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1.
Immunity ; 50(2): 462-476.e8, 2019 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-30770246

RESUMO

Although the fetal immune system is considered tolerogenic, preterm infants can suffer from severe intestinal inflammation, including necrotizing enterocolitis (NEC). Here, we demonstrate that human fetal intestines predominantly contain tumor necrosis factor-α (TNF-α)+CD4+CD69+ T effector memory (Tem) cells. Single-cell RNA sequencing of fetal intestinal CD4+ T cells showed a T helper 1 phenotype and expression of genes mediating epithelial growth and cell cycling. Organoid co-cultures revealed a dose-dependent, TNF-α-mediated effect of fetal intestinal CD4+ T cells on intestinal stem cell (ISC) development, in which low T cell numbers supported epithelial development, whereas high numbers abrogated ISC proliferation. CD4+ Tem cell frequencies were higher in inflamed intestines from preterm infants with NEC than in healthy infant intestines and showed enhanced TNF signaling. These findings reveal a distinct population of TNF-α-producing CD4+ T cells that promote mucosal development in fetal intestines but can also mediate inflammation upon preterm birth.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Feto/imunologia , Memória Imunológica/imunologia , Intestinos/imunologia , Fator de Necrose Tumoral alfa/imunologia , Animais , Linfócitos T CD4-Positivos/metabolismo , Células Epiteliais/citologia , Células Epiteliais/imunologia , Células Epiteliais/metabolismo , Feminino , Feto/metabolismo , Humanos , Recém-Nascido , Mucosa Intestinal/embriologia , Mucosa Intestinal/crescimento & desenvolvimento , Mucosa Intestinal/imunologia , Intestinos/embriologia , Intestinos/crescimento & desenvolvimento , Camundongos Endogâmicos C57BL , Gravidez , Células-Tronco/citologia , Células-Tronco/imunologia , Células-Tronco/metabolismo , Células Th1/imunologia , Células Th1/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
2.
Blood ; 137(13): 1713-1718, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33512430

RESUMO

Plasma cells no longer express a B-cell antigen receptor and are hence deprived of signals crucial for survival throughout B-cell development. Instead, normal plasma cells, as well as their malignant myeloma counterparts, heavily rely on communication with the bone marrow (BM) microenvironment for survival. The plasma cell heparan sulfate proteoglycan (HSPG) syndecan-1 (CD138) and HSPGs in the BM microenvironment act as master regulators of this communication by co-opting specific growth and survival factors from the BM niche. This designates syndecan-1/HSPGs and their synthesis machinery as potential treatment targets in multiple myeloma.


Assuntos
Heparitina Sulfato/metabolismo , Mieloma Múltiplo/patologia , Plasmócitos/patologia , Proteoglicanas/metabolismo , Sindecana-1/metabolismo , Animais , Medula Óssea/metabolismo , Medula Óssea/patologia , Humanos , Mieloma Múltiplo/metabolismo , Plasmócitos/metabolismo , Microambiente Tumoral
3.
Haematologica ; 108(3): 797-810, 2023 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-36226498

RESUMO

BCL-2 family proteins are frequently aberrantly expressed in mantle cell lymphoma (MCL). Recently, the BCL-2-specific inhibitor venetoclax has been approved by the US Food and Drug Administration for chronic lymphocytic leukemia (CLL) and acute myeloid leukemia (AML). In MCL, venetoclax has shown promising efficacy in early clinical trials; however, a significant subset of patients is resistant. By conducting a kinome-centered CRISPR-Cas9 knockout sensitizer screen, we identified casein kinase 2 (CK2) as a major regulator of venetoclax resistance in MCL. Interestingly, CK2 is over-expressed in MCL and high CK2 expression is associated with poor patient survival. Targeting of CK2, either by inducible short hairpin RNA (shRNA)-mediated knockdown of CK2 or by the CK2-inhibitor silmitasertib, did not affect cell viability by itself, but strongly synergized with venetoclax in both MCL cell lines and primary samples, also if combined with ibrutinib. Furthermore, targeting of CK2 reduced MCL-1 levels, which involved impaired MCL-1 translation by inhibition of eIF4F complex assembly, without affecting BCL-2 and BCL-XL expression. Combined, this results in enhanced BCL-2 dependence and, consequently, venetoclax sensitization. In cocultures, targeting of CK2 overcame stroma-mediated venetoclax resistance of MCL cells. Taken together, our findings indicate that targeting of CK2 sensitizes MCL cells to venetoclax through downregulation of MCL-1. These novel insights provide a strong rationale for combining venetoclax with CK2 inhibition as therapeutic strategy for MCL patients.


