed interferon-γ (IFNγ)-enrichment with overexpression of indoleamine 2,3-dioxygenase 1 (IDO1), an immunosuppressive enzyme, in more than half of pDLBCL (5/8), but less in mDLBCL (3/19) and CHL (1/19). Fluorescence in situ hybridization showed a higher frequency of 9p24.1-altered cells in CHL (54%; interquartile range [IQR], 42%-89%), but lower in pDLBCL (18%; IQR, 12%-23%) and mDLBCL (5%; IQR, 0%-30%). Notably, immunohistochemical expression of PDL1 was higher in pDLBCL than in mDLBCL, suggesting IFNγ-mediated upregulation. DLBCL with EBV latency type III (n = 13) exhibited lower tumor PDL1 expression and reduced IDO1-enriched microenvironment. Multivariate analysis of the total cohort revealed that both EBV latency type III and Eastern Cooperative Oncology Group performance status ≥2 were independently associated with shorter overall survival. EBV+ large B-cell lymphoma spectrum was reclassified into four molecular groups: (1) EBV latency type III suggestive of immune senescence (n = 10, 22%), (2) high proportion of 9p24.1-alteration (n = 9, 20%); (3) high IFNγ signature score (n = 9, 20%), and (4) low IFNγ signature score (n = 18, 39%). Moreover, these groups were identified using surrogate immunohistochemical markers: EBNA2, PDL1, and IDO1. In conclusion, the molecular studies assessing tumor-host interaction enhances understanding of the EBV+ large B-cell lymphoma spectrum and benefits pathological diagnosis and clinical management.

… only put on molecular data and PD-L1, without including the tumor microenvironment. … modifications involving 9p24.1 is partly driving overexpression of PD-L1 in EBV-positive DLBCL …

BACKGROUND Recently, unprecedented clinical efficacy was observed during treatment of many solid tumors because of the introduction of programmed cell death 1 (PD-1)/programmed cell death 1 ligand 1 (PD-L1) immune checkpoint inhibitors. Preliminary clinical data indicates that checkpoint inhibition also represents a promising therapeutic strategy for certain lymphoid malignancies. However, PD-1/PD-L1 expression levels on neoplastic cells and in the tumor microenvironment vary among subtypes and their prognostic implications remain uncertain. MAIN BODY Here, we review the clinicopathological significance of PD-1/PD-L1 expression in lymphomas. Increased infiltration of PD-1+ tumor-infiltrating lymphocytes (TILs) is a favorable prognostic factor in diffuse large B-cell lymphoma (DLBCL) but not in Hodgkin's lymphoma (HL). Higher numbers of PD-1+ TILs were observed in follicular lymphoma (FL) than in other subtypes of B-cell lymphoma; however, its prognostic significance remains controversial. Infiltration of PD-L1+ immune cells showed a trend toward better overall survival in nasal natural killer (NK)/T-cell lymphoma and adult T-cell leukemia/lymphoma, more likely to be classified as activated macrophages and dendritic cells in microenvironment but its biological effect is not clarified. Peripheral PD-1+ T cells could be detected in blood samples from DLBCL and chronic lymphocytic leukemia (CLL) and correlated with disease progression and poor prognosis. PD-1+ neoplastic T cells were more frequently observed in cutaneous T-cell lymphoma, including Sézary syndrome and mycosis fungoides, which may be involved in the progression of epithelial-derived T lymphoma. Studies on PD-L1 expression in neoplastic cells mostly focused on DLBCL. PD-L1+ neoplastic cells were observed only in a small subset of DLBCL, mainly associated with activated B cell (ABC) subtypes and Epstein-Barr virus (EBV) positivity; however, its prognostic role remains controversial. In either T or B lymphoma, elevated serum or plasma levels of soluble PD-L1 represent adverse prognostic factors. Notably, in clinical trials of classical HL, the frequency of 9p24.1 chromosome alterations increases the abundance of PD-1 ligand expression, appearing to predict responses to anti-PD-1/PD-L1 therapy. The cytogenetic alterations affecting chromosome 9p24.1 including the CIITA rearrangement were also frequently observed in certain specific subtypes of large B-cell lymphomas. CONCLUSIONS The clinical roles of PD-1/PD-L1 expression vary between subtypes of lymphoma. Future studies should delineate the prognostic and predictive roles of PD-1 and PD-L1 expression.

