RESUMEN
NUT midline carcinoma (NMC), a subtype of squamous cell cancer, is one of the most aggressive human solid malignancies known. NMC is driven by the creation of a translocation oncoprotein, BRD4-NUT, which blocks differentiation and drives growth of NMC cells. BRD4-NUT forms distinctive nuclear foci in patient tumors, which we found correlate with â¼100 unprecedented, hyperacetylated expanses of chromatin that reach up to 2 Mb in size. These "megadomains" appear to be the result of aberrant, feed-forward loops of acetylation and binding of acetylated histones that drive transcription of underlying DNA in NMC patient cells and naïve cells induced to express BRD4-NUT. Megadomain locations are typically cell lineage-specific; however, the cMYC and TP63 regions are targeted in all NMCs tested and play functional roles in tumor growth. Megadomains appear to originate from select pre-existing enhancers that progressively broaden but are ultimately delimited by topologically associating domain (TAD) boundaries. Therefore, our findings establish a basis for understanding the powerful role played by large-scale chromatin organization in normal and aberrant lineage-specific gene transcription.
Asunto(s)
Carcinoma de Células Escamosas/fisiopatología , Regulación Neoplásica de la Expresión Génica , Proteínas Nucleares/metabolismo , Proteínas Oncogénicas/metabolismo , Factores de Transcripción/metabolismo , Proteínas de Ciclo Celular , Línea Celular Tumoral , Elementos de Facilitación Genéticos , Humanos , Proteínas de Neoplasias , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Estructura Terciaria de Proteína , Factores de Transcripción/genéticaRESUMEN
Molibresib is an orally bioavailable, selective, small molecule BET protein inhibitor. Results from a first time in human study in solid tumors resulted in the selection of a 75 mg once daily dose of the besylate formulation of molibresib as the recommended Phase 2 dose (RP2D). Here we present the results of Part 2 of our study, investigating safety, pharmacokinetics, pharmacodynamics and clinical activity of molibresib at the RP2D for nuclear protein in testis carcinoma (NC), small cell lung cancer, castration-resistant prostate cancer (CRPC), triple-negative breast cancer, estrogen receptor-positive breast cancer and gastrointestinal stromal tumor. The primary safety endpoints were incidence of adverse events (AEs) and serious AEs; the primary efficacy endpoint was overall response rate. Secondary endpoints included plasma concentrations and gene set enrichment analysis (GSEA). Molibresib 75 mg once daily demonstrated no unexpected toxicities. The most common treatment-related AEs (any grade) were thrombocytopenia (64%), nausea (43%) and decreased appetite (37%); 83% of patients required dose interruptions and 29% required dose reductions due to AEs. Antitumor activity was observed in NC and CRPC (one confirmed partial response each, with observed reductions in tumor size), although predefined clinically meaningful response rates were not met for any tumor type. Total active moiety median plasma concentrations after single and repeated administration were similar across tumor cohorts. GSEA revealed that gene expression changes with molibresib varied by patient, response status and tumor type. Investigations into combinatorial approaches that use BET inhibition to eliminate resistance to other targeted therapies are warranted.
Asunto(s)
Benzodiazepinas/uso terapéutico , Neoplasias/tratamiento farmacológico , Proteínas Nucleares/metabolismo , Neoplasias Testiculares/tratamiento farmacológico , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Benzodiazepinas/administración & dosificación , Benzodiazepinas/efectos adversos , Benzodiazepinas/farmacocinética , Femenino , Humanos , Masculino , Persona de Mediana Edad , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Receptores de Superficie Celular/antagonistas & inhibidores , Adulto JovenRESUMEN
BACKGROUND: Bromodomain and extra-terminal (BET) proteins are epigenetic readers that can drive carcinogenesis and therapy resistance. RO6870810 is a novel, small-molecule BET inhibitor. METHODS: We conducted a Phase 1 study of RO6870810 administered subcutaneously for 21 or 14 days of 28- or 21-day cycles, respectively, in patients with the nuclear protein of the testis carcinoma (NC), other solid tumours, or diffuse large B-cell lymphoma (DLBCL) with MYC deregulation. RESULTS: Fatigue (42%), decreased appetite (35%) and injection-site erythema (35%) were the most common treatment-related adverse events. Pharmacokinetic parameters demonstrated linearity over the dose range tested and support once-daily dosing. Pharmacodynamic assessments demonstrated sustained decreases in CD11b levels in peripheral blood mononuclear cells. Objective response rates were 25% (2/8), 2% (1/47) and 11% (2/19) for patients with NC, other solid tumours and DLBCL, respectively. Responding tumours had evidence of deregulated MYC expression. CONCLUSIONS: This trial establishes the safety, favourable pharmacokinetics, evidence of target engagement and preliminary single-agent activity of RO6870810. Responses in patients with NC, other solid tumours and DLBCL provide proof-of-principle for BET inhibition in MYC-driven cancers. The results support further exploration of RO6870810 as monotherapy and in combinations. CLINICAL TRIALS REGISTRATION: NCT01987362.
