Genetic and Functional Drivers of Diffuse Large B Cell Lymphoma.

Diffuse large B cell lymphoma (DLBCL) is the most common form of blood cancer and is characterized by a striking degree of genetic and clinical heterogeneity. This heterogeneity poses a major barrier to understanding the genetic basis of the disease and its response to therapy. Here, we performed an integrative analysis of whole-exome sequencing and transcriptome sequencing in a cohort of 1,001 DLBCL patients to comprehensively define the landscape of 150 genetic drivers of the disease. We characterized the functional impact of these genes using an unbiased CRISPR screen of DLBCL cell lines to define oncogenes that promote cell growth. A prognostic model comprising these genetic alterations outperformed current established methods: cell of origin, the International Prognostic Index comprising clinical variables, and dual MYC and BCL2 expression. These results comprehensively define the genetic drivers and their functional roles in DLBCL to identify new therapeutic opportunities in the disease.

Follicular lymphoma triggers phenotypic and functional remodeling of the human lymphoid stromal cell landscape.

Lymphoid stromal cells (LSCs) are essential organizers of immune responses. We analyzed tonsillar tissue by combining flow cytometry, in situ imaging, RNA sequencing, and functional assays, defining three distinct human LSC subsets. The integrin CD49a designated perivascular stromal cells exhibiting features of local committed LSC precursors and segregated cytokine and chemokine-producing fibroblastic reticular cells (FRCs) supporting B and T cell survival. The follicular dendritic cell transcriptional profile reflected active responses to B cell and non-B cell stimuli. We therefore examined the effect of B cell stimuli on LSCs in follicular lymphoma (FL). FL B cells interacted primarily with CD49a+ FRCs. Transcriptional analyses revealed LSC reprogramming in situ downstream of the cytokines tumor necrosis factor (TNF) and transforming growth factor ß (TGF-ß), including increased expression of the chemokines CCL19 and CCL21. Our findings define human LSC populations in healthy tissue and reveal bidirectional crosstalk between LSCs and malignant B cells that may present a targetable axis in lymphoma.

Genome-wide Screens Identify Lineage- and Tumor-Specific Genes Modulating MHC-I- and MHC-II-Restricted Immunosurveillance of Human Lymphomas.

Tumors frequently subvert major histocompatibility complex class I (MHC-I) peptide presentation to evade CD8+ T cell immunosurveillance, though how this is accomplished is not always well defined. To identify the global regulatory networks controlling antigen presentation, we employed genome-wide screening in human diffuse large B cell lymphomas (DLBCLs). This approach revealed dozens of genes that positively and negatively modulate MHC-I cell surface expression. Validated genes clustered in multiple pathways including cytokine signaling, mRNA processing, endosomal trafficking, and protein metabolism. Genes can exhibit lymphoma subtype- or tumor-specific MHC-I regulation, and a majority of primary DLBCL tumors displayed genetic alterations in multiple regulators. We established SUGT1 as a major positive regulator of both MHC-I and MHC-II cell surface expression. Further, pharmacological inhibition of two negative regulators of antigen presentation, EZH2 and thymidylate synthase, enhanced DLBCL MHC-I presentation. These and other genes represent potential targets for manipulating MHC-I immunosurveillance in cancers, infectious diseases, and autoimmunity.

A Hyper-IgM Syndrome Mutation in Activation-Induced Cytidine Deaminase Disrupts G-Quadruplex Binding and Genome-wide Chromatin Localization.

Precise targeting of activation-induced cytidine deaminase (AID) to immunoglobulin (Ig) loci promotes antibody class switch recombination (CSR) and somatic hypermutation (SHM), whereas AID targeting of non-Ig loci can generate oncogenic DNA lesions. Here, we examined the contribution of G-quadruplex (G4) nucleic acid structures to AID targeting in vivo. Mice bearing a mutation in Aicda (AIDG133V) that disrupts AID-G4 binding modeled the pathology of hyper-IgM syndrome patients with an orthologous mutation, lacked CSR and SHM, and had broad defects in genome-wide AIDG133V chromatin localization. Genome-wide analyses also revealed that wild-type AID localized to MHCII genes, and AID expression correlated with decreased MHCII expression in germinal center B cells and diffuse large B cell lymphoma. Our findings indicate a crucial role for G4 binding in AID targeting and suggest that AID activity may extend beyond Ig loci to regulate the expression of genes relevant to the physiology and pathology of activated B cells.