Introduction: The activity of immune checkpoint blockade including anti-cytotoxic T-lymphocyte associated protein-4 and anti-programmed cell death protein-1 monoclonal antibodies in hematologic malignancies is limited outside of classical Hodgkin lymphoma and primary mediastinal B-cell lymphoma. Tumor mutational burden (TMB), programmed death ligand-1 (PD-L1) expression, and microsatellite instability-high (MSI-H) are well-established biomarkers predicting response to checkpoint blockade in solid malignancies. In addition, tumors with high TMB, defined as ≥10 mutations/megabase (mut/Mb), and/or MSI-H are Food and Drug Administration (FDA) approved tissue agnostic biomarkers for treatment with pembrolizumab. The frequencies of high TMB, MSI-H, and expression pattern of PD-L1 across specific hematologic malignancies are undefined. Methods: Patients with hematologic malignancies who had next generation sequencing (NGS) performed by Foundation One Heme were identified. TMB and MSI were measured by NGS. TMB was classified as high if ≥10 mut/Mb and low if <10 mut/Mb. When available, PD-L1 expression on tumor cells by immunohistochemistry (IHC) was also collected. PD-L1 IHC was performed with either Ventana (SP142) PD-L1 antibody or Dako (22C3) PD-L1 antibody. Scores were classified as high (≥ 50%), low (1-49%), and negative (<1%) based on the percent of tumor cells staining positive for PD-L1. Pathology reports were retrospectively re-reviewed to determine the diagnosis. Results: A total of 390 patients with hematologic malignancies with NGS were identified. Forty eight of the 390 samples (12%) had a high TMB (Table 1). Twenty five of 45 (56%) patients with DLBCL had a high TMB (Table 2). The TMB was low in all myeloid malignancies tested. None of the 302 samples tested were MSI-H. PD-L1 IHC was performed on 86 samples. Eleven (13%) had high expression, 26 (30%) had low expression, and 49 (57%) had no expression of PD-L1 on the tumor cells (Table 1). The majority of samples with PD-L1 expression were mature lymphomas (81%). TMB and PD-L1 score had a significant linear relationship (R = 0.22,p= 0.04, 95% CI 0.01 - 0.41) (Figure 1). Conclusion: This study provides detailed characteristics of TMB, MSI status, and PD-L1 expression for hematologic malignancies. Notably, a subset of lymphomas had high TMB and/or PD-L1 expression. Biomarker driven studies of checkpoint blockade in hematologic malignancies with high TMB and/or PD-L1 expression are warranted. Sokol: Foundation Medicine:Current Employment;Roche:Current equity holder in publicly-traded company.Kurzrock:Medimmune:Research Funding;Foundation Medicine:Research Funding;Konica Minolta:Research Funding;IDbyDNA:Current equity holder in private company;Pfizer:Consultancy, Research Funding;Sequenom:Research Funding;Bicara Therapeutics, Inc.:Consultancy;Incyte:Research Funding;Takeda:Research Funding;TopAlliance:Research Funding;Boehringer Ingelheim:Research Funding;CureMatch Inc:Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees;Grifols:Research Funding;Guardant:Research Funding;X Biotech:Consultancy;Neomed:Consultancy;Actuate Therapeutics:Consultancy;Roche:Consultancy;Merck Serono:Research Funding;Genentech:Research Funding;Debiopharm:Research Funding;CureMetrix:Membership on an entity's Board of Directors or advisory committees;OmniSeq:Research Funding;TD2/Volastra:Consultancy;Turning Point Therapeutics:Consultancy.Goodman:EUSA Pharma:Consultancy;Seattle Genetics:Consultancy.