Assuntos
Antineoplásicos , Linfoma de Célula do Manto , Humanos , Adulto , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Linfoma de Célula do Manto/tratamento farmacológico , Linfoma de Célula do Manto/genética , Linfoma de Célula do Manto/metabolismo , Caseína Quinase II/genética , Caseína Quinase II/metabolismo , Regulação para Baixo , Linhagem Celular Tumoral , Proteínas Proto-Oncogênicas c-bcl-2 , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/uso terapêutico
4.
Am J Hum Genet ; 100(2): 281-296, 2017 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-28132690

RESUMO

EXTL3 regulates the biosynthesis of heparan sulfate (HS), important for both skeletal development and hematopoiesis, through the formation of HS proteoglycans (HSPGs). By whole-exome sequencing, we identified homozygous missense mutations c.1382C>T, c.1537C>T, c.1970A>G, and c.2008T>G in EXTL3 in nine affected individuals from five unrelated families. Notably, we found the identical homozygous missense mutation c.1382C>T (p.Pro461Leu) in four affected individuals from two unrelated families. Affected individuals presented with variable skeletal abnormalities and neurodevelopmental defects. Severe combined immunodeficiency (SCID) with a complete absence of T cells was observed in three families. EXTL3 was most abundant in hematopoietic stem cells and early progenitor T cells, which is in line with a SCID phenotype at the level of early T cell development in the thymus. To provide further support for the hypothesis that mutations in EXTL3 cause a neuro-immuno-skeletal dysplasia syndrome, and to gain insight into the pathogenesis of the disorder, we analyzed the localization of EXTL3 in fibroblasts derived from affected individuals and determined glycosaminoglycan concentrations in these cells as well as in urine and blood. We observed abnormal glycosaminoglycan concentrations and increased concentrations of the non-sulfated chondroitin disaccharide D0a0 and the disaccharide D0a4 in serum and urine of all analyzed affected individuals. In summary, we show that biallelic mutations in EXTL3 disturb glycosaminoglycan synthesis and thus lead to a recognizable syndrome characterized by variable expression of skeletal, neurological, and immunological abnormalities.


Assuntos
Anormalidades Musculoesqueléticas/genética , N-Acetilglucosaminiltransferases/genética , Osteocondrodisplasias/genética , Alelos , Linhagem Celular , Linhagem Celular Tumoral , Condroitina/sangue , Condroitina/urina , Variações do Número de Cópias de DNA , Estudo de Associação Genômica Ampla , Glicosaminoglicanos/metabolismo , Humanos , Anormalidades Musculoesqueléticas/diagnóstico , Mutação de Sentido Incorreto , Osteocondrodisplasias/diagnóstico , Imunodeficiência Combinada Severa/diagnóstico , Imunodeficiência Combinada Severa/genética
5.
Blood ; 131(9): 982-994, 2018 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-29212806

RESUMO

Multiple myeloma (MM) is characterized by the expansion of malignant plasma cells in the bone marrow (BM). Most MMs display aberrant Wnt/ß-catenin signaling, which drives proliferation; however, they lack oncogenic Wnt pathway mutations, suggesting activation by autocrine Wnt ligands and/or paracrine Wnts from the BM microenvironment. Expression of the heparan sulfate (HS) proteoglycan syndecan-1 is a hallmark of MM. Syndecan-1 is a critical player in the complex reciprocal interaction between MM cells and their BM niche, mediating growth factor/cytokine binding and signaling by its HS chains. Here, by means of CRISPR/Cas9-mediated knockout and doxycycline-inducible short hairpin RNA-mediated knockdown of EXT1, a critical enzyme for HS polymerization, we demonstrate that the HS chains decorating syndecan-1 mediate aberrant Wnt pathway activation in MM. HS-deficient MM cells exhibited strongly decreased autocrine Wnt/ß-catenin pathway activity and reduced Wnt pathway-dependent proliferation. In addition, we demonstrate that Wnts bind to the HS side chains of syndecan-1 and that this binding contributes to paracrine Wnt pathway activation through the Wnt receptor Frizzled (Fzd). Furthermore, in an HS-dependent fashion, syndecan-1 also binds osteoblast-produced R-spondin, which represses Fzd degradation by activation of LGR4, an R-spondin receptor aberrantly expressed on MM cells. Costimulation with R-spondin and its binding to HS chains decorating syndecan-1 are indispensable for optimal stimulation of Wnt signaling in MM. Taken together, our results identify syndecan-1 as a crucial component of the Wnt signalosome in MM cells, binding Wnts and R-spondins to promote aberrant Wnt/ß-catenin signaling and cell growth, and suggest HS and its biosynthetic enzymes as potential targets in the treatment of MM.


