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With recent advances in adeno-associated virus (AAV)-based gene therapy, efficacy and toxicity screening have become essential for developing gene therapeutic drugs for retinal diseases. Retinal organoids from human pluripotent stem cells (hPSCs) offer a more accessible and reproducible human test platform for evaluating AAV-based gene therapy. In this study, hPSCs were differentiated into retinal organoids composed of various types of retinal cells. The transduction efficiencies of AAV2 and AAV8, which are widely used in clinical trials of inherited retinal diseases, were analyzed using retinal organoids. These results suggest that retinal organoids derived from hPSCs serve as suitable screening platforms owing to their diverse retinal cell types and similarity to the human retina. In summary, we propose an optimal stepwise protocol that includes the generation of retinal organoids and analysis of AAV transduction efficacy, providing a comprehensive approach for evaluating AAV-based gene therapy for retinal diseases.
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Serpin family E member 1 (SERPINE1), a serine proteinase inhibitor, serves as an important regulator of extracellular matrix remodeling. Emerging evidence suggests that SERPINE1 has diverse roles in cancer and is associated with poor prognosis. However, the mechanism via which SERPINE1 is induced in cancer has not been fully determined. In order to examine the molecular mechanism of SERPINE1 expression, the present study took advantage of the isogenic pair of lung cancer cells with epithelial or mesenchymal features. Using genetic perturbation and following biochemical analysis, the present study demonstrated that SERPINE1 expression was upregulated in mesenchymal lung cancer cells and promoted cellular invasiveness. Yesassociated protein (YAP)dependent SERPINE1 expression was modulated by treatment with a Rhoassociated protein kinase inhibitor, Y27632. Moreover, TGFß treatment supported YAPdependent SERPINE1 expression, and an enhanced TGFß response in mesenchymal lung cancer cells promoted SERPINE1 expression. TGFßmediated SERPINE1 expression was significantly attenuated by knockdown of YAP or transcriptional coactivator with PDZbinding motif, suggesting that crosstalk between the TGFß and YAP pathways underlies SERPINE1 expression in mesenchymal cancer cells.
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Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Neoplasias Pulmonares/genética , Inibidor 1 de Ativador de Plasminogênio/genética , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Células A549 , Proteínas Adaptadoras de Transdução de Sinal/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Retroalimentação Fisiológica , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Pulmão/patologia , Neoplasias Pulmonares/patologia , Células-Tronco Mesenquimais/patologia , Fatores de Transcrição/genética , Proteínas com Motivo de Ligação a PDZ com Coativador Transcricional , Fator de Crescimento Transformador beta/genética , Regulação para Cima , Proteínas de Sinalização YAPRESUMO
The acquisition of chemoresistance remains a major cause of cancer mortality due to the limited accessibility of targeted or immune therapies. However, given that severe alterations of molecular features during epithelial-to-mesenchymal transition (EMT) lead to acquired chemoresistance, emerging studies have focused on identifying targetable drivers associated with acquired chemoresistance. Particularly, AXL, a key receptor tyrosine kinase that confers resistance against targets and chemotherapeutics, is highly expressed in mesenchymal cancer cells. However, the underlying mechanism of AXL induction in mesenchymal cancer cells is poorly understood. Our study revealed that the YAP signature, which was highly enriched in mesenchymal-type lung cancer, was closely correlated to AXL expression in 181 lung cancer cell lines. Moreover, using isogenic lung cancer cell pairs, we also found that doxorubicin treatment induced YAP nuclear translocation in mesenchymal-type lung cancer cells to induce AXL expression. Additionally, the concurrent activation of TGFß signaling coordinated YAP-dependent AXL expression through SMAD4. These data suggest that crosstalk between YAP and the TGFß/SMAD axis upon treatment with chemotherapeutics might be a promising target to improve chemosensitivity in mesenchymal-type lung cancer.
