Subjects with chronic lymphocytic leukaemia-like B-cell clones with stereotyped B-cell receptors frequently show MDS-associated phenotypes on myeloid cells

An increasing body of evidence suggests the potential occurrence of antigen encounter by the cell of origin in chronic lymphocytic leukaemia (CLL) and CLL-like monoclonal B-cell lymphocytosis (MBL). However, the scenario in which this event might occur remains unknown. In order to gain insight into this scenario we investigated the molecular, cytogenetic and haematological features of 223 CLL-like (n = 84) and CLL (n = 139) clones with stereotyped (n = 32) versus non-stereotyped (n = 191) immunoglobulin heavy chain variable region (IGHV) amino acid sequences. Overall, stereotyped CLL-like MBL and CLL clones showed a unique IGHV profile, associated with higher IGHV1 and lower IGHV3 gene family usage (P = 0·03), longer IGHV complementary determining region 3 (HCDR3) sequences (P = 0·007) and unmutated IGHV (P 0·05), it did show a significant association with the presence of myelodysplastic syndrome (MDS)-associated immunophenotypes on peripheral blood neutrophils and/or monocytes (P = 0·01). Altogether our results point to the potential involvement of different selection forces in the expansion of stereotyped vs. non-stereotyped CLL and CLL-like MBL clones, the former being potentially favoured by an underlying altered haematopoiesis.Funded by: Red Temática de Investigación Cooperativa en Cáncer (RTICC) of Instituto de Salud Carlos III – FONDOS FEDER. Grant Numbers: RD06/0020/0035, RD12/0036/0048, FIS PI06/0824-FEDER, PS09/02430-FEDER; FIS PI12/00905-FEDER Fondo de Investigación Sanitaria of Instituto de Salud Carlos III. Grant Numbers: GRS206/A/08, SAN/1778/2009; Gerencia Regional de Salud, Consejería de Educación and Consejería de Sanidad of Castilla y León. Grant Numbers: FS/1-2010, FS/19-2013; Fundación Memoria D. Samuel Solórzano, Universidad de Salamanca; Fundação para a Ciência e Tecnologia of Portugal. Grant Number: SFRH/BD/31609/2006; Fundación Científica de la Asociación Española contra el Cáncer. Grant Number: AECC-2008.Peer Reviewe

High-resolution copy number analysis of paired normal-tumor samples from diffuse large B cell lymphoma

Copy number analysis can be useful for assessing prognosis in diffuse large B cell lymphoma (DLBCL). We analyzed copy number data from tumor samples of 60 patients diagnosed with DLBCL de novo and their matched normal samples. We detected 63 recurrent copy number alterations (CNAs), including 33 gains, 30 losses, and nine recurrent acquired copy number neutral loss of heterozygosity (CNN-LOH). Interestingly, 20 % of cases acquired CNN-LOH of 6p21 locus, which involves the HLA region. In normal cells, there were no CNAs but we observed CNN-LOH involving some key lymphoma regions such as 6p21 and 9p24.1 (5 %) and 17p13.1 (2.5 %) in DLBCL patients. Furthermore, a model with some specific CNA was able to predict the subtype of DLBCL, 1p36.32 and 10q23.31 losses being restricted to germinal center B cell-like (GCB) DLBCL. In contrast, 8p23.3 losses and 11q24.3 gains were strongly associated with the non-GCB subtype. A poor prognosis was associated with biallelic inactivation of TP53 or 18p11.32 losses, while prognosis was better in cases carrying 11q24.3 gains. In summary, CNA abnormalities identify specific DLBCL groups, and we describe CNN-LOH in germline cells from DLBCL patients that are associated with genes that probably play a key role in DLBCL development.This work was supported by research funding from the Health Council of Castilla y León (GRS265/A/08), the Health Research Program (PS09/01382), and the Red Temática de Investigación Cooperativa en Cáncer (RTICC) grant RD12/0036 (groups 0069, 0029, 0036, 0058, and 0060) included in the National Plan I+D+I supported by the Instituto Carlos III and the Fondo Europeo de Desarrollo Regional (FEDER), the Spanish Ministry of Economy and Competitiveness, and the European Regional Development Fund (ERDF) 'Una manera de hacer Europa' (Innocampus; CEI-2010-1-0010). ES was supported by CM10/00078-Río Hortega, an ISCIII contract, FEHH grant 2013–2014 and JR14/00025-Juan Rodés, an ISCIII contract. IS was supported by the Subprograma Juan de la Cierva (JCI-2011-10232) and a Miguel Servet contract (CP13/00159).Peer Reviewe

