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1.
Haematologica ; 105(10): 2432-2439, 2020 10 01.
Article in English | MEDLINE | ID: mdl-33054083

ABSTRACT

Myeloid and lymphoid malignancies are postulated to have distinct pathogenetic mechanisms. The recent observation that patients with a myeloproliferative neoplasm have an increased risk of developing lymphoproliferative malignancy has challenged this assumption. We collected a nationwide cohort of patients with both malignancies. Patients diagnosed in 1990-2015 were identified through the national Danish Pathology Registry. We identified 599 patients with myeloproliferative neoplasm and a concomitant or subsequent diagnosis of lymphoma. Histopathological review of the diagnostic samples from each patient led to a final cohort of 97 individuals with confirmed dual diagnoses of myeloproliferative neoplasm and lymphoma. The age range at diagnosis was 19-94 years (median: 71 years). To avoid the inclusion of cases of therapy-induced myeloproliferative neoplasm occurring in patients previously treated for lymphoma, only patients with myeloproliferative neoplasm diagnosed unequivocally before the development of lymphoma were included. The average time interval between the diagnoses of the two malignancies was 1.5 years. In the majority of patients (90%) both diagnoses were established within 5 years from each other. Among the lymphoma entities, the frequency of peripheral T-cell lymphomas was markedly increased. Interestingly, all but one of the T-cell lymphomas were of angioimmunoblastic type. These findings suggest that myeloproliferative neoplasm and lymphoproliferative malignancy developing in the same patient may have common pathogenetic events, possibly already at progenitor level. We believe that the molecular characterization of the newly developed biorepository will help to highlight the mechanisms driving the genesis and clonal evolution of these hematopoietic malignancies.


Subject(s)
Hematologic Diseases , Hematologic Neoplasms , Lymphoma, T-Cell, Peripheral , Myeloproliferative Disorders , Adult , Aged , Aged, 80 and over , Cohort Studies , Hematologic Neoplasms/diagnosis , Hematologic Neoplasms/epidemiology , Hematologic Neoplasms/etiology , Humans , Middle Aged , Myeloproliferative Disorders/diagnosis , Myeloproliferative Disorders/epidemiology , Young Adult
2.
Cancer Epidemiol ; 61: 133-138, 2019 08.
Article in English | MEDLINE | ID: mdl-31254794

ABSTRACT

PURPOSE: Hardly anything is known about the aetiology of thymoma. This paper presents data regarding tobacco smoking and alcohol consumption in relation to thymoma from the first case-control study performed on this rare tumour. METHODS: A European multi-centre case-control study including incident cases aged 35-69 years with thymoma between 1995 and 1997, was conducted in seven countries. A set of controls, used in seven parallel case-control studies by the same research group was used, including population-based controls from five countries and hospital controls with colon cancer from two countries. Altogether 103 cases, accepted by a reference pathologist, 712 colon cancer controls, and 2071 population controls were interviewed. RESULTS: Tobacco smoking was moderately related with thymoma (OR 1.4, 95% CI 0.9-2.2), and a tendency to dose-response was shown (p = 0.04), with an increased risk for heavy smokers defined as ≥41 pack-years (OR 2.1, 95% CI 1.1-3.9). A high consumption of spirits defined as ≥25 g of alcohol per day was associated with an increased risk of thymoma (OR 2.4, 95% CI 1.1-5.4), whereas no association was found with beer or wine. CONCLUSIONS: Tobacco smoking and a high intake of spirits were indicated as risk factors for thymoma.


Subject(s)
Alcohol Drinking/adverse effects , Thymoma/etiology , Tobacco Smoking/adverse effects , Adult , Aged , Case-Control Studies , Female , Humans , Male , Middle Aged , Risk Factors
3.
Blood Adv ; 2(18): 2400-2411, 2018 09 25.
Article in English | MEDLINE | ID: mdl-30254104

ABSTRACT

Despite the recent progress in treatment of multiple myeloma (MM), it is still an incurable malignant disease, and we are therefore in need of new risk stratification tools that can help us to understand the disease and optimize therapy. Here we propose a new subtyping of myeloma plasma cells (PCs) from diagnostic samples, assigned by normal B-cell subset associated gene signatures (BAGS). For this purpose, we combined fluorescence-activated cell sorting and gene expression profiles from normal bone marrow (BM) Pre-BI, Pre-BII, immature, naïve, memory, and PC subsets to generate BAGS for assignment of normal BM subtypes in diagnostic samples. The impact of the subtypes was analyzed in 8 available data sets from 1772 patients' myeloma PC samples. The resulting tumor assignments in available clinical data sets exhibited similar BAGS subtype frequencies in 4 cohorts from de novo MM patients across 1296 individual cases. The BAGS subtypes were significantly associated with progression-free and overall survival in a meta-analysis of 916 patients from 3 prospective clinical trials. The major impact was observed within the Pre-BII and memory subtypes, which had a significantly inferior prognosis compared with other subtypes. A multiple Cox proportional hazard analysis documented that BAGS subtypes added significant, independent prognostic information to the translocations and cyclin D classification. BAGS subtype analysis of patient cases identified transcriptional differences, including a number of differentially spliced genes. We identified subtype differences in myeloma at diagnosis, with prognostic impact and predictive potential, supporting an acquired B-cell trait and phenotypic plasticity as a pathogenetic hallmark of MM.