The expression of programmed cell death ligand 1 (PD‐L1) is a biomarker for immunotherapy, but approved detection method is absent in diffuse large B‐cell lymphoma (DLBCL). Here, we performed three methods including immunohistochemistry (IHC) (clone SP263 and SP142), RNAscope, and fluorescence in situ hybridization (FISH) to evaluate PD‐L1 status on a cohort of DLBCL including 94 of DLBCL‐NOS, 25 of primary mediastinal large B‐cell lymphoma (PMBCL) and 7 of double‐hit lymphoma (DHL). SP263 with 25% for immune cell (IC) or combined cell and SP142 with 10% for tumor cell (TC), 20% for both of IC and combined cell were proved to have corresponding survival prognostic. Combined+ showed comparable prognostic value with TC+ and IC+. SP263 and SP142 showed strong concordance (k = 0.788) with combined+ rates of 33.3% (42/126) and 34.9% (44/126), respectively. In DLBCL‐NOS, TC+ by SP263 preferred to non‐GCB and immunoblastic variant DLBCL‐NOS (P = 0.029 and P = 0.004). Combined+ (SP263 and SP142) were associated with more than one extranodal site involved (P = 0.006, P = 0.042), higher ECOG PS scores (P = 0.001, P < 0.001), high IPI risk (P = 0.012, P = 0.005), and poor treatment response (P = 0.095, P = 0.002). IC+ by SP263 and SP142 were both independent risk factors (P = 0.027, P = 0.037). 9p24.1 locus amplification and gain were identified in 4.3% and 7.6% DLBCL‐NOS and indicated shorter overall survival (P = 0.004). Positive rate of PD‐L1 by RNAscope was 36.5%, while no clinical significance shown. PD‐L1 positive rates were all higher in PMBCL and DHL than in DLBCL‐NOS by SP263, SP142, RNAscope, and FISH (P = 0.001, P < 0.001, P = 0.005 and P < 0.001, respectively). In conclusion, combined PD‐L1 expression by IHC was potentially reliable and convenient as a predicting biomarker. SP263 staining was easier to evaluate and recognized more PD‐L1‐stained cells, but SP142 presented a better prognostic indicator. FISH and RNAscope could be used as supplementary assays. PMBCL itself was a sensitive cohort for immunotherapy.

Treatment of cancer has transformed with the introduction of checkpoint inhibitors. However, the majority of solid tumor patients do not respond to checkpoint blockade. In contrast, the response rate to programmed cell death 1 (PD-1) blockade in relapsed/refractory classical Hodgkin lymphoma (cHL) is 65% to 84% which is the highest among all cancers. Currently, checkpoint inhibitors are only approved for cHL and primary mediastinal B-cell lymphoma as the responses to single-agent checkpoint blockade in other hematologic malignancies is disappointingly low. Various established biomarkers such as programmed cell death 1 ligand 1 (PD-L1) protein surface expression, mismatch repair (MMR) status, and tumor mutational burden (TMB) are routinely used in clinical decision-making in solid tumors. In this review, we will explore these biomarkers in the context of hematologic malignancies. We review characteristic 9p24.1 structural alteration in cHL and primary mediastinal B-cell lymphoma (PMBCL) as a basis for response to PD-1 inhibition, as well as the role of antigen presentation pathways. We also explore the reported frequencies of MMR deficiency in various hematologic malignancies and investigate TMB as a predictive marker.

ABSTRACT Diffuse large B-cell lymphoma (DLBCL) is the most common type of lymphoma with high mutation burdens but a low response rate to immune checkpoint inhibitors. In this study, we performed targeted next-generation sequencing and fluorescent multiplex immunohistochemistry, and investigated the clinical significance and immunological effect of mutation numbers in 424 DLBCL patients treated with standard immunochemotherapy. We found that KMT2D and TP53 nonsynonymous mutations (MUT) were significantly associated with increased nonsynonymous mutation numbers, and that high mutation numbers (MUThigh) were associated with significantly poorer clinical outcome in germinal center B-cell-like DLBCL with wild-type TP53. To understand the underlying mechanisms, we identified a gene-expression profiling signature and the association of MUThigh with decreased T cells in DLBCL patients with wild-type TP53. On the other hand, in overall cohort, MUThigh was associated with lower PD-1 expression in T cells and PD-L1 expression in macrophages, suggesting a positive role of MUThigh in immune responses. Analysis in a whole-exome sequencing dataset of 304 patients deposited by Chapuy et al. validated the correlation of MUT-KMT2D with genomic complexity and the significantly poorer survival associated with higher numbers of genomic single nucleotide variants in activated B-cell–like DLBCL with wild-type TP53. Together, these results suggest that KMT2D inactivation or epigenetic dysregulation has a role in driving DLBCL genomic instability, and that genomic complexity has adverse impact on clinical outcome in DLBCL patients with wild-type TP53 treated with standard immunochemotherapy. The oncoimmune data in this study have important implications for biomarker and therapeutic studies in DLBCL.

