RESUMEN
Over 50% of patients with systemic LCH are not cured with front-line therapies, and data to guide salvage options are limited. We describe 58 patients with LCH who were treated with clofarabine. Clofarabine monotherapy was active against LCH in this cohort, including heavily pretreated patients with a systemic objective response rate of 92.6%, higher in children (93.8%) than adults (83.3%). BRAFV600E+ variant allele frequency in peripheral blood is correlated with clinical responses. Prospective multicentre trials are warranted to determine optimal dosing, long-term efficacy, late toxicities, relative cost and patient-reported outcomes of clofarabine compared to alternative LCH salvage therapy strategies.
Asunto(s)
Clofarabina , Histiocitosis de Células de Langerhans , Humanos , Clofarabina/uso terapéutico , Clofarabina/administración & dosificación , Histiocitosis de Células de Langerhans/tratamiento farmacológico , Masculino , Femenino , Adulto , Adolescente , Niño , Persona de Mediana Edad , Preescolar , Adulto Joven , Anciano , Recurrencia , Proteínas Proto-Oncogénicas B-raf/genética , Lactante , Resultado del Tratamiento , Terapia Recuperativa , Nucleótidos de Adenina/uso terapéutico , Nucleótidos de Adenina/administración & dosificación , Nucleótidos de Adenina/efectos adversos , Arabinonucleósidos/uso terapéutico , Arabinonucleósidos/administración & dosificación , Arabinonucleósidos/efectos adversosRESUMEN
SMARCA4 encodes one of two mutually exclusive ATPase subunits in the BRG/BRM associated factor (BAF) complex that is recruited by transcription factors (TFs) to drive chromatin accessibility and transcriptional activation. SMARCA4 is among the most recurrently mutated genes in human cancer, including â¼30% of germinal center (GC)-derived Burkitt lymphomas. In mice, GC-specific Smarca4 haploinsufficiency cooperated with MYC over-expression to drive lymphomagenesis. Furthermore, monoallelic Smarca4 deletion drove GC hyperplasia with centroblast polarization via significantly increased rates of centrocyte recycling to the dark zone. Mechanistically, Smarca4 loss reduced the activity of TFs that are activated in centrocytes to drive GC-exit, including SPI1 (PU.1), IRF family, and NF-κB. Loss of activity for these factors phenocopied aberrant BCL6 activity within murine centrocytes and human Burkitt lymphoma cells. SMARCA4 therefore facilitates chromatin accessibility for TFs that shape centrocyte trajectories, and loss of fine-control of these programs biases toward centroblast cell-fate, GC hyperplasia and lymphoma.
Asunto(s)
Haploinsuficiencia , Linfoma de Células B , Animales , Humanos , Ratones , Cromatina , ADN Helicasas/genética , Hiperplasia , Linfoma de Células B/genética , Proteínas Nucleares/genética , Factores de Transcripción/genéticaRESUMEN
ABSTRACT: Posttransplant lymphoproliferative disease (PTLD) in pediatric solid organ transplant (SOT) recipients is characterized by uncontrolled proliferation of Epstein-Barr virus-infected (EBV+) B cells due to decreased immune function. This study evaluated the feasibility, safety, clinical and immunobiological outcomes in pediatric SOT recipients with PTLD treated with rituximab and third-party latent membrane protein-specific T cells (LMP-TCs). Newly diagnosed (ND) patients without complete response to rituximab and all patients with relapsed/refractory (R/R) disease received LMP-TCs. Suitable LMP-TC products were available for all eligible subjects. Thirteen of 15 patients who received LMP-TCs were treated within the prescribed 14-day time frame. LMP-TC therapy was generally well tolerated. Notable adverse events included 3 episodes of rejection in cardiac transplant recipients during LMP-TC therapy attributed to subtherapeutic immunosuppression and 1 episode of grade 3 cytokine release syndrome. Clinical outcomes were associated with disease severity. Overall response rate (ORR) after LMP-TC cycle 1 was 70% (7/10) for the ND cohort and 20% (1/5) for the R/R cohort. For all cohorts combined, the best ORR for LMP-TC cycles 1 and 2 was 53% and the 2-year overall survival was 70.7%. vßT-cell receptor sequencing showed persistence of adoptively transferred third-party LMP-TCs for up to 8 months in the ND cohort. This study establishes the feasibility of administering novel T-cell therapies in a cooperative group clinical trial and demonstrates the potential for positive outcomes without chemotherapy for ND patients with PTLD. This trial was registered at www.clinicaltrials.gov as #NCT02900976 and at the Children's Oncology Group as ANHL1522.
