Sonia Blanco-Prieto

On the identification of biomarkers for non-small cell lung cancer in serum and pleural effusion

The current imperative need for new biomarkers of non-small cell lung cancer (NSCLC) prompted us to compare the proteome of serum and pleural effusion samples from cancer patients with those with benign lung diseases as pneumonia or tuberculosis. Samples were prefractionated through affinity chromatography prior to 2D-DIGE to detect proteins with altered expression in cancer patients. Overall, we identified more potential biomarkers in pleural effusion, which is closer to the affected organ, than in serum. Nevertheless, in both cases principal component analysis demonstrated that the pattern of significantly altered proteins discriminates between disease groups. The biomarker candidates comprise proteins increased in malignant pleural effusions as gelsolin and the metalloproteinase inhibitor 2, and others with lower levels as S100-A8 and S100-A9. The most interesting protein was the pigment epithelium-derived factor (PEDF), which is related to angiogenesis inhibition, and was significantly overexpressed both in serum and pleural effusion from NSCLC patients. More than 12 PEDF isoforms were specifically immunodetected in both fluids in 2-D blots, most of them overexpressed in NSCLC. Thus, further validation would be ideally directed to quantify individual PEDF isoforms, as it may be only one or some of them the ones altered in the cancer process.

Pretreatment levels of the serum biomarkers CEA, CYFRA 21-1, SCC and the soluble EGFR and its ligands EGF, TGF-alpha, HB-EGF in the prediction of outcome in erlotinib treated non-small-cell lung cancer patients.

The aim of this study has been to investigate the potential of serum biomarkers used in clinical practice (CEA, CYFRA 21–1, SCC) together with the serum epidermal growth factor receptor (EGFR) and its associated ligands (EGF, TGF-α, HB-EGF) as outcome predictors of non-small cell lung cancer (NSCLC) patients treated with the TKI erlotinib. The pretreatment levels of these markers were evaluated through immunoassays carried out in 58 patients. The progression-free survival (PFS) and overall survival (OS) were assessed by the Kaplan-Meier method and differences between groups were compared by means of the Log-Rank test. Association of risk factors with survival was evaluated using the univariate and multivariate Cox modelling procedures. Higher CEA (>5xa0ng/mL) and sEGFR (>56.87xa0ng/mL) concentrations associated significantly with a higher overall survival. The pre-treatment sEGFR serum levels constituted an independent prognostic factor. The EGFR gene mutational status and the sEGFR level combination was the single to associate significantly with longer progression-free survival periods, in circumstances in which the EGFR gene was mutated and increased protein serum levels were detected. The overall survival as assessed through a Cox analysis revealed similar death hazards with respect to low sEGFR levels combined both with non-mutated EGFR genotypes and low CEA serum levels. Our results suggest that the pre-treatment CEA and sEGFR serum levels may provide a comparable source of information to that supplied by the EGFR gene mutational status with respect to the prognosis of erlotinib treated NSCLC patients. A combined sEGFR and CEA level appraisal could be of considerable value to select patients to undergo EGFR-TKI treatments.

Secreted clusterin in colon tumor cell models and its potential as diagnostic marker for colorectal cancer.

We studied the specific changes of the secreted protein clusterin and its cytoplasmic precursor regarding colorectal tumorigenesis, using in vitro differentiation of Caco-2 cells. In tumor-like stage, we observed an overexpression of both precursor and secreted clusterin, corroborated in the cell line SW-480. Noticeably, SW-620 cells (from a tumoral node, thus with metastatic capacity) did not show overexpression of either precursor or secreted clusterin, suggesting a downregulation related to local metastasis. We further investigated clusterin in serum, finding a significant increase in colorectal cancer patients, with 81% sensitivity, 79% specificity, and an area under the ROC curve of 0.85.

