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Patients with cancer of unknown primary (CUP) carry the double burden of an aggressive disease and reduced access to therapies. Experimental models are pivotal for CUP biology investigation and drug testing. We derived two CUP cell lines (CUP#55 and #96) and corresponding patient-derived xenografts (PDXs), from ascites tumor cells. CUP cell lines and PDXs underwent histological, immune-phenotypical, molecular, and genomic characterization confirming the features of the original tumor. The tissue-of-origin prediction was obtained from the tumor microRNA expression profile and confirmed by single-cell transcriptomics. Genomic testing and fluorescence in situ hybridization analysis identified FGFR2 gene amplification in both models, in the form of homogeneously staining region (HSR) in CUP#55 and double minutes in CUP#96. FGFR2 was recognized as the main oncogenic driver and therapeutic target. FGFR2-targeting drug BGJ398 (infigratinib) in combination with the MEK inhibitor trametinib proved to be synergic and exceptionally active, both in vitro and in vivo. The effects of the combined treatment by single-cell gene expression analysis revealed a remarkable plasticity of tumor cells and the greater sensitivity of cells with epithelial phenotype. This study brings personalized therapy closer to CUP patients and provides the rationale for FGFR2 and MEK targeting in metastatic tumors with FGFR2 pathway activation.
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Background: Malignant pleural mesothelioma (MPM) is an aggressive malignancy associated to asbestos exposure. One of the most frequent genetic alteration in MPM patients is CDKN2A/ARF loss, leading to aberrant activation of the Rb pathway. In MPM cells, we previously demonstrated the therapeutic efficacy of targeting this signaling with the CDK4/6 inhibitor palbociclib in combination with PI3K/mTOR inhibitors. Here, we investigated whether such combination may have an impact on cell energy metabolism. Methods: The study was performed in MPM cells of different histotypes; metabolic analyses were conducted by measuring GLUT-1 expression and glucose uptake/consumption, and by SeaHorse technologies. Results: MPM cell models differed for their ability to adapt to metabolic stress conditions, such as glucose starvation and hypoxia. Independently of these differences, combined treatments with palbociclib and PI3K/mTOR inhibitors inhibited cell proliferation more efficaciously than single agents. The drugs alone reduced glucose uptake/consumption as well as glycolysis, and their combination further enhanced these effects under both normoxic and hypoxic conditions. Moreover, the drug combinations significantly impaired mitochondrial respiration as compared with individual treatments. These metabolic effects were mediated by the concomitant inhibition of Rb/E2F/c-myc and PI3K/AKT/mTOR signaling. Conclusions: Dual blockade of glycolysis and respiration contributes to the anti-tumor efficacy of palbociclib-PI3K/mTOR inhibitors combination.
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Proliferação de Células/efeitos dos fármacos , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Metabolismo Energético/efeitos dos fármacos , Mesotelioma Maligno/metabolismo , Neoplasias Pleurais/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Linhagem Celular Tumoral , Glicólise/efeitos dos fármacos , Humanos , Mesotelioma Maligno/tratamento farmacológico , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase/farmacologia , Piperazinas/farmacologia , Neoplasias Pleurais/tratamento farmacológico , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Piridinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismoRESUMO
The importance of the immune system as a potent anti-tumor defense has been consolidated in recent times, and novel immune-related therapies are today demonstrating a strong clinical benefit in the setting of several solid neoplasms. Tumor-infiltrating lymphocytes reflect the attempt of the host to eradicate malignancies, and during the last decades, they have been shown to possess an interesting prognostic utility for breast cancer, especially in case of HER2 positive and triple-negative molecular subtypes. In parallel, the clinical evaluation of tumor-infiltrating lymphocytes has been shown to effectively predict treatment outcomes in both neoadjuvant and adjuvant settings. Currently, tumor-infiltrating lymphocytes are promising further predictive utility in view of novel immune-related therapeutic strategies which are coming into the clinical setting launching a solid rationale for the future next-generation treatment options. In this scenario, tumor-infiltrating lymphocytes might represent an important resource for the selection of the most appropriate therapeutic strategy, as well as further evaluations of the molecular mechanisms underlying tumor-infiltrating lymphocytes and the immunoediting process would eventually provide new insights to augment therapeutic success. Considering these perspectives, we review the potential utility of tumor-infiltrating lymphocytes in the definition of breast cancer prognosis and in the prediction of treatment outcomes, along with the new promising molecular-based therapeutic discoveries.