Assuntos
Mieloma Múltiplo/metabolismo , Proteínas de Neoplasias/metabolismo , Sindecana-1/metabolismo , Trombospondinas/metabolismo , Proteínas Wnt/metabolismo , Via de Sinalização Wnt , Linhagem Celular Tumoral , Receptores Frizzled/genética , Receptores Frizzled/metabolismo , Heparitina Sulfato/genética , Heparitina Sulfato/metabolismo , Humanos , Mieloma Múltiplo/genética , Mieloma Múltiplo/patologia , Proteínas de Neoplasias/genética , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Sindecana-1/genética , Trombospondinas/genética , Proteínas Wnt/genética , beta Catenina/genética , beta Catenina/metabolismo
6.
Haematologica ; 105(2): 424-434, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31123031

RESUMO

The 2016 World Health Organization classification defines diffuse large B-cell lymphoma (DLBCL) subtypes based on Epstein-Barr virus (EBV) infection and oncogenic rearrangements of MYC/BCL2/BCL6 as drivers of lymphomagenesis. A subset of DLBCL, however, is characterized by activating mutations in MYD88/CD79B We investigated whether MYD88/CD79B mutations could improve the classification and prognostication of DLBCL. In 250 primary DLBCL, MYD88/CD79B mutations were identified by allele-specific polymerase chain reaction or next-generation-sequencing, MYC/BCL2/BCL6 rearrangements were analyzed by fluorescence in situ hybridization, and EBV was studied by EBV-encoded RNA in situ hybridization. Associations of molecular features with clinicopathologic characteristics, outcome, and prognosis according to the International Prognostic Index (IPI) were investigated. MYD88 and CD79B mutations were identified in 29.6% and 12.3%, MYC, BCL2, and BCL6 rearrangements in 10.6%, 13.6%, and 20.3%, and EBV in 11.7% of DLBCL, respectively. Prominent mutual exclusivity between EBV positivity, rearrangements, and MYD88/CD79B mutations established the value of molecular markers for the recognition of biologically distinct DLBCL subtypes. MYD88-mutated DLBCL had a significantly inferior 5-year overall survival than wild-type MYD88 DLBCL (log-rank; P=0.019). DLBCL without any of the studied aberrations had superior overall survival compared to cases carrying ≥1 aberrancy (log-rank; P=0.010). MYD88 mutations retained their adverse prognostic impact upon adjustment for other genetic and clinical variables by multivariable analysis and improved the prognostic performance of the IPI. This study demonstrates the clinical utility of defining MYD88-mutated DLBCL as a distinct molecular subtype with adverse prognosis. Our data call for sequence analysis of MYD88 in routine diagnostics of DLBCL to optimize classification and prognostication, and to guide the development of improved treatment strategies.


Assuntos
Infecções por Vírus Epstein-Barr , Linfoma Difuso de Grandes Células B , Fator 88 de Diferenciação Mieloide/genética , Infecções por Vírus Epstein-Barr/genética , Herpesvirus Humano 4 , Humanos , Hibridização in Situ Fluorescente , Linfoma Difuso de Grandes Células B/diagnóstico , Linfoma Difuso de Grandes Células B/genética , Mutação , Prognóstico
7.
Proc Natl Acad Sci U S A ; 114(2): 376-381, 2017 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-28028233

RESUMO

The unrestrained growth of tumor cells is generally attributed to mutations in essential growth control genes, but tumor cells are also affected by, or even addicted to, signals from the microenvironment. As therapeutic targets, these extrinsic signals may be equally significant as mutated oncogenes. In multiple myeloma (MM), a plasma cell malignancy, most tumors display hallmarks of active Wnt signaling but lack activating Wnt-pathway mutations, suggesting activation by autocrine Wnt ligands and/or paracrine Wnts emanating from the bone marrow (BM) niche. Here, we report a pivotal role for the R-spondin/leucine-rich repeat-containing G protein-coupled receptor 4 (LGR4) axis in driving aberrant Wnt/ß-catenin signaling in MM. We show that LGR4 is expressed by MM plasma cells, but not by normal plasma cells or B cells. This aberrant LGR4 expression is driven by IL-6/STAT3 signaling and allows MM cells to hijack R-spondins produced by (pre)osteoblasts in the BM niche, resulting in Wnt (co)receptor stabilization and a dramatically increased sensitivity to auto- and paracrine Wnts. Our study identifies aberrant R-spondin/LGR4 signaling with consequent deregulation of Wnt (co)receptor turnover as a driver of oncogenic Wnt/ß-catenin signaling in MM cells. These results advocate targeting of the LGR4/R-spondin interaction as a therapeutic strategy in MM.


