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1.
Circ Res ; 129(8): 804-820, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34433292
2.
Blood ; 136(21): 2442-2456, 2020 11 19.
Artigo em Inglês | MEDLINE | ID: mdl-32589720

RESUMO

The interaction of menin (MEN1) and MLL (MLL1, KMT2A) is a dependency and provides a potential opportunity for treatment of NPM1-mutant (NPM1mut) and MLL-rearranged (MLL-r) leukemias. Concomitant activating driver mutations in the gene encoding the tyrosine kinase FLT3 occur in both leukemias and are particularly common in the NPM1mut subtype. In this study, transcriptional profiling after pharmacological inhibition of the menin-MLL complex revealed specific changes in gene expression, with downregulation of the MEIS1 transcription factor and its transcriptional target gene FLT3 being the most pronounced. Combining menin-MLL inhibition with specific small-molecule kinase inhibitors of FLT3 phosphorylation resulted in a significantly superior reduction of phosphorylated FLT3 and transcriptional suppression of genes downstream of FLT3 signaling. The drug combination induced synergistic inhibition of proliferation, as well as enhanced apoptosis, compared with single-drug treatment in models of human and murine NPM1mut and MLL-r leukemias harboring an FLT3 mutation. Primary acute myeloid leukemia (AML) cells harvested from patients with NPM1mutFLT3mut AML showed significantly better responses to combined menin and FLT3 inhibition than to single-drug or vehicle control treatment, whereas AML cells with wild-type NPM1, MLL, and FLT3 were not affected by either of the 2 drugs. In vivo treatment of leukemic animals with MLL-r FLT3mut leukemia reduced leukemia burden significantly and prolonged survival compared with results in the single-drug and vehicle control groups. Our data suggest that combined menin-MLL and FLT3 inhibition represents a novel and promising therapeutic strategy for patients with NPM1mut or MLL-r leukemia and concurrent FLT3 mutation.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Leucemia Mieloide Aguda/tratamento farmacológico , Proteína de Leucina Linfoide-Mieloide/antagonistas & inibidores , Proteínas de Neoplasias/antagonistas & inibidores , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Tirosina Quinase 3 Semelhante a fms/antagonistas & inibidores , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Técnicas de Cocultura , Sinergismo Farmacológico , Humanos , Leucemia Mieloide Aguda/genética , Camundongos , Camundongos Endogâmicos NOD , Proteína Meis1/biossíntese , Proteína Meis1/genética , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Nucleofosmina , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Processamento de Proteína Pós-Traducional , Distribuição Aleatória , Transcrição Gênica/efeitos dos fármacos , Tirosina Quinase 3 Semelhante a fms/biossíntese , Tirosina Quinase 3 Semelhante a fms/genética
3.
Eur J Immunol ; 49(11): 2083-2094, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31393597

RESUMO

Transcutaneous immunization (TCI) is a novel vaccination strategy that utilizes skin-associated lymphatic tissue to induce immune responses. Employing T-cell epitopes and the TLR7 agonist imiquimod onto intact skin mounts strong primary, but limited memory CTL responses. To overcome this limitation, we developed a novel imiquimod-containing vaccination platform (IMI-Sol) rendering superior primary CD8+ and CD4+ T-cell responses. However, it has been unclear whether IMI-Sol per se is restricted in terms of memory formation and tumor protection. In our present work, we demonstrate that the combined administration of IMI-Sol and CD40 ligation unleashes fullblown specific T-cell responses in the priming and memory phase, strongly enhancing antitumor protection in mice. Interestingly, these effects were entirely CD4+ T cell independent, bypassing the necessity of helper T cells. Moreover, blockade of CD70 in vivo abrogated the boosting effect of CD40 ligation, indicating that the adjuvant effect of CD40 in TCI is mediated via CD70 on professional APCs. Furthermore, this work highlights the so far underappreciated importance of the CD70/CD27 interaction as a promising adjuvant target in TCI. Summing up, we demonstrate that the novel formulation IMI-Sol represents a powerful vaccination platform when applied in combination with sufficient adjuvant thereby overcoming current limitations of TCI.