Immunization with Components of the Viral Fusion Apparatus Elicits Antibodies That Neutralize Epstein-Barr Virus in B Cells and Epithelial Cells.

Epstein-Barr virus (EBV) causes infectious mononucleosis and is associated with epithelial-cell cancers and B cell lymphomas. An effective EBV vaccine is not available. We found that antibodies to the EBV glycoprotein gH/gL complex were the principal components in human plasma that neutralized infection of epithelial cells and that antibodies to gH/gL and gp42 contributed to B cell neutralization. Immunization of mice and nonhuman primates with nanoparticle vaccines that displayed components of the viral-fusion machinery EBV gH/gL or gH/gL/gp42 elicited antibodies that potently neutralized both epithelial-cell and B cell infection. Immune serum from nonhuman primates inhibited EBV-glycoprotein-mediated fusion of epithelial cells and B cells and targeted an epitope critical for virus-cell fusion. Therefore, unlike the leading EBV gp350 vaccine candidate, which only protects B cells from infection, these EBV nanoparticle vaccines elicit antibodies that inhibit the virus-fusion apparatus and provide cell-type-independent protection from virus infection.

Unique Immune Cell Coactivators Specify Locus Control Region Function and Cell Stage.

Locus control region (LCR) functions define cellular identity and have critical roles in diseases such as cancer, although the hierarchy of structural components and associated factors that drive functionality are incompletely understood. Here we show that OCA-B, a B cell-specific coactivator essential for germinal center (GC) formation, forms a ternary complex with the lymphoid-enriched OCT2 and GC-specific MEF2B transcription factors and that this complex occupies and activates an LCR that regulates the BCL6 proto-oncogene and is uniquely required by normal and malignant GC B cells. Mechanistically, through OCA-B-MED1 interactions, this complex is required for Mediator association with the BCL6 promoter. Densely tiled CRISPRi screening indicates that only LCR segments heavily bound by this ternary complex are essential for its function. Our results demonstrate how an intimately linked complex of lineage- and stage-specific factors converges on specific and highly essential enhancer elements to drive the function of a cell-type-defining LCR.

Deficiency of factor-inhibiting HIF creates a tumor-promoting immune microenvironment.

Hypoxia signaling influences tumor development through both cell-intrinsic and -extrinsic pathways. Inhibiting hypoxia-inducible factor (HIF) function has recently been approved as a cancer treatment strategy. Hence, it is important to understand how regulators of HIF may affect tumor growth under physiological conditions. Here we report that in aging mice factor-inhibiting HIF (FIH), one of the most studied negative regulators of HIF, is a haploinsufficient suppressor of spontaneous B cell lymphomas, particular pulmonary B cell lymphomas. FIH deficiency alters immune composition in aged mice and creates a tumor-supportive immune environment demonstrated in syngeneic mouse tumor models. Mechanistically, FIH-defective myeloid cells acquire tumor-supportive properties in response to signals secreted by cancer cells or produced in the tumor microenvironment with enhanced arginase expression and cytokine-directed migration. Together, these data demonstrate that under physiological conditions, FIH plays a key role in maintaining immune homeostasis and can suppress tumorigenesis through a cell-extrinsic pathway.

KMT2D acetylation by CREBBP reveals a cooperative functional interaction at enhancers in normal and malignant germinal center B cells.

