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1.
Int J Mol Sci ; 22(6)2021 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-33808720

RESUMO

Using a murine model of chronic ischemic cardiomyopathy caused by an old myocardial infarction (MI), we have previously found that three doses of 1 × 106 c-kit positive cardiac cells (CPCs) are more effective than a single dose of 1 × 106 cells. The goal of this study was to determine whether the beneficial effects of three doses of CPCs (1 × 106 cells each) can be fully replicated by a single combined dose of 3 × 106 CPCs. Mice underwent a 60-min coronary occlusion; after 90 days of reperfusion, they received three echo-guided intraventricular infusions at 5-week intervals: (1) vehicle × 3; (2) one combined dose of CPCs (3 × 106) and vehicle × 2; or (3) three doses of CPCs (1 × 106 each). In the combined-dose group, left ventricular ejection fraction (LVEF) improved after the 1st CPC infusion, but not after the 2nd and 3rd (vehicle) infusions. In contrast, in the multiple-dose group, LVEF increased after each CPC infusion; at the final echo, LVEF averaged 35.2 ± 0.6% (p < 0.001 vs. the vehicle group, 27.3 ± 0.2%). At the end of the study, the total cumulative change in EF from pretreatment values was numerically greater in the multiple-dose group (6.6 ± 0.6%) than in the combined-dose group (4.8 ± 0.8%), although the difference was not statistically significant (p = 0.08). Hemodynamic studies showed that several parameters of LV function in the multiple-dose group were numerically greater than in the combined-dose group (p = 0.08 for the difference in LVEF). Compared with vehicle, cardiomyocyte cross-sectional area was reduced only in the multiple-dose group (-32.7%, 182.6 ± 15.1 µm2 vs. 271.5 ± 27.2 µm2, p < 0.05, in the risk region and -28.5%, 148.5 ± 12.1 µm2 vs. 207.6 ± 20.5 µm2, p < 0.05, in the noninfarcted region). LV weight/body weight ratio and LV weight/tibia length ratios were significantly reduced in both cell treated groups vs. the vehicle group, indicating the attenuation of LV hypertrophy; however, the lung weight/body weight ratio was significantly reduced only in the multiple-dose group, suggesting decreased pulmonary congestion. Taken together, these results indicate that in mice with chronic ischemic cardiomyopathy, the beneficial effects of three doses of CPCs on LV function and hypertrophy cannot be fully replicated with a single dose, notwithstanding the fact that the total number of cells delivered with one or three doses is the same. Thus, it is the multiplicity of doses, and not the total number of cells, that accounts for the superiority of the repeated-dose paradigm. This study supports the idea that the efficacy of cell therapy in heart failure can be augmented by repeated administrations.


Assuntos
Cardiomiopatias/etiologia , Dosagem de Genes , Isquemia Miocárdica/complicações , Miócitos Cardíacos/metabolismo , Proteínas Proto-Oncogênicas c-kit/genética , Animais , Biomarcadores , Biópsia , Pesos e Medidas Corporais , Cardiomiopatias/diagnóstico , Cardiomiopatias/metabolismo , Cardiomiopatias/terapia , Células Cultivadas , Modelos Animais de Doenças , Ecocardiografia , Fibrose , Testes de Função Cardíaca , Hemodinâmica , Hipertrofia Ventricular Esquerda/etiologia , Hipertrofia Ventricular Esquerda/metabolismo , Hipertrofia Ventricular Esquerda/patologia , Camundongos , Infarto do Miocárdio/etiologia , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Isquemia Miocárdica/etiologia , Proteínas Proto-Oncogênicas c-kit/metabolismo
2.
Int J Mol Sci ; 22(5)2021 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-33806359

RESUMO

In systemic mastocytosis (SM), qualitative and serial quantitative assessment of the KIT D816V mutation is of diagnostic and prognostic relevance. We investigated peripheral blood and bone marrow samples of 161 patients (indolent SM (ISM), n = 40; advanced SM, AdvSM, n = 121) at referral and during follow-up for the KIT D816V variant allele frequency (VAF) at the DNA-level and the KIT D816V expressed allele burden (EAB) at the RNA-level. A round robin test with four participating laboratories revealed an excellent correlation (r > 0.99, R2 > 0.98) between three different DNA-assays. VAF and EAB strongly correlated in ISM (r = 0.91, coefficient of determination, R2 = 0.84) but only to a lesser extent in AdvSM (r = 0.71; R2 = 0.5). However, as compared to an EAB/VAF ratio ≤2 (cohort A, 77/121 patients, 64%) receiver operating characteristic (ROC) analysis identified an EAB/VAF ratio of >2 (cohort B, 44/121 patients, 36%) as predictive for an advanced phenotype and a significantly inferior median survival (3.3 vs. 11.7 years; p = 0.005). In terms of overall survival, Cox-regression analysis was only significant for the EAB/VAF ratio >2 (p = 0.006) but not for VAF or EAB individually. This study demonstrates for the first time that the transcriptional activity of KIT D816V may play an important role in the pathophysiology of SM.


