Enhancing cell death in B-cell malignancies through targeted inhibition of Bcl-3.

The t(14;19)(q32;q13) is a rare recurring translocation found in B-cell lymphoproliferative malignancies, involving the Bcl-3 gene. This chromosomal translocation is often found in patients under the age of 50 and causes a more progressive disease. The Bcl-3 gene encodes a protein belonging to the IκB family of proteins, which tightly regulates NFκB signaling by acting as an activator or repressor of transcription. Previously, we developed a second-generation Bcl-3 inhibitor that could directly interfere with Bcl-3 signaling pathway, resulting in reduced melanoma cell proliferation, invasion, and migration. The present study aimed to investigate the effect of a Bcl-3 inhibitor on B-cell lymphoma and leukemia cells. It was found that treatment of cells with this inhibitor caused a decrease in cell proliferation and cell survival. Furthermore, Bcl-3 inhibition in B-cell malignant cells resulted in the loss of mitochondrial membrane potential and functionality, as well as the increased expression of cleaved caspase 3, indicating that cell death occurs through the intrinsic apoptotic pathway. This observation is further supported by reduced expression of cIAP1 protein 1 (cIAP1) upon treatment of cancer cells. Given the current lack of clinical advancements targeting Bcl-3 in oncology, this opens a novel avenue for the development and investigation of highly specific therapeutic interventions against B-cell malignancies.

Spatial tumor immune microenvironment phenotypes in ovarian cancer.

Immunotherapy has largely failed in ovarian carcinoma (OC), likely due to that the vast tumor heterogeneity and variation in immune response have hampered clinical trial outcomes. Tumor-immune microenvironment (TIME) profiling may aid in stratification of OC tumors for guiding treatment selection. Here, we used Digital Spatial Profiling combined with image analysis to characterize regions of spatially distinct TIME phenotypes in OC to assess whether immune infiltration pattern can predict presence of immuno-oncology targets. Tumors with diffuse immune infiltration and increased tumor-immune spatial interactions had higher presence of IDO1, PD-L1, PD-1 and Tim-3, while focal immune niches had more CD163 macrophages and a preliminary worse outcome. Immune exclusion was associated with presence of Tregs and Fibronectin. High-grade serous OC showed an overall stronger immune response and presence of multiple targetable checkpoints. Low-grade serous OC was associated with diffuse infiltration and a high expression of STING, while endometrioid OC had higher presence of CTLA-4. Mucinous and clear cell OC were dominated by focal immune clusters and immune-excluded regions, with mucinous tumors displaying T-cell rich immune niches.

Quantification and Profiling of Early and Late Differentiation Stage T Cells in Mantle Cell Lymphoma Reveals Immunotherapeutic Targets in Subsets of Patients.

With the aim to advance the understanding of immune regulation in MCL and to identify targetable T-cell subsets, we set out to combine image analysis and spatial omic technology focused on both early and late differentiation stages of T cells. MCL patient tissue (n = 102) was explored using image analysis and GeoMx spatial omics profiling of 69 proteins and 1812 mRNAs. Tumor cells, T helper (TH) cells and cytotoxic (TC) cells of early (CD57-) and late (CD57+) differentiation stage were analyzed. An image analysis workflow was developed based on fine-tuned Cellpose models for cell segmentation and classification. TC and CD57+ subsets of T cells were enriched in tumor-rich compared to tumor-sparse regions. Tumor-sparse regions had a higher expression of several key immune suppressive proteins, tentatively controlling T-cell expansion in regions close to the tumor. We revealed that T cells in late differentiation stages (CD57+) are enriched among MCL infiltrating T cells and are predictive of an increased expression of immune suppressive markers. CD47, IDO1 and CTLA-4 were identified as potential targets for patients with T-cell-rich MCL TIME, while GITR might be a feasible target for MCL patients with sparse T-cell infiltration. In subgroups of patients with a high degree of CD57+ TC-cell infiltration, several immune checkpoint inhibitors, including TIGIT, PD-L1 and LAG3 were increased, emphasizing the immune-suppressive features of this highly differentiated T-cell subset not previously described in MCL.