Asunto(s)
Infecciones por Virus de Epstein-Barr , Trastornos Linfoproliferativos , Humanos , Niño , Rituximab/farmacología , Rituximab/uso terapéutico , Herpesvirus Humano 4 , Infecciones por Virus de Epstein-Barr/complicaciones , Infecciones por Virus de Epstein-Barr/tratamiento farmacológico , Linfocitos T , Trastornos Linfoproliferativos/tratamiento farmacológico , Trastornos Linfoproliferativos/etiología , Trastornos Linfoproliferativos/diagnósticoRESUMEN
Neurodegenerative diseases (ND) are characterized by progressive loss of neuronal function. Mechanisms of ND pathogenesis are incompletely understood, hampering the development of effective therapies. Langerhans cell histiocytosis (LCH) is an inflammatory neoplastic disorder caused by hematopoietic progenitors expressing mitogen-activated protein kinase (MAPK)-activating mutations that differentiate into senescent myeloid cells that drive lesion formation. Some individuals with LCH subsequently develop progressive and incurable neurodegeneration (LCH-ND). Here, we showed that LCH-ND was caused by myeloid cells that were clonal with peripheral LCH cells. Circulating BRAFV600E+ myeloid cells caused the breakdown of the blood-brain barrier (BBB), enhancing migration into the brain parenchyma where they differentiated into senescent, inflammatory CD11a+ macrophages that accumulated in the brainstem and cerebellum. Blocking MAPK activity and senescence programs reduced peripheral inflammation, brain parenchymal infiltration, neuroinflammation, neuronal damage and improved neurological outcome in preclinical LCH-ND. MAPK activation and senescence programs in circulating myeloid cells represent targetable mechanisms of LCH-ND.
Asunto(s)
Histiocitosis de Células de Langerhans , Proteínas Proto-Oncogénicas B-raf , Humanos , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas B-raf/metabolismo , Histiocitosis de Células de Langerhans/genética , Histiocitosis de Células de Langerhans/patología , Histiocitosis de Células de Langerhans/terapia , Encéfalo/metabolismo , Células Mieloides/metabolismo , Diferenciación CelularRESUMEN
Neurodegenerative diseases (ND) are characterized by progressive loss of neuronal function. Mechanisms of ND pathogenesis are incompletely understood, hampering the development of effective therapies. Langerhans cell histiocytosis (LCH) is an inflammatory neoplastic disorder caused by hematopoietic progenitors expressing MAPK activating mutations that differentiate into senescent myeloid cells that drive lesion formation. Some patients with LCH subsequently develop progressive and incurable neurodegeneration (LCH-ND). Here, we show that LCH-ND is caused by myeloid cells that are clonal with peripheral LCH cells. We discovered that circulating BRAF V600E + myeloid cells cause the breakdown of the blood-brain barrier (BBB), enhancing migration into the brain parenchyma where they differentiate into senescent, inflammatory CD11a + macrophages that accumulate in the brainstem and cerebellum. Blocking MAPK activity and senescence programs reduced parenchymal infiltration, neuroinflammation, neuronal damage and improved neurological outcome in preclinical LCH-ND. MAPK activation and senescence programs in circulating myeloid cells represent novel and targetable mechanisms of ND.
RESUMEN
Sepsis-induced coagulopathy leading to disseminated microvascular thrombosis is associated with high mortality and has no existing therapy. Despite the high prevalence of Gram-positive bacterial sepsis, especially methicillin-resistant Staphylococcus aureus (MRSA), there is a paucity of published Gram-positive pediatric sepsis models. Large animal models replicating sepsis-induced coagulopathy are needed to test new therapeutics before human clinical trials. HYPOTHESIS: Our objective is to develop a pediatric sepsis-induced coagulopathy swine model that last 70 hours. METHODS AND MODELS: Ten 3 weeks old piglets, implanted with telemetry devices for continuous hemodynamic monitoring, were IV injected with MRSA (n = 6) (USA300, Texas Children's Hospital 1516 strain) at 1 × 109 colony forming units/kg or saline (n = 4). Fluid resuscitation was given for heart rate greater than 50% or mean arterial blood pressure less than 30% from baseline. Acetaminophen and dextrose were provided as indicated. Point-of-care complete blood count, prothrombin time (PT), activated thromboplastin time, d-dimer, fibrinogen, and specialized coagulation assays were performed at pre- and post-injection, at 0, 24, 48, 60, and 70 hours. Piglets were euthanized and necropsies performed. RESULTS: Compared with the saline treated piglets (control), the septic piglets within 24 hours had significantly lower neurologic and respiratory scores. Over time, PT, d-dimer, and fibrinogen increased, while platelet counts and activities of factors V, VII, protein C, antithrombin, and a disintegrin and metalloproteinase with thrombospondin-1 motifs (13th member of the family) (ADAMTS-13) decreased significantly in septic piglets compared with control. Histopathologic examination showed minor focal organ injuries including microvascular thrombi and necrosis in the kidney and liver of septic piglets. INTERPRETATIONS AND CONCLUSIONS: We established a 70-hour swine model of MRSA sepsis-induced coagulopathy with signs of consumptive coagulopathy, disseminated microvascular thrombosis, and early organ injuries with histological minor focal organ injuries. This model is clinically relevant to pediatric sepsis and can be used to study dysregulated host immune response and coagulopathy to infection, identify potential early biomarkers, and to test new therapeutics.
