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1.
Gut ; 2023 Nov 24.
Artigo em Inglês | MEDLINE | ID: mdl-38050079

RESUMO

OBJECTIVES: Cholangiocarcinoma (CCA) is a heterogeneous malignancy with high mortality and dismal prognosis, and an urgent clinical need for new therapies. Knowledge of the CCA epigenome is largely limited to aberrant DNA methylation. Dysregulation of enhancer activities has been identified to affect carcinogenesis and leveraged for new therapies but is uninvestigated in CCA. Our aim is to identify potential therapeutic targets in different subtypes of CCA through enhancer profiling. DESIGN: Integrative multiomics enhancer activity profiling of diverse CCA was performed. A panel of diverse CCA cell lines, patient-derived and cell line-derived xenografts were used to study identified enriched pathways and vulnerabilities. NanoString, multiplex immunohistochemistry staining and single-cell spatial transcriptomics were used to explore the immunogenicity of diverse CCA. RESULTS: We identified three distinct groups, associated with different etiologies and unique pathways. Drug inhibitors of identified pathways reduced tumour growth in in vitro and in vivo models. The first group (ESTRO), with mostly fluke-positive CCAs, displayed activation in estrogen signalling and were sensitive to MTOR inhibitors. Another group (OXPHO), with mostly BAP1 and IDH-mutant CCAs, displayed activated oxidative phosphorylation pathways, and were sensitive to oxidative phosphorylation inhibitors. Immune-related pathways were activated in the final group (IMMUN), made up of an immunogenic CCA subtype and CCA with aristolochic acid (AA) mutational signatures. Intratumour differences in AA mutation load were correlated to intratumour variation of different immune cell populations. CONCLUSION: Our study elucidates the mechanisms underlying enhancer dysregulation and deepens understanding of different tumourigenesis processes in distinct CCA subtypes, with potential significant therapeutics and clinical benefits.

2.
Tumour Biol ; 39(11): 1010428317725925, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29110582

RESUMO

Cholangiocarcinoma is a rare type of cancer which is an increasingly discernible health threat. The disease is usually very difficult in diagnosis and various treatment modalities are typically not effective. Cholangiocarcinoma is a complex and very heterogeneous malignancy characterized by tumor location, different risk factors, molecular profiling, and prognosis. Cancer cell lines represent an important tool for investigation in various aspects of tumor biology and molecular therapeutics. We established two cell lines, KKU-452 and KKU-023, which were derived from patients residing in the endemic area of liver fluke infection in Thailand. Both of tumor tissues have gross pathology of perihilar and intrahepatic mass-forming cholangiocarcinoma. Two cell lines were characterized for their biological, molecular and genetic properties. KKU-452 and KKU-023 cells are both adherent cells with epithelium morphology, but have some differences in their growth pattern (a doubling time of 17.9 vs 34.8 h, respectively) and the expression of epithelial bile duct markers, CK7 and CK19. Cytogenetic analysis of KKU-452 and KKU-023 cells revealed their highly complex karyotypes; hypertriploid and hypotetraploid, respectively, with multiple chromosomal aberrations. Both cell lines showed mutations in p53 but not in KRAS. KKU-452 showed a very rapid migration and invasion properties in concert with low expression of E-cadherin and high expression of N-cadherin, whereas KKU-023 showed opposite characters. KKU-023, but not KKU-452, showed in vivo tumorigenicity in xenografted nude mice. Those two established cholangiocarcinoma cell lines with unique characters may be valuable for better understanding the process of carcinogenesis and developing new therapeutics for the patients.


Assuntos
Neoplasias dos Ductos Biliares/patologia , Linhagem Celular Tumoral , Colangiocarcinoma/patologia , Animais , Neoplasias dos Ductos Biliares/genética , Separação Celular , Colangiocarcinoma/genética , Doenças Endêmicas , Fasciolíase , Feminino , Xenoenxertos , Humanos , Camundongos , Pessoa de Meia-Idade , Tailândia
3.
Curr Opin Gastroenterol ; 31(3): 258-63, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25693006

RESUMO

PURPOSE OF REVIEW: We review the genetic, epigenetic and transcriptional landscape of liver fluke (Opisthorchis viverrini, Ov)-related cholangiocarcinoma (CCA). Its distinct alterations, as compared with non-Ov-related CCA may help shed light on its underlying molecular mechanisms. RECENT FINDINGS: Recent whole-exome and targeted sequencing not only confirmed frequent mutations in known CCA-related genes including TP53 (44%), KRAS (16.7%) and SMAD4 (16.7%), but also revealed mutations in novel CCA-related genes associated with chromatin remodeling [BAP1 (2.8%), ARID1A (17.6%), MLL3 (13%) and IDH1/2 (2.8%)], WNT signaling [RNF43 (9.3%) and PEG3 (5.6%)] and KRAS/G protein signaling [GNAS (9.3%) and ROBO2 (9.3%)]. Interestingly, there is a significant difference in the frequency of mutated genes between Ov-related CCA and non-Ov-related CCA, such as p53 and IDH1/2, reflecting the impact of cause on pathogenesis. Altered DNA methylation and transcriptional profiles associated with xenobiotic metabolism and pro-inflammatory responses were also found in Ov-related CCA. SUMMARY: Liver fluke-induced chronic inflammation plays a crucial role in cholangiocarcinogenesis, resulting in distinct signatures of genetic, epigenetic and transcriptional alterations. These alterations, when contrasted with non-Ov-related CCA, indicate a unique pathogenic process in Ov-related CCA and may have potential clinical implications on diagnostics, therapeutics and prevention.


