Implicación de la expresión condicional de HGAL en modelos murinos de linfoma de células grandes B difuso

[ES] Los linfomas de células grandes B difusos (LCGBD) son tumores clínica y genéticamente heterogéneos. La desregulación de diversos procesos biológicos específicos de las células B, como la señalización del receptor de células B (BCR) y la regulación de la motilidad, contribuyen a la linfomagénesis. La proteína HGAL (del inglés human germinal associated lymphoma) es una proteína adaptadora específica de las células B que controla su motilidad y la señalización del BCR. En las células B normales, el gen HGAL se expresa en el centro germinal y rápidamente es reprimido. La mayoría de los LCGBD, principalmente los del centro germinal (GCB), aunque en menor medida los de tipo activado (ABC), expresan HGAL. Para investigar las consecuencias de la expresión constitutiva de HGAL in vivo, generamos ratones que expresaban condicionalmente HGAL humano en diferentes etapas del desarrollo hematopoyético usando 3 enfoques restringidos mediados por la recombinasa Cre para iniciar la expresión de HGAL en las células madre hematopoyéticas, las células pro-B o las células B del centro germinal. Tras la estimulación inmunológica, se observaron centros germinales más grandes en ratones en los que se inició la expresión de HGAL en células B del centro germinal, lo que puso de evidencia su implicación en el proceso tumorogénico: en el examen macroscópico estos animales presentaban una marcada esplenomegalia causada por grandes tumores, independientemente del promotor que controla la expresión de HGAL. Los tres modelos de ratón desarrollaron LCGBD con una frecuencia del 12% al 30% a partir de los 13 meses, lo que condicionó una supervivencia más corta: 21 meses, 20 meses y 16 meses de supervivencia específica media las cohortes de Rosa26HGAL/Aid-Cre, Rosa26HGAL/Mb1-Cre y Rosa26HGAL/Sca1-Cre, respectivamente. Los estudios inmunohistoquímicos revelaron que todos los linfomas eran del tipo BGC, con infiltración nodular difusa de los órganos afectados por grandes células linfoides pleomórficas que expresaban B220, PAX5, HGAL, PNA e IRF4. Además, en estos modelos murinos la sobreexpresión condicionada de HGAL dio lugar a una evidente afectación extraganglionar con afectación del hígado y de los ganglios linfáticos en los ratones Rosa26HGAL/Sca1-Cre, del riñón y del páncreas en los ratones Rosa26HGAL/Mb1-Cre y del riñón y del hígado en los ratones Rosa26HGAL/Aid- Cre. El estudio de microarrays puso de manifiesto la presencia de una marcada expresión diferencial entre las células con linfoma y los esplenocitos B normales en la que destacaba una sobreexpresión del regulador de la señalización de proteína G1 (RGS1), relacionado con la regulación de la migración celular y de RAB10. El análisis de los genes expresados diferencialmente entre las células esplénicas B de los tres modelos murinos en comparación con ratones control de la misma camada reveló un enriquecimiento en genes comunes a los tres modelos entre los que destacaba una sobreexpresión en los linfomas de la señalización de mTORC1, la respuesta de interferón, las dianas E2F y la señalización de la vía IL-6/JAK/STAT3. La secuenciación del exoma completo de 8 muestras de tumores murinos reveló la presencia de 124 mutaciones sin-sentido o de sentido erróneo en 110 genes, con una media de 15 por tumor (número similar a la mediana de 17 alteraciones genéticas por tumor encontradas en los LCGBD humanos). Algunos de los genes mutados ya habían sido descritos previamente en tumores humanos del tipo LCGBD, con 18 mutaciones detectadas previamente, de forma recurrente, en un 0.7% de los LCGBD humanos : PIM1, GNA13, FAS, PDF4DIP, NFKBIA y PTPN6. Posteriormente, la secuenciación del ARN mostró que todos los tumores murinos analizados expresaban BCL6. Es posible que la ausencia de expresión de BCL6 en algunos tumores con expresión de HGAL pudiera deberse a la estimulación mejorada del BCR con HGAL en estos tumores, tal y como se ha descrito en algunos estudios que muestran una disminución en la expresión de proteína proteica de BCL6. El análisis del reordenamiento de las inmunoglobulinas confirmó que todos los tumores de células B eran clonales y tenían mutaciones somáticas independientemente del impulsor del promotor encargado de la expresión de HGAL. Nuestros resultados demuestran que la expresión forzada constitutiva de HGAL conduce al desarrollo de LCGBD de tipo BCG. [EN] Diffuse large B-cell lymphomas (DLBCL) are clinically and genetically heterogeneous tumors. Dysregulation of various biological processes specific to B cells, such as B cell receptor (BCR) signaling and regulation of motility, contribute to lymphomagenesis. Human germinal associated lymphoma (HGAL) protein is a specific adapter protein of B cells that controls their motility and BCR signaling. In normal B cells, the HGAL gene is expressed in the germinal center and is rapidly repressed. Most DLBCL, mainly those of the germinal center (GCB), although to a lesser extent those of the activated type (ABC), express HGAL. To investigate the consequences of constitutive expression of HGAL in vivo, we generated mice conditionally expressing human HGAL at different stages of hematopoietic development using 3-restricted Cre recombinase-mediated approaches to initiate HGAL expression in hematopoietic stem cells, pro-B or B cells of the germinal center. After immunological stimulation, larger germinal centers were observed in mice in which the expression of HGAL was initiated in B cells of the germinal center, which evidenced its involvement in the tumorigenic process: in the macroscopic examination these animals presented a marked splenomegaly caused by large tumors, regardless of the promoter that controls HGAL expression. The three mouse models developed DLBCL with a frequency of 12% to 30% from 13 months, leading to a shorter survival: 21 months, 20 months and 16 months of median specific survival in the Rosa26HGAL/Aid-Cre, Rosa26HGAL/Mb1-Cre and Rosa26HGAL/Sca1-Cre cohorts, respectively. Immunohistochemical studies revealed that all the lymphomas were of the SLNB type, with diffuse nodular infiltration of the affected organs by large pleomorphic lymphoid cells expressing B220, PAX5, ALGH, PNA, and IRF4. In addition, in these mouse models conditioned overexpression of HGAL resulted in evident extranodal involvement with involvement of the liver and lymph nodes in Rosa26HGAL/Sca1-Cre mice, kidney and pancreas in Rosa26HGAL/Mb1-Cre mice and kidney and liver in Rosa26HGAL/Aid-Cre mice. The microarray study revealed the presence of a marked differential expression between cells with lymphoma and normal B splenocytes, in which an overexpression of the regulator of signaling protein G1 (RGS1), related to the regulation of cell migration and RAB10. Analysis of differentially expressed genes between spleen B cells of the three mouse models compared with littermate control mice revealed an enrichment in genes common to all three models notable among them was an overexpression of B signaling in lymphomas of mTORC1, interferon response, E2F targets, and IL-6/JAK/STAT3 signaling pathway. Whole-exome sequencing of 8 murine tumor samples revealed the presence of 124 nonsense or missense mutations in 110 genes, with a mean of 15 per tumor (like the median of 17 genetic alterations per tumor found in the human DLBCL). Some of the mutated genes had already been previously described in human LCGBD-type tumors, with 18 mutations previously detected, recurrently, in 0.7% of human DLBCL: PIM1, GNA13, FAS, PDF4DIP, NFKBIA, and PTPN6. Subsequently, RNA sequencing showed that all murine tumors analyzed expressed BCL6. It is possible that the absence of BCL6 expression in some tumors with HGAL expression could be due to the enhanced stimulation of the BCR with HGAL in these tumors, as has been described in some studies showing a decrease in the expression of HGAL protein. Immunoglobulin rearrangement analysis confirmed that all B-cell tumors were clonal and had somatic mutations, regardless of the promoter drive responsible for HGAL expression. Our results demonstrate that constitutive forced expression of HGAL leads to the development of GCB-DLBCL. Our results demonstrate that constitutive forced expression of HGAL leads to the development of DLBCL

