RESUMEN
Apart from the lethal midline carcinoma (NUT carcinoma), NUTM1 translocation has also been reported in mesenchymal tumors, but is exceedingly rare. Here, we describe a series of 8 NUTM1 -rearranged sarcomas to further characterize the clinicopathologic features of this emerging entity. This cohort included 2 males and 6 females with age ranging from 24 to 64 years (mean: 51 y; median: 56 y). Tumors occurred in the colon (2), abdomen (2), jejunum (1), esophagus (1), lung (1) and infraorbital region (1). At diagnosis, 6 patients presented with metastatic disease. Tumor size ranged from 1 to 10.5 cm (mean: 6 cm; median: 5.5 cm). Histologically, 4 tumors were composed of primitive small round cells to epithelioid cells intermixed with variable spindle cells, while 3 tumors consisted exclusively of small round cells to epithelioid cells and 1 tumor consisted predominantly of high-grade spindle cells. The neoplastic cells were arranged in solid sheets, nests, or intersecting fascicles. Mitotic activity ranged from 1 to 15/10 HPF (median: 5/10 HPF). Other features included rhabdoid phenotype (4/8), pronounced nuclear convolutions (2/8), prominent stromal hyalinization (2/8), focally myxoid stroma (1/8), foci of osteoclasts (1/8), and necrosis (1/8). By immunohistochemistry, all tumors showed diffuse and strong nuclear staining of NUT protein, with variable expression of pancytokeratin (AE1/AE3) (2/8), CK18 (1/8), CD99 (3/8), NKX2.2 (2/8), cyclin D1 (2/8), desmin (2/8), BCOR (2/8), S100 (1/8), TLE1 (1/8), and synaptophysin (1/8). Seven of 8 tumors demonstrated NUTM1 rearrangement by fluorescence in situ hybridization analysis. RNA-sequencing analysis identified MXD4::NUTM1 (3/7), MXI1::NUTM1 (3/7), and MGA::NUTM1 (1/7) fusions, respectively. DNA-based methylation profiling performed in 2 cases revealed distinct methylation cluster differing from those of NUT carcinoma and undifferentiated small round cell and spindle cell sarcomas. At follow-up (range: 4 to 24 mo), 1 patient experienced recurrence at 8.5 months, 4 patients were alive with metastatic disease (5, 10, 11, and 24 mo after diagnosis), 3 patients remained well with no signs of recurrence or metastasis (4, 6, and 12 mo after diagnosis). Our study further demonstrated that NUTM1 -rearranged sarcoma had a broad range of clinicopathologic spectrum. NUT immunohistochemistry should be included in the diagnostic approach of monotonous undifferentiated small round, epithelioid to high-grade spindle cell malignancies that difficult to classify by conventional means. DNA-based methylation profiling might provide a promising tool in the epigenetic classification of undifferentiated sarcomas.
