Publications by authors named "Rikhia Chakraborty"

22 Publications

  • Page 1 of 1

Defining the Inflammatory Plasma Proteome in Pediatric Hodgkin Lymphoma.

Cancers (Basel) 2020 Dec 2;12(12). Epub 2020 Dec 2.

Department of Pediatrics, Baylor College of Medicine, Texas Children's Cancer and Hematology Centers, Houston, TX 77030, USA.

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.
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http://dx.doi.org/10.3390/cancers12123603DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7761312PMC
December 2020

Overcoming T cell exhaustion in LCH: PD-1 blockade and targeted MAPK inhibition are synergistic in a mouse model of LCH.

Blood 2020 Oct 19. Epub 2020 Oct 19.

1Texas Children's Cancer Center, Texas Children's Hospital, United States.

Langerhans cell histiocytosis (LCH) is an inflammatory myeloid neoplasia characterized by granulomatous lesions containing pathological CD207+ dendritic cells (DCs) with persistent mitogen-activated protein kinase (MAPK) pathway activation. Standard of care chemotherapy strategies is 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 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. Intra-lesional CD8+ T cells showed blunted expression of Tc1/Tc2 cytokines and impaired effector function. In contrast, intra-lesional regulatory T cells (Tregs) 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 cell 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 highlight combined MAPK and checkpoint inhibition as a potential therapeutic strategy.
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http://dx.doi.org/10.1182/blood.2020005867DOI Listing
October 2020

MAP-Kinase-Driven Hematopoietic Neoplasms: A Decade of Progress in the Molecular Age.

Cold Spring Harb Perspect Med 2020 Jun 29. Epub 2020 Jun 29.

Human Oncology and Pathogenesis Program, Department of Medicine, Leukemia Service, Department of Medicine, Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, New York 10065, USA.

Mutations in members of the mitogen-activated protein kinase (MAPK) pathway are extensively studied in epithelial malignancies, with mutations being one of the most common alterations activating this pathway. However, mutations are overall quite rare in hematological malignancies. Studies over the past decade have identified high-frequency , , and other kinase alterations in two groups of MAPK-driven hematopoietic neoplasms: hairy cell leukemia (HCL) and the systemic histiocytoses. Despite HCL and histiocytoses sharing common molecular alterations, these are phenotypically distinct malignancies that differ in respect to clinical presentation and suspected cell of origin. The purpose of this review is to highlight the molecular advancements over the last decade in the histiocytic neoplasms and HCL and discuss the impact these insights have had on our understanding of the molecular pathophysiology, cellular origins, and therapy of these enigmatic diseases as well as perspectives for future research directions.
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http://dx.doi.org/10.1101/cshperspect.a034892DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7770072PMC
June 2020

Circulating CD1c+ myeloid dendritic cells are potential precursors to LCH lesion CD1a+CD207+ cells.

Blood Adv 2020 01;4(1):87-99

Texas Children's Cancer Center, Texas Children's Hospital, Houston, TX.

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.
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http://dx.doi.org/10.1182/bloodadvances.2019000488DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6960472PMC
January 2020

CNS Langerhans cell histiocytosis: Common hematopoietic origin for LCH-associated neurodegeneration and mass lesions.

Cancer 2018 06 6;124(12):2607-2620. Epub 2018 Apr 6.

Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, Texas.

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.
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http://dx.doi.org/10.1002/cncr.31348DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6289302PMC
June 2018

RAF/MEK/extracellular signal-related kinase pathway suppresses dendritic cell migration and traps dendritic cells in Langerhans cell histiocytosis lesions.

J Exp Med 2018 01 20;215(1):319-336. Epub 2017 Dec 20.

Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY

Langerhans cell histiocytosis (LCH) is an inflammatory myeloid neoplasia characterized by granulomatous lesions containing pathological CD207 dendritic cells (DCs) with constitutively activated mitogen-activated protein kinase (MAPK) pathway signaling. Approximately 60% of LCH patients harbor somatic V600E mutations localizing to CD207 DCs within lesions. However, the mechanisms driving V600E LCH cell accumulation in lesions remain unknown. Here we show that sustained extracellular signal-related kinase activity induced by V600E inhibits C-C motif chemokine receptor 7 (CCR7)-mediated DC migration, trapping DCs in tissue lesions. Additionally, V600E increases expression of BCL2-like protein 1 (BCL2L1) in DCs, resulting in resistance to apoptosis. Pharmacological MAPK inhibition restores migration and apoptosis potential in a mouse LCH model, as well as in primary human LCH cells. We also demonstrate that MEK inhibitor-loaded nanoparticles have the capacity to concentrate drug delivery to phagocytic cells, significantly reducing off-target toxicity. Collectively, our results indicate that MAPK tightly suppresses DC migration and augments DC survival, rendering DCs in LCH lesions trapped and resistant to cell death.
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http://dx.doi.org/10.1084/jem.20161881DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5748846PMC
January 2018

p53 Nongenotoxic Activation and mTORC1 Inhibition Lead to Effective Combination for Neuroblastoma Therapy.

Clin Cancer Res 2017 Nov 18;23(21):6629-6639. Epub 2017 Aug 18.

Department of Pediatrics, Section of Hematology-Oncology, Texas Children's Cancer and Hematology Centers, Baylor College of Medicine, Houston, Texas.

mTORC1 inhibitors are promising agents for neuroblastoma therapy; however, they have shown limited clinical activity as monotherapy, thus rational drug combinations need to be explored to improve efficacy. Importantly, neuroblastoma maintains both an active p53 and an aberrant mTOR signaling. Using an orthotopic xenograft model and modulating p53 levels, we investigated the antitumor effects of the mTORC1 inhibitor temsirolimus in neuroblastoma expressing normal, decreased, or mutant p53, both as single agent and in combination with first- and second-generation MDM2 inhibitors to reactivate p53. Nongenotoxic p53 activation suppresses mTOR activity. Moreover, p53 reactivation via RG7388, a second-generation MDM2 inhibitor, strongly enhances the antitumor activity of temsirolimus. Single-agent temsirolimus does not elicit apoptosis, and tumors rapidly regrow after treatment suspension. In contrast, our combination therapy triggers a potent apoptotic response in wild-type p53 xenografts and efficiently blocks tumor regrowth after treatment completion. We also found that this combination uniquely led to p53-dependent suppression of survivin whose ectopic expression is sufficient to rescue the apoptosis induced by our combination. Our study supports a novel highly effective strategy that combines RG7388 and temsirolimus in wild-type p53 neuroblastoma, which warrants testing in early-phase clinical trials. .
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http://dx.doi.org/10.1158/1078-0432.CCR-17-0668DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5959272PMC
November 2017

New somatic BRAF splicing mutation in Langerhans cell histiocytosis.

Mol Cancer 2017 07 6;16(1):115. Epub 2017 Jul 6.

EA4340, Versailles SQY University, Paris-Saclay University, Boulogne, France.

Langerhans cell histiocytosis (LCH) is an inflammatory myeloid neoplasia with constitutive activation of the MAPKinase RAS-RAF-MEK-ERK cell signaling pathway. We analyzed 9 LCH cases without BRAF and MAP2K1 mutations by whole exome sequencing. We identified a new somatic BRAF splicing mutation in 2 cases. Both cases were childhood single system (SS) LCH cases, with self-healing outcome of the bone lesions. This mutant consisted in a 9 base pair duplication (c.1511_1517 + 2 duplication), encoding for a predicted mutant protein with insertion of 3 amino acids (p.Arg506_Lys507insLeuLeuArg) in the N-terminal lobe of the kinase domain of BRAF. Transient expression of the c.1511_1517 + 2dup BRAF mutant in HEK293 cells enhanced MAPKinase pathway activation, and was not inhibited by vemurafenib but was inhibited by PLX8394, a second-generation BRAF inhibitor able to inhibit signaling of BRAF monomers and dimers. Future LCH molecular screening panel should include this new mutation to better define its prevalence in LCH and its restriction to autoregressive bone SS LCH.
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http://dx.doi.org/10.1186/s12943-017-0690-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5498996PMC
July 2017

Activating MAPK1 (ERK2) mutation in an aggressive case of disseminated juvenile xanthogranuloma.

