BCL3 rearrangements in B-cell lymphoid neoplasms occur in two breakpoint clusters associated with different diseases.

The t(14;19)(q32;q13) often juxtaposes BCL3 with immunoglobulin heavy chain (IGH) resulting in overexpression of the gene. In contrast to other oncogenic translocations, BCL3 rearrangement (BCL3-R) has been associated with a broad spectrum of lymphoid neoplasms. Here we report an integrative whole-genome sequence, transcriptomic, and DNA methylation analysis of 13 lymphoid neoplasms with BCL3-R. The resolution of the breakpoints at single base-pair revealed that they occur in two clusters at 5' (n=9) and 3' (n=4) regions of BCL3 associated with two different biological and clinical entities. Both breakpoints were mediated by aberrant class switch recombination of the IGH locus. However, the 5' breakpoints (upstream) juxtaposed BCL3 next to an IGH enhancer leading to overexpression of the gene whereas the 3' breakpoints (downstream) positioned BCL3 outside the influence of the IGH and were not associated with its expression. Upstream BCL3-R tumors had unmutated IGHV, trisomy 12, and mutated genes frequently seen in chronic lymphocytic leukemia (CLL) but had an atypical CLL morphology, immunophenotype, DNA methylome, and expression profile that differ from conventional CLL. In contrast, downstream BCL3-R neoplasms were atypical splenic or nodal marginal zone lymphomas (MZL) with mutated IGHV, complex karyotypes and mutated genes typical of MZL. Two of the latter four tumors transformed to a large B-cell lymphoma. We designed a novel fluorescence in situ hybridization assay that recognizes the two different breakpoints and validated these findings in 17 independent tumors. Overall, upstream or downstream breakpoints of BCL3-R are mainly associated with two subtypes of lymphoid neoplasms with different (epi)genomic, expression, and clinicopathological features resembling atypical CLL and MZL, respectively.

Cardiovascular events in children with brain injury: A systematic review.

BACKGROUND: Brain injury is a leading cause of morbidity and mortality in the pediatric population. Neurogenic stunned cardiomyopathy is a complication associated with several neurological conditions that can lead to worse outcomes. It presents as alterations in blood pressure, cardiac rhythm disturbances and the increase in cardiac injury biomarkers. This systematic review aims to assess the hemodynamic consequences of brain injury in the pediatric population to identify better management strategies and improve outcomes. METHODS: An electronic literature search was performed in Pubmed, Scopus and WebOfScience, up until October 3rd, 2022. The selected articles underwent quality assessment using the National Heart, Lung and Blood Institute tools for cohort and case-control studies. RESULTS: This systematic review includes thirteen articles on the effects of brain injury in arterial pressure, rhythm disturbances and biomarkers of myocardial injury. These studies showed the following key results: both hypotension and hypertension are associated with worse outcomes; brain injury could be related to longer QTc intervals; neurogenic stunned cardiomyopathy was a common found after brain injury. CONCLUSION: This is the first systematic review to report cardiovascular abnormalities arising from brain injury in children. An early arterial pressure, electrocardiographic and echocardiographic evaluation, as well as the measure of serum biomarkers for myocardial injury, can be critical in identifying poor prognostic factors. Further research is required to understand the implications of our findings in clinical practice.

Unraveling the genetics of transformed splenic marginal zone lymphoma.

The genetic mechanisms associated with splenic marginal zone lymphoma (SMZL) transformation are not well defined. We studied 41 patients with SMZL that eventually underwent large B-cell lymphoma transformation. Tumor material was obtained either only at diagnosis (9 patients), at diagnosis and transformation (18 patients), and only at transformation (14 patients). Samples were categorized in 2 groups: (1) at diagnosis (SMZL, n = 27 samples), and (2) at transformation (SMZL-T, n = 32 samples). Using copy number arrays and a next-generation sequencing custom panel, we identified that the main genomic alterations in SMZL-T involved TNFAIP3, KMT2D, TP53, ARID1A, KLF2, 1q gains, and losses of 9p21.3 (CDKN2A/B) and 7q31-q32. Compared with SMZL, SMZL-T had higher genomic complexity, and higher incidence of TNFAIP3 and TP53 alterations, 9p21.3 (CDKN2A/B) losses, and 6p gains. SMZL and SMZL-T clones arose by divergent evolution from a common altered precursor cell that acquired different genetic alterations in virtually all evaluable cases (92%, 12 of 13 cases). Using whole-genome sequencing of diagnostic and transformation samples in 1 patient, we observed that the SMZL-T sample carried more genomic aberrations than the diagnostic sample, identified a translocation t(14;19)(q32;q13) present in both samples, and detected a focal B2M deletion due to chromothripsis acquired at transformation. Survival analysis showed that KLF2 mutations, complex karyotype, and International Prognostic Index score at transformation were predictive of a shorter survival from transformation (P = .001; P = .042; and P = .007; respectively). In summary, SMZL-T are characterized by higher genomic complexity than SMZL, and characteristic genomic alterations that could represent key players in the transformation event.