Asunto(s)
Biomarcadores de Tumor , Reordenamiento Génico , Proteínas de Neoplasias , Proteínas Nucleares , Sarcoma , Humanos , Masculino , Persona de Mediana Edad , Femenino , Adulto , Sarcoma/genética , Sarcoma/patología , Sarcoma/química , Proteínas Nucleares/genética , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/análisis , Adulto Joven , Proteínas de Neoplasias/genética , Inmunohistoquímica , Hibridación Fluorescente in Situ , Neoplasias de los Tejidos Blandos/genética , Neoplasias de los Tejidos Blandos/patología , Fenotipo , Predisposición Genética a la Enfermedad , Proteína Homeobox Nkx-2.2 , Factores de Transcripción , Proteínas de HomeodominioRESUMEN
BACKGROUND: CIC-rearranged sarcomas (CRS) represent a new entity of undifferentiated small round cell sarcoma belonging to the Ewing-like sarcomas family. CRS are the most common type. Fusion partners for the CIC gene include DUX4, FOXO4, and the recently recognizedNUTM1. Rare cases of CIC::NUTM1 sarcoma in pediatric patients have recently been reported in brain, kidney, bone, and soft tissues. However, such cases have not been identified in the soft tissues of the limbs. CASE PRESENTATION: We reported a case of CIC::NUTM1 sarcoma located in the right upper limb of an 18-year-old man. The tumor displayed morphologic features typical of CIC::DUX4 sarcomas, with small- to medium-sized round cells, a lobular pattern, focal spindling, myxoid stroma, and patchy necrosis. The tumor diffusely expressed NUTM1, was positive for WT1cter at weak to moderate intensity, and was focally positive for CD99, while it was negative for keratins, EMA, P40, MyoD1, myogenin, NKX2.2, BCOR, and pan-TRK. Fluorescence in situ hybridization analyses revealed cleavage of the CIC and NUTM1 genes. CONCLUSION: CIC::NUTM1 sarcomas represent a novel molecular variant of CRS with a preference for the central nervous system and younger pediatric persons. Its morphology and phenotype may be mistaken for NUT carcinomas, and the behavior is more progressive than other forms of CRS. For this rare and newly discovered gene fusion variant, it is necessary to integrate molecular and immunohistochemical findings with morphologic features in the diagnosis of undifferentiated neoplasms.
Asunto(s)
Proteínas Represoras , Neoplasias de los Tejidos Blandos , Humanos , Masculino , Adolescente , Neoplasias de los Tejidos Blandos/genética , Neoplasias de los Tejidos Blandos/patología , Proteínas Represoras/genética , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/análisis , Proteínas de Fusión Oncogénica/genética , Sarcoma/genética , Sarcoma/patología , Sarcoma/diagnóstico , Extremidad Superior/patología , Reordenamiento Génico , Proteína Homeobox Nkx-2.2 , Hibridación Fluorescente in Situ , Factores de Transcripción , Proteínas de HomeodominioRESUMEN
A female in her 60's presented with a left-sided breast mass. A core needle biopsy specimen showed diffuse proliferation of a round cell tumor, which was positive for vimentin, NKX2.2, BCOR, and focal CD99 on immunohistochemistry (IHC). No fusion genes of the Ewing family sarcomas were detected. With a tentative diagnosis of primary breast sarcoma (PBS), total mastectomy was performed after chemotherapy. The resected tissues showed proliferation of round or spindle-shaped tumor cells with a high nuclear-to-cytoplasmic ratio, exhibiting solid and fascicular arrangements but no epithelial component or organoid pattern. While IHC indicated no particular histological diagnosis, genomic examination revealed gene alterations in MED12 p.G44D, MLL2 (KMT2D) p.T1496fs*27, and EGFR variant III (vIII). Moreover, a retrospective IHC study showed overexpression of EGFRvIII. A malignant phyllodes tumor (PT) with extensive sarcomatous overgrowth was indicated as an integrative diagnosis. This is a rare case of a malignant PT harboring EGFRvIII. The present case provides an importance of accurate diagnosis and genomic analysis of rare breast tumors, as malignant PT and PBS are different in its treatment strategy and prognosis.