Assuntos
Neoplasias dos Ductos Biliares/genética , Colangiocarcinoma/genética , Regulação Neoplásica da Expressão Gênica/genética , Opisthorchis/isolamento & purificação , Receptores Imunológicos/genética , Animais , Neoplasias dos Ductos Biliares/parasitologia , Neoplasias dos Ductos Biliares/patologia , Colangiocarcinoma/parasitologia , Colangiocarcinoma/patologia , Proteínas de Ligação a DNA , Exoma , Variação Genética , Humanos , Análise de Sequência de DNA , Ativação Transcricional/genética
4.
Heliyon ; 10(11): e32352, 2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38961933

RESUMO

Previous studies have indicated the efficacy of momordin Ic (MIc), a plant-derived triterpenoid, against several types of cancers, implying its potential for further development. However, comprehensive insights into the molecular mechanisms and targets of MIc in cholangiocarcinoma (CCA) are lacking. This study aimed to investigate the actions of MIc against CCA at the molecular level. Network pharmacology analysis was first employed to predict the mechanisms and targets of MIc. The results unveiled the potential involvement of MIc in apoptosis and cell migration, pinpointing Src and FAK as key targets. Subsequently, cell-based assays, in accordance with FAK/Src-associated metastasis, were conducted, demonstrating the ability of MIc to attenuate the metastatic behaviours of KKU-452 cells. The in vitro results further indicated the capability of MIc to suppress the epithelial-mesenchymal transition (EMT) process, notably by downregulating EMT regulators, including N-cadherin, vimentin, ZEB2 and FOXC1/2 expression. Furthermore, MIc suppressed the activation of the FAK/Src signalling pathway, influencing critical downstream factors such as MMP-9, VEGF, ICAM-1, and c-Myc. Molecular docking simulations also suggested that MIc could interact with FAK and Src domains and restrain kinases from being activated by hindering ATP binding. In conclusion, this study employs a comprehensive approach encompassing network pharmacology analysis, in vitro assays, and molecular docking to unveil the mechanisms and targets of MIc in CCA. MIc mitigates metastatic behaviours and suppresses key pathways, offering a promising avenue for future therapeutic strategies against this aggressive cancer.

5.
Heliyon ; 10(10): e31112, 2024 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-38799762

RESUMO

Purpose: To assess the impact of concurrent inhibition of the FGFR and PI3K/mTOR signaling pathways on oncogenic characteristics in cholangiocarcinoma (CCA) cells, including proliferation, autophagy, and cell death. Materials and methods: KKU-213A, KKU-100, and KKU-213C cells were treated with either infigratinib or PKI-402 alone or in combination. Cell viability and cell death were evaluated using the sulforhodamine B (SRB) assay and acridine orange/ethidium bromide (AO/EB) staining. Cell cycle progression and apoptotic cell death were analyzed by flow cytometry. Western blotting was performed to assess the expression of proteins involved in cell cycle regulation and autophagy. Additionally, AO staining was employed to assess autophagic induction. Results: The combination of infigratinib and PKI-402 showed a remarked synergistic suppression in cell viability in both CCA cell lines compared to treatment with single inhibitors. This antiproliferative effect was associated with cell cycle arrest in the G2-M phase and a decrease in the expression of cyclin A and cyclin B1 in CCA cells. Furthermore, the combination treatment induced apoptotic cell death to a greater extent than treatment with a single inhibitor. Infigratinib enhanced the induction of autophagy by PKI-402, as evidenced by marked increases of autophagic vacuoles stained acridine orange, levels of LC3B-II and suppression of levels of p-mTOR and. Notably, inhibition of autophagic flux by chloroquine prevented cell death induced by the combination treatment. Conclusions: This study demonstrated that concurrent inhibition of the key FGFR/PI3K/mTOR pathways in CCA carcinogenesis enhances the suppression of CCA cells. The present findings indicate potential clinical implications for using combination treatment modalities in CCA therapy.

6.
BMC Complement Med Ther ; 24(1): 178, 2024 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-38689275

RESUMO

BACKGROUND: Gastrointestinal cancer (GIC) ranks as the highest cause of cancer-related deaths globally. GIC patients are often diagnosed at advanced stages, limiting effective treatment options. Chemotherapy, the common GIC recommendation, has significant disadvantages such as toxicity and adverse effects. Natural products contain substances with diverse pharmacological characteristics that promise for use in cancer therapeutics. In this study, the flower of renowned Asian medicinal plant, Shorea roxburghii was collected and extracted to investigate its phytochemical contents, antioxidant, and anticancer properties on GIC cells. METHODS: The phytochemical contents of Shorea roxburghii extract were assessed using suitable methods. Phenolic content was determined through the Folin-Ciocalteu method, while flavonoids were quantified using the aluminum chloride (AlCl3) method. Antioxidant activity was evaluated using the FRAP and DPPH assays. Cytotoxicity was assessed in GIC cell lines via the MTT assay. Additionally, intracellular ROS levels and apoptosis were examined through flow cytometry techniques. The correlation between GIC cell viability and phytochemicals, 1H-NMR analysis was conducted. RESULTS: Among the four different solvent extracts, ethyl acetate extract had the highest phenolic and flavonoid contents. Water extract exhibited the strongest reducing power and DPPH scavenging activity following by ethyl acetate. Interestingly, ethyl acetate extract demonstrated the highest inhibitory activity against three GIC cell lines (KKU-213B, HepG2, AGS) with IC50 values of 91.60 µg/ml, 39.38 µg/ml, and 35.59 µg/ml, while showing less toxicity to normal fibroblast cells. Ethyl acetate extract induced reactive oxygen species and apoptosis in GIC cell lines by downregulating anti-apoptotic protein Bcl-2. Metabolic profiling-based screening revealed a positive association between reduced GIC cell viability and phytochemicals like cinnamic acid and its derivatives, ferulic acid and coumaric acid. CONCLUSIONS: This study highlights the potential of natural compounds in Shorea roxburghii in the development of more effective and safer anticancer agents as options for GIC as well as shedding light on new avenues for cancer treatment.