Assuntos
Mastocitose Sistêmica/genética , Mutação , Proteínas Proto-Oncogênicas c-kit/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Substituição de Aminoácidos , Medula Óssea/metabolismo , DNA/sangue , DNA/genética , DNA/metabolismo , Feminino , Frequência do Gene , Humanos , Masculino , Mastocitose Sistêmica/sangue , Mastocitose Sistêmica/metabolismo , Pessoa de Meia-Idade , Fenótipo , Prognóstico , Proteínas Proto-Oncogênicas c-kit/metabolismo , RNA/sangue , RNA/genética , RNA/metabolismo , Transcrição Genética
3.
Int J Mol Sci ; 22(6)2021 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-33804174

RESUMO

Systemic mastocytosis (SM) is a rare clonal hematologic neoplasm, driven, in almost all cases, by the activating KIT D816V mutation that leads to the growth and accumulation of neoplastic mast cells. While patients with advanced forms of SM have a poor prognosis, the introduction of KIT inhibitors (e.g., midostaurin, and avapritinib) has changed their outlook. Because of the heterogenous nature of advanced SM (advSM), successive iterations of response criteria have tried to capture different dimensions of the disease, including measures of mast cell burden (percentage of bone marrow mast cells and serum tryptase level), and mast cell-related organ damage (referred to as C findings). Historically, response criteria have been anchored to reversion of one or more organ damage finding(s) as a minimal criterion for response. This is a central principle of the Valent criteria, Mayo criteria, and International Working Group-Myeloproliferative Neoplasms Research and Treatment and European Competence Network on Mastocytosis (IWG-MRT-ECNM) consensus criteria. Irrespective of the response criteria, an ever-present challenge is how to apply response criteria in patients with SM and an associated hematologic neoplasm, where the presence of both diseases complicates assignment of organ damage and adjudication of response. In the context of trials with the selective KIT D816V inhibitor avapritinib, pure pathologic response (PPR) criteria, which rely solely on measures of mast cell burden and exclude consideration of organ damage findings, are being explored as more robust surrogate of overall survival. In addition, the finding that avapritinib can elicit complete molecular responses of KIT D816V allele burden, establishes a new benchmark for advSM and motivates the inclusion of definitions for molecular response in future criteria. Herein, we also outline how the concept of PPR can inform a proposal for new response criteria which use a tiered evaluation of pathologic, molecular, and clinical responses.


Assuntos
Proliferação de Células/genética , Mastocitose Sistêmica/tratamento farmacológico , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas c-kit/genética , Inibidores Enzimáticos/uso terapêutico , Feminino , Humanos , Mastócitos/efeitos dos fármacos , Mastócitos/patologia , Mastocitose Sistêmica/genética , Mastocitose Sistêmica/patologia , Pirazóis/uso terapêutico , Pirróis/uso terapêutico , Estaurosporina/análogos & derivados , Estaurosporina/uso terapêutico , Triazinas/uso terapêutico
4.
Int J Mol Sci ; 22(4)2021 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-33562323

RESUMO

Aggressive chemotherapy treatment may lead to male infertility. Prepubertal boys do not produce sperm at this age, however, they have spermatogonial stem cells in their testes. Here, we examined the effect of intraperitoneal injection of cyclophosphamide (CP) on the capacity of immature mice (IM) to develop spermatogenesis in vivo and in vitro [using methylcellulose culture system (MCS)]. Our results show a significant decrease in testicular weight, total number of testicular cells, and the number of Sertoli, peritubular, premeiotic, and meiotic/post-meiotic cells, but an increase in the percentages of damaged seminiferous tubules in CP-treated IM compared to control. The functionality of Sertoli cells was significantly affected. The addition of testosterone to isolated cells from seminiferous tubules of CP-treated IM significantly increased the percentages of premeiotic (CD9-positive cells) and meiotic/post-meiotic cells (ACROSIN-positive cells) developed in MCS compared to control. The addition of FSH did not affect developed cells in MCS compared to control, but in combination with testosterone, it significantly decreased the percentages of CD9-positive cells and ACROSIN-positive cells. The addition of IL-1 did not affect developed cells in MCS compared to control, but in combination with testosterone, it significantly increased the percentages of VASA-positive cells and BOULE-positive cells compared to IL-1 or testosterone. Addition of TNF significantly increased only CD9-positive cells in MCS compared to control, but in combination with testosterone, it significantly decreased ACROSIN-positive cells compared to testosterone. Our results show a significant impairment of spermatogenesis in the testes of CP-treated IM, and that spermatogonial cells from these mice proliferate and differentiate to meiotic/post-meiotic cells under in vitro culture conditions.


Assuntos
Ciclofosfamida/toxicidade , Citocinas/farmacologia , Hormônios/farmacologia , Infertilidade Masculina/patologia , Tamanho do Órgão/efeitos dos fármacos , Espermatogênese , Espermatogônias/patologia , Animais , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/genética , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Técnicas In Vitro , Infertilidade Masculina/induzido quimicamente , Infertilidade Masculina/metabolismo , Integrina alfa6/genética , Integrina alfa6/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos ICR , Mutagênicos/toxicidade , Proteínas Proto-Oncogênicas c-kit/genética , Proteínas Proto-Oncogênicas c-kit/metabolismo , Espermatogônias/efeitos dos fármacos , Espermatogônias/metabolismo , Tetraspanina 29/genética , Tetraspanina 29/metabolismo
5.
Am J Hematol ; 96(4): 508-525, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33524167