Assuntos
Ligante CD27/imunologia , Ligante de CD40/administração & dosagem , Imiquimode/administração & dosagem , Melanoma Experimental/terapia , Neoplasias Cutâneas/terapia , Linfócitos T Citotóxicos/efeitos dos fármacos , Administração Cutânea , Aloenxertos , Animais , Ligante CD27/genética , Citotoxicidade Imunológica/efeitos dos fármacos , Expressão Gênica , Rejeição de Enxerto , Imunização/métodos , Memória Imunológica/efeitos dos fármacos , Imunoterapia/métodos , Melanoma Experimental/genética , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Glicoproteínas de Membrana/agonistas , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina/administração & dosagem , Pele/efeitos dos fármacos , Pele/imunologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/patologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/patologia , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Auxiliares-Indutores/patologia , Receptor 7 Toll-Like/agonistas , Receptor 7 Toll-Like/genética , Receptor 7 Toll-Like/imunologia
5.
Front Immunol ; 14: 1269012, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37809078

RESUMO

AML is a malignant disease of hematopoietic progenitor cells with unsatisfactory treatment outcome, especially in patients that are ineligible for intensive chemotherapy. Immunotherapy, comprising checkpoint inhibition, T-cell engaging antibody constructs, and cellular therapies, has dramatically improved the outcome of patients with solid tumors and lymphatic neoplasms. In AML, these approaches have been far less successful. Discussed reasons are the relatively low mutational burden of AML blasts and the difficulty in defining AML-specific antigens not expressed on hematopoietic progenitor cells. On the other hand, epigenetic dysregulation is an essential driver of leukemogenesis, and non-selective hypomethylating agents (HMAs) are the current backbone of non-intensive treatment. The first clinical trials that evaluated whether HMAs may improve immune checkpoint inhibitors' efficacy showed modest efficacy except for the anti-CD47 antibody that was substantially more efficient against AML when combined with azacitidine. Combining bispecific antibodies or cellular treatments with HMAs is subject to ongoing clinical investigation, and efficacy data are awaited shortly. More selective second-generation inhibitors targeting specific chromatin regulators have demonstrated promising preclinical activity against AML and are currently evaluated in clinical trials. These drugs that commonly cause leukemia cell differentiation potentially sensitize AML to immune-based treatments by co-regulating immune checkpoints, providing a pro-inflammatory environment, and inducing (neo)-antigen expression. Combining selective targeted epigenetic drugs with (cellular) immunotherapy is, therefore, a promising approach to avoid unintended effects and augment efficacy. Future studies will provide detailed information on how these compounds influence specific immune functions that may enable translation into clinical assessment.


Assuntos
Leucemia Mieloide Aguda , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Imunoterapia , Azacitidina/uso terapêutico , Resultado do Tratamento , Epigênese Genética
6.
Hamostaseologie ; 43(3): 215-218, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-34327693

RESUMO

Immune thrombotic thrombocytopenic purpura (iTTP) is a rare autoimmune disorder characterized by severely reduced activity of the von Willebrand factor (VWF)-cleaving protease ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) due to autoantibodies. This leads to the development of pathogenic multimers of VWF, causing a thrombotic microangiopathy with decreased number of platelets, hemolysis, and life-threatening tissue ischemia of mostly brain, heart, and kidneys. Standard treatment of iTTP involves daily plasma exchange to remove ultra large multimers of VWF, inhibitors, substituting ADAMTS13, and the accompaniment of an immunosuppressive treatment with steroids. Recently, caplacizumab was approved for iTTP. Caplacizumab is a nanobody binding the A1 domain of VWF, blocking its interaction with glycoprotein Ib-IX-V platelet receptor and therefore preventing platelet aggregation. VWF activities may serve as therapeutic drug monitoring of caplacizumab, whereas ADAMTS13 activities may be used for biomarkers to guide caplacizumab treatment modalities and overall treatment duration. Additional immunosuppressive treatment by inhibiting autoantibody formation (e.g., the use of Rituximab, a chimeric monoclonal antibody directed against the B-cell antigen CD20) is a further treatment option. Infections are well-known causes for an acute episode for patients with iTTP. The novel SARS-CoV-2 virus is mainly associated with acute respiratory distress as well as diffuse endothelial inflammation and increased coagulopathy. However, little is known about an infection with SARS-CoV-2 virus triggering iTTP relapses. We herein report the case of an acute iTTP episode accompanying a SARS-CoV-2 infection.


Assuntos
COVID-19 , Púrpura Trombocitopênica Idiopática , Púrpura Trombocitopênica Trombótica , Trombose , Humanos , Púrpura Trombocitopênica Trombótica/diagnóstico , Púrpura Trombocitopênica Trombótica/terapia , Fator de von Willebrand/metabolismo , COVID-19/complicações , SARS-CoV-2 , Púrpura Trombocitopênica Idiopática/diagnóstico , Púrpura Trombocitopênica Idiopática/tratamento farmacológico , Imunossupressores/uso terapêutico , Trombose/tratamento farmacológico , Autoanticorpos , Proteína ADAMTS13/uso terapêutico
7.
Naunyn Schmiedebergs Arch Pharmacol ; 394(11): 2233-2244, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34410453