Assuntos
Neoplasias Gastrointestinais , Extratos Vegetais , Humanos , Extratos Vegetais/farmacologia , Extratos Vegetais/química , Linhagem Celular Tumoral , Neoplasias Gastrointestinais/tratamento farmacológico , Apoptose/efeitos dos fármacos , Antioxidantes/farmacologia , Antineoplásicos Fitogênicos/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Metabolômica , Compostos Fitoquímicos/farmacologia , Flavonoides/farmacologia , Flavonoides/análise
7.
Cancers (Basel) ; 16(4)2024 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-38398194

RESUMO

Cholangiocarcinomas (CCA) pose a complex challenge in oncology due to diverse etiologies, necessitating tailored therapeutic approaches. This review discusses the risk factors, molecular pathology, and current therapeutic options for CCA and explores the emerging strategies encompassing targeted therapies, immunotherapy, novel compounds from natural sources, and modulation of gut microbiota. CCA are driven by an intricate landscape of genetic mutations, epigenetic dysregulation, and post-transcriptional modification, which differs based on geography (e.g., for liver fluke versus non-liver fluke-driven CCA) and exposure to environmental carcinogens (e.g., exposure to aristolochic acid). Liquid biopsy, including circulating cell-free DNA, is a potential diagnostic tool for CCA, which warrants further investigations. Currently, surgical resection is the primary curative treatment for CCA despite the technical challenges. Adjuvant chemotherapy, including cisplatin and gemcitabine, is standard for advanced, unresectable, or recurrent CCA. Second-line therapy options, such as FOLFOX (oxaliplatin and 5-FU), and the significance of radiation therapy in adjuvant, neoadjuvant, and palliative settings are also discussed. This review underscores the need for personalized therapies and demonstrates the shift towards precision medicine in CCA treatment. The development of targeted therapies, including FDA-approved drugs inhibiting FGFR2 gene fusions and IDH1 mutations, is of major research focus. Investigations into immune checkpoint inhibitors have also revealed potential clinical benefits, although improvements in survival remain elusive, especially across patient demographics. Novel compounds from natural sources exhibit anti-CCA activity, while microbiota dysbiosis emerges as a potential contributor to CCA progression, necessitating further exploration of their direct impact and mechanisms through in-depth research and clinical studies. In the future, extensive translational research efforts are imperative to bridge existing gaps and optimize therapeutic strategies to improve therapeutic outcomes for this complex malignancy.

8.
Int J Mol Med ; 51(3)2023 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-36660943

RESUMO

13­cis­retinoic acid (13CRA), a Food and Drug Administration­approved drug for severe acne, is currently being investigated for its potential use in skin cancer prevention. 13CRA has been reported to exhibit antitumor effects against various types of cancer cells, both in vitro and in vivo. However, to the best of our knowledge, no information is yet available regarding the effects of 13CRA on cholangiocarcinoma (CCA), a malignancy of the bile duct epithelia. Currently, there are no reliably effective therapeutic options for metastatic CCA. The present study thus aimed to evaluate the effects of 13CRA on the self­renewal, migration, invasion and adhesion of CCA cells, and also investigated the underlying mechanisms. The results revealed that 13CRA suppressed cell proliferation via the inhibition of the self­renewal ability of CCA cells. 13CRA induced cell cycle arrest at the G2/M phase in KKU­100 and KKU­213B CCA cells through the regulation of cell cycle­regulatory genes and proteins. 13CRA reduced the cell migratory ability of both cell lines via the modulation of the genes and proteins associated with epithelial­mesenchymal transition. 13CRA also inhibited the invasive and adhesive abilities of CCA cells via the suppression of genes and proteins associated with the invasion and adhesion of CCA cells. On the whole, these results suggested that 13CRA exerts suppressive effects on CCA cell proliferation, migration, adhesion and invasion.


