RESUMO
Lung cancer is the leading cause of cancer-related deaths worldwide. KRAS is the main oncogenic driver in lung cancer that can be activated by gene mutation or amplification, but whether long non-coding RNAs (lncRNAs) regulate its activation remains unknown. Through gain and loss of function approaches, we identified that lncRNA HIF1A-As2, a KRAS-induced lncRNA, is required for cell proliferation, epithelial-mesenchymal transition (EMT) and tumor propagation in non-small cell lung cancer (NSCLC) in vitro and in vivo. Integrative analysis of HIF1A-As2 transcriptomic profiling reveals that HIF1A-As2 modulates gene expression in trans, particularly regulating transcriptional factor genes including MYC. Mechanistically, HIF1A-As2 epigenetically activates MYC by recruiting DHX9 on MYC promoter, consequently stimulating the transcription of MYC and its target genes. In addition, KRAS promotes HIF1A-As2 expression via the induction of MYC, suggesting HIF1A-As2 and MYC form a double-regulatory loop to strengthen cell proliferation and tumor metastasis in lung cancer. Inhibition of HIF1A-As2 by LNA GapmeR antisense oligonucleotides (ASO) significantly improves sensitization to 10058-F4 (a MYC-specific inhibitor) and cisplatin treatment in PDX and KRASLSLG12D-driven lung tumors, respectively.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , MicroRNAs , RNA Longo não Codificante , Humanos , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Linhagem Celular Tumoral , Carcinoma Pulmonar de Células não Pequenas/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Retroalimentação , Neoplasias Pulmonares/genética , MicroRNAs/genética , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismoRESUMO
[This corrects the article DOI: 10.1371/journal.pone.0067581.].
RESUMO
AMP-activated protein kinase (AMPK) is a critical sensor of energy status that coordinates cell growth with energy balance. In non-small cell lung cancer (NSCLC) the role of AMPKα is controversial and its contribution to lung carcinogenesis is not well-defined. Furthermore, it remains largely unknown whether long non-coding RNAs (lncRNAs) are involved in the regulation of AMPK-mediated pathways. Here, we found that loss of AMPKα in combination with activation of mutant KRASG12D increased lung tumour burden and reduced survival in KrasLSLG12D/+/AMPKαfl/fl mice. In agreement, functional in vitro studies revealed that AMPKα silencing increased growth and migration of NSCLC cells. In addition, we identified an AMPKα-modulated lncRNA, KIMAT1 (ENSG00000228709), which in turn regulates AMPKα activation by stabilizing the lactate dehydrogenase B (LDHB). Collectively, our study indicates that AMPKα loss promotes KRAS-mediated lung tumorigenesis and proposes a novel KRAS/KIMAT1/LDHB/AMPKα axis that could be exploited for therapeutic purposes.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Neoplasias Pulmonares/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Animais , Carcinogênese , Modelos Animais de Doenças , Humanos , CamundongosRESUMO
Wild-type KRAS (KRASWT) amplification has been shown to be a secondary means of KRAS activation in cancer and associated with poor survival. Nevertheless, the precise role of KRASWT overexpression in lung cancer progression is largely unexplored. Here, we identify and characterize a KRAS-responsive lncRNA, KIMAT1 (ENSG00000228709) and show that it correlates with KRAS levels both in cell lines and in lung cancer specimens. Mechanistically, KIMAT1 is a MYC target and drives lung tumorigenesis by promoting the processing of oncogenic microRNAs (miRNAs) through DHX9 and NPM1 stabilization while halting the biogenesis of miRNAs with tumor suppressor function via MYC-dependent silencing of p21, a component of the Microprocessor Complex. KIMAT1 knockdown suppresses not only KRAS expression but also KRAS downstream signaling, thereby arresting lung cancer growth in vitro and in vivo. Taken together, this study uncovers a role for KIMAT1 in maintaining a positive feedback loop that sustains KRAS signaling during lung cancer progression and provides a proof of principle that interfering with KIMAT1 could be a strategy to hamper KRAS-induced tumorigenesis.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , MicroRNAs/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , RNA Longo não Codificante/genética , Células A549 , Animais , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/terapia , Linhagem Celular Tumoral , Feminino , Perfilação da Expressão Gênica/métodos , Ontologia Genética , Humanos , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/terapia , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Nucleofosmina , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