Immune checkpoint blockade (ICB) has revolutionized the prognosis of several advanced-stage solid tumors. However, its success has been far more limited in hematological malignancies and is mostly restricted to classical Hodgkin lymphoma (cHL) and primary mediastinal B cell lymphoma (PMBCL). In patients with non-Hodgkin lymphoma (NHL), response to PD-1/PD-L1 ICB monotherapy has been relatively limited, although some subtypes are more sensitive than others. Numerous predictive biomarkers have been investigated in solid malignancies, such as PD-L1 expression, tumor mutational burden (TMB) and microsatellite instability (MSI), among others. This review aims to appraise the current knowledge on PD-1/PD-L1 ICB efficacy in lymphoma when used either as monotherapy or combined with other agents, and describes potential biomarkers of response in this specific setting.

Purpose: The growth of non–Hodgkin lymphomas can be influenced by tumor–immune system interactions. Cytotoxic T-lymphocyte antigen 4 (CTLA-4) is a negative regulator of T-cell activation that serves to dampen antitumor immune responses. Blocking anti–CTLA-4 monoclonal antibodies improves host resistance to immunogenic tumors, and the anti–CTLA-4 antibody ipilimumab (MDX-010) has clinical activity against melanoma, prostate, and ovarian cancers. Experimental Design: We did a phase I trial of ipilimumab in patients with relapsed/refractory B-cell lymphoma to evaluate safety, immunologic activity, and potential clinical efficacy. Treatment consisted of ipilimumab at 3 mg/kg and then monthly at 1 mg/kg × 3 months (dose level 1), with subsequent escalation to 3 mg/kg monthly × 4 months (dose level 2). Results: Eighteen patients were treated, 12 at the lower dose level and 6 at the higher dose level. Ipilimumab was generally well tolerated, with common adverse events attributed to it, including diarrhea, headache, abdominal pain, anorexia, fatigue, neutropenia, and thrombocytopenia. Two patients had clinical responses; one patient with diffuse large B-cell lymphoma had an ongoing complete response (&amp;gt;31 months), and one with follicular lymphoma had a partial response lasting 19 months. In 5 of 16 cases tested (31%), T-cell proliferation to recall antigens was significantly increased (&amp;gt;2-fold) after ipilimumab therapy. Conclusions: Blockade of CTLA-4 signaling with the use of ipilimumab is well tolerated at the doses used and has antitumor activity in patients with B-cell lymphoma. Further evaluation of ipilimumab alone or in combination with other agents in B-cell lymphoma patients is therefore warranted. (Clin Cancer Res 2009;15(20):6446–53)

