Development and extensive analytical validation of deep amplicon sequencing for detecting KRAS and NRAS mutations in metastatic colorectal cancer samples.

The presence of wild-type RAS alleles, as determined by genotyping codons 12, 13, 59, 61, 117, and 146, is a prerequisite for personalized anti-EGFR treatment of metastatic colorectal cancer (mCRC) patients. Here we describe analytical validation of in-house developed massively parallel sequencing technology (MPS) in comparison to the in vitro diagnostics (IVD) certified qPCR method. DNA extracted from FFPE samples from CRC patients (n=703) and reference standards (n=33) were tested for KRAS and NRAS mutations in 6 codons of exons 2, 3, and 4 using deep amplicon sequencing (DAS) on a MiSeq benchtop sequencer (Illumina). Two different amplicon lengths and two different library preparation methods (long-RAS and short-RAS) were tested in order to evaluate their impact on DAS performance. In parallel, identical tumor DNA was tested by the following IVD assays: therascreen KRAS RGQ PCR Kit (Qiagen), cobas® KRAS Mutation Test (Roche Diagnostics), and SNaPshot assay (Thermo Fisher Scientific). Both DAS assays detected all the mutations present in reference standards and external quality control samples, except for the artificially generated KRAS codon 146 mutation. The DAS assays performed sufficient analytical specificity and sensitivity (≥0.95). The use of shorter amplicons prolonged the preparation steps but significantly improved the sequencing success rate of FFPE-derived DNA. RAS mutation frequencies in the Czech CRC patients were similar to previous reports, although rare mutations were also detected. DAS with short amplicons is a good strategy for routine assessment of somatic mutations in low-quality FFPE-derived DNA.

Autoimmune anti-N-methyl-D-aspartate receptor encephalitis - paraneoplastic syndrome of ovarian teratoma.

OBJECTIVE: Case presentation and subsequent dia-gnostic and therapeutic procedure of autoimmune encephalitis caused by the presence of ovarian teratoma. CASE REPORT: We describe the case of a young woman with symptoms of an acute psychotic attack unresponsive to antipsychotic treatment. Anti-N-methyl-D-aspartate receptor encephalitis was dia-gnosed within the interdisciplinary cooperation at the University Hospital in Olomouc. CONCLUSION: This type of rare and potentially fatal paraneoplastic limbic encephalitis occurs predominantly in young women. Due to the high variability of neuropsychiatric symptoms, the dia-gnosis of the disease is very difficult, and therefore patients are often primarily incorrectly treated for other neurological or psychiatric diseases. The most prognostically important part is early dia-gnosis and adequate therapy.

CD38-negative relapse in multiple myeloma after daratumumab-based chemotherapy.

We present a case report of a patient relapsing after anti-CD38 treatment (daratumumab). The phenotype of the disease changed during this treatment, and the myeloma clone became CD38 negative and daratumumab refractory. We expected clonal shift, however, based on immunophenotyping, cytogenetics and arrayCGH; the clone was identical as before daratumumab-based treatment with the exception of CD38 negativity. We suggest that the downregulation or loss of CD38 might be an epigenetic "escape mechanism" of malignant plasma cells from antibody-based treatment. The aim of our study was to point out the pitfalls of immunophenotyping and cytogenetics in both assessing the minimal residual disease and clone detection after monoclonal antibody-based therapy.

Prospective study of signalling pathways in myeloma bone disease with regard to activity of the disease, extent of skeletal involvement and correlation to bone turnover markers.

AIMS: The aim of our study was to address the utility of serum levels of selected parameters of myeloma bone disease (MBD) signalling with regard to the pathogenesis of multiple myeloma (MM), activity, markers of bone turnover and extent of skeletal changes. PATIENTS AND METHODS: We assessed prospectively 77 individuals with monoclonal gammopathies - 46 patients with active MM (AMM), 12 patients with smouldering MM (SMM) and 19 individuals with monoclonal gammopathy of undetermined significance (MGUS) to determine the role of HGF, MIP-1α, Syndecan-1, osteoprotegerin, Activin A, DKK1, Annexin A2 and NF-κB. RESULTS: We found significant differences of most of the parameters between MGUS and AMM, and with respect to the activity of MM assessed by International Staging System. Most of the parameters of MBD signalling correlated with traditional markers of bone turnover. CONCLUSIONS: All the signalling pathways were activated in MM with more pronounced osteoclastogenesis in comparison with bone formation but not in MGUS regardless of its risk category, suggesting that MBD is not activated in MGUS until the process of transformation into MM. The parameters of MBD signalling might precede the increase of conventional parameters of bone turnover suggesting their possible role in early indication of anti-resorption therapy.

