RESUMEN
Cancer cells depend on nicotinamide adenine dinucleotide phosphate (NADPH) to combat oxidative stress and support reductive biosynthesis. One major NADPH production route is the oxidative pentose phosphate pathway (committed step: glucose-6-phosphate dehydrogenase, G6PD). Alternatives exist and can compensate in some tumors. Here, using genetically-engineered lung cancer mouse models, we show that G6PD ablation significantly suppresses KrasG12D/+;Lkb1-/- (KL) but not KrasG12D/+;P53-/- (KP) lung tumorigenesis. In vivo isotope tracing and metabolomics reveal that G6PD ablation significantly impairs NADPH generation, redox balance, and de novo lipogenesis in KL but not KP lung tumors. Mechanistically, in KL tumors, G6PD ablation activates p53, suppressing tumor growth. As tumors progress, G6PD-deficient KL tumors increase an alternative NADPH source from serine-driven one carbon metabolism, rendering associated tumor-derived cell lines sensitive to serine/glycine depletion. Thus, oncogenic driver mutations determine lung cancer dependence on G6PD, whose targeting is a potential therapeutic strategy for tumors harboring KRAS and LKB1 co-mutations.
Asunto(s)
Glucosafosfato Deshidrogenasa , Homeostasis , Neoplasias Pulmonares , NADP , Oxidación-Reducción , Proteínas Serina-Treonina Quinasas , Proteínas Proto-Oncogénicas p21(ras) , Glucosafosfato Deshidrogenasa/metabolismo , Glucosafosfato Deshidrogenasa/genética , Animales , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , NADP/metabolismo , Ratones , Humanos , Línea Celular Tumoral , Lipogénesis/genética , Proteína p53 Supresora de Tumor/metabolismo , Proteína p53 Supresora de Tumor/genética , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Quinasas de la Proteína-Quinasa Activada por el AMP/metabolismo , Vía de Pentosa Fosfato/genética , Proteínas Quinasas Activadas por AMP/metabolismo , Masculino , Ratones Noqueados , Femenino , MutaciónRESUMEN
Lipid metabolism disorders are a major cause of several chronic metabolic diseases which seriously affect public health. Salusinα, a vasoactive peptide, has been shown to attenuate lipid metabolism disorders, although its mechanism of action has not been reported. To investigate the effects and potential mechanisms of Salusinα on lipid metabolism, Salusinα was overexpressed or knocked down using lentiviral vectors. Hepatocyte steatosis was induced by free fatty acid (FFA) after lentiviral transfection into HepG2 cells. The degree of lipid accumulation was assessed using Oil Red O staining and by measuring several biochemical indices. Subsequently, bioinformatics was used to analyze the signaling pathways that may have been involved in lipid metabolism disorders. Finally, semiquantitative PCR and western blotting were used to verify the involvement of the liver kinase B1 (LKB1)/AMPK pathway. Compound C, an inhibitor of AMPK, was used to confirm this mechanism's involvement further. The results showed that Salusinα significantly attenuated lipid accumulation, inflammation and oxidative stress. In addition, Salusinα increased the levels of LKB1 and AMPK, which inhibited the expression of sterol regulatory element binding protein1c, fatty acid synthase and acetylCoA carboxylase. The addition of Compound C abrogated the Salusinαmediated regulation of AMPK on downstream signaling molecules. In summary, overexpression of Salusinα activated the LKB1/AMPK pathway, which in turn inhibited lipid accumulation in HepG2 cells. This provides insights into the potential mechanism underlying the mechanism by which Salusinα ameliorates lipid metabolism disorders while identifying a potential therapeutic target.
