Utility of KRAS mutation detection using circulating cell‐free DNA from patients with colorectal cancer

In this study, we evaluated the clinical utility of detecting KRAS mutations in circulating cell‐free (ccf)DNA of metastatic colorectal cancer patients. We prospectively recruited 94 metastatic colorectal cancer patients. Circulating cell‐free DNA was extracted from plasma samples and analyzed for the presence of seven KRAS point mutations. Using the Invader Plus assay with peptide nucleic acid clamping method and digital PCR, KRAS mutations were detected in the ccfDNA in 35 of 39 patients previously determined to have primary tumors containing KRAS mutations using the Luminex method, and in 5 of 55 patients with tumors containing wild‐type KRAS. Curative resection was undertaken in 7 of 34 patients with primary and ccfDNA KRAS mutations, resulting in the disappearance of the mutation from the cell‐free DNA in five of seven patients. Three of these patients had tumor recurrence and KRAS mutations in their ccfDNA reappeared. Epidermal growth factor receptor blockade was administered to 24 of the KRAS tumor wild‐type patients. Of the 24 patients with wild‐type KRAS in their primary tumors, three patients had KRAS mutations in their ccfDNA and did not respond to treatment with epidermal growth factor receptor (EGFR) blockade. We also detected a new KRAS mutation in five patients during chemotherapy with EGFR blockade, before disease progression was detectable with imaging. The detection of KRAS mutations in ccfDNA is an attractive approach for predicting both treatment response and acquired resistance to EGFR blockade, and for detecting disease recurrence.     

Analysis of colorectal cancer‐related mutations by liquid biopsy: Utility of circulating cell‐free DNA and circulating tumor cells

We recruited 56 colorectal cancer patients and compared the mutational spectrum of tumor tissue DNA, circulating cell‐free DNA (ccfDNA) and circulating tumor cell (CTC) DNA (ctcDNA) to evaluate the potential of liquid biopsy to detect heterogeneity of cancer. Tumor tissue DNA, ccfDNA, and ctcDNA were extracted from each patient and analyzed using next‐generation sequencing (NGS) and digital PCR. To maximize yields of CTC, three antibodies were used in the capture process. From 34 untreated patients, 53 mutations were detected in tumor tissue DNA using NGS. Forty‐seven mutations were detected in ccfDNA, including 20 not detected in tissues. Sixteen mutations were detected in ctcDNA, including five not detected in tissues. In 12 patients (35.3%), mutations not found in tumor tissues were detected by liquid biopsy: nine (26.5%) in ccfDNA only and three (8.8%) in ctcDNA only. Combination analysis of the two liquid biopsy samples increased the sensitivity to detect heterogeneity. From 22 stage IV patients with RAS mutations in their primary tumors, RAS mutations were detected in 14 (63.6%) ccfDNA and in eight (36.4%) ctcDNA using digital PCR. Mutations not detected in primary tumors can be identified in ccfDNA and in ctcDNA, indicating the potential of liquid biopsy in complementing gene analysis. Combination analysis improves sensitivity. Sensitivity to detect cancer‐specific mutations is higher in ccfDNA compared with ctcDNA.     

Subclonal evolution of a classical Hodgkin lymphoma from a germinal center B‐cell‐derived mantle cell lymphoma

Composite lymphomas (CL) represent the occurrence of two distinct lymphomas in the same patient. Often, CL share a common cellular origin, thus representing a unique model to investigate the multistep genetic path leading to lymphomagenesis in general and to the specific development of each distinct lymphoma component in particular. Here, we present the molecular analysis of a case consisting of an unusual Hodgkin lymphoma (HL) and a mantle cell lymphoma (MCL), intimately admixed within one another in lymph nodes and bone marrow yet phenotypically distinct, in a patient who first presented with splenic/leukemic MCL two years earlier. MCL and Hodgkin and Reed/Sternberg (HRS) cells harbored identical immunoglobulin (Ig) V_H gene rearrangements with shared somatic mutations, proving their common clonal origin from a (post‐)germinal center (GC) B cell. This also demonstrates the (post‐)GC origin of MCL with mutated IgV genes. Both lymphomas carried the same CCND1/IGH translocation and, unexpectedly for HL, expressed cyclin D1 and OCT2. Thus, HRS cells are able to preserve IGH locus activity (otherwise usually silenced in HL) to promote expression of an oncogene translocated into this locus. Both lymphoma populations further showed an identical TP53 function‐impairing mutation, and later acquired a TP53 heterozygous deletion independently from one another (convergent evolution). The surprisingly close genetic relationship of the lymphomas, together with their histological intermingling and the clinical history of the patient, suggests subclonal evolution of HL from MCL as a plausible pathway in alternative to that so far described in CL, i.e. separate development from a common precursor.     