Asunto(s)
Azepinas/administración & dosificación , Azepinas/efectos adversos , Linfoma de Células B Grandes Difuso/tratamiento farmacológico , Proteínas de Neoplasias/metabolismo , Neoplasias/tratamiento farmacológico , Proteínas Nucleares/metabolismo , Proteínas/antagonistas & inhibidores , Adulto , Anciano , Anciano de 80 o más Años , Azepinas/sangre , Azepinas/farmacocinética , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Linfoma de Células B Grandes Difuso/sangre , Masculino , Persona de Mediana Edad , Neoplasias/sangre , Neoplasias/metabolismo , Bibliotecas de Moléculas Pequeñas/administración & dosificación , Bibliotecas de Moléculas Pequeñas/efectos adversos , Bibliotecas de Moléculas Pequeñas/farmacocinéticaRESUMEN
To investigate the mechanism that drives dramatic mistargeting of active chromatin in NUT midline carcinoma (NMC), we have identified protein interactions unique to the BRD4-NUT fusion oncoprotein compared with wild-type BRD4. Using cross-linking, affinity purification, and mass spectrometry, we identified the EP300 acetyltransferase as uniquely associated with BRD4 through the NUT fusion in both NMC and non-NMC cell types. We also discovered ZNF532 associated with BRD4-NUT in NMC patient cells but not detectable in 293T cells. EP300 and ZNF532 are both implicated in feed-forward regulatory loops leading to propagation of the oncogenic chromatin complex in BRD4-NUT patient cells. Adding key functional significance to our biochemical findings, we independently discovered a ZNF532-NUT translocation fusion in a newly diagnosed NMC patient. ChIP sequencing of the major players NUT, ZNF532, BRD4, EP300, and H3K27ac revealed the formation of ZNF532-NUT-associated hyperacetylated megadomains, distinctly localized but otherwise analogous to those found in BRD4-NUT patient cells. Our results support a model in which NMC is dependent on ectopic NUT-mediated interactions between EP300 and components of BRD4 regulatory complexes, leading to a cascade of misregulation.
Asunto(s)
Carcinoma de Células Escamosas/patología , Cromatina/metabolismo , Proteína p300 Asociada a E1A/metabolismo , Neoplasias Pulmonares/patología , Proteínas Nucleares/metabolismo , Proteínas de Fusión Oncogénica/genética , Proteínas Oncogénicas/metabolismo , Proteínas Represoras/metabolismo , Factores de Transcripción/metabolismo , Carcinoma de Células Escamosas/genética , Proteínas de Ciclo Celular , Línea Celular Tumoral , Proliferación Celular/genética , Células Epiteliales/patología , Femenino , Células HEK293 , Humanos , Hibridación Fluorescente in Situ , Neoplasias Pulmonares/genética , Persona de Mediana Edad , Complejos Multiproteicos/genética , Proteínas de Neoplasias , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Dominios Proteicos/genética , Interferencia de ARN , ARN Interferente Pequeño/genética , Proteínas Represoras/genética , Factores de Transcripción/genética , Dedos de Zinc/genéticaRESUMEN
NUT carcinoma (NC) shows very aggressive clinical behavior, occurs predominantly in the thorax and head and neck region of children and adults, and is defined by the presence of NUT (aka NUTM1) rearrangement, mostly BRD4-NUTM1 fusion resulting from t(15;19)(q13; p13.1). So-called "NUT variants" harbor alternate fusions between NUTM1 and BRD3, NSD3, ZNF532, or unknown partners. Rare cases of pediatric tumors with CIC-NUTM1 fusion were recently reported in somatic soft tissue, brain, and kidney. However, such cases have not been identified in adult patients and the presence of a fusion between CIC, characteristic of CIC-rearranged sarcoma, and NUTM1-a defining feature of NC-poses a diagnostic challenge. We herein report a case of malignant epithelioid neoplasm with myoepithelial features harboring CIC-NUTM1 fusion arising in soft tissue of the head in a 60-year-old man. Immunohistochemistry revealed strong expression of NUT, but only weak ETV4 staining and negativity for keratins, EMA, p40, CD99, and WT1. SMARCB1 expression was retained. Fluorescence in situ hybridization and targeted next-generation sequencing identified a CIC-NUTM1 fusion resulting from t(15;19)(q14;q13.2). In light of morphologic features that overlap with those of NC from typical anatomical sites we have seen previously, the tumor was best classified as falling within the NC spectrum rather than CIC-associated sarcoma. This case highlights the emerging diagnostic challenges generated by newly detected gene fusions of unknown clinical and biologic significance. Careful integration of cytogenetic, molecular, and immunohistochemical findings with morphologic appearances in the diagnostic workup of undifferentiated neoplasms is essential.