RESUMO

Skin fibrosis is a complex biological remodeling process occurring in disease like systemic sclerosis, morphea, or eosinophilic fasciitis. Since the knowledge about the underlying pathomechanisms is still incomplete, there is currently no therapy, which prevents or reverses skin fibrosis sufficiently. The present study investigates the role of polo-like kinase 2 (PLK2) and the pro-fibrotic cytokine osteopontin (OPN) in the pathogenesis of cutaneous fibrosis and demonstrates the antifibrotic effects of systemic mesalazine treatment in vivo. Isolated primary dermal fibroblasts of PLK2 wild-type (WT) and knockout (KO) mice were characterized in vitro. Skin thickness and histoarchitecture were studied in paraffin-embedded skin sections. The effects of mesalazine treatment were examined in isolated fibroblasts and PLK2 KO mice, which were fed 100 µg/g mesalazine for 6 months via the drinking water. Compared to WT, PLK2 KO fibroblasts displayed higher spontaneous myofibroblast differentiation, reduced proliferation rates, and overexpression of the fibrotic cytokine OPN. In vitro, 72 h of treatment with 10 mmol/L mesalazine induced phenotype conversion in PLK2 KO fibroblasts and attenuated OPN expression by inhibiting ERK1/2. In vivo, dermal myofibroblast differentiation, collagen accumulation, and skin thickening were prevented by mesalazine in PLK2 KO. Plasma creatinine levels indicated good tolerability of systemic long-term mesalazine treatment. The current study reveals a spontaneous fibrotic skin phenotype and ERK1/2-dependent OPN overexpression in PLK2 KO mice. We provide experimental evidence for the antifibrotic effectiveness of systemic mesalazine treatment to prevent fibrosis of the skin, suggesting further investigation in experimental and clinical settings.


Assuntos
Fibroblastos/efeitos dos fármacos , Mesalamina/farmacologia , Proteínas Serina-Treonina Quinases/genética , Pele/efeitos dos fármacos , Animais , Anti-Inflamatórios não Esteroides/administração & dosagem , Anti-Inflamatórios não Esteroides/farmacologia , Anti-Inflamatórios não Esteroides/toxicidade , Diferenciação Celular/efeitos dos fármacos , Colágeno/metabolismo , Creatinina/sangue , Modelos Animais de Doenças , Feminino , Fibroblastos/patologia , Fibrose/prevenção & controle , Masculino , Mesalamina/administração & dosagem , Mesalamina/toxicidade , Camundongos , Camundongos Knockout , Osteopontina/genética , Pele/patologia
8.
Naunyn Schmiedebergs Arch Pharmacol ; 394(3): 533-543, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33064167

RESUMO

Cardiovascular diseases are exacerbated and driven by cardiac fibrosis. TGFß induces fibroblast activation and differentiation into myofibroblasts that secrete excessive extracellular matrix proteins leading to stiffening of the heart, concomitant cardiac dysfunction, and arrhythmias. However, effective pharmacotherapy for preventing or reversing cardiac fibrosis is presently unavailable. Therefore, drug repurposing could be a cost- and time-saving approach to discover antifibrotic interventions. The aim of this study was to investigate the antifibrotic potential of mesalazine in a cardiac fibroblast stress model. TGFß was used to induce a profibrotic phenotype in a human cardiac fibroblast cell line. After induction, cells were treated with mesalazine or solvent control. Fibroblast proliferation, key fibrosis protein expression, extracellular collagen deposition, and mechanical properties were subsequently determined. In response to TGFß treatment, fibroblasts underwent a profound phenoconversion towards myofibroblasts, determined by the expression of fibrillary αSMA. Mesalazine reduced differentiation nearly by half and diminished fibroblast proliferation by a third. Additionally, TGFß led to increased cell stiffness and adhesion, which were reversed by mesalazine treatment. Collagen 1 expression and deposition-key drivers of fibrosis-were significantly increased upon TGFß stimulation and reduced to control levels by mesalazine. SMAD2/3 and ERK1/2 phosphorylation, along with reduced nuclear NFκB translocation, were identified as potential modes of action. The current study provides experimental pre-clinical evidence for antifibrotic effects of mesalazine in an in vitro model of cardiac fibrosis. Furthermore, it sheds light on possible mechanisms of action and suggests further investigation in experimental and clinical settings.


