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BACKGROUND: We investigated the role of A2B-adenosine receptor in regulating immunosuppressive metabolic stress in the tumor microenvironment. Novel A2B-adenosine receptor antagonist PBF-1129 was tested for antitumor activity in mice and evaluated for safety and immunologic efficacy in a phase I clinical trial of patients with non-small cell lung cancer. METHODS: The antitumor efficacy of A2B-adenosine receptor antagonists and their impact on the metabolic and immune tumor microenvironment were evaluated in lung, melanoma, colon, breast, and epidermal growth factor receptor-inducible transgenic cancer models. Employing electron paramagnetic resonance, we assessed changes in tumor microenvironment metabolic parameters, including pO2, pH, and inorganic phosphate, during tumor growth and evaluated the immunologic effects of PBF-1129, including its pharmacokinetics, safety, and toxicity, in patients with non-small cell lung cancer. RESULTS: Levels of metabolic stress correlated with tumor growth, metastasis, and immunosuppression. Tumor interstitial inorganic phosphate emerged as a correlative and cumulative measure of tumor microenvironment stress and immunosuppression. A2B-adenosine receptor inhibition alleviated metabolic stress, downregulated expression of adenosine-generating ectonucleotidases, increased expression of adenosine deaminase, decreased tumor growth and metastasis, increased interferon γ production, and enhanced the efficacy of antitumor therapies following combination regimens in animal models (anti-programmed cell death 1 protein vs anti-programmed cell death 1 protein plus PBF-1129 treatment hazard ratio = 11.74 [95% confidence interval = 3.35 to 41.13], n = 10, P < .001, 2-sided F test). In patients with non-small cell lung cancer, PBF-1129 was well tolerated, with no dose-limiting toxicities; demonstrated pharmacologic efficacy; modulated the adenosine generation system; and improved antitumor immunity. CONCLUSIONS: Data identify A2B-adenosine receptor as a valuable therapeutic target to modify metabolic and immune tumor microenvironment to reduce immunosuppression, enhance the efficacy of immunotherapies, and support clinical application of PBF-1129 in combination therapies.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Humanos , Animales , Ratones , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Receptor de Adenosina A2B/metabolismo , Microambiente Tumoral , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/patología , Terapia de Inmunosupresión , Adenosina/metabolismo , Fosfatos , Línea Celular TumoralRESUMEN
OBJECTIVES: To describe characteristics, treatment and outcomes of non-small cell lung cancer (NSCLC) patients with MET alterations (MET exon 14 [METex14] skipping or MET amplification [METamp]) in real-world clinical care. METHODS: This non-interventional cohort study used real-world data extracted from electronic medical records from academic oncology sites in Israel, The Netherlands, Taiwan, and the USA. Patients had confirmed diagnosis of advanced (Stage IIIB-IV) NSCLC harboring MET alterations (date of diagnosis = index date) between 1 Jan 2010 and 30 Sept 2018. Medical history was assessed prior to and at the index date (baseline period), and outcomes from first date of treatment to death, loss to follow-up, or end of study period. RESULTS: A total of 117 patients were included (METex14 n = 70; METamp n = 47); testing methods were heterogeneous. Concomitant oncogenic mutations were more common in the METamp cohort than METex14. Patients in the METex14 cohort were older than those in METamp, and a larger proportion were never smokers. Anticancer first-line therapies received by patients (METex14; METamp) included chemotherapy only (44%; 41%), MET inhibitors (33%; 29%), immune checkpoint inhibitor (ICI) mono-(12%; 15%) and combination-therapy (8%; 3%). Second-line therapies included chemotherapy (35%; 30%) and MET inhibitors (30%; 39%). In the METex14 cohort, objective response rate (ORR) was generally low (first-line 28%; second-line 30%); no patients who received ICIs had a response. In the METamp cohort, ORR was 36% in first-line and 22% in second-line. Median (95% confidence interval) overall survival from start of first-line therapy was 12.0 months (6.8, 19.2) in the METex14 cohort and 22.0 months (9.8, 31.2) in METamp. CONCLUSIONS: Heterogeneous treatments reflect the changing landscape and availability of new treatments, as well as the high unmet medical need in older, METex14 patients who had more advanced disease at diagnosis. MET-targeted therapies could be beneficial in patients with these rare MET alterations.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Anciano , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/epidemiología , Carcinoma de Pulmón de Células no Pequeñas/genética , Estudios de Cohortes , Exones , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/epidemiología , Neoplasias Pulmonares/genética , MutaciónRESUMEN
STK11 (liver kinase B1, LKB1) is the fourth most frequently mutated gene in lung adenocarcinoma, with loss of function observed in up to 30% of all cases. Our previous work identified a 16-gene signature for LKB1 loss of function through mutational and nonmutational mechanisms. In this study, we applied this genetic signature to The Cancer Genome Atlas (TCGA) lung adenocarcinoma samples and discovered a novel association between LKB1 loss and widespread DNA demethylation. LKB1-deficient tumors showed depletion of S-adenosyl-methionine (SAM-e), which is the primary substrate for DNMT1 activity. Lower methylation following LKB1 loss involved repetitive elements (RE) and altered RE transcription, as well as decreased sensitivity to azacytidine. Demethylated CpGs were enriched for FOXA family consensus binding sites, and nuclear expression, localization, and turnover of FOXA was dependent upon LKB1. Overall, these findings demonstrate that a large number of lung adenocarcinomas exhibit global hypomethylation driven by LKB1 loss, which has implications for both epigenetic therapy and immunotherapy in these cancers. SIGNIFICANCE: Lung adenocarcinomas with LKB1 loss demonstrate global genomic hypomethylation associated with depletion of SAM-e, reduced expression of DNMT1, and increased transcription of repetitive elements.