RESUMO

OVERVIEW: Systemic mastocytosis (SM) results from a clonal proliferation of abnormal mast cells (MC) in extra-cutaneous organs. DIAGNOSIS: The major criterion is presence of multifocal clusters of spindled MC in the bone marrow. Minor diagnostic criteria include elevated serum tryptase level, abnormal MC CD25 expression, and presence of KITD816V mutation. RISK STRATIFICATION: Establishing SM subtype as per the World Health Organization classification system is an important first step. Broadly, patients either have indolent/smoldering SM (ISM/SSM) or advanced SM, the latter includes aggressive SM (ASM), SM with associated hematological neoplasm (SM-AHN), and mast cell leukemia (MCL). Identification of poor-risk mutations (ie, ASXL1, RUNX1, SRSF2, NRAS) further refines the risk stratification. Recently, clinical and hybrid clinical-molecular risk models have been developed to more accurately assign prognosis in SM patients. MANAGEMENT: Treatment goals for ISM patients are primarily directed towards anaphylaxis prevention/symptom control/osteoporosis treatment. Patients with advanced SM frequently need MC cytoreductive therapy to ameliorate disease-related organ dysfunction. High response rates have been seen with small-molecule inhibitors that target mutant-KIT, including midostaurin (Food and Drug Administration approved) or avapritinib (investigational). Other options for MC cytoreduction include cladribine or interferon-α, although head-to-head comparisons are lacking. Treatment of SM-AHN primarily targets the AHN component, particularly if an aggressive disease such as acute myeloid leukemia is present. Allogeneic stem cell transplant can be considered in such patients, or in those with relapsed/refractory advanced SM. Imatinib has a limited therapeutic role in SM; effective cytoreduction is limited to those with imatinib-sensitive KIT mutations.


Assuntos
Mastocitose Sistêmica , Adulto , Algoritmos , Animais , Medula Óssea/patologia , Gerenciamento Clínico , Modelos Animais de Doenças , Drogas em Investigação/uso terapêutico , Mutação com Ganho de Função , Neoplasias Hematológicas/epidemiologia , Humanos , Hidroxiureia/uso terapêutico , Subunidade alfa de Receptor de Interleucina-2/análise , Estimativa de Kaplan-Meier , Leucemia de Mastócitos/epidemiologia , Leucemia de Mastócitos/etiologia , Mastócitos/química , Mastócitos/patologia , Mastocitose Sistêmica/diagnóstico , Mastocitose Sistêmica/tratamento farmacológico , Mastocitose Sistêmica/epidemiologia , Mastocitose Sistêmica/genética , Camundongos , Camundongos Transgênicos , Mutação de Sentido Incorreto , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas c-kit/genética , Medição de Risco , Triptases/sangue
6.
Am J Hum Genet ; 108(2): 284-294, 2021 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-33421400

RESUMO

Mastocytosis is a rare myeloid neoplasm characterized by uncontrolled expansion of mast cells, driven in >80% of affected individuals by acquisition of the KIT D816V mutation. To explore the hypothesis that inherited variation predisposes to mastocytosis, we performed a two-stage genome-wide association study, analyzing 1,035 individuals with KIT D816V positive disease and 17,960 healthy control individuals from five European populations. After quality control, we tested 592,007 SNPs at stage 1 and 75 SNPs at stage 2 for association by using logistic regression and performed a fixed effects meta-analysis to combine evidence across the two stages. From the meta-analysis, we identified three intergenic SNPs associated with mastocytosis that achieved genome-wide significance without heterogeneity between cohorts: rs4616402 (pmeta = 1.37 × 10-15, OR = 1.52), rs4662380 (pmeta = 2.11 × 10-12, OR = 1.46), and rs13077541 (pmeta = 2.10 × 10-9, OR = 1.33). Expression quantitative trait analyses demonstrated that rs4616402 is associated with the expression of CEBPA (peQTL = 2.3 × 10-14), a gene encoding a transcription factor known to play a critical role in myelopoiesis. The role of the other two SNPs is less clear: rs4662380 is associated with expression of the long non-coding RNA gene TEX41 (peQTL = 2.55 × 10-11), whereas rs13077541 is associated with the expression of TBL1XR1, which encodes transducin (ß)-like 1 X-linked receptor 1 (peQTL = 5.70 × 10-8). In individuals with available data and non-advanced disease, rs4616402 was associated with age at presentation (p = 0.009; beta = 4.41; n = 422). Additional focused analysis identified suggestive associations between mastocytosis and genetic variation at TERT, TPSAB1/TPSB2, and IL13. These findings demonstrate that multiple germline variants predispose to KIT D816V positive mastocytosis and provide novel avenues for functional investigation.


Assuntos
Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Mastocitose/genética , Polimorfismo de Nucleotídeo Único , Proteínas Proto-Oncogênicas c-kit/genética , Sistema y+ de Transporte de Aminoácidos/genética , Proteínas Estimuladoras de Ligação a CCAAT/genética , DNA Intergênico , Feminino , Humanos , Interleucina-13/genética , Íntrons , Masculino , RNA Longo não Codificante/genética , Receptores Citoplasmáticos e Nucleares/genética , Proteínas Repressoras/genética , Telomerase/genética , Triptases/genética
7.
J Cancer Res Clin Oncol ; 147(4): 1065-1075, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33389076