Molecular analysis of the polymorphisms of thymidylate synthase on cell‐free circulating DNA in blood of patients with advanced colorectal carcinoma

As a key enzyme in folate metabolism, the thymidylate synthase (TS) is important for the synthesis of nucleotides. Its polymorphisms may affect the TS gene expression and the susceptibility for Fluoropyrimidine (FU)‐based chemotherapies. In this study, we assessed the relationship between the TS genotypes and clinical outcome to 5‐FU‐based chemotherapy, and examined whether cell‐free circulating DNA is applicable for these molecular analyses. We combined the variable number tandem repeat (VNTR) and single nucleotide (SNP) polymorphisms of the TS promoter and the deletion variants (1494del6) in the 3′UTR with the occurrence of loss of heterozygosity (LOH) at the microsatellite markers D18S59, D18S1140, and D18S976 mapped up‐ and downstream to the TS locus. Cell‐free blood DNA, tumor tissues, and leukocytes of 51 patients with advanced colorectal cancer were used. Genotyping revealed linkage disequilibrium between TS promoter and 3′UTR (p = 0.03) in blood and leukocytes. Inverse associations of the response to therapy with the number of polymorphisms (p = 0.05) and the 494del6 polymorphism (p < 0.02) were detected on serum DNA. The quantification of serum DNA showed significant correlations with the 1494del6 variant (p = 0.006) in tumor tissues and the combined polymorphisms in leukocytes (p < 0.04). In contrast to the low LOH frequency in blood, LOH spanned more than one marker in the primary tumor of the majority of the patients suggesting the loss of the entire TS locus. Our data provide insight into the molecular diversity of the regulation of the TS gene expression. This is the first study that compares multi‐variant TS genotypes in different clinical specimens.     

Molecular and clinical analysis of Chinese patients with anaplastic lymphoma kinase (ALK)‐rearranged non‐small cell lung cancer

Anaplastic lymphoma kinase (ALK) fusions have been recognized as a therapeutic target in non‐small cell lung cancer (NSCLC). However, molecular signatures and clinical characteristics of the Chinese population with ALK‐rearranged NSCLC are not well elucidated. In the present study, we carried out targeted next‐generation sequencing on tissue and plasma ctDNA samples in 1688 patients with NSCLC. Overall, ALK fusions were detected in 70 patients (4.1%), and the frequencies of ALK fusions detected in tissue and plasma samples were 5.1% and 3.3%, respectively. Additionally, the prevalence of breakpoint locations for EML4‐ALK fusions in ctDNA was significantly correlated with that in tumor tissues (R² = .91, P = .045). According to age, the incidence rates of ALK fusions among young (age <45 years), middle‐aged (between 45 and 70 years) and elderly (>70 years) patients were significantly different (P < .001). In 70 ALK‐rearranged cases, coexistence of epidermal growth factor receptor (EGFR) alterations and ALK fusions was detected in 12 cases (17.1%) and EGFR mutations tended to coexist with non‐EML4‐ALK rearrangements. Notably, novel ALK fusion partners, including TRIM66, SWAP70, WNK3, ERC1, TCF12 and FBN1 were identified in the present study. Among EML4‐ALK fusion variants, patients with variant V1 were younger than patients with variant V3 (P = .023), and TP53 mutations were more frequently concurrent with variant V3 compared with variant V1 (P = .009). In conclusion, these findings provide new insights into the molecular‐clinical profiles of patients with ALK‐rearranged NSCLC that may improve the treatment strategy of this population.     

