Peter de Nully Brown

Long-term progression-free survival of mantle cell lymphoma after intensive front-line immunochemotherapy with in vivo–purged stem cell rescue: a nonrandomized phase 2 multicenter study by the Nordic Lymphoma Group

Mantle cell lymphoma (MCL) is considered incurable. Intensive immunochemotherapy with stem cell support has not been tested in large, prospective series. In the 2nd Nordic MCL trial, we treated 160 consecutive, untreated patients younger than 66 years in a phase 2 protocol with dose-intensified induction immunochemotherapy with rituximab (R) + cyclophosphamide, vincristine, doxorubicin, prednisone (maxi-CHOP), alternating with R + high-dose cytarabine. Responders received high-dose chemotherapy with BEAM or BEAC (carmustine, etoposide, cytarabine, and melphalan/cyclophosphamide) with R-in vivo purged autologous stem cell support. Overall and complete response was achieved in 96% and 54%, respectively. The 6-year overall, event-free, and progression-free survival were 70%, 56%, and 66%, respectively, with no relapses occurring after 5 years. Multivariate analysis showed Ki-67 to be the sole independent predictor of event-free survival. The nonrelapse mortality was 5%. The majority of stem cell products and patients assessed with polymerase chain reaction (PCR) after transplantation were negative. Compared with our historical control, the Nordic MCL-1 trial, the event-free, overall, and progression-free survival, the duration of molecular remission, and the proportion of PCR-negative stem cell products were significantly increased (P < .001). Intensive immunochemotherapy with in vivo purged stem cell support can lead to long-term progression-free survival of MCL and perhaps cure. Registered at www.isrctn.org as #ISRCTN 87866680.

CHOP-like chemotherapy with or without rituximab in young patients with good-prognosis diffuse large-B-cell lymphoma: 6-year results of an open-label randomised study of the MabThera International Trial (MInT) Group

BACKGROUND The MInT study was the first to show improved 3-year outcomes with the addition of rituximab to a CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisone)-like regimen in young patients with good-prognosis diffuse large-B-cell lymphoma. Extended follow-up was needed to establish long-term effects. METHODS In the randomised open-label MInT study, patients from 18 countries (aged 18-60 years with none or one risk factor according to the age-adjusted International Prognostic Index [IPI], stage II-IV disease or stage I disease with bulk) were randomly assigned to receive six cycles of a CHOP-like chemotherapy with or without rituximab. Bulky and extranodal sites received additional radiotherapy. Randomisation was done centrally with a computer-based tool and was stratified by centre, bulky disease, age-adjusted IPI, and chemotherapy regimen by use of a modified minimisation algorithm that incorporated a stochastic component. Patients and investigators were not masked to treatment allocation. The primary endpoint was event-free survival. Analyses were by intention to treat. This observational study is a follow-up of the MInT trial, which was stopped in 2003, and is registered at ClinicalTrials.gov, number NCT00400907. FINDINGS The intention-to-treat population included 410 patients assigned to chemotherapy alone and 413 assigned to chemotherapy plus rituximab. After a median follow-up of 72 months (range 0·03-119), 6-year event-free survival was 55·8% (95% CI 50·4-60·9; 166 events) for patients assigned to chemotherapy alone and 74·3% (69·3-78·6; 98 events) for those assigned to chemotherapy plus rituximab (difference between groups 18·5%, 11·5-25·4, log-rank p<0·0001). Multivariable analyses showed that event-free survival was affected by treatment group, presence of bulky disease, and age-adjusted IPI and that overall survival was affected by treatment group and presence of bulky disease only. After chemotherapy and rituximab, a favourable subgroup (IPI=0, no bulk) could be defined from a less favourable subgroup (IPI=1 or bulk, or both; event-free survival 84·3% [95% CI 74·2-90·7] vs 71·0% [65·1-76·1], log-rank p=0·005). 18 (4·4%, 95% CI 2·6-6·9) second malignancies occurred in the chemotherapy-alone group and 16 (3·9%, 2·2-6·2) in the chemotherapy and rituximab group (Fishers exact p=0·730). INTERPRETATION Rituximab added to six cycles of CHOP-like chemotherapy improved long-term outcomes for young patients with good-prognosis diffuse large-B-cell lymphoma. The definition of two prognostic subgroups allows a more refined therapeutic approach to these patients than does assessment by IPI alone. FUNDING Hoffmann-La Roche.