Clinicopathological correlations of the immunoprofile in diffuse large B-cell lymphoma NOS - a single institutions experience.

The experimental platform in hematooncology is still searching for more valid prognostic and predictive factors on clinical, morphological and molecular levels. The bridge closer to daily practice is so-called translation medicine and from this point of view we have tried to sort diffuse large B-cell lymphoma not otherwise specified. The applied methodological approaches are morphology, indirect immunohistochemistry on formaline-fixed, parrafin-embeded tissue, Hans classifier sorting, expression of Bcl-2, CD5, CD20, CD30 and NfκB proteins in comparison with the clinical (Ann Arbor stage, IPI, aa-IPI, PFS, OS), laboratory and cytogenetic results (complex and simplex karyotypes). Statistical analysis included Cox regressive analysis, Mann-Whitney and Kruska-Wallis test. The interval of PFS and OS has been assessed according to Kaplan-Meier analysis. According to Hans classifier 11 cases (18.7 %) could not be sorted exactly into GCB/nonGCB- like subgroups. All relapsing cases bear negative expression of CD10 and 28 cases of non-relapsing cases showed positive expression of CD10. The "third" - GCB-like/nonGCB-like unsortable subgroups shared a very similar course of PFS with the nonGCB-like subgroup and a worse clinical course of OS. Statistically nonsignificantly better response to chemotherapy was shown by cases with positive Bcl-2 expression of more than 30 %. Statistically nonsignificantly better OS and PFS was shown by cases with a proliferation index Ki67 more than 70 %. The study detected 17 cases (28.8 %) with a nuclear expression of p50 and one case with nuclear expression of p65 (1.7 %) which may imply the possibility of NfκB signaling pathway activation. A statistically nonsignificant realationship of p50 expression and OS/PFS was indicated.

Cutaneous bacillary angiomatosis due to Bartonella quintana in a renal transplant recipient.

Bacillary angiomatosis (BA) is a disorder of neovascular proliferation involving skin and other organs of immunosuppressed patients caused by Bartonella species. BA has been recognized in both immunocompetent and immunodeficient patients, mostly in human immunodeficiency virus (HIV)-infected persons, much more rare in those with other immunodeficiencies, including organ transplantation. Diagnosis is based on serologic analysis, culture and molecular biology [detection of Bartonella species deoxyribonucleic acid (DNA) in tissue biopsy extracts by real-time polymerase chain reaction (PCR)]. All immunosuppressed patients with BA should be treated with antibiotics because of potentially life-threatening course of the disease. We report the first case of cutaneous bacillary angiomatosis due to Bartonella quintana in renal transplant recipient. This presentation demonstrates that BA should be considered a differential diagnosis in immunocompromised patients presenting with fever and cutaneous angioma-like lesions.

Epstein-Barr virus, the germinal centre and the development of Hodgkin's lymphoma.

The relationship between Epstein-Barr virus (EBV) and the germinal centre (GC) of the asymptomatic host remains an enigma. The occasional appearance of EBV-positive germinal centres in some patients, particularly those with a history of immunosuppression, suggests that EBV numbers in the GC are subject to immune control. The relationship, if any, between lymphoid hyperplasia with EBV-positive germinal centres and subsequent or concurrent lymphomagenesis remains to be clarified. As far as the development of EBV-associated Hodgkin's lymphoma is concerned, the suppression of virus replication, mediated by LMP1 on the one hand, and the loss of B-cell receptor signalling on the other, appears to be an important pathogenic mechanism. A further important emerging concept is that alterations in the microenvironment of the EBV-infected B-cell may be important for lymphomagenesis.

Primitive neuroectodermal tumor (PNET) of the lung in an adult woman.

Primary primitive neuroectodermal tumors (PNETs) are extremely rare in the lung and especially in adult women. We describe a case of PNET of the lung with aggressive behavior in 31-year-old woman. Diagnosis was based on histopathological and immunohistochemical studies, and confirmed by molecular genetic analysis of chromosome rearrangements in the EWSR1 gene region. Clinical follow-up, post-mortem findings, and differential diagnosis are also discussed.

