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1.
J Cell Mol Med ; 26(5): 1684-1698, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-35118811

RESUMO

Uterine leiomyoma (UL) is the most common gynaecologic tumour, affecting an estimated 70 to 80% of women. Leiomyomas develop from the transformation of myometrial stem cells into leiomyoma stem (or tumour-initiating) cells. These cells undergo self-renewal and differentiation to mature cells, both are necessary for the maintenance of tumour stem cell niche and tumour growth, respectively. Wnt/ß-catenin and TGF-ß/SMAD pathways, both overactive in UL, promote stem cell self-renewal, crosstalk between stem and mature cells, cellular proliferation, extracellular matrix (ECM) accumulation and drive overall UL growth. Recent evidence suggests that simvastatin, an antihyperlipidemic drug, may have anti-leiomyoma properties. Herein, we investigated the effects of simvastatin on UL stem cells. We isolated leiomyoma stem cells by flow cytometry using DyeCycle Violet staining and Stro-1/CD44 surface markers. We found that simvastatin inhibits proliferation and induces apoptosis in UL stem cells. In addition, it also suppressed the expression of the stemness markers Nanog, Oct4 and Sox2. Simvastatin significantly decreased the production of the key ECM proteins, collagen 1 and fibronectin. Finally, it inhibited genes and/or proteins expression of TGF-ß1, 2 and 3, SMAD2, SMAD4, Wnt4, ß-Catenin, LRP6, AXIN2 and Cyclin D1 in UL stem cells, all are key drivers of the TGF-ß3/SMAD2 and Wnt4/ß-Catenin pathways. Thus, we have identified a novel stem cell-targeting anti-leiomyoma simvastatin effect. Further studies are needed to replicate these findings in vivo.


Assuntos
Leiomioma , Neoplasias Uterinas , Proliferação de Células/genética , Feminino , Humanos , Leiomioma/tratamento farmacológico , Leiomioma/genética , Sinvastatina/farmacologia , Fator de Crescimento Transformador beta3/metabolismo , Neoplasias Uterinas/genética , Neoplasias Uterinas/metabolismo , Neoplasias Uterinas/patologia , beta Catenina/metabolismo
2.
Int J Mol Sci ; 23(4)2022 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-35216305

RESUMO

The absence of standardized molecular profiling to differentiate uterine leiomyosarcomas versus leiomyomas represents a current diagnostic challenge. In this study, we aimed to search for a differential molecular signature for these myometrial tumors based on artificial intelligence. For this purpose, differential exome and transcriptome-wide research was performed on histologically confirmed leiomyomas (n = 52) and leiomyosarcomas (n = 44) to elucidate differences between and within these two entities. We identified a significantly higher tumor mutation burden in leiomyosarcomas vs. leiomyomas in terms of somatic single-nucleotide variants (171,863 vs. 81,152), indels (9491 vs. 4098), and copy number variants (8390 vs. 5376). Further, we discovered alterations in specific copy number variant regions that affect the expression of some tumor suppressor genes. A transcriptomic analysis revealed 489 differentially expressed genes between these two conditions, as well as structural rearrangements targeting ATRX and RAD51B. These results allowed us to develop a machine learning approach based on 19 differentially expressed genes that differentiate both tumor types with high sensitivity and specificity. Our findings provide a novel molecular signature for the diagnosis of leiomyoma and leiomyosarcoma, which could be helpful to complement the current morphological and immunohistochemical diagnosis and may lay the foundation for the future evaluation of malignancy risk.


Assuntos
Leiomioma , Leiomiossarcoma , Neoplasias Uterinas , Inteligência Artificial , Diagnóstico Diferencial , Feminino , Humanos , Leiomioma/diagnóstico , Leiomioma/genética , Leiomioma/metabolismo , Leiomiossarcoma/diagnóstico , Leiomiossarcoma/genética , Leiomiossarcoma/metabolismo , Transcriptoma , Neoplasias Uterinas/diagnóstico , Neoplasias Uterinas/genética , Neoplasias Uterinas/metabolismo
3.
J Reprod Immunol ; 150: 103475, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-35074692

RESUMO

Decidual natural killer cells (dNK) have been the focus of many studies because of their unique roles in both the anti-tumor immune response and healthy placental formation. Revealing the immunological mechanisms by which they interact with their target cells may lead to a better understanding of immune evasion of certain tumor cells, including abnormal cells of the different forms of gestational trophoblast disease and miscarriages of immunologic origin. Efforts to perform functional immunological studies on dNK cells have been limited by difficulty obtaining sufficent quantities of cells and sustaining the dNK phenotype. A novel protocol was developed to isolate and culture dNK cells from fresh, term placentas and complete hydatidiform moles.The placental samples were collected from healthy women undergoing scheduled elective cesarean delivery. The molar samples were collected after evacuation and curettage. Tissue samples were made into single cell suspensions using mechanical and enzymatic degradation, followed by fluorescence-activated cell sorting (FACS) using surface markers. The dNK cells were then expanded in cell culture. Their surface markers and cytotoxicity were reassessed by flow cytometry and functional assays. The protocol produces high quantities of enriched dNK cells which can be sustained in cell culture for at least a month, preserving their phenotype and funcionality for a week.