CD163+ macrophages in mantle cell lymphoma induce activation of prosurvival pathways and immune suppression.

ABSTRACT: Mantle cell lymphoma (MCL) is dependent on a supportive tumor immune microenvironment (TIME) in which infiltration of CD163+ macrophages has a negative prognostic impact. This study explores how abundance and spatial localization of CD163+ cells are associated with the biology of MCL, using spatial multiomic investigations of tumor and infiltrating CD163+ and CD3+ cells. A total of 63 proteins were measured using GeoMx digital spatial profiling in tissue microarrays from 100 diagnostic MCL tissues. Regions of interest were selected in tumor-rich and tumor-sparse tissue regions. Molecular profiling of CD163+ macrophages, CD20+ MCL cells, and CD3+ T-cells was performed. To validate protein profiles, 1811 messenger RNAs were measured in CD20+ cells and 2 subsets of T cells. Image analysis was used to extract the phenotype and position of each targeted cell, thereby allowing the exploration of cell frequencies and cellular neighborhoods. Proteomic investigations revealed that CD163+ cells modulate their immune profile depending on their localization and that the immune inhibitory molecules, V-domain immunoglobulin suppressor of T-cell activation and B7 homolog 3, have higher expression in tumor-sparse than in tumor-rich tissue regions and that targeting should be explored. We showed that MCL tissues with more abundant infiltration of CD163+ cells have a higher proteomic and transcriptional expression of key components of the MAPK pathway. Thus, the MAPK pathway may be a feasible therapeutic target in patients with MCL with CD163+ cell infiltration. We further showed the independent and combined prognostic values of CD11c and CD163 beyond established risk factors.

Empowering macrophages: the cancer fighters within the tumour microenvironment in mantle cell lymphoma.

In Mantle Cell Lymphoma (MCL), the role of macrophages within the tumour microenvironment (TME) has recently gained attention due to their impact on prognosis and response to therapy. Despite their low absolute number in MCL tumour tissue, recent findings reveal an association between the levels of macrophages and prognosis, consistent with trends observed in other lymphoma subtypes. M2-like macrophages, identified by markers such as CD163, contribute to angiogenesis and suppression of the immune response. Clinical trials with MCL patients treated with chemoimmunotherapy and targeted treatments underscore the adverse impact of high levels of M2-like macrophages. Immunomodulatory drugs like lenalidomide reduce the levels of MCL-associated CD163+ macrophages and enhance macrophage phagocytic activity. Similarly, clinical approaches targeting the CD47 "don't eat me" signalling, in combination with the anti-CD20-antibody rituximab, demonstrate increased macrophage activity and phagocytosis of MCL tumour cells. Cell-based therapies such as chimeric antigen receptor (CAR) T-cell have shown promise but various challenges persist, leading to a potential interest in CAR-macrophages (CAR-M). When macrophages are recruited to the TME, they offer advantages including phagocytic function and responsiveness to microenvironment alterations, suggesting their potential as a manipulable and inducible alternative when CAR T-cell therapies fails in the complex landscape of MCL treatment.

Predictive and Prognostic Biomarkers in Patients With Mycosis Fungoides and Sézary Syndrome (BIO-MUSE): Protocol for a Translational Study.