Subject(s)
B-Lymphocyte Subsets/metabolism , Multiple Myeloma/diagnosis , Multiple Myeloma/mortality , Phenotype , B-Lymphocyte Subsets/immunology , Biomarkers, Tumor , Gene Expression Profiling , Humans , Immunophenotyping , Multiple Myeloma/etiology , Prognosis , Survival Analysis , Transcriptome
4.
Blood Adv ; 2(5): 559-564, 2018 03 13.
Article in English | MEDLINE | ID: mdl-29523528

ABSTRACT

Stringent complete remission (sCR) of acute myeloid leukemia is defined as normal hematopoiesis after therapy. Less sCR, including non-sCR, was introduced as insufficient blood platelet, neutrophil, or erythrocyte recovery. These latter characteristics were defined retrospectively as postremission transfusion dependency and were suggested to be of prognostic value. In the present report, we evaluated the prognostic impact of achieving sCR and non-sCR in the Danish National Acute Leukaemia Registry, including 769 patients registered with classical CR (ie, <5% blasts in the postinduction bone marrow analysis). Individual patients were classified as having sCR (n = 360; 46.8%) or non-sCR (n = 409; 53.2%) based on data from our national laboratory and transfusion databases. Survival analysis revealed that patients achieving sCR had superior overall survival (hazard ratio [HR], 1.34; 95% confidence interval [CI], 1.10-1.64) as well as relapse-free survival (HR, 1.25; 95% CI, 1.03-1.51) compared with those with non-sCR after adjusting for covariates. Cox regression analysis regarding the impact of the stringent criteria for blood cell recovery identified these as significant and independent variables. In conclusion, this real-life register study supports the international criteria for response evaluation on prognosis and, most importantly, documents each of the 3 lineage recovery criteria as contributing independently.


Subject(s)
Leukemia, Myeloid, Acute/diagnosis , Adult , Aged , Cell Lineage , Denmark/epidemiology , Humans , Leukemia, Myeloid, Acute/epidemiology , Leukemia, Myeloid, Acute/mortality , Leukemia, Myeloid, Acute/therapy , Middle Aged , Prognosis , Registries , Remission Induction/methods , Survival Analysis
5.
PLoS One ; 13(3): e0193249, 2018.
Article in English | MEDLINE | ID: mdl-29513759

ABSTRACT

Diagnostic and prognostic evaluation of chronic lymphocytic leukemia (CLL) involves blood cell counts, immunophenotyping, IgVH mutation status, and cytogenetic analyses. We generated B-cell associated gene-signatures (BAGS) based on six naturally occurring B-cell subsets within normal bone marrow. Our hypothesis is that by segregating CLL according to BAGS, we can identify subtypes with prognostic implications in support of pathogenetic value of BAGS. Microarray-based gene-expression samples from eight independent CLL cohorts (1,024 untreated patients) were BAGS-stratified into pre-BI, pre-BII, immature, naïve, memory, or plasma cell subtypes; the majority falling within the memory (24.5-45.8%) or naïve (14.5-32.3%) categories. For a subset of CLL patients (n = 296), time to treatment (TTT) was shorter amongst early differentiation subtypes (pre-BI/pre-BII/immature) compared to late subtypes (memory/plasma cell, HR: 0.53 [0.35-0.78]). Particularly, pre-BII subtype patients had the shortest TTT among all subtypes. Correlates derived for BAGS subtype and IgVH mutation (n = 405) revealed an elevated mutation frequency in late vs. early subtypes (71% vs. 45%, P < .001). Predictions for BAGS subtype resistance towards rituximab and cyclophosphamide varied for rituximab, whereas all subtypes were sensitive to cyclophosphamide. This study supports our hypothesis that BAGS-subtyping may be of tangible prognostic and pathogenetic value for CLL patients.


Subject(s)
B-Lymphocyte Subsets/metabolism , Bone Marrow/metabolism , Leukemia, Lymphocytic, Chronic, B-Cell/classification , Leukemia, Lymphocytic, Chronic, B-Cell/metabolism , Adult , Aged , Aged, 80 and over , Antineoplastic Agents, Alkylating/therapeutic use , Antineoplastic Agents, Immunological/therapeutic use , Cyclophosphamide/therapeutic use , Drug Resistance, Neoplasm/physiology , Female , Gene Expression Regulation, Neoplastic , Humans , Leukemia, Lymphocytic, Chronic, B-Cell/therapy , Male , Microarray Analysis , Middle Aged , Prognosis , Proof of Concept Study , Retrospective Studies , Rituximab/therapeutic use , Survival Analysis , Time-to-Treatment
6.
Cytometry B Clin Cytom ; 94(1): 121-128, 2018 01.
Article in English | MEDLINE | ID: mdl-29024461