RESUMEN
Overall survival after reduced-intensity conditioning (RIC) allogeneic hematopoietic cell transplantation (HCT) using alemtuzumab, fludarabine, and melphalan is associated with high rates of mixed chimerism (MC) and secondary graft failure (GF). We hypothesized that peritransplantation alemtuzumab levels or specific patterns of inflammation would predict these risks. We assessed samples from the Bone Marrow Transplant Clinical Trials Network 1204 (NCT01998633) to study the impact of alemtuzumab levels and cytokine patterns on MC and impending or established secondary GF (defined as donor chimerism <5% after initial engraftment and/or requirement of cellular intervention). Thirty-three patients with hemophagocytic lymphohistiocytosis (n = 25) and other IEIs (n = 8) who underwent HCTs with T-cell-replete grafts were included. Patients with day 0 alemtuzumab levels ≤0.32 µg/mL had a markedly lower incidence of MC, 14.3%, vs 90.9% in patients with levels >0.32 µg/mL (P = .008). Impending or established secondary GF was only observed in patients with day 0 alemtuzumab levels >0.32 µg/mL (P = .08). Unexpectedly, patients with impending or established secondary GF had lower CXCL9 levels. The cumulative incidence of impending or established secondary GF in patients with a day 14+ CXCL9 level ≤2394 pg/mL (day 14+ median) was 73.6% vs 0% in patients with a level >2394 pg/mL (P = .002). CXCL9 levels inversely correlated with alemtuzumab levels. These data suggest a model in which higher levels of alemtuzumab at day 0 deplete donor T cells, inhibit the graft-versus-marrow reaction (thereby suppressing CXCL9 levels), and adversely affect sustained engraftment in the nonmyeloablative HCT setting. This trial was registered at www.clinicaltrials.gov as #NCT01998633.
Asunto(s)
Anticuerpos Monoclonales Humanizados , Trasplante de Células Madre Hematopoyéticas , Humanos , Alemtuzumab/uso terapéutico , Anticuerpos Monoclonales Humanizados/uso terapéutico , Melfalán/uso terapéutico , Trasplante de Células Madre Hematopoyéticas/efectos adversos , Donantes de Tejidos , Quimiocina CXCL9RESUMEN
Langerhans cell histiocytosis (LCH) is a rare inflammatory myeloid neoplasia with a broad spectrum of clinical manifestations. The activation of the MAP kinase pathway plays an integral role in its pathogenesis with genetic alterations found in the majority of cases that most frequently involve a somatic mutation of the oncogenic BRAFV600E variant. In this study we investigated the prevalence of the BRAFV600E mutation and its clinical relevance in adult Greek patients with LCH. Among 37 patients studied, the BRAFV600E mutation was identified in 12 out of 31 (38.7%), whereas in six patients (19.3%) the results were in conclusive. The presence of the mutation did not correlate with age at diagnosis, organ involvement, disease extent, response to initial treatment, development of diabetes insipidus and relapse risk. In our series the prevalence of the BRAFV600E mutation is at the lower range of the relative percentage found in children, but in line to that obtained in previous studies of adult patients with LCH that have found an up to 50% prevalence of the BRAFV600E mutation in these patients. Further studies with a larger number of adults are needed to identify the exact prevalence of mutations in the RAS-RAF-MEK-ERK pathway and their role on clinical parameters and disease outcomes.Supplemental data for this article is available online at https://doi.org/10.1080/08880018.2022.2029988 .
