RESUMO
BACKGROUND: Although Huaier granules can be used as prospective anti-cholangiocarcinoma drugs, the mechanism of action of Huaier granules in cholangiocarcinoma is not clear. The anti-cholangiocarcinoma effect of Huaier granules was validated in cell line research. In vitro experiments were conducted to investigate the signalling pathways affected by Huaier in CCA cells. METHODS AND RESULTS: Real-time quantitative PCR (RTâqPCR) and Western blot analysis were performed to analyse gene expression in CCA cells. MTT assays, scratch tests, and Transwell assays were used to explore the effects on the proliferation and metastasis of CCA cells. Chromatin immunoprecipitation assays were performed to reveal the potential underlying mechanisms involved. Twist1 was upregulated in human CCA tissues. In addition, its expression levels were negatively related to FBP1 expression levels. Mechanistically, Twist1 can bind to the region of the FBP1 promoter to reduce its expression. Huaier plays an indispensable role in suppressing Twist1 expression to inhibit the Twist1/FBP1/Wnt/ß-catenin axis. Then, we verified the effect of Huaier in vitro. CONCLUSIONS: These findings suggested that Huaier granules were capable of inhibiting CCA development through regulating the Twist1/FBP1/Wnt/ß-catenin signalling axis and provided a novel orientation for the development of novel anti-CCA drugs.
Assuntos
Neoplasias dos Ductos Biliares , Proliferação de Células , Colangiocarcinoma , Regulação Neoplásica da Expressão Gênica , Proteínas Nucleares , Proteína 1 Relacionada a Twist , Via de Sinalização Wnt , beta Catenina , Humanos , Proteína 1 Relacionada a Twist/metabolismo , Proteína 1 Relacionada a Twist/genética , Colangiocarcinoma/metabolismo , Colangiocarcinoma/genética , Colangiocarcinoma/patologia , Colangiocarcinoma/tratamento farmacológico , Via de Sinalização Wnt/efeitos dos fármacos , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Linhagem Celular Tumoral , Neoplasias dos Ductos Biliares/metabolismo , Neoplasias dos Ductos Biliares/genética , Neoplasias dos Ductos Biliares/patologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , beta Catenina/metabolismo , Movimento Celular/efeitos dos fármacos , Movimento Celular/genéticaRESUMO
Lysine methyltransferase 5A (KMT5A) is the sole mammalian enzyme known to catalyse the monomethylation of histone H4 lysine 20 and nonhistone proteins such as p53, which are involved in the occurrence and progression of numerous cancers. The present study aimed to determine the function of KMT5A in inducing docetaxel (DTX) resistance in patients with breast carcinoma by evaluating glucose metabolism and the underlying mechanism involved. The upregulation or downregulation of KMT5Arelated proteins was examined after KMT5A knockdown in breast cancer (BRCA) cells by Tandem Mass Tag proteomics. Through differential protein expression and pathway enrichment analysis, the upregulated key gluconeogenic enzyme fructose1,6bisphosphatase 1 (FBP1) was discovered. Loss of FBP1 expression is closely related to the development and prognosis of cancers. A dualluciferase reporter gene assay confirmed that KMT5A inhibited the expression of FBP1 and that overexpression of FBP1 could enhance the chemotherapeutic sensitivity to DTX through the suppression of KMT5A expression. The KMT5A inhibitor UNC0379 was used to verify that DTX resistance induced by KMT5A through the inhibition of FBP1 depended on the methylase activity of KMT5A. According to previous literature and interaction network structure, it was revealed that KMT5A acts on the transcription factor twist family BHLH transcription factor 1 (TWIST1). Then, it was verified that TWSIT1 promoted the expression of FBP1 by using a dualluciferase reporter gene experiment. KMT5A induces chemotherapy resistance in BRCA cells by promoting cell proliferation and glycolysis. After the knockdown of the KMT5A gene, the FBP1 related to glucose metabolism in BRCA was upregulated. KMT5A knockdown expression and FBP1 overexpression synergistically inhibit cell proliferation and block cells in the G2/M phase. KMT5A inhibits the expression of FBP1 by methylating TWIST1 and weakening its promotion of FBP1 transcription. In conclusion, KMT5A was shown to affect chemotherapy resistance by regulating the cell cycle and positively regulate glycolysismediated chemotherapy resistance by inhibiting the transcription of FBP1 in collaboration with TWIST1. KMT5A may be a potential therapeutic target for chemotherapy resistance in BRCA.
Assuntos
Neoplasias da Mama , Docetaxel , Resistencia a Medicamentos Antineoplásicos , Frutose-Bifosfatase , Regulação Neoplásica da Expressão Gênica , Proteínas Nucleares , Proteína 1 Relacionada a Twist , Humanos , Neoplasias da Mama/genética , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Neoplasias da Mama/metabolismo , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Frutose-Bifosfatase/genética , Frutose-Bifosfatase/metabolismo , Docetaxel/farmacologia , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Proliferação de Células/efeitos dos fármacos , Metilação de DNARESUMO
Crohn's disease (CD) is marked by recurring intestinal inflammation and tissue injury, often resulting in fibrostenosis and bowel obstruction, necessitating surgical intervention with high recurrence rates. To elucidate the mechanisms underlying fibrostenosis in CD, we analyzed the transcriptome of cells isolated from the transmural ileum of patients with CD, including a trio of lesions from each patient: non-affected, inflamed, and stenotic ileum samples, and compared them with samples from patients without CD. Our computational analysis revealed that profibrotic signals from a subset of monocyte-derived cells expressing CD150 induced a disease-specific fibroblast population, resulting in chronic inflammation and tissue fibrosis. The transcription factor TWIST1 was identified as a key modulator of fibroblast activation and extracellular matrix (ECM) deposition. Genetic and pharmacological inhibition of TWIST1 prevents fibroblast activation, reducing ECM production and collagen deposition. Our findings suggest that the myeloid-stromal axis may offer a promising therapeutic target to prevent fibrostenosis in CD.