ABSTRACT

The diagnostic criteria for CLL rely on morphology and immunophenotype. Current approaches have limitations affecting reproducibility and there is no consensus on the role of new markers. The aim of this project was to identify reproducible criteria and consensus on markers recommended for the diagnosis of CLL. ERIC/ESCCA members classified 14 of 35 potential markers as "required" or "recommended" for CLL diagnosis, consensus being defined as >75% and >50% agreement, respectively. An approach to validate "required" markers using normal peripheral blood was developed. Responses were received from 150 participants with a diagnostic workload >20 CLL cases per week in 23/150 (15%), 5-20 in 82/150 (55%), and <5 cases per week in 45/150 (30%). The consensus for "required" diagnostic markers included: CD19, CD5, CD20, CD23, Kappa, and Lambda. "Recommended" markers potentially useful for differential diagnosis were: CD43, CD79b, CD81, CD200, CD10, and ROR1. Reproducible criteria for component reagents were assessed retrospectively in 14,643 cases from 13 different centers and showed >97% concordance with current approaches. A pilot study to validate staining quality was completed in 11 centers. Markers considered as "required" for the diagnosis of CLL by the participants in this study (CD19, CD5, CD20, CD23, Kappa, and Lambda) are consistent with current diagnostic criteria and practice. Importantly, a reproducible approach to validate and apply these markers in individual laboratories has been identified. Finally, a consensus "recommended" panel of markers to refine diagnosis in borderline cases (CD43, CD79b, CD81, CD200, CD10, and ROR1) has been defined and will be prospectively evaluated. © 2017 International Clinical Cytometry Society.


Subject(s)
Leukemia, Lymphocytic, Chronic, B-Cell/diagnosis , Biomarkers, Tumor/metabolism , Flow Cytometry/methods , Humans , Immunophenotyping/methods , Leukemia, Lymphocytic, Chronic, B-Cell/metabolism , Pilot Projects , Reproducibility of Results , Retrospective Studies
7.
Leuk Lymphoma ; 58(11): 2695-2704, 2017 11.
Article in English | MEDLINE | ID: mdl-28393658

ABSTRACT

The origin of multiple myeloma depends on interactions with stromal cells in the course of normal B-cell differentiation and evolution of immunity. The concept of the present study is that genes involved in MM pathogenesis, such as immune response genes, can be identified by screening for single-nucleotide polymorphisms (SNPs) involved in the immune response and a subsequent statistical analysis that focusses on the association of SNPs, certain haplotypes or SNP-SNP interactions with MM risk and prognosis. We genotyped 348 Danish patients and 355 controls for 13 SNPs located in the TNFA, IL-4, IL-6, IL-10 and CHI3L1 gene promoters. The occurrence of single polymorphisms, haplotypes and SNP-SNP interactions were statistically analyzed for association with disease risk and outcome following high-dose therapy. Identified genes that carried SNPs or haplotypes that were identified as risk or prognostic factors were studied for expression in normal B-cell subsets and myeloma plasma cells. We observed a significantly reduced risk when harboring the TNFA-238A allele (OR = 0.51 (0.29-0.86)) and interactions between the TNFA-1031T/C * and IL-10 -3575T/A (p = .007) as well as the TNFA-308G/A * and IL-10-1082G/A (p = .008) allels. By statistical approaches, we observed association between prognosis and the TNFA-857CC genotype (HR = 2.80 (1.29-6.10)) and IL-10-1082GG + GA genotypes (HR = 1.93 (1.07-3.49)) and interactions between IL-6-174G/C and IL-10-3575T/A (p = .001) and between TNFA-308G/A and IL-4-1098T/G (p= .005). The 'risk genes' were analyzed for expression in normal B-cell subsets (N = 6) from seven healthy donors and we found TNFA and IL-6 expressed both in naïve and in memory B cells when compared to preBI, II, immature and plasma cells. The 'prognosis genes' CHI3L1, IL-6 and IL-10 were differential expressed in malignant plasma cells when comparing poor and good prognosis groups based on to the TC classification. In summary, these findings suggest that TNFA, IL-4, IL-6, IL-10 and CHI3L1 might be important players in MM pathogenesis during disease initiation and drug resistance in multiple myeloma.


Subject(s)
Cytokines/genetics , Inflammation Mediators/metabolism , Multiple Myeloma/genetics , Polymorphism, Single Nucleotide , Promoter Regions, Genetic/genetics , Alleles , Chitinase-3-Like Protein 1/genetics , Female , Gene Expression , Gene Frequency , Genotype , Haplotypes , Humans , Interleukin-10/genetics , Interleukin-4/genetics , Interleukin-6/genetics , Male , Middle Aged , Risk Factors , Survival Analysis , Tumor Necrosis Factor-alpha/genetics
8.
PLoS One ; 12(1): e0169286, 2017.
Article in English | MEDLINE | ID: mdl-28052121

ABSTRACT

BACKGROUND: Toxicity of the oral and gastrointestinal mucosa induced by high-dose melphalan is a clinical challenge with no documented prophylactic interventions or predictive tests. The aim of this study was to describe molecular changes in human oral mucosa and to identify biomarkers correlated with the grade of clinical mucositis. METHODS AND FINDINGS: Ten patients with multiple myeloma (MM) were included. For each patient, we acquired three buccal biopsies, one before, one at 2 days, and one at 20 days after high-dose melphalan administration. We also acquired buccal biopsies from 10 healthy individuals that served as controls. We analyzed the biopsies for global gene expression and performed an immunohistochemical analysis to determine HLA-DRB5 expression. We evaluated associations between clinical mucositis and gene expression profiles. Compared to gene expression levels before and 20 days after therapy, at two days after melphalan treatment, we found gene regulation in the p53 and TNF pathways (MDM2, INPPD5, TIGAR), which favored anti-apoptotic defense, and upregulation of immunoregulatory genes (TREM2, LAMP3) in mucosal dendritic cells. This upregulation was independent of clinical mucositis. HLA-DRB1 and HLA-DRB5 (surface receptors on dendritic cells) were expressed at low levels in all patients with MM, in the subgroup of patients with ulcerative mucositis (UM), and in controls; in contrast, the subgroup with low-grade mucositis (NM) displayed 5-6 fold increases in HLA-DRB1 and HLA-DRB5 expression in the first two biopsies, independent of melphalan treatment. Moreover, different splice variants of HLA-DRB1 were expressed in the UM and NM subgroups. CONCLUSIONS: Our results revealed that, among patients with MM, immunoregulatory genes and genes involved in defense against apoptosis were affected immediately after melphalan administration, independent of the presence of clinical mucositis. Furthermore, our results suggested that the expression levels of HLA-DRB1 and HLA-DRB5 may serve as potential predictive biomarkers for mucositis severity.