Assuntos
Glicoproteínas de Membrana/metabolismo , Mieloma Múltiplo/metabolismo , Osteoblastos/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Proteínas Wnt/metabolismo , Via de Sinalização Wnt/fisiologia , Animais , Linhagem Celular Tumoral , Células HEK293 , Humanos , Interleucina-6/metabolismo , Ligantes , Camundongos , Ligação Proteica/fisiologia , Fator de Transcrição STAT3/metabolismo , beta Catenina/metabolismo
8.
Haematologica ; 104(12): 2337-2348, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31699794

RESUMO

More than 50 subtypes of B-cell non-Hodgkin lymphoma (B-NHL) are recognized in the most recent World Health Organization classification of 2016. The current treatment paradigm, however, is largely based on 'one-size-fits-all' immune-chemotherapy. Unfortunately, this therapeutic strategy is inadequate for a significant number of patients. As such, there is an indisputable need for novel, preferably targeted, therapies based on a biologically driven classification and risk stratification. Sequencing studies identified mutations in the MYD88 gene as an important oncogenic driver in B-cell lymphomas. MYD88 mutations constitutively activate NF-κB and its associated signaling pathways, thereby promoting B-cell proliferation and survival. High frequencies of the hotspot MYD88(L265P) mutation are observed in extranodal diffuse large B-cell lymphoma and Waldenström macroglobulinemia, thereby demonstrating this mutation's potential as a disease marker. In addition, the presence of mutant MYD88 predicts survival outcome in B-NHL subtypes and it provides a therapeutic target. Early clinical trials targeting MYD88 have shown encouraging results in relapsed/refractory B-NHL. Patients with these disorders can benefit from analysis for the MYD88 hotspot mutation in liquid biopsies, as a minimally invasive method to demonstrate treatment response or resistance. Given these clear clinical implications and the crucial role of MYD88 in lymphomagenesis, we expect that analysis of this gene will increasingly be used in routine clinical practice, not only as a diagnostic classifier, but also as a prognostic and therapeutic biomarker directing precision medicine. This review focuses on the pivotal mechanistic role of mutated MYD88 and its clinical implications in B-NHL.


Assuntos
Regulação Neoplásica da Expressão Gênica , Leucemia de Células B/patologia , Linfoma de Células B/patologia , Mutação , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/metabolismo , Humanos , Leucemia de Células B/genética , Leucemia de Células B/metabolismo , Linfoma de Células B/genética , Linfoma de Células B/metabolismo , Prognóstico
9.
Gastroenterology ; 153(4): 1040-1053.e4, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28716720

RESUMO

BACKGROUND & AIMS: Resistance of metastatic human colorectal cancer cells to drugs that block epidermal growth factor (EGF) receptor signaling could be caused by aberrant activity of other receptor tyrosine kinases, activating overlapping signaling pathways. One of these receptor tyrosine kinases could be MET, the receptor for hepatocyte growth factor (HGF). We investigated how MET signaling, and its interaction with CD44 (a putative MET coreceptor regulated by Wnt signaling and highly expressed by intestinal stem cells [ISCs] and adenomas) affects intestinal homeostasis, regeneration, and adenoma formation in mini-gut organoids and mice. METHODS: We established organoid cultures from ISCs stimulated with HGF or EGF and assessed intestinal differentiation by immunohistochemistry. Mice with total epithelial disruption of MET (AhCre/Metfl/fl/LacZ) or ISC-specific disruption of MET (Lgr5Creert2/Metfl/fl/LacZ) and control mice (AhCre/Met+/+/LacZ, Lgr5Creert2/Met+/+/LacZ) were exposed to 10 Gy total body irradiation; intestinal tissues were collected, and homeostasis and regeneration were assessed by immunohistochemistry. We investigated adenoma organoid expansion stimulated by HGF or EGF using adenomas derived from Lgr5Creert2/Metfl/fl/Apcfl/fl and Lgr5Creert2/Met+/+/Apcfl/fl mice. The same mice were evaluated for adenoma prevalence and size. We also quantified adenomas in AhCre/Metfl/fl/Apcfl/+ mice compared with AhCre/Met+/+/Apcfl/+ control mice. We studied expansion of organoids generated from crypts and adenomas, stimulated by HGF or EGF, that were derived from mice expressing different CD44 splice variants (Cd44+/+, Cd44-/-, Cd44s/s, or Cd44v4-10/v4-10 mice). RESULTS: Crypts incubated with EGF or HGF expanded into self-organizing mini-guts with similar levels of efficacy and contained all differentiated cell lineages. MET-deficient mice did not have defects in intestinal homeostasis. Total body irradiation reduced numbers of proliferating crypts in AhCre/Metfl/fl/LacZ mice. Lgr5Creert2/Metfl/fl/LacZ mice had impaired regeneration of MET-deficient ISCs. Adenoma organoids stimulated with EGF or HGF expanded to almost twice the size of nonstimulated organoids. MET-deficient adenoma organoids did not respond to HGF stimulation, but did respond to EGF. ISC-specific disruption of Met (Lgr5Creert2/Metfl/fl/Apcfl/fl mice) caused a twofold increase in apoptosis in microadenomas, resulting in an approximately 50% reduction of microadenoma numbers and significantly reduced average adenoma size. Total epithelial disruption of Met (AhCre/Metfl/fl/Apcfl/+ mice) resulted in an approximate 50% reduction in (micro)adenoma numbers. Intestinal crypts from Cd44-/- mice did not expand to the same extent as crypts from Cd44+/+ mice on stimulation with HGF, but had the same response to EGF. The negative effect on HGF-mediated growth was overcome by expression of CD44v4-10, but not by CD44s. Similarly, HGF-mediated expansion of adenoma organoids required CD44v4-10. CONCLUSIONS: In studies of intestinal organoid cultures and mice with inducible deletion of MET, we found HGF receptor signaling to regulate intestinal homeostasis and regeneration, as well as adenoma formation. These activities of MET are promoted by the stem cell CD44 isoform CD44v4-10. Our findings provide rationale for targeting signaling via MET and CD44 during anti-EGF receptor therapy of patients with colorectal cancer or in patients resistant to EGF receptor inhibitors.