Asunto(s)
Biomarcadores de Tumor/genética , Canales de Cloruro/genética , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Sarcoma/genética , Proteínas de Ciclo Celular , Femenino , Reordenamiento Génico/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Hibridación Fluorescente in Situ , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias , Proteínas de Fusión Oncogénica/genética , Sarcoma/diagnóstico , Sarcoma/patología , Factores de Transcripción/genéticaRESUMEN
NUT carcinoma (NC) is a rare, aggressive subtype of squamous cell carcinoma defined by rearrangement of the NUTM1 (aka NUT) gene. NC is driven by NUT-fusion oncoproteins resulting from chromosomal translocation, most commonly BRD4-NUT. This is a nearly uniformly lethal cancer affecting patients of all ages, but predominantly teens and young adults. The cell of origin is unknown, but NC most commonly arises within the thorax and head and neck. NC typically consists of sheets of monomorphic primitive round cells that can exhibit focal abrupt squamous differentiation. Diagnosis of NC is easy, and can be established by positive NUT nuclear immunohistochemical staining. Though characterization of the NUTM1-fusion gene is desirable by molecular analysis, it is not required for the diagnosis. The increasingly widespread availability of the NUT diagnostic test is leading to increasing diagnoses of this vastly underdiagnosed disease. The NUT midline carcinoma registry (www.NMCRegistry.org) serves as a central repository that has provided the main source of clinical and outcomes data for NC. Currently there is no effective therapy for NC, however small molecules directly targeting the BRD4 portion of BRD4-NUT, termed BET bromodomain inhibitors, have shown activity.
Asunto(s)
Carcinoma de Células Escamosas , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Humanos , Proteínas de Neoplasias , Proteínas de Fusión Oncogénica/genéticaRESUMEN
AIMS: NUT midline carcinoma (NMC) is a rare undifferentiated and aggressive carcinoma that locates characteristically to the midline of the head and neck, and mediastinum. NMC is characterized by chromosomal rearrangements of the gene NUT, at 15q14. The BRD4 gene on 19q13 is the most common translocation partner forming a fusion oncogene, BRD4-NUT. By the end of 2014, the International NUT Midline Carcinoma Registry had 48 patients treated for NMC. Laryngeal NMC are exceedingly rare, and we report a case series of seven cases. METHODS AND RESULTS: We searched for cases in files of different hospitals as well as a thorough search of the English language literature. The diagnosis of NMC is made by demonstration of NUT rearrangement either by immunohistochemistry, fluorescence in-situ hybridization (FISH) or reverse transcription-polymerase chain reaction (RT-PCR). We found three previously published cases, and in this series add four cases of our own. CONCLUSIONS: NMC consists of monomorphic, often discohesive, cells with an epithelioid appearance and distinct nucleoli. The tumours typically show abrupt squamous differentiation. The mean age of the patients was 34 years, hence significantly lower than that for conventional laryngeal carcinoma. All tumours were located in the supraglottis and five patients died of the disease after 3, 7, 8, 9 and 11 months. Laryngeal NMC may be underdiagnosed, and an increased awareness among pathologists is warranted. NMC has characteristic morphological features, and positive immunostaining with the NUT antibody is diagnostic. Its aggressive behaviour demands a very intense treatment strategy and the need for its recognition is emphasized further by new promising treatment strategies.
Asunto(s)
Carcinoma/patología , Neoplasias Laríngeas/patología , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Adolescente , Adulto , Anciano , Carcinoma/diagnóstico , Carcinoma/genética , Niño , Preescolar , Femenino , Humanos , Neoplasias Laríngeas/diagnóstico , Neoplasias Laríngeas/genética , Masculino , Persona de Mediana Edad , Proteínas de NeoplasiasRESUMEN
A subset of poorly differentiated squamous cell carcinomas, NUT midline carcinomas (NMC) are characterized by a translocation t(15;19)(q13;p13) [ 1 ]. The prognosis is generally dismal [ 2 ] and therapeutic success has been limited to exceptional cases [ 3 ]. We present two cases of pediatric NMC from two different institutions treated according to a multimodal sarcoma approach involving surgery, chemotherapy, and focal radiotherapy. One patient has remained in complete continuous remission for over 6 years, while the other is in CR in early follow-up off therapy. Our proposed multimodal strategy apparently meets the aggressive biologic nature of NMC and should be considered for further evaluation in this context potentially in the setting of a clinical trial.