Asunto(s)
Quinasas de la Proteína-Quinasa Activada por el AMP , Proteínas Quinasas Activadas por AMP , Lipogénesis , Proteínas Serina-Treonina Quinasas , Transducción de Señal , Humanos , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Proteínas Quinasas Activadas por AMP/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Células Hep G2 , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Péptidos y Proteínas de Señalización Intercelular/genética , Metabolismo de los Lípidos/efectos de los fármacos , Metabolismo de los Lípidos/genética , Trastornos del Metabolismo de los Lípidos/metabolismo , Trastornos del Metabolismo de los Lípidos/genética , Trastornos del Metabolismo de los Lípidos/tratamiento farmacológico , Lipogénesis/genética , Lipogénesis/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Transducción de Señal/efectos de los fármacos , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/metabolismo , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/genéticaRESUMEN
BACKGROUND: Liver kinase B1 (LKB1) is frequently mutated in lung adenocarcinoma, and its loss contributes to tumor progression. METHODS: To identify LKB1 downstream genes that promote lung adenocarcinoma aggressiveness, we performed bioinformatical analysis using publicly available datasets. RESULTS: Rab3B was upregulated in LKB1-depleted lung adenocarcinoma cells and suppressed by LKB1 overexpression. CREB protein was enriched at the promoter of Rab3B in lung cancer cells. Silencing of CREB abrogated the upregulation of Rab3B upon LKB1 loss. Immunohistochemistry revealed the elevated expression of Rab3B in lung adenocarcinomas relative to adjacent normal tissues. Upregulation of Rab3B was significantly associated with lymph node metastasis, advanced tumor stage, and reduced overall survival in lung adenocarcinoma patients. Knockdown of Rab3B suppressed and overexpression of Rab3B promoted the proliferation, colony formation, and migration of lung adenocarcinoma cells in vitro. In a mouse xenograft model, Rab3B depletion restrained and Rab3B overexpression augmented the growth of lung adenocarcinoma tumors. Mechanistically, Rab3B interacted with DDX6 and enhanced its protein stability. Ectopic expression of DDX6 significantly promoted the proliferation, colony formation, and migration of lung adenocarcinoma cells. DDX6 knockdown phenocopied the effects of Rab3B depletion on lung adenocarcinoma cells. Additionally, DDX6 overexpression partially rescued the aggressive phenotype of Rab3B-depleted lung adenocarcinoma cells. CONCLUSION: LKB1 deficiency promotes Rab3B upregulation via a CREB-dependent manner. Rab3B interacts with and stabilizes DDX6 protein to accelerate lung adenocarcinoma progression. The Rab3B-DDX6 axis may be potential therapeutic target for lung adenocarcinoma.
Asunto(s)
Adenocarcinoma del Pulmón , ARN Helicasas DEAD-box , Regulación Neoplásica de la Expresión Génica , Neoplasias Pulmonares , Animales , Femenino , Humanos , Masculino , Ratones , Adenocarcinoma del Pulmón/genética , Adenocarcinoma del Pulmón/metabolismo , Adenocarcinoma del Pulmón/patología , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , ARN Helicasas DEAD-box/metabolismo , ARN Helicasas DEAD-box/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Estabilidad ProteicaRESUMEN
The tumor suppressor LKB1 is a serine/threonine protein kinase that is frequently mutated in human lung adenocarcinoma (LUAD). LKB1 regulates a complex signaling network that is known to control cell polarity and metabolism; however, the pathways that mediate the tumor-suppressive activity of LKB1 are incompletely defined. To identify mechanisms of LKB1-mediated growth suppression, we developed a spheroid-based cell culture assay to study LKB1-dependent growth. We then performed genome-wide CRISPR screens in spheroidal culture and found that LKB1 suppresses growth, in part, by activating the PIKFYVE lipid kinase. Finally, we used chemical inhibitors and a pH-sensitive reporter to determine that LKB1 impairs growth by promoting the internalization of wild-type EGFR in a PIKFYVE-dependent manner.
Asunto(s)
Quinasas de la Proteína-Quinasa Activada por el AMP , Fosfatidilinositol 3-Quinasas , Proteínas Serina-Treonina Quinasas , Esferoides Celulares , Humanos , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Quinasas de la Proteína-Quinasa Activada por el AMP/metabolismo , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Esferoides Celulares/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfatidilinositol 3-Quinasas/genética , Proliferación Celular , Línea Celular Tumoral , Sistemas CRISPR-Cas , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/metabolismo , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas/genéticaRESUMEN
The role of the mitochondrial electron transport chain (ETC) in regulating ferroptosis is not fully elucidated. Here, we reveal that pharmacological inhibition of the ETC complex I reduces ubiquinol levels while decreasing ATP levels and activating AMP-activated protein kinase (AMPK), the two effects known for their roles in promoting and suppressing ferroptosis, respectively. Consequently, the impact of complex I inhibitors on ferroptosis induced by glutathione peroxidase 4 (GPX4) inhibition is limited. The pharmacological inhibition of complex I in LKB1-AMPK-inactivated cells, or genetic ablation of complex I (which does not trigger apparent AMPK activation), abrogates the AMPK-mediated ferroptosis-suppressive effect and sensitizes cancer cells to GPX4-inactivation-induced ferroptosis. Furthermore, complex I inhibition synergizes with radiotherapy (RT) to selectively suppress the growth of LKB1-deficient tumors by inducing ferroptosis in mouse models. Our data demonstrate a multifaceted role of complex I in regulating ferroptosis and propose a ferroptosis-inducing therapeutic strategy for LKB1-deficient cancers.