TP53 mutations in circulating free DNA from Egyptian patients with non-Hodgkin’s lymphoma

Background

P53 protein plays important role in the maintenance of genome stability in mammalian cells; it acts in many processes including cell-cycle checkpoint, DNA repair, apoptosis, and angiogenesis. Mutations of P53 have been reported as common mutations in solid tumours, including non-Hodgkin lymphoma, NHL, and have been implicated in drug resistance, aggression and poor prognosis. Chronic infection with hepatitis C, HCV, has been associated in some studies with increased risk of NHL. HCV is a widespread infection in the Egyptian population. Circulating free DNA (CFDNA) has been shown to be a good source of liver tissue-derived DNA in African and Asian patients with chronic liver disease or hepatocellular carcinoma, HCC. Our previous results have shown TP53 mutations in 5% of CFDNA and 10% of tumours of HCC, with underlying HCV.

Objective

Since previous studies have shown p53 mutations in the DNAs extracted from the NH lymphoid tumours, we have assessed the presence of p53 mutations from exons 5 to 9 in CFDNA in patients with NHL, from Alexandria, Egypt, where HCV is highly prevalent, in a first attempt case-control preliminary study.

Methods

CFDNA was extracted from sera of 20 cases with NHL and 20 negative control individuals. The retrieved serum DNAs were screened for TP53 mutations from exons 5 to 9 using direct sequencing and a PCR-restriction digestion analysis (RFLP). Concentrations of CFDNA were measured using Fluoremetric assay.

Results

Concentrations of CFDNA were significantly higher among NHL patients compared to the negative control individuals indicating a very high release or turn-over of DNA from the tumour into the blood stream among NHL patients. Mutations of p53 determined in NHL cases (30%) were of Arg-176 (1/20: 5%), Phe-238 (1/20: 5%), Ser-249 (2/20; 10%), Lys-249 (1/20: 5%) and Phe-250 (1/20: 5%). No mutations were detected among controls. However, Arg-213 polymorphism was found in 2 cases of NHL (10%) and in 1 case of controls (5%).

Conclusion

Our findings of higher DNA concentrations with some p53 mutations in CFDNA from patients with NHL that match the previous reported p53 mutations from tumour DNA may hold promises that CFDNA may serve as a convenient source of tumour-derived DNA to serve as a promising tool of a non-invasive, low-cost new strategy for earlier detection, diagnosis and follow up of the disease. A large-scale prospective study utilizing CFDNA and DNA from tumours of NHL patients will be required to validate this first trial of utilizing CFDNA from NHL patients.     

Combination of RERG and ZNF671 methylation rates in circulating cell‐free DNA: A novel biomarker for screening of nasopharyngeal carcinoma

Nasopharyngeal carcinoma (NPC) is a prevalent malignancy in Southeast Asia, hence, identifying easily detectable biomarkers for NPC screening is essential for better diagnosis and prognosis. Using genome‐wide and targeted analyses based on next‐generation sequencing approaches, we previously showed that gene promoters are hypermethylated in NPC tissues. To confirm whether DNA methylation rates of genes could be used as biomarkers for NPC screening, 79 histologically diagnosed NPC patients and 29 noncancer patients were recruited. A convenient quantitative analysis of DNA methylation using real‐time PCR (qAMP) was carried out, involving pretreatment of tissue DNA, and circulating cell‐free DNA (ccfDNA) from nonhemolytic plasma, with methylation‐sensitive and/or methylation‐dependent restriction enzymes. The qAMP analyses revealed that methylation rates of RERG, ZNF671, ITGA4, and SHISA3 were significantly higher in NPC primary tumor tissues compared to noncancerous tissues, with sufficient diagnostic accuracy of the area under receiver operating characteristic curves (AUC). Interestingly, higher methylation rates of RERG in ccfDNA were statistically significant and yielded a very good AUC; however, those of ZNF671, ITGA4, and SHISA3 were not significant. Furthermore, the combination of methylation rates of RERG and ZNF671 in ccfDNA showed higher diagnostic accuracy than either of them individually. In conclusion, the methylation rates of specific genes in ccfDNA can serve as novel biomarkers for early detection and screening of NPC.     