RESUMO

PURPOSE: Imatinib, a small-molecule tyrosine kinase inhibitor, has shown good clinical activity by inhibiting adenosine triphosphate (ATP) binding to the receptor. Unfortunately, majority of patients eventually develop drug resistance, which limits the long-term benefits of the tyrosine kinase inhibitors and poses a significant challenge in the clinical management of GIST. The aim of our study was to explore the feasibility of blocking KIT dimerisation upstream of the phosphorylation in imatinib-resistant GIST. METHOD: KITMAb was prepared using hybridoma technique. The biological function of KITMAb was examined in KIT-dimer-expressing cells constructed by transfecting with liposomes using enzyme linked immunosorbent assay (ELISA), immunohistochemistry, western blot, MTT, Annexin V/FITC, and flow cytometry assay, respectively. RESULTS: KIT-dimer was expressed in 293 cells transfected with c-kit mutated-type pcDNA3.1. Treatment of KIT-dimer-expressing cells with the KITMAb significantly decreased the expression of both KIT-dimer and other phosphorylated proteins of KIT downstream signalling pathway. Furthermore, KITMAb slowed down cell growth and reduced the proportion of cells in the proliferative phase (S + G2-M). Finally, we also found that KITMAb treatment accelerated cell apoptosis. These results indicate that KITMAb strongly inhibits KIT receptor dimerisation-mediated signalling pathway and cell growth responses in vitro. CONCLUSIONS: We demonstrate c-kit mutation-driven KIT auto-dimerisation prior to tyrosine kinase phosphorylation as same as the procedure in ligand-dependent signalling pathway and describe a monoclonal antibody, KITMAb, with strong affinity to the dimerisation domain of KIT that blocks the important step in both the KIT signalling pathways. Further, the results suggest that treatment with KITMAb may be potentially therapeutic in imatinib-resistant GIST.


Assuntos
Anticorpos Monoclonais/farmacologia , Biomarcadores Tumorais/metabolismo , Regulação Neoplásica da Expressão Gênica , Mutação , Neoplasias/patologia , Multimerização Proteica , Proteínas Proto-Oncogênicas c-kit/metabolismo , Apoptose , Biomarcadores Tumorais/genética , Ciclo Celular , Proliferação de Células , Humanos , Neoplasias/genética , Neoplasias/imunologia , Neoplasias/metabolismo , Proteínas Proto-Oncogênicas c-kit/química , Proteínas Proto-Oncogênicas c-kit/genética , Proteínas Proto-Oncogênicas c-kit/imunologia , Células Tumorais Cultivadas
8.
Phys Chem Chem Phys ; 23(5): 3361-3376, 2021 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-33502401

RESUMO

The stability of c-KIT G-quadruplex DNA via ligands has been a significant concern in the growing field of cancer therapy. Thus, it is very important to understand the mechanism behind the high binding affinity of the small drug molecules on the c-KIT G-quadruplex DNA. In this study, we have investigated the binding mode and pathway of the APTO-253 ligand on the c-KIT G-quadruplex DNA employing a total of 10 µs all atom molecular dynamics simulations and further 8.82 µs simulations via the umbrella sampling method using both OL15 and BSC1 latest force fields for DNA structures. From the cluster structure analysis, mainly three binding pathways i.e., top, bottom and side loop stacking modes are identified. Moreover, RMSD, RMSF and 2D-RMSD values indicate that the c-KIT G-quadruplex DNA and APTO-253 molecules are stable throughout the simulation run. Furthermore, the number of hydrogen bonds in each tetrad and the distance between the two central K+ cations confirm that the c-KIT G-quadruplex DNA maintains its conformation in the process of complex formation with the APTO-253 ligand. The binding free energies and the minimum values in the potential of mean forces suggest that the binding processes are energetically favorable. Furthermore, we have found that the bottom stacking mode is the most favorable binding mode among all the three modes for the OL15 force field. However, for the BSC1 force field, both the top and bottom binding modes of the APTO-253 ligand in c-KIT G-quadruplex DNA are comparable to each other. To investigate the driving force for the complex formation, we have noticed that the van der Waals (vdW) and π-π stacking interactions are mainly responsible. Our detailed studies provide useful information for the discovery of novel drugs in the field of stabilization of G-quadruplex DNAs.


Assuntos
DNA/metabolismo , Quadruplex G , Imidazóis/metabolismo , Fenantrolinas/metabolismo , Proteínas Proto-Oncogênicas c-kit/genética , Sítios de Ligação , DNA/química , DNA/genética , Humanos , Ligação de Hidrogênio , Imidazóis/química , Simulação de Dinâmica Molecular , Fenantrolinas/química , Eletricidade Estática
9.
Int J Mol Sci ; 22(1)2021 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-33401724

RESUMO

Mastocytosis is a rare and complex disease characterized by expansion of clonal mast cells (MC) in skin and/or various internal organ systems. Involvement of internal organs leads to the diagnosis of systemic mastocytosis (SM). The WHO classification divides SM into indolent SM, smoldering SM and advanced SM variants, including SM with an associated hematologic neoplasm, aggressive SM, and MC leukemia. Historically, genetic analysis of individuals with pure cutaneous mastocytosis (CM) and SM have focused primarily on cohort studies of inherited single nucleotide variants and acquired pathogenic variants. The most prevalent pathogenic variant (mutation) in patients with SM is KIT p.D816V, which is detectable in most adult patients. Other somatic mutations have also been identified-especially in advanced SM-in TET2, SRSF2, ASXL1, RUNX1, CBL and JAK2, and shown to impact clinical and cellular phenotypes. Although only small patient cohorts have been analyzed, disease associations have also been identified in several germline variants within genes encoding certain cytokines or their receptors (IL13, IL6, IL6R, IL31, IL4R) and toll-like receptors. More recently, an increased prevalence of hereditary alpha-tryptasemia (HαT) caused by increased TPSAB1 copy number encoding alpha-tryptase has been described in patients with SM. Whereas HαT is found in 3-6% of general Western populations, it is identified in up to 17% of patients with SM. In the current manuscript we review the prevalence, functional role and clinical impact of various germline and somatic genetic variants in patients with mastocytosis.