Circulating clonotypic B cells in multiple myeloma and monoclonal gammopathy of undetermined significance

This is an open-access paper.The B-cell compartment in which multiple myeloma stem cells reside remains unclear. We investigated the potential presence of mature, surface-membrane immunoglobulin-positive B lymphocytes clonally related to the tumor bone marrow plasma cells among different subsets of peripheral blood B cells from ten patients (7 with multiple myeloma and 3 with monoclonal gammopathies of undetermined significance). The presence of clonotypic immunoglobulin heavy chain gene rearrangements was determined in multiple highly-purified fractions of peripheral blood B-lymphocytes including surface-membrane IgM+ CD27- naïve B-lymphocytes, plus surface-membrane IgG+, IgA+ and IgM+ memory CD27+ B cells, and normal circulating plasma cells, in addition to (mono)clonal plasma cells, by a highly-specific and sensitive allele-specific oligonucleotide polymerase chain reaction directed to the CDR3 sequence of the rearranged IGH gene of tumor plasma cells from individual patients. Our results showed systematic absence of clonotypic rearrangements in all the different B-cell subsets analyzed, including M-compo-nent isotype-matched memory B-lymphocytes, at frequencies <0.03 cells/mL (range: 0.0003-0.08 cells/mL); the only exception were the myeloma plasma cells detected and purified from the peripheral blood of four of the seven myeloma patients. These results indicate that circulating B cells from patients with multiple myeloma and monoclonal gammopathies of undetermined significance are usually devoid of clonotypic B cells while the presence of immunophenotypically aberrant myeloma plasma cells in peripheral blood of myeloma patients is a relatively frequent finding.This work was supported by grants from European Union FP6 STREP MSCNet (N. E06005FF), Cooperative Research Thematic Network on Cancer (RTICs; RTICC RD06/0020/0035-FEDER, RD06/0020/0006, RD12/0036/0048, RD12/0036/0069 and G03/136), Instituto de Salud Carlos III/Subdirección General de Investigación Sanitaria Ministerio de Sanidad y Consumo (FIS: PI060339; 02/0905; 01/0089/01-02;PS09/01897, and PI06/0824-FEDER), Asociacion Española Contra el Cancer AECC (GCB120981SAN) and Gerencia Regional de Salud de Castilla y León; Ayuda de Excelencia de Castilla y León, Consejería de Educación (EDU/894/2009, GR37) Junta de Castilla y León, Valladolid, Spain. LST received a CAPES/Ministério da Educação scholarship from the Brazilian Government.Peer Reviewe

Anti-trbc1 antibody-based flow cytometric detection of t-cell clonality: Standardization of sample preparation and diagnostic implementation

© 2021 by the authors.A single antibody (anti-TRBC1; JOVI-1 antibody clone) against one of the two mutually exclusive T-cell receptor β-chain constant domains was identified as a potentially useful flow-cytometry (FCM) marker to assess Tαβ-cell clonality. We optimized the TRBC1-FCM approach for detecting clonal Tαβ-cells and validated the method in 211 normal, reactive and pathological samples. TRBC1 labeling significantly improved in the presence of CD3. Purified TRBC1+ and TRBC1− monoclonal and polyclonal Tαβ-cells rearranged TRBJ1 in 44/47 (94%) and TRBJ1+TRBJ2 in 48 of 48 (100%) populations, respectively, which confirmed the high specificity of this assay. Additionally, TRBC1+/TRBC1− ratios within different Tαβ-cell subsets are provided as reference for polyclonal cells, among which a bimodal pattern of TRBC1-expression profile was found for all TCRVβ families, whereas highly-variable TRBC1+/TRBC1− ratios were observed in more mature vs. naïve Tαβ-cell subsets (vs. total T-cells). In 112/117 (96%) samples containing clonal Tαβ-cells in which the approach was validated, monotypic expression of TRBC1 was confirmed. Dilutional experiments showed a level of detection for detecting clonal Tαβ-cells of ≤10−4 in seven out of eight pathological samples. These results support implementation of the optimized TRBC1-FCM approach as a fast, specific and accurate method for assessing T-cell clonality in diagnostic-FCM panels, and for minimal (residual) disease detection in mature Tαβ+ leukemia/lymphoma patients.This work was supported by the CB16/12/00400 (CIBERONC) and PI20-01346 grants, from the Instituto de Salud Carlos III, Ministerio de Ciencia e Innovación (Madrid, Spain) and FONDOS FEDER, and by the EuroFlow Foundation (Leiden, The Netherlands). N. Muñoz-García is supported by a pre-doctoral grant (Ref. IBPredoc17/00012) from IBSAL (Salamanca, Spain). M. Lima, N. Villamor, A.W. Langerak, J.J.M. van Dongen, A. Orfao, and J. Almeida are members of the EuroFlow Consortiu