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Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Neoplasias Pulmonares/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Células A549 , Proteínas Adaptadoras de Transdução de Sinal/genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Proteínas Proto-Oncogênicas/genética , Receptores Proteína Tirosina Quinases/genética , Fatores de Transcrição/genética , Fator de Crescimento Transformador beta/genética , Proteínas de Sinalização YAP , Receptor Tirosina Quinase AxlRESUMO
Erastin, a synthetic lethal compound against cancer expressing an oncogenic RAS, inhibits cystine/glutamate antiporters and causes ferroptosis. However, despite recent evidence for the mechanisms underlying ferroptosis, molecular biomarkers of erastin-dependent ferroptosis have not been identified. Here, we employed isogenic lung cancer cell models to show that a redox imbalance leads to glutathione depletion and ferroptosis. Subsequent transcriptome analysis of pan-cancer cell lines revealed that the activity of transcription factors, including NRF2 and AhR, serve as important markers of erastin resistance. Based on the integrated expression of genes in the nuclear receptor meta-pathway (NRM), we constructed an NRM model and validated its robustness using an independent pharmacogenomics dataset. The NRM model was further evaluated by sensitivity tests on nine cancer cell lines for which erastin sensitivities had not been determined. Our pharmacogenomics approach has the potential to pave the way for the efficient classification of patients for therapeutic intervention using erastin.
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Ferroptose , Glutationa , Humanos , Piperazinas , Receptores Citoplasmáticos e NuclearesRESUMO
An efficient gene-editing technique for use in human pluripotent stem cells (hPSCs) has great potential value in regenerative medicine, as well as in drug discovery based on isogenic human disease models. However, the extremely low efficiency of gene editing in hPSCs remains as a major technical hurdle. Previously, we demonstrated that YM155, a survivin inhibitor developed as an anti-cancer drug, induces highly selective cell death in undifferentiated hPSCs. In this study, we demonstrated that the high cytotoxicity of YM155 in hPSCs, which is mediated by selective cellular uptake of the drug, is due to the high expression of SLC35F2 in these cells. Knockout of SLC35F2 with CRISPR-Cas9, or depletion with siRNAs, made the hPSCs highly resistant to YM155. Simultaneous editing of a gene of interest and transient knockdown of SLC35F2 following YM155 treatment enabled the survival of genome-edited hPSCs as a result of temporary YM155 resistance, thereby achieving an enriched selection of clonal populations with gene knockout or knock-in. This precise and efficient genome editing approach took as little as 3 weeks and required no cell sorting or the introduction of additional genes, to be a more feasible approach for gene editing in hPSCs due to its simplicity.
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Células-Tronco Pluripotentes , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Resistência a Medicamentos/genética , Edição de Genes , Genoma Humano , HumanosRESUMO
Despite the continual discovery of promising new cancer targets, drug discovery is often hampered by the poor druggability of these targets. As such, repurposing FDA-approved drugs based on cancer signatures is a useful alternative to cancer precision medicine. Here, we adopted an in silico approach based on large-scale gene expression signatures to identify drug candidates for lung cancer metastasis. Our clinicogenomic analysis identified GALNT14 as a putative driver of lung cancer metastasis, leading to poor survival. To overcome the poor druggability of GALNT14 in the control of metastasis, we utilized the Connectivity Map and identified bortezomib (BTZ) as a potent metastatic inhibitor, bypassing the direct inhibition of the enzymatic activity of GALNT14. The antimetastatic effect of BTZ was verified both in vitro and in vivo. Notably, both BTZ treatment and GALNT14 knockdown attenuated TGFß-mediated gene expression and suppressed TGFß-dependent metastatic genes. These results demonstrate that our in silico approach is a viable strategy for the use of undruggable targets in cancer therapies and for revealing the underlying mechanisms of these targets.