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Neoplasias da Mama/terapia , Linfócitos do Interstício Tumoral/imunologia , Neoplasias da Mama/classificação , Neoplasias da Mama/imunologia , Feminino , Humanos , Terapia Neoadjuvante , Prognóstico , Resultado do TratamentoRESUMO
BACKGROUND: HER-2 represents a relatively new therapeutic target for non small cell lung cancer (NSCLC) patients. The incidence for reported HER-2 overexpression/amplification/mutations ranges from 2 to 20% in NSCLC. Moreover, HER-2 amplification is a potential mechanism of resistance to tyrosine kinase inhibitors of the epidermal growth factor receptor (EGFR-TKI) (about 10% of cases). T-DM1, trastuzumab emtansine is an antibody-drug conjugate composed by the monoclonal antibody trastuzumab and the microtubule polymerization inhibitor DM1. The activity of T-DM1 has been studied in breast cancer but the role of T-DM1 in lung cancer remains unexplored. METHODS: Antiproliferative and proapoptotic effects of T-DM1 have been investigated in different NSCLC cell lines by MTT, crystal violet staining, morphological study and Western blotting. HER-2 expression and cell cycle were evaluated by flow cytometry and Western blotting. Antibody dependent cell cytotoxicity (ADCC) was measured with a CytoTox assay. Xenografted mice model has been generated using a NSCLC cell line to evaluate the effect of T-DM1 on tumor growth. Moreover, a morphometric and immunohistochemical analysis of tumor xenografts was conducted. RESULTS: In this study we investigated the effect of T-DM1 in a panel of NSCLC cell lines with different HER-2 expression levels, in H1781 cell line carrying HER-2 mutation and in gefitinib resistant HER-2 overexpressing PC9/HER2cl1 cell clone. T-DM1 efficiently inhibited proliferation with arrest in G2-M phase and induced cell death by apoptosis in cells with a significant level of surface expression of HER-2. Antibody-dependent cytotoxicity assay documented that T-DM1 maintained the same activity of trastuzumab. Our data also suggest that targeting HER-2 with T-DM1 potentially overcomes gefitinib resistance. In addition a correlation between cell density/tumor size with both HER-2 expression and T-DM1 activity was established in vitro and in an in vivo xenograft model. CONCLUSIONS: Our results indicate that targeting HER-2 with T-DM1 may offer a new therapeutic approach in HER-2 over-expressing lung cancers including those resistant to EGFR TKIs.
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Anticorpos Monoclonais Humanizados/farmacologia , Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Imunoconjugados/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Maitansina/análogos & derivados , Receptor ErbB-2/genética , Animais , Anticorpos Monoclonais Humanizados/química , Antineoplásicos/química , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feminino , Gefitinibe , Expressão Gênica , Humanos , Imunoconjugados/química , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Maitansina/química , Maitansina/farmacologia , Camundongos , Camundongos Nus , Quinazolinas/farmacologia , Receptor ErbB-2/metabolismo , Trastuzumab , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The approval of immune checkpoint inhibitors (ICIs) has revolutionized the management of metastatic renal cell carcinoma (RCC), introducing several ICI-based combinations as the new standard of care for affected patients. Nonetheless, monotherapy with antiangiogenic tyrosine kinase inhibitors (TKIs), such as pazopanib or sunitinib, still represents a first-line treatment option for selected patients belonging to the favorable risk group according to the International mRCC Database Consortium (IMDC) model. After TKI monotherapy, the main second-line option is represented by ICI monotherapy with the anti-Programmed Death Receptor 1(PD-1) nivolumab. To date, the expected clinical outcomes are similar with pazopanib or sunitinib and there is no clear indication for selecting one TKI over the other. Moreover, their impact on subsequent ICI treatment outcomes is not well defined, yet. Based on these premises, we investigated the immunomodulatory activity of these drugs in vitro and in vivo.Both TKIs induced Programmed Cell Death Ligand-1 (PD-L1) expression and soluble PD-L1 release in RCC cells, and hampered T cell activation, reducing cytokine production and the proportion of activated T cells. Nevertheless, in a syngeneic co-culture system with peripheral blood mononuclear cells (PBMCs) and tumor cells, incubation with anti-PD-1 antibody following TKIs treatment significantly restored T cell function, potentiating the cytotoxic effects against tumor cells. Pazopanib and sunitinib followed by anti-PD-1 antibody produced a comparable inhibition of tumor growth in a RCC syngeneic mouse model. Our findings suggest that pazopanib and sunitinib, showing similar immunomodulatory effects, may have a comparable impact on the subsequent effectiveness of PD-1/PD-L1 blockade.