BACKGROUND: Cutaneous T-cell lymphoma (CTCL) is a rare group of lymphomas that primarily affects the skin. Mycosis fungoides (MF) is the most common form of CTCL and Sézary syndrome (SS) is more infrequent. Early stages (IA-IIA) have a favorable prognosis, while advanced stages (IIB-IVB) have a worse prognosis. Around 25% of patients with early stages of the disease will progress to advanced stages. Malignant skin-infiltrating T-cells in CTCL are accompanied by infiltrates of nonmalignant T-cells and other immune cells that produce cytokines that modulate the inflammation. Skin infection, often with Staphylococcus aureus, is frequent in advanced stages and can lead to sepsis and death. S. aureus has also been reported to contribute to the progression of the disease. Previous reports indicate a shift from Th1 to Th2 cytokine production and dysfunction of the skin barrier in CTCL. Treatment response is highly variable and often unpredictable, and there is a need for new predictive and prognostic biomarkers. OBJECTIVE: This prospective translational study aims to identify prognostic biomarkers in the blood and skin of patients with MF and SS. METHODS: The Predictive and Prognostic Biomarkers in Patients With MF and SS (BIO-MUSE) study aims to recruit 120 adult patients with MF or SS and a control group of 20 healthy volunteers. The treatments will be given according to clinical routine. The sampling of each patient will be performed every 3 months for 3 years. The blood samples will be analyzed for lactate dehydrogenase, immunoglobulin E, interleukins, thymus and activation-regulated chemokine, and lymphocyte subpopulations. The lymphoma microenvironment will be investigated through digital spatial profiling and single-cell RNA sequencing. Microbiological sampling and analysis of skin barrier function will be performed. The life quality parameters will be evaluated. The results will be evaluated by the stage of the disease. RESULTS: Patient inclusion started in 2021 and is still ongoing in 2023, with 18 patients and 20 healthy controls enrolled. The publication of selected translational findings before the publication of the main results of the trial is accepted. CONCLUSIONS: This study aims to investigate blood and skin with a focus on immune cells and the microbiological environment to identify potential new prognostic biomarkers in MF and SS. TRIAL REGISTRATION: ClinicalTrials.gov NCT04904146; https://www.clinicaltrials.gov/study/NCT04904146. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/55723.

MYC protein is a high-risk factor in mantle cell lymphoma and identifies cases beyond morphology, proliferation and TP53/p53 - a Nordic Lymphoma Group study.

The transcription factor MYC is a well-described oncogene with an important role in lymphomagenesis, but its significance for clinical outcome in mantle cell lymphoma (MCL) remains to be determined. We performed an investigation of the expression of MYC protein in a cohort of 251 MCL patients complemented by analyses of structural aberrations and mRNA, in a sub-cohort of patients. Fourteen percent (n=35) of patients showed high MYC protein expression with >20% positive cells (MYChigh), among whom only one translocation was identified, and 86% (n=216) of patients showed low MYC protein expression. Low copy number gains of MYC were detected in ten patients, but with no correlation to MYC protein levels. However, MYC mRNA levels correlated significantly to MYC protein levels with a R2 value of 0.76. Patients with a MYChigh tumor had both an independent inferior overall survival and an inferior progression-free survival (hazard ratio [HR]=2.03, 95% confidence interval [95% CI]: 1.2-3.4 and HR=2.2, 95% CI: 1.04-4.6, respectively) when adjusted for additional high-risk features. Patients with MYChigh tumors also tended to have additional high-risk features and to be older at diagnosis. A subgroup of 13 patients had concomitant MYChigh expression and TP53/p53 alterations and a substantially increased risk of progression (HR=16.9, 95% CI: 7.4-38.3) and death (HR=7.8, 95% CI: 4.4-14.1) with an average overall survival of only 0.9 years. In summary, we found that at diagnosis a subset of MCL patients (14%) overexpressed MYC protein, and had a poor prognosis but that MYC rearrangements were rare. Tumors with concurrent MYC overexpression and TP53/p53 alterations pinpointed MCL patients with a dismal prognosis with a median overall survival of less than 3 years. We propose that MYC needs to be assessed beyond the current high-risk factors in MCL in order to identify cases in need of alternative treatment.

Soluble CD163 predicts outcome in both chemoimmunotherapy and targeted therapy-treated mantle cell lymphoma.