Calprotectin: a novel biomarker for the diagnosis of pleural effusion

Background:Novel non-invasive biomarkers for the precise diagnosis of malignancy in pleural effusion (PE) are needed. The aim of this study was to determine the diagnostic accuracy of calprotectin for predicting malignancy in patients with exudative PE.Methods:Calprotectin concentration was measured in 156 individuals diagnosed with exudative PE (67 malignant and 89 benign). Calprotectin accuracy for discriminating between malignant and benign PE was evaluated using receiver operating characteristic (ROC) curves. Univariate and multivariate logistic regression were performed to test the association between calprotectin levels and malignant PE.Results:Calprotectin levels were significantly lower in malignant pleural fluid (257.2u2009ngu2009ml−1, range: 90.7–736.4) than benign effusions (2627.1u2009ngu2009ml−1, range: 21–9530.1). The area under the curve was 0.963. A cutoff point of ⩽736.4u2009ngu2009ml−1 rendered a sensitivity of 100%, with a specificity of 83.15%, which could prove useful to delimit those patients with negative cytology tests that should be referred for more invasive diagnostic procedures. Logistic regression demonstrated a strong association between calprotectin and malignancy (adjusted OR 663.14).Conclusion:Calprotectin predicts malignancy in pleural fluid with high accuracy and could be a good complement to cytological methods.

Serum calprotectin, CD26 and EGF to establish a panel for the diagnosis of lung cancer.

Lung cancer is the most lethal neoplasia, and an early diagnosis is the best way for improving survival. Symptomatic patients attending Pulmonary Services could be diagnosed with lung cancer earlier if high-risk individuals are promptly separated from healthy individuals and patients with benign respiratory pathologies. We searched for a convenient non-invasive serum test to define which patients should have more immediate clinical tests. Six cancer-associated molecules (HB-EGF, EGF, EGFR, sCD26, VEGF, and Calprotectin) were investigated in this study. Markers were measured in serum by specific ELISAs, in an unselected population that included 72 lung cancer patients of different histological types and 56 control subjects (healthy individuals and patients with benign pulmonary pathologies). Boosted regression and random forests analysis were conducted for the selection of the best candidate biomarkers. A remarkable discriminatory capacity was observed for EGF, sCD26, and especially for Calprotectin, these three molecules constituting a marker panel boasting a sensitivity of 83% and specificity of 87%, resulting in an associated misclassification rate of 15%. Finally, an algorithm derived by logistic regression and a nomogram allowed generating classification scores in terms of the risk of a patient of suffering lung cancer. In conclusion, we propose a non-invasive test to identify patients at high-risk for lung cancer from a non-selected population attending a Pulmonary Service. The efficacy of this three-marker panel must be tested in a larger population for lung cancer.

Preoperative serum CA 72.4 as prognostic factor of recurrence and death, especially at TNM stage II, for colorectal cancer

BackgroundNowadays, evaluation of colorectal cancer prognosis and decision-making for treatment continues to be based primarily on TNM tumour stage. Administration of adjuvant chemotherapy is especially challenging for stage II patients that can have very different disease-related outcomes. Therefore, more reliable prognostic markers need to be developed to improve the selection of stage II patients at high risk for recurrence. Our purpose is to assess the prognostic value of preoperative serum CA 72.4 to improve the risk stratification of CRC patients.MethodsPreoperative sera collected from 71 unselected patients between January 1994 and February 1997 was assayed for CA 72.4 and CEA levels. Patients were followed-up for at least 30xa0months or until relapse. Survival curves were estimated by the Kaplan-Meier method and the prognostic value was determined using Log-Rank test and Cox regression analysis.ResultsPreoperative CA 72.4 levels above 7 U/mL correlate with a worse prognosis, with associated recurrence and death percentages exceeding the displayed by CEA. In a multivariate analysis, its combination with CEA proved the most important independent factor predicting survival. Remarkably, at stage II CA 72.4 also discriminates better than CEA those patients that will relapse or die from those with a favourable prognosis; however, CEA has not a negligible effect on survival.ConclusionsThe most outstanding finding of the present work is the correct classification of nearly every patient with bad prognosis (relapse or death) at TNM stage II when CEA and CA 72.4 are used altogether. This could improve the decision-making involved in the treatment of stage II colon cancer. Certainly further large-scale studies must be performed to determine whether CA 72.4 can be effectively used in the clinical setting.