Assuntos
Citocinas/genética , Mastocitose Sistêmica/genética , Polimorfismo Genético , Proteínas Proto-Oncogênicas c-kit/genética , Receptor 2 Toll-Like/genética , Humanos , Interleucina-13/genética , Interleucina-6/genética , Interleucinas/genética , Mastócitos/patologia , Mastocitose Sistêmica/diagnóstico , Mastocitose Sistêmica/metabolismo , Mastocitose Sistêmica/fisiopatologia , Proteínas do Tecido Nervoso/genética , Fosfolipase C gama/genética , Proteínas Proto-Oncogênicas c-kit/metabolismo , Receptores Acoplados a Proteínas-G/genética , Receptores de Neuropeptídeos/genética , Receptor 2 Toll-Like/metabolismo
10.
Ann Hematol ; 100(5): 1203-1212, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33474629

RESUMO

Core binding factor acute myeloid leukemia (CBF-AML), including cases with KIT mutation, is currently defined as a low-risk AML. However, some patients have poor response to treatment, and the prognostic significance of KIT mutation is still controversial. This study aimed to explore the prognostic significance of different KIT mutation subtypes and minimal residual disease (MRD) in CBF-AML. We retrospectively evaluated continuous patients diagnosed with CBF-AML in our center between January 2014 and April 2019. Of the 215 patients, 147 (68.4%) and 68 (31.6%) patients were RUNX1-RUNX1T1- and CBFB-MYH11 positive, respectively. KIT mutations were found in 71 (33.0%) patients; of them, 38 (53.5%) had D816/D820 mutations. After excluding 10 patients who died or were lost to follow-up within a half year, 42.0% (n = 86) of the remaining 205 patients received allogeneic hematopoietic stem cell transplantation (allo-HSCT). An MRD > 0.1% at the end of two cycles of consolidation predicted relapse (P < 0.001). Multivariate analysis showed that D816 or D820 mutations and MRD > 0.1% at the end of two cycles of consolidation were independent adverse factors affecting relapse-free survival (RFS) and overall survival (OS). Allo-HSCT could improve RFS (74.4% vs. 34.6%, P < 0.001) and OS (78.1% vs. 52.3%, P = 0.002). In conclusion, high-risk CBF-AML patients must be identified before treatment. D816/D820 mutation, MRD > 0.1% at the end of two cycles of consolidation chemotherapy predicted poor survivals, and allo-HSCT can improve the survival of properly identified patients.


Assuntos
Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/genética , Proteínas Proto-Oncogênicas c-kit/genética , Adolescente , Adulto , Idoso , Feminino , Transplante de Células-Tronco Hematopoéticas , Humanos , Leucemia Mieloide Aguda/terapia , Masculino , Pessoa de Meia-Idade , Mutação , Neoplasia Residual , Prognóstico , Estudos Retrospectivos , Adulto Jovem
11.
Expert Opin Pharmacother ; 22(4): 439-447, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33307872

RESUMO

INTRODUCTION: 90% of gastrointestinal stromal tumors (GISTs) harbor an activating mutation in the KIT or PDGFRα oncogene, and these are known to confer imatinib sensitivity. AREAS COVERED: The author reviews the data regarding the current management of GIST, mechanisms of resistance to imatinib, and new drugs currently in clinical development and provides his unique perspectives on the subject matter. EXPERT OPINION: Several studies have shown that the response to imatinib in GIST patients mainly depends on the mutational status of KIT or PDGFRα. Moreover, most, if not all, patients treated with imatinib for advanced GIST will develop a secondary progressive disease under the treatment. In most cases, such progressions are the result of acquired resistance due to the occurrence of secondary c-KIT mutations, especially in GISTs with primary exon 11 mutations. Sunitinib and regorafenib are inhibitors of multiple tyrosine kinases, including KIT, PDGFRα, PDGFRß, and VEGFRs, and are approved for the management of imatinib- and imatinib/sunitinib-refractory GIST patients, respectively. Clearly, better knowledge of the molecular mechanisms underlying the resistance to imatinib as well as the development of a new class of broad-spectrum tyrosine kinase inhibitors such as avapritinib and ripretinib will provide new individualized therapeutic strategies for GIST patients.