Conditional expression of HGAL leads to the development of diffuse large B-cell lymphoma in mice

Diffuse large B-cell lymphomas (DLBCLs) are clinically and genetically heterogeneous tumors. Deregulation of diverse biological processes specific to B cells, such as B-cell receptor (BCR) signaling and motility regulation, contribute to lymphomagenesis. Human germinal center associated lymphoma (HGAL) is a B-cell–specific adaptor protein controlling BCR signaling and B lymphocyte motility. In normal B cells, it is expressed in germinal center (GC) B lymphocytes and promptly downregulated upon further differentiation. The majority of DLBCL tumors, primarily GC B-cell types, but also activated types, express HGAL. To investigate the consequences of constitutive expression of HGAL in vivo, we generated mice that conditionally express human HGAL at different stages of hematopoietic development using 3 restricted Cre-mediated approaches to initiate expression of HGAL in hematopoietic stem cells, pro-B cells, or GC B cells. Following immune stimulation, we observed larger GCs in mice in which HGAL expression was initiated in GC B cells. All 3 mouse strains developed DLBCL at a frequency of 12% to 30% starting at age 13 months, leading to shorter survival. Immunohistochemical studies showed that all analyzed tumors were of the GC B-cell type. Exon sequencing revealed mutations reported in human DLBCL. Our data demonstrate that constitutive enforced expression of HGAL leads to DLBCL development