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Antineoplásicos/farmacologia , Bortezomib/farmacologia , Transformação Celular Neoplásica/genética , Neoplasias Pulmonares/tratamento farmacológico , N-Acetilgalactosaminiltransferases/genética , Células A549 , Linhagem Celular Tumoral , Reposicionamento de Medicamentos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Pulmonares/patologia , Metástase Neoplásica/tratamento farmacológico , Medicina de Precisão/métodos , Interferência de RNA , RNA Interferente Pequeno/genéticaRESUMO
The role of Situin 1 (SIRT1) in tumorigenesis is still controversial due to its wide range of substrates, including both oncoproteins and tumor suppressors. A recent study has demonstrated that SIRT1 interferes in the Kirsten rat sarcoma viral oncogene homolog (KRAS)-driven activation of the Raf-mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) kinase (MEK)-ERK pathway, thereby inhibiting tumorigenesis. However, the molecular mechanism of SIRT1 as a tumor suppressor in RAS-driven tumorigenesis has been less clearly determined. This study presents evidence that the ectopic expression of SIRT1 attenuates RAS- or MEK-driven ERK activation and reduces cellular proliferation and transformation in vitro. The attenuation of ERK activation by SIRT1 results from prompt dephosphorylation of ERK, while MEK activity remains unchanged. We identified that MKP1, a dual specific phosphatase for MAPK, was deacetylated by SIRT1. Deacetylation of MKP1 by direct interaction with SIRT1 increased the binding affinity to ERK which in turn facilitated inactivation of ERK. Taken together, these results suggest that SIRT1 would act as a tumor suppressor by modulating RAS-driven ERK activity through MKP1 deacetylation.
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Aims: A subpopulation of cancer cells, termed cancer stem cells (CSCs), has stemness properties, such as self-renewal and differentiation, which drive cancer recurrence and tumor resistance. CSCs possess enhanced protection capabilities to maintain reduced intracellular levels of reactive oxygen species (ROS) compared with nonstem-like cancer cells. This study investigated whether reductive stress could regulate self-renewal activity in breast CSCs. Results: We found that manifestation of stemness in breast cancer stem-like cells was associated with an elevated production of reduced glutathione (GSH) maintained by upregulation of glutamate cysteine ligase catalytic subunit (GCLC) and consequently, lowered ROS levels. This was accompanied by upregulation of phospho-AMP-activated protein kinase, FoxO3a, and Bmi-1. Notably, expression of nuclear factor erythroid-derived 2-like 2 (Nrf2) protein was substantially increased in cells undergoing sphere formation. We noticed that expression of Bmi-1 was inhibited after introduction of Nrf2 short interfering RNA into MCF-7 mammosphere cells. Silencing of Nrf2 expression suppressed the xenograft growth of subcutaneously or orthotopically injected human breast cancer cells. Innovation: Association between Nrf2 and self-renewal signaling in CSCs has been reported, but the underlying molecular mechanism remains largely unresolved. This study demonstrates the Nrf2-mediated signaling pathway in maintenance of reductive stress in breast CSCs. Conclusion: Nrf2 overactivation in breast CSCs upregulates GCLC expression and consequently enhances GSH biosynthesis with concurrent reduction in intracellular ROS accumulation, thereby provoking the reductive stress. The consequent upregulation of nuclear FoxO3a and its binding to the promoter of the gene encoding Bmi-1 account for the self-renewal activity of breast cancer stem-like cells and their growth in a xenograft mouse model.