Asunto(s)
Proteínas Quinasas Activadas por AMP , Complejo I de Transporte de Electrón , Ferroptosis , Animales , Femenino , Humanos , Ratones , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Proteínas Quinasas Activadas por AMP/metabolismo , Proteínas Quinasas Activadas por AMP/genética , Línea Celular Tumoral , Complejo I de Transporte de Electrón/metabolismo , Complejo I de Transporte de Electrón/genética , Ferroptosis/genética , Ferroptosis/efectos de los fármacos , Mitocondrias/metabolismo , Mitocondrias/genética , Mitocondrias/efectos de los fármacos , Neoplasias/genética , Neoplasias/patología , Neoplasias/metabolismo , Neoplasias/tratamiento farmacológico , Fosfolípido Hidroperóxido Glutatión Peroxidasa/metabolismo , Fosfolípido Hidroperóxido Glutatión Peroxidasa/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Transducción de Señal , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: This study aimed to systematically analyze the effect of a serine/threonine kinase (STK11) mutation (STK11mut) on therapeutic efficacy and prognosis in patients with non-small cell lung cancer (NSCLC). METHODS: Candidate articles were identified through a search of relevant literature published on or before April 1, 2023, in PubMed, Embase, Cochrane Library, CNKI and Wanfang databases. The extracted and analyzed data included the hazard ratios (HRs) of PFS and OS, the objective response rate (ORR) of immune checkpoint inhibitors (ICIs), and the positive rates of PD-L1 expression. The HR of PFS and OS and the merged ratios were calculated using a meta-analysis. The correlation between STK11mut and clinical characteristics was further analyzed in NSCLC datasets from public databases. RESULTS: Fourteen retrospective studies including 4317 patients with NSCLC of whom 605 had STK11mut were included. The meta-analysis revealed that the ORR of ICIs in patients with STK11mut was 10.1% (95%CI 0.9-25.2), and the positive rate of PD-L1 expression was 41.1% (95%CI 25.3-57.0). STK11mut was associated with poor PFS (HR = 1.49, 95%CI 1.28-1.74) and poor OS (HR = 1.44, 95%CI 1.24-1.67). In the bioinformatics analysis, PFS and OS in patients with STK11 alterations were worse than those in patients without alterations (p < 0.001, p = 0.002). Nutlin-3a, 5-fluorouracil, and vinorelbine may have better sensitivity in patients with STK11mut than in those with STK11wt. CONCLUSIONS: Patients with STK11-mutant NSCLC had low PD-L1 expression and ORR to ICIs, and their PFS and OS were worse than patients with STK11wt after comprehensive treatment. In the future, more reasonable systematic treatments should be explored for this subgroup of patients with STK11-mutant NSCLC.
Asunto(s)
Quinasas de la Proteína-Quinasa Activada por el AMP , Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Humanos , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Antígeno B7-H1 , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Neoplasias Pulmonares/genética , Mutación , Pronóstico , Proteínas Serina-Treonina Quinasas/genética , Estudios RetrospectivosRESUMEN
The tumor suppressor Liver Kinase B1 (LKB1) is a multifunctional serine/threonine protein kinase that regulates cell metabolism, polarity, and growth and is associated with Peutz-Jeghers Syndrome and cancer predisposition. The LKB1 gene comprises 10 exons and 9 introns. Three spliced LKB1 variants have been documented, and they reside mainly in the cytoplasm, although two possess a nuclear-localization sequence (NLS) and are able to shuttle into the nucleus. Here, we report the identification of a fourth and novel LKB1 isoform that is, interestingly, targeted to the mitochondria. We show that this mitochondria-localized LKB1 (mLKB1) is generated from alternative splicing in the 5' region of the transcript and translated from an alternative initiation codon encoded by a previously unknown exon 1b (131 bp) hidden within the long intron 1 of LKB1 gene. We found by replacing the N-terminal NLS of the canonical LKB1 isoform, the N-terminus of the alternatively spliced mLKB1 variant encodes a mitochondrial transit peptide that allows it to localize to the mitochondria. We further demonstrate that mLKB1 colocalizes histologically with mitochondria-resident ATP Synthase and NAD-dependent deacetylase sirtuin-3, mitochondrial (SIRT3) and that its expression is rapidly and transiently upregulated by oxidative stress. We conclude that this novel LKB1 isoform, mLKB1, plays a critical role in regulating mitochondrial metabolic activity and oxidative stress response.