Diagnostic and prognostic role of cell‐free DNA testing for colorectal cancer patients

Circulating cell‐free DNA (cfDNA) was found in increased amounts in cancer patients and tumor‐associated molecular alteration can be detected in cancer patient's samples. For this reason, the cfDNA analysis is actually considered as a new concept of liquid biopsy. We evaluated the presence and integrity of plasma cfDNA by ALU‐based qPCR and the methylation profile of OSMR and SFRP1 genes promoter in a large cohort of colorectal cancer (CRC) patients (n = 114) in comparison to healthy subjects (n = 56) and patients with adenomatous lesions (n = 22). Moreover, we studied the prognosis value focusing on histopathological staging and survival. The cfDNA concentration and the integrity index were increased in CRC patients. The ALU83 and ALU244 fragment dosage showed a moderate discriminant capacity between CRC patients and controls and CRC and adenoma patients. Especially, cfDNA was significantly higher in CRC patients at advanced histopathological stage. In addition, the increased cfDNA level was associated with poor prognosis. A comparison of methylation profile in matched tissue and plasma on 25 CRC patients was performed and only three mismatched cases were observed. A lower methylation quantification was observed in cfDNA than tissue DNA. The cfDNA methylation frequency was statistically different in controls, adenoma and CRC patients and this frequency increased with the histopathological stage of tumor. The adenoma and CRC patients methylated cfDNA showed a higher quantity of ALU83 and ALU244. An integrated approach, combining the detection of ALU fragments and cancer type‐specific epigenetic alteration, can improve diagnostic efficiency and better define the prognostic value for CRC disease.     

Tumor volume determines the feasibility of cell‐free DNA sequencing for mutation detection in non‐small cell lung cancer

Next‐generation sequencing (NGS) and digital PCR technologies allow analysis of the mutational profile of circulating cell‐free DNA (cfDNA) in individuals with advanced lung cancer. We have now evaluated the feasibility of cfDNA sequencing for mutation detection in patients with non‐small cell lung cancer at earlier stages. A total of 150 matched tumor and serum samples were collected from non‐small cell lung cancer patients at stages IA–IIIA. Amplicon sequencing with DNA extracted from tumor tissue detected frequent mutations in EGFR (37% of patients), TP53 (39%), and KRAS (10%), consistent with previous findings. In contrast, NGS of cfDNA identified only EGFR, TP53, and PIK3CA mutations in three, five, and one patient, respectively, even though adequate amounts of cfDNA were extracted (median of 4936 copies/mL serum). Next‐generation sequencing showed a high accuracy (98.8%) compared with droplet digital PCR for cfDNA mutation detection, suggesting that the low frequency of mutations in cfDNA was not due to a low assay sensitivity. Whereas the yield of cfDNA did not differ among tumor stages, the cfDNA mutations were detected in seven patients at stages IIA–IIIA and at T2b or T3. Tumor volume was significantly higher in the cfDNA mutation‐positive patients than in the negative patients at stages T2b–T4 (159.1 ± 58.0 vs. 52.5 ± 9.9 cm³, P = 0.014). Our results thus suggest that tumor volume is a determinant of the feasibility of mutation detection with cfDNA as the analyte.     

Reversal of tumor acidosis by systemic buffering reactivates NK cells to express IFN‐γ and induces NK cell‐dependent lymphoma control without other immunotherapies

Like other immune cells, natural killer (NK) cells show impaired effector functions in the microenvironment of tumors, but little is known on the underlying mechanisms. Since lactate acidosis, a hallmark of malignant tissue, was shown to contribute to suppression of effective antitumor immune responses, we investigated the impact of tissue pH and lactate concentration on NK‐cell functions in an aggressive model of endogenously arising B‐cell lymphoma. The progressive loss of IFN‐γ production by NK cells observed during development of this disease could be ascribed to decreased pH values and lactate accumulation in the microenvironment of growing tumors. Interestingly, IFN‐γ expression by lymphoma‐derived NK cells could be restored by transfer of these cells into a normal micromilieu. Likewise, systemic alkalization by oral delivery of bicarbonate to lymphoma‐developing mice was capable of enhancing IFN‐γ expression in NK cells and increasing the NK‐cell numbers in the lymphoid organs where tumors were growing. By contrast, NK‐cell cytotoxicity was dampened in vivo by tumor‐dependent mechanisms that seemed to be different from lactate acidosis and could not be restored in a normal milieu. Most importantly, alkalization and the concomitant IFN‐γ upregulation in NK cells were sufficient to significantly delay tumor growth without any other immunotherapy. This effect was strictly dependent on NK cells.     