Oncotarget 2017 Jul;8(28):46065-46070

Texas Children's Cancer Center, Texas Children's Hospital, Houston, TX 77030, USA.

Juvenile xanthogranuloma (JXG) is a rare histiocytic disorder that is usually benign and self-limiting. We present a case of atypical, aggressive JXG harboring a novel mitogen-activated protein kinase (MAPK) pathway mutation in the MAPK1 gene, which encodes mitogen-activated protein kinase 1 or extracellular signal-regulated 2 (ERK2). Our analysis revealed that the mutation results in constitutive ERK activation that is resistant to BRAF or MEK inhibitors but susceptible to an ERK inhibitor. These data highlight the importance of identifying specific MAPK pathway alterations as part of the diagnostic workup for patients with histiocytic disorders rather than initiating empiric treatment with MEK inhibitors.
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http://dx.doi.org/10.18632/oncotarget.17521DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5542249PMC
July 2017

Evaluation of maternal and perinatal characteristics on childhood lymphoma risk: A population-based case-control study.

Pediatr Blood Cancer 2017 05 28;64(5). Epub 2016 Nov 28.

Department of Pediatrics, Section of Hematology-Oncology, Baylor College of Medicine, Houston, Texas.

Background: Lymphoma is one of the most common pediatric malignancies; however, there are few well-established risk factors. Therefore, we investigated if maternal and perinatal characteristics influenced the risk of childhood lymphoma.

Procedure: Information on cases (n = 374) diagnosed with lymphoma and born in Texas for the period 1995-2011 was obtained from the Texas Cancer Registry. Birth certificate controls were randomly selected at a ratio of 10 controls per 1 case for the same period of 1995-2011. Unconditional logistic regression was used to generate unadjusted (OR) and adjusted odds ratios (aOR) and 95% confidence intervals (CI) for the following histologic subtypes: Hodgkin (HL), Burkitt (BL), and non-BL non-HLs (non-BL NHLs).

Results: Overall, our findings indicate specific maternal and perinatal characteristics influence childhood lymphoma risk. Mexico-born mothers were more likely to have offspring who developed BL compared to mothers born in the United States (U.S.; aOR: 2.15; 95% CI: 1.06-4.36). Further, mothers who resided at time of delivery in a county on the U.S.-Mexico border were more likely to give birth to offspring who developed non-BL NHL (aOR: 1.72; 95% CI: 1.11-2.67) compared to mothers not living on the U.S.-Mexico border at time of infant birth. Last, infants born large-for-gestational-age experienced a twofold increase in BL risk (aOR: 2.00; 95% CI: 1.10-3.65).

Conclusions: In this population-based assessment, we confirmed previously reported risk predictors of childhood lymphoma, including sex of infant, while highlighting novel risk factors that warrant assessment in future studies.
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http://dx.doi.org/10.1002/pbc.26321DOI Listing
May 2017

Alternative genetic mechanisms of BRAF activation in Langerhans cell histiocytosis.

Blood 2016 11 11;128(21):2533-2537. Epub 2016 Oct 11.

Texas Children's Cancer Center, Texas Children's Hospital, Houston, TX.

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 ∼75% 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 β3-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.
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http://dx.doi.org/10.1182/blood-2016-08-733790DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5123197PMC
November 2016

Langerhans Cell Histiocytosis: Emerging Insights and Clinical Implications.