The Second Oncogenic Hit Determines the Cell Fate of ETV6-RUNX1 Positive Leukemia

© 2021 Rodríguez-Hernández, Casado-García, Isidro-Hernández, Picard, Raboso-Gallego, Alemán-Arteaga, Orfao, Blanco, Riesco, Prieto-Matos, García Criado, García Cenador, Hock, Enver, Sanchez-Garcia and Vicente-Dueñas.ETV6-RUNX1 is almost exclusively associated with childhood B-cell acute lymphoblastic leukemia (B-ALL), but the consequences of ETV6-RUNX1 expression on cell lineage decisions during B-cell leukemogenesis are completely unknown. Clinically silent ETV6-RUNX1 preleukemic clones are frequently found in neonatal cord blood, but few carriers develop B-ALL as a result of secondary genetic alterations. The understanding of the mechanisms underlying the first transforming steps could greatly advance the development of non-toxic prophylactic interventions. Using genetic lineage tracing, we examined the capacity of ETV6-RUNX1 to instruct a malignant phenotype in the hematopoietic lineage by cell-specific Cre-mediated activation of ETV6-RUNX1 from the endogenous Etv6 gene locus. Here we show that, while ETV6-RUNX1 has the propensity to trigger both T- and B-lymphoid malignancies, it is the second hit that determines tumor cell identity. To instigate leukemia, both oncogenic hits must place early in the development of hematopoietic/precursor cells, not in already committed B-cells. Depending on the nature of the second hit, the resulting B-ALLs presented distinct entities that were clearly separable based on their gene expression profiles. Our findings give a novel mechanistic insight into the early steps of ETV6-RUNX1+ B-ALL development and might have major implications for the potential development of ETV6-RUNX1+ B-ALL prevention strategies.Research in CV-D group has been funded by Instituto de Salud Carlos III through the project “PI17/00167” and by a “Miguel Servet Grant” (CPII19/00024—AES 2017–2020), co-funded by European Regional Development Fund/European Social Fund (“A way to make Europe”/“Investing in your future”). Research in the IS-G group is partially supported by FEDER and SAF2015-64420-R MINECO/FEDER, UE, RTI2018-093314-B-I00 MCIU/AEI/FEDER, UE, and by Junta de Castilla y León (UIC-017, CSI001U16, CSI234P18, and CSI144P20). The IS-G lab is a member of the EuroSyStem and the DECIDE Network funded by the European Union under the FP7 program. CV-D and IS-G have been supported by the German Federal Office for Radiation Protection (BfS)–Germany (FKZ: 3618S32274). IS-G has been supported by the Fundacion Unoentrecienmil (CUNINA project). HH was supported by a Hyundai Hope on Wheels scholar grant. GR-H was supported by FSE-Conserjería de Educación de la Junta de Castilla y León (CSI001-15). AC-G and MI-H are supported by FSE-Conserjería de Educación de la Junta de Castilla y León 2019 and 2020 (ESF—European Social Fund) fellowship, respectively (REF. CSI067-18 and CSI021-19). JR-G was supported by a scholarship from the University of Salamanca, co-financed by Banco Santander and ESF. SA-A was supported by RTI2018-093314-B-I00 MCIU/AEI/FEDER fellowship (PRE2019-088887)

The Second Oncogenic Hit Determines the Cell Fate of ETV6-RUNX1 Positive Leukemia