[Use of archival formalin-fixed, paraffin-embedded (FFPE) tissue samples for molecular genetic analysis in diffuse large B-cell lymphoma (DLBCL)]. / Vyuzití archivovaných parafínových blocku s formalinem fixovanou tkání (FFPE) pro molekulárne genetickou analýzu u difuzního velkobunecného lymfomu (DLBCL).

The currently valid molecular genetic subclassification of patients with diffuse large B-cell lymphoma (DLBCL) into three prognostic subgroups based on expression profiling has been the objective of numerous genetic studies. In routine clinical practice, however, expression profiling technology remains unavailable for the most of centers. Apart from the technology, in some cases molecular genetic laboratories have problems obtaining high-quality material, i.e. fresh tissues, for RNA isolation to determine gene expression. One possibility is to determine the gene expression from RNA obtained by isolation from formalin-fixed, paraffin-embedded (FFPE) tissue. This pilot study aimed at isolating RNA from FFPE in patients diagnosed with DLBCL and verifying the potential use of such RNA for the expression analysis of 7 selected genes. Although the study showed that it is possible to isolate RNA and determine the expression of the selected genes from archival material, the values of relative expression of some genes in the set were too variable to be used for unambiguous prognostic classification. It was confirmed that retrospective analyses of selected genes may be performed with sufficient material obtained, and that properly archived blocks may be used for molecular biology analyses even after 8 years.

Cellular senescence marker p16INK4a and NFKB1 gene polymorphisms in lower gastro-intestinal acute graft versus host disease.

BACKGROUND: Lower gastrointestinal (GI) graft versus host disease (GVHD) represents a severe complication in allogeneic hematopoietic stem cell transplant (HSCT) recipients with high rates of transplant-related mortality. Deregulated innate immunity reactions are the features of its pathogenesis. Cellular senescence has been considered a program of the innate immunity. We focused on lower GI GVHD from the perspective of cellular senescence. OBJECTIVE: We analyzed the impact of p16INK4a expression, a hallmark of cellular senescence, in intestinal biopsies of patients with lower GI GVHD symptoms and NFKB1 gene polymorphisms (rs3774937 C/T and rs3774959 A/G) on HSCT outcome. STUDY DESIGN: Fifty-two single-center patients who presented with symptoms of lower GI GVHD were analyzed in a retrospective manner. Two SNPs located in the NFKB1 gene regions (rs3774937 C/T and rs3774959 A/G) were genotyped from the peripheral blood samples collected before the start of the conditioning. All patients underwent proctosigmoidoscopy with biopsy of the mucosa. The expression of p16INK4a was analyzed in normal intestinal crypts and stroma. RESULTS: Fifty-two patients (50% male) received HSCT for hematological diseases (acute leukemias in 67%) and developed lower GI symptoms. Patients with p16INK4a expression in the intestinal stroma were in lower risk of developing histological grade 3-4 aGVHD (RR 0.18 [95% CI 0.05-0.65]; p = 0.009). The multivariate linear regression confirmed the independent effect of p16INK4a expression on time of the lower GI aGVHD symptoms onset (Coef. 38.9 [95% CI 12.7-65.1]; p = 0.005). The NFKB1 rs3774937 CC and TT/TC genotype were present in 40 and 80% of patients with p16INK4a expression, respectively (p = 0.04). The rs3774959 AA and GG/AG genotype were present among 43 and 82% of patients with p16INK4a expression, respectively (p = 0.02). Expression of p16INK4a was associated with no clinical variable but NFKB1 genotype. CONCLUSIONS: Our results address possible new mechanisms that may lead to better understanding of HSCT-related immune complications. Cellular senescence may bring novel approaches in GVHD diagnostics and therapy.

Epstein-Barr Virus and the Pathogenesis of Diffuse Large B-Cell Lymphoma.