Assuntos
Adenoma/metabolismo , Proliferação de Células , Transformação Celular Neoplásica/metabolismo , Receptores de Hialuronatos/metabolismo , Neoplasias Intestinais/metabolismo , Intestinos/enzimologia , Proteínas Proto-Oncogênicas c-met/metabolismo , Regeneração , Células-Tronco/enzimologia , Adenoma/genética , Adenoma/patologia , Animais , Diferenciação Celular , Linhagem da Célula , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/efeitos da radiação , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Células Cultivadas , Fator de Crescimento Epidérmico/farmacologia , Receptores ErbB/metabolismo , Genótipo , Fator de Crescimento de Hepatócito/farmacologia , Homeostase , Receptores de Hialuronatos/genética , Neoplasias Intestinais/genética , Neoplasias Intestinais/patologia , Intestinos/efeitos dos fármacos , Intestinos/patologia , Intestinos/efeitos da radiação , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fenótipo , Proteínas Proto-Oncogênicas c-met/genética , Regeneração/efeitos dos fármacos , Regeneração/efeitos da radiação , Transdução de Sinais , Células-Tronco/efeitos dos fármacos , Células-Tronco/patologia , Células-Tronco/efeitos da radiação , Fatores de Tempo , Técnicas de Cultura de Tecidos , Carga Tumoral
11.
Haematologica ; 103(3): 486-496, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29269521

RESUMO

Although it is known that B-cell lymphomas occur more frequently in immunocompromised patients, thus far such an association has not been clearly established for T-cell lymphomas. Of the 251 patients who were diagnosed with a T-cell non-Hodgkin lymphoma in our center between 1999 and 2014, at least 25 were identified in immunocompromised patients. Herein, we retrospectively analyzed the clinical and pathological characteristics of these 25 cases. In addition, we searched the literature and present an overview of 605 previously published cases. The actual number of patients with B-cell chronic lymphocytic leukemia and patients on immunosuppressive drugs for inflammatory bowel disease or rheumatoid arthritis in the total cohort of 251 patients diagnosed with T-cell non-Hodgkin lymphoma was much higher than the number of patients expected to have these diseases in this cohort, based on their prevalence in the general population. This, together with the large number of additional cases found in the literature, suggest that the risk of developing T-cell non-Hodgkin lymphoma is increased in immunocompromised patients. Compared to T-cell non-Hodgkin lymphoma in the general population, these lymphomas are more often located extranodally, present at a younger age and appear to have a poor outcome. The observations made in the study herein should raise awareness of the possible development of T-cell non-Hodgkin lymphoma in immunodeficient patients, and challenge the prolonged use of immunosuppressive drugs in patients who are in clinical remission of their autoimmune disease.