Assuntos
Mola Hidatiforme , Neoplasias Uterinas , Decídua , Feminino , Humanos , Mola Hidatiforme/metabolismo , Células Matadoras Naturais , Placenta , Gravidez , Neoplasias Uterinas/metabolismo
4.
Bioengineered ; 13(2): 2076-2086, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35038949

RESUMO

Choriocarcinoma is one of the most aggressive gestational trophoblastic neoplasias (GTN). Methotrexate (MTX) resistance is the main cause of treatment failure in choriocarcinoma. However, the mechanism of MTX resistance in choriocarcinoma is poorly known. This study aims to explore the function of Lectin galactoside-binding soluble 3 binding protein (LGALS3BP) in MTX-resistance in choriocarcinoma cells. Gradual dose escalation of MTX was used to establish MTX-resistant choriocarcinoma cells (JAR-MTX and JEG3-MTX cell lines). RNA-sequencing was used to explore the differentially expressed genes. Plasmids or SiRNA transfection was used to regulate the expression of LGALS3BP. ELISA was used to detect the concentrations of LGALS3BP in the serum of MTX-sensitive and MTX-resistant patients. qRT-PCR, Western blot, and CCK-8 assay were used to determine the effects of LGALS3BP on MTX-resistance in JAR and JEG3 cells. The results showed the relative resistance index (RI) of MTX is 791.50 and 1040.04 in JAR-MTX and JEG3-MTX, respectively. LGALS3BP was up-regulated in MTX-resistant cells compared to original cells in both RNA and protein level. The concentrations of LGALS3BP were higher in the sera of MTX-resistant patients than in MTX-sensitive patients. Knocking down LGALS3BP can reverse the MTX-resistance in JAR-MTX and JEG3-MTX cells. In summary, we preliminarily established two MTX-resistant cells, and performed RNA-sequencing, and found LGALS3BP may play important role in MTX-resistance. Our work not only provides a research tool (MTX-resistant cells) for other researchers, but gives some hint on how MTX resistance is regulated.


Assuntos
Antígenos de Neoplasias/metabolismo , Biomarcadores Tumorais/metabolismo , Coriocarcinoma/metabolismo , Resistencia a Medicamentos Antineoplásicos , Metotrexato/farmacologia , Proteínas de Neoplasias/metabolismo , Neoplasias Uterinas/metabolismo , Antígenos de Neoplasias/genética , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Coriocarcinoma/tratamento farmacológico , Coriocarcinoma/genética , Coriocarcinoma/patologia , Feminino , Humanos , Proteínas de Neoplasias/genética , Gravidez , Neoplasias Uterinas/tratamento farmacológico , Neoplasias Uterinas/genética , Neoplasias Uterinas/patologia
5.
FASEB J ; 36(2): e22101, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35032343

RESUMO

Tetrabromobisphenol A (TBBPA), a derivative of BPA, is a ubiquitous environmental contaminant with weak estrogenic properties. In women, uterine fibroids are highly prevalent estrogen-responsive tumors often with excessive accumulation of extracellular matrix (ECM) and may be the target of environmental estrogens. We have found that BPA has profibrotic effects in vitro, in addition to previous reports of the in vivo fibrotic effects of BPA in mouse uterus. However, the role of TBBPA in fibrosis is unclear. To investigate the effects of TBBPA on uterine fibrosis, we developed a 3D human uterine leiomyoma (ht-UtLM) spheroid culture model. Cell proliferation was evaluated in 3D ht-UtLM spheroids following TBBPA (10-6 -200 µM) administration at 48 h. Fibrosis was assessed using a Masson's Trichrome stain and light microscopy at 7 days of TBBPA (10-3  µM) treatment. Differential expression of ECM and fibrosis genes were determined using RT² Profiler™ PCR arrays. Network and pathway analyses were conducted using Ingenuity Pathway Analysis. The activation of pathway proteins was analyzed by a transforming growth factor-beta (TGFB) protein array. We found that TBBPA increased cell proliferation and promoted fibrosis in 3D ht-UtLM spheroids with increased deposition of collagens. TBBPA upregulated the expression of profibrotic genes and corresponding proteins associated with the TGFB pathway. TBBPA activated TGFB signaling through phosphorylation of TGFBR1 and downstream effectors-small mothers against decapentaplegic -2 and -3 proteins (SMAD2 and SMAD3). The 3D ht-UtLM spheroid model is an effective system for studying environmental agents on human uterine fibrosis. TBBPA can promote fibrosis in uterine fibroid through TGFB/SMAD signaling.


Assuntos
Fibrose/induzido quimicamente , Fibrose/metabolismo , Leiomioma/induzido quimicamente , Bifenil Polibromatos/administração & dosagem , Fator de Crescimento Transformador beta/metabolismo , Neoplasias Uterinas/induzido quimicamente , Neoplasias Uterinas/metabolismo , /métodos , Proliferação de Células/efeitos dos fármacos , Estrogênios/metabolismo , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Feminino , Humanos , Leiomioma/metabolismo , Fosforilação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
6.
Genes (Basel) ; 13(1)2022 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-35052473

RESUMO

The transcription factor ZBED1 is highly expressed in trophoblast cells, but its functions in the processes of trophoblast and placental biology remain elusive. Here, we characterized the role of ZBED1 in trophoblast cell differentiation using an in vitro BeWo cell model. We demonstrate that ZBED1 is enhanced in its expression early after forskolin-induced differentiation of BeWo cells and regulates many of the genes that are differentially expressed as an effect of forskolin treatment. Specifically, genes encoding markers for the differentiation of cytotrophoblast into syncytiotrophoblast and factors essential for trophoblast cell fusion and invasion were negatively regulated by ZBED1, indicating that ZBED1 might be important for maintaining a steady pool of cytotrophoblast cells. In addition, ZBED1 affected genes involved in the regulation of trophoblast cell survival and apoptosis, in agreement with the observed increase in apoptosis upon knockdown of ZBED1 in forskolin-treated BeWo cells. In addition, genes implicated in the differentiation, recruitment, and function of innate immune cells by the placenta were affected by ZBED1, further suggesting a role for this protein in the regulation of maternal immune tolerance. In conclusion, our study implicates ZBED1 in major biological processes of placental biology.