Assuntos
Neoplasias dos Ductos Biliares , Colangiocarcinoma , Humanos , Isotretinoína/farmacologia , Isotretinoína/uso terapêutico , Linhagem Celular Tumoral , Colangiocarcinoma/genética , Proliferação de Células , Movimento Celular/genética , Ductos Biliares Intra-Hepáticos/metabolismo , Ductos Biliares Intra-Hepáticos/patologia , Neoplasias dos Ductos Biliares/patologia
9.
In Vivo ; 37(4): 1628-1637, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37369494

RESUMO

BACKGROUND/AIM: Increasing evidence has revealed FGFR2 as an attractive therapeutic target for cancer including cholangiocarcinoma (CCA). The present study investigated the oncogenic mechanisms by which FGF10 ligand activates FGFR2 in CCA cells and determined whether FGFR inhibitors could suppress FGF10-mediated migration of CCA cells. MATERIALS AND METHODS: Effects of FGF10 on the proliferation, migration, and invasion of KKU-M213A cells were assessed using clonogenic and transwell assays. Protein expression levels of FGFR2 and pro-angiogenic factors were determined via immunoblotting and antibody array analysis. FGFR2 knockdown using a small interfering RNA was used to validate the role of FGF10 in promoting cell migration via FGFR2. The effects of infigratinib (FGFR inhibitor) on cell viability, were determined in KKU-100, KKU-M213A, KKU-452 cells. Moreover, the efficacy of the FGFR inhibitor in suppressing migration via FGF10/FGFR2 stimulation was assessed in KKU-M213A cells. RESULTS: FGF10 significantly increased the expression of phospho-FGFR/FGFR2 and promoted the proliferation, migration, and invasion of KKU-M213A cells. FGF10 increased the expression levels of p-Akt, p-mTOR, VEGF, Slug, and pro-angiogenic proteins related to metastasis. Cell migration mediated by FGF10 was markedly decreased in FGFR2-knockdown cells. Moreover, FGF10/FGFR2 promoted the migration of cells, which was suppressed by the FGFR inhibitor. CONCLUSION: FGF10/FGFR2 activates the Akt/mTOR and VEGF/Slug pathways, which are associated with the stimulation of migration and invasion in CCA. Moreover, the FGF10/FGFR2 signaling was inhibited by an FGFR inhibitor resulting suppression of cell migration, which warrants further studies on their clinical utility for CCA treatment.


Assuntos
Neoplasias dos Ductos Biliares , Colangiocarcinoma , Humanos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ligantes , Fator A de Crescimento do Endotélio Vascular , Linhagem Celular Tumoral , Colangiocarcinoma/tratamento farmacológico , Colangiocarcinoma/genética , Serina-Treonina Quinases TOR , Inibidores de Proteínas Quinases/uso terapêutico , Ductos Biliares Intra-Hepáticos/metabolismo , Neoplasias dos Ductos Biliares/tratamento farmacológico , Neoplasias dos Ductos Biliares/genética , Proliferação de Células , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/uso terapêutico , Fator 10 de Crescimento de Fibroblastos/farmacologia , Fator 10 de Crescimento de Fibroblastos/uso terapêutico
10.
In Vivo ; 37(4): 1638-1648, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37369462

RESUMO

BACKGROUND/AIM: Serine/threonine kinase 11 (STK11), a tumor suppressor, controls 5' AMP-activated protein kinase (AMPK) signaling in a variety of cellular functions. Mutated STK11 has been identified as a novel driver gene that promotes cancer progression. The purpose of this study was to investigate the alteration of STK11 and its correlation with clinicopathological data in cholangiocarcinoma (CCA). MATERIALS AND METHODS: Gene mutation and expression analyses were performed using cBioportal and Gene Expression Profiling Interactive Analysis version 2 (GEPIA2). qRT-PCR was performed to measure STK11 mRNA levels and immunohistochemistry was performed to investigate STK11 protein expression in CCA tissues. RESULTS: The results from publicly available cancer datasets showed that 2.7% of CCA cases had STK11 mutations. Most of STK11 gene mutations are of the truncating type and result in low STK11 mRNA and protein expression. We detected a correlation between STK11 mutation status and the tendency for shorter patient survival. The results of qRT-PCR revealed that STK11 mRNA levels were statistically significantly lower in CCA patients with mutated STK11 compared to those with wild-type STK11 (p-value=0.013). Immunohistochemical staining showed high STK11 expression in 43.8% and low expression in 56.2% of CCA tissues examined. Low STK11 protein expression resulted in poor prognosis compared with high STK11 expression, especially in CCA papillary carcinoma. Univariate and multivariate analysis revealed that high STK11 expression was associated with a decreased hazard ratio of patient survival rates (HR=0.696, p-value=0.06 and HR=0.666, p-value=0.04, respectively). CONCLUSION: Alteration of STK11 mutational or mRNA/protein status might be used as a potential predictive biomarker for the prognosis of the clinical outcomes in CCA patients.


Assuntos
Neoplasias dos Ductos Biliares , Colangiocarcinoma , Humanos , Colangiocarcinoma/genética , Colangiocarcinoma/patologia , Prognóstico , Ductos Biliares Intra-Hepáticos/metabolismo , Neoplasias dos Ductos Biliares/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Quinases Proteína-Quinases Ativadas por AMP
11.
J Pharm Pharmacol ; 74(7): 996-1005, 2022 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-35640567

RESUMO

OBJECTIVES: Strategies that induce apoptosis of malignant cells are recognized as effective cancer treatments. This study evaluated the apoptosis-inducing ability of momordin Ic against cholangiocarcinoma (CCA) cells and the respective underlying mechanisms. METHODS: Quantification of apoptotic cells was performed using Annexin V/7-AAD double dye staining followed by flow cytometry. The effect of momordin Ic on the expression of epidermal growth factor receptor (EGFR) and its downstream signalling molecules was determined via Western blot analysis. The RT2 Profiler PCR Array was used to determine the expression of cell death-associated genes. Expression levels of apoptosis-related proteins were examined using an apoptosis antibody array. KEY FINDINGS: Momordin Ic potently limited the ability of CCA cells to thrive by promoting apoptotic cell death. This apoptosis-inducing activity was accompanied with suppression of expression of EGFR, p-EGFR, c-Myc and other downstream EGFR signalling-related molecules. Additional molecular analyses demonstrated that momordin Ic modified the expression profile of cell death-associated genes in CCA cells. Moreover, significant upregulation of apoptosis-activating proteins and downregulation of apoptosis-inhibiting protein were also observed after exposure to momordin Ic. CONCLUSIONS: These results suggest that momordin Ic has a potential therapeutic opportunity for CCA treatment by acting as an EGFR suppressant.