Subject(s)
Gene Expression Regulation , Melphalan/adverse effects , Multiple Myeloma/drug therapy , Stomatitis/chemically induced , Stomatitis/genetics , Aged , Biopsy , Dose-Response Relationship, Drug , Female , Follow-Up Studies , HLA-DRB5 Chains/immunology , Hematopoietic Stem Cell Transplantation , Humans , Immunohistochemistry , Male , Melphalan/administration & dosage , Middle Aged , Mouth Mucosa/immunology , Mouth Mucosa/metabolism , Mouth Mucosa/pathology , Stomatitis/blood , Stomatitis/immunology
9.
Haematologica ; 101(12): 1451-1459, 2016 12.
Article in English | MEDLINE | ID: mdl-27903712

ABSTRACT

The concept of the myeloma stem cell may have important therapeutic implications, yet its demonstration has been hampered by a lack of consistency in terms and definitions. Here, we summarize the current documentation and propose single-cell in vitro studies for future translational studies. By the classical approach, a CD19-/CD45low/-/CD38high/CD138+ malignant plasma cell, but not the CD19+/CD38low/- memory B cell compartment, is enriched for tumorigenic cells that initiate myeloma in xenografted immunodeficient mice, supporting that myeloma stem cells are present in the malignant PC compartment. Using a new approach, analysis of c-DNA libraries from CD19+/CD27+/CD38- single cells has identified clonotypic memory B cell, suggested to be the cell of origin. This is consistent with multiple myeloma being a multistep hierarchical process before or during clinical presentation. We anticipate that further characterization will require single cell geno- and phenotyping combined with clonogenic assays. To implement such technologies, we propose a revision of the concept of a myeloma stem cell by including operational in vitro assays to describe the cellular components of origin, initiation, maintenance, and evolution of multiple myeloma. These terms are in accordance with recent (2012) consensus statements on the definitions, assays, and nomenclature of cancer stem cells, which is technically precise without completely abolishing established terminology. We expect that this operational model will be useful for future reporting of parameters used to identify and characterize the multiple myeloma stem cells. We strongly recommend that these parameters include validated standard technologies, reproducible assays, and, most importantly, supervised prospective sampling of selected biomaterial which reflects clinical stages, disease spectrum, and therapeutic outcome. This framework is key to the characterization of the cellular architecture of multiple myeloma and its use in precision medicine.


Subject(s)
Multiple Myeloma/etiology , Multiple Myeloma/metabolism , Neoplastic Stem Cells/metabolism , Animals , Antineoplastic Agents/pharmacology , Antineoplastic Agents/therapeutic use , B-Lymphocytes/metabolism , B-Lymphocytes/pathology , Biomarkers , Cell Plasticity , Cell Self Renewal , Drug Resistance, Neoplasm , Genetic Variation , Humans , Multiple Myeloma/drug therapy , Multiple Myeloma/pathology , Neoplastic Stem Cells/drug effects , Neoplastic Stem Cells/pathology , Phenotype
10.
Exp Hematol ; 44(10): 982-990.e11, 2016 10.
Article in English | MEDLINE | ID: mdl-27297329

ABSTRACT

The rare memory B cells in thymus (Thy) are considered the cells of origin for primary mediastinal large B-cell lymphoma. The objectives of the present study were to characterize the normal memory B-cell compartment in Thy and to support its association with primary mediastinal B-cell lymphoma. Seven paired human tissue samples from Thy and sternum bone marrow (BM) were harvested during cardiac surgery. B-cell subsets were phenotyped by Euroflow standard and fluorescence-activated cell sorting for microarray analysis on the Human Exon 1.0 ST Arrays platform. Differentially expressed genes between Thy and BM memory B cells were identified and correlated with the molecular subclasses of diffuse large B-cell lymphoma. Within Thy, 4% (median; range 2%-14%) of the CD45(+) hematopoietic cells were CD19(+) B cells, with a major fraction being CD27(+)/CD38(-) memory B cells (median 80%, range 76%-93%). The BM contained 14% (median; range 3%-27%), of which only a minor fraction (median 5%, range 2%-10%) were memory B cells. Global gene expression analysis of the memory B-cell subsets from the two compartments identified 133 genes upregulated in Thy, including AICDA, REL, STAT1, TNF family, SLAMF1, CD80, and CD86. In addition, exons 4 and 5 in the 3' end of AICDA were more highly expressed in Thy than in BM. The Thy memory B-cell gene profile was overexpressed in primary mediastinal B-cell lymphoma compared with other diffuse large B-cell lymphoma subclasses. The present study describes a Thy memory B-cell subset and its gene profile correlated with primary mediastinal B-cell lymphomas, suggesting origin from Thy memory B cells.