Allele and haplotype frequencies of HLA-A, -B, -C, -DRB1, -DQB1 and -DQA1 in Castile and Leon region from North West of Spain

HLA studies have been used to determine the admixture of different populations within the Iberian Peninsula including neighbouring regions with shared origins, such as Portugal and Castile and Leon. These studies certainly can be used to study human migration that could establish populations currently settled according to genetic distant analysis based on the HLA diversity and language variety.This work was supported by the “Gerencia Regional de Salud de Castilla y Leon” (GRS 2080/A/19, 2019) and (GRS COVID 70/A/20, 2020)

Genetic complexity impacts the clinical outcome of follicular lymphoma patients

© The Author(s) 2021.Follicular lymphoma (FL) is the second most common non-Hodgkin lymphoma (NHL, 20–30%) after diffuse large B-cell lymphoma (DLBCL). Despite the introduction of rituximab and the high response rate to first-line treatment, approximately 20% of the FL patients relapse or progress within 2 years of receiving first-line therapy. Therefore, the major challenge is finding biomarkers that identify high-risk patients at diagnosis.This work was partially supported by the Instituto de Salud Carlos III (ISCIII), Spanish Ministry of Economy and Competitiveness PI15/01393, PI18/00410, CIBERONC-CB16/12/00233, and “Una manera de hacer Europa” (Innocampus; CEI-2010-1-0010)”, the Education Council or Health Council of the Junta de Castilla y León (CAS102P17, GRS 1180/A/15), Spanish Association Against Cancer (AECC; PROYE18020BEA), and Gilead Sciences (GLD17/00334). CJ, MES, and AMe are supported by the ISCII (CD19/00030, CPII18/00028, and FI19/00320). MGA, IPC, and CJ were supported by the Spanish Society of Hematology Foundation (FEHH). All Spanish funding is co-sponsored by the European Union FEDER program

Molecular profiling of immunoglobulin heavy-chain gene rearrangements unveils new potential prognostic markers for multiple myeloma patients

Multiple myeloma is a heterogeneous disease whose pathogenesis has not been completely elucidated. Although B-cell receptors play a crucial role in myeloma pathogenesis, the impact of clonal immunoglobulin heavy-chain features in the outcome has not been extensively explored. Here we present the characterization of complete heavy-chain gene rearrangements in 413 myeloma patients treated in Spanish trials, including 113 patients characterized by next-generation sequencing. Compared to the normal B-cell repertoire, gene selection was biased in myeloma, with significant overrepresentation of IGHV3, IGHD2 and IGHD3, as well as IGHJ4 gene groups. Hypermutation was high in our patients (median: 8.8%). Interestingly, regarding patients who are not candidates for transplantation, a high hypermutation rate (≥7%) and the use of IGHD2 and IGHD3 groups were associated with improved prognostic features and longer survival rates in the univariate analyses. Multivariate analysis revealed prolonged progression-free survival rates for patients using IGHD2/IGHD3 groups (HR: 0.552, 95% CI: 0.361−0.845, p = 0.006), as well as prolonged overall survival rates for patients with hypermutation ≥7% (HR: 0.291, 95% CI: 0.137−0.618, p = 0.001). Our results provide new insights into the molecular characterization of multiple myeloma, highlighting the need to evaluate some of these clonal rearrangement characteristics as new potential prognostic markers