Assuntos
Neoplasias dos Ductos Biliares , Colangiocarcinoma , Apoptose , Neoplasias dos Ductos Biliares/tratamento farmacológico , Neoplasias dos Ductos Biliares/patologia , Ductos Biliares Intra-Hepáticos/patologia , Linhagem Celular Tumoral , Proliferação de Células , Colangiocarcinoma/tratamento farmacológico , Colangiocarcinoma/patologia , Receptores ErbB , Humanos , Extratos Vegetais
12.
Asian Pac J Cancer Prev ; 23(1): 115-123, 2022 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-35092379

RESUMO

OBJECTIVE: To investigate the anti-tumor effect of licochalcone A (LCA) on proliferation and migration in cholangiocarcinoma (CCA) cells and to elucidate their underlying mechanisms. METHODS: Human CCA cells, KKU-100, KKU-213, KKU-214, KKU-156, and KKU-452 were used to study effect of LCA on proliferation and migration by a cytotoxicity assay, wound healing assay. Reactive oxygen species levels were evaluated using DHE-fluorescent probes. Proteins associated with cancer survival and progression were analyzed by immune blotting assay. RESULTS: LCA suppressed proliferation and induced cell death in CCA cells including KKU-100, KKU-213, KKU-214, KKU-156, and KKU-452. The CCAs cells were suppressed in association with LCA-induced accumulation of intracellular reactive oxygen species (ROS). Increased formation of ROS was causally related with suppression of Nrf2 and its down-stream antioxidant and cytoprotective enzymes. These effects may lead to the expression of Bax and release of cytochrome c and ensuring cell death.  Interestingly, LCA could also inhibit cell migration and cell cycle arrest at low concentrations. These effects were associated with down-regulation of NF-kB, STAT3 and their down-stream proteins, cyclin D1, VEGF, and ICAM-1. CONCLUSIONS: These results suggest that LCA has potential therapeutic activity in suppression of CCA cells.


Assuntos
Antineoplásicos/farmacologia , Neoplasias dos Ductos Biliares/tratamento farmacológico , Chalconas/farmacologia , Colangiocarcinoma/tratamento farmacológico , Fator 2 Relacionado a NF-E2/efeitos dos fármacos , NF-kappa B/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Humanos , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos
13.
Oncol Lett ; 23(6): 179, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35464301

RESUMO

All-trans-retinoic acid (ATRA) has been clinically used to treat acute promyelocytic leukemia and is being studied to treat other types of cancer; however, the therapeutic role and mechanism of ATRA against cholangiocarcinoma (CCA) remain unclear. The present study investigated the cytotoxic effect and underlying mechanisms of ATRA on CCA cell lines. Cell viability was evaluated by sulforhodamine B assay. Intracellular reactive oxygen species (ROS) levels were assessed by dihydroethidium assay. Apoptosis analysis was performed by flow cytometry. The pathways of apoptotic cell death induction were examined using enzymatic caspase activity assay. Proteins associated with apoptosis were evaluated by western blotting. The effects on gene expression were analyzed by reverse transcription-quantitative PCR analysis. ATRA induced a concentration- and time-dependent toxicity in CCA cells. Furthermore, when the cytotoxicity of ATRA against retinoic acid receptor (RAR)-deficient cells was assessed, it was revealed that ATRA cytotoxicity was RARB-dependent. Following ATRA treatment, there was a significant accumulation of cellular ROS and ATRA-induced ROS generation led to an increase in the expression levels of apoptosis-inducing proteins and intrinsic apoptosis. Pre-treatment with ROS scavengers could diminish the apoptotic effect of ATRA, suggesting that ROS and mitochondria may have an essential role in the induction of apoptosis. Furthermore, following ATRA treatment, an increase in cellular ROS content was associated with suppressing nuclear factor erythroid 2-related factor 2 (NFE2L2 or NRF2) and NRF2-downstream active genes. ATRA also suppressed cisplatin-induced NRF2 expression, suggesting that the enhancement of cisplatin cytotoxicity by ATRA may be associated with the downregulation of NRF2 signaling. In conclusion, the results of the present study demonstrated that ATRA could be repurposed as an alternative drug for CCA therapy.