Subject(s)
B-Lymphocyte Subsets/metabolism , B-Lymphocytes/metabolism , Lymphoma, Large B-Cell, Diffuse/diagnosis , Lymphoma, Large B-Cell, Diffuse/metabolism , Aged , B-Lymphocyte Subsets/immunology , B-Lymphocyte Subsets/pathology , B-Lymphocytes/immunology , B-Lymphocytes/pathology , Biomarkers , Female , Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Humans , Immunologic Memory , Immunophenotyping , Lymphocyte Count , Lymphoma, Large B-Cell, Diffuse/genetics , Lymphoma, Large B-Cell, Diffuse/immunology , Male , Middle Aged , Organ Specificity/genetics , Phenotype , Signal Transduction
11.
Acta Haematol ; 135(2): 81-7, 2016.
Article in English | MEDLINE | ID: mdl-26505268

ABSTRACT

BACKGROUND/AIMS: Bone marrow aspiration (BMA) is an essential procedure in the examination of hematological disorders, but there is limited evidence as to whether the aspiration rate affects specimen quality. We aimed to assess the specimen quality and pain intensity using slow (S-technique) or rapid (R-technique) aspiration. METHODS: This was a single-center, prospective, randomized patient- and assessor-blinded study of 482 patients scheduled for BMA. Specimen quality was evaluated by grading bone marrow (BM) cellularity and counting the number of marrow particles. Pain was assessed using a visual analog scale (VAS). RESULTS: We found a significant difference between the 2 groups with regard to the quality of specimens. For cellularity, the odds ratio (OR) for having a poor quality aspirate using the S-technique versus the R-technique was 3.05 [confidence interval (CI) 1.79-5.31]. For BM particles, the quality of specimens with the S-technique proved to be poor compared with the R-technique (OR 2.52; CI 1.51-4.28). We found a statistically significant difference of 1 VAS point (p < 0.001) of the median pain intensity in favor of the S-technique. CONCLUSION: Even though the pain intensity is significantly higher with the R-technique, the median difference is relatively small. We propose that the R-technique is preferable to the S-technique due to better specimen quality.


Subject(s)
Bone Marrow Cells/cytology , Bone Marrow Examination/methods , Pain/physiopathology , Specimen Handling/methods , Adolescent , Adult , Aged , Aged, 80 and over , Cross-Over Studies , Female , Humans , Male , Middle Aged , Odds Ratio , Prospective Studies , Young Adult
12.
PLoS One ; 10(10): e0139329, 2015.
Article in English | MEDLINE | ID: mdl-26448050

ABSTRACT

BACKGROUND: Malignant B-cell clones are affected by both acquired genetic alterations and by inherited genetic variations changing the inflammatory tumour microenvironment. METHODS: We investigated 50 inflammatory response gene polymorphisms in 355 B-cell non-Hodgkin's lymphoma (B-NHL) samples encompassing 216 diffuse large B cell lymphoma (DLBCL) and 139 follicular lymphoma (FL) and 307 controls. The effect of single genes and haplotypes were investigated and gene-expression analysis was applied for selected genes. Since interaction between risk genes can have a large impact on phenotype, two-way gene-gene interaction analysis was included. RESULTS: We found inherited SNPs in genes critical for inflammatory pathways; TLR9, IL4, TAP2, IL2RA, FCGR2A, TNFA, IL10RB, GALNT12, IL12A and IL1B were significantly associated with disease risk and SELE, IL1RN, TNFA, TAP2, MBL2, IL5, CX3CR1, CHI3L1 and IL12A were, associated with overall survival (OS) in specific diagnostic entities of B-NHL. We discovered noteworthy interactions between DLBCL risk alleles on IL10 and IL4RA and FL risk alleles on IL4RA and IL4. In relation to OS, a highly significant interaction was observed in DLBCL for IL4RA (rs1805010) * IL10 (rs1800890) (HR = 0.11 (0.02-0.50)). Finally, we explored the expression of risk genes from the gene-gene interaction analysis in normal B-cell subtypes showing a different expression of IL4RA, IL10, IL10RB genes supporting a pathogenetic effect of these interactions in the germinal center. CONCLUSIONS: The present findings support the importance of inflammatory genes in B-cell lymphomas. We found association between polymorphic sites in inflammatory response genes and risk as well as outcome in B-NHL and suggest an effect of gene-gene interactions during the stepwise oncogenesis.