Assuntos
Fusão Celular , Coriocarcinoma/patologia , Regulação da Expressão Gênica , Placenta/patologia , Fatores de Transcrição/metabolismo , Trofoblastos/patologia , Neoplasias Uterinas/patologia , Diferenciação Celular , Coriocarcinoma/genética , Coriocarcinoma/metabolismo , Feminino , Humanos , Placenta/metabolismo , Gravidez , Fatores de Transcrição/genética , Trofoblastos/metabolismo , Células Tumorais Cultivadas , Neoplasias Uterinas/genética , Neoplasias Uterinas/metabolismo
7.
Reprod Sci ; 29(3): 781-790, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-34642915

RESUMO

Uterine leiomyosarcoma (LMS) contributes to a significant proportion of uterine cancer deaths. It is a rare and high-risk gynecological cancer. LMS is challenging to the treatment due to the resistance of several therapies. The activation of the Hedgehog (HH) pathway has been reported in several types of female cancers. Uterine LMS presents an upregulation of the crucial HH signaling pathway members such as SMO and GLI1. Although targeting the HH pathway exhibited a potent inhibitory effect on the phenotype of uterine LMS in vitro, the effect of the HH inhibitors on LMS growth in vivo has not been identified. The present study aimed to assess the effect of Hedgehog pathway inhibitors (SMO-LDE225 and GLI-Gant61) as a therapeutic option in the xenograft model of uterine LMS. The results demonstrated that LDE225 treatment did not show any inhibitory effect on LMS tumor growth; however, treatment with GLI inhibitor (Gant61) induced a remarkable tumor regression with a significant decrease in Ki67 expression, compared to control (p < 0.01). Moreover, administration of Gant61 decreased the expression of GLI1, GLI target genes BMP4 and c-MYC (p < 0.05), indicating that the HH pathway is implicated in the LMS experimental model. In conclusion, our studies demonstrate for the first time that GLI inhibitor (Gant61), but not SMO inhibitor (LDE225), shows a potent inhibitory effect on LMS tumor growth and concomitantly suppresses the expression of GLI1- and GLI-targeted genes using the xenograft model of uterine LMS.


Assuntos
Compostos de Bifenilo/farmacologia , Proteínas Hedgehog/antagonistas & inibidores , Proteínas Hedgehog/metabolismo , Leiomiossarcoma/tratamento farmacológico , Piridinas/farmacologia , Pirimidinas/farmacologia , Neoplasias Uterinas/tratamento farmacológico , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Leiomiossarcoma/metabolismo , Camundongos , Camundongos Nus , Fenótipo , Transdução de Sinais , Neoplasias Uterinas/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
8.
G3 (Bethesda) ; 12(1)2022 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-34791179

RESUMO

Uterine cancer is the fourth most common cancer among women, projected to affect 66,000 US women in 2021. Uterine cancer often arises in the inner lining of the uterus, known as the endometrium, but can present as several different types of cancer, including endometrioid cancer, serous adenocarcinoma, and uterine carcinosarcoma. Previous studies have analyzed the genetic changes between normal and cancerous uterine tissue to identify specific genes of interest, including TP53 and PTEN. Here we used Gaussian Mixture Models to build condition-specific gene coexpression networks for endometrial cancer, uterine carcinosarcoma, and normal uterine tissue. We then incorporated uterine regulatory edges and investigated potential coregulation relationships. These networks were further validated using differential expression analysis, functional enrichment, and a statistical analysis comparing the expression of transcription factors and their target genes across cancerous and normal uterine samples. These networks allow for a more comprehensive look into the biological networks and pathways affected in uterine cancer compared with previous singular gene analyses. We hope this study can be incorporated into existing knowledge surrounding the genetics of uterine cancer and soon become clinical biomarkers as a tool for better prognosis and treatment.


Assuntos
Cistadenocarcinoma Seroso , Neoplasias do Endométrio , Neoplasias Uterinas , Biomarcadores , Cistadenocarcinoma Seroso/genética , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/patologia , Feminino , Humanos , Neoplasias Uterinas/genética , Neoplasias Uterinas/metabolismo , Neoplasias Uterinas/patologia , Útero/metabolismo
9.
Reprod Sci ; 29(4): 1086-1101, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-33844188

RESUMO

Uterine leiomyoma (UL), as the most prevalent type of women's health disorders, is a benign tumor that originates from the smooth muscle cell layer of the uterus. A great number of associated complications are observed including infertility, miscarriage, bleeding, pain, dysmenorrhea, menorrhagia, and dyspareunia. Although the etiology of UL is largely undefined, environmental and genetic factors are witnessed to engage in the UL development. As long non-coding RNAs (lncRNAs) are involved in various types of cellular functions, in recent years, a great deal of attention has been drawn to them and their possible roles in UL pathogenesis. Moreover, they have illustrated their potential to be promising candidates for UL treatment. In this review paper, firstly, an overview of UL pathogenesis is presented. Then, the regulation of lncRNAs in UL and their possible mechanisms in cancer development are reviewed. Eventually, therapeutic approaches targeting lncRNAs in various cancers and UL are explored.