14.
Integr Cancer Ther ; 21: 15347354221124861, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36154723

RESUMO

Owing to the crucial role of Src in cancer metastasis, interruption of Src and its signaling has been considered a promising strategy for cancer metastasis treatment. Cucurbitacin B, a dietary triterpenoid, has been shown to possess anti-proliferative and apoptosis-inducing activities in cholangiocarcinoma (CCA) cells via suppressing the activation of FAK which is a main downstream Src effector. We hypothesized that cucurbitacin B might act as a Src suppressant which conferring anti-metastasis effect against CCA cells. To investigate this, the role of Src in regulating metastasis behavior of CCA cells and the effect of cucurbitacin B on Src-mediated metastatic phenotype of these cells were determined. The results showed that activation of Src significantly enhanced the migratory and invasive abilities of CCA cells. Molecular analysis revealed that Src-facilitated metastasis behavior of CCA cells occurred by modifying expression of a wide range of metastasis-related genes in the cells. Consistent with gene expression results, activation of Src significantly induced the protein expression of 2 important metastasis-associated molecules, MMP-9 and VEGF. Cucurbitacin B markedly suppressed activation of Src and its key effector, FAK. As a consequence, the alteration of expression profiles of metastasis-associated genes induced by Src activator in CCA cells was diminished by cucurbitacin B treatment. The compound also down-regulated Src-induced expression of MMP-9 and VEGF proteins in the cells. Moreover, molecular docking analysis revealed that cucurbitacin B could interact with Src kinase domain and possibly restrain the kinase from being activated by hindering the ATP binding. In conclusion, cucurbitacin B exhibited anti-metastatic property in CCA cells via negatively influencing Src and Src-related oncogenic signaling. This compound may therefore be a potential therapeutic drug for further development as an anti-Src agent for treatment of metastatic CCA.


Assuntos
Neoplasias dos Ductos Biliares , Colangiocarcinoma , Triterpenos , Trifosfato de Adenosina/farmacologia , Trifosfato de Adenosina/uso terapêutico , Neoplasias dos Ductos Biliares/tratamento farmacológico , Neoplasias dos Ductos Biliares/patologia , Ductos Biliares Intra-Hepáticos/metabolismo , Ductos Biliares Intra-Hepáticos/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Colangiocarcinoma/tratamento farmacológico , Colangiocarcinoma/patologia , Humanos , Metaloproteinase 9 da Matriz , Simulação de Acoplamento Molecular , Triterpenos/farmacologia , Triterpenos/uso terapêutico , Fator A de Crescimento do Endotélio Vascular/metabolismo , Quinases da Família src/farmacologia , Quinases da Família src/uso terapêutico
15.
Life Sci ; 296: 120427, 2022 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-35218764

RESUMO

AIM: To investigate the oncogenic role of FGFR2 in carcinogenesis in cholangiocarcinoma (CCA) cells. In addition, the feasibility of using FGFR inhibitors in combination with standard chemotherapy was also explored for the chemosensitizing effect in CCA cells. MAIN METHODS: Five CCA cell lines were used to screen FGFR2 expression by Western immunoblotting. Two CCA cell lines, KKU-100 and KKU-213A, were knocked down of the FGFR2 gene using siRNA. Cell viability was assessed by the MTS cell proliferation assay. Reproductive cell death was assessed by clonogenic assay. The effects on cell migration and invasion were analyzed by the Transwell chamber method. Cell cycle analysis was performed by flow cytometry. Cell angiogenesis was assessed by HUVEC tube formation and human angiogenesis antibody array analysis. Proteins associated with proliferative and metastatic properties were evaluated by Western blotting. KEY FINDINGS: Knockdown of FGFR2 suppressed cell growth and colony formation in CCA cells in association with G2/M cell cycle arrest and downregulation of STAT3, cyclin A and cyclin B1. Silencing FGFR2 enhanced the suppressive effect of gemcitabine (Gem) on cell migration and invasion. The combination of infigratinib, an FGFR inhibitor, and Gem, interrupted cell growth, migration, and invasion via downregulation of FGFR/AKT/mTOR pathways and the EMT-associated proteins vimentin and slug. Moreover, the combination also suppressed tube formation together with decreased expression of the proangiogenic factor VEGF. SIGNIFICANCE: Inhibition of FGFRs by infigratinib enhanced the antitumor effect of Gem in CCA cells through downregulation of the FGFR/AKT/mTOR, FGFR/STAT3 and EMT signaling pathways.


Assuntos
Neoplasias dos Ductos Biliares/tratamento farmacológico , Colangiocarcinoma/tratamento farmacológico , Desoxicitidina/análogos & derivados , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo , Antimetabólitos Antineoplásicos/farmacologia , Neoplasias dos Ductos Biliares/metabolismo , Neoplasias dos Ductos Biliares/patologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Colangiocarcinoma/metabolismo , Colangiocarcinoma/patologia , Desoxicitidina/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/fisiologia , Humanos , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/patologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/genética , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo , Gencitabina
16.
PeerJ ; 10: e12750, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35070505