Assuntos
Doença de Crohn , Fibroblastos , Fibrose , Monócitos , Proteína 1 Relacionada a Twist , Doença de Crohn/metabolismo , Doença de Crohn/patologia , Doença de Crohn/imunologia , Humanos , Fibroblastos/metabolismo , Fibroblastos/patologia , Proteína 1 Relacionada a Twist/metabolismo , Proteína 1 Relacionada a Twist/genética , Monócitos/metabolismo , Monócitos/patologia , Monócitos/imunologia , Masculino , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Feminino , Matriz Extracelular/metabolismo , Matriz Extracelular/patologia , Receptores de Superfície Celular/metabolismo , Receptores de Superfície Celular/genética , Íleo/patologia , Íleo/metabolismo , Íleo/imunologia , Comunicação Celular , Adulto , Endopeptidases/metabolismo , Endopeptidases/genética , Animais , CamundongosRESUMO
Cutaneous melanoma is a lethal skin cancer variant with pronounced aggressiveness and metastatic potential. However, few targeted medications inhibit the progression of melanoma. Ganoderma lucidum, which is a type of mushroom, is widely used as a non-toxic alternative adjunct therapy for cancer patients. This study determines the effect of WSG, which is a water-soluble glucan that is derived from G. lucidum, on melanoma cells. The results show that WSG inhibits cell viability and the mobility of melanoma cells. WSG induces changes in the expression of epithelial-to-mesenchymal transition (EMT)-related markers. WSG also downregulates EMT-related transcription factors, Snail and Twist. Signal transduction assays show that WSG reduces the protein levels in transforming growth factor ß receptors (TGFßRs) and consequently inhibits the phosphorylation of intracellular signaling molecules, such as FAK, ERK1/2 and Smad2. An In vivo study shows that WSG suppresses melanoma growth in B16F10-bearing mice. To enhance transdermal drug delivery and prevent oxidation, two highly biocompatible compounds, polyvinyl alcohol (PVA) and polyvinylpyrrolidone (PVP), are used to synthesize a dissolvable microneedle patch that is loaded with WSG (MN-WSG). A functional assay shows that MN-WSG has an effect that is comparable to that of WSG alone. These results show that WSG has significant potential as a therapeutic agent for melanoma treatment. MN-WSG may allow groundbreaking therapeutic approaches and offers a novel method for delivering this potent compound effectively.
Assuntos
Reishi , Fatores de Transcrição da Família Snail , Animais , Camundongos , Reishi/química , Fatores de Transcrição da Família Snail/metabolismo , Humanos , Melanoma/tratamento farmacológico , Melanoma/patologia , Melanoma/metabolismo , Linhagem Celular Tumoral , Proteína 1 Relacionada a Twist/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/metabolismo , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/patologia , Melanoma Experimental/metabolismo , Movimento Celular/efeitos dos fármacos , Antineoplásicos/farmacologia , Antineoplásicos/química , Álcool de Polivinil/química , Polissacarídeos/farmacologia , Polissacarídeos/química , Transdução de Sinais/efeitos dos fármacosRESUMO
Epithelial-mesenchymal transition (EMT) plays a pivotal role in the development and progression of many cancers. Partial EMT (pEMT) could represent a critical step in tumor migration and dissemination. Sarcomatoid renal cell carcinoma (sRCC) is an aggressive form of renal cell carcinoma (RCC) composed of a carcinomatous (sRCC-Ca) and sarcomatous (sRCC-Sa) component. The role of (p)EMT in the progression of RCC to sRCC remains unclear. The aim of this study was to investigate the involvement of (p)EMT in RCC and sRCC. Tissue samples from 10 patients with clear cell RCC (ccRCC) and 10 patients with sRCC were selected. The expression of main EMT markers (miR-200 family, miR-205, SNAI1/2, TWIST1/2, ZEB1/2, CDH1/2, VIM) was analyzed by qPCR in ccRCC, sRCC-Ca, and sRCC-Sa and compared to non-neoplastic tissue and between both groups. Expression of E-cadherin, N-cadherin, vimentin and ZEB2 was analyzed using immunohistochemistry. miR-200c was downregulated in sRCC-Ca compared to ccRCC, while miR-200a was downregulated in sRCC-Sa compared to ccRCC. CDH1 was downregulated in sRCC-Sa when compared to any other group. ZEB2 was downregulated in ccRCC and sRCC compared to corresponding non-neoplastic kidney. A positive correlation was observed between CDH1 expression and miR-200a/b/c. Our results suggest that full EMT is not present in sRCC. Instead, discreet molecular differences exist between ccRCC, sRCC-Ca, and sRCC-Sa, possibly representing distinct intermediary states undergoing pEMT.