Assuntos
Leiomioma , RNA Longo não Codificante , Neoplasias Uterinas , Feminino , Humanos , Leiomioma/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Neoplasias Uterinas/metabolismo
10.
Cancer Res ; 82(4): 721-733, 2022 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-34921012

RESUMO

Uterine serous carcinoma (USC) is a highly aggressive endometrial cancer subtype with limited therapeutic options and a lack of targeted therapies. While mutations to PPP2R1A, which encodes the predominant protein phosphatase 2A (PP2A) scaffolding protein Aα, occur in 30% to 40% of USC cases, the clinical actionability of these mutations has not been studied. Using a high-throughput screening approach, we showed that mutations in Aα results in synthetic lethality following treatment with inhibitors of ribonucleotide reductase (RNR). In vivo, multiple models of Aα mutant uterine serous tumors were sensitive to clofarabine, an RNR inhibitor (RNRi). Aα-mutant cells displayed impaired checkpoint signaling upon RNRi treatment and subsequently accumulated more DNA damage than wild-type (WT) cells. Consistently, inhibition of PP2A activity using LB-100, a catalytic inhibitor, sensitized WT USC cells to RNRi. Analysis of The Cancer Genome Atlas data indicated that inactivation of PP2A, through loss of PP2A subunit expression, was prevalent in USC, with 88% of patients with USC harboring loss of at least one PP2A gene. In contrast, loss of PP2A subunit expression was rare in uterine endometrioid carcinomas. While RNRi are not routinely used for uterine cancers, a retrospective analysis of patients treated with gemcitabine as a second- or later-line therapy revealed a trend for improved outcomes in patients with USC treated with RNRi gemcitabine compared with patients with endometrioid histology. Overall, our data provide experimental evidence to support the use of ribonucleotide reductase inhibitors for the treatment of USC. SIGNIFICANCE: A drug repurposing screen identifies synthetic lethal interactions in PP2A-deficient uterine serous carcinoma, providing potential therapeutic avenues for treating this deadly endometrial cancer.


Assuntos
Cistadenocarcinoma Seroso/genética , Proteína Fosfatase 2/genética , Ribonucleotídeo Redutases/genética , Mutações Sintéticas Letais/genética , Neoplasias Uterinas/genética , Animais , Antimetabólitos Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Clofarabina/farmacologia , Cistadenocarcinoma Seroso/tratamento farmacológico , Cistadenocarcinoma Seroso/metabolismo , Feminino , Humanos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Proteína Fosfatase 2/metabolismo , Ratos Sprague-Dawley , Ribonucleotídeo Redutases/antagonistas & inibidores , Ribonucleotídeo Redutases/metabolismo , Mutações Sintéticas Letais/efeitos dos fármacos , Carga Tumoral/efeitos dos fármacos , Carga Tumoral/genética , Neoplasias Uterinas/tratamento farmacológico , Neoplasias Uterinas/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
11.
Dis Markers ; 2021: 1484227, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34745385

RESUMO

Uterine carcinosarcoma (UCS) is a highly invasive malignant tumor that originated from the uterine epithelium. Many studies suggested that the abnormal changes of alternative splicing (AS) of pre-mRNA are related to the occurrence and metastasis of the tumor. This study investigates the mechanism of alternative splicing events (ASEs) in the tumorigenesis and metastasis of UCS. RNA-seq of UCS samples and alternative splicing event (ASE) data of UCS samples were downloaded from The Cancer Genome Atlas (TCGA) and TCGASpliceSeq databases, several times. Firstly, we performed the Cox regression analysis to identify the overall survival-related alternative splicing events (OSRASEs). Secondly, a multivariate model was applied to approach the prognostic values of the risk score. Afterwards, a coexpressed network between splicing factors (SFs) and OSRASEs was constructed. In order to explore the relationship between the potential prognostic signaling pathways and OSRASEs, we fabricated a network between these pathways and OSRASEs. Finally, validations from multidimension platforms were used to explain the results unambiguously. 1,040 OSRASEs were identified by Cox regression. Then, 6 OSRASEs were incorporated in a multivariable model by Lasso regression. The area under the curve (AUC) of the receiver operator characteristic (ROC) curve was 0.957. The risk score rendered from the multivariate model was corroborated to be an independent prognostic factor (P < 0.001). In the network of SFs and ASEs, junction plakoglobin (JUP) noteworthily regulated RALGPS1-87608-AT (P < 0.001, R = 0.455). Additionally, RALGPS1-87608-AT (P = 0.006) showed a prominent relationship with distant metastasis. KEGG pathways related to prognosis of UCS were selected by gene set variation analysis (GSVA). The pyrimidine metabolism (P < 0.001, R = -0.470) was the key pathway coexpressed with RALGPS1. We considered that aberrant JUP significantly regulated RALGPS1-87608-AT and the pyrimidine metabolism pathway might play a significant part in the metastasis and prognosis of UCS.


Assuntos
Biomarcadores Tumorais/genética , Carcinossarcoma/genética , Neoplasias Uterinas/genética , Biomarcadores Tumorais/metabolismo , Carcinossarcoma/metabolismo , Carcinossarcoma/patologia , Feminino , Humanos , Metástase Neoplásica , Análise de Sobrevida , Neoplasias Uterinas/metabolismo , Neoplasias Uterinas/patologia , gama Catenina/genética , gama Catenina/metabolismo
12.
Gynecol Oncol ; 163(3): 524-530, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34625284