The outcome for patients with mantle cell lymphoma (MCL) has drastically improved with new treatments directed toward the tumor immune microenvironment, where macrophages play an important role. In MCL, the presence of M2 macrophages defined by CD163 expression in diagnostic biopsies has been associated with a worse prognosis. An alternative way to assess the abundance of M2 macrophages is by measuring the level of soluble CD163 in serum (sCD163). We aimed to investigate the prognostic value of sCD163 in 131 patients with MCL. We found that high sCD163 at diagnosis was associated with shorter progression-free survival (PFS) and shorter overall survival (OS) in 81 patients who were newly diagnosed and subsequently treated with chemoimmunotherapy. The same was seen in a cohort of 50 patients with relapsed MCL that were mainly treated within the phase 2 Philemon-trial with rituximab, ibrutinib, and lenalidomide. In patients who were newly diagnosed and had low levels of sCD163, 5-year survival was 97%. There was a moderate correlation between sCD163 and tissue CD163. The association with a poor prognosis was independent of MCL international prognostic index, Ki67, p53 status, and blastoid morphology, as assessed in a multivariable Cox proportional hazards model. In this study, high sCD163 was associated with both shorter PFS and shorter OS, showing that high levels of the M2 macrophage marker sCD163 is an independent negative prognostic factor in MCL, both in the chemoimmunotherapy and ibrutinib/lenalidomide era. In addition, low sCD163 levels identify patients with MCL with a very good prognosis.

Exploring new prognostic biomarkers in Mantle Cell Lymphoma: a comparison of the circSCORE and the MCL35 score.

Mantle cell lymphoma (MCL) is a biologically and clinically heterogeneous disease, emphasizing the need for prognostic biomarkers. In this study we aimed at comparing the prognostic value of two RNA-based risk scores, circSCORE and MCL35, in 149 patients from the MCL2 (ISRCTN87866680) and MCL3 (NCT00514475) patient cohorts. Both risk scores provided significant stratification of high versus low risk for progression free survival (PFS) and overall survival (OS). The circSCORE retained significant prognostic value in adjusted multivariable Cox regressions for PFS, but not for OS. Furthermore, circSCORE added significant prognostic value to MIPI in the pooled cohort (MCL2 and MCL3) for PFS and OS, and for PFS in MCL3 alone, outperforming Ki67 and MCL35. We suggest a new, combined MIPI-circSCORE with improved prognostic value, and with potential for future clinical implementation, if validated in a larger, independent cohort.

Phenotypic characterization of spatial immune infiltration niches in non-small cell lung cancer.

The immune microenvironment of non-small cell lung cancer (NSCLC) is heterogeneous, which impedes the prediction of response to immune checkpoint inhibitors. We have mapped the expression of 49 proteins to spatial immune niches in 33 NSCLC tumors and report key differences in phenotype and function associated with the spatial context of immune infiltration. Tumor-infiltrating leukocytes (TIL), identified in 42% of tumors, had a similar proportion of lymphocyte antigens compared to stromal leukocytes (SL) but displayed significantly higher levels of functional, mainly immune suppressive, markers including PD-L1, PD-L2, CTLA-4, B7-H3, OX40L, and IDO1. In contrast, SL expressed higher levels of the targetable T-cell activation marker CD27, which increased with a longer distance to the tumor. Correlation analysis confirmed that metabolic-driven immune regulatory mechanisms, including ARG1 and IDO1, are present in the TIL. Tertiary lymphoid structures (TLS) were identified in 30% of patients. They displayed less variation in the expression profile and with significantly higher levels of pan lymphocyte and activation markers, dendritic cells, and antigen presentation compared to other immune niches. TLS also had higher CTLA-4 expression than non-structured SL, which may indicate immune dysfunction. Neither the presence of TIL nor TLS was associated with improved clinical outcomes. The apparent discrimination in functional profiles of distinct immune niches, independent of the overall level of leukocytes, illustrates the importance of spatial profiling to deconvolute how the immune microenvironment can dictate a therapeutic response and to identify biomarkers in the context of immunomodulatory treatment.

Spatially Guided and Single Cell Tools to Map the Microenvironment in Cutaneous T-Cell Lymphoma.