Methylation patterns in cell‐free plasma DNA reflect removal of the primary tumor and drug treatment of breast cancer patients

Abnormal DNA methylation is a feature of most types of cancer, which is reflected in cell‐free circulating DNA in plasma. It is, however, unknown whether surgical removal of the tumor and subsequent therapy induces changes in plasma DNA methylation, which can be used to monitor treatment. In this pilot study, methylation in cell‐free plasma DNA of 20 breast cancer patients was determined by the previously developed MethDet‐56 technique. Samples at three time points were analyzed—before surgery (baseline), after surgery (to evaluate the effects of resection) and after surgery on tamoxifen therapy (to determine the effects of treatment). Methylation patterns of healthy controls were used as a reference for all comparisons. Seven promoters were differentially methylated (p < 0.05) in at least one comparison; three changed after surgery; another one changed after beginning of tamoxifen treatment; and four were differentially methylated in baseline versus combined treatment samples. Increased methylation of PR PROX, MDGI, PAX 5 and RARβ2 at baseline (presurgery) diminished toward the healthy controls with the lowest methylation in the combined treatment group. Surgery alone decreased methylation in PAX 5 and RARβ2, whereas tamoxifen treatment changed methylation only in the B promoter of ESR1. Methylation patterns in cell‐free plasma DNA change after surgery and tamoxifen treatment, most significantly—after combined treatment. The baseline (presurgery) patterns become similar to those of healthy controls, suggesting that methylation patterns in cell‐free plasma DNA may be used to monitor treatment.     

Prognostic role of PIK3CA mutations of cell‐free DNA in early‐stage triple negative breast cancer

PIK3CA is an oncogene that encodes the p110α component of phosphatidylinositol 3‐kinase (PI3K); it is the second most frequently mutated gene following the TP53 gene. In the clinical setting, PIK3CA mutations may have favorable prognostic value for hormone receptor‐positive breast cancer patients and, during the past few years, PIK3CA mutations of cell‐free DNA (cfDNA) have attracted attention as a potential noninvasive biomarker of cancer. However, there are few reports on the clinical implications of PIK3CA mutations for TNBC patients. We investigated the PIK3CA major mutation status of cfDNA as a noninvasive biomarker of cancer using droplet digital polymerase chain reaction (ddPCR), which has high level sensitivity and specificity for cancer mutation, in early‐stage 49 triple negative breast cancer (TNBC) patients. A total of 12 (24.4%) of 49 patients had PIK3CA mutations of cfDNA. In a median follow up of 54.4 months, the presence of PIK3CA mutations of cfDNA had significant impacts on relapse‐free survival (RFS; P = 0.0072) and breast cancer‐specific survival (BCSS; P = 0.016), according to the log‐lank test. In a Cox proportional hazards model, the presence of PIK3CA mutations of cfDNA had significant prognostic value in the univariate and multivariate analysis. Additionally, the presence of PIK3CA mutations of cfDNA was significantly correlated with positive androgen receptor phosphorylated form depending on PI3K signaling pathway (pAR) which is independent favorable prognostic factors of TNBC. We demonstrated that the presence of PIK3CA major mutations of cfDNA could be a discriminatory predictor of RFS and BCSS in early‐stage TNBC patients and it was associated with PI3K pathway‐dependent AR phosphorylation.     

Bendamustine is effective therapy in patients with rituximab‐refractory,indolent B‐cell non‐Hodgkin lymphoma

BACKGROUND:

Bendamustine hydrochloride is a novel alkylating agent. In this multicenter study, the authors evaluated the efficacy and toxicity of single‐agent bendamustine in patients with rituximab‐refractory, indolent B‐cell lymphoma.

METHODS:

Eligible patients (N = 100, ages 31‐84 years) received bendamustine at a dose of 120 mg/m² by intravenous infusion on Days 1 and 2 every 21 days for 6 to 8 cycles. Histologies included follicular (62%), small lymphocytic (21%), and marginal zone (16%) lymphomas. Patients had received a median of 2 previous regimens (range, 0‐6 previous regimens), and 36%were refractory to their most recent chemotherapy regimen. Primary endpoints included overall response rate (ORR) and duration of response (DOR). Secondary endpoints were safety and progression‐free survival (PFS).

RESULTS:

An ORR of 75% (a 14% complete response rate, a 3% unconfirmed complete response rate, and a 58% partial response rate) was observed. The median DOR was 9.2 months, and median PFS was 9.3 months. Six deaths were considered to be possibly treatment related. Grade 3 or 4 (determined using National Cancer Institute Common Toxicity Criteria [version 3.0.19]. reversible hematologic toxicities included neutropenia (61%), thrombocytopenia (25%), and anemia (10%). The most frequent nonhematologic adverse events (any grade) included nausea (77%), infection (69%), fatigue (64%), diarrhea (42%), vomiting (40%), pyrexia (36%), constipation (31%), and anorexia (24%).