Asunto(s)
Biomarcadores de Tumor , Neoplasias de la Mama , Receptores ErbB , Inmunohistoquímica , Mutación , Tumor Filoide , Humanos , Femenino , Tumor Filoide/genética , Tumor Filoide/patología , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Persona de Mediana Edad , Receptores ErbB/genética , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/análisis , Proteína Homeobox Nkx-2.2 , Proteínas de Unión al ADN/genética , Proteínas de Homeodominio , Proteínas Nucleares , Complejo Mediador , Factores de Transcripción , Proteínas de NeoplasiasRESUMEN
Rapid information processing in the central nervous system requires the myelination of axons by oligodendrocytes. The transcription factor Sox2 and its close relative Sox3 redundantly regulate the development of myelin-forming oligodendrocytes, but little is known about the underlying molecular mechanisms. Here, we characterized the expression profile of cultured oligodendroglial cells during early differentiation and identified Bcas1, Enpp6, Zfp488 and Nkx2.2 as major downregulated genes upon Sox2 and Sox3 deletion. An analysis of mice with oligodendrocyte-specific deletion of Sox2 and Sox3 validated all four genes as downstream targets in vivo. Additional functional assays identified regulatory regions in the vicinity of each gene that are responsive to and bind both Sox proteins. Bcas1, Enpp6, Zfp488 and Nkx2.2 therefore likely represent direct target genes and major effectors of Sox2 and Sox3. Considering the preferential expression and role of these genes in premyelinating oligodendrocytes, our findings suggest that Sox2 and Sox3 impact oligodendroglial development at the premyelinating stage with Bcas1, Enpp6, Zfp488 and Nkx2.2 as their major effectors.
Asunto(s)
Diferenciación Celular , Proteína Homeobox Nkx-2.2 , Oligodendroglía , Factores de Transcripción SOXB1 , Factores de Transcripción , Animales , Ratones , Diferenciación Celular/genética , Regulación del Desarrollo de la Expresión Génica , Proteínas de Homeodominio/metabolismo , Proteínas de Homeodominio/genética , Oligodendroglía/metabolismo , Oligodendroglía/citología , Hidrolasas Diéster Fosfóricas/metabolismo , Hidrolasas Diéster Fosfóricas/genética , Factores de Transcripción SOXB1/metabolismo , Factores de Transcripción SOXB1/genética , Factores de Transcripción SOXC/metabolismo , Factores de Transcripción SOXC/genética , Factores de Transcripción/metabolismo , Factores de Transcripción/genéticaRESUMEN
Ewing sarcoma is an uncommon neoplasm considered in the differential diagnosis of tumors with "small round cell" morphology, but its occurrence in the gynecologic tract has only been sporadically documented. Herein, we describe the largest cohort of Ewing sarcoma localized to the female genital tract to date, and emphasize their clinicopathologic resemblance to more common gynecologic neoplasms. Ewing sarcoma (n=21) was retrospectively identified from 5 institutions. The average patient age was 35 (range 6-61) years. Tumor sites included uterus (n=8), cervix (n=4), vulva (n=5), vagina (n=1), broad ligament (n=1), inguinal area (n=1), and pelvis (n=1). Nine of 18 cases in which slides were available for review demonstrated only classic round cell morphology, with the remainder showing a variable combination and prominence of variant ovoid/spindle or epithelioid appearance. Tumors showed diffuse membranous reactivity for CD99 (20/20) and were positive for NKX2.2 (8/8, diffuse) and cyclin D1 (7/7, of which 3/7 were patchy/multifocal and 4/7 were diffuse). They were negative for ER (0/6) and CD10 (0/6). Three cases were initially diagnosed as endometrial stromal sarcomas. EWSR1 rearrangement was confirmed in 20/21 by fluorescence in situ hybridization (n=15) and/or sequencing (n=8). Of the eight tumors that underwent sequencing, 6 harbored FLI1 , 1 ERG, and 1 FEV as the fusion partner. Of 11 patients with available follow-up, 5 died of disease, 1 developed lung metastases and 5 are alive with no evidence of disease. Ewing sarcoma of the gynecologic tract is a rare, aggressive entity that shares some morphologic and immunohistochemical features with other more common gynecologic neoplasms. In addition to the typical round cell appearance, variant spindled/ovoid to epithelioid morphology may also be observed and should prompt consideration of this entity with appropriate immunohistochemical and/or molecular studies.