Mycosis fungoides (MF) and Sézary syndrome (SS) are two closely related clinical variants of cutaneous T-cell lymphomas (CTCL). Previously demonstrated large patient-to-patient and intra-patient disease heterogeneity underpins the importance of personalized medicine in CTCL. Advanced stages of CTCL are characterized by dismal prognosis, and the early identification of patients who will progress remains a clinical unmet need. While the exact molecular events underlying disease progression are poorly resolved, the tumor microenvironment (TME) has emerged as an important driver. In particular, the Th1-to-Th2 shift in the immune response is now commonly identified across advanced-stage CTCL patients. Herein, we summarize the role of the TME in CTCL evolution and the latest studies in deciphering inter- and intra-patient heterogeneity. We introduce spatially resolved omics as a promising technology to advance immune-oncology efforts in CTCL. We propose the combined implementation of spatially guided and single-cell omics technologies in paired skin and blood samples. Such an approach will mediate in-depth profiling of phenotypic and molecular changes in reactive immune subpopulations and malignant T cells preceding the Th1-to-Th2 shift and reveal mechanisms underlying disease progression from skin-limited to systemic disease that collectively will lead to the discovery of novel biomarkers to improve patient prognostication and the design of personalized treatment strategies.

SOX11 regulates SWI/SNF complex components as member of the adrenergic neuroblastoma core regulatory circuitry.

The pediatric extra-cranial tumor neuroblastoma displays a low mutational burden while recurrent copy number alterations are present in most high-risk cases. Here, we identify SOX11 as a dependency transcription factor in adrenergic neuroblastoma based on recurrent chromosome 2p focal gains and amplifications, specific expression in the normal sympatho-adrenal lineage and adrenergic neuroblastoma, regulation by multiple adrenergic specific (super-)enhancers and strong dependency on high SOX11 expression in adrenergic neuroblastomas. SOX11 regulated direct targets include genes implicated in epigenetic control, cytoskeleton and neurodevelopment. Most notably, SOX11 controls chromatin regulatory complexes, including 10 SWI/SNF core components among which SMARCC1, SMARCA4/BRG1 and ARID1A. Additionally, the histone deacetylase HDAC2, PRC1 complex component CBX2, chromatin-modifying enzyme KDM1A/LSD1 and pioneer factor c-MYB are regulated by SOX11. Finally, SOX11 is identified as a core transcription factor of the core regulatory circuitry (CRC) in adrenergic high-risk neuroblastoma with a potential role as epigenetic master regulator upstream of the CRC.

Overexpression of the key metabolic protein CPT1A defines mantle cell lymphoma patients with poor response to standard high-dose chemotherapy independent of MIPI and complement established highrisk factors.

The variable outcome to standard immunochemotherapy for mantle cell lymphoma (MCL) patients is a clinical challenge. Established risk factors, including high MCL International Prognostic Index (MIPI), high proliferation (Ki-67), non-classic (blastoid/pleomorphic) morphology, and mutated TP53, only partly identify patients in need of alternative treatment. Deepened understanding of biological factors that influence time to progression and relapse would allow for an improved stratification, and identification of novel targets for high-risk patients. We performed gene expression analyses to identify pathways and genes associated with outcome in a cohort of homogeneously treated patients. In addition to deregulated proliferation, we show that thermogenesis, fatty acid degradation and oxidative phosphorylation are altered in patients with poor survival, and that high expression of carnitine palmitoyltransferase 1A (CPT1A), an enzyme involved in fatty acid degradation, can specifically identify high-risk patients independent of the established high-risk factors. We suggest that complementary investigations of metabolism may increase the accuracy of patient stratification and that immunohistochemistry- based assessment of CPT1A can contribute to defining high-risk MCL.

Clinical and biological impact of SAMHD1 expression in mantle cell lymphoma.