Epstein-Barr virus (EBV), defined as a group I carcinogen by the World Health Organization (WHO), is present in the tumour cells of patients with different forms of B-cell lymphoma, including Burkitt lymphoma, Hodgkin lymphoma, post-transplant lymphoproliferative disorders, and, most recently, diffuse large B-cell lymphoma (DLBCL). Understanding how EBV contributes to the development of these different types of B-cell lymphoma has not only provided fundamental insights into the underlying mechanisms of viral oncogenesis, but has also highlighted potential new therapeutic opportunities. In this review, we describe the effects of EBV infection in normal B-cells and we address the germinal centre model of infection and how this can lead to lymphoma in some instances. We then explore the recent reclassification of EBV+ DLBCL as an established entity in the WHO fifth edition and ICC 2022 classifications, emphasising the unique nature of this entity. To that end, we also explore the unique genetic background of this entity and briefly discuss the potential role of the tumour microenvironment in lymphomagenesis and disease progression. Despite the recent progress in elucidating the mechanisms of this malignancy, much work remains to be done to improve patient stratification, treatment strategies, and outcomes.

A unique case of AH-dominant type nodular pulmonary amyloidosis presenting as a spontaneous pneumothorax: a case report and review of the literature.

Amyloidosis is a rare metabolic disorder primarily brought on by misfolding of an autologous protein, which causes its local or systemic deposition in an aberrant fibrillar form. It is quite rare for pulmonary tissue to be impacted by amyloidosis; of the three forms it can take when involving pulmonary tissue, nodular pulmonary amyloidosis is the most uncommon. Nodular pulmonary amyloidosis rarely induces clinical symptoms, and most often, it is discovered accidentally during an autopsy or via imaging techniques. Only one case of nodular pulmonary amyloidosis, which manifested as a spontaneous pneumothorax, was found in the literature. In terms of more precise subtyping, nodular amyloidosis is typically AL or mixed AL/AH type. No publications on AH-dominant type of nodular amyloidosis were found in the literature. We present a case of an 81 years-old male with nodular pulmonary AH-dominant type amyloidosis who presented with spontaneous pneumothorax. For a deeper understanding of the subject, this study also provides a review of the literature on cases with nodular pulmonary amyloidosis in relation to precise amyloid fibril subtyping. Since it is often a difficult process, accurate amyloid type identification is rarely accomplished. However, this information is very helpful for identifying the underlying disease process (if any) and outlining the subsequent diagnostic and treatment steps. Even so, it is crucial to be aware of this unit and make sure it is taken into consideration when making a differential diagnosis of pulmonary lesions.

Standardised uptake value of 18F-FDG on staging PET/CT in newly diagnosed patients with different subtypes of non-Hodgkin's lymphoma.

OBJECTIVES: Positron emission tomography using 2-[fluorine-18]-fluoro-2-deoxy-D-glucose ((18) F-FDG) is considered to be the most beneficial imaging method for staging patients with non-Hodgkin's lymphoma (NHL). The intensity of (18) F-FDG accumulation may be determined by calculating the so-called standardised uptake value (SUV). The study aimed at assessing the benefit of SUV(max) determination in staging (18) F-FDG PET/CT in untreated patients with NHL. METHODS: One hundred and forty-nine initial staging (18) F-FDG PET/CT scans performed in patients with NHL between January 2007 and August 2009 were assessed, and the SUV(max) was determined. RESULTS: The highest mean and median values of SUV(max) were observed in patients with diffuse large B-cell lymphoma (DLBCL), the lowest mean and median values were found in small lymphocytic lymphoma. The overlap in SUV(max) < 10 between DLBCL and the other subgroups of NHL was very significant. Statistically, no correlation was found between the lactate dehydrogenase and SUV(max) values. On the other hand, a correlation of the Ki-67 proliferative index of tumour cells and SUV(max) was revealed (r = 0.409, P < 0.001). The geometric mean of SUV(max) in patients with Ki-67 ≤ 60 and those with Ki-67 > 60 was 8.8 and 14.3, respectively (P < 0.001). CONCLUSIONS: The results confirm that SUV(max) is not beneficial for making a more precise diagnosis in most patients with NHL. Correlation of SUV(max) with the Ki-67 values suggests that SUV(max) might have a prognostic values in NHL.

Low Plasma Citrate Levels and Specific Transcriptional Signatures Associated with Quiescence of CD34+ Progenitors Predict Azacitidine Therapy Failure in MDS/AML Patients.