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Inibidores da Angiogênese , Carcinoma de Células Renais , Neoplasias Renais , Receptor de Morte Celular Programada 1 , Inibidores de Proteínas Quinases , Pirimidinas , Sulfonamidas , Sunitinibe , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/patologia , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/patologia , Animais , Humanos , Camundongos , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/uso terapêutico , Inibidores da Angiogênese/administração & dosagem , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Sunitinibe/farmacologia , Sunitinibe/uso terapêutico , Linhagem Celular Tumoral , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Sulfonamidas/farmacologia , Sulfonamidas/uso terapêutico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , Indazóis/farmacologia , Indazóis/uso terapêutico , Indazóis/administração & dosagem , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Feminino , Ensaios Antitumorais Modelo de Xenoenxerto/métodos , Nivolumabe/farmacologia , Nivolumabe/uso terapêutico , Nivolumabe/administração & dosagem , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/metabolismoRESUMO
Osimertinib, a tyrosine kinase inhibitor targeting mutant EGFR, has received approval for initial treatment in patients with Non-Small Cell Lung Cancer (NSCLC). While effective in both first- and second-line treatments, patients eventually develop acquired resistance. Metabolic reprogramming represents a strategy through which cancer cells may resist and adapt to the selective pressure exerted by the drug. In the current study, we investigated the metabolic adaptations associated with osimertinib-resistance in NSCLC cells under low glucose culture conditions. We demonstrated that, unlike osimertinib-sensitive cells, osimertinib-resistant cells were able to survive under low glucose conditions by increasing the rate of glucose and glutamine uptake and by shifting towards mitochondrial metabolism. Inhibiting glucose/pyruvate contribution to mitochondrial respiration, glutamine deamination to glutamate, and oxidative phosphorylation decreased the proliferation and survival abilities of osimertinib-resistant cells to glucose starvation. Our findings underscore the remarkable adaptability of osimertinib-resistant NSCLC cells in a low glucose environment and highlight the pivotal role of mitochondrial metabolism in mediating this adaptation. Targeting the metabolic adaptive responses triggered by glucose shortage emerges as a promising strategy, effectively inhibiting cell proliferation and promoting cell death in osimertinib-resistant cells.
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The EGFR tyrosine kinase inhibitor osimertinib has been approved for the first-line treatment of EGFR-mutated Non-Small Cell Lung Cancer (NSCLC) patients. Despite its efficacy, patients develop resistance. Mechanisms of resistance are heterogeneous and not fully understood, and their characterization is essential to find new strategies to overcome resistance. Ceramides are well-known regulators of apoptosis and are converted into glucosylceramides (GlcCer) by glucosylceramide synthase (GCS). A higher content of GlcCers was observed in lung pleural effusions from NSCLC patients and their role in osimertinib-resistance has not been documented. The aim of this study was to determine the therapeutic potential of inhibiting GCS in NSCLC EGFR-mutant models resistant to osimertinib in vitro and in vivo. Lipidomic analysis showed a significant increase in the intracellular levels of glycosylceramides, including GlcCers in osimertinib resistant clones compared to sensitive cells. In resistant cells, the GCS inhibitor PDMP caused cell cycle arrest, inhibition of 2D and 3D cell proliferation, colony formation and migration capability, and apoptosis induction. The intratumoral injection of PDMP completely suppressed the growth of OR xenograft models. This study demonstrated that dysregulation of ceramide metabolism is involved in osimertinib-resistance and targeting GCS may be a promising therapeutic strategy for patients progressed to osimertinib.