SAMHD1 is a deoxynucleoside triphosphate triphosphohydrolase (dNTPase) that restricts viral replication in infected cells and limits the sensitivity to cytarabine by hydrolysing its active metabolite, as recently shown in acute myeloid leukemia. Cytarabine is an essential component in the Nordic mantle cell lymphoma protocols (MCL2 and MCL3) for induction and high-dose chemotherapy treatment before autologous stem cell transplantation for younger patients with mantle cell lymphoma (MCL). We here investigated the expression of SAMHD1 in a population-based cohort of MCL (N = 150). SAMHD1 was highly variably expressed in MCL (range, 0.4% to 100% of positive tumor cells). Cases with blastoid/pleomorphic morphology had higher SAMHD1 expression (P = 0.028) and SAMHD1 was also correlated to tumor cell proliferation (P = 0.016). SAMHD1 expression showed moderate correlation to the expression of the transcriptional regulator SOX11 (P = 0.036) but genetic silencing of SOX11 and SAMHD1 by siRNA in MCL cell lines did not suggest mutual regulation. We hypothesized that expression of SAMHD1 could predict short time to progression in patients treated with Cytarabine as part of high-dose chemotherapy. Despite the correlation with known biological adverse prognostic factors, neither low or high SAMHD1 expression correlated to PFS or OS in patients treated according to the Nordic MCL2 or MCL3 protocols (N = 158).

Expression patterns and prognostic potential of circular RNAs in mantle cell lymphoma: a study of younger patients from the MCL2 and MCL3 clinical trials.

Mantle cell lymphoma (MCL) is characterized by marked differences in outcome, emphasizing the need for strong prognostic biomarkers. Here, we explore expression patterns and prognostic relevance of circular RNAs (circRNAs), a group of endogenous non-coding RNA molecules, in MCL. We profiled the circRNA expression landscape using RNA-sequencing and explored the prognostic potential of 40 abundant circRNAs in samples from the Nordic MCL2 and MCL3 clinical trials, using NanoString nCounter Technology. We report a circRNA-based signature (circSCORE) developed in the training cohort MCL2 that is highly predictive of time to progression (TTP) and lymphoma-specific survival (LSS). The dismal outcome observed in the large proportion of patients assigned to the circSCORE high-risk group was confirmed in the independent validation cohort MCL3, both in terms of TTP (HR 3.0; P = 0.0004) and LSS (HR 3.6; P = 0.001). In Cox multiple regression analysis incorporating MIPI, Ki67 index, blastoid morphology and presence of TP53 mutations, circSCORE retained prognostic significance for TTP (HR 3.2; P = 0.01) and LSS (HR 4.6; P = 0.01). In conclusion, circRNAs are promising prognostic biomarkers in MCL and circSCORE improves identification of high-risk disease among younger patients treated with cytarabine-containing chemoimmunotherapy and autologous stem cell transplant.

Serum proteome modulations upon treatment provides biological insight on response to treatment in relapsed mantle cell lymphoma.

BACKGROUND: The possibility to monitor patient's serum proteome during treatment can provide deepened understanding of the biology associated with response to specific drugs. Non-invasive serum sampling provides an opportunity for sustainable repetitive sampling of patients, which allows for more frequent evaluation of the biological response and enhanced flexibility in treatment selection in contrast to tissue biopsies. AIM: To pin-point biologically relevant changes in pre- and on-treatment serum proteome samples in relapsed mantle cell lymphoma (MCL) patients, leading to insight into mechanisms behind response to treatment in sub-groups of patients. METHODS: Pre- and on-treatment serum samples from relapsed MCL patients treated with a triple combination therapy of rituximab, ibrutinib and lenalidomide were available for the study, together with detailed clinicopathological information. A microarray technology targeting 158 serum proteins using 371 antibody-fragments was used to compare the serum proteome at the two time-points. RESULTS: Proteins modulated by the treatment were shown to be associated to a MCL sub-group with ATM/TP53 alterations, which emphasizes the importance of treatment stratification. Absolute values of serum protein levels in on-treatment samples were highly variable and showed no correlation to outcome. To circumvent the challenge of variability in absolute serum protein levels, the velocity of change of individual serum proteins was used to identify proteins associated with clinical response. Increased values of TGF-ß1, CD40 and complement component 4 comparing pre- and on-treatment samples were associated with remaining minimal residual disease (MRD) and increased BTK was associated with short progression-free survival (PFS). CONCLUSION: We show that the genetic sub-type of MCL affects the biological response to treatment in serum and that the change in defined serum proteins reveals the biology associated with clinical response.

Targeted genomic investigations in a population-based cohort of mantle cell lymphoma reveal novel clinically relevant targets.