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Quinasas de la Proteína-Quinasa Activada por el AMP/fisiología , Adenocarcinoma/genética , Metilación de ADN , Neoplasias Pulmonares/genética , S-Adenosilmetionina/metabolismo , Quinasas de la Proteína-Quinasa Activada por el AMP/genética , Adenocarcinoma/metabolismo , Línea Celular , Supervivencia Celular , Análisis por Conglomerados , Biología Computacional , Islas de CpG , Bases de Datos Genéticas , Epigénesis Genética , Genes ras , Humanos , Neoplasias Pulmonares/metabolismo , Metionina , Mutación , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteínas Proto-Oncogénicas p21(ras)/genética , Secuencias Repetitivas de Ácidos NucleicosRESUMEN
We have identified a non-canonical role of0 Notch3 in response to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) therapy, whereby Notch3 associates with ß-catenin, resulting in increased catenin beta-1 (CTNNB1, best known as ß-catenin) stability and increased survival of drug persister cells (DPCs). Furthermore, combined treatment of an EGFR TKI with a ß-catenin inhibitor demonstrated improved therapeutic outcomes in xenograft models.
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EGFR tyrosine kinase inhibitors cause dramatic responses in EGFR-mutant lung cancer, but resistance universally develops. The involvement of ß-catenin in EGFR TKI resistance has been previously reported, however, the precise mechanism by which ß-catenin activation contributes to EGFR TKI resistance is not clear. Here, we show that EGFR inhibition results in the activation of ß-catenin signaling in a Notch3-dependent manner, which facilitates the survival of a subset of cells that we call "adaptive persisters". We previously reported that EGFR-TKI treatment rapidly activates Notch3, and here we describe the physical association of Notch3 with ß-catenin, leading to increased stability and activation of ß-catenin. We demonstrate that the combination of EGFR-TKI and a ß-catenin inhibitor inhibits the development of these adaptive persisters, decreases tumor burden, improves recurrence free survival, and overall survival in xenograft models. These results supports combined EGFR-TKI and ß-catenin inhibition in patients with EGFR mutant lung cancer.
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Carcinoma de Pulmón de Células no Pequeñas/genética , Neoplasias Pulmonares/genética , Mutación/genética , Inhibidores de Proteínas Quinasas/farmacología , Receptor Notch3/metabolismo , Transducción de Señal , beta Catenina/metabolismo , Animales , Carcinoma de Pulmón de Células no Pequeñas/sangre , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Proteínas de Unión al ADN/metabolismo , Resistencia a Antineoplásicos/efectos de los fármacos , Transición Epitelial-Mesenquimal/efectos de los fármacos , Receptores ErbB/genética , Humanos , Neoplasias Pulmonares/sangre , Neoplasias Pulmonares/patología , Ratones Endogámicos NOD , Ratones SCID , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Fenotipo , Inhibidor 1 de Activador Plasminogénico/sangre , Estabilidad Proteica/efectos de los fármacos , Factores de Transcripción/metabolismo , beta Catenina/antagonistas & inhibidoresRESUMEN
Mutations in the epidermal growth factor receptor (EGFR) are the most common actionable genetic abnormalities yet discovered in lung cancer. However, targeting these mutations with kinase inhibitors is not curative in advanced disease and has yet to demonstrate an impact on potentially curable, early-stage disease, with some data suggesting adverse outcomes. Here, we report that treatment of EGFR-mutated lung cancer cell lines with erlotinib, while showing robust cell death, enriches the ALDH(+) stem-like cells through EGFR-dependent activation of Notch3. In addition, we demonstrate that erlotinib treatment increases the clonogenicity of lung cancer cells in a sphere-forming assay, suggesting increased stem-like cell potential. We demonstrate that inhibition of EGFR kinase activity leads to activation of Notch transcriptional targets in a γ secretase inhibitor-sensitive manner and causes Notch activation, leading to an increase in ALDH high(+) cells. We also find a kinase-dependent physical association between the Notch3 and EGFR receptors and tyrosine phosphorylation of Notch3. This could explain the worsened survival observed in some studies of erlotinib treatment at early-stage disease, and suggests that specific dual targeting might overcome this adverse effect.