Assuntos
Imunossupressores/efeitos adversos , Linfoma de Células T/induzido quimicamente , Adulto , Idoso , Feminino , Humanos , Hospedeiro Imunocomprometido , Síndromes de Imunodeficiência/complicações , Linfoma de Células T/epidemiologia , Linfoma de Células T/etiologia , Linfoma de Células T/mortalidade , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Análise de Sobrevida , Adulto Jovem
12.
Blood ; 126(18): 2098-109, 2015 Oct 29.
Artigo em Inglês | MEDLINE | ID: mdl-26289642

RESUMO

Expression of the forkhead transcription factor FOXP1 is essential for early B-cell development, whereas downregulation of FOXP1 at the germinal center (GC) stage is required for GC B-cell function. Aberrantly high FOXP1 expression is frequently observed in diffuse large B-cell lymphoma and mucosa-associated lymphoid tissue lymphoma, being associated with poor prognosis. Here, by gene expression analysis upon ectopic overexpression of FOXP1 in primary human memory B cells (MBCs) and B-cell lines, combined with chromatin immunoprecipitation and sequencing, we established that FOXP1 directly represses expression of PRDM1, IRF4, and XBP1, transcriptional master regulators of plasma cell (PC) differentiation. In accordance, FOXP1 is prominently expressed in primary human naive and MBCs, but expression strongly decreases during PC differentiation. Moreover, as compared with immunoglobulin (Ig) M(+) MBCs, IgG(+) MBCs combine lower expression of FOXP1 with an enhanced intrinsic PC differentiation propensity, and constitutive (over)expression of FOXP1 in B-cell lines and primary human MBCs represses their ability to differentiate into PCs. Taken together, our data indicate that proper control of FOXP1 expression plays a critical role in PC differentiation, whereas aberrant expression of FOXP1 might contribute to lymphomagenesis by blocking this terminal B-cell differentiation.


Assuntos
Linfócitos B/citologia , Fatores de Transcrição Forkhead/metabolismo , Plasmócitos/citologia , Proteínas Repressoras/metabolismo , Linfócitos B/metabolismo , Diferenciação Celular , Linhagem Celular , Células Cultivadas , Fatores de Transcrição Forkhead/genética , Humanos , Imunoglobulina G/análise , Imunoglobulina M/análise , Plasmócitos/metabolismo , Proteínas Repressoras/genética , Ativação Transcricional , Regulação para Cima
13.
Haematologica ; 102(3): 573-583, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-27909217

RESUMO

The forkhead transcription factor FOXP1 is generally regarded as an oncogene in activated B cell-like diffuse large B-cell lymphoma. Previous studies have suggested that a small isoform of FOXP1 rather than full-length FOXP1, may possess this oncogenic activity. Corroborating those studies, we herein show that activated B cell-like diffuse large B-cell lymphoma cell lines and primary activated B cell-like diffuse large B-cell lymphoma cells predominantly express a small FOXP1 isoform, and that the 5'-end of the Foxp1 gene is a common insertion site in murine lymphomas in leukemia virus- and transposon-mediated insertional mutagenesis screens. By combined mass spectrometry, (quantative) reverse transcription polymerase chain reaction/sequencing, and small interfering ribonucleic acid-mediated gene silencing, we determined that the small FOXP1 isoform predominantly expressed in activated B cell-like diffuse large B-cell lymphoma lacks the N-terminal 100 amino acids of full-length FOXP1. Aberrant overexpression of this FOXP1 isoform (ΔN100) in primary human B cells revealed its oncogenic capacity; it repressed apoptosis and plasma cell differentiation. However, no difference in potency was found between this small FOXP1 isoform and full-length FOXP1. Furthermore, overexpression of full-length FOXP1 or this small FOXP1 isoform in primary B cells and diffuse large B-cell lymphoma cell lines resulted in similar gene regulation. Taken together, our data indicate that this small FOXP1 isoform and full-length FOXP1 have comparable oncogenic and transcriptional activity in human B cells, suggesting that aberrant expression or overexpression of FOXP1, irrespective of the specific isoform, contributes to lymphomagenesis. These novel insights further enhance the value of FOXP1 for the diagnostics, prognostics, and treatment of diffuse large B-cell lymphoma patients.