Assuntos
Biomarcadores Tumorais , Carcinoma de Células Renais , Transição Epitelial-Mesenquimal , Neoplasias Renais , MicroRNAs , Vimentina , Homeobox 2 de Ligação a E-box com Dedos de Zinco , Humanos , Transição Epitelial-Mesenquimal/genética , Carcinoma de Células Renais/patologia , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/metabolismo , Neoplasias Renais/patologia , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , MicroRNAs/genética , Masculino , Pessoa de Meia-Idade , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Feminino , Vimentina/metabolismo , Vimentina/genética , Homeobox 2 de Ligação a E-box com Dedos de Zinco/genética , Homeobox 2 de Ligação a E-box com Dedos de Zinco/metabolismo , Idoso , Caderinas/genética , Caderinas/metabolismo , Regulação Neoplásica da Expressão Gênica , Antígenos CD/genética , Antígenos CD/metabolismo , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismo , Fatores de Transcrição da Família Snail/genética , Fatores de Transcrição da Família Snail/metabolismo , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Transformação Celular Neoplásica/metabolismo , Adulto , Proteínas NuclearesRESUMO
OBJECTIVE: To evaluate and compare the expression of E-cadherin, Snail1 and Twist1 in pleomorphic adenomas (PAs), adenoid cystic carcinomas (AdCCa) and carcinoma ex-pleomorphic adenomas (CaexPA) of salivary glands, as well as investigate possible associations with clinicopathological parameters. STUDY DESIGN: E-cadherin, Snail1 and Twist1 antibody immunostaining were analyzed semiquantitatively in 20 PAs, 20 AdCCas and 10 CaexPAs. Cases were classified as low and high expression for analysis of the association with clinicopathological parameters. RESULTS: Compared to PAs, AdCCas and CaexPAs exhibited higher nuclear expression of Snail1 (p = 0.021 and p = 0.028, respectively) and Twist1 (p = 0.009 and p = 0.001). Membranous and cytoplasmic expression of E-cadherin were positively correlated in PAs, AdCCas and CaexPAs (r = 0.645, p = 0.002; r = 0.824, p < 0.001; r = 0.677, p = 0.031). In PAs, positive correlation was found between nuclear expression of Snail1 and membrane expression of E-cadherin (r = 0.634; p = 0.003), as well as between nuclear expression of Snail1 and Twist1 (r = 0.580; p = 0.007). Negative correlations were detected between membrane expression of E-cadherin and cytoplasmic expression of Snail1 in AdCCas (r = - 0.489; p = 0.029). CONCLUSIONS: E-cadherin, Twist1, and Snail1 may participate in modulating events related to cell differentiation and adhesion in PAs and to biological behavior in AdCCas and CaexPAs, which indicates the involvement of EMT in these processes. Furthermore, the expression of these proteins in these carcinomas may reflect the plasticity feature of EMT.
Assuntos
Adenoma Pleomorfo , Caderinas , Carcinoma Adenoide Cístico , Transição Epitelial-Mesenquimal , Proteínas Nucleares , Neoplasias das Glândulas Salivares , Fatores de Transcrição da Família Snail , Proteína 1 Relacionada a Twist , Humanos , Neoplasias das Glândulas Salivares/patologia , Neoplasias das Glândulas Salivares/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Caderinas/metabolismo , Feminino , Masculino , Proteína 1 Relacionada a Twist/metabolismo , Pessoa de Meia-Idade , Carcinoma Adenoide Cístico/patologia , Carcinoma Adenoide Cístico/metabolismo , Proteínas Nucleares/metabolismo , Adulto , Adenoma Pleomorfo/metabolismo , Adenoma Pleomorfo/patologia , Idoso , Fatores de Transcrição Twist/metabolismo , Imuno-Histoquímica , Fatores de Transcrição/metabolismo , Biomarcadores Tumorais/metabolismoRESUMO
INTRODUCTION: Radiation-induced pulmonary fibrosis (RIPF) is a chronic, progressive, irreversible lung interstitial disease that develops after radiotherapy. Although several previous studies have focused on the mechanism of epithelial-mesenchymal transition (EMT) in lung epithelial cells, the essential factors involved in this process remain poorly understood. The DNA-dependent protein kinase catalytic subunit (DNA-PKcs) exhibits strong repair capacity when cells undergo radiation-induced damage; whether DNA-PKcs regulates EMT during RIPF remains unclear. OBJECTIVES: To investigate the role and molecular mechanism of DNA-PKcs in RIPF and provide an important theoretical basis for utilising DNA-PKcs-targeted drugs for preventing RIPF. METHODS: DNA-PKcs knockout (DPK-/-) mice were generated via the Cas9/sgRNA technique and subjected to whole chest ionizing radiation (IR) at a 20 Gy dose. Before whole chest IR, the mice were intragastrically administered the DNA-PKcs-targeted drug VND3207. Lung tissues were collected at 1 and 5 months after IR. RESULTS: The expression of DNA-PKcs is low in pulmonary fibrosis (PF) patients. DNA-PKcs deficiency significantly exacerbated RIPF by promoting EMT in lung epithelial cells. Mechanistically, DNA-PKcs deletion by shRNA or inhibitor NU7441 maintained the protein stability of Twist1. Furthermore, AKT1 mediated the interaction between DNA-PKcs and Twist1. High Twist1 expression and EMT-associated changes caused by DNA-PKcs deletion were blocked by insulin-like growth factor-1 (IGF-1), an AKT1 agonist. The radioprotective drug VND3207 prevented IR-induced EMT and alleviated RIPF in mice by stimulating the kinase activity of DNA-PKcs. CONCLUSION: Our study clarified the critical role and mechanism of DNA-PKcs in RIPF and showed that it could be a potential target for preventing RIPF.