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Receptores ErbB/antagonistas & inhibidores , Neoplasias Pulmonares/patología , Células Madre Neoplásicas/patología , Receptores Notch/metabolismo , Transducción de Señal , Línea Celular Tumoral , Receptores ErbB/metabolismo , Clorhidrato de Erlotinib , Humanos , Neoplasias Pulmonares/metabolismo , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Quinazolinas/farmacología , Receptor Notch3 , Transcripción GenéticaRESUMEN
INTRODUCTION: Inactivation of serine/threonine kinase 11 (STK11 or LKB1) is common in lung cancer, and understanding the pathways and phenotypes altered as a consequence will aid the development of targeted therapeutic strategies. Gene and protein expressions in a murine model of v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (Kras)-mutant lung cancer have been studied to gain insight into the biology of these tumors. However, the molecular consequences of LKB1 loss in human lung cancer have not been fully characterized. METHODS: We studied gene expression profiles associated with LKB1 loss in resected lung adenocarcinomas, non-small-cell lung cancer cell lines, and murine tumors. The biological significance of dysregulated genes was interpreted using gene set enrichment and transcription factor analyses and also by integration with somatic mutations and proteomic data. RESULTS: Loss of LKB1 is associated with consistent gene expression changes in resected human lung cancers and cell lines that differ substantially from the mouse model. Our analysis implicates novel biological features associated with LKB1 loss, including altered mitochondrial metabolism, activation of the nuclear respiratory factor 2 (NRF2) transcription factor by kelch-like ECH-associated protein 1 (KEAP1) mutations, and attenuation of the phosphatidylinositiol 3-kinase and v-akt murine thymoma viral oncogene homolog (PI3K/AKT) pathway. Furthermore, we derived a 16-gene classifier that accurately predicts LKB1 mutations and loss by nonmutational mechanisms. In vitro, transduction of LKB1 into LKB1-mutant cell lines results in attenuation of this signature. CONCLUSION: Loss of LKB1 defines a subset of lung adenocarcinomas associated with characteristic molecular phenotypes and distinctive gene expression features. Studying these effects may improve our understanding of the biology of these tumors and lead to the identification of targeted treatment strategies.
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Adenocarcinoma/genética , Carcinoma de Pulmón de Células no Pequeñas/genética , Neoplasias Pulmonares/genética , Factor 2 Relacionado con NF-E2/metabolismo , Fosfatidilinositol 3-Quinasa/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transcriptoma , Quinasas de la Proteína-Quinasa Activada por el AMP , Proteínas Quinasas Activadas por AMP/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Proteínas de la Ataxia Telangiectasia Mutada/genética , Línea Celular Tumoral , Proteínas del Citoesqueleto/genética , Receptores ErbB/genética , Factor de Transcripción de la Proteína de Unión a GA/metabolismo , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Proteína 1 Asociada A ECH Tipo Kelch , Ratones , Mitocondrias/metabolismo , Familia de Multigenes , Mutación , Fosforilación , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas p21(ras) , ARN Mensajero/metabolismo , Transducción de Señal , Proteína p53 Supresora de Tumor/genética , Proteínas ras/genéticaRESUMEN
Heregulin activation of the endogenous receptor tyrosine kinase ErbB-4 in ZR-75-1 breast cancer cells provokes tyrosine phosphorylation of Hdm2 in a manner that is sensitive to inhibition of alpha- or gamma-secretase activity, indicating that liberation of the tyrosine kinase intracellular domain (ICD) fragment is required. Similar results are obtained when Erbb-4 is exogenously expressed in 32D cells, which do not otherwise express any ErbB family members. Expression of the ErbB-4 ICD fragment leads to its constitutive association with Mdm2 and tyrosine phosphorylation of Mdm2, a protein that is predominantly localized in the nucleus and that regulates p53 levels. When the ErbB-4 ICD fragment was expressed in H1299 cells, it promoted Hdm2 ubiquitination and increased the levels of p53 and p21, a transcriptional target of p53. In addition, expression of the ICD fragment increased p53 activity toward the p21 promoter in a luciferase reporter assay.