Assuntos
Linfócitos B/metabolismo , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Regulação Neoplásica da Expressão Gênica , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Ativação Transcricional , Processamento Alternativo , Sequência de Aminoácidos , Animais , Linfócitos B/citologia , Linfócitos B/imunologia , Caspase 3/metabolismo , Caspase 7/metabolismo , Ciclo Celular/genética , Diferenciação Celular , Linhagem Celular Tumoral , Proliferação de Células/genética , Sobrevivência Celular/genética , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/química , Humanos , Memória Imunológica , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Camundongos , Mutagênese Insercional , Plasmócitos/citologia , Plasmócitos/imunologia , Plasmócitos/metabolismo , Isoformas de Proteínas , Proteínas Repressoras/química
14.
Blood ; 124(23): 3431-40, 2014 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-25267198

RESUMO

The forkhead transcription factor FOXP1 is involved in B-cell development and function and is generally regarded as an oncogene in activated B-cell-like subtype of diffuse large B-cell lymphoma (DLBCL) and mucosa-associated lymphoid tissue lymphoma, lymphomas relying on constitutive nuclear factor κB (NF-κB) activity for survival. However, the mechanism underlying its putative oncogenic activity has not been established. By gene expression microarray, upon overexpression or silencing of FOXP1 in primary human B cells and DLBCL cell lines, combined with chromatin immunoprecipitation followed by next-generation sequencing, we established that FOXP1 directly represses a set of 7 proapoptotic genes. Low expression of these genes, encoding the BH3-only proteins BIK and Harakiri, the p53-regulatory proteins TP63, RASSF6, and TP53INP1, and AIM2 and EAF2, is associated with poor survival in DLBCL patients. In line with these findings, we demonstrated that FOXP1 promotes the expansion of primary mature human B cells by inhibiting caspase-dependent apoptosis, without affecting B-cell proliferation. Furthermore, FOXP1 is dependent upon, and cooperates with, NF-κB signaling to promote B-cell expansion and survival. Taken together, our data indicate that, through direct repression of proapoptotic genes, (aberrant) expression of FOXP1 complements (constitutive) NF-κB activity to promote B-cell survival and can thereby contribute to B-cell homeostasis and lymphomagenesis.


Assuntos
Proteínas Reguladoras de Apoptose/genética , Linfócitos B/fisiologia , Fatores de Transcrição Forkhead/fisiologia , Regulação Leucêmica da Expressão Gênica , NF-kappa B/fisiologia , Proteínas Repressoras/fisiologia , Transcrição Gênica , Proteínas Reguladoras de Apoptose/metabolismo , Sobrevivência Celular/genética , Transformação Celular Neoplásica/genética , Células Cultivadas , Regulação para Baixo , Perfilação da Expressão Gênica , Humanos , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/patologia , Análise em Microsséries , Regulação para Cima
16.
Blood ; 122(14): 2412-24, 2013 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-23940282

RESUMO

Ibrutinib (PCI-32765) is a highly potent oral Bruton tyrosine kinase (BTK) inhibitor in clinical development for treating B-cell lymphoproliferative diseases. Patients with chronic lymphocytic leukemia (CLL) often show marked, transient increases of circulating CLL cells following ibrutinib treatments, as seen with other inhibitors of the B-cell receptor (BCR) pathway. In a phase 1 study of ibrutinib, we noted similar effects in patients with mantle cell lymphoma (MCL). Here, we characterize the patterns and phenotypes of cells mobilized among patients with MCL and further investigate the mechanism of this effect. Peripheral blood CD19(+)CD5(+) cells from MCL patients were found to have significant reduction in the expression of CXCR4, CD38, and Ki67 after 7 days of treatment. In addition, plasma chemokines such as CCL22, CCL4, and CXCL13 were reduced 40% to 60% after treatment. Mechanistically, ibrutinib inhibited BCR- and chemokine-mediated adhesion and chemotaxis of MCL cell lines and dose-dependently inhibited BCR, stromal cell, and CXCL12/CXCL13 stimulations of pBTK, pPLCγ2, pERK, or pAKT. Importantly, ibrutinib inhibited migration of MCL cells beneath stromal cells in coculture. We propose that BTK is essential for the homing of MCL cells into lymphoid tissues, and its inhibition results in an egress of malignant cells into peripheral blood. This trial was registered at www.clinicaltrials.gov as #NCT00114738.


Assuntos
Antineoplásicos/uso terapêutico , Linfócitos B/efeitos dos fármacos , Quimiotaxia de Leucócito/efeitos dos fármacos , Linfoma de Célula do Manto/sangue , Pirazóis/uso terapêutico , Pirimidinas/uso terapêutico , Adenina/análogos & derivados , Antígenos CD19/biossíntese , Linfócitos B/metabolismo , Western Blotting , Antígenos CD5/biossíntese , Adesão Celular/efeitos dos fármacos , Citometria de Fluxo , Humanos , Linfoma de Célula do Manto/tratamento farmacológico , Piperidinas , Inibidores de Proteínas Quinases/uso terapêutico
17.
Blood ; 122(17): 3010-9, 2013 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-24014238