Oncology (Williston Park) 2016 Feb;30(2):122-32, 139

Langerhans cell histiocytosis is a disorder characterized by lesions that include CD207+ dendritic cells along with an inflammatory infiltrate. Langerhans cell histiocytosis has a highly variable clinical presentation, ranging from a single lesion to potentially fatal disseminated disease. The uncertainty as to whether Langerhans cell histiocytosis is a reactive or a neoplastic disease has resulted in a long-standing debate on this question, and the limited understanding of the pathogenesis of the disease has impeded clinical improvement for patients. The current standard of care for multisystem Langerhans cell histiocytosis, empirically derived chemotherapy with vinblastine and prednisone, cures fewer than 50% of patients, and optimal therapies for relapse and neurodegenerative disease remain uncertain. Recent research advances support a model in which Langerhans cell histiocytosis arises due to pathologic activation of the mitogen-activated protein kinase (MAPK) pathway in myeloid precursors. Redefinition of Langerhans cell histiocytosis as a myeloid neoplastic disorder driven by hyperactive ERK supports the potential of chemotherapy with efficacy against immature myeloid cells, as well as mutation-specific targeted therapy.
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February 2016

Differentiating skin-limited and multisystem Langerhans cell histiocytosis.

J Pediatr 2014 Nov 21;165(5):990-6. Epub 2014 Oct 21.

Texas Children's Cancer Center, Baylor College of Medicine, Houston, TX.

Objective: To identify features associated with multisystem involvement and therapeutic failure in patients with skin Langerhans cell histiocytosis (LCH).

Study Design: We reviewed medical records of 71 consecutive patients with LCH with skin involvement evaluated at Texas Children's Hospital and analyzed clinical features, laboratory results, and the presence of circulating cells with the BRAF-V600E mutation with respect to initial staging and clinical outcomes.

Results: Skin disease in patients older than 18 months of age at diagnosis was associated with the presence of multisystem disease (OR, 9.65; 95% CI, 1.17-79.4). Forty percent of patients referred for presumed skin-limited LCH had underlying multisystem involvement, one-half of these with risk-organ involvement. Patients with skin-limited LCH had a 3-year progression-free survival of 89% after initial therapy, and none developed multisystem disease. Patients with skin/multisystem involvement had a 3-year progression-free survival of 44% with vinblastine/prednisone therapy, and risk-organ involvement did not correlate with failure to achieve nonactive disease. Circulating cells with BRAF-V600E were detected at higher frequency in patients with multisystem involvement (8 of 11 skin/multisystem vs 1 of 13 skin-limited; P = .002).

Conclusion: Skin-limited LCH necessitates infrequent therapeutic intervention and has a lower risk of progression relative to skin plus multisystem LCH. The less-aggressive clinical course and lack of circulating cells with the BRAF-V600E mutation in skin-limited LCH suggest a different mechanism of disease origin compared with multisystem or risk-organ disease.
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http://dx.doi.org/10.1016/j.jpeds.2014.07.063DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4254414PMC
November 2014

Mutually exclusive recurrent somatic mutations in MAP2K1 and BRAF support a central role for ERK activation in LCH pathogenesis.

Blood 2014 Nov 8;124(19):3007-15. Epub 2014 Sep 8.

Texas Children's Cancer Center, Texas Children's Hospital, Houston, TX; Department of Molecular and Human Genetics, Human Genome Sequencing Center, Division of Pediatric Hematology-Oncology, Department of Pediatrics, Program in Translational Biology and Molecular Medicine, and.

Langerhans cell histiocytosis (LCH) is a myeloproliferative disorder characterized by lesions composed of pathological CD207(+) dendritic cells with an inflammatory infiltrate. BRAFV600E remains the only recurrent mutation reported in LCH. In order to evaluate the spectrum of somatic mutations in LCH, whole exome sequencing was performed on matched LCH and normal tissue samples obtained from 41 patients. Lesions from other histiocytic disorders, juvenile xanthogranuloma, Erdheim-Chester disease, and Rosai-Dorfman disease were also evaluated. All of the lesions from histiocytic disorders were characterized by an extremely low overall rate of somatic mutations. Notably, 33% (7/21) of LCH cases with wild-type BRAF and none (0/20) with BRAFV600E harbored somatic mutations in MAP2K1 (6 in-frame deletions and 1 missense mutation) that induced extracellular signal-regulated kinase (ERK) phosphorylation in vitro. Single cases of somatic mutations of the mitogen-activated protein kinase (MAPK) pathway genes ARAF and ERBB3 were also detected. The ability of MAPK pathway inhibitors to suppress MAPK kinase and ERK phosphorylation in cell culture and primary tumor models was dependent on the specific LCH mutation. The findings of this study support a model in which ERK activation is a universal end point in LCH arising from pathological activation of upstream signaling proteins.
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http://dx.doi.org/10.1182/blood-2014-05-577825DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4224195PMC
November 2014

BRAF-V600E expression in precursor versus differentiated dendritic cells defines clinically distinct LCH risk groups.

