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2.
Nature ; 574(7779): 538-542, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31645727

RESUMO

The most common causes of chronic liver disease are excess alcohol intake, viral hepatitis and non-alcoholic fatty liver disease, with the clinical spectrum ranging in severity from hepatic inflammation to cirrhosis, liver failure or hepatocellular carcinoma (HCC). The genome of HCC exhibits diverse mutational signatures, resulting in recurrent mutations across more than 30 cancer genes1-7. Stem cells from normal livers have a low mutational burden and limited diversity of signatures8, which suggests that the complexity of HCC arises during the progression to chronic liver disease and subsequent malignant transformation. Here, by sequencing whole genomes of 482 microdissections of 100-500 hepatocytes from 5 normal and 9 cirrhotic livers, we show that cirrhotic liver has a higher mutational burden than normal liver. Although rare in normal hepatocytes, structural variants, including chromothripsis, were prominent in cirrhosis. Driver mutations, such as point mutations and structural variants, affected 1-5% of clones. Clonal expansions of millimetres in diameter occurred in cirrhosis, with clones sequestered by the bands of fibrosis that surround regenerative nodules. Some mutational signatures were universal and equally active in both non-malignant hepatocytes and HCCs; some were substantially more active in HCCs than chronic liver disease; and others-arising from exogenous exposures-were present in a subset of patients. The activity of exogenous signatures between adjacent cirrhotic nodules varied by up to tenfold within each patient, as a result of clone-specific and microenvironmental forces. Synchronous HCCs exhibited the same mutational signatures as background cirrhotic liver, but with higher burden. Somatic mutations chronicle the exposures, toxicity, regeneration and clonal structure of liver tissue as it progresses from health to disease.


Assuntos
Células Clonais/citologia , Células Clonais/patologia , Fibrose/genética , Fibrose/patologia , Fígado/citologia , Fígado/metabolismo , Mutação , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Células Clonais/metabolismo , Análise Mutacional de DNA , Hepatócitos/citologia , Hepatócitos/metabolismo , Hepatócitos/patologia , Humanos , Fígado/patologia , Masculino , Pessoa de Meia-Idade , Filogenia , Células-Tronco/citologia , Células-Tronco/metabolismo , Células-Tronco/patologia
3.
Blood ; 134(18): 1517-1527, 2019 10 31.
Artigo em Inglês | MEDLINE | ID: mdl-31515249

RESUMO

Mycosis fungoides (MF) is a mature T-cell lymphoma currently thought to develop primarily in the skin by a clonal expansion of a transformed, resident memory T cell. However, this concept does not explain the key characteristics of MF, such as the debut with multiple, widespread skin lesions or inability of skin-directed therapies to provide cure. The testable inference of the mature T-cell theory is the clonality of MF with respect to all rearranged T-cell receptor (TCR) genes. Here, we used a whole-exome sequencing approach to detect and quantify TCR-α, ß, and γ clonotypes in tumor cell clusters microdissected from MF lesions. This method allowed us to calculate the tumor cell fraction of the sample and therefore an unequivocal identification of the TCR clonotypes as neoplastic. Analysis of TCR sequences from 29 patients with MF stage I to IV proved the existence of multiple T-cell clones within the tumor cell fraction, with a considerable variation between patients and between lesions from the same patient (median, 11 clones; range, 2-80 clones/sample). We have also detected multiple neoplastic clones in the peripheral blood in all examined patients. Based on these findings, we propose that circulating neoplastic T-cell clones continuously replenish the lesions of MF, thus increasing their heterogeneity by a mechanism analogous to the consecutive tumor seeding. We hypothesize that circulating neoplastic clones might be a promising target for therapy and could be exploited as a potential biomarker in MF.


Assuntos
Micose Fungoide/patologia , Células Neoplásicas Circulantes/patologia , Neoplasias Cutâneas/patologia , Células Clonais/patologia , Humanos
4.
Ann Clin Lab Sci ; 49(4): 468-473, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31471335