Asunto(s)
Histiocitosis de Células de Langerhans , Proteínas Proto-Oncogénicas B-raf , Adulto , Niño , Grecia/epidemiología , Histiocitosis de Células de Langerhans/epidemiología , Histiocitosis de Células de Langerhans/genética , Humanos , Mutación , Prevalencia , Proteínas Proto-Oncogénicas B-raf/genéticaRESUMEN
Hemophagocytic lymphohistiocytosis (HLH) is a syndrome characterized by pathologic immune activation in which prompt recognition and initiation of immune suppression is essential for survival. Children with HLH have many overlapping clinical features with critically ill children with sepsis and systemic inflammatory response syndrome (SIRS) in whom alternative therapies are indicated. To determine whether plasma biomarkers could differentiate HLH from other inflammatory conditions and to better define a core inflammatory signature of HLH, concentrations of inflammatory plasma proteins were compared in 40 patients with HLH to 47 pediatric patients with severe sepsis or SIRS. Fifteen of 135 analytes were significantly different in HLH plasma compared with SIRS/sepsis, including increased interferon-γ (IFN-γ)-regulated chemokines CXCL9, CXCL10, and CXCL11. Furthermore, a 2-analyte plasma protein classifier including CXCL9 and interleukin-6 was able to differentiate HLH from SIRS/sepsis. Gene expression in CD8+ T cells and activated monocytes from blood were also enriched for IFN-γ pathway signatures in peripheral blood cells from patients with HLH compared with SIRS/sepsis. This study identifies differential expression of inflammatory proteins as a diagnostic strategy to identify critically ill children with HLH, and comprehensive unbiased analysis of inflammatory plasma proteins and global gene expression demonstrates that IFN-γ signaling is uniquely elevated in HLH. In addition to demonstrating the ability of diagnostic criteria for HLH and sepsis or SIRS to identify groups with distinct inflammatory patterns, results from this study support the potential for prospective evaluation of inflammatory biomarkers to aid in diagnosis of and optimizing therapeutic strategies for children with distinctive hyperinflammatory syndromes.
Asunto(s)
Linfohistiocitosis Hemofagocítica , Sepsis , Niño , Diagnóstico Diferencial , Humanos , Interferón gamma , Linfohistiocitosis Hemofagocítica/diagnóstico , Proteoma , Sepsis/diagnóstico , Síndrome de Respuesta Inflamatoria Sistémica/diagnósticoRESUMEN
Langerhans cell histiocytosis (LCH) is an inflammatory myeloid neoplasia characterized by granulomatous lesions containing pathological CD207+ dendritic cells (DCs) with persistent MAPK pathway activation. Standard-of-care chemotherapies are inadequate for most patients with multisystem disease, and optimal strategies for relapsed and refractory disease are not defined. The mechanisms underlying development of inflammation in LCH lesions, the role of inflammation in pathogenesis, and the potential for immunotherapy are unknown. Analysis of the immune infiltrate in LCH lesions identified the most prominent immune cells as T lymphocytes. Both CD8+ and CD4+ T cells exhibited "exhausted" phenotypes with high expression of the immune checkpoint receptors. LCH DCs showed robust expression of ligands to checkpoint receptors. Intralesional CD8+ T cells showed blunted expression of Tc1/Tc2 cytokines and impaired effector function. In contrast, intralesional regulatory T cells demonstrated intact suppressive activity. Treatment of BRAFV600ECD11c LCH mice with anti-PD-1 or MAPK inhibitor reduced lesion size, but with distinct responses. Whereas MAPK inhibitor treatment resulted in reduction of the myeloid compartment, anti-PD-1 treatment was associated with reduction in the lymphoid compartment. Notably, combined treatment with MAPK inhibitor and anti-PD-1 significantly decreased both CD8+ T cells and myeloid LCH cells in a synergistic fashion. These results are consistent with a model that MAPK hyperactivation in myeloid LCH cells drives recruitment of functionally exhausted T cells within the LCH microenvironment, and they highlight combined MAPK and checkpoint inhibition as a potential therapeutic strategy.