Assuntos
Antineoplásicos/uso terapêutico , Neoplasias Gastrointestinais/tratamento farmacológico , Tumores do Estroma Gastrointestinal/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Tumores do Estroma Gastrointestinal/genética , Tumores do Estroma Gastrointestinal/patologia , Humanos , Proteínas Proto-Oncogênicas c-kit/genética
12.
Nucleic Acids Res ; 49(1): 416-431, 2021 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-33313902

RESUMO

G-Quadruplexes are non-B form DNA structures present at regulatory regions in the genome, such as promoters of proto-oncogenes and telomeres. The prominence in such sites suggests G-quadruplexes serve an important regulatory role in the cell. Indeed, oxidized G-quadruplexes found at regulatory sites are regarded as epigenetic elements and are associated with an interlinking of DNA repair and transcription. PARP-1 binds damaged DNA and non-B form DNA, where it covalently modifies repair enzymes or chromatin-associated proteins respectively with poly(ADP-ribose) (PAR). PAR serves as a signal in regulation of transcription, chromatin remodeling, and DNA repair. PARP-1 is known to bind G-quadruplexes with stimulation of enzymatic activity. We show that PARP-1 binds several G-quadruplex structures with nanomolar affinities, but only a subset promote PARP-1 activity. The G-quadruplex forming sequence found in the proto-oncogene c-KIT promoter stimulates enzymatic activity of PARP-1. The loop-forming characteristics of the c-KIT G-quadruplex sequence regulate PARP-1 catalytic activity, whereas eliminating these loop features reduces PARP-1 activity. Oxidized G-quadruplexes that have been suggested to form unique, looped structures stimulate PARP-1 activity. Our results support a functional interaction between PARP-1 and G-quadruplexes. PARP-1 enzymatic activation by G-quadruplexes is dependent on the loop features and the presence of oxidative damage.


Assuntos
Quadruplex G , Poli(ADP-Ribose) Polimerase-1/metabolismo , Catálise , Dano ao DNA , Ativação Enzimática , Guanina/análogos & derivados , Guanina/química , Humanos , Oxirredução , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas c-kit/genética , Proteínas Recombinantes/metabolismo , Relação Estrutura-Atividade , Especificidade por Substrato
13.
Am J Clin Pathol ; 155(2): 239-266, 2021 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-33313644

RESUMO

OBJECTIVES: The 2019 Workshop of the Society for Hematopathology/European Association for Haematopathology received and reviewed cases covering the spectrum of mastocytosis and related diseases, including morphologic mimics, focusing on recent updates and relevant findings for pathologists. METHODS: The workshop panel reviewed 99 cases of cutaneous and systemic mastocytosis (SM) and SM and associated hematologic neoplasms (SM-AHN). RESULTS: Despite a common theme of KIT mutation (particularly D816V), mastocytosis is a heterogeneous neoplasm with a wide variety of presentations. This spectrum, including rare subtypes and extramedullary organ involvement, is discussed and illustrated by representative cases. CONCLUSIONS: In the age of targeted treatment aimed at KIT, the accurate diagnosis and classification of mastocytosis has major implications for therapy and further interventions. Understanding the clinical, pathologic, and genetic findings of mastocytosis is crucial for selecting the proper tests to perform and subsequent arrival at a correct diagnosis in this rare disease.


Assuntos
Mastocitose , Adolescente , Adulto , Idoso , Antineoplásicos/uso terapêutico , Criança , Diagnóstico Diferencial , Feminino , Testes Genéticos , Humanos , Imuno-Histoquímica , Lactente , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mielomonocítica Crônica/diagnóstico , Masculino , Mastócitos/patologia , Mastocitose/diagnóstico , Mastocitose/tratamento farmacológico , Mastocitose/genética , Mastocitose/patologia , Mastocitose Sistêmica/tratamento farmacológico , Mastocitose Sistêmica/patologia , Pessoa de Meia-Idade , Mutação , Oncogenes , Prognóstico , Proteínas Proto-Oncogênicas c-kit/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-kit/genética , Triptases/isolamento & purificação , Adulto Jovem
14.
Methods Mol Biol ; 2158: 323-336, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32857384

RESUMO

Genetic lineage tracing is accomplished using bi-transgenic mice, where one allele is altered to express Cre recombinase, and another allele encodes a Cre-dependent genetic reporter protein. Once Cre is activated (constitutive or in response to tamoxifen), the marker gene-expressing cells become indelibly labeled by the reporter protein. Therefore, daughter cells derived from labeled cells are permanently labeled even if the marker gene that drove Cre recombinase expression is no longer expressed in these cells. This system is commonly used to label putative progenitor cells and determine the fate of their progeny. Here, we describe the use of c-kit-based genetic lineage-tracing mouse line as an example and discuss caveats for performing these types of experiments.


Assuntos
Linhagem da Célula/genética , Rastreamento de Células/métodos , Células-Tronco/química , Células-Tronco/metabolismo , Animais , Expressão Gênica , Genes Reporter , Ligação Genética , Proteínas de Fluorescência Verde/genética , Integrases/genética , Integrases/metabolismo , Camundongos , Camundongos Transgênicos , Modelos Animais , Miócitos Cardíacos/química , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , Proteínas Proto-Oncogênicas c-kit/análise , Proteínas Proto-Oncogênicas c-kit/genética , Células-Tronco/citologia , Tamoxifeno/farmacologia
15.
Medicine (Baltimore) ; 99(50): e21948, 2020 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-33327223

