RESUMO
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
Robinow syndrome is a rare disease caused by variants of seven WNT pathway genes. Craniofacial features include widening of the nasal bridge and jaw hypoplasia. We used the chicken embryo to test whether two missense human FZD2 variants (1301G>T, p.Gly434Val; 425C>T, p.Pro142Lys) were sufficient to change frontonasal mass development. In vivo, the overexpression of retroviruses with wild-type or variant human FZD2 inhibited upper beak ossification. In primary cultures, wild-type and variant human FZD2 significantly inhibited chondrogenesis, with the 425C>T variant significantly decreasing activity of a SOX9 luciferase reporter compared to that for the wild type or 1301G>T. Both variants also increased nuclear shuttling of ß-catenin (CTNNB1) and increased the expression of TWIST1, which are inhibitory to chondrogenesis. In canonical WNT luciferase assays using frontonasal mass cells, the variants had dominant-negative effects on wild-type FZD2. In non-canonical assays, the 425C>T variant failed to activate the reporter above control levels and was unresponsive to exogenous WNT5A. This is the first single amino acid change to selectively alter ligand binding in a FZD receptor. Therefore, FZD2 missense variants are pathogenic and could lead to the altered craniofacial morphogenesis seen in Robinow syndrome.
Assuntos
Condrogênese , Anormalidades Craniofaciais , Receptores Frizzled , Animais , Embrião de Galinha , Humanos , Bico , beta Catenina/metabolismo , Núcleo Celular/metabolismo , Condrogênese/genética , Anormalidades Craniofaciais/genética , Anormalidades Craniofaciais/patologia , Nanismo , Receptores Frizzled/genética , Receptores Frizzled/metabolismo , Deformidades Congênitas dos Membros , Crânio/patologia , Crânio/embriologia , Proteína 1 Relacionada a Twist/metabolismo , Proteína 1 Relacionada a Twist/genética , Anormalidades Urogenitais , Via de Sinalização WntRESUMO
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
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
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
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
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
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
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
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
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
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
BACKGROUND: Diagnosis of early mycosis fungoides (eMF) is challenging and often delayed as many of its clinical and histopathologic features may mimic various benign inflammatory dermatoses (BIDs). The products of the thymocyte selection-associated high mobility group box (TOX), twist family BHLH transcription factor 1 (TWIST1), signal transducer and activator of transcription 4 (STAT4), and special AT-rich sequence-binding protein 1 (SATB1) genes function as transcription factors and are involved in the pathogenesis of MF. OBJECTIVES: We aim to determine the diagnostic value of TOX, TWIST1, STAT4, and SATB1 protein expressions in eMF. METHODS: This non-randomized, controlled, prospective analytic study was conducted by performing immunohistochemistry staining with TOX, TWIST1, STAT4, and SATB1 polyclonal antibodies in lesional skin biopsies of eMF and BID patients. Nuclear staining of lymphocytes was compared between eMF and BIDs, and the capacity of these antibodies to predict eMF was determined. RESULTS: Immunostainings with anti-TWIST1 showed an increase in protein expression (p = 0.003) and showed a decrease with anti-SATB1 antibodies in eMF compared to BIDs (p = 0.005) while anti-TOX and anti-STAT4 antibodies did not exhibit significant differences (p = 0.384; p = 0.150). Receiver operating characteristic analysis showed that immunohistochemical evaluations of TWIST1 and SATB1 protein expressions can differentiate eMF (area under the curve [AUC]: 0.728, 95% confidence interval [CI]: 0.605-0.851, p = 0.002; AUC: 0.686, 95% CI: 0.565-0.807, p = 0.013). CONCLUSIONS: TWIST1 and SATB1 are potential diagnostic markers for the histologic diagnosis of eMF.
Assuntos
Proteínas de Ligação à Região de Interação com a Matriz , Micose Fungoide , Neoplasias Cutâneas , Humanos , Proteínas de Ligação à Região de Interação com a Matriz/metabolismo , Micose Fungoide/patologia , Proteínas Nucleares/metabolismo , Estudos Prospectivos , Neoplasias Cutâneas/patologia , Fator de Transcrição STAT4/metabolismo , Proteína 1 Relacionada a Twist/metabolismoRESUMO
Jagged1 (JAG1) is a Notch ligand that correlates with tumor progression. Not limited to its function as a ligand, JAG1 can be cleaved, and its intracellular domain translocates to the nucleus, where it functions as a transcriptional cofactor. Previously, we showed that JAG1 intracellular domain (JICD1) forms a protein complex with DDX17/SMAD3/TGIF2. However, the molecular mechanisms underlying JICD1-mediated tumor aggressiveness remains unclear. Here, we demonstrate that JICD1 enhances the invasive phenotypes of glioblastoma cells by transcriptionally activating epithelial-to-mesenchymal transition (EMT)-related genes, especially TWIST1. The inhibition of TWIST1 reduced JICD1-driven tumor aggressiveness. Although SMAD3 is an important component of transforming growth factor (TGF)-ß signaling, the JICD1/SMAD3 transcriptional complex was shown to govern brain tumor invasion independent of TGF-ß signaling. Moreover, JICD1-TWIST1-MMP2 and MMP9 axes were significantly correlated with clinical outcome of glioblastoma patients. Collectively, we identified the JICD1/SMAD3-TWIST1 axis as a novel inducer of invasive phenotypes in cancer cells.