Nordic MCL2 trial update: six-year follow-up after intensive immunochemotherapy for untreated mantle cell lymphoma followed by BEAM or BEAC + autologous stem-cell support: still very long survival but late relapses do occur

Mantle cell lymphoma (MCL) is a heterogenic non‐Hodgkin lymphoma entity, with a median survival of about 5 years. In 2008 we reported the early – based on the median observation time of 4 years – results of the Nordic Lymphoma Group MCL2 study of frontline intensive induction immunochemotherapy and autologous stem cell transplantation (ASCT), with more than 60% event‐free survival at 5 years, and no subsequent relapses reported. Here we present an update after a median observation time of 6·5 years. The overall results are still excellent, with median overall survival and response duration longer than 10 years, and a median event‐free survival of 7·4 years. However, six patients have now progressed later than 5 years after end of treatment. The international MCL Prognostic Index (MIPI) and Ki‐67‐expression were the only independent prognostic factors. Subdivided by the MIPI‐Biological Index (MIPI + Ki‐67, MIPI‐B), more than 70% of patients with low‐intermediate MIPI‐B were alive at 10 years, but only 23% of the patients with high MIPI‐B. These results, although highly encouraging regarding the majority of the patients, underline the need of a risk‐adapted treatment strategy for MCL. The study was registered at www.isrctn.org as ISRCTN 87866680.

Routine Bone Marrow Biopsy Has Little or No Therapeutic Consequence for Positron Emission Tomography/Computed Tomography–Staged Treatment-Naive Patients With Hodgkin Lymphoma

PURPOSE To investigate whether bone marrow biopsy (BMB) adds useful information to [(18)F]fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) staging in patients with Hodgkin lymphoma (HL). PATIENTS AND METHODS Newly diagnosed patients with HL undergoing a pretherapeutic staging that encompasses both PET/CT and BMB were included in this retrospective study. The pattern of skeletal FDG uptake was categorized as uni-, bi-, or multifocal (≥ three lesions). Clinical stage, risk assessment, and treatment plan were determined with and without the contribution of BMB results according to the Ann Arbor classification and the guidelines from the German Hodgkin Study Group. RESULTS A total of 454 patients with HL were included of whom 82 (18%) had focal skeletal PET/CT lesions and 27 (6%) had positive BMB. No patients with positive BMB were assessed as having stage I to II disease by PET/CT staging. BMB upstaged five patients, assessed as being stage III before BMB; none of the 454 patients would have been allocated to another treatment on the basis of BMB results. Focal skeletal PET/CT lesions identified positive and negative BMBs with a sensitivity and specificity of 85% and 86%, respectively. The positive and negative predictive values of focal skeletal PET/CT lesions for BMB results were 28% and 99%, respectively. CONCLUSION A consistent finding of this study was the absence of positive BMBs in PET/CT-assessed stage I to II disease. The omission of staging BMB would not have changed the risk assessment or treatment strategy in this cohort of 454 newly diagnosed patients with HL.

In B-cell chronic lymphocytic leukaemia chromosome 17 abnormalities and not trisomy 12 are the single most important cytogenetic abnormalities for the prognosis: A cytogenetic and immunophenotypic study of 480 unselected newly diagnosed patients

Of 560 consecutive, newly diagnosed untreated patients with B CLL submitted for chromosome study, G-banded karyotypes could be obtained in 480 cases (86%). Of these, 345 (72%) had normal karyotypes and 135 (28%) had clonal chromosome abnormalities: trisomy 12 (+12) was found in 40 cases, 20 as +12 alone (+12single), 20 as +12 with additional abnormalities (+12complex). Other frequent findings included abnormalities of 14q, chromosome 17, 13q and 6q. The immunophenotype was typical for CLL in 358 patients (CD5+, Slg(weak), mainly FMC7-) and atypical for CLL in 122 patients (25%) (CD5-, or Slg(strong) or FMC7+). Chromosome abnormalities were found significantly more often in patients with atypical (48%) than in patients with typical CLL phenotype (22%) (P < 0.00005). Also +12complex, 14q+, del6q, and abnormalities of chromosome 17 were significantly more frequent in patients with atypical CLL phenotype, whereas +12single was found equally often in patients with typical and atypical CLL phenotype. The cytomorphology of most of the +12 patients was that of classical CLL irrespective of phenotype. In univariate survival analysis the following cytogenetic findings were significantly correlated to a poor prognosis: chromosome 17 abnormalities, 14q+, an abnormal karyotype, +12complex, more than one cytogenetic event, and the relative number of abnormal mitoses. In multivariate survival analysis chromosome 17 abnormalities were the only cytogenetic findings with independent prognostic value irrespective of immunophenotype. We conclude that in patients with typical CLL immunophenotype, chromosome abnormalities are somewhat less frequent at the time of diagnosis than hitherto believed. +12single is compatible with classical CLL, and has no prognostic influence whereas chromosome 17 abnormalities signify a poor prognosis. In patients with an atypical CLL immunophenotype, chromosome abnormalities including +12complex, 14q+, del 6q and chromosome 17 are found in about 50% of the patients, and in particular chromosome 17 abnormalities suggest a poor prognosis.