Assuntos
Cardiotônicos/uso terapêutico , Mesalamina/uso terapêutico , Miocárdio/patologia , Actinas/metabolismo , Cardiotônicos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Colágeno Tipo I/metabolismo , Reposicionamento de Medicamentos , MAP Quinases Reguladas por Sinal Extracelular/antagonistas & inibidores , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fibrose , Humanos , Mesalamina/farmacologia , Miocárdio/metabolismo , Miofibroblastos/efeitos dos fármacos , Miofibroblastos/metabolismo , NF-kappa B/metabolismo , Proteína Smad2/antagonistas & inibidores , Proteína Smad2/metabolismo , Proteína Smad3/antagonistas & inibidores , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta
9.
FEBS Open Bio ; 10(7): 1210-1218, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32421922

RESUMO

Atrial fibrillation (AF) is regularly accompanied by cardiac fibrosis and concomitant heart failure. Due to the heterogeneous nature and complexity of fibrosis, the knowledge about the underlying mechanisms is limited, which prevents effective pharmacotherapy. A deeper understanding of cardiac fibroblasts is essential to meet this need. We previously described phenotypic and functional differences between atrial fibroblasts from patients in sinus rhythm and with AF. Herein, we established and characterized a novel human atrial fibroblast line, which displays typical fibroblast morphology and function comparable to primary cells but with improved proliferation capacity and low spontaneous myofibroblast differentiation. These traits make our model suitable for the study of fibrosis mechanisms and for drug screening aimed at developing effective antifibrotic pharmacotherapy.


Assuntos
Fibroblastos/metabolismo , Fibrose/metabolismo , Átrios do Coração/metabolismo , Modelos Biológicos , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Fibroblastos/patologia , Fibrose/patologia , Átrios do Coração/patologia , Humanos
10.
J Dermatol Sci ; 87(3): 300-306, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28666747

RESUMO

BACKGROUND: The epidermal application of the Toll Like Receptor 7 agonist imiquimod and a T-cell peptide epitope (transcutaneous immunization, TCI) mediates systemic peptide-specific cytotoxic T-cell (CTL) responses and leads to tumor protection in a prophylactic tumor setting. However, it does not accomplish memory formation or permanent defiance of tumors in a therapeutic set-up. As a distinct immunologic approach, CTLA-4 blockade augments systemic immune responses and has shown long-lasting effects in preclinical experiments as well as in clinical trials. OBJECTIVE: The study investigates the vaccination capacity of TCI in combination with the checkpoint inhibitor CTLA-4 in matters of primary response, memory formation and tumor protection and characterizes the role of regulatory T cells (Tregs). METHODS: After performing TCI with IMI-Sol (containing 5% Imiquimod) and the model epitope SIINFEKL, 6-8 week old C57BL/6 mice received anti-CTLA-4 antibody either s.c or i.p. The CTL responses and frequency of peptide specific CD8+ T-cells were then evaluated on day 8. To determine anti-tumor effects, a therapeutic tumor challenge with B16 OVA melanoma was performed. RESULTS: The combination of s.c. anti-CTLA-4 antibody and TCI leads to an enhanced systemic cytotoxic response, to memory formation and allows significantly improved survival in a tumor setting with B16 OVA melanoma. Towards the mechanism, we show that in this vaccination protocol the CTLA-4 antibody acts mainly Treg-independent. CONCLUSION: We demonstrate that the combination of TCI with IMI-Sol and anti-CTLA-4 can confer potent immune responses and tumor-protection. These results might contribute to the development of advanced vaccination approaches targeting tumors or persistent infectious diseases.


Assuntos
Adjuvantes Imunológicos/farmacologia , Antineoplásicos Imunológicos/farmacologia , Antígeno CTLA-4/antagonistas & inibidores , Melanoma Experimental/terapia , Neoplasias Cutâneas/terapia , Linfócitos T Citotóxicos/imunologia , Linfócitos T Reguladores/imunologia , Adjuvantes Imunológicos/uso terapêutico , Aminoquinolinas/farmacologia , Aminoquinolinas/uso terapêutico , Animais , Antineoplásicos Imunológicos/uso terapêutico , Antígeno CTLA-4/imunologia , Sinergismo Farmacológico , Citometria de Fluxo , Humanos , Imiquimode , Memória Imunológica/efeitos dos fármacos , Imunoterapia/métodos , Melanoma Experimental/imunologia , Melanoma Experimental/mortalidade , Glicoproteínas de Membrana/antagonistas & inibidores , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina/farmacologia , Ovalbumina/uso terapêutico , Fragmentos de Peptídeos/farmacologia , Fragmentos de Peptídeos/uso terapêutico , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/mortalidade , Linfócitos T Citotóxicos/metabolismo , Linfócitos T Reguladores/metabolismo , Receptor 7 Toll-Like/antagonistas & inibidores , Ensaios Antitumorais Modelo de Xenoenxerto
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