PURPOSE Treatment options are limited for patients with relapsed/refractory diffuse large B-cell lymphoma (DLBCL). Tumor cells can exploit the programmed death-1 checkpoint pathway to evade immune surveillance. In the current study, we evaluated the efficacy and safety of programmed death-1 blockade by nivolumab in patients with relapsed/refractory DLBCL. METHODS In this phase II, open-label study, patients with relapsed/refractory DLBCL who were ineligible for autologous hematopoietic cell transplantation (auto-HCT) or who had experienced failure with auto-HCT received nivolumab 3 mg/kg every 2 weeks. We assessed the efficacy and safety of nivolumab as well as genetic alterations of 9p24.1. RESULTS Among 121 treated patients, patients in the auto-HCT-failed cohort (n = 87) received a median of four nivolumab doses and a median of three doses were administered to those in the auto-HCT-ineligible cohort (n = 34). At a median follow-up of 9 months in the auto-HCT-failed cohort and 6 months in the auto-HCT-ineligible cohort, independently assessed objective response rates were 10% and 3%, and median durations of response were 11 and 8 months, respectively. Median progression-free survival and overall survival were 1.9 and 12.2 months in the auto-HCT-failed cohort and 1.4 and 5.8 months in the auto-HCT-ineligible cohort respectively. All three patients with complete remission-3% of the auto-HCT-failed cohort-had durable response (11 or more, 14 or more, and 17 months). Treatment-related grade 3 and 4 adverse events were reported in 24% of patients. The most common were neutropenia (4%), thrombocytopenia (3%), and increased lipase (3%). Of all evaluable samples for 9p24.1 analysis, 16% exhibited low-level copy gain and 3% had amplification. CONCLUSION Nivolumab monotherapy is associated with a favorable safety profile but a low overall response rate among patients with DLBCL who are ineligible for auto-HCT or who experienced failure with auto-HCT. Genetic alterations of 9p24.1 are infrequent in DLBCL.

… , PD-L1, and CTLA-4 gene polymorphisms, along with their mRNA and protein expression, in DLBCL … In an early phase I trial, nivolumab achieved an overall response rate of 36% and a …

Highlights • PD-1 may probably be the more attractive ICI target rather than PD-L1 and CTLA-4 in NHL cases.• In T-NHLs, MF and ENKTL had the best responses to PD-1 inhibitors.• In B-NHLs, RT and PMBCL had the best responses to PD-1 inhibitors.• Neither FL nor DLBCL had satisfactory responses to ICI monotherapy.• ICIs have favorable safety profile and adequate toleration in NHL cases.

&lt;div&gt;Abstract&lt;p&gt;PD-1/L1 and CTLA-4 blockade immunotherapies have been approved for 13 types of cancers and are being studied in diffuse large B-cell lymphoma (DLBCL), the most common aggressive B-cell lymphoma. However, whether both PD-1 and CTLA-4 checkpoints are active and clinically significant in DLBCL is unknown. Whether PD-1 ligands expressed by tumor cells or by the microenvironment of DLBCL are critical for the PD-1 immune checkpoint is unclear. We performed immunophenotypic profiling for 405 patients with &lt;i&gt;de novo&lt;/i&gt; DLBCL using a MultiOmyx immunofluorescence platform and simultaneously quantitated expression/coexpression of 13 immune markers to identify prognostic determinants. In both training and validation cohorts, results demonstrated a central role of the tumor immune microenvironment, and when its functionality was impaired by deficiency in tumor-infiltrating T cells and/or natural killer cells, high PD-1 expression (but not CTLA-4) on CD8&lt;sup&gt;+&lt;/sup&gt; T cells, or PD-L1 expression on T cells and macrophages, patients had significantly poorer survival after rituximab–CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisone) immunochemotherapy. In contrast, tumor-cell PD-L2 expression was associated with superior survival, as well as PD-L1&lt;sup&gt;+&lt;/sup&gt;CD20&lt;sup&gt;+&lt;/sup&gt; cells proximal (indicates interaction) to PD-1&lt;b&gt;&lt;sup&gt;+&lt;/sup&gt;&lt;/b&gt;CD8&lt;sup&gt;+&lt;/sup&gt; T cells in patients with low PD-1&lt;b&gt;&lt;sup&gt;+&lt;/sup&gt;&lt;/b&gt; percentage of CD8&lt;sup&gt;+&lt;/sup&gt; T cells. Gene-expression profiling results suggested the reversibility of T-cell exhaustion in PD-1&lt;sup&gt;+&lt;/sup&gt;/PD-L1&lt;sup&gt;+&lt;/sup&gt; patients with unfavorable prognosis and implication of &lt;i&gt;LILRA&lt;/i&gt;/&lt;i&gt;B, IDO1, CHI3L1&lt;/i&gt;, and &lt;i&gt;SOD2&lt;/i&gt; upregulation in the microenvironment dysfunction with PD-L1 expression. This study comprehensively characterized the DLBCL immune landscape, deciphered the differential roles of various checkpoint components in rituximab–CHOP resistance in DLBCL patients, and suggests targets for PD-1/PD-L1 blockade and combination immunotherapies.&lt;/p&gt;&lt;/div&gt;