Heterozygous inactivating mutations of the KMT2D methyltransferase and the CREBBP acetyltransferase are among the most common genetic alterations in B cell lymphoma and co-occur in 40 to 60% of follicular lymphoma (FL) and 30% of EZB/C3 diffuse large B cell lymphoma (DLBCL) cases, suggesting they may be coselected. Here, we show that combined germinal center (GC)-specific haploinsufficiency of Crebbp and Kmt2d synergizes in vivo to promote the expansion of abnormally polarized GCs, a common preneoplastic event. These enzymes form a biochemical complex on select enhancers/superenhancers that are critical for the delivery of immune signals in the GC light zone and are only corrupted upon dual Crebbp/Kmt2d loss, both in mouse GC B cells and in human DLBCL. Moreover, CREBBP directly acetylates KMT2D in GC-derived B cells, and, consistently, its inactivation by FL/DLBCL-associated mutations abrogates its ability to catalyze KMT2D acetylation. Genetic and pharmacologic loss of CREBBP and the consequent decrease in KMT2D acetylation lead to reduced levels of H3K4me1, supporting a role for this posttranslational modification in modulating KMT2D activity. Our data identify a direct biochemical and functional interaction between CREBBP and KMT2D in the GC, with implications for their role as tumor suppressors in FL/DLBCL and for the development of precision medicine approaches targeting enhancer defects induced by their combined loss.

USP36 inhibits apoptosis by deubiquitinating cIAP1 and survivin in colorectal cancer cells.

Chemotherapeutic agents for treating colorectal cancer (CRC) primarily induce apoptosis in tumor cells. The ubiquitin-proteasome system is critical for apoptosis regulation. Deubiquitinating enzymes (DUBs) remove ubiquitin from substrates to reverse ubiquitination. Although over 100 DUB members have been discovered, the biological functions of only a small proportion of DUBs have been characterized. Here, we aimed to systematically identify the DUBs that contribute to the development of CRC. Among the DUBs, ubiquitin-specific protease 36 (USP36) is upregulated in CRC. We showed that the knockdown of USP36 induces intrinsic and extrinsic apoptosis. Through gene silencing and coimmunoprecipitation techniques, we identified survivin and cIAP1 as USP36 targets. Mechanistically, USP36 binds and removes lysine-11-linked ubiquitin chains from cIAP1 and lysine-48-linked ubiquitin chains from survivin to abolish protein degradation. Overexpression of USP36 disrupts the formation of the XIAP-second mitochondria-derived activator of caspase complex and promotes receptor-interacting protein kinase 1 ubiquitination, validating USP36 as an inhibitor to intrinsic and extrinsic apoptosis through deubiquitinating survivin and cIAP1. Therefore, our results suggest that USP36 is involved in CRC progression and is a potential therapeutic target.

The fifth edition of the WHO classification of mature B-cell neoplasms: open questions for research.

The fifth edition of the World Health Organization Classification of Haematolymphoid Tumours (WHO-HAEM5) is the product of an evidence-based evolution of the revised fourth edition with wide multidisciplinary consultation. Nonetheless, while every classification incorporates scientific advances and aims to improve upon the prior version, medical knowledge remains incomplete and individual neoplasms may not be easily subclassified in a given scheme. Thus, optimal classification requires ongoing study, and there are certain aspects of some entities and subtypes that require further refinements. In this review, we highlight a selection of these challenging areas to prompt more research investigations. These include (1) a 'placeholder term' of splenic B-cell lymphoma/leukaemia with prominent nucleoli (SBLPN) to accommodate many of the splenic lymphomas previously classified as hairy cell leukaemia variant and B-prolymphocytic leukaemia, a clear new start to define their pathobiology; (2) how best to classify BCL2 rearrangement negative follicular lymphoma including those with BCL6 rearrangement, integrating the emerging new knowledge on various germinal centre B-cell subsets; (3) what is the spectrum of non-IG gene partners of MYC translocation in diffuse large B-cell lymphoma/high-grade B-cell lymphoma and how they impact MYC expression and clinical outcome; how best to investigate this in a routine clinical setting; and (4) how best to define high-grade B-cell lymphoma not otherwise specified and high-grade B-cell lymphoma with 11q aberrations to distinguish them from their mimics and characterise their molecular pathogenetic mechanism. Addressing these questions would provide more robust evidence to better define these entities/subtypes, improve their diagnosis and/or prognostic stratification, leading to better patient care. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

CEBPA facilitates LOXL2 and LOXL3 transcription to promote BCL-2 stability and thus enhances the growth and metastasis of lung carcinoma cells in vitro.