RESUMO

OBJECTIVE: To explore the clonal origin pattern of cardiac myxoma and its relationship with recurrence of the disease. METHODS: 20 female patients diagnosed with cardiac myxoma underwent appropriate surgery and were followed-up after the treatment. The DNA of tumor tissues and pairing normal tissues from 20 patients were taken, with polymerase chain reaction (PCR) assay being used to amplify the HUMARA gene on X-chromosome, which could hint the tumor cloning state. Cases were divided into a polyclonal origin group and monoclonal origin group, according to the PCR result. The recurrence rate in the two groups was compared using Fisher's exact probability method. RESULTS: All tumors were successfully removed. PCR assay showed that the hybrid rate in tumors was 90.0% (18/20). Among them, 88.9%(16/18) of cases were identified as polyclonal origin and 11.1%(2/18) were identified as monoclonal origin. After 4 years of follow-up, the recurrence rate was 12.5(2/16) in the polyclonal origin group and 0%(0/2) in monoclonal origin group, with significant difference between the two groups (P<0.05). CONCLUSION: Cardiac myxoma is mostly of polyclonal original, and its polyclonal origin characteristics may contribute to tumor recurrence.


Assuntos
Células Clonais/patologia , Neoplasias Cardíacas/patologia , Mixoma/patologia , Recidiva Local de Neoplasia/patologia , Feminino , Neoplasias Cardíacas/genética , Heterozigoto , Humanos , Mixoma/genética , Receptores Androgênicos/genética
5.
Virchows Arch ; 475(6): 693-699, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31267202

RESUMO

The bone is a frequent localization for lung non-small cell cancer metastasis; decalcification is required to permit tissue section. Pre-analytical conditions can influence the detection of immunohistochemical markers. The aim of our work is to evaluate PD-L1 expression in samples with delayed fixation and in decalcified tissue with chelating agent or acid at different time. Tumor-expressing PD-L1 and placental tissue were fixed at different times or decalcified with an acid decalcifier or EDTA for different durations. For 22C3 antibody, when tissues were decalcified with DC3, there was a significant decrease in the percentage of tumor cells or placental villi stained which after 4 h (p = 0.035 at 4 h). When EDTA is used for 22C3 antibody, there was a slight decrease in the percentage of stained tumor cells or villi but although there was a trend (p = 0.058 at 20 h), this was never statistically significant. For E1L3N antibody, when tissues were decalcified either with DC3 or EDTA, there was no significant decrease for the proportion of stained tumor cells or placental villi, neither for staining intensity for the first 24 h. The proportion of placental villi and tumor stained or intensity of staining was not significantly lower for any sample after delayed fixation also at 24 h for both PD-L1 clones. Delayed fixation does not affect the proportion of stained cell and intensity with PD-L1 immunohistochemistry. Decalcification also performed with EDTA lower the proportion and intensity of stained cells with PD-L1 immunohistochemistry.


Assuntos
Antígeno B7-H1/metabolismo , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Fixação de Tecidos , Anticorpos Monoclonais/imunologia , Biomarcadores Tumorais/metabolismo , Células Clonais/patologia , Feminino , Humanos , Imuno-Histoquímica/métodos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Gravidez , Coloração e Rotulagem/métodos , Fatores de Tempo , Fixação de Tecidos/métodos
6.
BMC Cancer ; 19(1): 666, 2019 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-31277602

RESUMO

BACKGROUND: Cancer is a rapidly evolving, multifactorial disease that accumulates numerous genetic and epigenetic alterations. This results in molecular and phenotypic heterogeneity within the tumor, the complexity of which is further amplified through specific interactions between cancer cells. We aimed to dissect the molecular mechanisms underlying the cooperation between different clones. METHODS: We produced clonal cell lines derived from the MDA-MB-231 breast cancer cell line, using the UbC-StarTrack system, which allowed tracking of multiple clones by color: GFP C3, mKO E10 and Sapphire D7. Characterization of these clones was performed by growth rate, cell metabolic activity, wound healing, invasion assays and genetic and epigenetic arrays. Tumorigenicity was tested by orthotopic and intravenous injections. Clonal cooperation was evaluated by medium complementation, co-culture and co-injection assays. RESULTS: Characterization of these clones in vitro revealed clear genetic and epigenetic differences that affected growth rate, cell metabolic activity, morphology and cytokine expression among cell lines. In vivo, all clonal cell lines were able to form tumors; however, injection of an equal mix of the different clones led to tumors with very few mKO E10 cells. Additionally, the mKO E10 clonal cell line showed a significant inability to form lung metastases. These results confirm that even in stable cell lines heterogeneity is present. In vitro, the complementation of growth medium with medium or exosomes from parental or clonal cell lines increased the growth rate of the other clones. Complementation assays, co-growth and co-injection of mKO E10 and GFP C3 clonal cell lines increased the efficiency of invasion and migration. CONCLUSIONS: These findings support a model where interplay between clones confers aggressiveness, and which may allow identification of the factors involved in cellular communication that could play a role in clonal cooperation and thus represent new targets for preventing tumor progression.


Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Células Clonais/metabolismo , Heterogeneidade Genética , Animais , Apoptose , Comunicação Celular , Linhagem Celular Tumoral , Movimento Celular , Sobrevivência Celular , Células Clonais/patologia , Técnicas de Cocultura , Citocinas/análise , Elementos de DNA Transponíveis/genética , Feminino , Expressão Gênica , Xenoenxertos , Humanos , Camundongos , Camundongos Nus , Transplante de Neoplasias , Peixe-Zebra
7.
Nature ; 571(7765): 355-360, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31270458

RESUMO

Defining the transcriptomic identity of malignant cells is challenging in the absence of surface markers that distinguish cancer clones from one another, or from admixed non-neoplastic cells. To address this challenge, here we developed Genotyping of Transcriptomes (GoT), a method to integrate genotyping with high-throughput droplet-based single-cell RNA sequencing. We apply GoT to profile 38,290 CD34+ cells from patients with CALR-mutated myeloproliferative neoplasms to study how somatic mutations corrupt the complex process of human haematopoiesis. High-resolution mapping of malignant versus normal haematopoietic progenitors revealed an increasing fitness advantage with myeloid differentiation of cells with mutated CALR. We identified the unfolded protein response as a predominant outcome of CALR mutations, with a considerable dependency on cell identity, as well as upregulation of the NF-κB pathway specifically in uncommitted stem cells. We further extended the GoT toolkit to genotype multiple targets and loci that are distant from transcript ends. Together, these findings reveal that the transcriptional output of somatic mutations in myeloproliferative neoplasms is dependent on the native cell identity.


Assuntos
Genótipo , Mutação , Transtornos Mieloproliferativos/genética , Transtornos Mieloproliferativos/patologia , Neoplasias/genética , Neoplasias/patologia , Transcriptoma/genética , Animais , Antígenos CD34/metabolismo , Calreticulina/genética , Linhagem Celular , Proliferação de Células , Células Clonais/classificação , Células Clonais/metabolismo , Células Clonais/patologia , Endorribonucleases/metabolismo , Hematopoese/genética , Células-Tronco Hematopoéticas/classificação , Células-Tronco Hematopoéticas/metabolismo , Células-Tronco Hematopoéticas/patologia , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Camundongos , Modelos Moleculares , Transtornos Mieloproliferativos/classificação , NF-kappa B/metabolismo , Neoplasias/classificação , Células-Tronco Neoplásicas/citologia , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Mielofibrose Primária/genética , Mielofibrose Primária/patologia , Proteínas Serina-Treonina Quinases/metabolismo , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , Resposta a Proteínas não Dobradas/genética
8.
J Clin Lab Anal ; 33(7): e22951, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31187545

RESUMO

Del(20q) is the most frequently detected large structural genetic mosaicism alteration in the healthy aging population, occurring in approximately 0.1% of older persons. Age-related clonal hematopoiesis of copy number variations (CNVs) is linked to an increased risk of hematologic malignancies, but the clinical impact of hematopoietic stem cells (HSCs) harboring such CNVs, such as del(20q), is not clearly understood. Here, we report an acute myeloid leukemia (AML) patient with known del(20q) who acquired donor-derived del(20q) after allogeneic hematopoietic stem cell transplantation (HSCT). The HSCs with del(20q) engrafted and expanded over time, but the patient did not clinically progress to myeloid neoplasm. Although donor-derived del(20q) from a healthy donor has been reported previously, our case is the first to review the clonal dynamics of a del(20q) clone and its post-transplantation impact. Since recurrent hematology neoplasm-associated CNVs, including del(20q), may not be rare among aged HSCT donors, identifying the origin of such a CNV is necessary for clinical decisions when clonal abnormality appears after HSCT, even in recipients who previously had the same abnormality.