ETV6-RUNX1 is almost exclusively associated with childhood B-cell acute lymphoblastic leukemia (B-ALL), but the consequences of ETV6-RUNX1 expression on cell lineage decisions during B-cell leukemogenesis are completely unknown. Clinically silent ETV6-RUNX1 preleukemic clones are frequently found in neonatal cord blood, but few carriers develop B-ALL as a result of secondary genetic alterations. The understanding of the mechanisms underlying the first transforming steps could greatly advance the development of non-toxic prophylactic interventions. Using genetic lineage tracing, we examined the capacity of ETV6-RUNX1 to instruct a malignant phenotype in the hematopoietic lineage by cell-specific Cre-mediated activation of ETV6-RUNX1 from the endogenous Etv6 gene locus. Here we show that, while ETV6-RUNX1 has the propensity to trigger both T- and B-lymphoid malignancies, it is the second hit that determines tumor cell identity. To instigate leukemia, both oncogenic hits must place early in the development of hematopoietic/precursor cells, not in already committed B-cells. Depending on the nature of the second hit, the resulting B-ALLs presented distinct entities that were clearly separable based on their gene expression profiles. Our findings give a novel mechanistic insight into the early steps of ETV6-RUNX1+ B-ALL development and might have major implications for the potential development of ETV6-RUNX1+ B-ALL prevention strategies

Transient inhibition of the JAK/STAT pathway prevents B-ALL development in genetically predisposed mice

Preventing development of childhood B-cell acute lymphoblastic leukemia (B-ALL), a disease with devastating effects, is a longstanding and unsolved challenge. Heterozygous germline alterations in the PAX5 gene can lead to B-ALL upon accumulation of secondary mutations affecting the JAK/STAT signaling pathway. Preclinical studies have shown that this malignant transformation occurs only under immune stress such as exposure to infectious pathogens. Here we show in Pax5+/− mice that transient, early-life administration of clinically relevant doses of ruxolitinib, a JAK1/2 inhibitor, significantly mitigates the risk of B-ALL following exposure to infection; 1 of 29 animals treated with ruxolitinib developed B-ALL versus 8 of 34 untreated mice. Ruxolitinib treatment preferentially targeted Pax5+/− versus wild-type B-cell progenitors and exerted unique effects on the Pax5+/− B-cell progenitor transcriptional program. These findings provide the first in vivo evidence for a potential strategy to prevent B-ALL development.C. Cobaleda and C. Vicente-Dueñas labs are members of the EU COST Action LEGEND (CA16223). Research in C. Vicente-Dueñas group has been funded by Instituto de Salud Carlos III through the project " PI17/00167 and by a “Miguel Servet Grant” [CPII19/00024 - AES 2017-2020; co-funded by European Regional Development Fund (ERDF)/European Social Fund (ESF) "A way to make Europe"/"Investing in your future"]. J.J. Yang and K.E. Nichols receive funding from the American Lebanese Syrian Associated Charities (ALSAC) and R01CA241452 from the NCI. Research in ISG group is partially supported by FEDER and by SAF2015-64420-R MINECO/FEDER, UE, RTI2018-093314-B-I00 MCIU/AEI/FEDER, UE, 9659122185-122185-4-21 MCIU/AEI/FEDER, UE, by Junta de Castilla y León (UIC-017, CSI001U16, CSI234P18, and CSI144P20). M. Ramírez-Orellana and I. Sánchez-García have been supported by the Fundacion Unoentrecienmil (CUNINA project). C. Cobaleda, M. Ramírez-Orellana, and I. Sánchez-García have been supported by the Fundación Científica de la Asociación Española contra el Cáncer (PRYCO211305SANC). A. Casado-García (CSI067-18) and M. Isidro-Hernández (CSI021-19) are supported by FSE-Conserjería de Educación de la Junta de Castilla y León 2019 and 2020 (ESF, European Social Fund) fellowship, respectively. J. Raboso-Gallego is supported by a scholarship from University of Salamanca co-financed by Banco Santander and ESF. S. Alemán-Arteaga is supported by an Ayuda para Contratos predoctorales para la formación de doctores (PRE2019-088887)

A novel speech analysis algorithm to detect cognitive impairment in a Spanish population