Assuntos
Proteína Quinase Ativada por DNA , Transição Epitelial-Mesenquimal , Proteínas Nucleares , Proteínas Proto-Oncogênicas c-akt , Fibrose Pulmonar , Proteína 1 Relacionada a Twist , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Animais , Proteína Quinase Ativada por DNA/metabolismo , Proteína Quinase Ativada por DNA/genética , Camundongos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Proteína 1 Relacionada a Twist/metabolismo , Proteína 1 Relacionada a Twist/genética , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/etiologia , Ubiquitinação , Humanos , Camundongos Knockout , Proteínas de Ligação a DNARESUMO
Colorectal cancer (CRC) is notable for its high mortality and high metastatic characteristics. The shear force generated by bloodstream provides mechanical signals regulating multiple responses of cells, including metastatic cancer cells, dispersing in blood vessels. We, therefore, studied the effect of shear flow on circulating CRC cells in the present study. The CRC cell line SW620 was subjected to shear flow of 12.5 dynes/cm2 for 1 and 2 h separately. Resulting elevated caspase-9 and -3 indicated that shear flow initiated the apoptosis of SW620. Enlarged cell size associated with a higher level of cyclin D1 was coincident with the flow cytometric results indicating that the cell cycle was arrested at the G1 phase. An elevated phosphor-eNOSS1177 increased the production of nitric oxide and led to reactive oxygen species-mediated oxidative stress. Shear flow also regulated epithelial-mesenchymal transition (EMT) by increasing E-cadherin and ZO-1 while decreasing Snail and Twist1. The migration and invasion of sheared SW620 were also substantially decreased. Further investigations showed that mitochondrial membrane potential was significantly decreased, whereas mitochondrial mass and ATP production were not changed. In addition to the shear flow of 12.5 dynes/cm2, the expressions of EMT were compared at lower (6.25 dynes/cm2) and at higher (25 dynes/cm2) shear flow. The results showed that lower shear flow increased mesenchymal characteristics and higher shear flow increased epithelial characteristics. Shear flow reduces the malignancy of CRC in their metastatic dispersal that opens up new ways to improve cancer therapies by applying a mechanical shear flow device.
Assuntos
Apoptose , Movimento Celular , Neoplasias Colorretais , Transição Epitelial-Mesenquimal , Espécies Reativas de Oxigênio , Humanos , Neoplasias Colorretais/patologia , Neoplasias Colorretais/metabolismo , Linhagem Celular Tumoral , Espécies Reativas de Oxigênio/metabolismo , Estresse Mecânico , Potencial da Membrana Mitocondrial , Ciclina D1/metabolismo , Estresse Oxidativo , Caderinas/metabolismo , Óxido Nítrico/metabolismo , Caspase 9/metabolismo , Caspase 3/metabolismo , Proteína da Zônula de Oclusão-1/metabolismo , Proteína 1 Relacionada a Twist/metabolismoRESUMO
Breast cancer is the most common invasive neoplasm and the leading cause of cancer death in women worldwide. The main cause of mortality in cancer patients is invasion and metastasis, where the epithelial-mesenchymal transition (EMT) is a crucial player in these processes. Pharmacological therapy has plants as its primary source, including isoflavonoids. Brazilin is an isoflavonoid isolated from Haematoxilum brasiletto that has shown antiproliferative activity in several cancer cell lines. In this study, we evaluated the effect of Brazilin on canonical markers of EMT such as E-cadherin, vimentin, Twist, and matrix metalloproteases (MMPs). By Western blot, we evaluated E-cadherin, vimentin, and Twist expression and the subcellular localization by immunofluorescence. Using gelatin zymography, we determined the levels of secretion of MMPs. We used Transwell chambers coated with matrigel to determine the in vitro invasion of breast cancer cells treated with Brazilin. Interestingly, our results show that Brazilin increases 50% in E-cadherin expression and decreases 50% in vimentin and Twist expression, MMPs, and cell invasion in triple-negative breast cancer (TNBC) MDA-MB-231 and to a lesser extend in MCF7 ER+ breast cancer cells. Together, these findings position Brazilin as a new molecule with great potential for use as complementary or alternative treatment in breast cancer therapy in the future.
Assuntos
Benzopiranos , Neoplasias da Mama , Caderinas , Transição Epitelial-Mesenquimal , Feminino , Humanos , Benzopiranos/farmacologia , Neoplasias da Mama/patologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Neoplasias da Mama/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Metaloproteinases da Matriz/metabolismo , Metaloproteinases da Matriz/genética , Células MCF-7 , Invasividade Neoplásica/genética , Proteínas Nucleares , Neoplasias de Mama Triplo Negativas/patologia , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/genética , Proteína 1 Relacionada a Twist/metabolismo , Proteína 1 Relacionada a Twist/genética , Vimentina/metabolismo , Vimentina/genéticaRESUMO
Synovial chondromatosis (SC) is a disorder of the synovium characterized by the formation of osteochondral nodules within the synovium. This study aimed to identify the abnormally differentiated progenitor cells and possible pathogenic signaling pathways. Loose bodies and synovium were obtained from patients with SC during knee arthroplasty. Single-cell RNA sequencing was used to identify cell subsets and their gene signatures in SC synovium. Cells derived from osteoarthritis (OA) synovium were used as controls. Multi-differentiation and colony-forming assays were used to identify progenitor cells. The roles of transcription factors and signaling pathways were investigated through computational analysis and experimental verification. We identified an increased proportion of CD34+ sublining fibroblasts in SC synovium. CD34+CD31- cells and CD34-CD31- cells were sorted from SC synovium. Compared with CD34- cells, CD34+ cells had larger alkaline phosphatase (ALP)-stained area and calcified area after osteogenic induction. In addition, CD34+ cells exhibited a stronger tube formation ability than CD34- cells. Our bioinformatic analysis suggested the expression of TWIST1, a negative regulator of osteogenesis, in CD34- sublining fibroblasts and was regulated by the TGF-ß signaling pathway. The experiment showed that CD34+ cells acquired the TWIST1 expression during culture and the combination of TGF-ß1 and harmine, an inhibitor of Twist1, could further stimulate the osteogenesis of CD34+ cells. Overall, CD34+ synovial fibroblasts in SC synovium have multiple differentiation potentials, especially osteogenic differentiation potential, and might be responsible for the pathogenesis of SC.