Asunto(s)
Carcinoma de Células Escamosas/terapia , Neoplasias de la Lengua/terapia , Translocación Genética , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patología , Niño , Cromosomas Humanos Par 15/genética , Cromosomas Humanos Par 19/genética , Terapia Combinada/métodos , Humanos , Masculino , Neoplasias de la Lengua/genética , Neoplasias de la Lengua/patologíaRESUMEN
BACKGROUND: NUT midline carcinoma is a rare and aggressive genetically characterized subtype of squamous cell carcinoma frequently arising from the head and neck. The characteristics and optimal management of head and neck NUT midline carcinoma (HNNMC) are unclear. METHODS: A retrospective review of all known cases of HNNMC in the International NUT Midline Carcinoma Registry as of December 31, 2014, was performed. Forty-eight consecutive patients were treated from 1993 to 2014, and clinicopathologic variables and outcomes for 40 patients were available for analyses; they composed the largest HNNMC cohort studied to date. Overall survival (OS) and progression-free survival (PFS) according to patient characteristics and treatment were analyzed. RESULTS: This study identified a 5-fold increase in the diagnosis of HNNMC from 2011 to 2014. The median age was 21.9 years (range, 0.1-81.7 years); the male and female proportions were 40% and 60%, respectively; and 86% had bromodomain containing 4-nuclear protein in testis (BRD4-NUT) fusion. The initial treatment was initial surgery with or without adjuvant chemoradiation or adjuvant radiation (56%), initial radiation with or without chemotherapy (15%), or initial chemotherapy with or without surgery or radiation (28%). The median PFS was 6.6 months (range, 4.7-8.4 months). The median OS was 9.7 months (range, 6.6-15.6 months). The 2-year PFS rate was 26% (95% confidence interval [CI], 13%-40%). The 2-year OS rate was 30% (95% CI, 16%-46%). Initial surgery with or without postoperative chemoradiation or radiation (P = .04) and complete resection with negative margins (P = .01) were significant predictors of improved OS even after adjustments for age, tumor size, and neck lymphadenopathy. Initial radiation or chemotherapy and the NUT translocation type were not associated with outcomes. CONCLUSIONS: HNNMC portends a poor prognosis. Aggressive initial surgical resection with or without postoperative chemoradiation or radiation is associated with significantly enhanced survival. Chemotherapy or radiation alone is often inadequate. Cancer 2016;122:3632-40. © 2016 American Cancer Society.
Asunto(s)
Carcinoma/mortalidad , Carcinoma/terapia , Neoplasias de Cabeza y Cuello/mortalidad , Neoplasias de Cabeza y Cuello/terapia , Proteínas Nucleares/genética , Proteínas de Fusión Oncogénica/genética , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Carcinoma/genética , Carcinoma/patología , Quimioradioterapia/métodos , Niño , Preescolar , Supervivencia sin Enfermedad , Femenino , Neoplasias de Cabeza y Cuello/genética , Neoplasias de Cabeza y Cuello/patología , Humanos , Lactante , Recién Nacido , Masculino , Persona de Mediana Edad , Cuello/patología , Estudios Retrospectivos , Tasa de Supervivencia , Adulto JovenRESUMEN
Epigenetic proteins are intently pursued targets in ligand discovery. So far, successful efforts have been limited to chromatin modifying enzymes, or so-called epigenetic 'writers' and 'erasers'. Potent inhibitors of histone binding modules have not yet been described. Here we report a cell-permeable small molecule (JQ1) that binds competitively to acetyl-lysine recognition motifs, or bromodomains. High potency and specificity towards a subset of human bromodomains is explained by co-crystal structures with bromodomain and extra-terminal (BET) family member BRD4, revealing excellent shape complementarity with the acetyl-lysine binding cavity. Recurrent translocation of BRD4 is observed in a genetically-defined, incurable subtype of human squamous carcinoma. Competitive binding by JQ1 displaces the BRD4 fusion oncoprotein from chromatin, prompting squamous differentiation and specific antiproliferative effects in BRD4-dependent cell lines and patient-derived xenograft models. These data establish proof-of-concept for targeting protein-protein interactions of epigenetic 'readers', and provide a versatile chemical scaffold for the development of chemical probes more broadly throughout the bromodomain family.