Medulloblastoma (MB) is the most common aggressive paediatric brain tumour and, despite the recent progress in the treatments of MB patients, there is still an urgent need of complementary or alternative therapeutic options for MB infants. Cyclin Dependent Kinase inhibitors (CDKi) are at the front-line of novel targeted treatments for multiple cancers and the CDK4/6 specific inhibitor palbociclib has been pre-clinically identified as an effective option for MB cells. Herein, we identified the pan-CDKi dinaciclib as a promising alternative to palbociclib for the suppression of MB cells proliferation. We present evidence supporting dinaciclib's ability to inhibit MB cells in vitro proliferation at considerably lower doses than palbociclib. Sequencing data and pathway analysis suggested that dinaciclib is a potent cell death inducer in MB cells. We found that dinaciclib-triggered apoptosis is triggered by CDK9 inhibition and the resultant reduction in RNA pol II phosphorylation, which leads to the downregulation of the oncogenic marker MYC, and the anti-apoptotic protein MCL-1. Specifically, we demonstrated that MCL-1 is a key apoptotic mediator for MB cells and co-treatment of dinaciclib with BH3 mimetics boosts the therapeutic efficacy of dinaciclib. Together, these findings highlight the potential of multi-CDK inhibition by dinaciclib as an alternative option to CDK4/6 specific inhibition, frequently associated with drug resistance in patients.
Assuntos
Proliferação de Células/efeitos dos fármacos , Óxidos N-Cíclicos/farmacologia , Quinases Ciclina-Dependentes , Indolizinas/farmacologia , Meduloblastoma , Proteínas de Neoplasias , Inibidores de Proteínas Quinases/farmacologia , Compostos de Piridínio/farmacologia , Quinases Ciclina-Dependentes/antagonistas & inibidores , Quinases Ciclina-Dependentes/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Meduloblastoma/tratamento farmacológico , Meduloblastoma/enzimologia , Meduloblastoma/patologia , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/metabolismoRESUMO
Chromosomal instability (CIN) is a driver of clonal diversification and intratumor heterogeneity, providing genetic diversity that contributes to tumor progression. It is estimated that approximately 80% of solid cancers, including non-small cell lung cancer (NSCLC), exhibit features of CIN, which affects tumor growth and response to therapy. However, the molecular mechanisms connecting CIN to tumor progression are still poorly understood. Through an RNAi screen performed on genes involved in CIN and overexpressed in human lung adenocarcinoma samples, we identified the cytoskeleton-associated protein 2-like (CKAP2L) as a potential oncogene that promotes lung cancer proliferation and growth in vitro and in vivo. Mechanistically, CKAP2L directly interacted with RNA Pol II and regulated transcription elongation of key genes involved in spindle assembly checkpoint, chromosome segregation, cell cycle, and E2F signaling. Furthermore, depletion of CKAP2L increased the sensitivity of NSCLC cells to alvocidib, a pan-CDK inhibitor, leading to a significant reduction of cell proliferation and an increase in cell death. Altogether, these findings shed light on the molecular mechanisms through which CKAP2L, a protein involved in CIN, promotes cancer progression and suggest that its inhibition represents a novel therapeutic strategy in NSCLC. SIGNIFICANCE: These findings demonstrate the oncogenic function of CKAP2L through regulation of transcription elongation and suggest that targeting CKAP2L could enhance therapeutic response in patients with NSCLC.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/patologia , Proteínas do Citoesqueleto/fisiologia , Neoplasias Pulmonares/patologia , Elongação da Transcrição Genética , Células A549 , Animais , Carcinogênese/efeitos dos fármacos , Carcinogênese/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Linhagem Celular Tumoral , Proteínas do Citoesqueleto/química , Proteínas do Citoesqueleto/genética , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Neoplasias Pulmonares/genética , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Metástase Neoplásica , RNA Interferente Pequeno/farmacologia , RNA Interferente Pequeno/uso terapêutico , Elongação da Transcrição Genética/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Non-small-cell lung cancer (NSCLC) is the most prevalent form of lung cancer and has a poor five-year survival rate of 15%. Chemotherapy and targeted therapies have significantly improved patients' prognosis. Nevertheless, after a successful initial response, some patients relapse when cancer cells become resistant to drug treatments, representing an important clinical limitation. Therefore, investigating the mechanisms of drug resistance is of significant importance. Recently, considerable attention has been given to long non-coding RNAs (lncRNAs), a heterogeneous class of regulatory molecules that play essential roles in tumorigenesis by modulating genes and signalling pathways involved in cell growth, metastasis and drug response. In this article, we review recent research findings on the role of lncRNAs in drug resistance in NSCLC, highlighting their mechanisms of action.
Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Neoplasias Pulmonares/genética , RNA Longo não Codificante/metabolismo , Carcinoma Pulmonar de Células não Pequenas/genéticaRESUMO
Lung cancer is associated with a high mortality, with around 1.8 million deaths worldwide in 2018. Non-small-cell lung cancer (NSCLC) accounts for around 85% of cases and, despite improvement in the management of NSCLC, most patients are diagnosed at advanced stage and the five-year survival remains around 15%. This highlights a need to identify novel ways to treat the disease to reduce the burden of NSCLC. Long non-coding RNAs (lncRNAs) are non-coding RNA molecules longer than 200 nucleotides in length which play important roles in gene expression and signaling pathways. Recently, lncRNAs were implicated in cancer, where their expression is dysregulated resulting in aberrant functions. LncRNAs were shown to function as both tumor suppressors and oncogenes in a variety of cancer types. Although there are a few well characterized lncRNAs in NSCLC, many lncRNAs remain un-characterized and their mechanisms of action largely unknown. LncRNAs have success as therapies in neurodegenerative diseases, and having a detailed understanding of their function in NSCLC may guide novel therapeutic approaches and strategies. This review discusses the role of lncRNAs in NSCLC tumorigenesis, highlighting their mechanisms of action and their clinical potential.
RESUMO
A subset of lung adenocarcinomas is driven by the EML4-ALK translocation. Even though ALK inhibitors in the clinic lead to excellent initial responses, acquired resistance to these inhibitors due to on-target mutations or parallel pathway alterations is a major clinical challenge. Exploring these mechanisms of resistance, we found that EML4-ALK cells parental or resistant to crizotinib, ceritinib or alectinib are remarkably sensitive to inhibition of CDK7/12 with THZ1 and CDK9 with alvocidib or dinaciclib. These compounds robustly induce apoptosis through transcriptional inhibition and downregulation of anti-apoptotic genes. Importantly, alvocidib reduced tumour progression in xenograft mouse models. In summary, our study takes advantage of the transcriptional addiction hypothesis to propose a new treatment strategy for a subset of patients with acquired resistance to first-, second- and third-generation ALK inhibitors.