Asunto(s)
Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD8-positivos/efectos de los fármacos , Histiocitosis de Células de Langerhans/tratamiento farmacológico , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Inhibidores de Proteínas Quinasas/uso terapéutico , Animales , Linfocitos T CD4-Positivos/patología , Linfocitos T CD8-positivos/patología , Modelos Animales de Enfermedad , Sinergismo Farmacológico , Histiocitosis de Células de Langerhans/patología , Humanos , Ratones Endogámicos C57BL , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidoresRESUMEN
Hodgkin lymphoma (HL) histopathology is characterized by rare malignant Reed-Sternberg cells among an inflammatory infiltrate. We hypothesized that characteristics of inflammation in pediatric HL lesions would be reflected by the levels of inflammatory cytokines or chemokines in pre-therapy plasma of children with HL. The study objectives were to better define the inflammatory pre-therapy plasma proteome and identify plasma biomarkers associated with extent of disease and clinical outcomes in pediatric HL. Pre-therapy plasma samples were obtained from pediatric subjects with newly diagnosed HL and healthy pediatric controls. Plasma concentrations of 135 cytokines/chemokines were measured with the Luminex platform. Associations between protein concentration and disease characteristics were determined using multivariate permutation tests with false discovery control. Fifty-six subjects with HL (mean age: 13 years, range 3-18) and 47 controls were analyzed. The cytokine/chemokine profiles of subjects with HL were distinct from controls, and unique cytokines/chemokines were associated with high-risk disease (IL-10, TNF-α, IFN-γ, IL-8) and slow early response (CCL13, IFN-λ1, IL-8). TNFSF10 was significantly elevated among those who ultimately relapsed and was significantly associated with worse event-free survival. These biomarkers could be incorporated into biologically based risk stratification to optimize outcomes and minimize toxicities in pediatric HL.
RESUMEN
Cytokine storm syndromes (CSS) are severe hyperinflammatory conditions characterized by excessive immune system activation leading to organ damage and death. Hemophagocytic lymphohistiocytosis (HLH), a disease often associated with inherited defects in cell-mediated cytotoxicity, serves as a prototypical CSS for which the 5-year survival is only 60%. Frontline therapy for HLH consists of the glucocorticoid dexamethasone (DEX) and the chemotherapeutic agent etoposide. Many patients, however, are refractory to this treatment or relapse after an initial response. Notably, many cytokines that are elevated in HLH activate the JAK/STAT pathway, and the JAK1/2 inhibitor ruxolitinib (RUX) has shown efficacy in murine HLH models and humans with refractory disease. We recently reported that cytokine-induced JAK/STAT signaling mediates DEX resistance in T cell acute lymphoblastic leukemia (T-ALL) cells, and that this could be effectively reversed by RUX. On the basis of these findings, we hypothesized that cytokine-mediated JAK/STAT signaling might similarly contribute to DEX resistance in HLH, and that RUX treatment would overcome this phenomenon. Using ex vivo assays, a murine model of HLH, and primary patient samples, we demonstrate that the hypercytokinemia of HLH reduces the apoptotic potential of CD8 T cells leading to relative DEX resistance. Upon exposure to RUX, this apoptotic potential is restored, thereby sensitizing CD8 T cells to DEX-induced apoptosis in vitro and significantly reducing tissue immunopathology and HLH disease manifestations in vivo. Our findings provide rationale for combining DEX and RUX to enhance the lymphotoxic effects of DEX and thus improve the outcomes for patients with HLH and related CSS.
Asunto(s)
Apoptosis/efectos de los fármacos , Linfocitos T CD8-positivos/efectos de los fármacos , Síndrome de Liberación de Citoquinas/tratamiento farmacológico , Dexametasona/uso terapéutico , Inhibidores de las Cinasas Janus/uso terapéutico , Linfohistiocitosis Hemofagocítica/tratamiento farmacológico , Pirazoles/uso terapéutico , Transducción de Señal/efectos de los fármacos , Animales , Linfocitos T CD8-positivos/inmunología , Síndrome de Liberación de Citoquinas/etiología , Síndrome de Liberación de Citoquinas/fisiopatología , Citocinas/fisiología , Dexametasona/administración & dosificación , Dexametasona/farmacología , Modelos Animales de Enfermedad , Resistencia a Medicamentos/efectos de los fármacos , Quimioterapia Combinada , Humanos , Interleucina-2/farmacología , Inhibidores de las Cinasas Janus/administración & dosificación , Inhibidores de las Cinasas Janus/farmacología , Quinasas Janus , Coriomeningitis Linfocítica/complicaciones , Coriomeningitis Linfocítica/fisiopatología , Linfohistiocitosis Hemofagocítica/complicaciones , Linfohistiocitosis Hemofagocítica/enzimología , Linfohistiocitosis Hemofagocítica/inmunología , Ratones , Ratones Endogámicos C57BL , Nitrilos , Perforina/deficiencia , Pirazoles/administración & dosificación , Pirazoles/farmacología , Pirimidinas , Factor de Transcripción STAT5/fisiología , Organismos Libres de Patógenos EspecíficosRESUMEN