J Exp Med 2014 Apr 17;211(4):669-83. Epub 2014 Mar 17.

Department of Oncological Sciences, 2 Tisch Cancer Institute, and 3 Immunology Institute, Mount Sinai School of Medicine, New York, NY 10029.

Langerhans cell histiocytosis (LCH) is a clonal disorder with elusive etiology, characterized by the accumulation of CD207(+) dendritic cells (DCs) in inflammatory lesions. Recurrent BRAF-V600E mutations have been reported in LCH. In this study, lesions from 100 patients were genotyped, and 64% carried the BRAF-V600E mutation within infiltrating CD207(+) DCs. BRAF-V600E expression in tissue DCs did not define specific clinical risk groups but was associated with increased risk of recurrence. Strikingly, we found that patients with active, high-risk LCH also carried BRAF-V600E in circulating CD11c(+) and CD14(+) fractions and in bone marrow (BM) CD34(+) hematopoietic cell progenitors, whereas the mutation was restricted to lesional CD207(+) DC in low-risk LCH patients. Importantly, BRAF-V600E expression in DCs was sufficient to drive LCH-like disease in mice. Consistent with our findings in humans, expression of BRAF-V600E in BM DC progenitors recapitulated many features of the human high-risk LCH, whereas BRAF-V600E expression in differentiated DCs more closely resembled low-risk LCH. We therefore propose classification of LCH as a myeloid neoplasia and hypothesize that high-risk LCH arises from somatic mutation of a hematopoietic progenitor, whereas low-risk disease arises from somatic mutation of tissue-restricted precursor DCs.
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http://dx.doi.org/10.1084/jem.20130977DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3978272PMC
April 2014

Robust and cost effective expansion of human regulatory T cells highly functional in a xenograft model of graft-versus-host disease.

Haematologica 2013 Apr 14;98(4):533-7. Epub 2012 Dec 14.

Center for Cell and Gene Therapy, Methodist Hospital and Texas Children's Hospital, Houston, TX, USA.

The low frequency of naturally occurring regulatory T cells (nTregs) in peripheral blood and the suboptimal protocols available for their ex vivo expansion limit the development of clinical trials based on the adoptive transfer of these cells. We have, therefore, generated a simplified, robust and cost-effective platform for the large-scale expansion of nTregs using a gas permeable static culture flask (G-Rex) in compliance with Good Manufacturing Practice. More than 10(9) putative Tregs co-expressing CD25 and CD4 molecules (92 ± 5%) and FoxP3 (69 ± 19%) were obtained within 21 days of culture. Expanded Tregs showed potent regulatory activity in vitro (80 ± 13% inhibition of CD8(+) cell division) and in vivo (suppression or delay of graft-versus-host disease in a xenograft mouse model) indicating that the cost-effective and simplified production of nTregs we propose will facilitate the implementation of clinical trials based on their adoptive transfer.
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http://dx.doi.org/10.3324/haematol.2012.076430DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3659983PMC
April 2013

Changes in chemokine receptor expression of regulatory T cells after ex vivo culture.

J Immunother 2012 May;35(4):329-36

Center for Cell and Gene Therapy, Department of Pediatrics, Baylor College of Medicine, Methodist Hospital and Texas Children's Hospital, Houston, TX 77030, USA.