Asunto(s)
Azirinas/farmacología , Dihidropiridinas/farmacología , Modelos Moleculares , Proteínas Nucleares/antagonistas & inhibidores , Proteínas Nucleares/metabolismo , Factores de Transcripción/antagonistas & inhibidores , Factores de Transcripción/metabolismo , Secuencia de Aminoácidos , Animales , Azirinas/síntesis química , Azirinas/química , Sitios de Unión , Carcinoma de Células Escamosas/fisiopatología , Proteínas de Ciclo Celular , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cromatina/metabolismo , Dihidropiridinas/síntesis química , Dihidropiridinas/química , Femenino , Humanos , Ratones , Ratones Desnudos , Datos de Secuencia Molecular , Unión Proteica/efectos de los fármacos , Estructura Terciaria de Proteína , Proteínas Recombinantes/metabolismo , Alineación de Secuencia , Neoplasias Cutáneas/fisiopatología , EstereoisomerismoRESUMEN
14-3-3 proteins are ubiquitously expressed regulators of various cellular functions, including proliferation, metabolism, and differentiation, and altered 14-3-3 expression is associated with development and progression of cancer. We report a transforming 14-3-3 oncoprotein, which we identified through conventional cytogenetics and whole-transcriptome sequencing analysis as a highly recurrent genetic mechanism in a clinically aggressive form of uterine sarcoma: high-grade endometrial stromal sarcoma (ESS). The 14-3-3 oncoprotein results from a t(10;17) genomic rearrangement, leading to fusion between 14-3-3ε (YWHAE) and either of two nearly identical FAM22 family members (FAM22A or FAM22B). Expression of YWHAE-FAM22 fusion oncoproteins was demonstrated by immunoblot in t(10;17)-bearing frozen tumor and cell line samples. YWHAE-FAM22 fusion gene knockdowns were performed with shRNAs and siRNAs targeting various FAM22A exons in an t(10;17)-bearing ESS cell line (ESS1): Fusion protein expression was inhibited, with corresponding reduction in cell growth and migration. YWHAE-FAM22 maintains a structurally and functionally intact 14-3-3ε (YWHAE) protein-binding domain, which is directed to the nucleus by a FAM22 nuclear localization sequence. In contrast to classic ESS, harboring JAZF1 genetic fusions, YWHAE-FAM22 ESS display high-grade histologic features, a distinct gene-expression profile, and a more aggressive clinical course. Fluorescence in situ hybridization analysis demonstrated absolute specificity of YWHAE-FAM22A/B genetic rearrangement for high-grade ESS, with no fusions detected in other uterine and nonuterine mesenchymal tumors (55 tumor types, n = 827). These discoveries reveal diagnostically and therapeutically relevant models for characterizing aberrant 14-3-3 oncogenic functions.
Asunto(s)
Proteínas 14-3-3/metabolismo , Proteínas de Fusión Oncogénica/metabolismo , Sarcoma Estromático Endometrial/metabolismo , Sarcoma Estromático Endometrial/patología , Proteínas 14-3-3/genética , Núcleo Celular/metabolismo , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/patología , Cromosomas Humanos Par 10/genética , Cromosomas Humanos Par 17/genética , Proteínas Co-Represoras , Análisis Citogenético , Proteínas de Unión al ADN , Progresión de la Enfermedad , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Reordenamiento Génico/genética , Genoma Humano/genética , Humanos , Datos de Secuencia Molecular , Clasificación del Tumor , Proteínas de Neoplasias/metabolismo , Unión Proteica , Transporte de Proteínas , Sarcoma Estromático Endometrial/genética , Análisis de Secuencia de ADN , Transcriptoma , Translocación GenéticaRESUMEN
In a subset of poorly differentiated and highly aggressive carcinoma, a chromosomal translocation, t(15;19)(q13;p13), results in an in-frame fusion of the double bromodomain protein, BRD4, with a testis-specific protein of unknown function, NUT (nuclear protein in testis). In this study, we show that, after binding to acetylated chromatin through BRD4 bromodomains, the NUT moiety of the fusion protein strongly interacts with and recruits p300, stimulates its catalytic activity, initiating cycles of BRD4-NUT/p300 recruitment and creating transcriptionally inactive hyperacetylated chromatin domains. Using a patient-derived cell line, we show that p300 sequestration into the BRD4-NUT foci is the principal oncogenic mechanism leading to p53 inactivation. Knockdown of BRD4-NUT released p300 and restored p53-dependent regulatory mechanisms leading to cell differentiation and apoptosis. This study demonstrates how the off-context activity of a testis-specific factor could markedly alter vital cellular functions and significantly contribute to malignant cell transformation.
Asunto(s)
Cromatina/metabolismo , Proteína p300 Asociada a E1A/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Oncogénicas/metabolismo , Factores de Transcripción/metabolismo , Acetilación , Animales , Western Blotting , Células COS , Proteínas de Ciclo Celular , Línea Celular Tumoral , Chlorocebus aethiops , Humanos , Microscopía Fluorescente , Proteínas de Neoplasias , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Unión Proteica , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Recombinación Genética , Factores de Transcripción/genética , Translocación Genética , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
We present the first known case of a patient with BRD2::NUTM1-driven NUT carcinoma. A 59-year-old woman presented with poorly differentiated squamous cell lung cancer metastatic to the pleura. Eventually, a positive NUT immunohistochemistry, NUT fluorescence in situ hybridization, and RNA next-generation sequencing with a BRD2::NUTM1 fusion led to the diagnosis of NUT carcinoma. She received multiple lines of chemotherapy with response and is still alive at 2 years postdiagnosis. This report expands on the known fusions in NUT carcinoma and highlights potential differences in patient prognosis on the basis of gene fusion partners.