The immune microenvironment shapes transcriptional and genetic heterogeneity in chronic lymphocytic leukemia.

In chronic lymphocytic leukemia (CLL), B-cell receptor signaling, tumor-microenvironment interactions, and somatic mutations drive disease progression. To better understand the intersection between the microenvironment and molecular events in CLL pathogenesis, we integrated bulk transcriptome profiling of paired peripheral blood (PB) and lymph node (LN) samples from 34 patients. Oncogenic processes were upregulated in LN compared with PB and in immunoglobulin heavy-chain variable (IGHV) region unmutated compared with mutated cases. Single-cell RNA sequencing (scRNA-seq) distinguished 3 major cell states: quiescent, activated, and proliferating. The activated subpopulation comprised only 2.2% to 4.3% of the total tumor bulk in LN samples. RNA velocity analysis found that CLL cell fate in LN is unidirectional, starts in the proliferating state, transitions to the activated state, and ends in the quiescent state. A 10-gene signature derived from activated tumor cells was associated with inferior treatment-free survival (TFS) and positively correlated with the proportion of activated CD4+ memory T cells and M2 macrophages in LN. Whole exome sequencing (WES) of paired PB and LN samples showed subclonal expansion in LN in approximately half of the patients. Since mouse models have implicated activation-induced cytidine deaminase in mutagenesis, we compared AICDA expression between cases with and without clonal evolution but did not find a difference. In contrast, the presence of a T-cell inflamed microenvironment in LN was associated with clonal stability. In summary, a distinct minor tumor subpopulation underlies CLL pathogenesis and drives the clinical outcome. Clonal trajectories are shaped by the LN milieu, where T-cell immunity may contribute to suppressing clonal outgrowth. The clinical study is registered at clinicaltrials.gov as NCT00923507.

Genetic and phenotypic characterisation of HIV-associated aggressive B-cell non-Hodgkin lymphomas, which do not occur specifically in this population: diagnostic and prognostic implications.

The frequency of aggressive subtypes of B-cell non-Hodgkin lymphoma (B-NHL), such as high-grade B-cell lymphomas (HGBL) with MYC and BCL2 and/or BCL6 rearrangement (HGBL-DH/TH) or Burkitt-like lymphoma (BL) with 11q aberration, is not well known in the HIV setting. We aimed to characterise HIV-associated aggressive B-NHL according to the 2017 WHO criteria, and to identify genotypic and phenotypic features with prognostic impact. Seventy-five HIV-associated aggressive B-NHL were studied by immunohistochemistry (CD10, BCL2, BCL6, MUM1, MYC, and CD30), EBV-encoded RNAs (EBERs), and fluorescence in situ hybridisation (FISH) to evaluate the status of the MYC, BCL2, and BCL6 genes and chromosome 11q. The 2017 WHO classification criteria and the Hans algorithm, for the cell-of-origin classification of diffuse large B-cell lymphomas (DLBCL), were applied. In DLBCL cases, the frequencies of MYC and BCL6 rearrangements (14.9 and 27.7%, respectively) were similar to those described in HIV-negative patients, but BCL2 rearrangements were infrequent (4.3%). MYC expression was identified in 23.4% of DLBCL cases, and coexpression of MYC and BCL2 in 13.0%, which was associated with a worse prognosis. As for BL cases, the expression of MUM1 (30.4%) conferred a worse prognosis. Finally, the prevalence of HGBL-DH/TH and BL-like with 11q aberration are reported in the HIV setting. The phenotypic and genotypic characteristics of HIV-associated aggressive B-NHL are similar to those of the general population, except for the low frequency of BCL2 rearrangements in DLBCL. MYC and BCL2 coexpression in DLBCL, and MUM-1 expression in BL, have a negative prognostic impact on HIV-infected individuals.