Lung carcinoma (LC) is a complicated and highly heterogeneous disease with high morbidity and mortality. Both lysyl oxidase-like (LOXL) 2 and 3 act in cancer progression. This work endeavors to illustrate the influence of LOXL2/LOXL3 on LC progression and the underlying mechanisms. LOXL family genes and CCAAT enhancer binding protein A (CEBPA) were analyzed in the TCGA database for their expression patterns in LC patients and their correlations with the patient's prognosis. CEBPA, LOXL2, and LOXL3 expression levels were determined in LC cells. Gain- and loss-of-function assays were conducted, followed by assays for cell proliferation, epithelial-mesenchymal transition (EMT), apoptosis, invasion, and migration. The binding of CEBPA or B cell lymphoma protein (BCL)-2 to LOXL2/LOXL3 was verified. The ubiquitination level of BCL-2 and histone acetylation level of LOXL2/LOXL3 in LC cells were analyzed. Database analyses revealed that LC patients had high CEBPA, LOXL2, and LOXL3 expression, which were related to poor prognosis. LC cells also exhibited high CEBPA, LOXL2, and LOXL3 levels. LOXL2/LOXL3 knockdown subdued EMT, proliferation, migration, and invasion while enhancing the apoptosis of LC cells. LOXL2/LOXL3 could bind to CEBPA and BCL-2. LOXL2/LOXL3 knockdown upregulated BCL-2 ubiquitination level and diminished BCL-2 expression in LC cells. CEBPA recruited Tip60 to enhance histone acetylation and transcription of LOXL2/LOXL3 in LC cells. BCL-2 overexpression abolished the impacts of LOXL2/LOXL3 knockdown on LC cells. In conclusion, CEBPA boosts LOXL2 and LOXL3 transcription to facilitate BCL-2 stability by recruiting Tip60 and thus contributes to LC cell growth and metastasis.

High-multiplex single-cell imaging analysis reveals tumor immune contexture associated with clinical outcomes after CAR T cell therapy.

Chimeric antigen receptor (CAR) T cell therapy has made great progress in treating lymphoma, yet patient outcomes still vary greatly. The lymphoma microenvironment may be an important factor in the efficacy of CAR T therapy. In this study, we designed a highly multiplexed imaging mass cytometry (IMC) panel to simultaneously quantify 31 biomarkers from 13 patients with relapsed/refractory diffuse large B cell lymphoma (DLBCL) who received CAR19/22 T cell therapy. A total of 20 sections were sampled before CAR T cell infusion or after infusion when relapse occurred. A total of 35 cell clusters were identified, annotated, and subsequently redefined into 10 metaclusters. The CD4+ T cell fraction was positively associated with remission duration. Significantly higher Ki67, CD57, and TIM3 levels and lower CD69 levels in T cells, especially the CD8+/CD4+ Tem and Te cell subsets, were seen in patients with poor outcomes. Cellular neighborhood containing more immune cells was associated with longer remission. Fibroblasts and vascular endothelial cells resided much closer to tumor cells in patients with poor response and short remission after CAR T therapy. Our work comprehensively and systematically dissects the relationship between cell composition, state, and spatial arrangement in the DLBCL microenvironment and the outcomes of CAR T cell therapy, which is beneficial to predict CAR T therapy efficacy.

Bispecific antibodies and autologous chimeric antigen receptor T cell therapies for treatment of hematological malignancies.

In recent years, the therapeutic landscape for hematological malignancies has markedly advanced, particularly since the inaugural approval of autologous chimeric antigen receptor T cell (CAR-T) therapy in 2017 for relapsed/refractory acute lymphoblastic leukemia (ALL). Autologous CAR-T therapy involves the genetic modification of a patient's T cells to specifically identify and attack cancer cells, while bispecific antibodies (BsAbs) function by binding to both cancer cells and immune cells simultaneously, thereby triggering an immune response against the tumor. The subsequent approval of various CAR-T therapies and BsAbs have revolutionized the treatment of multiple hematological malignancies, highlighting high response rates and a subset of patients achieving prolonged disease control. This review explores the mechanisms underlying autologous CAR-T therapies and BsAbs, focusing on their clinical application in multiple myeloma, ALL, and non-Hodgkin lymphoma. We provide comprehensive insights into their individual efficacy, limitations concerning broad application, and the potential of combination therapies. These upcoming strategies aim to propel the field forward, paving the way for safer and more effective therapeutic interventions in hematological malignancies.