Subject(s)
Lymphoma, Follicular/genetics , Lymphoma, Large B-Cell, Diffuse/genetics , Aged , Alleles , Female , Genotype , Haplotypes , Humans , Interleukin-10/genetics , Interleukin-10/metabolism , Interleukin-10 Receptor beta Subunit/genetics , Interleukin-10 Receptor beta Subunit/metabolism , Interleukin-4/genetics , Interleukin-4/metabolism , Interleukin-4 Receptor alpha Subunit/genetics , Interleukin-4 Receptor alpha Subunit/metabolism , Linkage Disequilibrium , Lymphoma, Follicular/etiology , Lymphoma, Follicular/mortality , Lymphoma, Large B-Cell, Diffuse/etiology , Lymphoma, Large B-Cell, Diffuse/mortality , Male , Middle Aged , Odds Ratio , Polymorphism, Single Nucleotide , Proportional Hazards Models , Risk , Survival Analysis , Transcriptome
13.
J Clin Oncol ; 33(12): 1379-88, 2015 Apr 20.
Article in English | MEDLINE | ID: mdl-25800755

ABSTRACT

PURPOSE: Current diagnostic tests for diffuse large B-cell lymphoma use the updated WHO criteria based on biologic, morphologic, and clinical heterogeneity. We propose a refined classification system based on subset-specific B-cell-associated gene signatures (BAGS) in the normal B-cell hierarchy, hypothesizing that it can provide new biologic insight and diagnostic and prognostic value. PATIENTS AND METHODS: We combined fluorescence-activated cell sorting, gene expression profiling, and statistical modeling to generate BAGS for naive, centrocyte, centroblast, memory, and plasmablast B cells from normal human tonsils. The impact of BAGS-assigned subtyping was analyzed using five clinical cohorts (treated with cyclophosphamide, doxorubicin, vincristine, and prednisone [CHOP], n = 270; treated with rituximab plus CHOP [R-CHOP], n = 869) gathered across geographic regions, time eras, and sampling methods. The analysis estimated subtype frequencies and drug-specific resistance and included a prognostic meta-analysis of patients treated with first-line R-CHOP therapy. RESULTS: Similar BAGS subtype frequencies were assigned across 1,139 samples from five different cohorts. Among R-CHOP-treated patients, BAGS assignment was significantly associated with overall survival and progression-free survival within the germinal center B-cell-like subclass; the centrocyte subtype had a superior prognosis compared with the centroblast subtype. In agreement with the observed therapeutic outcome, centrocyte subtypes were estimated as being less resistant than the centroblast subtype to doxorubicin and vincristine. The centroblast subtype had a complex genotype, whereas the centrocyte subtype had high TP53 mutation and insertion/deletion frequencies and expressed LMO2, CD58, and stromal-1-signature and major histocompatibility complex class II-signature genes, which are known to have a positive impact on prognosis. CONCLUSION: Further development of a diagnostic platform using BAGS-assigned subtypes may allow pathogenetic studies to improve disease management.


Subject(s)
B-Lymphocyte Subsets/immunology , B-Lymphocytes/immunology , Lymphoma, Large B-Cell, Diffuse/classification , Lymphoma, Large B-Cell, Diffuse/immunology , B-Lymphocyte Subsets/cytology , B-Lymphocyte Subsets/drug effects , B-Lymphocytes/cytology , B-Lymphocytes/drug effects , Cyclophosphamide/pharmacology , Doxorubicin/pharmacology , Drug Resistance, Neoplasm , Humans , Lymphoma, Large B-Cell, Diffuse/drug therapy , Lymphoma, Large B-Cell, Diffuse/pathology , Phenotype , Prognosis , Proportional Hazards Models , Vincristine/pharmacology
14.
Hematol Oncol ; 33(4): 120-8, 2015 Dec.
Article in English | MEDLINE | ID: mdl-25052551

ABSTRACT

Clinical trials (CTs) are needed to improve the outcome for peripheral T-cell lymphomas (PTCL), and accrual into CTs is one of the main recommendations in international treatment guidelines. The use of risk-adapted strategies has been suggested as a way to optimize treatment outcome in PTCL. The aim of the present study was to evaluate CT eligibility and selected prognostic indices in a population-based PTCL cohort of 481 PTCL patients identified from the Danish Lymphoma Registry in the period 2000-2010. According to five predefined parameters (age, performance status, P-creatinine, P-ALAT and measurable tumour lesion), patients were subdivided into four groups: 'younger fit', 'elderly fit', 'frail' and 'not CT eligible'. International prognostic index (IPI), prognostic index for T-cell lymphoma (PIT) and anaplastic lymphoma kinase (ALK) protein expression were tested at subtype-specific level. Overall, 41% of the patients were considered eligible for interventional CTs implicating curatively intended multiagent chemotherapy, including, if considered appropriate, consolidating stem cell transplantation (SCT), as part of the upfront management strategy. Moreover, 28% was elderly fit and eligible for interventional CT, including those with SCT as part of the trial design. Approximately 7% were defined as 'too frail' for aggressive treatment schedules, whereas 24% were deemed not to be eligible for any CT. Both overall and progression-free survivals were effectively predicted by IPI and PIT (p < 0.001). ALK-positive anaplastic large cell lymphoma patients were significantly younger (median age 40 vs. 62, p < 0.001) and had a better outcome than their ALK-negative counterparts (p < 0.001). However, ALK expression lost its prognostic significance when adjusting for age. In a population-based cohort of adult Caucasian PTCL patients, approximately half were eligible for multiagent chemotherapy with or without consolidating SCT. Both IPI and PIT are useful prognostic indices in all 'primary nodal' PTCL entities. The prognostic value of ALK protein expression in anaplastic large cell lymphoma is significantly downsized when adjusting for age.