Bases psicopedagógicas para la estimulación temprana del niño sordo profundo

Previo análisis de los procesos cognitivos implicados en la comunicación y de las dimensiones del proceso comunicativo, pretende un análisis descriptivo de las estrategias y sistemas de comunicación que se establecen entre el niño sordo e interlocutores oyentes (su madre o un extraño). Así como sus deficits comunicativos como paso previo para la busqueda de formas alternativas de educación de sordos. Específicamente, el trabajo propone una serie de hipótesis en dos niveles: evolutivo y referido al tipo de interlocutor con el que se comunica el niño. Cuatro sujetos elegidos según los criterios: sordera profunda (-70 db), hijo de padres oyentes, ser sordo prelocutivo, no tener otros tastornos asociados y estar sometido a estimulación con programas orales. Descripción: suj. 1, niño de 2 años y 5 meses; suj. 2, niña de 3 años y 3 meses; suj. 3, niña de 3 años y 6 meses y suj. 4, niño de 3 años y 10 meses. Seguimiento de los sujetos durante 9 meses. 5 sesiones de grabación de una hora con una periodicidad aproximada de 3 meses. Sesión se registran interacciones madre-niño y extraño-niño en situaciones de juego libre y de comentario de cuentos o láminas. Efectúan entrevistas con la madre y evaluaciones cognitivas de los niños. Variables consideradas: V. Independientes: interlocutor del niño (madre o extraño), tiempo transcurrido desde la primera a la última sesión. V. Dependientes: funciones comunicativas mostradas por el niño (reguladora, declarativa, interrogadora, fáctica, rituales de saludación, expresiva, imitativa, autoreguladora, ininterpretables y otras), modalidades comunicativas o de expresión (acción-actividad, producción gestual, producción oral, combinaciones de producciones oral-signada y sintaxis oral-gestual). Se codifican 10 minutos (210 vueltas de magnetoscopio) considerados como de mayor riqueza comunicativa según un protocolo diseñado ad hoc. Observa en todos los sujetos un predominio de la función fáctica, debido, tal vez, a la carencia de un sistema de comunicación. Cierta relación entre la función fáctica y la reguladora. Falta de declarativos interrogativos debida a la carencia de medios lingüísticos. Esto pone de relieve la relación entre forma y función. El nivel de declarativos de estados internos, muy bajo, puede deberse al modelo de intervención, lo que origina pocos declarativos de atribución y de interrogación. En cuanto a la modalidad, observa una gran variabilidad. En general poca producción simbólica. Lo más parecido suelen ser asociaciones de gesto deíctico y gesto simbólico. Por último, el efecto del interlocutor es variable y desigual. En términos generales no hay diferencias del estilo comunicativo del niño hacia diferentes interlocutores. Es necesario dotar al sordo de un sistema comunicativo estructurado que le permita aprehender la realidad y expresarse. La intervención oral debe ir acompañada con el aprendizaje de lenguajes de signos o bimodales para facilitar el desarrollo cognitivo, lingüístico y social del niño. Concluye señalando la falta de investigaciones en muchos de los temas tratados.Castilla y LeónBiblioteca de Educación del Ministerio de Educación, Cultura y Deporte; Calle San Agustín, 5 - 3 Planta; 28014 Madrid; Tel. +34917748000; Fax +34917748026; [email protected]

The relevance of preferentially expressed antigen of melanoma (PRAME) as a marker of disease activity and prognosis in acute promyelocytic leukemia

[Background]: The gene for preferentially expressed antigen of melanoma (PRAME) has been shown to be over-expressed in acute promyelocytic leukemia, but its actual incidence and clinical impact are still unknown. [Design and Methods]: We studied PRAME expression at diagnosis using real-time quantitative polymerase chain reaction in 125 patients with acute promyelocytic leukemia enrolled in the Spanish PETHEMA-96 (n=45) and PETHEMA-99 (n=80) clinical trials. In addition, PRAME expression was evaluated as a marker of disease activity in 225 follow-up samples from 67 patients with acute promyelocytic leukemia. [Results]: At diagnosis, PRAME expression in patients with acute promyelocytic leukemia was significantly higher (p100-fold PRAME expression (86% vs. 74%; p=0.03), and this cut-off established two sub-groups with different relapse-free survival rates among patients with a white cell count 109/L, although differences were not statistically significant. In multivariate analysis, white cell count >109/L (p90% (p=0.001), and PRAME expression 10-fold increase in PRAME expression levels. [Conclusions]: Low PRAME expression defines a subgroup of patients with acute promyelocytic leukemia with a short relapse-free survival. This marker could be useful as a secondary marker for monitoring patients with acute promyelocytic leukemia. ©2008 Ferrata Storti Foundation.This work was partially supported by grants PI061351 from the Spanish “Fondo de Investigaciones Sanitarias de la Seguridad Social”, and 89/A/06 from the “Gerencia Regional de Salud, Junta Castilla y León”, CIC, IBMCC (USAL-CSIC), Spain and by funding from the CR-USA Foundation-Spanish National Research Council (CSIC) Cooperative Agreement.Peer Reviewe