Introduction The tumor microenvironment (TME) influences diffuse large B-cell lymphoma (DLBCL) progression, but the prognostic roles of tumor-infiltrating T-lymphocytes (TIL-T), tumor-associated macrophages (TAMs), and PD-L1 remain undefined. This study investigates the clinicopathological associations and prognostic impacts of TIL-T, TAMs, and PD-L1 expression in DLBCL. Methods This retrospective study evaluated 89 primary DLBCL cases, integrating clinicopathological data with automated immunohistochemical quantification of CD3, CD8, FOXP3, CD163, and PD-L1 expression in tumor hotspots and microenvironmental compartments. Prognostic associations of TIL-T, TAMs, and PD-L1 expression with PFS and OS were analyzed via Kaplan-Meier methods and Cox regression. Results High CD3+ infiltration correlated with lower Ki-67 expression, while elevated FOXP3+ levels linked to improved Eastern Cooperative Oncology Group Performance Status (ECOG). CD163+ TAMs varied by NCCN-IPI risk, ECOG, and cell of origin. Neoplastic PD-L1 (nPD-L1) positivity associated with higher NCCN-IPI scores, CD3+ T-cell infiltration, and CD163+ TAM enrichment. Microenvironmental PD-L1 (mPD-L1) correlated with age, ECOG, B symptoms, and infiltration of all T-cell subsets and TAMs. Survival analysis revealed prolonged overall survival (OS) with high CD3+, CD8+, FOXP3+ TIL-T, CD163+ TAMs, or mPD-L1 positivity, while progression-free survival (PFS) improved with CD3+ infiltration and mPD-L1. Univariate analysis identified B symptoms, extranodal involvement, and low TIL-T levels as OS risks, whereas ECOG 0 and mPD-L1+ were protective. Multivariate modeling confirmed B symptoms, extranodal disease, and CD3+ TIL-T as independent OS predictors; CD3+ TIL-T and B symptoms independently impacted PFS. Discussion The TME plays a crucial role in the biological behavior of DLBCL, particularly because TIL-T and TAMs are significantly associated with patient survival outcomes. These cell types may serve as critical biomarkers and provide novel immunotherapy targets in DLBCL.

Background & aims The immune checkpoint recently provides a new strategy for the immunotherapy of malignant tumors. However, the role in the immune microenvironment of DLBCL is not completely clear. Methods We detected the expression of PD-1, LAG-3, TIM-3, and TIGIT on TILs and on tumor cells among 174 DLBCL patients by IHC. Results In TILs, the positive rates of PD-1, LAG-3, TIM-3 and TIGIT were 79.3%, 78.8%, 62.7% and 69.5%, respectively.TIM-3 and TIGIT were expressed in 44.8% and 45.4% of tumor cells. The expression of TIM-3 in TILs was significantly correlated with the Ann-Arbor stage (P=0.039). There was a positive correlation Between PD-1 and LAG-3 or TIM-3 and TIGIT.In addition, LAG-3 expression in TILs was associated with inferior prognosis.Multivariate analysis showed that PS score and R-CHOP therapy were independent risk factors for OS and PFS in patients with DLBCL (P=0.000). Conclusions The expression level of TIM-3 is closely related to the Ann-Arbor stage, which may be expected to be a new index to evaluate the invasiveness of DLBCL. PD-1 was correlated with the expression of LAG-3, and the high expression of LAG-3 and LAG-3/PD-1 predicted the poor prognosis of DLBCL. Therefore, LAG-3 may become a new target of immunotherapy, or be used in combination with PD-1 inhibitors to improve the drug resistance of current patients with DLBCL.