RESUMO

BACKGROUND: Aromatase inhibitors have been used empirically to treat a subset of patients with hormone receptor positive uterine leiomyosarcomas(LMS) and carcinosarcomas (UCS) mainly supported by retrospective data. We evaluated the activity of anastrozole in two rare cohorts; patients with recurrent/metastatic LMS and UCS enrolled in PARAGON, a basket trial of anastrozole in estrogen receptor (ER+)/progesterone receptor positive (PR+) gynecological cancers. METHOD: An investigator-initiated, single-arm, prospective open-label trial of anastrozole 1 mg/day in patients with ER &/or PR + ve LMS or UCS with measurable disease, treated until progression or unacceptable toxicity. Primary endpoint was clinical benefit (complete/partial response + stable disease) rate (CBR) at 3 months. Secondary endpoints include progression-free survival (PFS), quality of life and toxicity. RESULTS: 39 eligible patients were enrolled, 32 with LMS and 7 with UCS. For the LMS cohort CBR at 3 months was 35% (95% CI: 21-53%) with a median duration of clinical benefit of 5.8 months. Best response was a partial response in one patient. Two patients remained on treatment for more than one year. The median progression-free survival was 2.8 months (95% CI: 2.6-4.9). For the UCS cohort CBR at 3 months was 43% (95% CI: 16-75%) with a median duration of clinical benefit of 5.6 months. Stable disease was seen in 3 patients but no objective responses were seen. The median progression-free survival was 2.7 months (95% CI, 1.1-8.2). Safety was acceptable with 5/39 evaluable patients showing grade 3 toxicities. CONCLUSION: Whilst objective response rates with anastrozole are low, the clinical benefit rate and good tolerance suggests that aromatase inhibitor therapy may have a role in a subset of patients with metastatic LMS and UCS.


Assuntos
Anastrozol/uso terapêutico , Carcinossarcoma/tratamento farmacológico , Leiomiossarcoma/tratamento farmacológico , Neoplasias Uterinas/tratamento farmacológico , Adulto , Idoso , Idoso de 80 Anos ou mais , Anastrozol/efeitos adversos , Inibidores da Aromatase/efeitos adversos , Inibidores da Aromatase/uso terapêutico , Carcinossarcoma/metabolismo , Carcinossarcoma/patologia , Feminino , Humanos , Leiomiossarcoma/metabolismo , Leiomiossarcoma/patologia , Pessoa de Meia-Idade , Metástase Neoplásica , Estudos Prospectivos , Qualidade de Vida , Receptores de Estrogênio/metabolismo , Receptores de Progesterona/metabolismo , Neoplasias Uterinas/metabolismo , Neoplasias Uterinas/patologia
13.
Endocrinology ; 162(12)2021 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-34614511

RESUMO

The Wnt/ß-catenin pathway is upregulated in uterine leiomyomas, the most common benign tumors in the female reproductive tract. Simvastatin is an antihyperlipidemic drug, and previous in vitro and in vivo reports showed that it may have therapeutic effects in treating leiomyomas. The objective of this study was to examine the effects of simvastatin on the Wnt/ß-catenin signaling pathway in leiomyoma. We treated primary and immortalized human leiomyoma cells with simvastatin and examined its effects using quantitative real-time polymerase chain reaction, Western blotting, and immunocytochemistry. We also examined the effects using human leiomyoma tissues from an ongoing randomized controlled trial in which women with symptomatic leiomyoma received simvastatin (40 mg) or placebo for 3 months prior to their surgery. The results of this study revealed that simvastatin significantly reduced the expression of Wnt4 and its co-receptor LRP5. After simvastatin treatment, levels of total ß-catenin and its active form, nonphosphorylated ß-catenin, were reduced in both cell types. Additionally, simvastatin reduced the expression of Wnt4 and total ß-catenin, as well as nonphosphorylated ß-catenin protein expression in response to estrogen and progesterone. Simvastatin also inhibited the expression of c-Myc, a downstream target of the Wnt/ß-catenin pathway. The effect of simvastatin on nonphosphorylated-ß-catenin, the key regulator of the Wnt/ß-catenin pathway, was recapitulated in human leiomyoma tissue. These results suggest that simvastatin may have a beneficial effect on uterine leiomyoma through suppressing the overactive Wnt/ß-catenin pathway.


Assuntos
Leiomioma/patologia , Sinvastatina/farmacologia , Neoplasias Uterinas/patologia , Via de Sinalização Wnt/efeitos dos fármacos , Adolescente , Adulto , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Células Cultivadas , Ensaios Clínicos Fase II como Assunto , Método Duplo-Cego , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Feminino , Humanos , Leiomioma/genética , Leiomioma/metabolismo , Pessoa de Meia-Idade , Cultura Primária de Células , Ensaios Clínicos Controlados Aleatórios como Assunto , Neoplasias Uterinas/genética , Neoplasias Uterinas/metabolismo , Adulto Jovem , beta Catenina/antagonistas & inibidores , beta Catenina/metabolismo
14.
Exp Oncol ; 43(3): 209-216, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34591419