Langerhans cell histiocytosis (LCH) is a myeloproliferative disorder that is characterized by the inflammatory lesions with pathogenic CD1a+CD207+ dendritic cells (DCs). BRAFV600E and other somatic activating MAPK gene mutations have been identified in differentiating bone marrow and blood myeloid cells, but the origin of the LCH lesion CD1a+CD207+ DCs and mechanisms of lesion formation remain incompletely defined. To identify candidate LCH CD1a+CD207+ DC precursor populations, gene-expression profiles of LCH lesion CD1a+CD207+ DCs were first compared with established gene signatures from human myeloid cell subpopulations. Interestingly, the CD1c+ myeloid DC (mDC) gene signature was most enriched in the LCH CD1a+CD207+ DC transcriptome. Additionally, the BRAFV600E allele was not only localized to CD1a+CD207- DCs and CD1a+CD207+ DCs, but it was also identified in CD1c+ mDCs in LCH lesions. Transcriptomes of CD1a+CD207- DCs were nearly indistinguishable from CD1a+CD207+ DCs (both CD1a+CD207low and CD1a+CD207high subpopulations). Transcription profiles of LCH lesion CD1a+CD207+ DCs and peripheral blood CD1c+ mDCs from healthy donors were compared to identify potential LCH DC-specific biomarkers: HLA-DQB2 expression was significantly increased in LCH lesion CD1a+CD207+ DCs compared with circulating CD1c+ mDCs from healthy donors. HLA-DQB2 antigen was identified on LCH lesion CD1a+CD207- DCs and CD1a+CD207+ DCs as well as on CD1c+(CD1a+CD207-) mDCs, but it was not identified in any other lesion myeloid subpopulations. HLA-DQB2 expression was specific to peripheral blood of patients with BRAFV600E+ peripheral blood mononuclear cells, and HLA-DQB2+CD1c+ blood cells were highly enriched for the BRAFV600E in these patients. These data support a model in which blood CD1c+HLA-DQB2+ mDCs with activated ERK migrate to lesion sites where they differentiate into pathogenic CD1a+CD207+ DCs.
Asunto(s)
Histiocitosis de Células de Langerhans , Leucocitos Mononucleares , Antígenos CD/genética , Antígenos CD1/genética , Biomarcadores , Células Dendríticas , Glicoproteínas , Histiocitosis de Células de Langerhans/diagnóstico , Histiocitosis de Células de Langerhans/genética , Humanos , Lectinas Tipo C/genética , Lectinas de Unión a Manosa/genética , Células MieloidesRESUMEN
BACKGROUND: Central nervous system Langerhans cell histiocytosis (CNS-LCH) brain involvement may include mass lesions and/or a neurodegenerative disease (LCH-ND) of unknown etiology. The goal of this study was to define the mechanisms of pathogenesis that drive CNS-LCH. METHODS: Cerebrospinal fluid (CSF) biomarkers including CSF proteins and extracellular BRAFV600E DNA were analyzed in CSF from patients with CNS-LCH lesions compared with patients with brain tumors and other neurodegenerative conditions. Additionally, the presence of BRAFV600E was tested in peripheral mononuclear blood cells (PBMCs) as well as brain biopsies from LCH-ND patients, and the response to BRAF-V600E inhibitor was evaluated in 4 patients with progressive disease. RESULTS: Osteopontin was the only consistently elevated CSF protein in patients with CNS-LCH compared with patients with other brain pathologies. BRAFV600E DNA was detected in CSF of only 2/20 (10%) cases, both with LCH-ND and active lesions outside the CNS. However, BRAFV600E+ PBMCs were detected with significantly higher frequency at all stages of therapy in LCH patients who developed LCH-ND. Brain biopsies of patients with LCH-ND demonstrated diffuse perivascular infiltration by BRAFV600E+ cells with monocyte phenotype (CD14+ CD33+ CD163+ P2RY12- ) and associated osteopontin expression. Three of 4 patients with LCH-ND treated with BRAF-V600E inhibitor experienced significant clinical and radiologic improvement. CONCLUSION: In LCH-ND patients, BRAFV600E+ cells in PBMCs and infiltrating myeloid/monocytic cells in the brain is consistent with LCH-ND as an active demyelinating process arising from a mutated hematopoietic precursor from which LCH lesion CD207+ cells are also derived. Therapy directed against myeloid precursors with activated MAPK signaling may be effective for LCH-ND. Cancer 2018;124:2607-20. © 2018 American Cancer Society.