Benefits and Risks of Sharing Genomic Data for Research: Comparing the Views of Rare Disease Patients, Informal Carers and Healthcare Professionals.

Assessing public and patients' expectations and concerns about genomic data sharing is essential to promote adequate data governance and engagement in rare diseases genomics research. This cross-sectional study compared the views of 159 rare disease patients, 478 informal carers and 63 healthcare professionals in Northern Portugal about the benefits and risks of sharing genomic data for research, and its associated factors. The three participant groups expressed significantly different views. The majority of patients (84.3%) and informal carers (87.4%) selected the discovery of a cure for untreatable diseases as the most important benefit. In contrast, most healthcare professionals revealed a preference for the development of new drugs and treatments (71.4%), which was the second most selected benefit by carers (48.3%), especially by the more educated (OR (95% CI): 1.58 (1.07-2.34)). Lack of security and control over information access and the extraction of information exceeding research objectives were the two most often selected risks by patients (72.6% and 50.3%, respectively) and carers (60.0% and 60.6%, respectively). Conversely, professionals were concerned with genomic data being used to discriminate citizens (68.3%), followed by the extraction of information exceeding research objectives (54.0%). The latter risk was more frequently expressed by more educated carers (OR (95% CI): 1.60 (1.06-2.41)) and less by those with blue-collar (OR (95% CI): 0.44 (0.25-0.77) and other occupations (OR (95% CI): 0.44 (0.26-0.74)). Developing communication strategies and consent approaches tailored to participants' expectations and needs can benefit the inclusiveness of genomics research that is key for patient-centred care.

Clinico-biological features and outcome of patients with splenic marginal zone lymphoma with histological transformation.

We describe 36 patients with splenic marginal zone lymphoma (SMZL) with transformation (SMZL-T), including 15 from a series of 84 patients with SMZL diagnosed at the Hospital Clinic of Barcelona (HCB) and 21 diagnosed with SMZL-T in other centres. In the HCB cohort, the cumulative incidence of transformation at 5 years was 15%. Predictors for transformation were cytopenias, hypoalbuminaemia, complex karyotype (CK) and both the Intergruppo Italiano Linfomi (ILL) and simplified Haemoglobin, Platelet count, lactate dehydrogenase (LDH) and extrahilar Lymphadenopathy (HPLL)/ABC scores (P < 0·05). The only independent predictor for transformation in multivariate analysis was CK [hazard ratio (HR) 4·025, P = 0·05]. Patients with SMZL-T had a significantly higher risk of death than the remainder (HR 3·89, P < 0·001). Of the 36 patients with SMZL-T, one developed Hodgkin lymphoma and 35 a diffuse large B-cell lymphoma, 71% with a non-germinal centre phenotype. The main features were B symptoms, lymphadenopathy, and high serum LDH. CK was observed in 12/22 (55%) SMZL-T and fluorescence in situ hybridisation detected abnormalities of MYC proto-oncogene, basic helix-loop-helix transcription factor (MYC), B-cell leukaemia/lymphoma 2 (BCL2) and/or BCL6 in six of 14 (43%). In all, 21 patients received immunochemotherapy, six chemotherapy, one radiotherapy and three splenectomy. The complete response (CR) rate was 61% and the median survival from transformation was 4·92 years. Predictors for a worse survival in multivariate analysis were high-risk International Prognostic Index (HR 5·294, P = 0·016) and lack of CR (HR 2·67, P < 0·001).

Genetic and phenotypic attributes of splenic marginal zone lymphoma.