RESUMO

BACKGROUND: Uterine leiomyosarcoma is a rare malignant smooth muscle tumor originating in the uterine wall that generally responds poorly to chemotherapy and radiation. AIM: We investigated the in vitro effects of a novel nutrient mixture containing lysine, proline, ascorbic acid, and green tea extract on the human leiomyosarcoma cell line SK-UT-1 by measuring cell proliferation, invasiveness, apoptosis, and expression of matrix metalloproteinases (MMP). We also tested the effects of nutrient mixture in vivo using nude mice. MATERIALS AND METHODS: Human leiomyosarcoma SK-UT-1 cells were treated with different concentrations of nutrient mixture. Cell proliferation was determined by MTT assay; MMP expression by gelatinase zymography; invasion by Matrigel assay; migration by scratch test; apoptosis using Live Green caspase kit. In vivo studies were conducted on 5-6 weeks old female nude mice inoculated subcutaneously with 3 â€¢ 106 SK-UT-1 cells. The mice were fed a regular diet or a diet supplemented with 0.5% nutrient mixture. After four weeks, the mice were sacrificed and the tumors were weighed and processed for histology. RESULTS: In vitro, nutrient mixture treatment was not toxic to SK-UT-1 cells at 250 µg/ml but exhibited 20% and 40% cytotoxicity at 500 and 1000 µg/ml respectively. Zymography did not show bands for either MMP-2 or MMP-9 in SK-UT-1 cells. However, treatment with phorbol myristate acetate stimulated the expression of MMP-9, both active and inactive forms in equal proportion. Nutrient mixture inhibited the secretion of both active and inactive forms in a dose dependent manner. Invasion through Matrigel and migration by scratch test were inhibited in a dose dependent fashion, with both invasion and migration inhibited at 250 µg/ml. Live Green Caspase apoptosis assay demonstrated slight apoptosis at 100 µg/ml and significant apoptosis at 250 to 1000 µg/ml. The results of in vitro studies were further confirmed in vivo by showing 50% decrease in tumor weight, 40% reduction in tumor burden compared to the tumors from mice fed regular diet. CONCLUSION: The results suggest a therapeutic potential for nutrient mixture in uterine leiomyosarcoma treatment.


Assuntos
Leiomiossarcoma/tratamento farmacológico , Metaloproteinase 2 da Matriz/química , Metaloproteinase 9 da Matriz/química , Inibidores de Metaloproteinases de Matriz/farmacologia , Nutrientes/farmacologia , Extratos Vegetais/farmacologia , Neoplasias Uterinas/tratamento farmacológico , Animais , Antioxidantes , Apoptose , Proliferação de Células , Suplementos Nutricionais , Feminino , Humanos , Leiomiossarcoma/metabolismo , Leiomiossarcoma/patologia , Camundongos , Camundongos Nus , Chá/química , Células Tumorais Cultivadas , Neoplasias Uterinas/metabolismo , Neoplasias Uterinas/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Nutrients ; 13(9)2021 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-34578952

RESUMO

Leiomyoma is the most common benign uterine tumor in reproductive-age women. Increasing numbers of studies are focusing on the effects of environmental exposure on the incidence and progression of tumors. One major step taken in the food industry is the addition of food preservatives to maintain freshness. Butylated hydroxytoluene (BHT) is a synthetic phenolic antioxidant, which is widely used as an additive to develop fat-soluble characteristics, as well as in cosmetics and rubber. Previous studies also highlighted that BHT may be related to increased fibrosis capacity and carcinogenic effects. In this study, we explored the effects of the commonly used food additive BHT on leiomyoma progression, and the related mechanism. The exposure of the ELT-3 leiomyoma cell line to BHT for 48 h increased the proliferative effect. Since leiomyoma progression is related to increases in extracellular matrix (ECM) accumulation and matrix metalloproteinase (MMP), BHT could effectively increase ECM-related protein expression, as well as MMP-2 and MMP-9 protein expression. This increase in ECM, in response to BHT, may be linked to the activation of the phosphoinositide 3-kinase (PI3K)/Akt and mitogen-activated protein kinase (MAPK) signaling pathway. Through PI3K inhibition, BHT's effect on leiomyoma progression could be partially modulated. These results suggest the harmful effect of BHT exposure on leiomyoma progression may relate to PI3K modulation. However, an in vivo study is necessary to confirm these findings.


Assuntos
Hidroxitolueno Butilado/metabolismo , Proliferação de Células/efeitos dos fármacos , Matriz Extracelular/metabolismo , Aditivos Alimentares/metabolismo , Leiomioma/metabolismo , Neoplasias Uterinas/metabolismo , Hidroxitolueno Butilado/efeitos adversos , Linhagem Celular Tumoral , Progressão da Doença , Matriz Extracelular/efeitos dos fármacos , Feminino , Aditivos Alimentares/efeitos adversos , Humanos
16.
Endocrinology ; 162(11)2021 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-34491311

RESUMO

The objective of this study was to determine the expression and functional role of a long noncoding RNA (lncRNA) MIAT (myocardial infarction-associated transcript) in leiomyoma pathogenesis. Leiomyoma compared with myometrium (n = 66) expressed significantly more MIAT that was independent of race/ethnicity and menstrual cycle phase but dependent on MED12 (mediator complex subunit 12) mutation status. Leiomyomas bearing the MED12 mutation expressed higher levels of MIAT and lower levels of microRNA 29 family (miR-29a, -b, and -c) compared with MED12 wild-type leiomyomas. Using luciferase reporter activity and RNA immunoprecipitation analysis, MIAT was shown to sponge the miR-29 family. In a 3-dimensional spheroid culture system, transient transfection of MIAT siRNA in leiomyoma smooth muscle cell (LSMC) spheroids resulted in upregulation of miR-29 family and downregulation of miR-29 targets, collagen type I (COL1A1), collagen type III (COL3A1), and TGF-ß3 (transforming growth factor ß-3). Treatment of LSMC spheroids with TGF-ß3 induced COL1A1, COL3A1, and MIAT levels, but repressed miR-29 family expression. Knockdown of MIAT in LSMC spheroids blocked the effects of TGF-ß3 on the induction of COL1A1 and COL3A1 expression. Collectively, these results underscore the physiological significance of MIAT in extracellular matrix accumulation in leiomyoma.