To better understand the molecular basis of resistance to azacitidine (AZA) therapy in myelodysplastic syndromes (MDS) and acute myeloid leukemia with myelodysplasia-related changes (AML-MRC), we performed RNA sequencing on pre-treatment CD34+ hematopoietic stem/progenitor cells (HSPCs) isolated from 25 MDS/AML-MRC patients of the discovery cohort (10 AZA responders (RD), six stable disease, nine progressive disease (PD) during AZA therapy) and from eight controls. Eleven MDS/AML-MRC samples were also available for analysis of selected metabolites, along with 17 additional samples from an independent validation cohort. Except for two patients, the others did not carry isocitrate dehydrogenase (IDH)1/2 mutations. Transcriptional landscapes of the patients' HSPCs were comparable to those published previously, including decreased signatures of active cell cycling and DNA damage response in PD compared to RD and controls. In addition, PD-derived HSPCs revealed repressed markers of the tricarboxylic acid cycle, with IDH2 among the top 50 downregulated genes in PD compared to RD. Decreased citrate plasma levels, downregulated expression of the (ATP)-citrate lyase and other transcriptional/metabolic networks indicate metabolism-driven histone modifications in PD HSPCs. Observed histone deacetylation is consistent with transcription-nonpermissive chromatin configuration and quiescence of PD HSPCs. This study highlights the complexity of the molecular network underlying response/resistance to hypomethylating agents.

Prognostic factors in patients with esophageal carcinoma treated with chemoradiation: single center experience.

BACKGROUND/AIMS: The trimodality therapy including chemotherapy, external beam radiation and surgery is widely accepted as the standard of care in patients with locoregional esophageal carcinoma. METHODOLOGY: We have performed a retrospective analysis of patients with locoregional esophageal carcinoma treated with chemoradiation. RESULTS: One-hundred and fifty-two consecutive patients with non-metastatic adenocarcinoma or squamous cell esophageal carcinoma treated with chemoradiation were included in the present analysis. The median survival of the whole group was 12 months. The estimated 3- and 5-year survival rates were 24% and 19%, respectively. On multivariate analysis, female sex, weight loss and serum albumin were independent negative predictors of survival. Among 140 patients who survived more than 3 months, weight loss, female sex and therapy with paclitaxel were negatively associated with prognosis, and among 109 patients surviving more than 6 months the dose of cisplatin and surgery were independent prognostic factors. Pathologic complete response was not predictive of prognosis. CONCLUSIONS: Long-term survival is obtained in only about 20% of patients with carcinoma of the esophagus treated with chemoradiation. Female sex, weight loss and low serum albumin are independent indicators of poor prognosis. Among treatment-related factors, higher dose of cisplatin and esophagectomy were independent predictors of better prognosis, while administration of paclitaxel was associated with poor prognosis.

Mass Spectrometry Amyloid Typing Is Reproducible across Multiple Organ Sites.

We have determined patient's amyloid subtype through immunohistochemical and proteomic analyses of formalin-fixed, paraffin-embedded (FFPE) tissue samples from two affected organs per patient. Amyloid typing, via immunohistochemistry (IHC) and laser microdissection followed by the combination of liquid chromatography with mass spectrometry (LMD-LC-MS), was performed using tissue samples of the human heart, liver, kidney, tongue, and small intestine from 11 patients, and the results were compared with clinical data. LMD-LC-MS correctly typed AL amyloidosis in all 22 FFPE tissue samples despite tissue origin. In contrast, IHC was successful only in the analysis of eight FFPE tissue samples with differences between the examined organs. In the majority of LMD-LC-MS typed samples, the level of IHC staining intensity for transthyretin and serum amyloid A was the same as that for Ig κ and Ig λ antibodies, suggesting low Ig κ or Ig λ antibodies reactivity and the additional antibody clones were essential for correct typing. Both methods used in the study were found to be suitable for amyloid typing, although LMD-LC-MS yielded more promising results than IHC.

Cardiac amyloidosis: from clinical suspicion to morphological diagnosis.

Amyloidosis is a heterogeneous group of diseases characterised by extracellular accumulation of amyloid in various tissues and organs of the body, leading to alteration and destruction of tissues. Heart involvement is the most important prognostic factor in patients with systemic amyloidosis and the diagnosis and typing of amyloid must be made properly. The clinical picture shows congestive heart failure with predominant right-sided heart failure symptoms in fully developed disease, various types of arrhythmias and characteristic electrocardiography and echocardiography findings. Blood and urine monoclonal protein studies and cardiac biomarkers belong to the spectrum of standard laboratory examinations. Cardiac cardiomyopathy is connected with amyloid based on immunoglobulin light chains, serum amyloid A, transthyretin, atrial natriuretic factor or apolipoprotein A1. In the routine diagnostic algorithm, biopsy specimens are examined using special histological staining, immunohistochemistry and immunofluorescence; proteomic analysis is only performed in specialised centres.

