Showing papers by "Luis M. Montuenga published in 2013"

A Prognostic DNA Methylation Signature for Stage I Non–Small-Cell Lung Cancer

Abstract: Purpose Non–small-cell lung cancer (NSCLC) is a tumor in which only small improvements in clinical outcome have been achieved. The issue is critical for stage I patients for whom there are no available biomarkers that indicate which high-risk patients should receive adjuvant chemotherapy. We aimed to find DNA methylation markers that could be helpful in this regard. Patients and Methods A DNA methylation microarray that analyzes 450,000 CpG sites was used to study tumoral DNA obtained from 444 patients with NSCLC that included 237 stage I tumors. The prognostic DNA methylation markers were validated by a single-methylation pyrosequencing assay in an independent cohort of 143 patients with stage I NSCLC. Results Unsupervised clustering of the 10,000 most variable DNA methylation sites in the discovery cohort identified patients with high-risk stage I NSCLC who had shorter relapse-free survival (RFS; hazard ratio [HR], 2.35; 95% CI, 1.29 to 4.28; P = .004). The study in the validation cohort of the signific...

Investigation of Complement Activation Product C4d as a Diagnostic and Prognostic Biomarker for Lung Cancer

Abstract: Lung cancer is the leading cause of cancer death worldwide (1). The US National Lung Screening Trial demonstrated that computed tomography (CT) screening reduces lung cancer mortality (2). In this context, the use of biomarkers may help in the implementation of population-based screening programs. Biomarkers could be used to identify populations at increased risk, to confirm the presence of malignant cells, or to monitor response to treatment. Numerous molecular markers have been proposed (3). Unfortunately, genetic heterogeneity has limited the success of these initiatives and, to date, no diagnostic marker has proven useful in lung cancer clinical practice. To overcome this limitation, an alternative approach would be to look not for cancer but for the immune response to cancer (4). Immune activation may generate host-derived markers more homogeneous than cancer-derived markers. Immune responses against intracellular and surface tumor antigens are well documented in patients with lung cancer (5). In particular, the complement system is activated in lung tumor cells (6–9). Complement is a central component of innate immunity that plays an essential role in immune surveillance and homeostasis (10). In the past years, our group has evaluated the role of complement in the control of lung cancer cell growth (9,11–13). We have recently reported that lung cancer cells produce C5a, a potent proinflammatory mediator that creates a favorable microenvironment for lung cancer progression (14). However, the pathway by which lung cancer cells activate complement and the value of complement activation fragments as diagnostic biomarkers remain unclear. In this study, we dissected the mechanisms by which complement is activated in lung cancer cells and evaluated the diagnostic performance of molecules released during complement activation. Our results indicate that lung tumors activate the classical complement pathway and generate C4d, a degradation product of this pathway. Moreover, our results suggest that the determination of C4d may be of value for the diagnosis and prognosis of lung cancer.

Phosphorylated tubulin adaptor protein CRMP‐2 as prognostic marker and candidate therapeutic target for NSCLC

Abstract: Collapsin response mediator protein-2 (CRMP-2) is the first described and most studied member of a family of proteins that mediate the addition of tubulin dimers to the growing microtubule. CRMPs have mainly been studied in the nervous system, but recently, they have been described in other tissues where they participate in vesicle transport, migration and mitosis. In this work, we aimed at studying the role of CRMP-2 in lung cancer cell division. We first explored the expression of CRMP-2 and phosphorylated (Thr 514) CRMP-2 in 91 samples obtained from patients with localized nonsmall cell lung cancer. We observed a significant correlation between high levels of nuclear phosphorylated CRMP-2 and poor prognosis in those patients. Interestingly, this association was only positive for untreated patients. To provide a mechanistic explanation to these findings, we used in vitro models to analyze the role of CRMP-2 and its phosphorylated forms in cell division. Thus, we observed by confocal microscopy and immunoprecipitation assays that CRMP-2 differentially colocalizes with the mitotic spindle during cell division. The use of phosphodefective or phosphomimetic mutants of CRMP-2 allowed us to prove that anomalies in the phosphorylation status of CRMP-2 result in changes in the mitotic tempo, and increments in the number of multinucleated cells. Finally, here we demonstrate that CRMP-2 phosphorylation impairment, or silencing induces p53 expression and promotes apoptosis through caspase 3 activation. These results pointed to CRMP-2 phosphorylation as a prognostic marker and potential new target to be explored in cancer therapy.

Multiscale in situ analysis of the role of dyskerin in lung cancer cells

Abstract: Dyskerin is one of the three subunits of the telomerase ribonucleoprotein (RNP) complex. Very little is known about the role of dyskerin in the biology of the telomeres in cancer cells. In this study, we use a quantitative, multiscale 3D image-based in situ method and several molecular techniques to show that dyskerin is overexpressed in lung cancer cell lines. Furthermore, we show that dyskerin expression correlates with telomere length both at the cell population level – cells with higher dyskerin expression have short telomeres – and at the single cell level – the shortest telomeres of the cell are spatially associated with areas of concentration of dyskerin proteins. Using this in vitro model, we also show that exogenous increase in dyskerin expression confers resistance to telomere shortening caused by a telomerase inactivating drug. Finally, we show that resistance is achieved by the recovery of telomerase activity associated with dyskerin. In summary, using a novel multiscale image-based in situ method, we show that, in lung cancer cell lines, dyskerin responds to continuous telomere attrition by increasing the telomerase RNP activity, which in turn provides resistance to telomere shortening.

Smokers with CT detected emphysema and no airway obstruction have decreased plasma levels of EGF, IL-15, IL-8 and IL-1ra.

Abstract: Rationale: Low-grade inflammation and emphysema have been shown to be associated with an increased risk of lung cancer. However, the systemic inflammatory response in patients with emphysema is still unknown. Objective: To compare the plasma cytokine profiles in two groups of current or former smokers without airway obstruction: a control group of individuals without computed tomography (CT) detected emphysema vs. a study group of individuals with CT detected emphysema. Methods: Subjects underwent a chest CT, spirometry, and determination of EGF, IL-15, IL-1ra, IL-8, MCP-1, MIP-1b, TGFa, TNFa, and VEGF levels in plasma. Cytokine levels in each group were compared adjusting for confounding factors. Results: 160 current smokers and former smokers without airway obstruction participated in the study: 80 without emphysema and 80 subjects with emphysema. Adjusted group comparisons revealed significant reductions in EGF (20.317, p=0.01), IL-15 (20.21, p=0.01), IL-8 (20.180, p=0.02) and IL-1ra (20.220, p=0.03) in subjects with emphysema and normal spirometry. Conclusions: Current or former smokers expressing a well-defined disease characteristic such as emphysema, has a specific plasma cytokine profile. This includes a decrease of cytokines mainly implicated in activation of apoptosis or decrease of immunosurveillance. This information should be taken into account when evaluated patients with tobacco respiratory diseases.