Asunto(s)
Biomarcadores de Tumor , Neoplasias de los Genitales Femeninos , Proteína EWS de Unión a ARN , Sarcoma de Ewing , Humanos , Femenino , Sarcoma de Ewing/genética , Sarcoma de Ewing/patología , Sarcoma de Ewing/diagnóstico , Sarcoma de Ewing/química , Neoplasias de los Genitales Femeninos/patología , Neoplasias de los Genitales Femeninos/genética , Neoplasias de los Genitales Femeninos/diagnóstico , Adulto , Diagnóstico Diferencial , Adolescente , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/análisis , Adulto Joven , Persona de Mediana Edad , Niño , Estudios Retrospectivos , Proteína EWS de Unión a ARN/genética , Inmunohistoquímica , Hibridación Fluorescente in Situ , Proteína Homeobox Nkx-2.2 , Factores de Transcripción/genética , Proteínas de Homeodominio/genética , Valor Predictivo de las Pruebas , Reordenamiento Génico , Antígeno 12E7/metabolismo , Células Epitelioides/patología , Células Epitelioides/química , Proteínas NuclearesRESUMEN
BACKGROUND: Ewing's sarcoma (ES) is an aggressive cancer of bone and soft tissue, most of which tend to occur in the bone. Extraosseous Ewing's sarcoma (EES) of the cervix is extremely rare. CASE PRESENTATION: In the present work, we reported a 39-year-old cervical EES patient with a 2.5*2.1*1.8 cm tumor mass. According to previous literatures, our case is the smallest tumor found in primary cervical ES ever. The patient initially came to our hospital due to vaginal bleeding, and then the gynecological examination found a neoplasm between the cervical canal and partially in the external cervical orifice. The diagnosis of EES was confirmed below: Hematoxylin & Eosin staining (H&E) revealed small round blue malignant cells in biopsy specimens. Immunohistochemistry (IHC) showed the positive staining for CD99, NKX2.2, and FLI1. Disruption of EWSR1 gene was found by fluorescence in situ hybridization (FISH), and the EWSR1-FLI1 gene fusion was determined by next-generation sequencing (NGS). The patient received laparoscopic wide hysterectomy, bilateral adnexectomy, pelvic lymphadenectomy, and postoperative adjuvant chemotherapy and remained disease free with regular follow-up for 1 year. CONCLUSIONS: Through a systematic review of previously reported cervical ES and this case, we highlighted the importance of FISH and NGS for the accuracy of ESS diagnosis, which could assist on the optimal treatment strategy. However, due to the rarity of the disease, there is no standard treatment schemes. Investigation on molecular pathological diagnosis and standardization of treatment regimens for cervical ES are critical to patients' prognosis.
Asunto(s)
Sarcoma de Ewing , Neoplasias del Cuello Uterino , Humanos , Femenino , Sarcoma de Ewing/patología , Sarcoma de Ewing/genética , Sarcoma de Ewing/diagnóstico , Neoplasias del Cuello Uterino/patología , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/diagnóstico , Neoplasias del Cuello Uterino/cirugía , Adulto , Proteínas de Fusión Oncogénica/genética , Proteína Homeobox Nkx-2.2 , Proteína EWS de Unión a ARN/genética , Hibridación Fluorescente in Situ , Factores de Transcripción/genética , Proteína Proto-Oncogénica c-fli-1/genética , Proteínas Nucleares , Proteínas de HomeodominioRESUMEN
Retroperitoneal Ewing sarcomas (RES) are very rare and mostly described in case reports. The purpose of this study was to retrospectively analyze the clinicopathology, molecular characteristics, biological behavior, and therapeutic information of 13 cases of primary RES with immunohistochemical staining, fluorescence in situ hybridization, RT-PCR and NGS sequencing detection techniques. The thirteen patients included eight males and five females with a mean age of 34 years. Morphologically, the tumors were comprised of small round or epithelial-like cells with vacuolated cytoplasm (6/13,46 %) arranged in diffuse, nested (8/13,62 %) and perivascular (7/13,54 %) patterns. Unusual morphologic patterns, such as meningioma-like swirling structures and sieve-like structures were relatively novel findings. Immunohistochemical studies showed CD99 (12/13; 92 %), CD56 (11/13; 85 %), NKX2.2 (9/13; 69 %), PAX7 (10/11;91 %) and CD117(6/9;67 %) to be positive.12 cases (92 %) demonstrated EWSR1 rearrangement and 3 cases displayed EWSR1::FLI1 fusion by FISH. ERCC4 splice-site variant, a novel pathogenic variant, was discovered for the first time via RNA sequencing. With a median follow-up duration of 14 months (6 to 79 months), 8/13 (62 %) patients died, while 5/13(38 %) survived. Three cases recurred, and five patients developed metastasis to the liver (2 cases), lung (2 cases) and bone (1 case). RES is an aggressive, high-grade tumor, prone to multiple recurrences and metastases, with distinctive morphologic, immunohistochemical, and molecular genetic features. ERCC4 splicing mutation, which is a novel pathogenic variant discovered for the first time, with possible significance for understanding the disease, as well as the development of targeted drugs.