Asunto(s)
Quinasas de la Proteína-Quinasa Activada por el AMP , Mitocondrias , Mutación , Estrés Oxidativo , Proteínas Serina-Treonina Quinasas , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Quinasas de la Proteína-Quinasa Activada por el AMP/metabolismo , Mitocondrias/genética , Mitocondrias/metabolismo , Estrés Oxidativo/genética , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Sirtuina 3/metabolismo , Señales de Clasificación de Proteína , Transporte de Proteínas , ATPasas de Translocación de Protón Mitocondriales/metabolismo , Empalme Alternativo , Codón IniciadorRESUMEN
OBJECTIVE: Intratumoral heterogeneity was found to be a significant factor causing resistance to lung cancer therapies, including immune checkpoint blockade. Lesser is known about spatial heterogeneity of the tumor microenvironment (TME) and its association with genetic properties of the tumor, which is of particular interest in the therapy-naïve setting. MATERIALS AND METHODS: We performed multi-region sampling (2-4 samples per tumor; total of 55 samples) from a cohort of 19 untreated stage IA-IIIB lung adenocarcinomas (n = 11 KRAS mutant, n = 1 ERBB2 mutant, n = 7 KRAS wildtype). For each sample the expression of 770 immunooncology-related genes was analyzed using the nCounter platform, while the mutational status was determined by hybrid capture-based next-generation sequencing (NGS) using a large panel covering more than 500 genes. RESULTS: Global unsupervised analyses revealed clustering of the samples into two groups corresponding to a 'hot' or 'cold' immunologic tumor contexture based on the abundance of immune cell infiltrates. All analyzed specific immune cell signatures (ICsig) showed a significantly higher intertumoral than intratumoral heterogeneity (p < 0.02), as most of the analyzed cases (14/19) showed a very homogenous spatial immune cell profile. PD-L1 exhibited a significantly higher intertumoral than intratumoral heterogeneity (p = 1.03e-13). We found a specific association with 'cold' TME for STK11 (11/14, p < 0.07), but not KRAS, TP53, LRP1B, MTOR, U2AF1 co-mutations, and validated this finding using The Cancer Genome Atlas (TCGA) data. CONCLUSION: Early-stage lung adenocarcinomas show considerable intertumoral, but limited intratumoral heterogeneity, which is clinically highly relevant as assessment before neoadjuvant treatment is based on small biopsies. STK11 mutations are specifically associated with a 'cold' TME, which could affect the efficacy of perioperative immunotherapy.
Asunto(s)
Quinasas de la Proteína-Quinasa Activada por el AMP , Adenocarcinoma del Pulmón , Evasión Inmune , Neoplasias Pulmonares , Adenocarcinoma del Pulmón/genética , Adenocarcinoma del Pulmón/inmunología , Adenocarcinoma del Pulmón/terapia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/inmunología , Neoplasias Pulmonares/terapia , Evasión Inmune/genética , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Microambiente Tumoral/genética , Microambiente Tumoral/inmunología , Humanos , Mutación , Estadificación de NeoplasiasRESUMEN
Contradictory characteristics of elevated mutational burden and a "cold" tumor microenvironment (TME) coexist in liver kinase B1 (LKB1)-mutant non-small cell lung cancers (NSCLC). The molecular basis underlying this paradox and strategies tailored to these historically difficult to treat cancers are lacking. Here, by mapping the single-cell transcriptomic landscape of genetically engineered mouse models with Kras versus Kras/Lkb1-driven lung tumors, we detected impaired tumor-intrinsic IFNγ signaling in Kras/Lkb1-driven tumors that explains the inert immune context. Mechanistic analysis showed that mutant LKB1 led to deficiency in the DNA damage repair process and abnormally activated PARP1. Hyperactivated PARP1 attenuated the IFNγ pathway by physically interacting with and enhancing the poly(ADP-ribosyl)ation of STAT1, compromising its phosphorylation and activation. Abrogation of the PARP1-driven program triggered synthetic lethality in NSCLC on the basis of the LKB1 mutation-mediated DNA repair defect, while also restoring phosphorylated STAT1 to favor an immunologically "hot" TME. Accordingly, PARP1 inhibition restored the disrupted IFNγ signaling and thus mounted an adaptive immune response to synergize with PD-1 blockade in multiple LKB1-deficient murine tumor models. Overall, this study reveals an unexplored interplay between the DNA repair process and adaptive immune response, providing a molecular basis for dual PARP1 and PD-1 inhibition in treating LKB1-mutant NSCLC. SIGNIFICANCE: Targeting PARP exerts dual effects to overcome LKB1 loss-driven immunotherapy resistance through triggering DNA damage and adaptive immunity, providing a rationale for dual PARP and PD-1 inhibition in treating LKB1-mutant lung cancers.
Asunto(s)
Inmunidad Adaptativa , Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Inhibidores de Poli(ADP-Ribosa) Polimerasas , Animales , Ratones , Inmunidad Adaptativa/efectos de los fármacos , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Receptor de Muerte Celular Programada 1/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Mutaciones Letales Sintéticas/efectos de los fármacos , Microambiente Tumoral , Quinasas de la Proteína-Quinasa Activada por el AMP/genéticaRESUMEN
The increasing morbidity and high mortality of intrahepatic cholangiocarcinoma (ICC) has led to the urgent need for new diagnostics and therapeutics. Liver kinase B1 (LKB1) exerts a tumor suppressor role in multiple malignances, while its regulatory role in exosomes secreted by ICC cells is obscure. In the present study, exosomes were extracted from cell culture supernatants of RBE and HCCC9810 ICC cells as well as plasma of patients with ICC by ultracentrifugation and the morphology of exosomes was identified by transmission electron microscopy. Notably, compared with that of intracellular LKB1, the protein level of exosomal LKB1 was decreased. Silencing intracellular LKB1 increased the protein levels of programmed death ligand 1 (PDL1), Slug and phosphorylatedAKT in exosomes, accompanied by decreased expression levels of exosomal LKB1. Exosomes with lower protein levels of LKB1 promoted the expression of the immune checkpoint PDL1, malignant phenotypes of ICC cells in vitro, and cancer metastasis in vivo. Moreover, the low level of exosomal LKB1 in plasma was tightly associated with the poor prognosis of patients with ICC. Collectively, exosomal LKB1 inhibits the immune checkpoint PDL1 and metastasis of ICC cells. These findings may provide new methods for the diagnosis and immune therapy of ICC.