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Proteína Forkhead Box O3/metabolismo , Neoplasias Mamárias Experimentais/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Células-Tronco Neoplásicas/metabolismo , Complexo Repressor Polycomb 1/metabolismo , Animais , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Feminino , Proteína Forkhead Box O3/genética , Humanos , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Fator 2 Relacionado a NF-E2/genética , Células-Tronco Neoplásicas/patologia , Oxirredução , Estresse Oxidativo , Complexo Repressor Polycomb 1/genética , Células Tumorais CultivadasRESUMO
Cell migration, an essential process for normal cell development and cancer metastasis, differs from a simple random walk: the mean-square displacement (ã(Δr)2(t)ã) of cells sometimes shows non-Fickian behavior, and the spatiotemporal correlation function (G(r, t)) of cells is often non-Gaussian. We find that this intriguing cell migration should be attributed to heterogeneity in a cell population, even one with a homogeneous genetic background. There are two limiting types of heterogeneity in a cell population: cellular heterogeneity and temporal heterogeneity. Cellular heterogeneity accounts for the cell-to-cell variation in migration capacity, while temporal heterogeneity arises from the temporal noise in the migration capacity of single cells. We illustrate that both cellular and temporal heterogeneity need to be taken into account simultaneously to elucidate cell migration. We investigate the two-dimensional migration of A549 lung cancer cells using time-lapse microscopy and find that the migration of A549 cells is Fickian but has a non-Gaussian spatiotemporal correlation. We find that when a theoretical model considers both cellular and temporal heterogeneity, the model reproduces all of the anomalous behaviors of cancer cell migration.
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Movimento Celular , Modelos Teóricos , Neoplasias/patologia , Processos Estocásticos , Algoritmos , Linhagem Celular Tumoral , HumanosRESUMO
Despite great potential for regenerative medicine, the high tumorigenic potential of human pluripotent stem cells (hPSCs) to form undesirable teratoma is an important technical hurdle preventing safe cell therapy. Various small molecules that induce the complete elimination of undifferentiated hPSCs, referred to as "stemotoxics," have been developed to facilitate tumor-free cell therapy, including the Survivin inhibitor YM155. In the present work, based on the chemical structure of YM155, total 26 analogs were synthesized and tested for stemotoxic activity toward human embryonic stem cells (hESCs) and induced PSCs (iPSCs). We found that a hydrogen bond acceptor in the pyrazine ring of YM155 derivatives is critical for stemotoxic activity, which is completely lost in hESCs lacking SLC35F2, which encodes a solute carrier protein. These results suggest that hydrogen bonding interactions between the nitrogens of the pyrazine ring and the SLC35F2 protein are critical for entry of YM155 into hPSCs, and hence stemotoxic activity.
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Even when targets responsible for chemoresistance are identified, drug development is often hampered due to the poor druggability of these proteins. We systematically analyzed therapy-resistance with a large-scale cancer cell transcriptome and drug-response datasets and predicted the candidate drugs based on the gene expression profile. Our results implicated the epithelial-mesenchymal transition as a common mechanism underlying resistance to chemotherapeutic drugs. Notably, we identified ITGB3, whose expression was abundant in both drug resistance and mesenchymal status, as a promising target to overcome chemoresistance. We also confirmed that depletion of ITGB3 sensitized cancer cells to conventional chemotherapeutic drugs by modulating the NF-κB signaling pathway. Considering the poor druggability of ITGB3 and the lack of feasible drugs to directly inhibit this protein, we took an in silico screening for drugs mimicking the transcriptome-level changes caused by knockdown of ITGB3. This approach successfully identified atorvastatin as a novel candidate for drug repurposing, paving an alternative path to drug screening that is applicable to undruggable targets.
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Resistencia a Medicamentos Antineoplásicos/genética , Transição Epitelial-Mesenquimal/genética , Integrina beta3/genética , Neoplasias Pulmonares/genética , Células A549 , Linhagem Celular Tumoral , Descoberta de Drogas/métodos , Humanos , NF-kappa B/genética , Farmacogenética/métodos , Transdução de Sinais/genéticaRESUMO
The selective survival advantage of culture-adapted human embryonic stem cells (hESCs) is a serious safety concern for their clinical application. With a set of hESCs with various passage numbers, we observed that a subpopulation of hESCs at late passage numbers was highly resistant to various cell death stimuli, such as YM155, a survivin inhibitor. Transcriptome analysis from YM155-sensitive (YM155S) and YM155-resistant (YM155R) hESCs demonstrated that BCL2L1 was highly expressed in YM155R hESCs. By matching the gene signature of YM155R hESCs with the Cancer Therapeutics Response Portal dataset, BH3 mimetics were predicted to selectively ablate these cells. Indeed, short-course treatment with a sub-optimal dose of BH3 mimetics induced the spontaneous death of YM155R, but not YM155S hESCs by disrupting the mitochondrial membrane potential. YM155S hESCs remained pluripotent following BH3 mimetics treatment. Therefore, the use of BH3 mimetics is a promising strategy to specifically eliminate hESCs with a selective survival advantage.
