RESUMO
Loss of LKB1 is associated with increased metastasis and poor prognosis in lung cancer, but the development of targeted agents is in its infancy. Here we report that a glutaminolytic enzyme, glutamate dehydrogenase 1 (GDH1), upregulated upon detachment via pleomorphic adenoma gene 1 (PLAG1), provides anti-anoikis and pro-metastatic signals in LKB1-deficient lung cancer. Mechanistically, the GDH1 product α-KG activates CamKK2 by enhancing its substrate AMPK binding, which contributes to energy production that confers anoikis resistance. The effect of GDH1 on AMPK is evident in LKB1-deficient lung cancer, where AMPK activation predominantly depends on CamKK2. Targeting GDH1 with R162 attenuated tumor metastasis in patient-derived xenograft model and correlation studies in lung cancer patients further validated the clinical relevance of our finding. Our study provides insight into the molecular mechanism by which GDH1-mediated metabolic reprogramming of glutaminolysis mediates lung cancer metastasis and offers a therapeutic strategy for patients with LKB1-deficient lung cancer.
Assuntos
Anoikis/fisiologia , Proteínas de Ligação a DNA/metabolismo , Glutamato Desidrogenase/metabolismo , Neoplasias Pulmonares/patologia , Proteínas Serina-Treonina Quinases/genética , Carcinoma de Pequenas Células do Pulmão/patologia , Células A549 , Quinases Proteína-Quinases Ativadas por AMP , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Quinase da Proteína Quinase Dependente de Cálcio-Calmodulina/metabolismo , Linhagem Celular Tumoral , Ativação Enzimática/fisiologia , Feminino , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos Nus , Camundongos SCID , Metástase Neoplásica/patologia , Transplante de Neoplasias , Transplante HeterólogoRESUMO
Metastasis is the leading cause of death in patients with breast, lung, and head and neck cancers. However, the molecular mechanisms underlying metastases in these cancers remain unclear. We found that the p90 ribosomal S6 kinase 2 (RSK2)-cAMP response element-binding protein (CREB) pathway is commonly activated in diverse metastatic human cancer cells, leading to up-regulation of a CREB transcription target Fascin-1. We also observed that the protein expression patterns of RSK2 and Fascin-1 correlate in primary human tumor tissue samples from head and neck squamous cell carcinoma patients. Moreover, knockdown of RSK2 disrupts filopodia formation and bundling in highly invasive cancer cells, leading to attenuated cancer cell invasion in vitro and tumor metastasis in vivo, whereas expression of Fascin-1 significantly rescues these phenotypes. Furthermore, targeting RSK2 with the small molecule RSK inhibitor FMK-MEA effectively attenuated the invasive and metastatic potential of cancer cells in vitro and in vivo, respectively. Taken together, our findings for the first time link RSK2-CREB signaling to filopodia formation and bundling through the up-regulation of Fascin-1, providing a proinvasive and prometastatic advantage to human cancers. Therefore, protein effectors of the RSK2-CREB-Fascin-1 pathway represent promising biomarkers and therapeutic targets in the clinical prognosis and treatment of metastatic human cancers.
Assuntos
Biomarcadores Tumorais/metabolismo , Proteína de Ligação a CREB/metabolismo , Proteínas de Transporte/biossíntese , Regulação Neoplásica da Expressão Gênica , Proteínas dos Microfilamentos/biossíntese , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Proteínas Quinases S6 Ribossômicas 90-kDa/metabolismo , Transdução de Sinais , Animais , Biomarcadores Tumorais/genética , Proteína de Ligação a CREB/genética , Proteínas de Transporte/genética , Linhagem Celular Tumoral , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Camundongos Nus , Proteínas dos Microfilamentos/genética , Metástase Neoplásica , Proteínas de Neoplasias/genética , Neoplasias/genética , Neoplasias/patologia , Pseudópodes/genética , Pseudópodes/metabolismo , Pseudópodes/patologia , Proteínas Quinases S6 Ribossômicas 90-kDa/genética , Regulação para Cima/genéticaRESUMO
The choroid plexuses (CPs) form the blood-cerebrospinal fluid (CSF) barrier (BCSFB) and play an important role in maintaining brain normal function and the brain response to injury. Many neurological disorders are associated with oxidative stress that can impact CP function. This study examined the effects of isothiocyanates, an abundant component in cruciferous vegetables, on H(2)O(2)-induced BCSFB disruption and CP cell death in vitro. It further examined the potential role of a transcription factor, nuclear factor erythroid 2-related factor 2 (Nrf2), in isothiocyanate-induced protection. Sulforaphane (SF) significantly reduced H(2)O(2)-induced BCSFB disruption as assessed by transepithelial electrical resistance (29 ± 7% reduction vs. 92 ± 2% decrease in controls) and [(3)H]mannitol permeability. Allyl-isothiocyanate (AITC) had a similar protective effect. H(2)O(2)-induced epithelial cell death was also reduced by these isothiocyanates. In primary CP cells, SF and AITC reduced cell death by 42 ± 3% and 53 ± 10%, respectively. Similar protection was found in a CP cell line Z310. Protection was only found with pretreatment for 12-48 h and not with acute exposure (1 h). The protective effects of SF and AITC were associated with Nrf2 nuclear translocation and upregulated expression of antioxidative systems regulated by Nrf2, including heme oxygenase-1, NAD(P)H quinine oxidoreductase, and cysteine/glutamate exchange transporter. Thus isothiocyanates, as diet or medicine, may be a method for protecting BCSFB in neurological disorders.