Assuntos
Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Proteínas de Fusão Oncogênica/genética , Inibidores de Proteínas Quinases/farmacologia , Transcrição Gênica/efeitos dos fármacos , Animais , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Feminino , Humanos , Camundongos , Proteínas de Fusão Oncogênica/antagonistas & inibidores , Inibidores de Proteínas Quinases/uso terapêuticoRESUMO
Non-small cell lung carcinoma (NSCLC) is leading cause of cancer-related deaths in the world. The Tumor Suppressor Candidate 3 (TUSC3) at chromosome 8p22 known to be frequently deleted in cancer is often found to be deleted in advanced stage of solid tumors. However, the role of TUSC3 still remains controversial in lung cancer and context-dependent in several cancers. Here we propose that miR-224/-520c-dependent TUSC3 deficiency enhances the metastatic potential of NSCLC through the alteration of three unfolded protein response pathways and HRD1-dependent ERAD. ATF6α-dependent UPR is enhanced whereas the affinity of HRD1 to its substrates, PERK, IRE1α and p53 is weakened. Consequently, the alteration of UPRs and the suppressed p53-NM23H1/2 pathway by TUSC3 deficiency is ultimately responsible for enhancing metastatic potential of lung cancer. These findings provide mechanistic insight of unrecognized roles of TUSC3 in cancer progression and the oncogenic role of HRD1-dependent ERAD in cancer metastasis.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Degradação Associada com o Retículo Endoplasmático/fisiologia , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Proteínas de Membrana/metabolismo , MicroRNAs/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Fator 6 Ativador da Transcrição/genética , Fator 6 Ativador da Transcrição/metabolismo , Animais , Carcinoma Pulmonar de Células não Pequenas/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Proliferação de Células/fisiologia , Degradação Associada com o Retículo Endoplasmático/genética , Endorribonucleases/genética , Endorribonucleases/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Hibridização In Situ , Neoplasias Pulmonares/genética , Proteínas de Membrana/genética , Camundongos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de Tumor/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Resposta a Proteínas não Dobradas/genética , Resposta a Proteínas não Dobradas/fisiologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
A plethora of tumours have characteristic oncogenic mutations which are the main causes of malignant transformation, exerting their effects through multiple signalling pathways. Downstream of such pathways, microRNAs are small non-coding RNAs that negatively regulate gene expression, assisting or antagonizing oncogenic signalling. The differential expression of microRNAs in cancer is well-documented and is considered a fundamental aspect of tumourigenesis. While data mapping the interaction between oncogenic lesions and microRNAs are accruing, we provide particular cases of such interaction. Except for notable, well-studied examples of microRNAs regulated by oncogenes, we examine the effect of this relationship in regard to tumour initiation, progression, metastasis and ultimately, its implications for the development of new therapeutics.
Assuntos
MicroRNAs/genética , Neoplasias/patologia , Neoplasias/terapia , Oncogenes , Progressão da Doença , Humanos , Neoplasias/genéticaRESUMO
Oncogenic KRAS induces tumor onset and development by modulating gene expression via different molecular mechanisms. MicroRNAs (miRNAs) are small non-coding RNAs that have been established as main players in tumorigenesis. By overexpressing wild type or mutant KRAS (KRASG12D) and using inducible human and mouse cell lines, we analyzed KRAS-regulated microRNAs in non-small-cell lung cancer (NSCLC). We show that miR-30c and miR-21 are significantly upregulated by both KRAS isoforms and induce drug resistance and enhance cell migration/invasion via inhibiting crucial tumor suppressor genes, such as NF1, RASA1, BID, and RASSF8. MiR-30c and miR-21 levels were significantly elevated in tumors from patients that underwent surgical resection of early stages NSCLC compared to normal lung and in plasma from the same patients. Systemic delivery of LNA-anti-miR-21 in combination with cisplatin in vivo completely suppressed the development of lung tumors in a mouse model of lung cancer. Mechanistically, we demonstrated that ELK1 is responsible for miR-30c and miR-21 transcriptional activation by direct binding to the miRNA proximal promoter regions. In summary, our study defines that miR-30c and miR-21 may be valid biomarkers for early NSCLC detection and their silencing could be beneficial for therapeutic applications.