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Acrilamidas , Carcinoma Pulmonar de Células não Pequenas , Glucosiltransferases , Indóis , Neoplasias Pulmonares , Pirimidinas , Humanos , Compostos de Anilina/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Receptores ErbB , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Mutação , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêuticoRESUMO
In this study, we investigated the effects and the underlying molecular mechanisms of the multi-kinase inhibitor sorafenib in a panel of breast cancer cell lines. Sorafenib inhibited cell proliferation and induced apoptosis through the mitochondrial pathway. These effects were neither correlated with modulation of MAPK and AKT pathways nor dependent on the ERα status. Sorafenib promoted an early perturbation of mitochondrial function, inducing a deep depolarization of mitochondrial membrane, associated with drop of intracellular ATP levels and increase of ROS generation. As a response to this stress condition, the energy sensor AMPK was rapidly activated in all the cell lines analyzed. In MCF-7 and SKBR3 cells, AMPK enhanced glucose uptake by up-regulating the expression of GLUT-1 glucose transporter, as also demonstrated by AMPKα1 RNA interference, and stimulated aerobic glycolysis thus increasing lactate production. Moreover, the GLUT-1 inhibitor fasentin blocked sorafenib-induced glucose uptake and potentiated its cytotoxic activity in SKBR3 cells. Persistent activation of AMPK by sorafenib finally led to the impairment of glucose metabolism both in MCF-7 and SKBR3 cells as well as in the highly glycolytic MDA-MB-231 cells, resulting in cell death. This previously unrecognized long-term effect of sorafenib was mediated by AMPK-dependent inhibition of the mTORC1 pathway. Suppression of mTORC1 activity was sufficient for sorafenib to hinder glucose utilization in breast cancer cells, as demonstrated by the observation that the mTORC1 inhibitor rapamycin induced a comparable down-regulation of GLUT-1 expression and glucose uptake. The key role of AMPK-dependent inhibition of mTORC1 in sorafenib mechanisms of action was confirmed by AMPKα1 silencing, which restored mTORC1 activity conferring a significant protection from cell death. This study provides insights into the molecular mechanisms driving sorafenib anti-tumoral activity in breast cancer, and supports the need for going on with clinical trials aimed at proving the efficacy of sorafenib for breast cancer treatment.
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Proteínas Quinases Ativadas por AMP/fisiologia , Antineoplásicos/farmacologia , Neoplasias da Mama/metabolismo , Metabolismo Energético/efeitos dos fármacos , Complexos Multiproteicos/antagonistas & inibidores , Proteínas de Neoplasias/fisiologia , Niacinamida/análogos & derivados , Compostos de Fenilureia/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/antagonistas & inibidores , Proteínas Quinases Ativadas por AMP/antagonistas & inibidores , Proteínas Quinases Ativadas por AMP/genética , Trifosfato de Adenosina/metabolismo , Anilidas/farmacologia , Divisão Celular/efeitos dos fármacos , Linhagem Celular Tumoral/efeitos dos fármacos , Regulação para Baixo , Feminino , Glucose/metabolismo , Transportador de Glucose Tipo 1/biossíntese , Transportador de Glucose Tipo 1/genética , Glicólise/efeitos dos fármacos , Humanos , Concentração Inibidora 50 , Alvo Mecanístico do Complexo 1 de Rapamicina , Mitocôndrias/metabolismo , Complexos Multiproteicos/fisiologia , Niacinamida/farmacologia , Fosforilação Oxidativa/efeitos dos fármacos , Interferência de RNA , RNA Interferente Pequeno/farmacologia , Sorafenibe , Serina-Treonina Quinases TOR/fisiologiaRESUMO
In the present study, a small set of reversible or irreversible 4-anilinoquinazoline EGFR inhibitors was tested in A549 cells at early (1h) and late (8h) time points after inhibitor removal from culture medium. A combination of assays was employed to explain the observed long-lasting inhibition of EGFR autophosphorylation. We found that EGFR inhibition at 8h can be due, besides to the covalent interaction of the inhibitor with Cys797, as for PD168393 (2) and its prodrug 4, to the intracellular accumulation of non-covalent inhibitors by means of an active cell uptake, as for 5 and 6. Compounds 5-6 showed similar potency and duration of inhibition of EGFR autophosphorylation as the covalent inhibitor 2, while being devoid of reactive groups forming covalent bonds with protein thiols.
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Receptores ErbB/antagonistas & inibidores , Quinazolinas , Compostos de Anilina/química , Compostos de Anilina/farmacocinética , Compostos de Anilina/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Química Farmacêutica , Humanos , Concentração Inibidora 50 , Estrutura Molecular , Fosforilação/efeitos dos fármacos , Quinazolinas/química , Quinazolinas/farmacocinética , Quinazolinas/farmacologia , Fatores de TempoRESUMO
Many cytokines control tumor development by directly lowering cancer cell proliferation and inducing apoptotic cell death, or indirectly by activating the antitumoral activity of specific immune cells such as NK or CD8+ T-lymphocytes [...].