Highly Sensitive Marker Panel for Guidance in Lung Cancer Rapid Diagnostic Units

While evidence for lung cancer screening implementation in Europe is awaited, Rapid Diagnostic Units have been established in many hospitals to accelerate the early diagnosis of lung cancer. We seek to develop an algorithm to detect lung cancer in a symptomatic population attending such unit, based on a sensitive serum marker panel. Serum concentrations of Epidermal Growth Factor, sCD26, Calprotectin, Matrix Metalloproteinases −1, −7, −9, CEA and CYFRA 21.1 were determined in 140 patients with respiratory symptoms (lung cancer and controls with/without benign pathology). Logistic Lasso regression was performed to derive a lung cancer prediction model, and the resulting algorithm was tested in a validation set. A classification rule based on EGF, sCD26, Calprotectin and CEA was established, able to reasonably discriminate lung cancer with 97% sensitivity and 43% specificity in the training set, and 91.7% sensitivity and 45.4% specificity in the validation set. Overall, the panel identified with high sensitivity stage I non-small cell lung cancer (94.7%) and 100% small-cell lung cancers. Our study provides a sensitive 4-marker classification algorithm for lung cancer detection to aid in the management of suspicious lung cancer patients in the context of Rapid Diagnostic Units.

Relevance of matrix metalloproteases in non-small cell lung cancer diagnosis

BackgroundThe need for novel biomarkers that could aid in non-small cell lung cancer (NSCLC) detection, together with the relevance of Matrix Metalloproteases (MMPs) -1, -2, -7, -9 and -10 in lung tumorigenesis, prompted us to assess the diagnostic usefulness of these MMPs and the Tissue Inhibitor of Metalloproteinase (TIMP) -1 in NSCLC patients.MethodsMarkers were evaluated in an initial study cohort (19 NSCLC cases and 19 healthy controls). Those that better performed were analyzed in a larger sample including patients with benign lung diseases. Serum MMPs and TIMP-1 were determined by multiplexed immunoassays. Logistic regression was employed for multivariate analysis of biomarker combinations.ResultsMMPs and TIMP-1 were elevated in the serum of NSCLC patients compared to healthy controls. MMP-1, -7 and -9 performed at best and were further evaluated in the sample including benign pathologies, corroborating the superiority of MMP-9 in NSCLC discrimination, also at early-stage NSCLC. The optimal diagnostic value was obtained with the model including MMP-9, gender, age and smoking history, that demonstrated an AUC of 0.787, 85.54% sensitivity and 64.89% specificity.ConclusionOur results suggest that MMP-9 is a potential biomarker for NSCLC diagnosis and its combined measurement with other biomarkers could improve NSCLC detection.

Levels of PEDF in pleural effusions from lung adenocarcinoma and benign disease patients

Anti-tumor properties assigned to PEDF, beside its role as an inhibitor of angiogenesis, make it a promising candidate in the search of new biomarkers for malignancy. In this study levels of PEDF were investigated in pleural effusions from lung adenocarcinoma and benign inflammatory disease patients. The mean PEDF concentration in the malignant group was slightly superior to that in patients suffering benign diseases (4.59 μg/mL vs 3.97 μg/mL), although the difference did not reach statistical significance (P 0.166). Pleural effusion PEDF levels were not related to gender, age, smoking habit or pleural effusion size. We also investigated the possible relationship of PEDF levels in pleural effusion regarding clinicopathological features. Correlations were found for monocytes (P 0.010) and polymorphonuclear leukocytes (P 0.023) with PEDF levels in pleural effusion of malignant origin.