Asunto(s)
Quinasas de la Proteína-Quinasa Activada por el AMP , Neoplasias de los Conductos Biliares , Colangiocarcinoma , Exosomas , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Quinasas de la Proteína-Quinasa Activada por el AMP/metabolismo , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Neoplasias de los Conductos Biliares/patología , Conductos Biliares Intrahepáticos/patología , Línea Celular Tumoral , Colangiocarcinoma/patología , Exosomas/metabolismo , HumanosRESUMEN
This study was aimed at exploring the mechanism of serine threonine protein kinase 11 (STK11)/Adenosine 5'-monophosphate-activated protein kinase (AMPK) signaling pathway after immunotherapy for esophageal squamous cell carcinoma (ESCC), providing basic information for the clinical treatment of ESCC. In this study, tissue specimens from 100 patients with ESCC who underwent surgical treatment in Taizhou People's Hospital (group A) and 20 patients with recurrent or metastatic ESCC who received second-line immunotherapy (group B) were collected. The real-time fluorescent quantitative polymerase chain reaction (PCR) (RT-qPCR) technology was used to detect the expression levels of STK11, interferon-γ (IFN-γ), interleukin 6 (IL-6), and vascular endothelial growth factor (VEGF) in the tissues. The immunohistochemical staining was used to detect the positive expression levels (PELs) of STK11 and AMPKα in the tissues, and immunofluorescence staining was used to detect the PELs Teff cells (CD3 and CD8), Treg cells (CD4 and FOXP3), and neutrophils (CD68 and CD163). RT-qPCR results showed that the expression levels of STK11 and IFN-γ in group A were obviously lower, and those of IL-6 and VEGF were much higher in contrast to group B (P < 0.05). The results of immunohistochemical staining showed that the number of STK11- and AMPKα-positive staining cells in group A was dramatically less than that in group B (P <0.05). The results of immunofluorescence staining revealed that the number of positive staining cells for Teff cells, Treg cells, and neutrophils in group A was also less dramatically than that in group B (P <0.05). In summary, immunotherapy can play a therapeutic effect on ESCC by regulating STK11/AMPK pathway and immune cell infiltration.
Asunto(s)
Quinasas de la Proteína-Quinasa Activada por el AMP , Proteínas Quinasas Activadas por AMP , Neoplasias Esofágicas , Carcinoma de Células Escamosas de Esófago , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Quinasas de la Proteína-Quinasa Activada por el AMP/inmunología , Proteínas Quinasas Activadas por AMP/genética , Proteínas Quinasas Activadas por AMP/inmunología , Línea Celular Tumoral , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/inmunología , Neoplasias Esofágicas/cirugía , Neoplasias Esofágicas/terapia , Carcinoma de Células Escamosas de Esófago/genética , Carcinoma de Células Escamosas de Esófago/inmunología , Carcinoma de Células Escamosas de Esófago/cirugía , Carcinoma de Células Escamosas de Esófago/terapia , Regulación Neoplásica de la Expresión Génica , Humanos , Inmunoterapia/métodos , Interleucina-6/inmunología , Proteínas Serina-Treonina Quinasas/genética , Transducción de Señal , Factor A de Crecimiento Endotelial Vascular/inmunologíaRESUMEN
High-grade serous ovarian cancer (HGSOC) is an aggressive malignancy often diagnosed at an advanced stage. Although most HGSOC patients respond initially to debulking surgery combined with cytotoxic chemotherapy, many ultimately relapse with platinum-resistant disease. Thus, improving outcomes requires new ways of limiting metastasis and eradicating residual disease. We identified previously that Liver kinase B1 (LKB1) and its substrate NUAK1 are implicated in EOC spheroid cell viability and are required for efficient metastasis in orthotopic mouse models. Here, we sought to identify additional signalling pathways altered in EOC cells due to LKB1 or NUAK1 loss-of-function. Transcriptome analysis revealed that inflammatory signalling mediated by NF-κB transcription factors is hyperactive due to LKB1-NUAK1 loss in HGSOC cells and spheroids. Upregulated NF-κB signalling due to NUAK1 loss suppresses reactive oxygen species (ROS) production and sustains cell survival in spheroids. NF-κB signalling is also activated in HGSOC precursor fallopian tube secretory epithelial cell spheroids, and is further enhanced by NUAK1 loss. Finally, immunohistochemical analysis of OVCAR8 xenograft tumors lacking NUAK1 displayed increased RelB expression and nuclear staining. Our results support the idea that NUAK1 and NF-κB signalling pathways together regulate ROS and inflammatory signalling, supporting cell survival during each step of HGSOC pathogenesis. We propose that their combined inhibition may be efficacious as a novel therapeutic strategy for advanced HGSOC.