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Células-Tronco Embrionárias Humanas/citologia , Fragmentos de Peptídeos/farmacologia , Proteínas Proto-Oncogênicas/farmacologia , Compostos de Anilina/farmacologia , Contagem de Células , Células Cultivadas , Células-Tronco Embrionárias Humanas/efeitos dos fármacos , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Imidazóis/farmacologia , Naftoquinonas/farmacologia , Estresse Fisiológico/efeitos dos fármacos , Sulfonamidas/farmacologia , Proteína bcl-X/metabolismoRESUMO
Understanding the molecular mechanisms that underlie the aggressive behavior and relapse of breast cancer may help in the development of novel therapeutic interventions. CUB-domain-containing protein 1 (CDCP1), a transmembrane adaptor protein, is highly maintained and required in the context of cellular metastatic potential in triple-negative breast cancer (TNBC). For this reason, gene expression levels of CDCP1 have been considered as a prognostic marker in TNBC. However, not rarely, transcript levels of genes do not reflect always the levels of proteins, due to the post-transcriptional regulation. Here we show that miR-17/20a control the FBXL14 E3 ligase, establishing FBXL14 as an upstream regulator of the CDCP1 pathway. FBXL14 acts as an novel interaction partner of CDCP1, and facilitates its ubiquitination and proteasomal degradation with an enhanced capacity to suppress CDCP1 protein stability that eventually prevents CDCP1 target genes involved in breast cancer metastasis. Our findings first time uncovers the regulatory mechanism of CDCP-1 protein stabilization, more predictable criteria than gene expression levels for prognosis of breast cancer patients.
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Antígenos CD/metabolismo , Moléculas de Adesão Celular/metabolismo , Proteínas F-Box/metabolismo , MicroRNAs/genética , Proteínas de Neoplasias/metabolismo , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Ubiquitina-Proteína Ligases/metabolismo , Animais , Antígenos CD/genética , Antígenos de Neoplasias , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Moléculas de Adesão Celular/genética , Linhagem Celular Tumoral , Movimento Celular , Proteínas F-Box/genética , Feminino , Células HEK293 , Humanos , Células MCF-7 , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos NOD , Camundongos SCID , Invasividade Neoplásica/genética , Proteínas de Neoplasias/genética , Transplante de Neoplasias , Prognóstico , Transplante Heterólogo , Neoplasias de Mama Triplo Negativas/mortalidade , Ubiquitina-Proteína Ligases/genéticaRESUMO
The Fluorescent Ubiquitination-based Cell Cycle Indicator (FUCCI) system can be used not only to study gene expression at a specific cell cycle stage, but also to monitor cell cycle transitions in real time. In this study, we used a single clone of FUCCI-expressing HeLa cells (FUCCI-HeLa cells) and monitored the cell cycle in individual live cells over time by determining the ratios between red fluorescence (RF) of RFP-Cdt1 and green fluorescence (GF) of GFP-Geminin. Cytotoxic and cytostatic compounds, the latter of which induced G2 or mitotic arrest, were identified based on periodic cycling of the RF/GF and GF/RF ratios in FUCCI-HeLa cells treated with anti-cancer drugs. With this cell cycle monitoring system, ten flavonoids were screened. Of these, apigenin and luteolin, which have a flavone backbone, were cytotoxic, whereas kaempferol, which has a flavonol backbone, was cytostatic and induced G2 arrest. In summary, we developed a system to quantitatively monitor the cell cycle in real time. This system can be used to identify novel compounds that modulate the cell cycle and to investigate structure-activity relationships.