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Citocinas , Neoplasias , Humanos , Citocinas/metabolismo , Linfócitos T CD8-Positivos , Células Matadoras Naturais , Neoplasias/tratamento farmacológicoRESUMO
Hepatoblastoma (HB) and pediatric hepatocellular carcinoma (HCC) are rare primary malignant liver cancers in children and young adults. HB is the most common and accounts for about 70 % cases; it is usually diagnosed during the first 3 years of life. Instead, pediatric HCC is uncommon, and it is associated with a poor prognosis. Overall, the prognosis of pediatric HCC is dismal with 5-year event-free survival of <30 % as compared to >80 % for HB. Surgery approaches, either resection or transplant, remain the best chance for the cure of pediatric HCC. However, chemotherapy can be helpful as an adjuvant or neoadjuvant treatment. International groups have done trials in pediatric HCC with a chemotherapy regimen, based on cisplatin and doxorubicin (PLADO) as for HB, but the efficacy is limited. Sorafenib, a multi-kinase inhibitor, following positive results in adults and in a pilot study in children, is now tested in conjunction with chemotherapy in the PHITT phase III clinical trial. Some studies have been exploring the genetic profiles of patients to find biological hallmarks that determine the aggressiveness of pediatric HCC. Pathways involved in growth and differentiation are dysregulated and as demonstrated in HB and adult HCC, an important role of the Wnt/CTNNB1 pathway in the pathogenesis of pediatric HCC is also emerging. An extended molecular analysis of tumor samples could give information about pathways as possible targets of biological and immunotherapeutic agents bringing new pharmacological options for the treatment of pediatric HCC.
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Antineoplásicos , Carcinoma Hepatocelular , Hepatoblastoma , Neoplasias Hepáticas , Criança , Adulto Jovem , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/patologia , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/patologia , Projetos Piloto , Hepatoblastoma/tratamento farmacológico , Hepatoblastoma/genética , Antineoplásicos/uso terapêuticoRESUMO
BACKGROUND: Cell-cycle regulators are mutated in approximately 40% of all cancer types and have already been linked to worse outcomes in non-small cell lung cancer adenocarcinomas treated with osimertinib. However, their exact role in osimertinib resistance has not been elucidated. OBJECTIVE: In this study, we aimed to evaluate how the CDK4/6-Rb axis may affect the sensitivity to osimertinib. METHODS: We genetically increased the level of CCND1 (Cyclin D1) and reduced the levels of CDKN2A (p16) in two different adenocarcinoma cell lines, PC9 and HCC827. We also retrospectively evaluated the outcome of patients with epidermal growth factor receptor-mutated advanced non-small cell lung cancer depending on their level of Cyclin D1 and p16. RESULTS: The modified clones showed higher proliferative capacity, modifications in cell-cycle phases, and higher migratory capacity than the parental cells. Cyclin D1-overexpressing clones were highly resistant to acute osimertinib treatment. CDKN2A knockdown conferred intrinsic resistance as well, although a longer time was required for adaption to the drug. In both cases, the resistant phenotype was epidermal growth factor receptor independent and associated with a higher level of Rb phosphorylation, which was unaffected by osimertinib treatment. Blocking the phosphorylation of Rb using abemaciclib, a CDK4/6 inhibitor, exerted an additive effect with osimertinib, increasing sensitivity to this drug and reverting the intrinsic resistant phenotype. In a group of 32 patients with epidermal growth factor receptor-mutated advanced non-small cell lung cancer, assessed for Cyclin D1 and p16 expression, we found that the p16-deleted group presented a lower overall response rate compared with the control group. CONCLUSIONS: We conclude that perturbation in cell-cycle regulators leads to intrinsic osimertinib resistance and worse patient outcomes.