Assuntos
Antígenos CD34 , Condromatose Sinovial , Fibroblastos , Osteogênese , Membrana Sinovial , Humanos , Antígenos CD34/metabolismo , Fibroblastos/metabolismo , Fibroblastos/patologia , Condromatose Sinovial/patologia , Condromatose Sinovial/metabolismo , Membrana Sinovial/patologia , Membrana Sinovial/metabolismo , Feminino , Masculino , Pessoa de Meia-Idade , Diferenciação Celular , Idoso , Proteína 1 Relacionada a Twist/metabolismo , Proteína 1 Relacionada a Twist/genética , Proteínas NuclearesRESUMO
Cancer-associated Fibroblasts (CAFs) exert a tumor-promoting effect in various cancers, including breast cancer. CAFs secrete exosomes containing miRNA and proteins, influencing the tumor microenvironment. In this study, we identified CAF-derived exosomes that transport functional miR-92a from CAFs to tumor cells, thereby intensifying the aggressiveness of breast cancer. CAFs downregulate the expression of G3BP2 in breast cancer cells, and a significant elevation in miR-92a levels in CAF-derived exosomes was observed. Both in vitro and in vivo experiments demonstrate that miR-92a enhances breast cancer cell migration and invasion by directly targeting G3BP2, functioning as a tumor-promoting miRNA. We validated that the RNA-binding proteins SNRPA facilitate the transfer of CAF-derived exosomal miR-92a to breast cancer cells. The reduction of G3BP2 protein by CAF-derived exosomes releases TWIST1 into the nucleus, promoting epithelial-mesenchymal transition (EMT) and further exacerbating breast cancer progression. Moreover, CAF-derived exosomal miR-92a induces tumor invasion and metastasis in mice. Overall, our study reveals that CAF-derived exosomal miR-92a serves as a promoter in the migration and invasion of breast cancer cells by reducing G3BP2 and may represent a potential novel tumor marker for breast cancer.
Assuntos
Neoplasias da Mama , Fibroblastos Associados a Câncer , Movimento Celular , Transição Epitelial-Mesenquimal , Exossomos , Regulação Neoplásica da Expressão Gênica , MicroRNAs , Invasividade Neoplásica , Animais , Humanos , Camundongos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Neoplasias da Mama/patologia , Neoplasias da Mama/metabolismo , Neoplasias da Mama/genética , Fibroblastos Associados a Câncer/metabolismo , Fibroblastos Associados a Câncer/patologia , Proteínas de Transporte/metabolismo , Proteínas de Transporte/genética , Linhagem Celular Tumoral , Exossomos/metabolismo , MicroRNAs/metabolismo , MicroRNAs/genética , Metástase Neoplásica , Proteínas de Ligação a Poli-ADP-Ribose/metabolismo , Proteínas de Ligação a Poli-ADP-Ribose/genética , Proteínas de Ligação a RNA/metabolismo , Proteína 1 Relacionada a Twist/metabolismo , Proteína 1 Relacionada a Twist/genéticaRESUMO
MicroRNAs are small RNA molecules that have a significant role in translational repression and gene silencing through binding to downstream target mRNAs. MiR-762 can stimulate the proliferation and metastasis of various types of cancer. Hippo pathway is one of the pathways that regulate tissue development and carcinogenesis. Dysregulation of this pathway plays a vital role in the progression of cancer. This study aimed to evaluate the possible correlation between miR-762, the Hippo signaling pathway, TWIST1, and SMAD3 in patients with lung cancer, as well as patients with chronic inflammatory diseases. The relative expression of miR-762, MST1, LATS2, YAP, TWIST1, and SMAD3 was determined in 50 lung cancer patients, 30 patients with chronic inflammatory diseases, and 20 healthy volunteers by real-time PCR. The levels of YAP protein and neuron-specific enolase were estimated by ELISA and electrochemiluminescence immunoassay, respectively. Compared to the control group, miR-762, YAP, TWIST1, and SMAD3 expression were significantly upregulated in lung cancer patients and chronic inflammatory patients, except SMAD3 was significantly downregulated in chronic inflammatory patients. MST1, LATS2, and YAP protein were significantly downregulated in all patients. MiR-762 has a significant negative correlation with MST1, LATS2, and YAP protein in lung cancer patients and with MST1 and LATS2 in chronic inflammatory patients. MiR-762 may be involved in the induction of malignant behaviors in lung cancer through suppression of the Hippo pathway. MiR-762, MST1, LATS2, YAP mRNA and protein, TWIST1, and SMAD3 may be effective diagnostic biomarkers in both lung cancer patients and chronic inflammatory patients. High YAP, TWIST1, SMA3 expression, and NSE level are associated with a favorable prognosis for lung cancer.
