Non-small-cell lung cancer (NSCLC) is the leading cause of cancer-related death. Developing minimally invasive techniques that can diagnose NSCLC, particularly at an early stage, may improve its outcome. Using microarray platforms, we previously identified 12 microRNAs (miRNAs) the aberrant expressions of which in primary lung tumors are associated with early-stage NSCLC. Here, we extend our previous research by investigating whether the miRNAs could be used as potential plasma biomarkers for NSCLC. We initially validated expressions of the miRNAs in paired lung tumor tissues and plasma specimens from 28 stage I NSCLC patients by real-time quantitative reverse transcription PCR, and then evaluated diagnostic value of the plasma miRNAs in a cohort of 58 NSCLC patients and 29 healthy individuals. The altered miRNA expressions were reproducibly confirmed in the tumor tissues. The miRNAs were stably present and reliably measurable in plasma. Of the 12 miRNAs, five displayed significant concordance of the expression levels in plasma and the corresponding tumor tissues (all r>0.850, all P<0.05). A logistic regression model with the best prediction was defined on the basis of the four genes (miRNA-21, -126, -210, and 486-5p), yielding 86.22% sensitivity and 96.55% specificity in distinguishing NSCLC patients from the healthy controls. Furthermore, the panel of miRNAs produced 73.33% sensitivity and 96.55% specificity in identifying stage I NSCLC patients. In addition, the genes have higher sensitivity (91.67%) in diagnosis of lung adenocarcinomas compared with squamous cell carcinomas (82.35%) (P<0.05). Altered expressions of the miRNAs in plasma would provide potential blood-based biomarkers for NSCLC.

Summary Adenocarcinoma is the most common type of lung cancer, the leading cause of cancer deaths in the world. Early detection is the key to improve the survival of lung adenocarcinoma patients. We have previously shown that microRNAs were stably present in sputum and could be applied to diagnosis of lung cancer. The aim of this study was to develop a panel of microRNAs that can be used as highly sensitive and specific sputum markers for early detection of lung adenocarcinoma. This study contained three phases: (1) marker discovery using microRNA profiling on paired normal and tumor lung tissues from 20 patients with lung adenocarcinoma; (2) marker optimization by real-time RT-qPCR on sputum of a case-control cohort consisting of 36 cancer patients and 36 health individuals; and (3) validation on an independent set of 64 lung cancer patients and 58 cancer-free subjects. From the surgical tissues, seven microRNAs with significantly altered expression were identified, of which “four” were overexpressed and “three” were underexpressed in all 20 tumors. On the sputum samples of the case-control cohort, four (miR-21, miR-486, miR-375, and miR-200b) of the seven microRNAs were selected, which in combination produced the best prediction in distinguishing lung adenocarcinoma patients from normal subjects with 80.6% sensitivity and 91.7% specificity. Validation of the marker panel in the independent populations confirmed the sensitivity and specificity that provided a significant improvement over any single one alone. The sputum markers demonstrated the potential of translation to laboratory settings for improving the early detection of lung adenocarcinoma.

Squamous cell carcinoma is a common form of lung cancer, the leading cause of cancer deaths in the world. Identifying early stage lung squamous cell carcinoma patients who would benefit most from effective therapies will reduce the mortality. We have previously shown that microRNAs (miRNAs) were stably present in sputum and potentially useful in diagnosis of lung cancer. The objective of this study was to develop a panel of miRNAs that can be used as a sputum-based test for early stage squamous cell carcinoma of the lungs. This study contained three phases: (1) marker discovery by profiling miRNA expression signatures on 15 lung squamous cell carcinoma and matched normal lung tissue samples with GeneChip miRNA Array; (2) marker optimization by real-time quantitative RT-PCR on sputum of a case-control cohort of 48 stage I lung squamous cell carcinoma patients and 48 healthy individuals; and (3) marker validation on an independent set including 67 lung squamous cell carcinoma patients and 55 healthy subjects. On the surgical tissues, six miRNAs were identified, of which three were overexpressed and three were underexpressed in all 15 tumors. On the sputum samples of the case-control cohort, three (miR-205, miR-210 and miR-708) of the six miRNAs were selected, which in combination produced the best prediction in distinguishing lung squamous cell carcinoma patients from normal subjects with 73% sensitivity and 96% specificity. Validation of the marker panel in the independent populations confirmed the sensitivity and specificity that provided a significant improvement over any single one alone. The sputum markers showed the potential to improve the early detection of lung squamous cell carcinomas.