Splenic marginal zone B-cell lymphoma (SMZL) is a heterogeneous clinico-biological entity. The clinical course is variable, multiple genes are mutated with no unifying mechanism, and essential regulatory pathways and surrounding microenvironments are diverse. We sought to clarify the heterogeneity of SMZL by resolving different subgroups and their underlying genomic abnormalities, pathway signatures, and microenvironment compositions to uncover biomarkers and therapeutic vulnerabilities. We studied 303 SMZL spleen samples collected through the IELSG46 multicenter international study (NCT02945319) by using a multiplatform approach. We carried out genetic and phenotypic analyses, defined self-organized signatures, validated the findings in independent primary tumor metadata and in genetically modified mouse models, and determined correlations with outcome data. We identified 2 prominent genetic clusters in SMZL, termed NNK (58% of cases, harboring NF-κB, NOTCH, and KLF2 modules) and DMT (32% of cases, with DNA-damage response, MAPK, and TLR modules). Genetic aberrations in multiple genes as well as cytogenetic and immunogenetic features distinguished NNK- from DMT-SMZLs. These genetic clusters not only have distinct underpinning biology, as judged by differences in gene-expression signatures, but also different outcomes, with inferior survival in NNK-SMZLs. Digital cytometry and in situ profiling segregated 2 basic types of SMZL immune microenvironments termed immune-suppressive SMZL (50% of cases, associated with inflammatory cells and immune checkpoint activation) and immune-silent SMZL (50% of cases, associated with an immune-excluded phenotype) with distinct mutational and clinical connotations. In summary, we propose a nosology of SMZL that can implement its classification and also aid in the development of rationally targeted treatments.

Molecular and functional profiling identifies therapeutically targetable vulnerabilities in plasmablastic lymphoma.

Plasmablastic lymphoma (PBL) represents a rare and aggressive lymphoma subtype frequently associated with immunosuppression. Clinically, patients with PBL are characterized by poor outcome. The current understanding of the molecular pathogenesis is limited. A hallmark of PBL represents its plasmacytic differentiation with loss of B-cell markers and, in 60% of cases, its association with Epstein-Barr virus (EBV). Roughly 50% of PBLs harbor a MYC translocation. Here, we provide a comprehensive integrated genomic analysis using whole exome sequencing (WES) and genome-wide copy number determination in a large cohort of 96 primary PBL samples. We identify alterations activating the RAS-RAF, JAK-STAT, and NOTCH pathways as well as frequent high-level amplifications in MCL1 and IRF4. The functional impact of these alterations is assessed using an unbiased shRNA screen in a PBL model. These analyses identify the IRF4 and JAK-STAT pathways as promising molecular targets to improve outcome of PBL patients.

Percutaneous patent ductus arteriosus closure: Twelve years of experience.

INTRODUCTION: Patent ductus arteriosus, a persistent communication between the descending thoracic aorta and the pulmonary artery, is one of the most common congenital heart defects. Transcatheter occlusion is an effective alternative to surgery and is currently standard of care for most patients. The authors present the results from a single center after twelve years of experience using this technique. METHODS: Retrospective analysis of medical records from all patients referred to a tertiary center for percutaneous ductus closure between January 2006 and September 2018. RESULTS: A total of 221 patients were referred, with a mean age of 5.5 years-old (16 patients were infants, with the youngest aged four months). A Nit-Occlud® coil was used 139 times (62.9%), an Amplatzer™ duct occluder 79 times (35.7%), and vascular plugs were used three times. Percutaneous closure was achieved in every treated patient, with 1.4% maintaining residual shunting. Although higher overall coil device implantation was noted, duct occluder usage has been greater since 2011. Of all the coils, 55% were either 4x4 or 5x4 mm, and 73% of all Amplatzer duct occluders were either 6x4 or 8x6 mm, which correlates to the majority of patients having a small to moderately sized ductus. No complications were noted during the procedure, with a 1.8% post-procedure complication rate (one device embolization after 48 hours and three cases of loss of arterial pulse). CONCLUSIONS: Percutaneous patent ductus arteriosus closure was safe and effective in this setting, with a low global complication rate and similar outcomes to most equivalent centers.

Pulmonary embolism in pediatric age: A retrospective study from a tertiary center.