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Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Humanos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Ciclina D1/genética , Ciclina D1/farmacologia , Ciclina D1/uso terapêutico , Estudos Retrospectivos , Resistencia a Medicamentos Antineoplásicos/genética , Linhagem Celular Tumoral , Receptores ErbB/metabolismo , Compostos de Anilina/farmacologia , Compostos de Anilina/uso terapêutico , Mutação , Inibidores de Proteínas Quinases/uso terapêuticoRESUMO
BACKGROUND: The epidermal growth factor receptor (EGFR) is an established target for anti-cancer treatment in different tumour types. Two different strategies have been explored to inhibit this pivotal molecule in epithelial cancer development: small molecules TKIs and monoclonal antibodies. ErbB/HER-targeting by monoclonal antibodies such as cetuximab and trastuzumab or tyrosine-kinase inhibitors as gefitinib or erlotinib has been proven effective in the treatment of advanced NSCLC. RESULTS: In this study we explored the potential of combining either erlotinib with cetuximab or trastuzumab to improve the efficacy of EGFR targeted therapy in EGFR wild-type NSCLC cell lines. Erlotinib treatment was observed to increase EGFR and/or HER2 expression at the plasma membrane level only in NSCLC cell lines sensitive to the drug inducing protein stabilization. The combined treatment had marginal effect on cell proliferation but markedly increased antibody-dependent, NK mediated, cytotoxicity in vitro. Moreover, in the Calu-3 xenograft model, the combination significantly inhibited tumour growth when compared with erlotinib and cetuximab alone. CONCLUSION: Our results indicate that erlotinib increases surface expression of EGFR and/or HER2 only in EGFR-TKI sensitive NSCLC cell lines and, in turns, leads to increased susceptibility to ADCC both in vitro and in a xenograft models. The combination of erlotinib with monoclonal antibodies represents a potential strategy to improve the treatment of wild-type EGFR NSCLC patients sensitive to erlotinib.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/imunologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/imunologia , Análise de Variância , Animais , Anticorpos Monoclonais Humanizados/administração & dosagem , Citotoxicidade Celular Dependente de Anticorpos/efeitos dos fármacos , Sítios de Ligação/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/química , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Cetuximab , Sinergismo Farmacológico , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/química , Receptores ErbB/genética , Receptores ErbB/metabolismo , Cloridrato de Erlotinib , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Pulmonares/química , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Estabilidade Proteica/efeitos dos fármacos , Quinazolinas/administração & dosagem , Receptor ErbB-2/antagonistas & inibidores , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Trastuzumab , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Background: Pancreatic ductal adenocarcinoma (PDAC) is a highly devastating disease with rising incidence and poor prognosis. The lack of reliable prognostic biomarkers hampers the individual evaluation of the survival and recurrence potential. Methods: Here, we investigate the value of plasma levels of two potential key players in molecular mechanisms underlying PDAC aggressiveness and immune evasion, soluble TGF-beta (sTGF-beta) and sPD-L1, in both metastatic and radically-resected PDAC. To this aim we prospectively enrolled 38 PDAC patients and performed appropriate statistical analyses in order to evaluate their correlation, and role in the prediction of disease relapse/progression, and patients' outcome. Results: Metastatic patients showed lower levels of circulating sTGF-beta and higher levels of sPD-L1 compared to radically-resected patients. Moreover, a decrease in sTGF-beta levels (but not sPD-L1) was significantly associated with disease relapse in radically-resected patients. We also observed lower sTGF-beta at disease progression after first-line chemotherapy in metastatic patients, though this change was not statistically significant. We found a significant correlation between the levels of sTGF-beta and sPD-L1 before first-line chemotherapy. Conclusions: These findings support the possible interaction of TGF-beta and PD-L1 pathways and suggest that sTGF-beta and sPD-L1 might synergize and be new potential blood-based biomarkers.
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BACKGROUND: The loss of the CDKN2A/ARF (cyclin-dependent kinase inhibitor 2A/alternative reading frame) gene is the most common alteration in malignant pleural mesothelioma (MPM), with an incidence of about 70%, thus representing a novel target for mesothelioma treatment. In the present study, we evaluated the antitumor potential of combining the standard chemotherapy regimen used for unresectable MPM with the CDK4/6 (cyclin-dependent kinase 4 or 6) inhibitor abemaciclib. METHODS: Cell viability, cell death, senescence, and autophagy induction were evaluated in two MPM cell lines and in a primary MPM cell culture. RESULTS: The simultaneous treatment of abemaciclib with cisplatin and pemetrexed showed a greater antiproliferative effect than chemotherapy alone, both in MPM cell lines and in primary cells. This combined treatment induced cellular senescence or autophagic cell death, depending on the cell type. More in detail, the induction of cellular senescence was related to the increased expression of p21, whereas autophagy induction was due to the impairment of the AKT/mTOR signaling. Notably, the effect of the combination was irreversible and no resumption in tumor cell proliferation was observed after drug withdrawal. CONCLUSION: Our results demonstrated the therapeutic potential of CDK4/6 inhibitors in combination with chemotherapy for the treatment of MPM and are consistent with the recent positive results in the MiST2 arm in abemaciclib-treated patients.