RESUMEN
INTRODUCTION: NUT carcinoma (NC) is an underdiagnosed and aggressive poorly differentiated or squamous cell cancer. A subset of NC is sensitive to chemotherapy, but the optimal regimen is unknown. Experts have recommended platinum- and ifosfamide-based therapy based on case reports. METHODS: Patients with pathologically confirmed NC with known survival outcomes after chemotherapy and consented to participate in a worldwide registry were studied. Results were summarized using descriptive methods. RESULTS: The study included 118 patients with NC. Median age was 34 (range: 1-82) years, 39% were women, and 61% harbored a BRD4::NUTM1 fusion. Patients received platinum (74%) or ifosfamide (26%, including regimens with both, 13%). Of 62 patients with nonmetastatic disease, 40% had a thoracic primary. Compared with platinum-based chemotherapy, patients who received ifosfamide-based chemotherapy had nominally higher progression-free survival (12 mo: 59% [95% CI: 32-87] versus 37% [95% CI: 22-52], hazard ratio = 0.68 [0.32, 1.42], p = 0.3) but not overall survival (OS). Among the 56 patients with metastatic disease, 80% had a thoracic primary. Ifosfamide had an objective response rate (ORR) of 75% (six of eight) and platinum had an ORR of 31% (11 of 36). Nevertheless, there was no difference in progression-free survival or OS. The 3-year OS of the entire cohort was 19% (95% CI: 10%-28%). Of the 11 patients alive greater than 3 years, all presented with nonmetastatic and operable or resectable disease. CONCLUSION: There is a numerically higher ORR for ifosfamide-based therapy compared with platinum-based therapy, with limited durability. OS at 3 years is only 19%, and development of effective therapies is an urgent unmet need for this patient population.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica , Humanos , Femenino , Masculino , Persona de Mediana Edad , Adulto , Anciano , Anciano de 80 o más Años , Adulto Joven , Adolescente , Niño , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Ifosfamida/administración & dosificación , Ifosfamida/uso terapéutico , Tasa de Supervivencia , Proteínas Nucleares/genética , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/mortalidadRESUMEN
NUT carcinoma (NC) is an aggressive squamous carcinoma defined by the BRD4-NUT fusion oncoprotein. Routinely effective systemic treatments are unavailable for most NC patients. The lack of an adequate animal model precludes identifying and leveraging cell-extrinsic factors therapeutically in NC. Here, we created a genetically engineered mouse model (GEMM) of NC that forms a Brd4::NUTM1 fusion gene upon tamoxifen induction of Sox2-driven Cre. The model displayed complete disease penetrance, with tumors arising from the squamous epithelium weeks after induction and all mice succumbing to the disease shortly thereafter. Closely resembling human NC (hNC), GEMM tumors (mNC) were poorly differentiated squamous carcinomas with high expression of MYC that metastasized to solid organs and regional lymph nodes. Two GEMM-derived cell lines were developed whose transcriptomic and epigenetic landscapes harbored key features of primary GEMM tumors. Importantly, GEMM tumor and cell line transcriptomes co-classified with those of human NC. BRD4-NUT also blocked differentiation and maintained the growth of mNC as in hNC. Mechanistically, GEMM primary tumors and cell lines formed large histone H3K27ac-enriched domains, termed megadomains, that were invariably associated with the expression of key NC-defining proto-oncogenes, Myc and Trp63. Small-molecule BET bromodomain inhibition (BETi) of mNC induced differentiation and growth arrest and prolonged survival of NC GEMMs, as it does in hNC models. Overall, tumor formation in the NC GEMM is definitive evidence that BRD4-NUT alone can potently drive the malignant transformation of squamous progenitor cells into NC. SIGNIFICANCE: The development of an immunocompetent model of NUT carcinoma that closely mimics the human disease provides a valuable global resource for mechanistic and preclinical studies to improve treatment of this incurable disease.
Asunto(s)
Carcinoma de Células Escamosas , Factores de Transcripción , Animales , Humanos , Ratones , Carcinoma de Células Escamosas/patología , Proteínas de Ciclo Celular/genética , Transformación Celular Neoplásica/genética , Proteínas Nucleares/metabolismo , Proteínas de Fusión Oncogénica/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
NUT carcinoma is an aggressive carcinoma driven by the BRD4-NUT fusion oncoprotein, which activates chromatin to promote expression of progrowth genes. BET bromodomain inhibitors (BETi) are a promising treatment for NUT carcinoma that can impede BRD4-NUT's ability to activate genes, but the efficacy of BETi as monotherapy is limited. Here, we demonstrated that enhancer of zeste homolog 2 (EZH2), which silences genes through establishment of repressive chromatin, is a dependency in NUT carcinoma. Inhibition of EZH2 with the clinical compound tazemetostat potently blocked growth of NUT carcinoma cells. Epigenetic and transcriptomic analysis revealed that tazemetostat reversed the EZH2-specific H3K27me3 silencing mark and restored expression of multiple tumor suppressor genes while having no effect on key oncogenic BRD4-NUT-regulated genes. Indeed, H3K27me3 and H3K27ac domains were found to be mutually exclusive in NUT carcinoma cells. CDKN2A was identified as the only gene among all tazemetostat-derepressed genes to confer resistance to tazemetostat in a CRISPR-Cas9 screen. Combined inhibition of EZH2 and BET synergized to downregulate cell proliferation genes, resulting in more pronounced growth arrest and differentiation than either inhibitor alone. In preclinical models, combined tazemetostat and BETi synergistically blocked tumor growth and prolonged survival of NUT carcinoma-xenografted mice, with complete remission without relapse in one cohort. Identification of EZH2 as a dependency in NUT carcinoma substantiates the reliance of NUT carcinoma tumor cells on epigenetic dysregulation of functionally opposite, yet highly complementary, chromatin regulatory pathways to maintain NUT carcinoma growth. SIGNIFICANCE: Repression of tumor suppressor genes, including CDKN2A, by EZH2 provides a mechanistic rationale for combining EZH2 and BET inhibitors for the clinical treatment of NUT carcinoma. See related commentary by Kazansky and Kentsis, p. 3827.