Asunto(s)
Neoplasias Encefálicas/diagnóstico , Histiocitosis de Células de Langerhans/diagnóstico , Enfermedades Neurodegenerativas/diagnóstico , Osteopontina/líquido cefalorraquídeo , Proteínas Proto-Oncogénicas B-raf/genética , Adolescente , Adulto , Biomarcadores/sangre , Biomarcadores/líquido cefalorraquídeo , Biopsia , Encéfalo/patología , Neoplasias Encefálicas/líquido cefalorraquídeo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Niño , Preescolar , Diagnóstico Diferencial , Femenino , Células Madre Hematopoyéticas/patología , Histiocitosis de Células de Langerhans/líquido cefalorraquídeo , Histiocitosis de Células de Langerhans/genética , Histiocitosis de Células de Langerhans/patología , Humanos , Lactante , Recién Nacido , Leucocitos Mononucleares/patología , Sistema de Señalización de MAP Quinasas , Masculino , Enfermedades Neurodegenerativas/líquido cefalorraquídeo , Enfermedades Neurodegenerativas/genética , Enfermedades Neurodegenerativas/patología , Proteínas Proto-Oncogénicas B-raf/antagonistas & inhibidores , Estudios Retrospectivos , Adulto JovenRESUMEN
Langerhans cell histiocytosis (LCH) is characterized by inflammatory lesions containing pathologic CD207+ dendritic cells with constitutively activated ERK. Mutually exclusive somatic mutations in MAPK pathway genes have been identified in â¼75% of LCH cases, including recurrent BRAF-V600E and MAP2K1 mutations. To elucidate mechanisms of ERK activation in the remaining 25% of patients, we performed whole-exome sequencing (WES, n = 6), targeted BRAF sequencing (n = 19), and/or whole-transcriptome sequencing (RNA-seq, n = 6) on 24 LCH patient samples lacking BRAF-V600E or MAP2K1 mutations. WES and BRAF sequencing identified in-frame BRAF deletions in the ß3-αC loop in 6 lesions. RNA-seq revealed one case with an in-frame FAM73A-BRAF fusion lacking the BRAF autoinhibitory regulatory domain but retaining an intact kinase domain. High levels of phospho-ERK were detected in vitro in cells overexpressing either BRAF fusion or deletion constructs and ex vivo in CD207+ cells from lesions. ERK activation was resistant to BRAF-V600E inhibition, but responsive to both a second-generation BRAF inhibitor and a MEK inhibitor. These results support an emerging model of universal ERK-activating genetic alterations driving pathogenesis in LCH. A personalized approach in which patient-specific alterations are identified may be necessary to maximize benefit from targeted therapies for patients with LCH.
Asunto(s)
Histiocitosis de Células de Langerhans/genética , Mutación , Proteínas de Fusión Oncogénica/genética , Proteínas Proto-Oncogénicas B-raf/genética , Adolescente , Adulto , Anciano , Niño , Preescolar , Activación Enzimática/genética , Femenino , Histiocitosis de Células de Langerhans/enzimología , Humanos , Lactante , Masculino , Proteínas de Fusión Oncogénica/metabolismo , Dominios Proteicos , Proteínas Proto-Oncogénicas B-raf/metabolismoRESUMEN
Accurate and high-throughput technologies are needed for identification of new therapeutic targets and for optimizing therapy in inflammatory bowel disease. Our aim was to assess multi-analyte protein-based assays of cytokines/chemokines using Luminex technology. We have reported that Luminex-based profiling was useful in assessing response to L-arginine therapy in the mouse model of dextran sulfate sodium colitis. Therefore, we studied prospectively collected samples from ulcerative colitis (UC) patients and control subjects. Serum, colon biopsies, and clinical information were obtained from subjects undergoing colonoscopy for evaluation of UC or for non-UC indications. In total, 38 normal controls and 137 UC cases completed the study. Histologic disease severity and the Mayo Disease Activity Index (DAI) were assessed. Serum and colonic tissue cytokine/chemokine profiles were measured by Luminex-based multiplex testing of 42 analytes. Only eotaxin-1 and G-CSF were increased in serum of patients with histologically active UC vs. controls. While 13 cytokines/chemokines were increased in active UC vs. controls in tissues, only eotaxin-1 was increased in all levels of active disease in both serum and tissue. In tissues, eotaxin-1 correlated with the DAI and with eosinophil counts. Increased eotaxin-1 levels were confirmed by real-time PCR. Tissue eotaxin-1 levels were also increased in experimental murine colitis induced by dextran sulfate sodium, oxazolone, or Citrobacter rodentium, but not in murine Helicobacter pylori infection. Our data implicate eotaxin-1 as an etiologic factor and therapeutic target in UC, and indicate that Luminex-based assays may be useful to assess IBD pathogenesis and to select patients for anti-cytokine/chemokine therapies.