RESUMO

INTRODUCTION: Systemic mastocytosis is a rare disease due to mast cell accumulation in various extracutaneous sites. Systemic mastocytosis with an associated clonal hematologic non-MC lineage disease is the second most common subtype of systemic mastocytosis. The most common mutation associated with both systemic mastocytosis and myeloid sarcoma is mutation in Kit. Here, we identified the novel KIT D816V and ARID1A G1254S mutations co-occurring in systemic mastocytosis with myeloid sarcoma. PATIENT CONCERNS: A 33-year old male patient presented multiple skin lesions for 10 years. Symptoms accelerated in 2017 with decreased body weight. Physical examination revealed enlarged lymph nodes in his neck, axilla and inguinal region; conjunctival hemorrhage; gingival hyperplasia. Skin biopsy showed mast cell infiltration. Flow cytometry detected CD2, CD25 and CD117 positive cells in lymph nodes. Codon 816 KIT mutation D816V and codon 1245 ARID1A mutation G1254S were found in peripheral blood. MPO, CD117, CD68 positive cells in lymph nodes indicated co-existing myeloid sarcoma. DIAGNOSIS: Systemic mastocytosis with an associated clonal hematologic non-MC lineage disease of myeloid sarcoma INTERVENTIONS:: Cytarabine and daunorubicin for myeloid sarcoma and dasatinib for systemic mastocytosis were initiated. Anti-histamine and anti-leukotrienes therapy were used to prevent NSAIDs-induced shock. Platelets were infused to treat bone marrow suppression. OUTCOMES: Patient was discharged after recovered from bone marrow suppression. Dasatinib continued on outpatient. CONCLUSION: This is the first case of patient with systemic mastocytosis and myeloid sarcoma simultaneously presenting extensive skin involvements. Mutations of Kit and Arid1a emphasis the importance to notice possibility of various tumors occurring in patients with multiple mutations. In addition, cysteine-leukotrienes-receptor antagonists should always be used to prevent anaphylactic shock due to mast cell activation.


Assuntos
Mastocitose Sistêmica/complicações , Proteínas Proto-Oncogênicas c-kit/genética , Sarcoma Mieloide/complicações , Pele/patologia , Adulto , Antibióticos Antineoplásicos , Antimetabólitos Antineoplásicos/uso terapêutico , Antígenos CD2/metabolismo , Citarabina/uso terapêutico , Proteínas de Ligação a DNA/genética , Dasatinibe/uso terapêutico , Daunorrubicina/uso terapêutico , Quimioterapia Combinada , Antagonistas dos Receptores Histamínicos/uso terapêutico , Humanos , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Antagonistas de Leucotrienos/uso terapêutico , Linfonodos/metabolismo , Linfonodos/patologia , Masculino , Mastocitose Sistêmica/tratamento farmacológico , Mastocitose Sistêmica/genética , Mastocitose Sistêmica/patologia , Mutação , Transfusão de Plaquetas/métodos , Inibidores de Proteínas Quinases/uso terapêutico , Sarcoma Mieloide/tratamento farmacológico , Sarcoma Mieloide/genética , Sarcoma Mieloide/patologia , Fatores de Transcrição/genética , Resultado do Tratamento
16.
Drugs Today (Barc) ; 56(9): 561-571, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33025950

RESUMO

Avapritinib is a tyrosine kinase inhibitor (TKI) that has recently received Food and Drug Administration (FDA) approval for the treatment of metastatic or unresectable gastrointestinal stromal tumors harboring a platelet-derived growth factor receptor alpha (PDGFRA) exon 18 mutation. Mutations in the activation loop of PDGFRA or KIT confer resistance to conventional TKIs due to structural changes in the receptor. Avapritinib was developed to selectively target these mutations, thereby offering a new treatment option for patients in whom imatinib, sunitinib, and regorafenib have failed. This review covers the basic science and preclinical studies that guided avapritinib's development, in addition to the data currently available from early clinical studies as well as those later-stage trials that led to its approval.


Assuntos
Tumores do Estroma Gastrointestinal/tratamento farmacológico , Pirazóis/uso terapêutico , Pirróis/uso terapêutico , Triazinas/uso terapêutico , Ensaios Clínicos como Assunto , Tumores do Estroma Gastrointestinal/genética , Humanos , Mutação , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas c-kit/genética , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Estados Unidos
17.
Am J Hematol ; 95(12): 1562-1571, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32936982

RESUMO

Hyperdiploidy (HRD) and specific immunoglobulin heavy locus (IGH) translocations are primary chromosomal abnormalities (CA) in multiple myeloma (MM). In this retrospective study of 794 MM patients we aimed to investigate clinical features and common CA including gain(1q) in separate subgroups defined by primary CA. In the entire group, we confirmed that gain(1q) was associated with short time to next treatment and adverse overall survival (OS). The impact was worse for four or more copies of 1q21 as compared to three copies. However, in a subgroup of patients with clonal gain(11q) and without known primary IGH translocations (CG11q), already three copies of 1q21 were associated with a poor outcome; in the absence of gain(1q), patients in this subgroup had a remarkably long median OS of more than nine years. These cases were associated with HRD, coexpression of CD56 and CD117, male gender, and IgG subtype. In non-CG11q patients, four or more copies of 1q21 (but not three copies) had a significant adverse impact on outcome. Several associations with CA and clinical findings were observed for the defined subgroups. As an example, we found a predominance of early tetraploidy, plasma cell leukemia, and female gender in the t(14;16) subgroup. Our results underscore the importance of subgrouping in MM.