Asunto(s)
Carcinoma , Proteínas Nucleares , Animales , Humanos , Ratones , Carcinoma/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Cromatina , Proteína Potenciadora del Homólogo Zeste 2/genética , Proteína Potenciadora del Homólogo Zeste 2/metabolismo , Genes Supresores de Tumor , Histonas/metabolismo , Recurrencia Local de Neoplasia/genética , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
NUT carcinoma (NC) is an aggressive carcinoma driven by the BRD4-NUT fusion oncoprotein, which activates chromatin to promote expression of pro-growth genes. BET bromodomain inhibitors (BETi) impede BRD4-NUT's ability to activate genes and are thus a promising treatment but limited as monotherapy. The role of gene repression in NC is unknown. Here, we demonstrate that EZH2, which silences genes through establishment of repressive chromatin, is a dependency in NC. Inhibition of EZH2 with the clinical compound tazemetostat (taz) potently blocked growth of NC cells. Epigenetic and transcriptomic analysis revealed that taz reversed the EZH2-specific H3K27me3 silencing mark, and restored expression of multiple tumor suppressor genes while having no effect on key oncogenic BRD4- NUT-regulated genes. CDKN2A was identified as the only gene amongst all taz-derepressed genes to confer resistance to taz in a CRISPR-Cas9 screen. Combined EZH2 inhibition and BET inhibition synergized to downregulate cell proliferation genes resulting in more pronounced growth arrest and differentiation than either inhibitor alone. In pre-clinical models, combined taz and BETi synergistically blocked growth and prolonged survival of NC-xenografted mice, with all mice cured in one cohort. STATEMENT OF SIGNIFICANCE: Identification of EZH2 as a dependency in NC substantiates the reliance of NC tumor cells on epigenetic dysregulation of functionally opposite, yet highly complementary chromatin regulatory pathways to maintain NC growth. In particular, repression of CDKN2A expression by EZH2 provides a mechanistic rationale for combining EZH2i with BETi for the clinical treatment of NC.
RESUMEN
Midline carcinomas associated with the nuclear protein in testis (NUT) gene rearrangement are rare, aggressive tumors that have been diagnosed most commonly in the head, neck, mediastinum, and upper aerodigestive tract. The ultrastructural features associated with this tumor have thus far received only brief comment and have never been illustrated. The authors provide a more extensive description and illustrate the electron microscopic findings in a typical case of NUT midline carcinoma, confirmed by cytogenetic and fluorescence in situ hybridization studies. This tumor was composed of cells displaying large, irregularly shaped nuclei with prominent compact nucleoli and abundant cytoplasm containing prominent bundles of tonofilaments, occasional clusters of pleomorphic granules, small numbers of lipid inclusions, and rare glycogen deposits. The cells exhibited stubby microvillous projections, were intermittently enveloped by basal lamina, and were interjoined by numerous well-formed desmosomal-type junctions and occasional junctional complexes. The authors propose that this constellation of ultrastructural features can prove helpful in discriminating NUT midline carcinoma from similar appearing entities.