Proteomics Identifies Circulating TIMP-1 as a Prognostic Biomarker for Diffuse Large B-Cell Lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease, although disease stratification using in-depth plasma proteomics has not been performed to date. By measuring more than 1000 proteins in the plasma of 147 DLBCL patients using data-independent acquisition mass spectrometry and antibody array, DLBCL patients were classified into four proteomic subtypes (PS-I-IV). Patients with the PS-IV subtype and worst prognosis had increased levels of proteins involved in inflammation, including a high expression of metalloproteinase inhibitor-1 (TIMP-1) that was associated with poor survival across two validation cohorts (n = 180). Notably, the combination of TIMP-1 with the international prognostic index (IPI) identified 64.00% to 88.24% of relapsed and 65.00% to 80.49% of deceased patients in the discovery and two validation cohorts, which represents a 24.00% to 41.67% and 20.00% to 31.70% improvement compared to the IPI score alone, respectively. Taken together, we demonstrate that DLBCL heterogeneity is reflected in the plasma proteome and that TIMP-1, together with the IPI, could improve the prognostic stratification of patients.

Efficient chimeric antigen receptor targeting of a central epitope of CD22.

Chimeric antigen receptor (CAR) T-cell therapy has had considerable success in the treatment of B-cell malignancies. Targeting the B-lineage marker CD19 has brought great advances to the treatment of acute lymphoblastic leukemia and B-cell lymphomas. However, relapse remains an issue in many cases. Such relapse can result from downregulation or loss of CD19 from the malignant cell population or expression of alternate isoforms. Consequently, there remains a need to target alternative B-cell antigens and diversify the spectrum of epitopes targeted within the same antigen. CD22 has been identified as a substitute target in cases of CD19-negative relapse. One anti-CD22 antibody-clone m971-targets a membrane-proximal epitope of CD22 and has been widely validated and used in the clinic. Here, we have compared m971-CAR with a novel CAR derived from IS7, an antibody that targets a central epitope on CD22. The IS7-CAR has superior avidity and is active and specific against CD22-positive targets, including B-acute lymphoblastic leukemia patient-derived xenograft samples. Side-by-side comparisons indicated that while IS7-CAR killed less rapidly than m971-CAR in vitro, it remains efficient in controlling lymphoma xenograft models in vivo. Thus, IS7-CAR presents a potential alternative candidate for the treatment of refractory B-cell malignancies.

Mediation of Ferroptosis Suppressor Protein 1 Expression via 4-Hydroxy-2-Nonenal Accumulation Contributes to Acquisition of Resistance to Apoptosis and Ferroptosis in Diffuse Large B-Cell Lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is the most common non-Hodgkin lymphoma. New therapeutic strategies are needed for the treatment of refractory DLBCL. 4-Hydroxy-2-nonenal (4-HNE) is a cytotoxic lipid peroxidation marker, which alters intracellular signaling and induces genetic mutations. Lipid peroxidation is associated with nonapoptotic cell death, called ferroptosis. However, the relationship between 4-HNE accumulation and feroptotic regulators in DLBCL has not been fully evaluated. Here, we aimed to evaluate the accumulation of lipid peroxide and the expression of ferroptosis suppressor protein 1 (FSP1) in DLBCL using immunohistochemistry. We found a significant increase in the expression of FSP1 in cases with nuclear 4-HNE accumulation (P = .021). Both nuclear and cytoplasmic 4-HNE accumulation and FSP1 positivity were independent predictors of worse prognosis. In vitro exposure to 4-HNE resulted in its concentration- and time-dependent intracellular accumulation and increased expression of FSP1. Furthermore, short-term (0.25 and 1.0 µM) or long-term (0.25 µM) exposure to 4-HNE induced resistance to not only apoptosis but also ferroptosis. Taken together, regulation of FSP1 through 4-HNE accumulation may attenuate resistance to cell death in treatment-resistant DLBCL and might help develop novel therapeutic strategies for refractory DLBCL.