CONCLUSIONS:

Single‐agent bendamustine produced a high rate of objective responses with acceptable toxicity in patients with recurrent, rituximab‐refractory indolent B‐cell lymphoma. Cancer 2010. © 2010 American Cancer Society.     

BCL2 mutation spectrum in B‐cell non‐Hodgkin lymphomas and patterns associated with evolution of follicular lymphoma

BCL2 is a target of somatic hypermutation in t(14;18) positive and also in a small fraction of t(14;18) negative diffuse large B‐cell lymphoma (DLBCL), suggesting an aberrant role of somatic hypermutation (ASHM). To elucidate the prevalence of BCL2 mutations in lymphomas other than DLBCL, we Sanger‐sequenced the hypermutable region of the BCL2 gene in a panel of 69 mature B‐cell lymphomas, including Richter's syndrome DLBCL, marginal‐zone lymphomas, post‐transplant lymphoproliferative disorders, HIV‐associated and common‐variable immunodeficiency‐associated DLBCL, all known to harbour ASHM‐dependent mutations in other genes, as well as 16 t(14,18) negative and 21 t(14;18) positive follicular lymphomas (FLs). We also investigated the pattern of BCL2 mutations in longitudinal samples from 10 FL patients relapsing to FL or transforming to DLBCL (tFL). By direct sequencing, we found clonally represented BCL2 mutations in 2/16 (13%) of t(14;18) negative FLs, 2/16 (13%) HIV‐DLBCLs, 1/9 (11%) of Richter's syndrome DLBCL, 1/17 (6%) of post‐transplant lymphoproliferative disorders and 1/2 (50%) common‐variable immunodeficiency‐associated DLBCL. The proportion of mutated cases was significantly lower than in FLs carrying the t(14;18) translocation (15/21, 71%). However, the absence of t(14;18) by FISH or PCR and the molecular features of the mutations strongly suggest that BCL2 represents an additional target of ASHM in these entities. Analysis of the BCL2 mutation pattern in clonally related FL/FL and FL/tFL samples revealed two distinct scenarios of genomic evolution: (i) direct evolution from the antecedent FL clone, with few novel clonal mutations acquired by the tFL major clone, and (ii) evolution from a common mutated long‐lived progenitor cell, which subsequently acquired distinct mutations in the FL and in the relapsed or transformed counterpart. Copyright © 2014 John Wiley & Sons, Ltd.     

Characterization of genomic imbalances in diffuse large B‐cell lymphoma by detailed SNP‐chip analysis

The pathogenesis of diffuse large B‐cell lymphomas (DLBCL) is only partly understood. We analyzed 148 DLBCL by single nucleotide polymorphism (SNP)‐chips to characterize genomic imbalances. Seventy‐nine cases were of the germinal center B‐cell like (GCB) type of DLBCL, 49 of the activated B‐cell like (ABC) subtype and 20 were unclassified DLBCL. Twenty‐four regions of recurrent genomic gains and 38 regions of recurrent genomic losses were identified over the whole cohort, with a median of 25 imbalances per case for ABC‐DLBCL and 19 per case for GCB‐DLBCL. Several recurrent copy number changes showed differential frequencies in the GCB‐ and ABC‐DLBCL subgroups, including gains of HDAC7A predominantly in GCB‐DLBCL (38% of cases) and losses of BACH2 and CASP8AP2 predominantly in ABC‐DLBCL (35%), hinting at disparate pathogenetic mechanisms in these entities. Correlating gene expression and copy number revealed a strong gene dosage effect in all tumors, with 34% of probesets showing a concordant expression change in affected regions. Two new potential tumor suppressor genes emerging from the analysis, CASP3 and IL5RA, were sequenced in ten and 16 candidate cases, respectively. However, no mutations were found, pointing to a potential haploinsufficiency effect of these genes, considering their reduced expression in cases with deletions. Our study thus describes differences and similarities in the landscape of genomic aberrations in the DLBCL subgroups in a large collection of cases, confirming already known targets, but also discovering novel copy number changes with possible pathogenetic relevance.