RESUMO

BACKGROUND: Genetic alterations in ARID1A were detected at a high frequency in cholangiocarcinoma (CCA). Growing evidence indicates that the loss of ARID1A expression leads to activation of the PI3K/AKT pathway and increasing sensitivity of ARID1A-deficient cells for treatment with the PI3K/AKT inhibitor. Therefore, we investigated the association between genetic alterations of ARID1A and the PI3K/AKT pathway and evaluated the effect of AKT inhibition on ARID1A-deficient CCA cells. METHODS: Alterations of ARID1A, PI3K/AKT pathway-related genes, clinicopathological data and overall survival of 795 CCA patients were retrieved from cBio Cancer Genomics Portal (cBioPortal) databases. The association between genetic alterations and clinical data were analyzed. The effect of the AKT inhibitor (MK-2206) on ARID1A-deficient CCA cell lines and stable ARID1A-knockdown cell lines was investigated. Cell viability, apoptosis, and expression of AKT signaling were analyzed using an MTT assay, flow cytometry, and Western blots, respectively. RESULTS: The analysis of a total of 795 CCA samples revealed that ARID1A alterations significantly co-occurred with mutations of EPHA2 (p < 0.001), PIK3CA (p = 0.047), and LAMA1 (p = 0.024). Among the EPHA2 mutant CCA tumors, 82% of EPHA2 mutant tumors co-occurred with ARID1A truncating mutations. CCA tumors with ARID1A and EPHA2 mutations correlated with better survival compared to tumors with ARID1A mutations alone. We detected that 30% of patients with PIK3CA driver missense mutations harbored ARID1A-truncated mutations and 60% of LAMA1-mutated CCA co-occurred with truncating mutations of ARID1A. Interestingly, ARID1A-deficient CCA cell lines and ARID1A-knockdown CCA cells led to increased sensitivity to treatment with MK-2206 compared to the control. Treatment with MK-2206 induced apoptosis in ARID1A-knockdown KKU-213A and HUCCT1 cell lines and decreased the expression of pAKTS473 and mTOR. CONCLUSION: These findings suggest a dependency of ARID1A-deficient CCA tumors with the activation of the PI3K/AKT-pathway, and that they may be more vulnerable to selective AKT pathway inhibitors which can be used therapeutically.


Assuntos
Neoplasias dos Ductos Biliares , Colangiocarcinoma , Humanos , Proteínas Proto-Oncogênicas c-akt , Fosfatidilinositol 3-Quinases/genética , Colangiocarcinoma/tratamento farmacológico , Inibidores de Proteínas Quinases/metabolismo , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase/farmacologia , Ductos Biliares Intra-Hepáticos/metabolismo , Neoplasias dos Ductos Biliares/tratamento farmacológico , Proteínas de Ligação a DNA/genética , Fatores de Transcrição/genética
17.
Cancer Treat Rev ; 95: 102170, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33735689

RESUMO

Cholangiocarcinomas (CCAs) are rare but aggressive tumours of the bile ducts, which are often diagnosed at an advanced stage and have poor outcomes on systemic therapy. Somatic alterations with therapeutic implications have been identified in almost half of CCAs, in particular in intrahepatic CCA (iCCA), the subtype arising from bile ducts within the liver. Among patients with CCA, fibroblast growth factor receptor 2 (FGFR2) fusions or rearrangements occur almost exclusively in iCCA, where they are estimated to be found in up to 10-15% of patients. Clinical trials for selective FGFR kinase inhibitors have shown consistent activity of these agents in previously treated patients with iCCA harbouring FGFR alterations. Current FGFR kinase inhibitors show differences in their structure, mechanisms of target engagement, and specificities for FGFR1, 2, 3 and 4 and other related kinases. These agents offer the potential to improve outcomes in FGFR-driven CCA, and the impact of variations in the molecular profiles of the FGFR inhibitors on efficacy, safety, acquired resistance mechanisms, and patients' health-related quality of life remains to be fully characterized. The most common adverse event associated with FGFR inhibitors is hyperphosphatemia, an on-target off-tumour effect of FGFR1 inhibition, and strategies to manage this include dose adjustment, chelators, and the use of a low phosphate diet. As FGFR inhibitors and other targeted agents enter the clinic for use in FGFR-driven CCA, molecular testing for actionable mutations and monitoring for the emergence of acquired resistance will be essential.


Assuntos
Antineoplásicos/uso terapêutico , Neoplasias dos Ductos Biliares/tratamento farmacológico , Colangiocarcinoma/tratamento farmacológico , Terapia de Alvo Molecular , Inibidores de Proteínas Quinases/uso terapêutico , Receptores de Fatores de Crescimento de Fibroblastos/antagonistas & inibidores , Animais , Neoplasias dos Ductos Biliares/metabolismo , Neoplasias dos Ductos Biliares/patologia , Colangiocarcinoma/metabolismo , Colangiocarcinoma/patologia , Humanos
18.
Asian Pac J Cancer Prev ; 22(1): 219-225, 2021 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-33507702

RESUMO

OBJECTIVE: Cholangiocarcinoma (CCA) is a malignant tumor with aggressive metastatic property resulted from dysregulation of metastasis-regulated signaling pathways. The aim of this study was to investigate the effect of cucurbitacin B on metastatic behavior of CCA cells through modulation of focal adhesion kinase (FAK) protein. METHODS: KKU-452 cells were treated with a specific FAK inhibitor, FAK inhibitor-14, or cucurbitacin B at various concentrations for 24 h. Cell viability was assessed by sulforhodamine B assay. The migratory and invasive abilities of the cells were investigated using wound healing and transwell invasion assays, respectively. The fibronectin-coated plate was used for adhesion assay. The effects of the test compounds on FAK activation and the expression of metastasis-associated proteins were determined by Western blot analysis. The amount of MMP-9 was evaluated using a commercial ELISA Kit. RESULTS: FAK inhibitor-14 and cucurbitacin B at concentrations which minimally affected KKU-452 cell viability could suppress FAK activation, evidently by decreased level of phospho-FAK protein after exposure to the compound. At these conditions, cucurbitacin B suppressed metastatic behavior including migration, invasion and adhesion abilities of CCA cells similar to FAK inhibitor-14. Further molecular studies demonstrated that FAK inhibitor-14 and cucurbitacin B downregulated the expression of metastasis-associated proteins including MMP-9, ICAM-1 and VEGF. Consequently, exposure to cucurbitacin B inhibited the production of MMP-9 enzyme in CCA cells similar to FAK inhibitor-14 treatment. CONCLUSION: FAK participated in regulation of metastatic behavior of KKU-452 CCA cells. Cucurbitacin B suppressed FAK activation in the cells which was associated with inhibition of metastasis essential steps and their related metastatic proteins. The compound may be developed as a novel therapeutic agent for CCA metastasis therapy.
.