Assuntos
Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/fisiopatologia , Plexo Corióideo/efeitos dos fármacos , Isotiocianatos/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Animais , Antioxidantes/fisiologia , Apoptose/efeitos dos fármacos , Linhagem Celular , Células Cultivadas , Plexo Corióideo/citologia , Plexo Corióideo/fisiopatologia , Peróxido de Hidrogênio/efeitos adversos , Peróxido de Hidrogênio/farmacologia , Técnicas In Vitro , Modelos Animais , Fator 2 Relacionado a NF-E2/fisiologia , Estresse Oxidativo/fisiologia , Ratos , Ratos Sprague-Dawley , Sulfóxidos , Tiocianatos/farmacologiaRESUMO
Platinum-based chemotherapeutics represent a mainstay of cancer therapy, but resistance limits their curative potential. Through a kinome RNAi screen, we identified microtubule-associated serine/threonine kinase 1 (MAST1) as a main driver of cisplatin resistance in human cancers. Mechanistically, cisplatin but no other DNA-damaging agents inhibit the MAPK pathway by dissociating cRaf from MEK1, while MAST1 replaces cRaf to reactivate the MAPK pathway in a cRaf-independent manner. We show clinical evidence that expression of MAST1, both initial and cisplatin-induced, contributes to platinum resistance and worse clinical outcome. Targeting MAST1 with lestaurtinib, a recently identified MAST1 inhibitor, restores cisplatin sensitivity, leading to the synergistic attenuation of cancer cell proliferation and tumor growth in human cancer cells and patient-derived xenograft models.
Assuntos
Antineoplásicos/farmacologia , Cisplatino/farmacologia , MAP Quinase Quinase 1/fisiologia , Proteínas Associadas aos Microtúbulos/fisiologia , Proteínas Serina-Treonina Quinases/fisiologia , Proteínas Proto-Oncogênicas c-raf/fisiologia , Animais , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Ativação Enzimática , Feminino , Humanos , CamundongosRESUMO
How mitochondrial glutaminolysis contributes to redox homeostasis in cancer cells remains unclear. Here we report that the mitochondrial enzyme glutamate dehydrogenase 1 (GDH1) is commonly upregulated in human cancers. GDH1 is important for redox homeostasis in cancer cells by controlling the intracellular levels of its product alpha-ketoglutarate and subsequent metabolite fumarate. Mechanistically, fumarate binds to and activates a reactive oxygen species scavenging enzyme glutathione peroxidase 1. Targeting GDH1 by shRNA or a small molecule inhibitor R162 resulted in imbalanced redox homeostasis, leading to attenuated cancer cell proliferation and tumor growth.
Assuntos
Glutamato Desidrogenase/biossíntese , Glutationa Peroxidase/biossíntese , Glutationa/metabolismo , Leucemia/genética , Mitocôndrias/enzimologia , Antioxidantes/metabolismo , Carcinogênese , Fumaratos/metabolismo , Regulação Neoplásica da Expressão Gênica , Glutamato Desidrogenase/antagonistas & inibidores , Glutamato Desidrogenase/genética , Glutationa Peroxidase/genética , Humanos , Ácidos Cetoglutáricos/metabolismo , Leucemia/enzimologia , Leucemia/patologia , Mitocôndrias/patologia , Oxirredução , Cultura Primária de Células , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/genética , Glutationa Peroxidase GPX1RESUMO
How invasive and metastatic tumor cells evade anoikis induction remains unclear. We found that knockdown of RSK2 sensitizes diverse cancer cells to anoikis induction, which is mediated through phosphorylation targets including apoptosis signal-regulating kinase 1 (ASK1) and cyclic AMP (cAMP) response element-binding protein (CREB). We provide evidence to show that RSK2 inhibits ASK1 by phosphorylating S83, T1109, and T1326 through a novel mechanism in which phospho-T1109/T1326 inhibits ATP binding to ASK1, while phospho-S83 attenuates ASK1 substrate MKK6 binding. Moreover, the RSK2âCREB signaling pathway provides antianoikis protection by regulating gene expression of protein effectors that are involved in cell death regulation, including the antiapoptotic factor protein tyrosine kinase 6 (PTK6) and the proapoptotic factor inhibitor-of-growth protein 3 (ING3). PTK6 overexpression or ING3 knockdown in addition to ASK1 knockdown further rescued the increased sensitivity to anoikis induction in RSK2 knockdown cells. These data together suggest that RSK2 functions as a signal integrator to provide antianoikis protection to cancer cells in both transcription-independent and -dependent manners, in part by signaling through ASK1 and CREB, and contributes to cancer cell invasion and tumor metastasis.