Asunto(s)
Quimiocina CCL11/metabolismo , Colitis Ulcerosa/metabolismo , Adulto , Animales , Colitis Ulcerosa/sangre , Colitis Ulcerosa/inducido químicamente , Femenino , Gastritis/sangre , Gastritis/metabolismo , Gastritis/microbiología , Helicobacter pylori/patogenicidad , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Oxazolona/toxicidad , Estudios ProspectivosRESUMEN
L-Arginine (L-Arg) is a semiessential amino acid that has altered availability in human ulcerative colitis (UC), a form of inflammatory bowel disease, and is beneficial in murine colitis induced by dextran sulfate sodium (DSS), a model with similarity to UC. We assessed the role of cationic amino acid transporter 2 (CAT2), the inducible transporter of L-Arg, in DSS colitis. Expression of CAT2 was upregulated in tissues from colitic mice and localized predominantly to colonic macrophages. CAT2-deficient (CAT2-/-) mice exposed to DSS exhibited worsening of survival, body weight loss, colon weight, and histological injury. These effects were associated with increased serum L-Arg and decreased tissue L-Arg uptake and inducible nitric oxide synthase protein expression. Clinical benefits of L-Arg supplementation in wild-type mice were lost in CAT2-/- mice. There was increased infiltration of macrophages, dendritic cells, granulocytes, and T cells in colitic CAT2-/- compared with wild-type mice. Cytokine profiling revealed increases in proinflammatory granulocyte colony-stimulating factor, macrophage inflammatory protein-1α, IL-15, and regulated and normal T cell-expressed and -secreted and a shift from an IFN-γ- to an IL-17-predominant T cell response, as well as an increase in IL-13, in tissues from colitic CAT2-/- mice. However, there were no increases in other T helper cell type 2 cytokines, nor was there a global increase in macrophage-derived proinflammatory cytokines. The increase in IL-17 derived from both CD4 and γδ T cells and was associated with colonic IL-6 expression. Thus CAT2 plays an important role in controlling inflammation and IL-17 activation in an injury model of colitis, and impaired L-Arg availability may contribute to UC pathogenesis.
Asunto(s)
Transportador de Aminoácidos Catiônicos 2/deficiencia , Colitis/inducido químicamente , Colitis/inmunología , Sulfato de Dextran , Interleucina-17/metabolismo , Linfocitos T/inmunología , Animales , Arginina/metabolismo , Transportador de Aminoácidos Catiônicos 2/genética , Transportador de Aminoácidos Catiônicos 2/fisiología , Colitis/fisiopatología , Interleucina-17/genética , Interleucina-23/genética , Interleucina-6/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , ARN Mensajero/análisis , Regulación hacia ArribaRESUMEN
Gastric cancer is the second leading cause of cancer mortality worldwide and is projected to rise to tenth in all-cause mortality in the near term. Early detection requires improved sensitivity and specificity of endoscopic imaging with novel methods. The objective of this study was to evaluate the utility of activatable molecular probes for the detection of gastric cancer both in vivo and ex vivo in a preclinical model. Smad4âº/â» mice, which develop spontaneous gastric neoplasia, were compared to normal wild-type controls. Cathepsin-activatable and matrix metalloproteinase (MMP)-activatable molecular probes were injected 24 hours and 6 hours before imaging, respectively. In vivo imaging was performed using quantitative tomographic near-infrared fluorescence (NIRF) imaging. For validation, ex vivo imaging and histologic examination were performed. Molecular imaging in vivo of Smad4âº/â» gastric cancer murine models revealed intense activation of both cathepsin B and MMP probes. Ex vivo imaging and histology confirmed that the detected neoplasms were adenocarcinomas and hyperplastic lesions. This study provides proof of principle that the cathepsin- and MMP-activatable molecular probes are activated in the Smad4âº/â» murine model of spontaneous gastric adenocarcinoma and can be imaged by both in vivo and ex vivo NIRF methods. The cathepsin probe also detects hyperplastic lesions.