Mantle cell lymphoma (MCL) is an aggressive B-cell neoplasm that follows a heterogeneous clinical course. Recurrent mutations have been described, but their applicability in the clinical setting is currently limited. The main reasons are challenges in the sequencing of DNA retrieved from formalin-fixed tissue commonly used for tissue collection in clinical biobanks. In this study, we sequenced 77 samples from a population-based de novo MCL cohort to investigate the utility of targeted sequencing in guiding personalized treatment approaches. Tumors were genetically variable, and a similar genetic landscape as previous studies using non-formalin fixed samples was identified, with recurrent mutations including ATM, KMT2D, and TP53. Novel alterations that can be considered actionable and/or indicative of treatment response were also identified. Our approach shows the potential benefits of using target sequencing of formalin fixed samples to facilitate treatment selection and individualized clinical decisions in MCL.

Kalirin-RAC controls nucleokinetic migration in ADRN-type neuroblastoma.

The migrational propensity of neuroblastoma is affected by cell identity, but the mechanisms behind the divergence remain unknown. Using RNAi and time-lapse imaging, we show that ADRN-type NB cells exhibit RAC1- and kalirin-dependent nucleokinetic (NUC) migration that relies on several integral components of neuronal migration. Inhibition of NUC migration by RAC1 and kalirin-GEF1 inhibitors occurs without hampering cell proliferation and ADRN identity. Using three clinically relevant expression dichotomies, we reveal that most of up-regulated mRNAs in RAC1- and kalirin-GEF1-suppressed ADRN-type NB cells are associated with low-risk characteristics. The computational analysis shows that, in a context of overall gene set poverty, the upregulomes in RAC1- and kalirin-GEF1-suppressed ADRN-type cells are a batch of AU-rich element-containing mRNAs, which suggests a link between NUC migration and mRNA stability. Gene set enrichment analysis-based search for vulnerabilities reveals prospective weak points in RAC1- and kalirin-GEF1-suppressed ADRN-type NB cells, including activities of H3K27- and DNA methyltransferases. Altogether, these data support the introduction of NUC inhibitors into cancer treatment research.

Infiltration of CD163-, PD-L1- and FoxP3-positive cells adversely affects outcome in patients with mantle cell lymphoma independent of established risk factors.

We characterised patients with mantle cell lymphoma (MCL) with poor prognosis based on differences in immune infiltration. Different expressions of the tumour cell markers Cyclin D1 and sex-determining region Y-box transcription factor 11 (SOX11), and the immune markers cluster of differentiation 3 (CD3), CD4, CD8, CD25, forkhead box protein P3 (FoxP3), T-box transcription factor TBX21 (T-bet), programmed cell death protein 1 (PD-1), programmed-death ligand 1 (PD-L1) and CD163 were investigated for all-cause mortality in 282 patients with MCL and time-to-progression (TTP) in 106 clinical trial patients. With increasing age, a significantly lower infiltration of CD3+ T lymphocytes was seen. T-cell infiltration was independent of cellular tumour antigen p53 (p53) expression, Ki-67, morphology and frequency of tumour cells. The all-cause mortality was higher in patients with PD-L1-expression above cut-off [hazard ratio (HR) 1·97, 95% confidence interval (CI) 1·18-3·25, adjusted for sex and MCL International Prognostic Index (MIPI)] and a higher frequency of CD163+ cells (continuously, HR 1·51, 95% CI 1·03-2·23, adjusting for age, sex, morphology, Ki-67 and p53). In patients treated within the Nordic Lymphoma Group MCL2/3 trials, TTP was shorter in patients with a higher frequency of FoxP3+ cells (HR 3·22, 95% CI 1·40-7·43) and CD163+ cells (HR 6·09, 95% CI 1·84-20·21), independent of sex and MIPI. When combined a higher frequency of CD163+ macrophages and PD-L1+ cells or high CD163+ macrophages and FoxP3+ regulatory T cells indicated worse outcome independent of established risk factors. The T-cell infiltrate was in turn independent of molecular characteristics of the malignant cells and decreased with age.