Assuntos
Glioblastoma , Humanos , Linhagem Celular Tumoral , Transição Epitelial-Mesenquimal/genética , Glioblastoma/genética , Proteínas de Homeodomínio/metabolismo , Ligantes , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Repressoras/metabolismo , Transdução de Sinais , Proteína Smad3/genética , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismoRESUMO
Tumor metastasis is the major cause of breast cancer morbidity and mortality. It has been reported that the F-box protein FBXO3 functions as an E3 ubiquitin ligase in regulating various biological processes, including host autoimmune, antiviral innate immunity, and inflammatory response. However, the role of FBXO3 in tumor metastasis remains elusive. We have previously shown that ΔNp63α is a common inhibitory target in oncogene-induced cell motility and tumor metastasis. In this study, we show that FBXO3 plays a vital role in PI3K-mediated breast cancer metastasis independent of its E3 ligase activity and ΔNp63α in breast cancer cells and in mouse. FBXO3 can bind to and stabilize USP4, leading to Twist1 protein stabilization and increased breast cancer cell migration and tumor metastasis. Mechanistically, FBXO3 disrupts the interaction between USP4 and aspartyl aminopeptidase (DNPEP), thereby protecting USP4 from DNPEP-mediated degradation. Furthermore, p110αH1047R facilitates the phosphorylation and stabilization of FBXO3 in an ERK1-dependent manner. Knockdown of either FBXO3 or USP4 leads to significant inhibition of PI3K-induced breast cancer metastasis. Clinically, elevated expression of p110α/FBXO3/USP4/Twist1 is associated with poor overall survival (OS) and recurrence-free survival (RFS) of breast cancer patients. Taken together, this study reveals that the FBXO3-USP4-Twist1 axis is pivotal in PI3K-mediated breast tumor metastasis and that FBXO3/USP4 may be potential therapeutic targets for breast cancer treatment.
Assuntos
Neoplasias da Mama , Melanoma , Neoplasias Cutâneas , Animais , Feminino , Humanos , Camundongos , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Fosfatidilinositol 3-Quinases/metabolismo , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismo , Proteases Específicas de Ubiquitina/metabolismo , UbiquitinaçãoRESUMO
Cyclin-dependent kinase (CDK) 4/6 inhibitors have significantly improved progression-free survival in hormone-receptor-positive (HR+), human-epidermal-growth-factor-receptor-type-2-negative (HER2-) metastatic luminal breast cancer (mLBC). Several studies have shown that in patients with endocrine-sensitive or endocrine-resistant LBC, the addition of CDK4/6 inhibitors to endocrine therapy significantly prolongs progression-free survival. However, the percentage of patients who are unresponsive or refractory to these therapies is as high as 40%, and no reliable and reproducible biomarkers have been validated to select a priori responders or refractory patients. The selection of mutant clones in the target oncoprotein is the main cause of resistance. Other mechanisms such as oncogene amplification/overexpression or mutations in other pathways have been described in several models. In this study, we focused on palbociclib, a selective CDK4/6 inhibitor. We generated a human MCF-7 luminal breast cancer cell line that was able to survive and proliferate at different concentrations of palbociclib and also showed cross-resistance to abemaciclib. The resistant cell line was characterized via RNA sequencing and was found to strongly activate the epithelial-to-mesenchymal transition. Among the top deregulated genes, we found a dramatic downregulation of the CDK4 inhibitor CDKN2B and an upregulation of the TWIST1 transcription factor. TWIST1 was further validated as a target for the reversal of palbociclib resistance. This study provides new relevant information about the mechanisms of resistance to CDK4/6 inhibitors and suggests potential new markers for patients' follow-up care during treatment.
Assuntos
Neoplasias da Mama , Humanos , Feminino , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Regulação para Cima , Quinase 4 Dependente de Ciclina , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Intervalo Livre de Progressão , Quinase 6 Dependente de Ciclina , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismoRESUMO
Mesenchymal stem/stromal cells (MSCs) have been evaluated in >1500 clinical trials, but outcomes remain suboptimal because of knowledge gaps in quality attributes that confer potency. We show that TWIST1 directly represses TSG6 expression that TWIST1 and TSG6 are inversely correlated across bone marrow-derived MSC (BM-MSC) donor cohorts and predict interdonor differences in their proangiogenic, anti-inflammatory, and immune suppressive activity in vitro and in sterile inflammation and autoimmune type 1 diabetes preclinical models. Transcript profiling of TWIST1HiTSG6Low versus TWISTLowTSG6Hi BM-MSCs revealed previously unidentified roles for TWIST1/TSG6 in regulating cellular oxidative stress and TGF-ß2 in modulating TSG6 expression and anti-inflammatory activity. TWIST1 and TSG6 levels also correlate to donor stature and predict differences in iPSC-derived MSC quality attributes. These results validate TWIST1 and TSG6 as biomarkers that predict interdonor differences in potency across laboratories and assay platforms, thereby providing a means to manufacture MSC products tailored to specific diseases.
Assuntos
Células-Tronco Mesenquimais , Humanos , Anti-Inflamatórios/farmacologia , Biomarcadores/metabolismo , Células da Medula Óssea/metabolismo , Diferenciação Celular , Fatores Imunológicos/metabolismo , Células-Tronco Mesenquimais/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismoRESUMO
Glioma cells are characterized by high migration ability, resulting in the aggressive growth of the tumors and poor prognosis of patients. Epithelial-to-mesenchymal transition (EMT) is one of the most important steps for tumor migration and metastasis and be elevated during glioma progression and recurrence. Twist1 is a basic helix-loop-helix transcription factor and a key transcription factor involved in the process of EMT. Twist1 is related to glioma mesenchymal change, invasion, heterogeneity, self-renewal of tumor stem cells, angiogenesis, etc., and may be used as a prognostic indicator and therapeutic target for glioma patients. This paper mainly reviews the structural characteristics, regulatory mechanisms, and apparent regulation of Twist1, as well as the roles of Twist1 during glioma progression and recurrence, providing new revelations for its use as a potential drug target and glioma treatment research.