INTRODUCTION: Pediatric pulmonary embolism (PE) is rare but associated with adverse outcomes. We aimed to characterize PE cases admitted in a tertiary hospital and to evaluate sensitivity of selected PE diagnostic prediction tools. METHODS: Retrospective, descriptive study of PE cases admitted from 2008 to 2020 using data collected from hospital records. Patients were grouped according to PE severity and setting (outpatients vs. inpatients). Links and correlation with demographic characteristics, risk factors, clinical presentation, management, and outcomes were analyzed. PE diagnostic prediction tools were applied. RESULTS: Twenty-nine PE episodes occurred in 27 patients, 62.9% female, mean age 14.1 years. Most PE were central and split between massive or submassive. One was diagnosed in autopsy. Twenty outpatients, all adolescents, were admitted for classic PE symptoms; in half of them the diagnosis had been previously missed. Risk factors included contraceptives (65%), thrombophilia (35%), obesity (20%) and auto-immunity (20%). Eight inpatients, diagnosed during cardiorespiratory deterioration (n = 5), or through incidental radiological findings (n = 3), were younger and had immobilization (87.5%), complex chronic diseases (75%), infections (75%) and central venous catheter (62.5%) as risk factors. Retrospectively, d -dimer testing and adult scores performed better than pediatric scores (sensitivity 92.9%-96% vs. 85.7%-92.9%). Both pediatric scores missed a case with a positive family history. DISCUSSION: Pediatric PE diagnosis is often delayed or missed. Development of pediatric prediction tools from validated adult scores merits being explored. We argue clinical presentation and risk factors may be different in inpatients and outpatients and propose broader reliance on family history.

BTK inhibitors, irrespective of ITK inhibition, increase efficacy of a CD19/CD3-bispecific antibody in CLL.

Bruton tyrosine kinase inhibitors (BTKis) are a preferred treatment of patients with chronic lymphocytic leukemia (CLL). Indefinite therapy with BTKis, although effective, presents clinical challenges. Combination therapy can deepen responses, shorten treatment duration, and possibly prevent or overcome drug resistance. We previously reported on a CD19/CD3-bispecific antibody (bsAb) that recruits autologous T-cell cytotoxicity against CLL cells in vitro. Compared with observations with samples from treatment-naïve patients, T cells from patients being treated with ibrutinib expanded more rapidly and exerted superior cytotoxic activity in response to the bsAb. In addition to BTK, ibrutinib also inhibits interleukin-2 inducible T-cell kinase (ITK). In contrast, acalabrutinib, does not inhibit ITK. Whether ITK inhibition contributes to the observed immune effects is unknown. To better understand how BTKis modulate T-cell function and cytotoxic activity, we cultured peripheral blood mononuclear cells (PBMCs) from BTKi-naive and ibrutinib- or acalabrutinib-treated CLL patients with CD19/CD3 bsAb in vitro. T-cell expansion, activation, differentiation, and cytotoxicity were increased in PBMCs from patients on treatment with either BTKi compared with that observed for BKTi-naïve patients. BTKi therapy transcriptionally downregulated immunosuppressive effectors expressed by CLL cells, including cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) and CD200. CTLA-4 blockade with ipilimumab in vitro increased the cytotoxic activity of the bsAb in BTKi-naïve but not BTKi-treated PBMCS. Taken together, BTKis enhance bsAb-induced cytotoxicity by relieving T cells of immunosuppressive restraints imposed by CLL cells. The benefit of combining bsAb immunotherapy with BTKis needs to be confirmed in clinical trials.

Select Antitumor Cytotoxic CD8+ T Clonotypes Expand in Patients with Chronic Lymphocytic Leukemia Treated with Ibrutinib.

PURPOSE: In chronic lymphocytic leukemia (CLL), the T-cell receptor (TCR) repertoire is skewed and tumor-derived antigens are hypothesized as drivers of oligoclonal expansion. Ibrutinib, a standard treatment for CLL, inhibits not only Bruton tyrosine kinase of the B-cell receptor signaling pathway, but also IL2-inducible kinase of the TCR signaling pathway. T-cell polarization and activation are affected by ibrutinib, but it is unknown whether T cells contribute to clinical response. EXPERIMENTAL DESIGN: High-throughput TCRß sequencing was performed in 77 longitudinal samples from 26 patients with CLL treated with ibrutinib. TCRß usage in CD4+ and CD8+ T cells and granzyme B expression were assessed by flow cytometric analysis. Antitumor cytotoxicity of T cells expanded with autologous CLL cells or with antigen-independent anti-CD3/CD28/CD137 beads was tested. RESULTS: The clonality of the TCR repertoire increased at the time of response. With extended treatment, TCR clonality remained stable in patients with sustained remission and decreased in patients with disease progression. Expanded clonotypes were rarely shared between patients, indicating specificity for private antigens. Flow cytometry demonstrated a predominance of CD8+ cells among expanded clonotypes. Importantly, bulk T cells from responding patients were cytotoxic against autologous CLL cells in vitro and selective depletion of major expanded clonotypes reduced CLL cell killing. CONCLUSIONS: In patients with CLL, established T-cell responses directed against tumor are suppressed by disease and reactivated by ibrutinib.See related commentary by Zent, p. 4465.