Incidence, clinical features and outcome of essential thrombocythaemia in a well defined geographical area

Abstract: In an attempt to characterise the clinical features, incidence and outcome of essential thrombocythaemia (ET) we report our experience in a large unselected series of patients from a well defined region. All new cases of ET in the County of Copenhagen were registered during the period 1977–98. We identified 96 cases of ET, yielding an age‐ and sex‐adjusted annual incidence rate of 0.59/100.000 and a point‐prevalence at last follow up of 11/100.000. The overall incidence rate was 0.31 and 1.00 per 100.000 population during the consecutive periods 1977–89 and 1990–98, respectively, corresponding to a 3.2‐fold increase. Median age at diagnosis was 67 yr (females 68 yr, males 66 yr, range 18–87 yr), and the female to male (F/M) ratio was 2.6:1. At diagnosis, 52% of the patients displayed no ET‐related symptoms and were discovered fortuitously by a routine platelet count. Forty‐eight percent presented with thrombohaemorrhagic phenomena, of which microvascular disturbances of the central nervous system (CNS), extremities and skin were most frequently observed (23%). Compared to patients diagnosed after 1989, patients diagnosed before 1990 had a significantly higher mean platelet count, white blood cell (WBC) count, lactate dehydrogenase (LDH) value and alkaline phosphatase value. With a median follow up of 70 months, 5‐yr survival was 76%, significantly lower than the expected survival of an age‐ and sex‐matched control group (p=0.0052). Thirty‐seven patients experienced a total of 55 thrombohaemorrhagic events during follow‐up, corresponding to an incidence of thrombosis and microvascular disturbances or haemorrhage of 8.1% per pt‐yr and 2.5% per pt‐yr, respectively. The number of patients experiencing thrombosis or microvascular disturbances was significantly higher among the 29 patients who never received acetylsalicylic acid (ASA) compared to the 67 patients who received ASA during follow up (45% vs. 21%; p=0.017). This study provides population‐based data suggesting the benefit of treatment with low‐dose ASA in a non‐selected population of patients with ET.

Increased circulating platelet–leukocyte aggregates in myeloproliferative disorders is correlated to previous thrombosis, platelet activation and platelet count

Abstract: Platelet–leukocyte adhesion may occur as a consequence of platelet activation and possibly plays a key role in the deposition of activated platelets and fibrin in the thrombotic plug. The aim of the present study was to assess by whole blood flow cytometry the presence of circulating platelet–leukocyte aggregates (PLA) and the platelet–leukocyte response to platelet agonist stimulation (ADP and TRAP) in 50 patients with chronic myeloproliferative disorders (MPD) and 30 controls. PLA were identified as platelet–granulocyte/monocyte aggregates (PGMA), platelet–monocyte aggregates (PMA) and defined as the percentage of leukocytes coexpressing the platelet‐specific marker glycoprotein Ib. Compared to controls the mean percentage of PGMA and PMA was increased in unstimulated whole blood from patients with MPD (7.98 vs. 1.76%; p<0.001 and 12.34 vs. 3.2%; p<0.001, respectively). The percentage of PGMA was correlated to the platelet count (r=0.46; p<0.001), percentage of P‐selectin (r=0.69; p<0.001) and thrombospondin (r=0.58; p<0.001) positive platelets and platelet expression of GPIV (r=0.33; p=0.02). The mean percentage of PGMA and PMA was significantly increased in ADP‐stimulated whole blood of patients (57.14 vs. 47.92%; p=0.009 and 54.91 vs. 45.89%; p<0.001, respectively). Compared to patients without a history of thrombosis, patients having experienced microvascular disturbances or a thrombotic event had a higher mean percentage of PGMA and PMA in non‐stimulated whole blood (10.07 vs. 6.34%; p=0.025 and 14.81 vs. 10.48%; p=0.021, respectively) and a higher percentage of PGMA in ADP stimulated whole blood (64.32 vs. 51.50%; p<0.01). These data document an increased frequency of PLA in non‐stimulated whole blood in MPD associated with a previous history of thrombosis or microvascular disturbances.