Activation-induced cytidine deaminase causes recurrent splicing mutations in diffuse large B-cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is the most common lymphoma. A major mutagenic process in DLBCL is aberrant somatic hypermutation (aSHM) by activation-induced cytidine deaminase (AID), which occurs preferentially at RCH/TW sequence motifs proximal to transcription start sites. Splice sequences are highly conserved, rich in RCH/TW motifs, and recurrently mutated in DLBCL. Therefore, we hypothesized that aSHM may cause recurrent splicing mutations in DLBCL. In a meta-cohort of > 1,800 DLBCLs, we found that 77.5% of splicing mutations in 29 recurrently mutated genes followed aSHM patterns. In addition, in whole-genome sequencing (WGS) data from 153 DLBCLs, proximal mutations in splice sequences, especially in donors, were significantly enriched in RCH/TW motifs (p < 0.01). We validated this enrichment in two additional DLBCL cohorts (N > 2,000; p < 0.0001) and confirmed its absence in 12 cancer types without aSHM (N > 6,300). Comparing sequencing data from mouse models with and without AID activity showed that the splice donor sequences were the top genomic feature enriched in AID-induced mutations (p < 0.0001). Finally, we observed that most AID-related splice site mutations are clonal within a sample, indicating that aSHM may cause early loss-of-function events in lymphomagenesis. Overall, these findings support that AID causes an overrepresentation of clonal splicing mutations in DLBCL.

Unveiling the PDK4-centered rituximab-resistant mechanism in DLBCL: the potential of the "Smart" exosome nanoparticle therapy.

BACKGROUND: Diffuse large B-cell lymphoma (DLBCL) represents a prevalent malignant tumor, with approximately 40% of patients encountering treatment challenges or relapse attributed to rituximab resistance, primarily due to diminished or absent CD20 expression. Our prior research identified PDK4 as a key driver of rituximab resistance through its negative regulation of CD20 expression. Further investigation into PDK4's resistance mechanism and the development of advanced exosome nanoparticle complexes may unveil novel resistance targets and pave the way for innovative, effective treatment modalities for DLBCL. METHODS: We utilized a DLBCL-resistant cell line with high PDK4 expression (SU-DHL-2/R). We infected it with short hairpin RNA (shRNA) lentivirus for RNA sequencing, aiming to identify significantly downregulated mRNA in resistant cells. Techniques including immunofluorescence, immunohistochemistry, and Western blotting were employed to determine PDK4's localization and expression in resistant cells and its regulatory role in phosphorylation of Histone deacetylase 8 (HDAC8). Furthermore, we engineered advanced exosome nanoparticle complexes, aCD20@ExoCTX/siPDK4, through cellular, genetic, and chemical engineering methods. These nanoparticles underwent characterization via Dynamic Light Scattering (DLS) and Transmission Electron Microscopy (TEM), and their cellular uptake was assessed through flow cytometry. We evaluated the nanoparticles' effects on apoptosis in DLBCL-resistant cells and immune cells using CCK-8 assays and flow cytometry. Additionally, their capacity to counteract resistance and exert anti-tumor effects was tested in a resistant DLBCL mouse model. RESULTS: We found that PDK4 initiates HDAC8 activation by phosphorylating the Ser-39 site, suppressing CD20 protein expression through deacetylation. The aCD20@ExoCTX/siPDK4 nanoparticles served as effective intracellular delivery mechanisms for gene therapy and monoclonal antibodies, simultaneously inducing apoptosis in resistant DLBCL cells and triggering immunogenic cell death in tumor cells. This dual action effectively reversed the immunosuppressive tumor microenvironment, showcasing a synergistic therapeutic effect in a subcutaneous mouse tumor resistance model. CONCLUSIONS: This study demonstrates that PDK4 contributes to rituximab resistance in DLBCL by modulating CD20 expression via HDAC8 phosphorylation. The designed exosome nanoparticles effectively overcome this resistance by targeting the PDK4/HDAC8/CD20 pathway, representing a promising approach for drug delivery and treating patients with Rituximab-resistant DLBCL.