Subject(s)
Lymphoma, T-Cell, Peripheral/therapy , Adolescent , Adult , Aged , Aged, 80 and over , Clinical Trials as Topic , Cohort Studies , Disease-Free Survival , Female , Humans , Lymphoma, T-Cell, Peripheral/drug therapy , Lymphoma, T-Cell, Peripheral/pathology , Male , Middle Aged , Prognosis , Registries , Sweden , Treatment Outcome , Young Adult
15.
BMC Immunol ; 15: 3, 2014 Jan 31.
Article in English | MEDLINE | ID: mdl-24483235

ABSTRACT

BACKGROUND: This report describes a method for the generation of global gene expression profiles from low frequent B-cell subsets by using fluorescence-activated cell sorting and RNA amplification. However, some of the differentiating compartments involve a low number of cells and therefore it is important to optimize and validate each step in the procedure. METHODS: Normal lymphoid tissues from blood, tonsils, thymus and bone marrow were immunophenotyped by the 8-colour Euroflow panel using multiparametric flow cytometry. Subsets of B-cells containing cell numbers ranging from 800 to 33,000 and with frequencies varying between 0.1 and 10 percent were sorted, subjected to mRNA purification, amplified by the NuGEN protocol and finally analysed by the Affymetrix platform. RESULTS: Following a step by step strategy, each step in the workflow was validated and the sorting/storage conditions optimized as described in this report. First, an analysis of four cancer cell lines on Affymetrix arrays, using either 100 ng RNA labelled with the Ambion standard protocol or 1 ng RNA amplified and labelled by the NuGEN protocol, revealed a significant correlation of gene expressions (r ≥ 0.9 for all). Comparison of qPCR data in samples with or without amplification for 8 genes showed that a relative difference between six cell lines was preserved (r ≥ 0.9). Second, a comparison of cells sorted into PrepProtect, RNAlater or directly into lysis/binding buffer showed a higher yield of purified mRNA following storage in lysis/binding buffer (p < 0.001). Third, the identity of the B-cell subsets validated by the cluster of differentiation (CD) membrane profile was highly concordant with the transcriptional gene expression (p-values <0.001). Finally, in normal bone marrow and tonsil samples, eight evaluated genes were expressed in accordance with the biology of lymphopoiesis (p-values < 0.001), which enabled the generation of a gene-specific B-cell atlas. CONCLUSION: A description of the implementation and validation of commercially available kits in the laboratory has been examined. This included steps for cell sorting, cell lysis/stabilization, RNA isolation, RNA concentration and amplification for microarray analysis. The workflow described in this report will enable the generation of microarray data from minor sorted B-cell subsets.


Subject(s)
B-Lymphocyte Subsets/metabolism , Gene Expression Profiling/methods , Antigens, CD/metabolism , Flow Cytometry , Humans , Lymphoid Tissue/cytology , Lymphoid Tissue/metabolism , Oligonucleotide Array Sequence Analysis , Organ Specificity/genetics , Real-Time Polymerase Chain Reaction , Reproducibility of Results
16.
Pediatr Transplant ; 18(1): E35-9, 2014 Feb.
Article in English | MEDLINE | ID: mdl-24168387

ABSTRACT

We present a very late onset relapse of PTLD 10 yr after allogeneic HSCT in a patient in third remission for ALL, nine yr after the first episode of PTLD. The recipient was conditioned with fractionated TBI 12 Gy, cyclophosphamide, and horse ATG. The first episode of PTLD with a large retroperitoneal tumor occurred one yr after transplantation; a residual tumor infiltrating spleen and colon was resected one yr later. Due to continual pathological signals in liver and lungs, persistent fever, and an M-component in peripheral blood, a new course of four rituximab doses was given, after which the fever settled, the PET scan normalized, and the M-component disappeared. Without any ongoing immunosuppressive therapy, PTLD relapsed nine yr later with large intra-abdominal lymph node masses causing ureteric obstruction with bilateral hydronephrosis. Pathological features were identical to the primary PTLD tumor: EBV related, of donor origin, positive for CD138 and CD79 alpha, but negative for CD20 and CD19. The transcription factor PAX5 was negative but BOB1 and OCT2 were positive, consistent with plasmablastic lymphoma. The relapse was successfully treated with a combination of low dose chemotherapy and rituximab. Five yr after end of treatment, the girl has moderately reduced renal function but otherwise remains well without evidence of disease.


Subject(s)
Hematopoietic Stem Cell Transplantation , Immunosuppressive Agents/therapeutic use , Lymphoproliferative Disorders/etiology , Lymphoproliferative Disorders/therapy , Precursor Cell Lymphoblastic Leukemia-Lymphoma/therapy , Adolescent , Antibodies, Monoclonal, Murine-Derived/administration & dosage , Antilymphocyte Serum/therapeutic use , Cyclophosphamide/therapeutic use , Female , Humans , Precursor Cell Lymphoblastic Leukemia-Lymphoma/complications , Retroperitoneal Neoplasms/therapy , Rituximab , Transplantation, Homologous
17.
Leuk Lymphoma ; 55(6): 1251-60, 2014 Jun.
Article in English | MEDLINE | ID: mdl-23998255

ABSTRACT

Recent findings have suggested biological classification of B-cell malignancies as exemplified by the "activated B-cell-like" (ABC), the "germinal-center B-cell-like" (GCB) and primary mediastinal B-cell lymphoma (PMBL) subtypes of diffuse large B-cell lymphoma and "recurrent translocation and cyclin D" (TC) classification of multiple myeloma. Biological classification of B-cell derived cancers may be refined by a direct and systematic strategy where identification and characterization of normal B-cell differentiation subsets are used to define the cancer cell of origin phenotype. Here we propose a strategy combining multiparametric flow cytometry, global gene expression profiling and biostatistical modeling to generate B-cell subset specific gene signatures from sorted normal human immature, naive, germinal centrocytes and centroblasts, post-germinal memory B-cells, plasmablasts and plasma cells from available lymphoid tissues including lymph nodes, tonsils, thymus, peripheral blood and bone marrow. This strategy will provide an accurate image of the stage of differentiation, which prospectively can be used to classify any B-cell malignancy and eventually purify tumor cells. This report briefly describes the current models of the normal B-cell subset differentiation in multiple tissues and the pathogenesis of malignancies originating from the normal germinal B-cell hierarchy.