Asunto(s)
Reordenamiento Génico/genética , Neoplasias Pulmonares/ultraestructura , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Preescolar , Humanos , Hibridación Fluorescente in Situ/métodos , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Masculino , Microscopía Electrónica , Proteínas de Neoplasias , Translocación Genética/genéticaRESUMEN
OBJECTIVE: To investigate the importance of expression of the NUT gene and its rearrangement in diagnosing NUT midline carcinoma (NMC) of the upper respiratory tract; and to evaluate the prevalence, histological features and differential diagnosis of NMC of the upper respiratory tract. METHODS: One-hundred and sixty-three small cell malignant tumors of the upper respiratory tract were reviewed at the Beijing Tongren Hospital, Capital Medical University over a 20-year period. These cases included poorly-differentiated squamous cell carcinomas (n = 31), undifferentiated carcinoma (n = 1), non-keratizing undifferentiated nasopharyneal carcinomas (n = 60), small cell neuroendocrine carcinomas (n = 6) and non-epithelial small round cell malignant tumors (n = 65). The clinical and pathologic features were investigated. All cases were subjected to Epstein-Barr virus encoded RNA (EBER) in situ hybridization and NUT monoclonal antibody immunohistochemical staining. Cases positive for NUT immunohistochemistry and negative for EBER in situ hybridization were submitted for fluorescent in situ hybridization (FISH) for rearrangements in both BRD4 and NUT genes, and immunohistochemical staining for a set of cytokeratins (AE1/AE3, CK7, CK8), p63,and neuroendocrine markers (NSE, Syn, CgA, S-100 protein, CD56). RESULTS: Three cases of poorly-differentiated squamous cell carcinomas and one case of undifferentiated carcinoma showed diffuse nuclear immunohistochemical staining with antibody against NUT. These positive cases approximately accounted for 12.5% (4/32) of this group, 4.1% (4/98) of the malignant epithelial carcinomas and 2.5% (4/163) of all small round cell malignant tumors in the study. The age of these patients were 42 - 59 years. Other groups were all negative for NUT immunohistochemistry. These four cases also stained for antibodies against cytokeratins and p63, but were negative for neuroendocrine markers and not associated with EBV infection. Only two of these four cases showed rearrangements of the NUT and BRD4 genes by FISH. These two patients died within one year. The other two patients that did not demonstrate NUT rearrangement by FISH were alive and did not have an aggressive clinical course, surviving 40 and 12 months respectively. CONCLUSIONS: NMC is a rare small round cell malignant tumor in the upper respiratory tract. Only in the groups of primary poorly differentiated squamous cell carcinoma and undifferentiated carcinoma were positive for NUT immunohistochemical staining and NUT rearrangement by FISH. NMC typically occurs in midline organs, and affects the sinonasal tract. It is not associated with EBV infection. There is difference in the clinical course and prognosis among NMC patients. NUT immunohistochemical staining and NUT gene rearrangement analysis can differentiate NMC from other small cell tumors in the upper respiratory tract.
Asunto(s)
Carcinoma de Células Escamosas/genética , Carcinoma/genética , Neoplasias del Seno Maxilar/genética , Neoplasias Nasales/genética , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Adulto , Anticuerpos Monoclonales/metabolismo , Carcinoma/metabolismo , Carcinoma/patología , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patología , Proteínas de Ciclo Celular , Femenino , Regulación Neoplásica de la Expresión Génica , Reordenamiento Génico , Humanos , Queratinas/metabolismo , Masculino , Neoplasias del Seno Maxilar/metabolismo , Neoplasias del Seno Maxilar/patología , Proteínas de la Membrana/metabolismo , Persona de Mediana Edad , Proteínas de Neoplasias , Neoplasias Nasales/metabolismo , Neoplasias Nasales/patología , Proteínas Nucleares/metabolismo , Proteínas Oncogénicas/metabolismo , Proteínas de Fusión Oncogénica/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
NUT carcinoma (aka NUT midline carcinoma) is a rare, still significantly underrecognized aggressive malignancy. Although historically considered a midline malignancy of children and young adults, NUT carcinoma can originate in almost any body site and in any age group. Beside the classic BRD4-NUTM1 fusion, less common fusion partners include BRD3, NSD3, ZNF532, and ZNF592. Other fusions, including CIC, MGA, MXD4, MXD1, and BCORL1 are associated with sarcomas or cancers of unknown histogenesis. Involvement of the Z4 zinc finger protein (ZNF) family members ZNF532 and ZNF592 is exceedingly rare with only 3 recently reported cases. We herein describe a ZNF532-NUTM1-rearranged NUT carcinoma presenting as a 7.5 cm mass in the left lower lung lobe of a 65-year-old woman. Histology revealed undifferentiated monotonous small round cells with focal epithelioid and rhabdoid elements within a variably myxoid stroma. Immunohistochemistry revealed paucity of keratins and variable p63 combined with extensive CD30 and PLAP expression, leading to initial diagnoses of combined small cell carcinoma, CD30-positive unclassified hematolymphoid malignancy and malignant germ cell neoplasm. Negativity for other more specific germ cell markers justified seeking a fourth opinion, which revealed diffuse expression of the NUT antibody. The diagnosis was then confirmed by fluorescence in situ hybridization. Targeted RNA sequencing revealed the ZNF532-NUTM1 fusion. Screening of 7 NUT carcinomas (5 with BRD4-NUTM1 and 2 with NSD3-NUTM1 fusions) for germ cell markers revealed focal SALL4 reactivity in 3 cases (combined with variable AFP expression in 2), but none expressed CD30 or PLAP. An aberrant germ cell immunophenotype should be considered in NUT carcinoma to avoid misinterpretation as genuine germ cell malignancy as both diseases predominantly affect the young population, frequently involve the mediastinum and can be associated with elevated serum AFP.