RESUMO

Chronic lymphocytic leukemia (CLL) cells multiply in secondary lymphoid tissue, but the mechanisms leading to their proliferation are still uncertain. In addition to B-cell receptor (BCR)-triggered signals, other microenvironmental factors might well be involved. In proliferation centers, leukemic B cells are in close contact with CD4(+)CD40L(+) T cells. Therefore, we here dissected the signals provided by autologous activated T cells (Tact) to CLL cells. Although the gene expression profile induced by Tact was highly similar to that induced by sole CD40 signaling, an obvious difference was that Tact induced proliferation of CLL cells. We determined that stimulation with only CD40L+IL-21 was sufficient to induce robust proliferation in CLL cells. We then defined an interleukin (IL)-21-induced gene signature in CLL, containing components of Janus kinase/signal transducer and activator of transcription and apoptosis pathways, and this signature could be detected in lymph node (LN) samples from patients. Finally, we could detect IL-21 RNA and protein in LN, and IL-21 production ex vivo by LN CD4(+)CXCR5(+) follicular helper T cells. These results indicate that in addition to BCR signaling, activated T cells might contribute to CLL cell proliferation via CD40 and IL-21. Targeting these signaling pathways might offer new venues for treatment of CLL.


Assuntos
Linfócitos B/imunologia , Linfócitos T CD4-Positivos/imunologia , Ligante de CD40/imunologia , Interleucinas/imunologia , Leucemia Linfocítica Crônica de Células B/imunologia , Linfonodos/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Antígenos de Diferenciação de Linfócitos B/genética , Antígenos de Diferenciação de Linfócitos B/imunologia , Linfócitos B/patologia , Linfócitos T CD4-Positivos/patologia , Antígenos CD40/genética , Antígenos CD40/imunologia , Ligante de CD40/genética , Comunicação Celular/imunologia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Interleucinas/genética , Janus Quinases/genética , Janus Quinases/imunologia , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/patologia , Linfonodos/patologia , Ativação Linfocitária , Cultura Primária de Células , Receptores de Antígenos de Linfócitos B/genética , Receptores de Antígenos de Linfócitos B/imunologia , Transdução de Sinais , Linfócitos T Auxiliares-Indutores/patologia
20.
Kidney Int ; 86(3): 558-69, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24717295

RESUMO

Chronic kidney diseases (CKDs) are characterized by tubular atrophy and interstitial fibrosis. We previously showed that in obstructive nephropathy de novo CD44 renal expression contributes to renal fibrosis but attenuates tubular damage/apoptosis. As CD44-standard (CD44s) has been linked to TGF-ß1-mediated actions and CD44-variant-3 (CD44v3) favors HGF-c-Met binding, we compared the functional properties of these CD44 isoforms in the progression of obstructive nephropathy, using specific CD44-variant knockout/knockin mice. The presence of CD44v3 diminished tubular damage during obstructive nephropathy, decreased apoptosis, and increased proliferation of tubular epithelial cells, and prevented renal fibrosis development. In contrast, expression of CD44s led to increased tubular damage and tubular epithelial cell apoptosis, and more renal fibrosis. A relative increase in renal ß-catenin expression, HGF production, and HGF/c-Met signaling, together with a relative inhibition of TGF-ß1 downstream signaling and TGF-ß type I receptor expression, was found in CD44v3 mice compared with CD44s littermates. In line with this, Wnt3a/HGF treatment of tubular cells resulted in higher ß-catenin/p-AKT levels in CD44v3(+) tubular epithelial cells, whereas TGF-ß1 induced a mild collagen I upregulation in CD44v3(+) mouse embryonic fibroblasts as compared with CD44s(+) cells. Thus, CD44s and CD44v3 exert opposite roles in the progression of obstructive nephropathy, with CD44v3-v10 being the protective isoform that delays evolution of the renal pathology.


Assuntos
Receptores de Hialuronatos/metabolismo , Nefropatias/metabolismo , Nefropatias/patologia , Túbulos Renais/metabolismo , Túbulos Renais/patologia , Animais , Apoptose , Doença Crônica , Colágeno Tipo I/metabolismo , Células Epiteliais , Fibroblastos/metabolismo , Fibrose , Fator de Crescimento de Hepatócito/metabolismo , Fator de Crescimento de Hepatócito/farmacologia , Receptores de Hialuronatos/genética , Nefropatias/etiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosforilação , Isoformas de Proteínas , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-met/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo I , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta1/metabolismo , Regulação para Cima , Obstrução Ureteral/complicações , Proteína Wnt3A/farmacologia , beta Catenina/metabolismo
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