Programmed death ligand 1 (PD-L1) is expressed by 20% to 57% of systemic diffuse large B cell lymphomas (DLBCLs). PD-L1 expression in primary cutaneous DLBCL (pcDLBCL) has not been studied so far. Sixteen paraffin-embedded tissue samples of pcDLBCL (13 leg type [LT], 3 others [OT]) were investigated for PD-1, PD-L1, and CD33 expression and the cellular composition of the tumor microenvironment, focusing on myeloid-derived suppressor cells (MDSCs) and tumor-associated macrophages. Membrane-bound PD-L1 expression by the tumor cells was observed in all samples, albeit to a variable extent (19.9%). As expected, most DLBCL-LT (10 cases) were classified as activated B cell like type, with a higher PD-L1 score (21.9%) compared with that of the germinal center B cell like type (7.7%). The surrounding infiltrate consisted predominately of CD163(+) M2 rather than CD68(+) macrophages (CD68:CD163=1:4 to 6). Moreover, a considerable proportion of CD33(+) MDSCs with PD-L1 coexpression was admixed. Tumor cells expressed CD33 to variable degrees (2% to 60%). The number of MDSCs or M2 macrophages did not correlate with pcDLBCL subtypes LT or OT. T cells were only a minor component of the tumor microenvironment. We propose that PD-L1(+) tumor cells and PD-L1(+) MDSCs shield the tumor against PD-1(+) tumor-infiltrating lymphocytes, consequently leading to inhibition and diminution of tumor-infiltrating lymphocytes. Moreover, we found a polarization to M2 macrophages, which may contribute to the poor prognosis of DLBCL patients. Thus, targeting of tumor cells and MDSCs using anti-PD-1/anti-PD-L1 or anti-CD33 antibodies might be a worthwhile new approach to treat this aggressive form of cutaneous B-cell lymphoma.

Background Anaplastic diffuse large B-cell lymphoma(A-DLBCL) is a rare morphological subtype characterized by the presence of polygonal, bizarre-shaped tumor cells. Our previous research found that A-DLBCL displays many genetic alterations and biological features that differ greatly from those of ordinary DLBCL. However, the status of tumor immune microenvironment components and checkpoint molecules in A-DLBCL remains unclear. Methods Thirty A-DLBCL patients were enrolled to study tumor immune microenvironment components and checkpoint molecules and their associations with clinicopathological features and prognosis. Results Patients with A-DLBCL presented higher expression of PD-L1 (40% vs 10%, P=0.004) than patients with ordinary DLBCL. FISH analysis showed that extra copies of PD-L1 were more frequent in A-DLBCL cases than in ordinary DLBCL cases (23.3% vs 4.0%, P=0.001). The numbers of PD-1+ TILs (tumor infiltrating lymphocytes) and CD8+T cells were significantly lower in A-DLBCL versus ordinary DLBCL. In contrast, the numbers of GATA3+ Th2 cells, FOXP3+ Tregs and CD33+ myeloid-derived suppressor cells (MDSCs) were significantly higher in A-DLBCL than in ordinary DLBCL. The associations between clinicopathological features and tumor immune microenvironment cell frequency were analyzed in A-DLBCL patients. Briefly, the number of PD-1+ TILs was lower and the number of CD33+ MDSCs was higher in patients with mutated TP53 compared to those with wild-type TP53. The number of FOXP3+ Tregs was much lower in patients with a noncomplete response (CR) to chemotherapy than in those with a complete response. The number of CD8+ T cells showed a decreasing trend in patients with high International Prognostic Index (IPI) scores and in those with concurrent MYC and BCL2 and/or BCL6 abnormalities. Univariate survival analysis showed that patients with PD-L1+, mPD-L1+(PD-L1+ nonmalignant stromal cells) or mPD-L1+ status had a significantly poorer overall survival (OS) than those with PD-L1- status. An increase in the number of CD3+ T cells, FOXP3+ Treg cells and T-bet+ Th1 cells was significantly associated with prolonged OS in patients with A-DLBCL. Conclusion Our study suggests that A-DLBCL displays a distinct pattern of tumor immune microenvironment components and checkpoint molecules that distinguish it from ordinary DLBCL. The analysis of tumor immune microenvironment components and checkpoint molecules could help in predicting the prognosis of A-DLBCL patients and determining therapeutic strategies targeting the tumor immune microenvironment.