Assuntos
Cromossomos Humanos Par 1/genética , Loci Gênicos , Cadeias Pesadas de Imunoglobulinas/genética , Mieloma Múltiplo/genética , Mieloma Múltiplo/mortalidade , Proteínas de Neoplasias/genética , Translocação Genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígeno CD56/genética , Intervalo Livre de Doença , Feminino , Humanos , Imunoglobulina G/genética , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/terapia , Proteínas Proto-Oncogênicas c-kit/genética , Taxa de Sobrevida
18.
Nat Commun ; 11(1): 4147, 2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32811837

RESUMO

Mutated receptor tyrosine kinases (MT-RTKs) such as internal tandem duplication of FMS-like tyrosine kinase 3 (FLT3 ITD) and a point mutation KIT D816V are driver mutations for acute myeloid leukemia (AML). Clathrin assembly lymphoid myeloid leukemia protein (CALM) regulates intracellular transport of RTKs, however, the precise role for MT-RTKs remains elusive. We here show that CALM knock down leads to severely impaired FLT3 ITD- or KIT D814V-dependent cell growth compared to marginal influence on wild-type FLT3- or KIT-mediated cell growth. An antipsychotic drug chlorpromazine (CPZ) suppresses the growth of primary AML samples, and human CD34+CD38- AML cells including AML initiating cells with MT-RTKs in vitro and in vivo. Mechanistically, CPZ reduces CALM protein at post transcriptional level and perturbs the intracellular localization of MT-RTKs, thereby blocking their signaling. Our study presents a therapeutic strategy for AML with MT-RTKs by altering the intracellular localization of MT-RTKs using CPZ.


Assuntos
Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Clorpromazina/farmacologia , Leucemia Mieloide Aguda/genética , Proteínas Monoméricas de Montagem de Clatrina/metabolismo , Proteínas Proto-Oncogênicas c-kit/genética , Tirosina Quinase 3 Semelhante a fms/genética , Tirosina Quinase 3 Semelhante a fms/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Proliferação de Células/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Feminino , Células HL-60 , Humanos , Leucemia Mieloide Aguda/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Proteínas Monoméricas de Montagem de Clatrina/genética , Mutação Puntual , Transdução de Sinais/efeitos dos fármacos , Sequências de Repetição em Tandem/genética , Transplante Heterólogo , Adulto Jovem
19.
Pol J Pathol ; 71(2): 120-126, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32729302

RESUMO

Thymic epithelial tumours are rare malignancies of the anterior superior mediastinum. Several studies have analysed the presence of c-KIT mutations in thymic carcinoma. Immunohistochemical c-KIT expression and mutations in exons 8, 9, 11, 13, 14, 17, and 18 of the KIT gene and in the promoter region of the TERT gene (chr5, 1,295,228C>T/A and 1,295,250C>T) were analysed by PCR based direct sequencing using representative formalin-fixed paraffin-embedded tumour samples of 18 thymic carcinomas. Of 18 patients, 4 test samples were excluded from the study due to inadequate DNA quality. Of 14 patients with thymic carcinomas, KIT and TERT mutation was not detected in any samples. C-KIT expression was associated with nearly a worse overall survival (median time 24.160-49.840, log-rank, p = 0.05). We showed that squamous cell carcinomas led to worse survival than other subtypes. As expected, TNM stage II was significantly correlated with better OS (p = 0.015). Thymic carcinoma is characterised by a KIT-positive and CD5-positive staining pattern. We report a worse overall survival for patients with c-kit expressing tumours. These data suggest a negative prognostic role for c-kit expression especially within the first 5 years.


Assuntos
Proteínas Proto-Oncogênicas c-kit/genética , Timoma , Neoplasias do Timo , Carcinoma de Células Escamosas , Humanos , Mutação , Timoma/genética , Neoplasias do Timo/genética
20.
Yonsei Med J ; 61(7): 562-571, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32608199

RESUMO

Melanoma, originating from epidermal melanocytes, is a heterogeneous disease that has the highest mortality rate among all types of skin cancers. Numerous studies have revealed the cause of this cancer as related to various somatic driver mutations, including alterations in KIT-a proto-oncogene encoding for a transmembrane receptor tyrosine kinase. Although accounting for only 3% of all melanomas, mutations in c-KIT are mostly derived from acral, mucosal, and chronically sun-damaged melanomas. As an important factor for cell differentiation, proliferation, and survival, inhibition of c-KIT has been exploited for clinical trials in advanced melanoma. Here, apart from the molecular background of c-KIT and its cellular functions, we will review the wide distribution of alterations in KIT with a catalogue of more than 40 mutations reported in various articles and case studies. Additionally, we will summarize the association of KIT mutations with clinicopathologic features (age, sex, melanoma subtypes, anatomic location, etc.), and the differences of mutation rate among subgroups. Finally, several therapeutic trials of c-KIT inhibitors, including imatinib, dasatinib, nilotinib, and sunitinib, will be analyzed for their success rates and limitations in advanced melanoma treatment. These not only emphasize c-KIT as an attractive target for personalized melanoma therapy but also propose the requirement for additional investigational studies to develop novel therapeutic trials co-targeting c-KIT and other cytokines such as members of signaling pathways and immune systems.


Assuntos
Antineoplásicos/uso terapêutico , Biomarcadores Tumorais/metabolismo , Resistencia a Medicamentos Antineoplásicos/genética , Melanoma/tratamento farmacológico , Melanoma/metabolismo , Proteínas Proto-Oncogênicas c-kit/uso terapêutico , Transdução de Sinais/efeitos dos fármacos , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/metabolismo , Feminino , Humanos , Masculino , Melanoma/genética , Membrana Mucosa/metabolismo , Mutação , Proteínas Proto-Oncogênicas c-kit/genética , Proteínas Proto-Oncogênicas c-kit/metabolismo , Receptores Proteína Tirosina Quinases/genética , Neoplasias Cutâneas/genética
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