Assuntos
Deleção Cromossômica , Cromossomos Humanos Par 20/genética , Células Clonais/patologia , Transplante de Células-Tronco Hematopoéticas , Leucemia Mieloide Aguda/genética , Doadores de Tecidos , Humanos , Hibridização in Situ Fluorescente , Masculino , Pessoa de Meia-Idade , Transplante Homólogo
9.
PLoS One ; 14(3): e0213815, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30870501

RESUMO

Testicular germ cell tumors (TGCTs) are unique amongst solid tumors in terms of the high cure rates using chemotherapy for metastatic disease. Nevertheless, TGCTs still kill approximately 400 men per year, at a median age of 30 years, in the United States. This young age of mortality dramatically amplifies the impact of these deaths for the patients and their often young families. Furthermore the high cure rate makes it difficult to conduct further clinical trials of non curable disease. TGCTs are characterized by a marked aneuploidy and the presence of gain of chromosomal region 12p. Genomic testing may offer the ability to identify potentially lethal TGCTs at the time of initial diagnosis. However sequencing based studies have shown a paucity of somatic mutations in TGCT genomes including those that drive refractory disease. Furthermore these studies may be limited by genetic heterogeneity in primary tumors and the evolution of sub populations during disease progression. Herein we applied a systematic approach combining DNA content flow cytometry, whole genome copy number and whole exome sequence analyses to interrogate tumor heterogeneity in primary and metastatic refractory TGCTs. We identified both known and novel somatic copy number aberrations (12p, MDM2, and RHBDD1) and mutations (XRCC2, PIK3CA, RITA1) including candidate markers for platinum resistance that were present in a primary tumor of mixed histology and that remained after tandem autologous stem cell transplant.


Assuntos
Biomarcadores Tumorais/genética , Cisplatino/farmacologia , Células Clonais/patologia , Resistencia a Medicamentos Antineoplásicos/genética , Genoma Humano , Neoplasias Embrionárias de Células Germinativas/genética , Neoplasias Testiculares/genética , Antineoplásicos/farmacologia , Células Clonais/efeitos dos fármacos , Células Clonais/metabolismo , Humanos , Masculino , Metástase Neoplásica , Neoplasias Embrionárias de Células Germinativas/tratamento farmacológico , Neoplasias Embrionárias de Células Germinativas/patologia , Neoplasias Testiculares/tratamento farmacológico , Neoplasias Testiculares/patologia , Células Tumorais Cultivadas
10.
Methods Mol Biol ; 1866: 37-48, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30725406

RESUMO

Unlike normal cells, transformed cells are unable to grow when methionine in the growth media is restricted. Reversion to methionine independence is a rare event in transformed and malignant cells. Methionine-independent revertants provide an excellent system to identify metabolic signatures and molecular characteristics associated with methionine dependency of transformed cells. Revertants maintain the genetic background and general growth behavior of the parental cell line, except that they proliferate under methionine restriction such as in methionine-free media supplemented with homocysteine. Here we describe a general approach to generate methionine-independent revertants using the example of the triple-negative breast cancer cell line MDA-MB-468. To validate and characterize reversion we describe assays to evaluate cell proliferation and anchorage-independent growth in soft agar.


Assuntos
Separação Celular/métodos , Células Clonais/patologia , Metionina/farmacologia , Neoplasias/patologia , Adesão Celular/efeitos dos fármacos , Linhagem Celular Transformada , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Humanos , Medições Luminescentes
11.
Methods Mol Biol ; 1956: 77-103, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30779031

RESUMO

Assessment of the presence of clonal lymphoproliferations via polymerase chain reaction (PCR)-based analysis of rearranged immunoglobulin (IG) or T-cell receptor (TR) genes is a valuable method in the diagnosis of suspect lymphoproliferative disorders. Additionally, this methodology can be used for evaluating dissemination of lymphoma cells and for studying the clonal relationship between multiple (different locations) or consecutive (over time) lymphomas. Here we describe an integrated approach to assess clonality via analysis of Ig heavy chain (IGH), Ig kappa (IGK), TCR beta (TRB), and TCR gamma (TRG) gene rearrangements, based on the standardized multiplex PCRs as originally developed by the European BIOMED-2 consortium. The described protocol covers the pre-analytical phase of DNA isolation (from formalin-fixed paraffin-embedded and fresh tissues, body fluids, peripheral blood, and bone marrow), the analytical phase of PCR GeneScan and heteroduplex analysis, and the post-analytical interpretation of the obtained profiles, following established guidelines.