Assuntos
Neoplasias Pulmonares , MicroRNAs , Humanos , Via de Sinalização Hippo , Transdução de Sinais , Neoplasias Pulmonares/genética , Linhagem Celular Tumoral , MicroRNAs/genética , MicroRNAs/metabolismo , Doença Crônica , Proliferação de Células/genética , Proteína Smad3/genética , Proteína Smad3/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismoRESUMO
Traditional clinical modalities for diagnosing bladder urothelial carcinoma (BUC) remain limited due to their invasive nature, significant costs, discomfort associated with cystoscopy, and low sensitivity to urine cytology. Therefore, there is an urgent need to identify highly sensitive, specific, and noninvasive biomarkers for the early detection of this neoplasm. Hypermethylated TWIST1/Vimentin promoter may be a noninvasive biomarker using urine sample. We assessed the TWIST1/Vimentin promoter methylation status in urine samples using the Methylated Human TWIST1 and Vimentin Gene Detection Kit (Jiangsu MicroDiag Biomedicine Co., Ltd., China). The samples were collected from five groups: group 1 consisted of patients with BUC, group 2 contained other patients with urologic tumors, group 3 consisted of patients with benign diseases (e.g., urinary tract infections, lithiasis, and benign prostatic hyperplasia), Group 4 included UTUC (upper tract urothelial carcinoma) patients and group5 comprised healthy individuals. The study encompassed 77 BUC patients, and we evaluated the degree of methylation of the TWIST1/Vimentin gene in their urine samples. Notably, TWIST1/Vimentin positivity was significantly elevated in comparison to groups 2, 3 and 5 (all p < 0.001) at a rate of 77.9%, but no significant difference was observed when compared to group 4. In the relationship between TWIST1/Vimentin methylation and clinicopathological features of BC patients from our center, we found there was no significant association between TWIST1/Vimentin status and proteinuria and/or hematuria, and hypermethylation of TWIST1 / VIM genes was found in both high and low tumor grade and in both non-muscle invasive bladder cancer (stages Tis, Ta, or T1) and muscle-invasive bladder cancer (stage T2 or above). In the multivariable analysis for cancer detection, a positive TWIST1/Vimentin methylation were significantly linked to a heightened risk of BC. Moreover, TWIST1/Vimentin promoter methylation demonstrated an ability to detect BUC in urine samples with a sensitivity of 78% and a specificity of 83%. Our findings reveal that hypermethylation of the TWIST1/Vimentin promoter occurs in bladder urothelial carcinoma, and its high sensitivity and specificity suggest its potential as a screening and therapeutic biomarker for urothelial carcinoma of the bladder.
Assuntos
Carcinoma de Células de Transição , Neoplasias da Bexiga Urinária , Humanos , Neoplasias da Bexiga Urinária/diagnóstico , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/patologia , Carcinoma de Células de Transição/patologia , Bexiga Urinária/patologia , Vimentina/genética , Biomarcadores Tumorais/metabolismo , Metilação de DNA/genética , Proteínas Nucleares/genética , Proteína 1 Relacionada a Twist/genéticaRESUMO
BACKGROUND: Luteolin, a common dietary flavonoid found in plants, has been shown to have anti-cancer properties. However, its exact mechanisms of action in non-small cell lung cancer (NSCLC) are still not fully understood, particularly its role in regulating broader genomic networks and specific gene targets. In this study, we aimed to elucidate the role of microRNAs (miRNAs) in NSCLC treated with luteolin, using A549 cells as a model system. MATERIALS AND METHODS: miRNA profiling was conducted on luteolin-treated A549 cells using Exiqon microarrays, with validation of selected miRNAs by qRT-PCR. Bioinformatic analysis identified the regulatory roles of miRNAs in biological processes and pathways following luteolin treatment. Computational algorithms were employed to identify potential target genes. A549 cells were transfected with miR-106a-5p mimic and inhibitor or their corresponding controls. The expression levels of 2 genes, twist basic helix-loop-helix transcription factor 1 (TWIST1) and matrix metallopeptidase 2 (MMP2), and cell migration were assessed. RESULTS: miRNA profiling identified 341 miRNAs, with 18 exhibiting significantly altered expression (P < 0.05). Subsequent qRT-PCR analysis confirmed altered expression of 6 selected miRNAs. KEGG and GO analyses revealed significant alterations in pathways and biological processes crucial for tumor biology. TWIST1 and MMP2, which both contain conserved miR-106a-5p binding sites, exhibited an inverse correlation with the expression levels of miR-106a-5p. Dual-luciferase reporter assays confirmed TWIST1 and MMP2 as direct targets of miR-106a-5p. Luteolin treatment led to a reduction in A549 cell migration, and this reduction was further amplified by the overexpression of miR-106a-5p. CONCLUSION: Luteolin inhibits A549 cell migration by modulating the miRNA landscape, shedding light on its mechanisms and laying the foundation for miRNA-based therapeutic approaches for NSCLC.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Movimento Celular , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares , Luteolina , Metaloproteinase 2 da Matriz , MicroRNAs , Proteínas Nucleares , Proteína 1 Relacionada a Twist , Regulação para Cima , Humanos , Luteolina/farmacologia , MicroRNAs/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/tratamento farmacológico , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismo , Células A549 , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Regulação para Cima/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genéticaRESUMO
Background/Aims: While DNA methylation and gastric microbiome are each associated with gastric cancer (GC), their combined role in predicting GC remains unclear. This study investigated the potential of a combined DNA methylation and gastric microbiome signature to predict Helicobacter pylori-negative GC. Methods: In this case-control study, we conducted quantitative methylation-specific polymerase chain reaction to measure the methylation levels of DKK3, SFRP1, EMX1, NKX6-1, MIR124-3, and TWIST1 in the gastric mucosa from 75 H. pylori-negative patients, including chronic gastritis (CG), intestinal metaplasia (IM), and GC. A combined analysis of DNA methylation and gastric microbiome, using 16S rRNA gene sequencing, was performed in 30 of 75 patients. Results: The methylation levels of DKK3, SFRP1, EMX1, MIR124-3, and TWIST1 were significantly higher in patients with GC than in controls (all q<0.05). MIR124-3 and TWIST1 methylation levels were higher in patients with IM than those with CG and also in those with GC than in those with IM (all q<0.05). A higher methylation level of TWIST1 was an independent predictor for H. pylori-negative GC after adjusting for age, sex, and atrophy (odds ratio [OR], 15.15; 95% confidence interval [CI], 1.58 to 145.46; p=0.018). The combination of TWIST1 methylation and GC microbiome index (a microbiome marker) was significantly associated with H. pylori-negative GC after adjusting for age, sex, and atrophy (OR, 50.00; 95% CI, 1.69 to 1,476; p=0.024). Conclusions: The combination of TWIST1 methylation and GC microbiome index may offer potential as a biomarker for predicting H. pylori-negative GC.