Spontaneous splenic rupture in two patients with hematologic malignancy.

BACKGROUND: Spontaneous splenic rupture (SSR) is a very rare complication described in several hundred patients, mainly as case reports. It is defined as a splenic rupture without antecedent injury. The authors of the present paper describe the only two SSR cases diagnosed at the Hemato-oncology department, coincidentally in one year. PATIENTS: The first patient was admitted to hospital because of planned chemotherapy for relapsed hairy cell leukemia. The second was directed to the Hemato-oncology outpatient department because of anemia and painful splenomegaly diagnosed by a physician. The diagnose of hematologic malignancy (diffuse large B-cell lymphoma) was determined subsequently on the basis of histological examination of the spleen. CONCLUSION: It is necessary to consider SSR not only in patients with known diagnosis of malignant disease but in the patients with negative anamnesis, too. The aim of the paper is to draw attention to the existence of this complication.

Clonality testing of lymphoproliferative disorders in a large cohort of primary and consultant biopsies.

BACKGROUND: Lymphoproliferative disease often presents the clinician and pathologist with a diagnostic dilemma, particularly in the early course of the disease. METHODS: We used modified BIOMED-2 protocols to detect monoclonal expansions of immunoglobulin heavy chain (IgH) and T-cell receptor (TCR) genes in 957 formalin-fixed paraffin-embedded samples from 717 patients. To eliminate false-positive results, heteroduplex analysis was used after PCR reactions. The impact of different fixatives on DNA quality and performance of PCR was assessed. RESULTS: In the class of B lymphomas we detected clonal IgH rearrangement in nearly 80% of cases and in the class of T lymphomas in 64% of cases. Performance of the assays was 94.7% and 92.5% for IgH and TCR clonality, respectively. Clonality rates in various B and T lymphomas were in concordance with previous studies. We also present 10 difficult cases where PCR analysis of IgH and TCR gene rearrangements significantly contributed to a decision on the correct diagnosis. CONCLUSION: These results confirm that the PCR-based analysis is suitable as a routine method and is helpful in establishing a diagnosis in morphologically unclear cases.

A Novel Non-Immunoglobulin (non-Ig)/BCL6 Translocation in Diffuse Large B-Cell Lymphoma Involving Chromosome 10q11.21 Loci and Review on Clinical Consequences of BCL6 Rearrangements.

BCL6 rearrangements (3q27) are the most common chromosomal abnormalities in diffuse large B-cell lymphoma (DLBCL), with numerous immunoglobulin (Ig) and non-Ig genes as partners. In DLBCL, the translocations occur predominantly in the "major breakpoint region" encompassing the first noncoding exon and a part of the first intron of BCL6; few cases with "alternative breakpoint cluster" located 245-285 kb 5' BCL6 were also described. The regulatory sequences of known Ig and non-Ig partners replace the 5' untranslated region of the BCL6 in the same transcriptional orientation. Contrary to Ig/BCL6 fusions typical by high BCL6 gene expression, in non-Ig/BCL6 translocations were observed unexpectedly low BCL6 mRNA levels. From the clinical point of view, the survival rate of DLBCL patients with non-Ig partners is inferior to those with Ig/BCL6 translocations, suggesting that non-Ig/BCL6 fusion is a poor prognostic indicator. Hereby we provide comprehensive information about known non-Ig translocation partners and clinical consequences of BCL6 rearrangements in DLBCL. Moreover, we describe a novel reciprocal translocation t(3;10) in refractory patient with DLBCL with the breaking points at 5' untranslated region of BCL6 and 5' untranslated region of the RASGEF1A gene on chromosome 10q11.21 loci; this rearrangement was associated with low BCL6 and RASGEF1A gene expressions. Our patient harbouring dual chromosomal rearrangement involving BCL2 and BCL6 genes relapsed three-times and died soon; thus, further supporting the notion that non-Ig/BCL6 fusion is a poor prognostic indicator of DLBCL. There is evidence of prognostic value of BCL6 rearrangements also in rituximab era.