Assuntos
Matriz Extracelular/metabolismo , Leiomioma/genética , MicroRNAs/genética , RNA Longo não Codificante/fisiologia , Neoplasias Uterinas/genética , Adulto , Células Cultivadas , Regulação para Baixo/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Leiomioma/metabolismo , Leiomioma/patologia , Pessoa de Meia-Idade , Família Multigênica/genética , Multimerização Proteica/genética , Neoplasias Uterinas/metabolismo , Neoplasias Uterinas/patologia
17.
Cancer Res ; 81(21): 5413-5424, 2021 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-34475109

RESUMO

Uterine leiomyosarcoma (ULMS) is a malignancy, which arises from the uterine smooth muscle. Because of its rarity, aggressive nature, and extremely poor prognosis, the molecular mechanisms driving ULMS remain elusive. To identify candidate cancer genes (CCG) driving ULMS, we conducted an in vivo Sleeping Beauty (SB) transposon mutagenesis screen in uterine myometrium-specific, PTEN knockout, KRAS mutant (PTEN KO/KRAS) mice. ULMS quickly developed in SB PTEN KO/KRAS mice, but not in PTEN KO/KRAS mice, demonstrating the critical importance of SB mutagenesis for driving ULMS in this model. Subsequent sequencing of SB insertion sites in these tumors identified 19 ULMS CCGs that were significantly enriched in known cancer genes. Among them, Zfp217 and Sfmbt2 functioned at early stages of tumor initiation and appeared to be oncogenes. Expression of ZNF217, the human homolog of ZFP217, was shown to be elevated in human ULMS compared with paired normal uterine smooth muscle, where it negatively correlated with patient prognosis. Inhibition of ZNF217 suppressed, whereas overexpression induced, proliferation, survival, migration, and stemness of human ULMS. In a second ex vivo ULMS SB metastasis screen, three CCGs were identified that may drive ULMS metastasis to the lung. One of these CCGs, Nrd1 (NRDC in humans), showed stronger expression in human metastatic tumors compared with primary ULMS and negatively associated with patient survival. NRDC knockdown impaired migration and adhesion without affecting cell proliferation, whereas overexpression had the opposite effect. Together, these results reveal novel mechanism driving ULMS tumorigenesis and metastasis and identify ZNF217 and NRDC as potential targets for ULMS therapy. SIGNIFICANCE: An in vivo Sleeping Beauty transposon mutagenesis screen identifies candidate cancer genes that drive initiation and progression of uterine leiomyosarcoma and may serve as therapeutic targets.


Assuntos
Biomarcadores Tumorais/genética , Elementos de DNA Transponíveis , Leiomiossarcoma/patologia , Neoplasias Pulmonares/secundário , Mutagênese Insercional , Mutação , Neoplasias Uterinas/patologia , Animais , Feminino , Humanos , Leiomiossarcoma/etiologia , Leiomiossarcoma/metabolismo , Neoplasias Pulmonares/etiologia , Neoplasias Pulmonares/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , PTEN Fosfo-Hidrolase/fisiologia , Proteínas Proto-Oncogênicas p21(ras)/fisiologia , Transposases/genética , Transposases/metabolismo , Neoplasias Uterinas/etiologia , Neoplasias Uterinas/metabolismo
18.
Nature ; 596(7872): 398-403, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34349258

RESUMO

One in four women suffers from uterine leiomyomas (ULs)-benign tumours of the uterine wall, also known as uterine fibroids-at some point in premenopausal life. ULs can cause excessive bleeding, pain and infertility1, and are a common cause of hysterectomy2. They emerge through at least three distinct genetic drivers: mutations in MED12 or FH, or genomic rearrangement of HMGA23. Here we created genome-wide datasets, using DNA, RNA, assay for transposase-accessible chromatin (ATAC), chromatin immunoprecipitation (ChIP) and HiC chromatin immunoprecipitation (HiChIP) sequencing of primary tissues to profoundly understand the genesis of UL. We identified somatic mutations in genes encoding six members of the SRCAP histone-loading complex4, and found that germline mutations in the SRCAP members YEATS4 and ZNHIT1 predispose women to UL. Tumours bearing these mutations showed defective deposition of the histone variant H2A.Z. In ULs, H2A.Z occupancy correlated positively with chromatin accessibility and gene expression, and negatively with DNA methylation, but these correlations were weak in tumours bearing SRCAP complex mutations. In these tumours, open chromatin emerged at transcription start sites where H2A.Z was lost, which was associated with upregulation of genes. Furthermore, YEATS4 defects were associated with abnormal upregulation of bivalent embryonic stem cell genes, as previously shown in mice5. Our work describes a potential mechanism of tumorigenesis-epigenetic instability caused by deficient H2A.Z deposition-and suggests that ULs arise through an aberrant differentiation program driven by deranged chromatin, emanating from a small number of mutually exclusive driver mutations.


Assuntos
Montagem e Desmontagem da Cromatina , Cromatina/genética , Cromatina/metabolismo , Histonas/deficiência , Leiomioma/genética , Mutação , Neoplasias Uterinas/genética , Carcinogênese/genética , Linhagem Celular , Cromatina/química , Células-Tronco Embrionárias/metabolismo , Epigênese Genética , Feminino , Regulação Neoplásica da Expressão Gênica , Histonas/genética , Histonas/metabolismo , Humanos , Leiomioma/metabolismo , Leiomioma/patologia , Ligases/genética , Complexo Repressor Polycomb 1/genética , Proteínas do Grupo Polycomb/genética , Fatores de Transcrição/genética , Neoplasias Uterinas/metabolismo , Neoplasias Uterinas/patologia
19.
Reproduction ; 162(5): 375-384, 2021 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-34423782