Assuntos
Metilação de DNA , Mucosa Gástrica , Microbioma Gastrointestinal , Helicobacter pylori , Neoplasias Gástricas , Humanos , Masculino , Feminino , Neoplasias Gástricas/microbiologia , Neoplasias Gástricas/genética , Pessoa de Meia-Idade , Estudos de Casos e Controles , Helicobacter pylori/genética , Mucosa Gástrica/microbiologia , Microbioma Gastrointestinal/genética , Proteína 1 Relacionada a Twist/genética , Idoso , MicroRNAs/análise , Proteínas Nucleares/genética , Gastrite/microbiologia , Gastrite/genética , Biomarcadores Tumorais/genética , RNA Ribossômico 16S/análise , RNA Ribossômico 16S/genética , Infecções por Helicobacter/microbiologia , Metaplasia/microbiologia , Metaplasia/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Adulto , Peptídeos e Proteínas de Sinalização Intercelular/genética , Proteínas de HomeodomínioRESUMO
MET amplification/mutations are important targetable oncogenic drivers in NSCLC, however, acquired resistance is inevitable and the majority of patients with targetable MET alterations fail to respond to MET tyrosine kinase inhibitors (TKIs). Furthermore, MET amplification is among the most common mediators of TKI resistance. As such, novel therapies to target MET pathway and overcome MET TKI resistance are clearly needed. Here we show that the epithelial-mesenchymal transition (EMT) transcription factor, TWIST1 is a key downstream mediator of HGF/MET induced resistance through suppression of p27 and targeting TWIST1 can overcome resistance. We found that TWIST1 is overexpressed at the time of TKI resistance in multiple MET-dependent TKI acquired resistance PDX models. We have shown for the first time that MET directly stabilized the TWIST protein leading to TKI resistance and that TWIST1 was required for MET-driven lung tumorigenesis as well as could induce MET TKI resistance when overexpressed. TWIST1 mediated MET TKI resistance through suppression of p27 expression and genetic or pharmacologic inhibition of TWIST1 overcame TKI resistance in vitro and in vivo. Our findings suggest that targeting TWIST1 may be an effective therapeutic strategy to overcome resistance in MET-driven NSCLC as well as in other oncogene driven subtypes in which MET amplification is the resistance mechanism.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Fator de Crescimento de Hepatócito , Neoplasias Pulmonares , Proteínas Nucleares , Inibidores de Proteínas Quinases , Proteínas Proto-Oncogênicas c-met , Proteína 1 Relacionada a Twist , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismo , Proteínas Proto-Oncogênicas c-met/genética , Proteínas Proto-Oncogênicas c-met/metabolismo , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Resistencia a Medicamentos Antineoplásicos/genética , Fator de Crescimento de Hepatócito/genética , Fator de Crescimento de Hepatócito/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Animais , Camundongos , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Transdução de Sinais/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Transição Epitelial-Mesenquimal/genética , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Oncogenes/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Lung adenocarcinoma (LUAD) is a common and deadly form of lung cancer, with high rates of metastasis and unsatisfactory clinical outcomes. Herein, we examined the influence of TMEM158 on the LUAD progression. A combination of bioinformatic analyses was used to assess the TMEM158 expression pattern, prognostic implications, and potential function in LUAD. The levels of TMEM158 and TWIST1 were evaluated in clinical samples from LUAD patients using Western blot analysis and qRT-PCR. To discover the function and underlying molecular pathways of TMEM158 in LUAD, we employed a combination of experimental approaches in vitro, such as flow cytometry analysis and colony formation, Co-IP, CCK-8, Transwell, and wound-healing assays. Elevated expression of TMEM158 in LUAD is associated with increased cancer aggressiveness and a poor prognosis. In vitro experiments demonstrated that high levels of TMEM158 promote cell proliferation, progression through the cell cycle, migration, and invasion while suppressing apoptosis. Knockdown of TMEM158 produced opposite effects. The underlying mechanism involves TMEM158 and TWIST1 directly interacting, stimulating the PI3K/AKT signaling pathway in LUAD cells. This investigation emphasizes the molecular functions of TMEM158 in LUAD progression and proposes targeting it as a promising treatment approach for managing LUAD.
