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Gains of 3q26 chromosome region, where the human telomerase RNA gene (hTERC) is located, have been previously documented in cervical carcinomas and preneoplastic lesions. The aim of our study was to define the value of 3q26 gains related to persistence-progression in cervical specimens with cytologic diagnosis for low-grade squamous intraepithelial lesions, using liquid-based cytology (ThinPrep; Hologic, Marlborough, MA) and fluorescence in situ hybridization. For these purposes, 55 patients were included in the study: 25 cases with a negative cytologic diagnosis for squamous intraepithelial lesion or malignancy (20 premenopausal and 5 postmenopausal women, used as control negative cases) and 30 low-grade squamous intraepithelial lesion cases. The follow-up was performed using cytology at 6, 12, and 24 months after the low-grade squamous intraepithelial lesion diagnosis. When the cytology result showed a high-grade lesion, colposcopy and biopsy were performed. Fluorescence in situ hybridization technique with a 3q26 2-color commercial probe was performed to determine the number of hTERC copies. There were no differences between premenopausal and postmenopausal normal cases. Low-grade squamous intraepithelial lesion cases with regression in the follow-up at 6, 12, and 24 months showed a percentage of cells with 3q26 gains similar to the control cases and lower than low-grade squamous intraepithelial lesion cases with persistence or progression (P < .05). Fluorescence in situ hybridization results were similar in preserved and frozen samples. However, in frozen samples, the number of cells suitable to be evaluated by fluorescence in situ hybridization was lower than in preserved (nonfrozen) cases. In conclusion, the determination by fluorescence in situ hybridization of 3q26 gains in low-grade squamous intraepithelial lesion cases could be useful to predict the persistence-progression of such cervical lesions using both preserved and frozen cervical material.

Introduction Transbronchial needle aspiration (TBNA) is a bronchoscopic technique that has been shown to be useful for sampling mediastinal lymph nodes. The yield of this technique can be increased by using endobronchial ultrasound (EBUS) to guide needle placement. The aim of the present study was to compare the yield of EBUS-guided TBNA to that of conventional TBNA in the analysis of mediastinal lymph nodes. Patients and Methods All patients undergoing either EBUS-guided or conventional TBNA for the diagnosis of mediastinal lymph nodes between January 2006 and May 2007 were studied consecutively. Histology results were used as a reference standard in patients treated surgically. In cases in which surgery was not indicated, the results of cytology or of clinical follow-up of at least 6 months duration were used. Results TBNA was performed in 117 patients and a total of 143 lymph nodes were punctured (mean shortest [SD] diameter, 17.9 [8] mm). The samples obtained were diagnostic in 58 patients (49.6%) and in 70 lymph nodes (49.0%). For paratracheal and hilar stations, the yield of EBUS-guided TBNA was superior to that of conventional TBNA (59.2% compared to 34.1%, P=.02). Conclusions Radial EBUS guidance increases the diagnostic yield of TBNA in paratracheal and hilar lymph node stations.

Aim: Disappointing results have been observed in advanced NSCLC patients receiving MET inhibitors based on histological or immunohistochemical criteria. The aim of this study was to assess the impact of MET intratumor heterogeneity on MET evaluation and potentially on adequate selection of nsNSCLC patients for clinical trials. Methods: Samples were obtained from a cohort of 120 patients. Of these, 47 were incorporated in a tissue microarray (TMA), selecting 4 histologically distinct areas of the tumor for heterogeneity studies. Assessment of Met expression was performed by IHC (SP44 Ventana) using H-score and MetMab criteria. FISH was used to assess gene copy number (GCN) (MET and CEP7 probes from Vysis). Using different scoring methods, correlation with clinico-pathologic and molecular features (KRAS, EGFR) and between Met expression and GCN was investigated. Heterogeneity of Met status between different tumor cores was examined. Results: Seventy percent of the patients were male and 55% were current smokers. Median age was 66 years and 36% were stage IV nsNSCLC. Of 127 samples, 90% were adenocarcinomas. Nineteen per cent and 11% harbored KRAS and EGFR mutations, respectively. Forty-eight per cent were MET-High according to MetMab criteria, and median H-score was 140 (range 0-400). Increased Met expression was associated with advanced stage (p = 0.001) and with former or no smoking history (p = 0.038). Mean MET GCN was 2.8 and 9 cases were considered FISH-positive (GCN > =5). Increased MET GCN was associated with solid histological pattern. Met H-score was directly associated with GCN (p = 0.036). Intra-class correlation coefficient between 4 tissue cores was 0.57, both for Met H-score and GCN. In three patients considered FISH-positive, 4 out of 12 cores were FISH-negative. Conclusions: Classification of Met-driven tumors is challenging due to lack of robustness of methods and criteria for defining Met positivity. MET heterogeneity may hinder adequate patient selection for clinical trials with MET inhibitors. Disclosure: All authors have declared no conflicts of interest.

Aim: Recently, c-ros oncogene 1, receptor tyrosine kinase (ROS1, 6q22) has been identified as a driver oncogene in non-small-cell lung carcinomas (NSCLC). Patients carrying ROS1 translocations are candidates to targeted therapy with crizotinib®. We aimed to determine the prevalence, patterns and fusion partners in ROS1-rearranged NSCLC samples and to characterize ROS1 copy number alterations (CNAs) by fluorescence in situ hybridization (FISH). Methods: A total of 216 NSCLC patients (median age 64 years, 66% males, 51% smokers, 73% adenocarcinomas (ADC), 35% stage 4), were screened for ROS1 abnormalities using a break-apart FISH probe (Abbott Molecular). To characterize ROS1 partners, clones from Human 32K BAC Re-Array Library were used to develop CD74, SLC34A2 and EZR probes. Immunohistochemistry (IHC) with ROS1 D4D6 antibody (Cell Signalling Technology) was also performed in ROS1-translocated cases. Correlation with clinico-pathologic information was investigated. Results: Four samples were ROS1 translocated (1.9%): three showed deletion of the non-translocated allele and one had a 5'ROS1 deletion. One harbored a SLC34A2-ROS1 fusion and another a CD74-ROS1. No fusion partner was identified in the other two samples. All cases were positive for ROS1 IHC staining, three presenting a cytoplasmic pattern and one with membranous pattern. ROS1-translocated tumors were ADC, all EGFR, KRAS, and ALK wt. We observed an association with non-smoking habit (P=0.036). Regarding CNAs in ROS1 non-rearranged cases, 43.5% had gains (3-6 copies) and 26.4% had deletions. Conclusions: We confirm the low prevalence of ROS1-translocated lung ADC in a Spanish population. Moreover, our study reveals a significant frequency of ROS1 deletion as a potentially relevant aberration, also in ROS1-translocated patients. Disclosure: All authors have declared no conflicts of interest.