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Cachexia is a metabolic syndrome consisting of massive loss of muscle mass and function that has a severe impact on the quality of life and survival of cancer patients. Up to 20% of lung cancer patients and up to 80% of pancreatic cancer patients are diagnosed with cachexia, leading to death in 20% of them. The main drivers of cachexia are cytokines such as interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), macrophage inhibitory cytokine 1 (MIC-1/GDF15) and transforming growth factor-beta (TGF-ß). Besides its double-edged role as a tumor suppressor and activator, TGF-ß causes muscle loss through myostatin-based signaling, involved in the reduction in protein synthesis and enhanced protein degradation. Additionally, TGF-ß induces inhibin and activin, causing weight loss and muscle depletion, while MIC-1/GDF15, a member of the TGF-ß superfamily, leads to anorexia and so, indirectly, to muscle wasting, acting on the hypothalamus center. Against this background, the blockade of TGF-ß is tested as a potential mechanism to revert cachexia, and antibodies against TGF-ß reduced weight and muscle loss in murine models of pancreatic cancer. This article reviews the role of the TGF-ß pathway and to a minor extent of other molecules including microRNA in cancer onset and progression with a special focus on their involvement in cachexia, to enlighten whether TGF-ß and such other players could be potential targets for therapy.
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Caquexia , Neoplasias Pancreáticas , Fator de Crescimento Transformador beta , Animais , Caquexia/metabolismo , Humanos , Camundongos , Neoplasias Pancreáticas/complicações , Neoplasias Pancreáticas/metabolismo , Qualidade de Vida , Fator de Crescimento Transformador beta/metabolismo , Fatores de Crescimento Transformadores , Neoplasias PancreáticasRESUMO
Hepatocellular carcinoma (HCC) is the most frequent primary liver cancer with a poor prognosis and limited treatment options. Considering that alterations of the CDK4/6-cyclin D-Rb pathway occur frequently in HCC, we tested the efficacy of two CDK4/6 inhibitors, abemaciclib and ribociclib, in combination with lenvatinib, a multi-kinase inhibitor approved as first-line therapy for advanced HCC, in a panel of HCC Rb-expressing cell lines. The simultaneous drug combinations showed a superior anti-proliferative activity as compared with single agents or sequential schedules of treatment, either in short or in long-term experiments. In addition, the simultaneous combination of abemaciclib with lenvatinib reduced 3D cell growth, and impaired colony formation and cell migration. Mechanistically, these growth-inhibitory effects were associated with a stronger down-regulation of c-myc protein expression. Depending on the HCC cell model, reduced activation of MAPK, mTORC1/p70S6K or src/FAK signaling was also observed. Abemaciclib combined with lenvatinib arrested the cells in the G1 cell cycle phase, induced p21 accumulation, and promoted a stronger increase of cellular senescence, associated with elevation of ß-galactosidase activity and accumulation of ROS, as compared with single treatments. After drug withdrawal, the capacity of forming colonies was significantly impaired, suggesting that the anti-tumor efficacy of abemaciclib and lenvatinib combination was persistent. Our pre-clinical results demonstrate the effectiveness of the simultaneous combination of CDK4/6 inhibitors with lenvatinib in HCC cell models, suggesting that this combination may be worthy of further investigation as a therapeutic approach for the treatment of advanced HCC.
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INTRODUCTION: ALK tyrosine kinase inhibitors (TKIs) are the standard treatment for advanced ALK-positive NSCLC. Nevertheless, drug resistance inevitably occurs. Here, we report a case of a patient with metastatic ALK-positive lung adenocarcinoma with an impressive resistance to sequential treatment with ALK TKIs mediated by YES1 and MYC amplification in a contest of epithelial-to-mesenchymal transition and high progressive chromosomal instability. METHODS: The patient received, after chemotherapy and 7 months of crizotinib, brigatinib and lorlatinib with no clinical benefit to both treatments. A study of resistance mechanisms was performed with whole exome sequencing on different biological samples; primary cell lines were established from pleural effusion after lorlatinib progression. RESULTS: At whole exome sequencing analysis, YES1 and MYC amplifications were observed both in the pericardial biopsy and the pleural effusion samples collected at brigatinib and lorlatinib progression, respectively. Increasing chromosomal instability from diagnostic biopsy to pleural effusion was also observed. The addition of dasatinib to brigatinib or lorlatinib restored the sensitivity in primary cell lines; data were confirmed also in H3122_ALK-positive model overexpressing both YES1 and MYC. CONCLUSIONS: In conclusion, YES1 and MYC amplifications are candidates to justify a rapid acquired resistance to crizotinib entailing primary brigatinib and lorlatinib resistance. In this context, a combination strategy of ALK TKI with dasatinib could be effective to overcome a rapid resistance.