Assuntos
Adenocarcinoma de Pulmão , Neoplasias Pulmonares , Humanos , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Oncogenes , Adenocarcinoma de Pulmão/genética , Neoplasias Pulmonares/genética , Proliferação de Células/genética , Movimento Celular/genética , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/genética , Proteínas Nucleares/genética , Proteína 1 Relacionada a Twist/genética , Proteínas de Membrana/genética , Proteínas Supressoras de TumorRESUMO
BACKGROUND: Transcription factors are important in the epithelial-mesenchymal transition process and are possibly related to the development of a more invasive tumor phenotype. Thus, the objective of this study was to analyze the expression and identify the localization of cellular markers related to the epithelial-mesenchymal transition process in salivary gland tumors. STUDY DESIGN: The expression and localization of E-CADERIN, N-CADERIN, SLUG, SNAIL, and TWIST were evaluated, using immunohistochemistry, in 48 salivary gland tumors, being 17 pleomorphic adenomas (PA), 14 adenoid cystic carcinomas (ACC), and 17 mucoepidermoid carcinomas (MEC). these proteins were compared to clinical and histopathologic parameters. normal gland tissues were included for immunohistochemical comparisons. RESULTS: ACC and MEC cases showed higher expression of SNAIL compared to PA. MEC showed high expression of SLUG and TWIST. Low expression of N-CADHERIN, SNAIL, and TWIST in ACC was frequent in T3 and T4. High expression of TWIST in MEC was more frequent at age ≥ 40 years A positive correlation was only observed between N-cadherin/SNAIL in ACC, between SNAIL/TWIST in MEC, and between SLUG/TWIST in PA. CONCLUSION: This study provided insight into EMT-related proteins (E-cadherin, N-cadherin, SNAIL, SLUG, and TWIST) and their contribution to the maintenance of morphogenesis and the development of the salivary gland tumors and showed a positive correlation among N-CADHERIN/SNAIL in ACC and SNAIL/TWIST in MEC.
Assuntos
Adenoma Pleomorfo , Carcinoma Adenoide Cístico , Carcinoma Mucoepidermoide , Neoplasias das Glândulas Salivares , Humanos , Adulto , Fatores de Transcrição da Família Snail , Proteínas Nucleares/genética , Neoplasias das Glândulas Salivares/patologia , Carcinoma Adenoide Cístico/patologia , Carcinoma Mucoepidermoide/patologia , Adenoma Pleomorfo/patologia , Caderinas/genética , Transição Epitelial-Mesenquimal/genética , Biomarcadores Tumorais , Proteína 1 Relacionada a Twist/genéticaRESUMO
Aberrant activation of the epithelial-mesenchymal transition (EMT) pathway drives the development of solid tumors, which is precisely regulated by core EMT-related transcription factors, including Twist1. However, the expression pattern and regulatory mechanism of Twist1 in the progression of bladder cancer is still unclear. In this study, we explore the role of Twist1 in the progression of bladder cancer. We discovered that the EMT regulon Twist1 protein, but not Twist1 mRNA, is overexpressed in bladder cancer samples using RT-qPCR, western blot and immunohistochemistry (IHC). Mechanistically, co-immunoprecipitation (Co-IP) coupled with liquid chromatography and tandem mass spectrometry identified USP5 as a binding partner of Twist1, and the binding of Twist1 to ubiquitin-specific protease 5 (USP5) stabilizes Twist through its deubiquitinase activity to activate the EMT. Further studies found that USP5 depletion reduces cell proliferation, invasion and the EMT in bladder cancer cells, and ectopic expression of Twist1 rescues the adverse effects of USP5 loss on cell invasion and the EMT. A xenograft tumor model was used to reconfirmed the inhibitor effect of silencing USP5 expression on tumorigenesis in vivo. In addition, USP5 protein levels are significantly elevated and positively associated with Twist1 levels in clinical bladder cancer samples. Collectively, our study revealed that USP5-Twist1 axis is a novel regulatory mechanism driving bladder cancer progression and that approaches targeting USP5 may become a promising cancer treatment strategy.
Assuntos
Proteína 1 Relacionada a Twist , Neoplasias da Bexiga Urinária , Humanos , Animais , Proteína 1 Relacionada a Twist/genética , Neoplasias da Bexiga Urinária/genética , Bexiga Urinária , Transformação Celular Neoplásica , Modelos Animais de Doenças , Proteases Específicas de UbiquitinaRESUMO
Anaplastic thyroid carcinoma (ATC) is an extremely difficult disease to tackle, with an overall patient survival of only a few months. The currently used therapeutic drugs, such as kinase inhibitors or immune checkpoint inhibitors, can prolong patient survival but fail to eradicate the tumor. In addition, the onset of drug resistance and adverse side-effects over time drastically reduce the chances of treatment. We recently showed that Twist1, a transcription factor involved in the epithelial mesenchymal transition (EMT), was strongly upregulated in ATC, and we wondered whether it might represent a therapeutic target in ATC patients. To investigate this hypothesis, the effects of harmine, a ß-carboline alkaloid shown to induce degradation of the Twist1 protein and to possess antitumoral activity in different cancer types, were evaluated on two ATC-derived cell lines, BHT-101 and CAL-62. The results obtained demonstrated that, in both cell lines, harmine reduced the level of Twist1 protein and reverted the EMT, as suggested by the augmentation of E-cadherin and decrease in fibronectin expression. The drug also inhibited cell proliferation and migration in a dose-dependent manner and significantly reduced the anchorage-independent growth of both ATC cell lines. Harmine was also capable of inducing apoptosis in BHT-101 cells, but not in CAL-62 ones. Finally, the activation of PI3K/Akt signaling, but not that of the MAPK, was drastically reduced in treated cells. Overall, these in vitro data suggest that harmine could represent a new therapeutic option for ATC treatment.