Asunto(s)
Proteína Homeobox Nkx-2.2 , Proteína EWS de Unión a ARN , Neoplasias Retroperitoneales , Sarcoma de Ewing , Humanos , Sarcoma de Ewing/genética , Sarcoma de Ewing/patología , Sarcoma de Ewing/diagnóstico , Sarcoma de Ewing/metabolismo , Masculino , Femenino , Adulto , Neoplasias Retroperitoneales/genética , Neoplasias Retroperitoneales/patología , Neoplasias Retroperitoneales/diagnóstico , Estudios Retrospectivos , Persona de Mediana Edad , Adulto Joven , Adolescente , Proteína EWS de Unión a ARN/genética , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Hibridación Fluorescente in Situ/métodos , Reordenamiento Génico , Inmunohistoquímica/métodos , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Proteínas de Fusión Oncogénica/genética , Niño , Proteínas Nucleares , Proteínas de Homeodominio , Proteínas de Pez CebraRESUMEN
OBJECTIVE: This study aimed to investigate the role of Nkx1-2, a transcription factor with the NK homeobox domain, in the regulation of fat production. METHODS: Gene expression was analyzed using quantitative real-time polymerase chain reaction or transcriptome sequencing. CRISPR/Cas9 technology was employed to generate nkx1.2 knockout zebrafish and nkx1.2-deleted 3T3-L1 cells. Lipid droplet production in zebrafish larvae was visually quantified using Nile red staining, whereas lipid droplets in 3T3-L1 cells were stained with Oil red O. The binding of Nkx1-2 to the promoter was verified through an electrophoretic mobility shift assay experiment. RESULTS: Nkx1-2 plays crucial roles in the regulation of fat production in zebrafish. Knockout of nkx1.2 in zebrafish leads to weight loss, accompanied by significantly reduced lipid droplet production and decreased visceral and liver fat content. Furthermore, genes related to lipid biosynthesis are significantly downregulated. In 3T3-L1 preadipocytes, Nkx1-2 induces differentiation into mature adipocytes by binding to the cebpa promoter, thereby activating its transcription. Additionally, the expression of nkx1.2 is regulated by the p38 MAPK, JNK, or Smad2/3 signaling pathways in 3T3-L1 cells. CONCLUSIONS: Our findings suggest that Nkx1-2 functions as a positive regulator of fat production, playing a critical role in adipocyte differentiation and lipid biosynthesis.