Public and patient involvement in health data governance (DATAGov): protocol of a people-centred, mixed-methods study on data use and sharing for rare diseases care and research.

INTRODUCTION: International policy imperatives for the public and patient involvement in the governance of health data coexist with conflicting cross-border policies on data sharing. This can challenge the planning and implementation of participatory data governance in healthcare services locally. Engaging with local stakeholders and understanding how their needs, values and preferences for governing health data can be articulated with policies made at the supranational level is crucial. This paper describes a protocol for a project that aims to coproduce a people-centred model for involving patients and the public in decision-making processes about the use and sharing of health data for rare diseases care and research. METHODS AND ANALYSIS: This multidisciplinary project draws on an explanatory sequential mixed-methods study. A hospital-based survey with patients, informal carers, health professionals and technical staff recruited at two reference centres for rare diseases in Portugal will be conducted first. The qualitative study will follow consisting of semi-structured interviews and scenario-based workshops with a subsample of the participant groups recruited at baseline. Quantitative data will be analysed using descriptive and inferential statistics. Inductive and deductive approaches will be combined to analyse the qualitative interviews. Data from scenario-based workshops will be iteratively compared using the constant comparison method to identify cross-cutting themes and categories. ETHICS AND DISSEMINATION: The Ethics Committee for Health from the University Hospital Centre São João/Faculty of Medicine of University of Porto approved the study protocol (Ref. 99/19). Research findings will be disseminated at academic conferences and science promotion events, and through public meetings involving patient representatives, practitioners, policy-makers and students, a project website and peer-reviewed journal publications.

Cytokeratin expression in plasmablastic lymphoma - a possible diagnostic pitfall in the routine work-up of tumours.

AIMS: Plasmablastic lymphoma (PBL) is a rare aggressive B-cell lymphoma that frequently arises at extranodal sites in the setting of immunosuppression. The diagnosis of PBL is complex, owing to a frequent solid or cohesive growth pattern, and an often unusual immunophenotype. Several case reports have described cytokeratin (CK) expression in PBL, introducing a diagnostic pitfall. The aim of this study was to determine the frequency of CK expression in PBL in the largest series available to date. METHODS AND RESULTS: By using immunohistochemistry in a cohort of 72 PBLs, we identified CK8/18 positivity in 11 of 72 cases (15%) and AE1/3 positivity in six of 65 cases (9%), clearly contrasting with a control series of non-PBL aggressive B-cell lymphomas (one of 96 diffuse large B-cell lymphomas), as well as with data in the literature describing only occasional CK expression in haematological neoplasms. CONCLUSIONS: Our data indicate CK expression in a substantial number (15%) of PBLs. In view of the particular morphological features of PBL and its frequent negativity for the common leukocyte antigen and B-cell markers, this feature represents a pitfall in the routine diagnostic work-up of PBL, and requires more extensive immunohistochemical and molecular characterisation of cases entering the differential diagnosis.

Genetics and epigenetics of leukemia and lymphoma: from knowledge to applications, meeting report of the Josep Carreras Leukaemia Research Institute.

The meeting, which brought together leading scientists and clinicians in the field of leukemia and lymphoma, was held at the new headquarters of the Josep Carreras Leukaemia Research Institute (IJC) in Badalona, Catalonia, Spain, September 19-20, 2019. Its purpose was to highlight the latest advances in our understanding of the molecular mechanisms driving blood cancers, and to discuss how this knowledge can be translated into an improved management of the disease. Special emphasis was placed on the role of genetic and epigenetic heterogeneity, and the exploitation of epigenetic regulation for developing biomarkers and novel treatment approaches.