Subject(s)
B-Lymphocytes/metabolism , B-Lymphocytes/pathology , Leukemia, B-Cell/diagnosis , Leukemia, B-Cell/genetics , Lymphoma, B-Cell/diagnosis , Lymphoma, B-Cell/genetics , Transcriptome , Flow Cytometry , Gene Expression Profiling , Humans , Immunophenotyping , Lymphoma, Large B-Cell, Diffuse/diagnosis , Lymphoma, Large B-Cell, Diffuse/genetics , Microarray Analysis/methods , Models, Statistical , Multiple Myeloma/diagnosis , Multiple Myeloma/genetics
18.
J Occup Environ Med ; 55(8): 924-31, 2013 Aug.
Article in English | MEDLINE | ID: mdl-23887697

ABSTRACT

OBJECTIVE: To evaluate the potential association between occupational exposure to chlorinated and petroleum solvents and mycosis fungoides (MF). METHODS: A questionnaire on lifetime job history was administered to 100 patients diagnosed with MF and 2846 controls. Odds ratios (ORs) were calculated as the measure of the association between exposure to each specific solvent and MF. RESULTS: In the total sample and in men, cases and controls did not differ in relation to exposure to any of the solvents studied. In women, an association with MF was seen for the highest level of estimated exposure to perchloroethylene (OR = 11.38; 95% confidence interval: 1.04 to 124.85) and for exposure less than the median to kerosene/fuel/gasoil (OR = 8.53; 95% confidence interval: 1.11 to 65.62). CONCLUSIONS: These results do not provide conclusive evidence that exposure to solvents may increase risk of MF because they were not found in men.


Subject(s)
Hydrocarbons, Chlorinated/adverse effects , Mycosis Fungoides/chemically induced , Occupational Exposure/adverse effects , Petroleum/adverse effects , Skin Neoplasms/chemically induced , Adult , Aged , Case-Control Studies , Female , Fuel Oils/adverse effects , Humans , Kerosene/adverse effects , Male , Middle Aged , Odds Ratio , Sex Factors , Solvents/adverse effects , Surveys and Questionnaires , Tetrachloroethylene/adverse effects
19.
Am J Clin Pathol ; 136(6): 960-9, 2011 Dec.
Article in English | MEDLINE | ID: mdl-22095383

ABSTRACT

The purpose of this study was to establish a procedure capable of isolating distinct B-cell subpopulations from human tonsils as a basis for subsequent molecular analyses. Overall, 5 distinct B-cell subpopulations were purified from fresh tonsils based on their fluorescence surface marker expression: naive B cells, centroblasts, centrocytes, memory B cells, and plasmablasts. The immunophenotypic identity of the subpopulations was verified by quantitative real-time reverse transcriptase-polymerase chain reaction using the proliferation marker MKI-67 and 6 B-cell-associated differentiation markers (BACH2, BCL6, PAX5, IRF4, PRDM1, and XBP1). Furthermore, within the centroblast compartment, large and small centroblasts could be distinguished and large centroblasts were shown to proliferate with a morphologic appearance of a "centroblast"-like cell but with lower gene expression of the germinal center markers BCL6 and BACH2 vs small centroblasts. This study has established a detailed and fast procedure for simultaneous sorting of up to 5 distinct maturation-associated B-cell subpopulations from human tonsils.


Subject(s)
B-Lymphocytes/cytology , Cell Separation/methods , Flow Cytometry/methods , Palatine Tonsil/cytology , Adolescent , Adult , Basic-Leucine Zipper Transcription Factors/analysis , Cell Differentiation , Child , Female , Germinal Center/cytology , Humans , Middle Aged , Proto-Oncogene Proteins/analysis , Repressor Proteins/analysis
20.
Ugeskr Laeger ; 173(34): 2047-8, 2011 Aug 22.
Article in Danish | MEDLINE | ID: mdl-21867658

ABSTRACT

A 28 month-old boy was hospitalized with pallor and weight stagnation. He had macrocytic anaemia and pancytopenia due to cobalamin deficiency and a rare homozygous mutation in the intrinsic factor gene. His sister showed similar symptoms at the age of 15 months. The heterozygous father had no symptoms, but did have a low cobalamin level. Gastroscopy with biopsies showed no pathology. All were given monthly cyanocobalamin injections which, however, caused leg cramps. Replacement with monthly hydroxocobalamin was successful.


Subject(s)
Intrinsic Factor/genetics , Vitamin B 12 Deficiency/genetics , Child, Preschool , Female , Growth , Homozygote , Humans , Hydroxocobalamin/administration & dosage , Infant , Male , Mutation , Vitamin B 12 Deficiency/drug therapy , Vitamin B Complex/administration & dosage
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