Assuntos
Rearranjo Gênico , Análise Heteroduplex/métodos , Transtornos Linfoproliferativos/genética , Reação em Cadeia da Polimerase/métodos , Receptores de Antígenos de Linfócitos T/genética , Linfócitos B/metabolismo , Linfócitos B/patologia , Sequência de Bases , Células Clonais/metabolismo , Células Clonais/patologia , DNA/genética , DNA/isolamento & purificação , Genes Codificadores dos Receptores de Linfócitos T , Humanos , Cadeias Pesadas de Imunoglobulinas/genética , Imunoglobulinas/genética , Transtornos Linfoproliferativos/diagnóstico , Transtornos Linfoproliferativos/patologia , Linfócitos T/metabolismo , Linfócitos T/patologia
12.
Med Sci (Paris) ; 35(2): 187-190, 2019 Feb.
Artigo em Francês | MEDLINE | ID: mdl-30774090

RESUMO

Careful sequencing studies on small samples of normal oesophageal epithelium reveal the presence of very abundant cellular clones harbouring mutations in known cancer genes (and elsewhere). The number and size of these clones increases with age. This surprising finding confirms previous studies on sun-exposed epidermis. It has important implications for the understanding of cancer initiation and will hopefully lead to conceptual and clinical advances.


Assuntos
Células Clonais/patologia , Epitélio/patologia , Células-Tronco Neoplásicas/patologia , Lesões Pré-Cancerosas/patologia , Nicho de Células-Tronco , Envelhecimento/patologia , Células Clonais/metabolismo , Células Epidérmicas/metabolismo , Células Epidérmicas/patologia , Humanos , Mutação/fisiologia , Lesões Pré-Cancerosas/genética , Neoplasias Cutâneas/etiologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Nicho de Células-Tronco/genética , Luz Solar/efeitos adversos
13.
Nature ; 565(7739): 312-317, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30602793

RESUMO

Clonal expansion in aged normal tissues has been implicated in the development of cancer. However, the chronology and risk dependence of the expansion are poorly understood. Here we intensively sequence 682 micro-scale oesophageal samples and show, in physiologically normal oesophageal epithelia, the progressive age-related expansion of clones that carry mutations in driver genes (predominantly NOTCH1), which is substantially accelerated by alcohol consumption and by smoking. Driver-mutated clones emerge multifocally from early childhood and increase their number and size with ageing, and ultimately replace almost the entire oesophageal epithelium in the extremely elderly. Compared with mutations in oesophageal cancer, there is a marked overrepresentation of NOTCH1 and PPM1D mutations in physiologically normal oesophageal epithelia; these mutations can be acquired before late adolescence (as early as early infancy) and significantly increase in number with heavy smoking and drinking. The remodelling of the oesophageal epithelium by driver-mutated clones is an inevitable consequence of normal ageing, which-depending on lifestyle risks-may affect cancer development.


Assuntos
Envelhecimento/genética , Envelhecimento/patologia , Epitélio , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/patologia , Mutação , Lesões Pré-Cancerosas/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Consumo de Bebidas Alcoólicas/genética , Biópsia , Contagem de Células , Transformação Celular Neoplásica/genética , Criança , Pré-Escolar , Células Clonais/metabolismo , Células Clonais/patologia , Variações do Número de Cópias de DNA , Epitélio/metabolismo , Epitélio/patologia , Evolução Molecular , Feminino , Interação Gene-Ambiente , Genoma Humano/genética , Humanos , Lactente , Estilo de Vida , Masculino , Pessoa de Meia-Idade , Acúmulo de Mutações , Proteína Fosfatase 2C/genética , Receptor Notch1/genética , Fatores de Risco , Análise de Sequência de DNA , Análise de Célula Única , Fumar/genética , Adulto Jovem
14.
Leukemia ; 33(1): 266-270, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30026571

RESUMO

Multiple myeloma (MM) patients have an 11-fold increased risk of developing myeloid neoplasms compared to the general population; however, acute lymphoblastic leukemia (ALL) is rarely observed. Given that both MM and the majority of ALL are of B cell origin, this raises the question of whether ALL in patients with MM arises from the same clone. We report 13 cases of B-cell ALL following therapy for MM. The interval from MM diagnosis to ALL onset was 5.4 years (range 3.3-10). The median age at the time of ALL diagnosis was 60 years (range 43-67). MM therapy included immunomodulatory agents in all patients and autologous hematopoietic cell transplantation in 10 (77%) patients preceding ALL diagnosis. ALL genetics showed a normal karyotype, TP53 mutation/deletion, and monosomy 7 or 7q deletion in 5, 3, and 2 cases, respectively. Analysis of paired samples of MM and ALL using whole exome sequencing demonstrated that the malignancies arose from different clones. Thus, ALL as a second primary malignancy following MM is not clonally related but could potentially represent a therapy-related leukemia.