Asunto(s)
Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Quinasas de la Proteína-Quinasa Activada por el AMP/fisiología , Mutación con Pérdida de Función , FN-kappa B/metabolismo , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Proteínas Quinasas/genética , Proteínas Quinasas/fisiología , Proteínas Represoras/genética , Proteínas Represoras/fisiología , Transducción de Señal/genética , Transducción de Señal/fisiología , Esferoides Celulares , Animales , Supervivencia Celular , Modelos Animales de Enfermedad , Femenino , Perfilación de la Expresión Génica , Humanos , Ratones , Terapia Molecular Dirigida , Trasplante de Neoplasias , Neoplasias Ováricas/patología , Neoplasias Ováricas/terapia , Especies Reactivas de Oxígeno/metabolismo , Esferoides Celulares/metabolismo , Esferoides Celulares/fisiología , Transcriptoma/genética , Células Tumorales CultivadasRESUMEN
In the bone marrow, classical and plasmacytoid dendritic cells (DC) develop from the macrophage-DC precursor (MDP) through a common DC precursor (CDP) step. This developmental process receives essential input from the niche in which it takes place, containing endothelial cells (EC) among other cell types. Here we show that targeted deletion of serine/threonine kinase 11 (Stk11) encoding tumor suppressor liver kinase b1 (Lkb1) in mouse ECs but not DCs, results in disrupted differentiation of MDPs to CDPs, severe reduction in mature DC numbers and spontaneous tumorigenesis. In wild type ECs, Lkb1 phosphorylates polypyrimidine tract binding protein 1 (Ptbp1) at threonine 138, which regulates stem cell factor (Scf) pre-mRNA splicing. In the absence of Lkb1, exon 6 of Scf is spliced out, leading to the loss of Scf secretion. Adeno-associated-virus-mediated delivery of genes encoding either soluble Scf or the phosphomimetic mutant Ptbp1T138E proteins rescued the defects of MDP to CDP differentiation and DC shortage in the endothelium specific Stk11 knockout mice. In summary, endothelial Stk11 expression regulates DC differentiation via modulation of Scf splicing, marking the Stk11-soluble-Scf axis as a potential cause of DC deficiency syndromes.
Asunto(s)
Proteínas Quinasas Activadas por AMP/genética , Diferenciación Celular/genética , Transformación Celular Neoplásica/genética , Células Dendríticas/metabolismo , Células Endoteliales/metabolismo , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Quinasas de la Proteína-Quinasa Activada por el AMP/metabolismo , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Western Blotting , Células de la Médula Ósea/metabolismo , Supervivencia Celular/genética , Células Cultivadas , Técnica del Anticuerpo Fluorescente , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Humanos , Ratones Noqueados , Ratones Transgénicos , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factor de Células Madre/genética , Factor de Células Madre/metabolismoRESUMEN
High-grade serous ovarian carcinoma (HGSOC) is the most aggressive gynecological malignancy, resulting in approximately 70% of ovarian cancer deaths. However, it is still unclear how genetic dysregulations and biological processes generate the malignant subtype of HGSOC. Here we show that expression levels of microtubule affinity-regulating kinase 3 (MARK3) are downregulated in HGSOC, and that its downregulation significantly correlates with poor prognosis in HGSOC patients. MARK3 overexpression suppresses cell proliferation and angiogenesis of ovarian cancer cells. The LKB1-MARK3 axis is activated by metabolic stress, which leads to the phosphorylation of CDC25B and CDC25C, followed by induction of G2/M phase arrest. RNA-seq and ATAC-seq analyses indicate that MARK3 attenuates cell cycle progression and angiogenesis partly through downregulation of AP-1 and Hippo signaling target genes. The synthetic lethal therapy using metabolic stress inducers may be a promising therapeutic choice to treat the LKB1-MARK3 axis-dysregulated HGSOCs.