Assuntos
Neoplasias dos Ductos Biliares/tratamento farmacológico , Movimento Celular , Colangiocarcinoma/tratamento farmacológico , Quinase 1 de Adesão Focal/antagonistas & inibidores , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Triterpenos/farmacologia , Apoptose , Neoplasias dos Ductos Biliares/metabolismo , Neoplasias dos Ductos Biliares/secundário , Proliferação de Células , Colangiocarcinoma/metabolismo , Colangiocarcinoma/patologia , Humanos , Células Tumorais Cultivadas
19.
Naunyn Schmiedebergs Arch Pharmacol ; 394(9): 1929-1940, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34086099

RESUMO

Chemotherapy is a palliative treatment for unresectable patients with cholangiocarcinoma (CCA). However, drug resistance is a major cause of the failure of this treatment. Derrischalcone (DC), a novel chalcone isolated from Derris indica fruit, has been shown pharmacologically active; though, the effect of DC on CCA is unknown. The present study investigated the cytotoxic, antiproliferative, anti-migration, and anti-invasion effects and underlying mechanisms of DC on CCA KKU-M156 and KKU-100 cells. Cytotoxicity and apoptosis were evaluated by acridine orange and ethidium bromide fluorescent staining. Reactive oxygen species (ROS) was measured by dihydroethidium assay. Cell proliferation and reproductive cell death were assessed by sulforhodamine B staining and colony-forming assay. Migration and invasion were determined by wound healing and transwell chamber assays. Protein expressions associated with cell death, proliferation, migration, and invasion were analyzed by western immunoblotting. We found that DC induced cytotoxicity and apoptosis in association with ROS formation and oxidative stress. Treatment with N-acetylcysteine suppressed ROS formation and attenuated DC-induced cytotoxic and apoptotic effects. DC increased the expression of p53, p21, Bax, and cytochrome c proteins in association with cell death. DC-induced antiproliferation, colony formation, anti-migration, and anti-invasion were associated with the suppression of Akt/mTOR/cyclin D1 and FAK signaling pathways. These findings suggest that the multi-targeting strategies with DC may be a novel treatment for cancer therapy.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Neoplasias dos Ductos Biliares/tratamento farmacológico , Chalconas/farmacologia , Colangiocarcinoma/tratamento farmacológico , Apoptose/efeitos dos fármacos , Neoplasias dos Ductos Biliares/patologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Colangiocarcinoma/patologia , Quinase 1 de Adesão Focal/metabolismo , Humanos , Millettia/química , Mitocôndrias/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo
20.
Naunyn Schmiedebergs Arch Pharmacol ; 394(10): 2049-2059, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34283274

RESUMO

Northeast Thailand has the highest incidence of cholangiocarcinoma (CCA) in the world. The lack of promising diagnostic markers and appropriate therapeutic drugs is the main problem for metastatic stage CCA patients who have a poor prognosis. N-cadherin, a cell adhesion molecule, is usually upregulated in cancers and has been proposed as an important mediator in epithelial-mesenchymal transition (EMT), one of the metastasis processes. Additionally, it has been shown that arctigenin, a seed isolated compound from Arctium lappa, can inhibit cancer cell progression via suppression of N-cadherin pathway. In this study, we investigated the protein expression of N-cadherin and its correlation with clinicopathological data of CCA patients, as well as the impact of arctigenin on KKU-213A and KKU-100 CCA cell lines and its underlying mechanisms. Immunohistochemistry results demonstrated that high expression of N-cadherin was significantly associated with severe CCA stage (p = 0.027), and shorter survival time (p = 0.002) of CCA patients. The mean overall survival times between low and high expression of N-cadherin were 31.6 and 14.8 months, respectively. Wound healing assays showed that arctigenin significantly inhibited CCA cell migration by downregulating N-cadherin whereas upregulating E-cadherin expression. Immunocytochemical staining revealed that arctigenin suppressed the expression of N-cadherin in both CCA cell lines. Furthermore, flow cytometry and western blot analysis revealed that arctigenin significantly reduced CCA cell viability and induced apoptosis via the Bax/Bcl-2/caspase-3 pathway. This research supports the use of N-cadherin as a prognostic marker for CCA and arctigenin as a potential alternative therapy for improving CCA treatment outcomes.


Assuntos
Antígenos CD/metabolismo , Antineoplásicos Fitogênicos/uso terapêutico , Neoplasias dos Ductos Biliares/tratamento farmacológico , Biomarcadores Tumorais/metabolismo , Caderinas/metabolismo , Colangiocarcinoma/tratamento farmacológico , Furanos/uso terapêutico , Lignanas/uso terapêutico , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias dos Ductos Biliares/metabolismo , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Colangiocarcinoma/metabolismo , Progressão da Doença , Feminino , Furanos/farmacologia , Humanos , Lignanas/farmacologia , Masculino , Pessoa de Meia-Idade , Prognóstico
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