Assuntos
Aberrações Cromossômicas , Células Clonais/patologia , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Fatores Imunológicos/efeitos adversos , Mieloma Múltiplo/terapia , Segunda Neoplasia Primária/etiologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/etiologia , Adulto , Idoso , Terapia Combinada , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/patologia , Segunda Neoplasia Primária/patologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Prognóstico , Transplante Homólogo
15.
Melanoma Res ; 29(1): 89-94, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30222690

RESUMO

The management of metastatic melanoma is a difficult matter. Nevertheless, the advent of target therapy has significantly improved patient outcome, provided that tumor molecular characteristics become available: the detection of drug-resistant clones can contribute to understanding the reasons for resistance onset, influencing the choice of subsequent therapy. This work aimed to provide a possible explanation for the early resistance to vemurafenib developed by a patient with melanoma, and concurrently to assess the extent, and role, of the tumor clonal heterogeneity. We analyzed tissue samples from different sites and time points: first/second primary, three lymph node metastases, and circulating melanoma cells (CMCs). We first investigated these samples by the routine Sanger sequencing for BRAF, NRAS, and KIT, and then, we focused on specific hotspots by droplet digital PCR. We detected a BRAF V600E mutation by Sanger sequencing in the second primary and distant lymph node metastases, but not in the first primary or sentinel lymph node. Interestingly, by droplet digital PCR, the V600E mutation was also detected in the first primary, and the V600K in the second primary and metastases. Moreover, we identified a rare KIT V569G mutation, appearing to be CMC exclusive. This finding confirms the potential of CMCs as a source of tumor material for genetic analysis, reflecting real-time systemic disease evolution and, most likely, the most aggressive, treatment-resistant clones. In summary, this work underlines the importance of CMCs in the early identification of tumor clones putatively responsible for therapy resistance.


Assuntos
Células Clonais/patologia , Melanoma/patologia , Células Neoplásicas Circulantes/patologia , Neoplasias Cutâneas/patologia , Humanos , Masculino , Melanoma/tratamento farmacológico , Melanoma/genética , Pessoa de Meia-Idade , Mutação , Prognóstico , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/genética
17.
Int J Hematol ; 109(2): 221-227, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30368656

RESUMO

Adult T-cell leukemia (ATL) is an aggressive mature T-cell malignancy with a poor prognosis. The anti-C-C motif chemokine receptor 4 (CCR4) antibody mogamulizumab (moga) reduces ATL cells and induces reconstitution of polyclonal T cells; however, ATL cases often remain resistant and moga sometimes causes fatal immunopathology. Epstein-Barr virus (EBV)-related B-cell lymphoma develops in severely immunocompromised subjects, and is particularly associated with impaired T-cell immunity. Here, we report an ATL patient who had received conventional chemotherapy plus moga, and subsequently developed EBV-related diffuse large B-cell lymphoma (DLBCL) of the central nervous system. Next-generation sequencing-based T-cell receptor repertoire analyses identified residual abnormal clones and revealed that reconstitution of polyclonal T cells was incomplete, even after moga treatment. Furthermore, a skin rash that developed after moga treatment was found to contain ATL clones. This case suggests that the limited therapeutic effects of moga and incomplete T-cell reconstitution are associated with severely impaired T-cell immunity and subsequent development of EBV-related DLBCL.


Assuntos
Herpesvirus Humano 4 , Linfoma Difuso de Grandes Células B/etiologia , Adulto , Anticorpos Monoclonais Humanizados/uso terapêutico , Neoplasias do Sistema Nervoso Central , Criança , Células Clonais/patologia , Humanos , Leucemia-Linfoma de Células T do Adulto/complicações , Leucemia-Linfoma de Células T do Adulto/tratamento farmacológico , Linfoma Difuso de Grandes Células B/virologia , Linfócitos T/imunologia , Linfócitos T/patologia
18.
Exp Mol Pathol ; 106: 78-88, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30503404