The tumour microenvironment (TME), which is modulated after immune‐chemotherapy, is involved in tumour growth and metastasis. Programmed cell death 1 (PD‐1) expressed on tumour‐infiltrating non‐malignant cells plays an important role in the TME through the PD‐1/programmed cell death ligand 1 (PD‐L1) signalling pathway. However, its impact in patients with relapsed or refractory (R/R) diffuse large B‐cell lymphoma (DLBCL) remains unclear.

Abstract PD-1/PD-L1 immune checkpoint inhibitors (ICIs) demonstrate promising therapeutic potential in diffuse large B-cell lymphoma (DLBCL). However, a subset of DLBCL patients exhibits primary or acquired resistance to PD-1 blockade. This study investigated the mechanistic role of β2-microglobulin (B2M) mutations in modulating PD-L1 expression and their contribution to ICI resistance in DLBCL. The association between B2M mutations and PD-L1 expression in DLBCL patient samples was analyzed via immunohistochemistry. A B2M-knockout DLBCL cell line model was established, followed by evaluation of PD-L1 and MHC-I expression changes through qRT-PCR, western blot, and flow cytometry. T cell activation (CD69/CD25 markers) and cytotoxic activity (lactate dehydrogenase release) were assessed in PBMC co-culture experiments. Mechanistic studies employing MHC-I inhibitors and IFN-γ stimulation were conducted to dissect the regulatory relationship between MHC-I and PD-L1. Co-culture systems with PD-1 blockade was performed to validate the impact of B2M on PD-1 inhibitor resistance. The results demonstrated that B2M knockout significantly downregulated both PD-L1 mRNA and protein levels, concomitant with impaired MHC-I complex assembly and reduced TAP1/2 expression. B2M-deficient cells failed to activate CD8+ T cells, evidenced by diminished CD69+/CD25+ surface expression and reduced cytotoxic efficiency. Notably, B2M ablation abolished IFN-γ-induced PD-L1 upregulation (p < 0.05), demonstrating that intact MHC-I functionality is a prerequisite for PD-L1 expression. Further co-culture systems with PD-1 blockade validated the functional roles of MHC-I and PD-L1 in modulating PD-1 inhibitor resistance mediated by B2M. In conclusion, this work indicated that B2M mutations synergistically drive DLBCL resistance to PD-1 inhibitors through dual mechanisms-suppressing PD-L1 expression and completely disabling antigen presentation. The study highlights that combined strategies restoring MHC-I expression and targeting PD-L1 may overcome immunotherapy resistance, offering a novel direction for precision therapy in DLBCL.

… Hence, understanding the basis underlying treatment resistance in DLBCL is a pressing need … , tumor microenvironment (TME) and host variabilities for treatment resistance in DLBCL. …

In tumor microenvironment, the programmed death 1 (PD-1) immune checkpoint has a crucial role of mechanism of T cell exhaustion leading to tumor evasion. Ligands of PD-1, programmed death ligand 1/2 (PD-L1/L2) are over-expressed in tumor cells and participate in prolonged tumor progression and survivals. Recently, clinical trials for patients who failed to obtain an optimal response prior to standardized chemotherapy in several solid cancers have been focused on targeting therapy against PD-1 to reduce disease progression rates and prolonged survivals. Since various inhibitors targeting the immune checkpoint in PD-1/PD-L1 pathway in solid cancers have been introduced, promising approach using anti-PD-1 antibodies were attempted in several types of hematologic malignances. In diffuse large B cell lymphoma (DLBCL) as the most common and aggressive B cell type of non-Hodgkin’s lymphoma, anti-PD-1 and anti-PD-L1 antibodies were studies in various clinical trials. In this review, we summarized the results of several studies associated with PD-1/PD-L1 pathway as an immune evasion mechanism and described clinical trials about targeting therapy against PD-1/PD-L1 pathway in DLBCL.