ObjectiveEarly detection of cognitive impairment in the elderly is crucial for diagnosis and appropriate care. Brief, cost-effective cognitive screening instruments are needed to help identify individuals who require further evaluation. This study presents preliminary data on a new screening technology using automated voice recording analysis software in a Spanish population.MethodData were collected from 174 Spanish-speaking individuals clinically diagnosed as cognitively normal (CN, n = 87) or impaired (mild cognitive impairment [MCI], n = 63; all-cause dementia, n = 24). Participants were recorded performing four common language tasks (Animal fluency, alternating fluency [sports and fruits], phonemic “F” fluency, and Cookie Theft Description). Recordings were processed via text-transcription and digital-signal processing techniques to capture neuropsychological variables and audio characteristics. A training sample of 122 subjects with similar demographics across groups was used to develop an algorithm to detect cognitive impairment. Speech and task features were used to develop five independent machine learning (ML) models to compute scores between 0 and 1, and a final algorithm was constructed using repeated cross-validation. A socio-demographically balanced subset of 52 participants was used to test the algorithm. Analysis of covariance (ANCOVA), covarying for demographic characteristics, was used to predict logistically-transformed algorithm scores.ResultsMean logit algorithm scores were significantly different across groups in the testing sample (p < 0.01). Comparisons of CN with impaired (MCI + dementia) and MCI groups using the final algorithm resulted in an AUC of 0.93/0.90, with overall accuracy of 88.4%/87.5%, sensitivity of 87.5/83.3, and specificity of 89.2/89.2, respectively.ConclusionFindings provide initial support for the utility of this automated speech analysis algorithm as a screening tool for cognitive impairment in Spanish speakers. Additional study is needed to validate this technology in larger and more diverse clinical populations

Lineage choice decisions in B-cell development and leukemia

Every tissue in the body is founded by a particular type of cell that is termed stem cell. These cells are generally presumed to be tissue-specific and, as such, are also responsible for maintenance of the tissue throughout the lifespan of an organism. There are two fundamental aspects to the regular behaviour of tissues during their genesis and thereafter. The offspring of stem cells divide in a manner that is controlled to the social benefit of the organism to generate/ensure the bulk of tissue that is required. These dividing cells mature: they acquire individual characteristics to enable them to perform a unique function within the tissue. Within cancer cells aspects of this normal physiology are distorted. This means the many strands to understanding of the behaviour of normal cells cannot be loosened from the strands of efforts to unravel what goes wrong in cancer.Research in C Vicente-Dueñas group is partially supported by FEDER, “Miguel Servet” Grant (CP14/00082 -AES 2013-2016) from the Instituto de Salud Carlos III (Ministerio de Economía y Competitividad), “Fondo de Investigaciones Sanitarias/Instituto de Salud Carlos III” (PI17/00167). Research in I Sánchez-García (ISG) group is partially supported by FEDER and by MINECO (SAF2012-32810, SAF2015-64420-R and Red de Excelencia Consolider OncoBIO SAF2014-57791-REDC), by Junta de Castilla y León (UIC-017, and CSI001U16). ISG lab is a member of the EuroSyStem and the DECIDE Network funded by the European Union under the FP7 program. ISG has been supported by the German Carreras Foundation (DJCLS R13/26). G Rodríguez-Hernández was supported by FSE-Conserjería de Educación de la Junta de Castilla y León (CSI001-15)

Inhibition of inflammatory signaling in Pax5 mutant cells mitigates B-cell leukemogenesis

© The Author(s) 2020.PAX5 is one of the most frequently mutated genes in B-cell acute lymphoblastic leukemia (B-ALL), and children with inherited preleukemic PAX5 mutations are at a higher risk of developing the disease. Abnormal profiles of inflammatory markers have been detected in neonatal blood spot samples of children who later developed B-ALL. However, how inflammatory signals contribute to B-ALL development is unclear. Here, we demonstrate that Pax5 heterozygosis, in the presence of infections, results in the enhanced production of the inflammatory cytokine interleukin-6 (IL-6), which appears to act in an autocrine fashion to promote leukemia growth. Furthermore, in vivo genetic downregulation of IL-6 in these Pax5 heterozygous mice retards B-cell leukemogenesis, and in vivo pharmacologic inhibition of IL-6 with a neutralizing antibody in Pax5 mutant mice with B-ALL clears leukemic cells. Additionally, this novel IL–6 signaling paradigm identified in mice was also substantiated in humans. Altogether, our studies establish aberrant IL6 expression caused by Pax5 loss as a hallmark of Pax5-dependent B-ALL and the IL6 as a therapeutic vulnerability for B-ALL characterized by PAX5 loss.We would also like to thank all members of our groups for useful suggestions and for their critical reading of the manuscript. Research at G.C.’s laboratory was supported by Italian Association for Cancer Research (grant IG-17593 to GC) and Fondazione Cariplo (grant 2018-0339 to CP). Research at CC’s laboratory was partially supported by FEDER, EU, MINECO (SAF2017-83061-R), the “Fundación Ramón Areces,” a Research Contract with the “Fundación Síndrome de Wolf-Hirschhorn o 4p-”, and institutional grants from the “Fundación Ramón Areces” and “Banco de Santander” to the CBMSO. Research in the CVD group is partially supported by FEDER, “Miguel Servet” Grant (CPII19/00024—AES 2017-2020) from the Instituto de Salud Carlos III (Ministerio de Economía y Competitividad), “Fondo de Investigaciones Sanitarias/Instituto de Salud Carlos III” (PI17/00167). Research in the ISG group is partially supported by FEDER and by SAF2015-64420-R MINECO/FEDER, UE, RTI2018-093314-B-I00 MCIU/AEI/FEDER, UE, by Junta de Castilla y León (UIC-017, CSI001U16, and CSI234P18), and by the German Jose Carreras Foundation (DJCLS R13/26; DJCLS 07R/2019). CVD, and ISG have been supported by the German Federal Office for Radiation Protection (BfS)-Germany (FKZ: 3618S32274). M.R.O., and ISG have been supported by the Fundacion Unoentrecienmil (CUNINA project). Research in the A.O. group is partially supported by "Fondo de Investigaciones Sanitarias/Instituto de Salud Carlos III" - FEDER-Ministerio de Economía y Competitividad (PI19/01183). AC-G and M.I.-H. are supported by FSE-Conserjería de Educación de la Junta de Castilla y León 2019 and 2020 (ESF- European Social Fund) fellowship, respectively. J.R.-G. is supported by a scholarship from University of Salamanca co-financed by Banco Santander and ESF

