RESUMEN
Lung carcinoma is still the most common malignancy worldwide. One of the major subtypes of non-small cell lung cancer (NSCLC) is adenocarcinoma (AC). As driver mutations and hence therapies differ in AC subtypes, we theorized that the expression and function of ABC drug transporters important in multidrug resistance (MDR) would correlate with characteristic driver mutations KRAS or EGFR. Cisplatin resistance (CR) was generated in A549 (KRAS) and PC9 (EGFR) cell lines and gene expression was tested. In three-dimensional (3D) multicellular aggregate cultures, both ABCB1 and ABCG2 transporters, as well as the WNT microenvironment, were investigated. ABCB1 and ABCG2 gene expression levels were different in primary AC samples and correlated with specific driver mutations. The drug transporter expression pattern of parental A549 and PC9, as well as A549-CR and PC9-CR, cell lines differed. Increased mRNA levels of ABCB1 and ABCG2 were detected in A549-CR cells, compared to parental A549, while the trend observed in the case of PC9 cells was different. Dominant alterations were observed in LEF1, RHOU and DACT1 genes of the WNT signalling pathway in a mutation-dependent manner. The study confirmed that, in lung AC-s, KRAS and EGFR driver mutations differentially affect both drug transporter expression and the cisplatin-induced WNT signalling microenvironment.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/genética , Carcinoma de Pulmón de Células no Pequeñas/genética , Resistencia a Múltiples Medicamentos/genética , Resistencia a Antineoplásicos/genética , Neoplasias Pulmonares/genética , Mutación/genética , Proteínas Proto-Oncogénicas p21(ras)/genética , Células A549 , Anciano , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Línea Celular , Línea Celular Tumoral , Cisplatino/farmacología , Receptores ErbB/genética , Femenino , Expresión Génica/genética , Células Endoteliales de la Vena Umbilical Humana , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Masculino , Persona de Mediana Edad , Microambiente Tumoral/genéticaRESUMEN
BACKGROUND: The predominant metastatic site of lung cancer (LC) is the brain. Although outdated, conventional cisplatin treatment is still the main therapeutic approach for patients with advanced non-small cell lung cancer (NSCLC), since targeted therapy that offers better tumor control is not always possible. In the present study brain metastasis associated cytokine expression was investigated in primary NSCLC adenocarcinoma (AC) tissues with known oncogenic mutations in the presence or absence of platina based and tyrosine kinase inhibitor (TKI) drugs. METHODS: Primary lung tumor samples were isolated, DNA was sequenced and then the samples were grouped based on mutation. Experiments were also performed using KRAS mutant A549 and EGFR mutant PC-9 cells. Drug response was analyzed in three dimensional (3D) tissue cultures. We assessed drug response and IL-6 and IL-8 cytokine expression in relation to cellular invasion using ATP dependent cell viability, qRT-PCR analysis, cytokine bead array, and migration assay. RESULTS: In 3D co-cultures, primary NSCLC derived cells harboring EGFR mutation responded better to erlotinib treatment than KRAS mutant or KRAS/EGFR wild type (WT) cancer cells. In contrast, under the same culture conditions KRAS/EGFR WT or KRAS mutant cancer cells are more sensitive to cisplatin than EGFR mutant cells. Drug response and pro-inflammatory cytokine production varied depending on the driver mutations. Cisplatin but not erlotinib increased both IL-6 and IL-8 secretion and only IL-6 increased cellular migration and proliferation. CONCLUSION: In vitro assays are available to determine the response to planned therapeutic approach of lung cancer subtypes. The sequence of administration of therapeutic drugs determines cytokine production and therefore therapeutic response.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Cisplatino/uso terapéutico , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Neoplasias Pulmonares/metabolismo , Mutación/fisiología , Células A549 , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Movimiento Celular/efectos de los fármacos , Movimiento Celular/genética , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Cisplatino/farmacología , Humanos , Interleucina-6/genética , Interleucina-8/genética , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/genética , Mutación/efectos de los fármacosRESUMEN
BACKGROUND: Type 2 alveolar epithelial cells (AT2s) behave as stem cells and show clonal proliferation upon alveolar injury followed by trans-differentiation (TD) into Type 1 alveolar epithelial cells (AT1s). In the present study we identified signaling pathways involved in the physiological AT2-to-AT1 TD process. METHODS: AT2 cells can be isolated from human lungs and cultured in vitro where they undergo TD into AT1s. In the present study we identified signaling pathways involved in the physiological AT2-to-AT1 TD process using Affymetrix microarray, qRT-PCR, fluorescence microscopy, and an in vitro lung aggregate culture. RESULTS: Affymetrix microarray revealed Wnt signaling to play a crucial role in the TD process. Wnt7a was identified as a ligand regulating the AT1 marker, Aquaporin 5 (AQP5). Artificial Neural Network (ANN) analysis of the Affymetrix data exposed ITGAV: Integrin alpha V (ITGAV), thrombospondin 1 (THBS1) and epithelial membrane protein 2 (EMP2) as Wnt signaling targets. CONCLUSIONS: Wnt signaling targets that can serve as potential alveolar epithelial repair targets in future therapies of the gas exchange surface after injury. As ITGAV is significantly increases during TD and is regulated by Wnt signaling, ITGAV might be a potential target to speed up the alveolar healing process.