Increased platelet activation and abnormal membrane glycoprotein content and redistribution in myeloproliferative disorders

Chronic myeloproliferative disorders (MPDs) are characterized by a high incidence of thrombohaemorrhagic complications, possibly caused by platelet dysfunction. In an attempt to define platelet functional abnormalities, we assessed the expression of activation‐dependent membrane proteins in unstimulated and agonist [ADP and thrombin receptor‐activating peptide (TRAP)]‐stimulated platelets using quantitative whole blood flow cytometry in samples from 50 MPD patients and 30 controls. The receptor densities of activation markers and glycoproteins (GPs) were quantified using standardized fluorescent beads. Compared with controls, the mean percentage of P‐selectin‐positive (15·3% vs. 7·2%; P < 0·001) and thrombospondin (TSP)‐positive (6·6% vs. 3·7%; P = 0·003) platelets was increased in unstimulated platelets from patients. Patients having experienced a thrombotic event had a higher mean percentage of TSP‐positive non‐stimulated platelets than patients without a history of thrombosis (9·0% vs. 4·6%; P = 0·02) and a higher GPIV molecules of equivalent fluorochrome (MEF) value (33113 vs. 24471 MEF; P = 0·02). Mean MEF values of monoclonal antibodies (mAbs) against GPIb (34055 vs. 38945 MEF; P < 0·001) and GPIIb/IIIa (1416 vs. 1648 MEF; P < 0·001) were significantly reduced among patients, whereas surface expression of GPIV was increased in patients (28273 vs. 16258 MEF; P < 0·001). In TRAP (10 µmol/l) stimulated whole blood, the MEF of P‐selectin (9611 vs. 13293 MEF; P = 0·004) and CD63 (2385 vs. 5177 MEF; P < 0·001) and the ratio of PAC‐1/GPIIb/IIIa MEF (0·98 vs. 2·00; P < 0·001) was reduced in patients, indicating either a reduced granule GP content or an intrinsic cellular defect in receptor‐mediated granule secretion and activation of the GPIIb/IIIa complex. Expressed as the relative change of MEF compared with unstimulated platelets, TRAP induced decrease of GPIb (7·8% vs. 45%; P < 0·001) and increase of GPIIb/IIIa (49·1% vs. 95·7%; P < 0·001) and GPIV expression (17·8% vs. 55·2%; P < 0·001) was attenuated in patients.

Histone deacetylase 1, 2, 6 and acetylated histone H4 in B- and T-cell lymphomas.

Aims:  Histone deacetylase (HDAC) inhibitors are novel therapeutics in the treatment of peripheral T‐cell lymphoma, unspecified (PTCL) and diffuse large B‐cell lymphoma (DLBCL), where, for unknown reasons, T‐cell malignancies appear to be more sensitive than B‐cell malignancies. The aim was to determine HDAC expression in DLBCL and PTCL which has not previously been investigated.

Genome-wide profiling identifies a DNA methylation signature that associates with TET2 mutations in diffuse large B-cell lymphoma

The discovery that the Ten-Eleven Translocation (TET) hydroxylases cause DNA demethylation has fundamentally changed the notion of how DNA methylation is regulated. Clonal analysis of the hematopoetic stem cell compartment suggests that TET2 mutations can be early events in hematologic cancers and recent investigations have shown TET2 mutations in diffuse large B-cell lymphoma. However, the detection rates and the types of TET2 mutations vary, and the relation to global methylation patterns has not been investigated. Here, we show TET2 mutations in 12 of 100 diffuse large B-cell lymphomas with 7% carrying loss-of-function and 5% carrying missense mutations. Genome-wide methylation profiling using 450K Illumina arrays identified 315 differentially methylated genes between TET2 mutated and TET2 wild-type cases. TET2 mutations are primarily associated with hypermethylation within CpG islands (70%; P<0.0001), and at CpG-rich promoters (60%; P<0.0001) of genes involved in hematopoietic differentiation and cellular development. Hypermethylated loci in TET2 mutated samples overlap with the bivalent (H3K27me3/H3K4me3) silencing mark in human embryonic stem cells (P=1.5×10−30). Surprisingly, gene expression profiling showed that only 11% of the hypermethylated genes were down-regulated, among which there were several genes previously suggested to be tumor suppressors. A meta-analysis suggested that the 35 hypermethylated and down-regulated genes are associated with the activated B-cell-like type of diffuse large B-cell lymphoma in other studies. In conclusion, our data suggest that TET2 mutations may cause aberrant methylation mainly of genes involved in hematopoietic development, which are silenced but poised for activation in human embryonic stem cells.