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Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Genes Supresores de Tumor , Neoplasias Ováricas , Proteínas Serina-Treonina Quinasas/genética , Estrés Fisiológico/genética , Biomarcadores de Tumor/genética , Línea Celular Tumoral , Proliferación Celular/genética , Regulación hacia Abajo/genética , Epigénesis Genética/genética , Femenino , Humanos , Neoplasias Ováricas/genética , Neoplasias Ováricas/patologíaRESUMEN
Nonalcoholic fatty liver disease (NAFLD) is a metabolic liver disease with a complex etiology, and is considered as one of the main causes of hepatocellular carcinoma (HCC). The incidence of NAFLD has presented an increasing trend annually as a result of disequilibrium in the dietary structure. However, no specific treatment has been approved for clinical therapy in NAFLD. Ginsenoside CK has been investigated given its various pharmacological activities, but its effects against NAFLD and the underlying mechanism are still unclear. In this study, fructose was used to simulate hepatic fatty degeneration in vivo, while palmitic acid (PA) and oleic acid (OA) were applied to induce lipid accumulation in vitro. The level of lipid accumulation in hepatic tissue and HepG2 cells was evaluated by Oil Red O staining. Detection of serum and liver biomarkers, western blotting, and real-time qPCR were conducted to assess the degree of hepatic steatosis. Our results indicated that ginsenoside CK could decrease the lipid deposition in HepG2 cells, retard the increase of body weight of fructose-fed mice, alleviate the lipid accumulation in serum and hepatic tissue and improve the hepatic inflammation and injury. Mechanically, ginsenoside CK modulated the expression of factors correlated with lipid synthesis and metabolism in vitro and in vivo via activating the phosphorylation of LKB1 and AMPK. Compound C, an inhibitor of AMPK, partially abrogated the beneficial effects of ginsenoside CK in HepG2 cells. In summary, ginsenoside CK acts as a LKB1/AMPK agonist to regulate the lipid metabolism and interfere with the progression of NAFLD.
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Quinasas de la Proteína-Quinasa Activada por el AMP/metabolismo , Proteínas Quinasas Activadas por AMP/metabolismo , Ginsenósidos/metabolismo , Hígado/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Enfermedad del Hígado Graso no Alcohólico/prevención & control , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Animales , Masculino , Ratones , Enfermedad del Hígado Graso no Alcohólico/genética , Transducción de SeñalRESUMEN
In non-small-cell lung cancer (NSCLC), concurrent mutations in the oncogene KRAS and tumor suppressor STK11 (also known as LKB1) confer an aggressive malignant phenotype, an unfavourability towards immunotherapy, and overall poor prognoses in patients. In a previous study, we showed that murine KRAS/LKB1 co-mutant tumors and human co-mutant cancer cells have an enhanced dependence on glutamine-fructose-6-phosphate transaminase 2 (GFPT2), a rate-limiting enzyme in the hexosamine biosynthesis pathway (HBP), which could be targeted to reduce survival of KRAS/LKB1 co-mutants. Here, we found that KRAS/LKB1 co-mutant cells also exhibit an increased dependence on N-acetylglucosamine-phosphate mutase 3 (PGM3), an enzyme downstream of GFPT2. Genetic or pharmacologic suppression of PGM3 reduced KRAS/LKB1 co-mutant tumor growth in both in vitro and in vivo settings. Our results define an additional metabolic vulnerability in KRAS/LKB1 co-mutant tumors to the HBP and provide a rationale for targeting PGM3 in this aggressive subtype of NSCLC.
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Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Neoplasias Pulmonares/genética , Terapia Molecular Dirigida , Fosfoglucomutasa/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/genética , Animales , Vías Biosintéticas/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Glutamina-Fructosa-6-Fosfato Transaminasa (Isomerizadora)/antagonistas & inhibidores , Glutamina-Fructosa-6-Fosfato Transaminasa (Isomerizadora)/metabolismo , Glicosilación/efectos de los fármacos , Hexosaminas/biosíntesis , Humanos , Neoplasias Pulmonares/patología , Ratones , Fosfoglucomutasa/antagonistas & inhibidores , Fosfoglucomutasa/genéticaRESUMEN
Previous studies showed that depleting Liver Kinase-B1 (LKB1) from astrocytes increased inflammatory factors lipocalin-2 (LCN2) and osteopontin (OPN) in EAE. A single nucleotide polymorphism (SNP) in STK11 (encoding LKB1) is a risk factor for MS, suggesting increased LCN2 or OPN contributes to risk. Serum LCN2 and OPN levels in African American female MS patients were higher than healthy controls, and while levels increased with disease duration in cases without the SNP, levels decreased with duration in cases with the SNP. Increased MS risk associated with the STK11 SNP may be due to higher LCN2 or OPN levels at early times.
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Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Lipocalina 2/sangre , Esclerosis Múltiple Recurrente-Remitente/sangre , Esclerosis Múltiple Recurrente-Remitente/genética , Adulto , Negro o Afroamericano/genética , Biomarcadores/sangre , Progresión de la Enfermedad , Femenino , Predisposición Genética a la Enfermedad/genética , Humanos , Persona de Mediana Edad , Osteopontina/sangre , Polimorfismo de Nucleótido SimpleRESUMEN
BACKGROUND/AIM: Pancreatic cancer is one of the most devastating malignancies worldwide. Because of the disappointing outcome of traditional treatment, new drug candidates are being investigated. This study analysed the effect of eupatilin on pancreatic cancer cells. MATERIALS AND METHODS: Cell viability assay, western blot, siRNA transfection, 2-deoxyglucose uptake assay, AMP/ADP/ATP assay, and fluorescent activated cell sorting were performed. RESULTS: Eupatilin decreased cell viability and activated AMPK in MIA-PaCa2 cells. Eupatilin decreased glucose uptake in pancreatic cancer, which led to cell starvation and AMPK activation. It is well known that AMPK induces p21 and cell cycle arrest by activating p53. In MIA-PaCa2 cells, p53 is mutated and wild-type p53 protein is suppressed. Treatment with eupatilin induced p21 expression but inhibited the expression of mutated p53. Eupatilin activated Tap73, a p53 family member, which can substitute wild-type p53's role. CONCLUSION: Eupatilin shows an anticancer effect against pancreatic cancer cells via glucose uptake inhibition, AMPK activation, and cell cycle arrest.