RESUMO

Uterine endometrial carcinoma is one of the common cancers in females. Cancer stem-like cells (CSCs)/cancer-initiating cells (CICs) are a small subpopulation of cancer cells that are tumorigenic and are resistant to treatments, thus they are focused as treatment targets. However, the heterogeneity of CSCs/CICs is still elusive, and we therefore analyzed CSCs/CICs at the clonal level. We previously established sphere-cultured CSCs/CICs from primary human uterine endometrial carcinoma, and we isolated several clones from CSCs/CICs in this study. Interestingly, we established two types of clones based on the growth pattern. The clones were termed sphere clones (S clones) and leukemia-like clones (LL clones). Functional analysis revealed that S clones are resistant to chemotherapy, whereas LL clones are sensitive to chemotherapy. On the other hand, S clones are less tumorigenic, while LL clones are highly tumorigenic. Transcriptome analysis using serial analysis of gene expression sequencing (SAGE-Seq) revealed distinctive gene expression profiles in S clone cells and LL clone cells. The results indicate that CSCs/CICs are composed of functionally heterogenic subpopulations including highly tumorigenic clones and treatment-resistant clones and that the characteristics of CSCs/CICs might be determined by the characteristics of different clones that compose CSCs/CICs.


Assuntos
Carcinoma Endometrioide/patologia , Neoplasias do Endométrio/patologia , Células-Tronco Neoplásicas/patologia , Animais , Carboplatina/farmacologia , Carcinoma Endometrioide/genética , Células Clonais/patologia , Meios de Cultura , DNA de Neoplasias/genética , Neoplasias do Endométrio/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Xenoenxertos , Humanos , Camundongos , Células-Tronco Neoplásicas/efeitos dos fármacos , Paclitaxel/farmacologia , Fenótipo , Análise de Sequência de DNA , Soro , Esferoides Celulares , Células Tumorais Cultivadas
19.
Int J Mol Sci ; 19(12)2018 Dec 03.
Artigo em Inglês | MEDLINE | ID: mdl-30513905

RESUMO

Recent advances in the field of cancer genome analysis revolutionized the picture we have of acute myeloid leukemia (AML). Pan-genomic studies, using either single nucleotide polymorphism arrays or whole genome/exome next generation sequencing, uncovered alterations in dozens of new genes or pathways, intimately connected with the development of leukemia. From a simple two-hit model in the late nineties, we are now building clonal stories that involve multiple unexpected cellular functions, leading to full-blown AML. In this review, we will address several seminal concepts that result from these new findings. We will describe the genetic landscape of AML, the association and order of events that define multiple sub-entities, both in terms of pathogenesis and in terms of clinical practice. Finally, we will discuss the use of this knowledge in the settings of new strategies for the evaluation of measurable residual diseases (MRD), using clone-specific multiple molecular targets.


Assuntos
Células Clonais/patologia , Hematopoese/genética , Leucemia Mieloide Aguda/genética , Neoplasia Residual/genética , Predisposição Genética para Doença , Humanos , Leucemia Mieloide Aguda/classificação , Modelos Biológicos
20.
Elife ; 72018 11 30.
Artigo em Inglês | MEDLINE | ID: mdl-30499773

RESUMO

Tumors often co-exist with T cells that recognize somatically mutated peptides presented by cancer cells on major histocompatibility complex I (MHC-I). However, it is unknown why the immune system fails to eliminate immune-recognizable neoplasms before they manifest as frank disease. To understand the determinants of MHC-I peptide immunogenicity in nascent tumors, we tested the ability of thousands of MHC-I ligands to cause tumor subclone rejection in immunocompetent mice by use of a new 'PresentER' antigen presentation platform. Surprisingly, we show that immunogenic tumor antigens do not lead to immune-mediated cell rejection when the fraction of cells bearing each antigen ('clonal fraction') is low. Moreover, the clonal fraction necessary to lead to rejection of immunogenic tumor subclones depends on the antigen. These data indicate that tumor neoantigen heterogeneity has an underappreciated impact on immune elimination of cancer cells and has implications for the design of immunotherapeutics such as cancer vaccines.


Assuntos
Células Clonais/patologia , Neoplasias/imunologia , Neoplasias/patologia , Animais , Apresentação do Antígeno/imunologia , Antígenos de Neoplasias/imunologia , Sequência de Bases , Efeito Espectador , Linhagem Celular Tumoral , Citotoxicidade Imunológica , Biblioteca Gênica , Imunocompetência , Complexo Principal de Histocompatibilidade/imunologia , Camundongos Endogâmicos C57BL , Peptídeos/imunologia , Receptores de Quinase C Ativada/imunologia , Linfócitos T/imunologia , Vacinação
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