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BACKGROUND: Gefitinib is a tyrosine kinase inhibitor (TKI) of the epidermal growth factor receptor (EGFR) especially effective in tumors with activating EGFR gene mutations while EGFR wild-type non small cell lung cancer (NSCLC) patients at present do not benefit from this treatment.The primary site of gefitinib metabolism is the liver, nevertheless tumor cell metabolism can significantly affect treatment effectiveness. RESULTS: In this study, we investigated the intracellular metabolism of gefitinib in a panel of EGFR wild-type gefitinib-sensitive and -resistant NSCLC cell lines, assessing the role of cytochrome P450 1A1 (CYP1A1) inhibition on gefitinib efficacy. Our results indicate that there is a significant difference in drug metabolism between gefitinib-sensitive and -resistant cell lines. Unexpectedly, only sensitive cells metabolized gefitinib, producing metabolites which were detected both inside and outside the cells. As a consequence of gefitinib metabolism, the intracellular level of gefitinib was markedly reduced after 12-24 h of treatment. Consistent with this observation, RT-PCR analysis and EROD assay showed that mRNA and activity of CYP1A1 were present at significant levels and were induced by gefitinib only in sensitive cells. Gefitinib metabolism was elevated in crowded cells, stimulated by exposure to cigarette smoke extract and prevented by hypoxic condition. It is worth noting that the metabolism of gefitinib in the sensitive cells is a consequence and not the cause of drug responsiveness, indeed treatment with a CYP1A1 inhibitor increased the efficacy of the drug because it prevented the fall in intracellular gefitinib level and significantly enhanced the inhibition of EGFR autophosphorylation, MAPK and PI3K/AKT/mTOR signalling pathways and cell proliferation. CONCLUSION: Our findings suggest that gefitinib metabolism in lung cancer cells, elicited by CYP1A1 activity, might represent an early assessment of gefitinib responsiveness in NSCLC cells lacking activating mutations. On the other hand, in metabolizing cells, the inhibition of CYP1A1 might lead to increased local exposure to the active drug and thus increase gefitinib potency.
Assuntos
Antineoplásicos/farmacologia , Citocromo P-450 CYP1A1/metabolismo , Receptores ErbB/antagonistas & inibidores , Quinazolinas/farmacologia , Carcinoma Pulmonar de Células não Pequenas , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Receptores ErbB/genética , Receptores ErbB/metabolismo , Gefitinibe , Humanos , Neoplasias Pulmonares , Mutação , FosforilaçãoRESUMO
A facile synthesis of model 4-oxopyrido[3',2':4,5]thieno[3,2-b]indole-3-carboxylic acids 9a-e was achieved via Stille arylation of 2-chloro-3-nitro-4-oxothieno[2,3-b]pyridine-5-carboxylate and a subsequent microwave-assisted phosphite-mediated Cadogan reaction. The new compounds were tested for their in vitro antimicrobial and antiproliferative activity. Compounds 9a-c and 9e exhibited very high potency against Gram positive Bacillus subtilis and Bacillus megaterium at concentrations 0.000015-0.007 µg/mL. They also displayed excellent activity towards other Gram positive bacilli and staphylococci and Gram negative Haemophilus influenzae, being in most cases superior or equal to commercial fluoroquinolones. Both 9a and 9c were inhibitors of the DNA gyrase activity. As concerns antitumor properties, compounds 9b-e showed growth inhibition of MCF-7 breast tumor and A549 non-small cell lung cancer cells with IC(50) 1.6-2.8 µM and 2.6-6.9 µM, respectively, coupled with absence of cytotoxicity towards normal cells. These compounds are promising as dual acting chemotherapeutics.