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Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Flavonoides/farmacología , Neoplasias Pancreáticas/tratamiento farmacológico , Proteína Tumoral p73/genética , Apoptosis/efectos de los fármacos , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Glucosa/metabolismo , Humanos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patología , Activación Transcripcional/efectos de los fármacos , Proteína p53 Supresora de Tumor/genéticaRESUMEN
OBJECTIVES: To report an atypical presentation of a pathogenic STK11 gene variant in siblings not fulfilling the clinical diagnostic criteria for Peutz-Jeghers Syndrome (PJS). CASE PRESENTATION: Two siblings presented with prepubertal gynaecomastia and bilateral macro-orchidism, without mucocutaneous pigmentation or gastrointestinal symptoms. There was no family history of PJS. Sibling 1 had unilateral gynaecomastia. Sibling 2 had bilateral gynaecomastia, advanced bone age and bilateral testicular microlithiasis, not indicative of a large-cell calcifying Sertoli cell tumour. Genetics revealed a paternally inherited heterozygous pathogenic STK11 variant (910C>T) in both siblings. The diagnosis was confirmed following the identification of multiple intestinal polyps in their father. CONCLUSIONS: Prepubertal gynaecomastia and prepubertal macro-orchidism (testicular enlargement without virilisation), always warrant endocrinological investigation, with PJS being an important differential diagnosis. Children may not fulfil the clinical criteria for a diagnosis of PJS at presentation. Genetic testing and gastroenterological investigation of parents may aid diagnosis.
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Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Síndrome de Peutz-Jeghers/diagnóstico , Niño , Preescolar , Ginecomastia/diagnóstico , Humanos , Masculino , Síndrome de Peutz-Jeghers/genética , Hermanos , Testículo/patologíaRESUMEN
INTRODUCTION: STK11 and KEAP1 mutations (STK11 mutant [STK11MUT] and KEAP1MUT) are among the most often mutated genes in lung adenocarcinoma (LUAD). Although STK11MUT has been associated with resistance to programmed death-(ligand)1 (PD-[L]1) inhibition in KRASMUT LUAD, its impact on immunotherapy efficacy in KRAS wild-type (KRASWT) LUAD is currently unknown. Whether KEAP1MUT differentially affects outcomes to PD-(L)1 inhibition in KRASMUT and KRASWT LUAD is also unknown. METHODS: Clinicopathologic and genomic data were collected from September 2013 to September 2020 from patients with advanced LUAD at the Dana-Farber Cancer Institute/Massachusetts General Hospital cohort and the Memorial Sloan Kettering Cancer Center/MD Anderson Cancer Center cohort. Clinical outcomes to PD-(L)1 inhibition were analyzed according to KRAS, STK11, and KEAP1 mutation status in two independent cohorts. The Cancer Genome Atlas transcriptomic data were interrogated to identify differences in tumor gene expression and tumor immune cell subsets, respectively, according to KRAS/STK11 and KRAS/KEAP1 comutation status. RESULTS: In the combined cohort (Dana-Farber Cancer Institute/Massachusetts General Hospital + Memorial Sloan Kettering Cancer Center/MD Anderson Cancer Center) of 1261 patients (median age = 61 y [range: 22-92], 708 women [56.1%], 1065 smokers [84.4%]), KRAS mutations were detected in 536 cases (42.5%), and deleterious STK11 and KEAP1 mutations were found in 20.6% (260 of 1261) and 19.2% (231 of 1202) of assessable cases, respectively. In each independent cohort and in the combined cohort, STK11 and KEAP1 mutations were associated with significantly worse progression-free (STK11 hazard ratio [HR] = 2.04, p < 0.0001; KEAP1 HR = 2.05, p < 0.0001) and overall (STK11 HR = 2.09, p < 0.0001; KEAP1 HR = 2.24, p < 0.0001) survival to immunotherapy uniquely among KRASMUT but not KRASWT LUADs. Gene expression ontology and immune cell enrichment analyses revealed that the presence of STK11 or KEAP1 mutations results in distinct immunophenotypes in KRASMUT, but not in KRASWT, lung cancers. CONCLUSIONS: STK11 and KEAP1 mutations confer worse outcomes to immunotherapy among patients with KRASMUT but not among KRASWT LUAD. Tumors harboring concurrent KRAS/STK11 and KRAS/KEAP1 mutations display distinct immune profiles in terms of gene expression and immune cell infiltration.