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Melanin is a pigment produced from tyrosine in melanocytes. Although melanin has a protective role against UVB radiation-induced damage, it is also associated with the development of melanoma and darker skin tone. Tyrosinase is a key enzyme in melanin synthesis, which regulates the rate-limiting step during conversion of tyrosine into DOPA and dopaquinone. To develop effective RNA interference therapeutics, we designed a melanin siRNA pool by applying multiple prediction programs to reduce human tyrosinase levels. First, 272 siRNAs passed the target accessibility evaluation using the RNAxs program. Then we selected 34 siRNA sequences with ΔG ≥-34.6 kcal/mol, i-Score value ≥65, and siRNA scales score ≤30. siRNAs were designed as 19-bp RNA duplexes with an asymmetric 3' overhang at the 3' end of the antisense strand. We tested if these siRNAs effectively reduced tyrosinase gene expression using qRT-PCR and found that 17 siRNA sequences were more effective than commercially available siRNA. Three siRNAs further tested showed an effective visual color change in MNT-1 human cells without cytotoxic effects, indicating these sequences are anti-melanogenic. Our study revealed that human tyrosinase siRNAs could be efficiently designed using multiple prediction algorithms.
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BACKGROUND: Glioma stem cells (GSCs) are a major cause of the frequent relapse observed in glioma, due to their high drug resistance and their differentiation potential. Therefore, understanding the molecular mechanisms governing the 'cancer stemness' of GSCs will be particularly important for improving the prognosis of glioma patients. METHODS: We previously established cancerous neural stem cells (CNSCs) from immortalized human neural stem cells (F3 cells), using the H-Ras oncogene. In this study, we utilized the EGFRviii mutation, which frequently occurs in brain cancers, to establish another CNSC line (F3.EGFRviii), and characterized its stemness under spheroid culture. RESULTS: The F3.EGFRviii cell line was highly tumorigenic in vitro and showed high ERK1/2 activity as well as expression of a variety of genes associated with cancer stemness, such as SOX2 and NANOG, under spheroid culture conditions. Through meta-analysis, PCR super-array, and subsequent biochemical assays, the induction of MEK partner-1 (MP1, encoded by the LAMTOR3 gene) was shown to play an important role in maintaining ERK1/2 activity during the acquisition of cancer stemness under spheroid culture conditions. High expression of this gene was also closely associated with poor prognosis in brain cancer. CONCLUSION: These data suggest that MP1 contributes to cancer stemness in EGFRviii-expressing glioma cells by driving ERK activity.
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Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Receptores ErbB/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neurais/metabolismo , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , PrognósticoRESUMO
Primed human pluripotent stem cells (hPSCs) are highly dependent on glycolysis rather than oxidative phosphorylation, which is similar to the metabolic switch that occurs in cancer cells. However, the molecular mechanisms that underlie this metabolic reprogramming in hPSCs and its relevance to pluripotency remain unclear. Cha et al. (2017) recently revealed that downregulation of SIRT2 by miR-200c enhances acetylation of glycolytic enzymes and glycolysis, which in turn facilitates cellular reprogramming, suggesting that SIRT2 is a key enzyme linking the metabolic switch and pluripotency in hPSCs. [BMB Reports 2017; 50(9): 435-436].