Asunto(s)
Células 3T3-L1 , Proteínas de Homeodominio , Factores de Transcripción , Proteínas de Pez Cebra , Pez Cebra , Animales , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismo , Ratones , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Proteínas de Homeodominio/metabolismo , Proteínas de Homeodominio/genética , Adipocitos/metabolismo , Adipogénesis/genética , Diferenciación Celular , Sistemas CRISPR-Cas , Transducción de Señal , Gotas Lipídicas/metabolismo , Regiones Promotoras Genéticas , Metabolismo de los Lípidos/genética , Larva/metabolismo , Proteína Homeobox Nkx-2.2RESUMEN
BACKGROUND: Current approaches to profile the single-cell transcriptomics of human pancreatic endocrine cells almost exclusively rely on freshly isolated islets. However, human islets are limited in availability. Furthermore, the extensive processing steps during islet isolation and subsequent single cell dissolution might alter gene expressions. In this work, we report the development of a single-nucleus RNA sequencing (snRNA-seq) approach with targeted islet cell enrichment for endocrine-population focused transcriptomic profiling using frozen archival pancreatic tissues without islet isolation. RESULTS: We cross-compared five nuclei isolation protocols and selected the citric acid method as the best strategy to isolate nuclei with high RNA integrity and low cytoplasmic contamination from frozen archival human pancreata. We innovated fluorescence-activated nuclei sorting based on the positive signal of NKX2-2 antibody to enrich nuclei of the endocrine population from the entire nuclei pool of the pancreas. Our sample preparation procedure generated high-quality single-nucleus gene-expression libraries while preserving the endocrine population diversity. In comparison with single-cell RNA sequencing (scRNA-seq) library generated with live cells from freshly isolated human islets, the snRNA-seq library displayed comparable endocrine cellular composition and cell type signature gene expression. However, between these two types of libraries, differential enrichments of transcripts belonging to different functional classes could be observed. CONCLUSIONS: Our work fills a technological gap and helps to unleash frozen archival pancreatic tissues for molecular profiling targeting the endocrine population. This study opens doors to retrospective mappings of endocrine cell dynamics in pancreatic tissues of complex histopathology. We expect that our protocol is applicable to enrich nuclei for transcriptomics studies from various populations in different types of frozen archival tissues.
Asunto(s)
Núcleo Celular , Proteína Homeobox Nkx-2.2 , Proteínas de Homeodominio , Islotes Pancreáticos , Proteínas Nucleares , Análisis de Secuencia de ARN , Análisis de la Célula Individual , Factores de Transcripción , Humanos , Islotes Pancreáticos/metabolismo , Islotes Pancreáticos/citología , Análisis de la Célula Individual/métodos , Análisis de Secuencia de ARN/métodos , Núcleo Celular/genética , Núcleo Celular/metabolismo , Perfilación de la Expresión Génica/métodos , Páncreas/metabolismo , Páncreas/citología , TranscriptomaRESUMEN
Oligodendrocyte differentiation and myelination in the central nervous system are controlled and coordinated by a complex gene regulatory network that contains several transcription factors, including Zfp488 and Nkx2.2. Despite the proven role in oligodendrocyte differentiation little is known about the exact mode of Zfp488 and Nkx2.2 action, including their target genes. Here, we used overexpression of Zfp488 and Nkx2.2 in differentiating CG4 cells to identify aspects of the oligodendroglial expression profile that depend on these transcription factors. Although both transcription factors are primarily described as repressors, the detected changes argue for an additional function as activators. Among the genes activated by both Zfp488 and Nkx2.2 was the G protein-coupled receptor Gpr37 that is important during myelination. In agreement with a positive effect on Gpr37 expression, downregulation of the G protein-coupled receptor was observed in Zfp488- and in Nkx2.2-deficient oligodendrocytes in the mouse. We also identified several potential regulatory regions of the Gpr37 gene. Although Zfp488 and Nkx2.2 both bind to one of the regulatory regions downstream of the Gpr37 gene in vivo, none of the regulatory regions was activated by either transcription factor alone. Increased activation by Zfp488 or Nkx2.2 was only observed in the presence of Sox10, a transcription factor continuously present in oligodendroglial cells. Our results argue that both Zfp488 and Nkx2.2 also act as transcriptional activators during oligodendrocyte differentiation and cooperate with Sox10 to allow the expression of Gpr37 as a modulator of the myelination process.