RESUMO

This study aimed to determine if the (pro)renin receptor (ATP6AP2) changes the cellular profile of choriocarcinomas from cytotrophoblast cells to terminally syncytialised cells and ascertain whether this impacts the invasive potential of choriocarcinoma cells. Additionally, we aimed to confirm that FURIN and/or site 1 protease (MBTPS1) cleave soluble ATP6AP2 (sATP6AP2) in BeWo choriocarcinoma cells and determine whether sATP6AP2 levels reflect the cellular profile of choriocarcinomas. BeWo choriocarcinoma cells were treated with ATP6AP2 siRNA, FURIN siRNA, DEC-RVKR-CMK (to inhibit FURIN activity), or PF 429242 (to inhibit MBTPS1 activity). Cells were also treated with forskolin, to induce syncytialisation, or vehicle and incubated for 48 h before collection of cells and supernatants. Syncytialisation was assessed by measuring hCG secretion (by ELISA) and E-cadherin protein levels (by immunoblot and immunocytochemistry). Cellular invasion was measured using the xCELLigence real-time cell analysis system and secreted sATP6AP2 levels measured by ELISA. Forskolin successfully induced syncytialisation and significantly increased both BeWo choriocarcinoma cell invasion (P < 0.0001) and sATP6AP2 levels (P = 0.02). Treatment with ATP6AP2 siRNA significantly inhibited syncytialisation (decreased hCG secretion (P = 0.005), the percent of nuclei in syncytia (P = 0.05)), forskolin-induced invasion (P = 0.046), and sATP6AP2 levels (P < 0.0001). FURIN siRNA and DEC-RVKR-CMK significantly decreased sATP6AP2 levels (both P < 0.0001). In conclusion, ATP6AP2 is important for syncytialisation of choriocarcinoma cells and thereby limits choriocarcinoma cell invasion. We postulate that sATP6AP2 could be used as a biomarker measuring the invasive potential of choriocarcinomas. Additionally, we confirmed that FURIN, not MBTPS1, cleaves sATP6AP2 in BeWo cells, but other proteases (inhibited by DEC-RVKR-CMK) may also be involved.


Assuntos
Coriocarcinoma , Receptores de Superfície Celular , Renina , Neoplasias Uterinas , ATPases Vacuolares Próton-Translocadoras , Coriocarcinoma/metabolismo , Colforsina/metabolismo , Colforsina/farmacologia , Feminino , Humanos , Gravidez , Receptores de Superfície Celular/metabolismo , Renina/metabolismo , Trofoblastos/metabolismo , Neoplasias Uterinas/metabolismo , ATPases Vacuolares Próton-Translocadoras/metabolismo
20.
Curr Issues Mol Biol ; 43(2): 802-817, 2021 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-34449552

RESUMO

Uterine fibroids are the most common mesenchymal uterine neoplasms; their prevalence is estimated in 40%-60% of women under 35 and in 70%-80% of women over 50 years of age. The current research aims to focus on the etiopathogenesis of uterine fibroids, the factors that affect their growth, and markers with diagnostic and prognostic properties. The MCM (minichromosome maintenance) protein family consists of peptides whose primary function is participation in the molecular mechanism of creating replication forks while regulating DNA synthesis. The aim of this work was to determine the proliferative potential of uterine fibroid cells based on the expression of the Ki-67 antigen and the MCMs-i.e., MCM-3, MCM-5, and MCM-7. In addition, the expression of estrogen (ER) and progesterone (PgR) receptors was evaluated and correlated with the expression of the abovementioned observations. Ultimately, received results were analyzed in terms of clinical and pathological data. MATERIALS AND METHODS: In forty-four cases of uterine fibroids, immunohistochemical reactions were performed. A tissue microarray (TMA) technique was utilized and analyzed cases were assessed in triplicate. Immunohistochemistry was performed using antibodies against Ki-67 antigen, ER, PgR, MCM-3, MCM-5, and MCM-8 on an automated staining platform. Reactions were digitalized by a histologic scanner and quantified utilizing dedicated software for nuclear analysis. Assessment was based on quantification expression of the three histiospots, each representing one case in TMA. RESULTS: In the study group (uterine fibroids), statistically significant stronger expression of all the investigated MCMs was observed, as compared to the control group. In addition, moderate and strong positive correlations were found between all tested proliferative markers. The expression of the MCM-7 protein also correlated positively with ER and PgR. With regard to clinical and pathological data, there was a negative correlation between the expression of MCMs and the number of both pregnancies and births. Significant reductions in MCM-5 and MCM-7 expression were observed in the group of women receiving oral hormonal contraceptives, while smoking women showed an increase in MCM-7, ER, and PgR. CONCLUSIONS: Uterine fibroid cells have greater proliferative potential, as evaluated by expression of the Ki-67 antigen and MCMs, than unaltered myometrial cells of the uterine corpus. The expression of MCM-7 was found to have strong or moderate correlations in all assessed relations. In the context of the clinical data, as well evident proliferative potential of MCMs, further studies are strongly recommended.


Assuntos
Proteínas de Ciclo Celular/biossíntese , Leiomioma/metabolismo , Componente 3 do Complexo de Manutenção de Minicromossomo/biossíntese , Componente 7 do Complexo de Manutenção de Minicromossomo/biossíntese , Neoplasias Uterinas/metabolismo , Adulto , Idoso , Biomarcadores Tumorais/metabolismo , Proteínas de Ciclo Celular/metabolismo , Receptor alfa de Estrogênio/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Leiomioma/patologia , Pessoa de Meia-Idade , Componente 3 do Complexo de Manutenção de Minicromossomo/metabolismo , Componente 7 do Complexo de Manutenção de Minicromossomo/metabolismo , Gravidez , Receptores de Progesterona/metabolismo , Neoplasias Uterinas/patologia
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