By controlling and limiting inflammatory conditions, naturally occurring regulatory T cells (Tregs), defined as circulating CD4(+)CD25(bright)FoxP3(+) cells, play critical roles in maintaining tolerance and preventing autoimmunity and thus have tremendous potential for adoptive immunotherapy. Because they represent a scanty subset of the CD4(+) T-lymphocyte subset, several approaches have been developed to isolate and expand ex vivo polyclonal Tregs. However, one limitation of the functional analyses performed on these cultured Tregs is the incomplete characterization of their tissue-trafficking properties. As this aspect provides crucial information for their therapeutic effects, we have here explored the chemokine receptor expression profile and function of Tregs cultured ex vivo with validated expansion protocols. Our data show that ex vivo cultured Tregs retained the expression of CCR7 but dramatically downregulated CCR5 as compared with freshly isolated Tregs. The differential chemokine receptors expression pattern corroborated with their respective steady state messenger RNA expression and also with their migration toward specific chemokines. Our analyses suggest that ex vivo cultured Tregs may display impaired or suboptimal migration to the inflamed tissues releasing RANTES and MIP-1α chemokines.
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http://dx.doi.org/10.1097/CJI.0b013e318255adccDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3650839PMC
May 2012

Activation of Wnt signaling arrests effector differentiation in human peripheral and cord blood-derived T lymphocytes.

J Immunol 2011 Nov 19;187(10):5221-32. Epub 2011 Oct 19.

Department of Pathology and Immunology, Baylor College of Medicine, Houston, TX 77030, USA.

The canonical Wnt/β-catenin signaling pathway plays an important role in thymocyte development and T cell migration, but little is known about its role in naive-to-effector differentiation in human peripheral T cells. We show that activation of Wnt/β-catenin signaling arrests human peripheral blood and cord blood T lymphocytes in the naive stage and blocks their transition into functional T effector cells. Wnt signaling was induced in polyclonally activated human T cells by treatment either with the glycogen synthase kinase 3β inhibitor TWS119 or the physiological Wnt agonist Wnt-3a, and these T cells preserved a naive CD45RA(+)CD62L(+) phenotype compared with control-activated T cells that progressed to a CD45RO(+)CD62L(-) effector phenotype, and this occurred in a TWS119 dose-dependent manner. TWS119-induced Wnt signaling reduced T cell expansion, as a result of a block in cell division, and impaired acquisition of T cell effector function, measured by degranulation and IFN-γ production in response to T cell activation. The block in T cell division may be attributed to the reduced IL-2Rα expression in TWS119-treated T cells that lowers their capacity to use autocrine IL-2 for expansion. Collectively, our data suggest that Wnt/β-catenin signaling is a negative regulator of naive-to-effector T cell differentiation in human T lymphocytes. The arrest in T cell differentiation induced by Wnt signaling might have relevant clinical applications such as to preserve the naive T cell compartment in Ag-specific T cells generated ex vivo for adoptive T cell immunotherapy.
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http://dx.doi.org/10.4049/jimmunol.1101585DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3208052PMC
November 2011

Redox regulation of interleukin-4 signaling.

Immunity 2008 Oct;29(4):551-64

Department of Cancer Biology, Lerner Research Institute, Cleveland Clinic, Cleveland, OH 44195, USA.

The physiologic control of cytokine receptor activation is primarily mediated by reciprocal activation of receptor-associated protein tyrosine kinases and protein tyrosine phosphatases (PTPs). Here, we show that immediately after ligand-dependent activation, interleukin (IL)-4 receptor generated reactive oxygen species (ROS) via phosphatidylinositol 3-kinase-dependent activation of NAD(P)H oxidase (NOX)1 and NOX5L. ROS, in turn, promoted IL-4 receptor activation by oxidatively inactivating PTP1B that physically associated with and deactivated IL-4 receptor. However, ROS were not required for the initiation of IL-4 receptor activation. ROS generated by other cytokine receptors, including those for erythropoietin, tumor necrosis factor-alpha, or IL-3, also promoted IL-4 signaling. These data indicate that inactivation of receptor-associated PTP activity by cytokine-generated ROS is a physiologic mechanism for the amplification of cytokine receptor activation in both cis and trans, revealing a role for ROS in cytokine crosstalk.
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http://dx.doi.org/10.1016/j.immuni.2008.07.019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2631209PMC
October 2008