An intact gut microbiome protects genetically predisposed mice against leukemia

[EN]The majority of childhood leukemias are precursor B-cell acute lymphoblastic leukemias (pB-ALLs) caused by a combination of prenatal genetic predispositions and oncogenic events occurring after birth. Although genetic predispositions are frequent in children (>1% to 5%), fewer than 1% of genetically predisposed carriers will develop pB-ALL. Although infectious stimuli are believed to play a major role in leukemogenesis, the critical determinants are not well defined. Here, by using murine models of pB-ALL, we show that microbiome disturbances incurred by antibiotic treatment early in life were sufficient to induce leukemia in genetically predisposed mice, even in the absence of infectious stimuli and independent of T cells. By using V4 and full-length 16S ribosomal RNA sequencing of a series of fecal samples, we found that genetic predisposition to pB-ALL (Pax5 heterozygosity or ETV6-RUNX1 fusion) shaped a distinct gut microbiome. Machine learning accurately (96.8%) predicted genetic predisposition using 40 of 3983 amplicon sequence variants as proxies for bacterial species. Transplantation of either wild-type (WT) or Pax51/- hematopoietic bone marrow cells into WT recipient mice revealed that the microbiome is shaped and determined in a donor genotype-specific manner. Gas chromatography-mass spectrometry (GC-MS) analyses of sera from WT and Pax51/- mice demonstrated the presence of a genotype-specific distinct metabolomic profile. Taken together, our data indicate that it is a lack of commensal microbiota rather than the presence of specific bacteria that promotes leukemia in genetically predisposed mice. Future large-scale longitudinal studies are required to determine whether targeted microbiome modification in children predisposed to pB-ALL could become a successful prevention strategy

Conditional expression of HGAL leads to the development of diffuse large B-cell lymphoma in mice

Diffuse large B-cell lymphomas (DLBCLs) are clinically and genetically heterogeneous tumors. Deregulation of diverse biological processes specific to B cells, such as B-cell receptor (BCR) signaling and motility regulation, contribute to lymphomagenesis. Human germinal center associated lymphoma (HGAL) is a B-cell–specific adaptor protein controlling BCR signaling and B lymphocyte motility. In normal B cells, it is expressed in germinal center (GC) B lymphocytes and promptly downregulated upon further differentiation. The majority of DLBCL tumors, primarily GC B-cell types, but also activated types, express HGAL. To investigate the consequences of constitutive expression of HGAL in vivo, we generated mice that conditionally express human HGAL at different stages of hematopoietic development using 3 restricted Cre-mediated approaches to initiate expression of HGAL in hematopoietic stem cells, pro-B cells, or GC B cells. Following immune stimulation, we observed larger GCs in mice in which HGAL expression was initiated in GC B cells. All 3 mouse strains developed DLBCL at a frequency of 12% to 30% starting at age 13 months, leading to shorter survival. Immunohistochemical studies showed that all analyzed tumors were of the GC B-cell type. Exon sequencing revealed mutations reported in human DLBCL. Our data demonstrate that constitutive enforced expression of HGAL leads to DLBCL development