Assuntos
Carcinogênese/genética , Neoplasias Pulmonares/genética , MicroRNAs/efeitos adversos , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Animais , Humanos , Neoplasias Pulmonares/fisiopatologia , CamundongosRESUMO
In NSCLC alterations in PDGF receptors are markers of worst prognosis and efficient targeting of these receptors is yet to be achieved. In this study, we explored PDGFR-regulated microRNAs demonstrating that miR-23b cluster and miR-125a-5p are downregulated by increased expression of PDGFR-α or PDGFR-ß in NSCLC cells. Mechanistically, the expression of these microRNAs is positively regulated by p53 and negatively modulated by NF-kB p65. Forced expression of miR-23b cluster or miR-125a-5p enhanced drug sensitivity and suppressed invasiveness of NSCLC cells by silencing several genes involved in oncogenic KRAS and NF-kB pathways, including SOS1, GRB2, IQGAP1, RALA, RAF-1, IKKß, AKT2, ERK2 and KRAS itself. Of note, an inverse correlation between miR-23b cluster, miR-125a-5p and respective target genes was also found in vivo in a large dataset of lung adenocarcinoma samples. Furthermore, in vivo delivery of miR-23b cluster or miR-125a-5p significantly repressed tumour growth in a highly aggressive NSCLC circulating tumour cell (CTC) patient derived explant (CDX) mouse model. In conclusion, our finding sheds light on the PDGFR signaling and endorses the possibility to employ miR-23b cluster and miR-125a-5p as therapeutic tools to silence simultaneously a range of redundant pathways and main effectors of tumorigenesis in NSCLC.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , MicroRNAs/metabolismo , Receptores do Fator de Crescimento Derivado de Plaquetas/metabolismo , Transdução de Sinais/genética , Animais , Antineoplásicos/farmacologia , Carcinogênese/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/terapia , Regulação para Baixo , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica , Terapia Genética , Humanos , Pulmão/patologia , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/terapia , Masculino , Camundongos , MicroRNAs/administração & dosagem , MicroRNAs/genética , Pessoa de Meia-Idade , Família Multigênica/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Fator de Transcrição RelA/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Lung cancer is considered the most deadly of all cancers, with limited therapeutic options. Although advanced drugs have been tried in clinic, the therapeutic success has largely been hampered due to rapid development of drug-resistance mechanisms. Recently, microRNAs (miRNAs), a class of small non-coding RNAs, have occupied center stage in cancer biology. miRNAs negatively regulate gene expression either by promoting degradation or by interfering with translation of messenger RNA targets. Several lines of evidence have confirmed the crucial role of miRNAs in carcinogenesis, and, importantly, in the acquisition of resistance to chemotherapeutics. Modulation of miRNA expression levels has been proven to increase the efficacy of genotoxic drugs in various preclinical cancer studies. Therefore, comprehensive understanding of the role(s) of these key players in drug resistance may provide novel opportunities to design effective combinatorial therapeutic strategies for cancer treatment. In this review, we highlight recent findings on miRNAs acting as oncomiRs and tumor suppressor genes in lung cancer. Moreover, we discuss the involvement of miRNAs in different mechanisms of drug resistance in this deadly disease.
RESUMO
BACKGROUND: Overexpression of microRNA-31 (miR-31) is implicated in the pathogenesis of esophageal squamous cell carcinoma (ESCC), a deadly disease associated with dietary zinc deficiency. Using a rat model that recapitulates features of human ESCC, the mechanism whereby Zn regulates miR-31 expression to promote ESCC is examined. METHODS: To inhibit in vivo esophageal miR-31 overexpression in Zn-deficient rats (n = 12-20 per group), locked nucleic acid-modified anti-miR-31 oligonucleotides were administered over five weeks. miR-31 expression was determined by northern blotting, quantitative polymerase chain reaction, and in situ hybridization. Physiological miR-31 targets were identified by microarray analysis and verified by luciferase reporter assay. Cellular proliferation, apoptosis, and expression of inflammation genes were determined by immunoblotting, caspase assays, and immunohistochemistry. The miR-31 promoter in Zn-deficient esophagus was identified by ChIP-seq using an antibody for histone mark H3K4me3. Data were analyzed with t test and analysis of variance. All statistical tests were two-sided. RESULTS: In vivo, anti-miR-31 reduced miR-31 overexpression (P = .002) and suppressed the esophageal preneoplasia in Zn-deficient rats. At the same time, the miR-31 target Stk40 was derepressed, thereby inhibiting the STK40-NF-κΒ-controlled inflammatory pathway, with resultant decreased cellular proliferation and activated apoptosis (caspase 3/7 activities, fold change = 10.7, P = .005). This same connection between miR-31 overexpression and STK40/NF-κΒ expression was also documented in human ESCC cell lines. In Zn-deficient esophagus, the miR-31 promoter region and NF-κΒ binding site were activated. Zn replenishment restored the regulation of this genomic region and a normal esophageal phenotype. CONCLUSIONS: The data define the in vivo signaling pathway underlying interaction of Zn deficiency and miR-31 overexpression in esophageal neoplasia and provide a mechanistic rationale for miR-31 as a therapeutic target for ESCC.