Transformation to diffuse large B-cell lymphoma and its impact on survival in patients with marginal zone lymphoma: A population-based study.

Some patients with marginal zone lymphoma (MZL) experience histological transformation to diffuse large B-cell lymphoma (DLBCL). Because of the paucity of long-term data on transformation, we conducted a population-based study to estimate the risk of transformation and its impact on survival in MZL. Using the Surveillance, Epidemiology and End Results database, we identified 23 221 patients with histology-proven MZL between 2000 and 2018. Competing risk method, Kaplan-Meier and Cox proportional hazards regression were performed to analyze time-to-event outcomes. Based on 420 events of transformation, the 10-year cumulative incidence rate of transformation is 2.23% (95% CI: 2.00%-2.46%) in MZL, 1.5% (95% CI: 1.3%-1.8%), 2.7% (95% CI: 2.3%-3.2%) and 5.8% (95% CI: 4.6%-7.1%) in extranodal, nodal and splenic MZL (EMZL, NMZL and SMZL), respectively. Patients with SMZL (subdistribution hazard ratio [SHR], 2.96; 95% CI: 2.21-3.96) or NMZL (SHR, 1.49; 95% CI: 1.17-1.90) have a higher risk of transformation than those with EMZL. For each MZL subtype, patients with transformation had a significantly shorter overall survival. Patients with transformation >18 months since MZL diagnosis had longer OS than those who presented within 18 months (5-year rate, 87.4% [95% CI: 83.7%-91.2%] vs 47.9% [95% CI: 38.8%-59.0%]; P < .001). Compared to patients with matched de novo DLBCL, those whose DLBCL was transformed from MZL had a shorter OS (5-year rate, 56.6% [95% CI: 51.9%-61.8%] vs 46.1% [95% CI: 40.9%-51.9%]; P < .001). We concluded that patients with SMZL had the highest risk of transformation. Regardless of MZL subtype, transformation resulted in significantly increased mortality.

Ajania pacifica (Nakai) K. Bremer and Humphries Extract Limits MYC Expression to Induce Apoptosis in Diffuse Large B Cell Lymphoma.

The proto-oncogene MYC is frequently dysregulated in patients with diffuse large B-cell lymphoma (DLBCL) and plays a critical role in disease progression. To improve the clinical outcomes of patients with DLBCL, the development of strategies to target MYC is crucial. The use of medicinal plants for developing anticancer drugs has garnered considerable attention owing to their diverse mechanisms of action. In this study, 100 plant extracts of flora from the Republic of Korea were screened to search for novel agents with anti-DLBCL effects. Among them, Ajania pacifica (Nakai) K. Bremer and Humphries extract (APKH) efficiently suppressed the survival of DLBCL cells, while showing minimal toxicity toward normal murine bone marrow cells. APKH suppressed the expression of anti-apoptotic BCL2 family members, causing an imbalance between the pro-apoptotic and anti-apoptotic BCL2 members. This disrupted mitochondrial membrane potential, cytochrome c release, and pro-caspase-3 activation and eventually led to DLBCL cell death. Importantly, MYC expression was markedly downregulated by APKH and ectopic expression of MYC in DLBCL cells abolished the pro-apoptotic effects of APKH. These results demonstrate that APKH exerts anti-DLBCL effects by inhibiting MYC expression. Moreover, when combined with doxorubicin, an essential component of the CHOP regimen (cyclophosphamide, doxorubicin, vincristine, and prednisone), APKH synergistically enhanced the therapeutic effect of doxorubicin. This indicates that APKH may overcome drug resistance, which is common in patients with refractory/relapsed DLBCL. To identify compounds with anti-DLBCL activities in APKH, the chemical profile analysis of APKH was performed using UPLC-QTOF/MSe analysis and assessed for its anticancer activity. Based on the UPLC-QTOF/MSe chemical profiling, it is conceivable that APKH may serve as a novel agent targeting MYC and sensitizing drug-resistant DLBCL cells to CHOP chemotherapy. Further studies to elucidate how the compounds in APKH exert tumor-suppressive role in DLBCL are warranted.