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Células-Tronco Pluripotentes/metabolismo , Sirtuína 2/metabolismo , Acetilação , Reprogramação Celular/genética , Reprogramação Celular/fisiologia , Glicólise/genética , Glicólise/fisiologia , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Fosforilação Oxidativa , Células-Tronco Pluripotentes/citologia , Processamento de Proteína Pós-Traducional/genética , Processamento de Proteína Pós-Traducional/fisiologia , Sirtuína 2/genéticaRESUMO
Mesenchymal-type cancers after epithelial mesenchymal transition (EMT) were recently shown to acquire chemoresistance through expressing EMT specific transcription factors. However, druggable (or actionable) target(s) for chemoresistance in mesenchymal-type lung cancers remain unidentified. Here, we used a public clinical genomic database and mesenchymal lung cancer cells (MLCC) model derived from the A549 lung adenocarcinoma cell line to demonstrate that BCL2 expression, which is highly induced in mesenchymal-type lung cancers, as a predictor of poor prognosis in mesenchymal lung cancer patients and association with acquired chemoradioresistance. Thereby, combination treatment with BH3 mimetics, such as ABT-263 and ABT-737, clearly attenuated chemoresistance in MLCCs. BCL2 expression in MLCCs was induced by ERK1 activity through the upregulation of the MEK1/ERK1 scaffold protein MEK partner-1 (MP1). Interfering with the MEK1/MP1/ERK1 axis using a MEK1 inhibitor or MP1 depletion repressed BCL2 expression and sensitized MLCCs to chemoradiotherapy. Taken together, our results suggest that targeting druggable proteins in the MEK1/MP1/ERK1/BCL2 axis, such as MEK1 or BCL2, with currently available FDA approved drugs is a currently feasible approach to improve clinical outcomes of mesenchymal lung cancer patients.
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Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adenocarcinoma/terapia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Quimiorradioterapia , Resistencia a Medicamentos Antineoplásicos , Neoplasias Pulmonares/terapia , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Fragmentos de Peptídeos/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Proto-Oncogênicas/farmacologia , Tolerância a Radiação , Células A549 , Adenocarcinoma/enzimologia , Adenocarcinoma/genética , Adenocarcinoma/patologia , Adenocarcinoma de Pulmão , Compostos de Anilina/farmacologia , Benzamidas/farmacologia , Compostos de Bifenilo/farmacologia , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Difenilamina/análogos & derivados , Difenilamina/farmacologia , Relação Dose-Resposta a Droga , Etoposídeo/farmacologia , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , MAP Quinase Quinase 1/antagonistas & inibidores , MAP Quinase Quinase 1/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Mimetismo Molecular , Nitrofenóis/farmacologia , Piperazinas/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/genética , Interferência de RNA , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/efeitos da radiação , Sulfonamidas/farmacologia , Transfecção , Regulação para CimaRESUMO
Ell3 is an RNA polymerase II transcription elongation factor that acts as a negative regulator of p53 expression, and regulates cell proliferation and survival. Recent studies by our group have demonstrated that ectopic expression of Ell3 in breast cancer cell lines enhances cell proliferation, potentiates cancer stem cell properties, and promotes 5-Fluorouracil (5-FU) resistance. In the present study, the underlying mechanism for the induction of 5-FU resistance was investigated in Ell3 over-expressing MCF-7 cells (Ell3 OE cells). By comparing the gene expression profiles of Ell3 OE cells with control cells, the present data revealed that Lipocalin2 (LCN2) and Wnt signaling activity are associated with 5-FU resistance of Ell3 OE. siRNA-mediated suppression of LCN2 reversed 5-FU resistance in Ell3 OE cells. Chemical inhibition of Wnt signaling also reversed 5-FU resistance in Ell3 OE cells. Furthermore, the expression levels of survivin, which is a direct transcriptional target of Wnt/ß-catenin and an inhibitor of apoptosis, were markedly elevated when Ell3 OE cells were treated with 5-FU, as detected by western blot analysis. These findings suggest that enhanced expression of LCN2 and activation of the Wnt signaling pathway may induce 5-FU resistance in Ell3 OE cells as a means of evading apoptosis.