Paolo Carbognani

Determination of aldehydes in exhaled breath of patients with lung cancer by means of on-fiber-derivatisation SPME–GC/MS

A number of volatile organic compounds (VOCs) have been identified and used in preliminary clinical studies of the early diagnosis of lung cancer. The aim of this study was to evaluate the potential of aldehydes (known biomarkers of oxidative stress) in the diagnosis of patients with non-small cell lung cancer (NSCLC). We used an on-fiber-derivatisation SPME sampling technique coupled with GC/MS analysis to measure straight aldehydes C3-C9 in exhaled breath. Linearity was established over two orders of magnitude (range: 3.3-333.3×10(-12) M); the LOD and LOQ of all the aldehydes were respectively 1×10(-12) M and 3×10(-12) M. Accuracy was within 93% and precision calculated as % RSD was 7.2-15.1%. Aldehyde stability in a Bio-VOC(®) tube stored at +4°C was 10-17 h, but this became >10 days using a specific fiber storage device. Finally, exhaled aldehydes were measured in 38 asymptomatic non-smokers (controls) and 40 NSCLC patients. The levels of all of the aldehydes were increased in the NSCLC patients without any significant effect of smoking habits and little effect of age. The good discriminant power of the aldehyde pattern (90%) was confirmed by multivariate analysis. These results show that straight aldehydes may be promising biomarkers associated with NSCLC, and increase the sensitivity and specificity of previously identified VOC patterns.

Management of postintubation membranous tracheal rupture

BACKGROUND Postintubation tracheobronchial laceration is a rare complication of general anesthesia. A renewed interest in this disorder induced us to review our experience on its treatment, focusing on the evolution of the surgical approach, and describing a technical variation of the transcervical approach. METHODS From January 1994 to December 2002 we treated 13 patients with diagnosis of postintubation tracheobronchial laceration. The treatment was nonsurgical in 3 patients (1-cm-long tear) and surgical in the other cases. Two lesions extending to the main bronchi were repaired through a right thoracotomy as well as four lesions limited to the trachea observed before January 2001. After this date we used the transcervical approach for entirely intratracheal lesions: in three cases we performed an anterior transverse tracheotomy and in one case a transverse and midline vertical incision (T tracheotomy). RESULTS Both conservative and surgical therapy were successful in all the cases. Two patients in the thoracotomy group had a transient right vocal cord palsy. No morbidity was observed with the cervical approach. Normal healing of the sutures was evidenced by an endoscopic follow-up 30 days later. CONCLUSIONS In our experience nonsurgical treatment is advisable in small (length < 2 cm) uncomplicated tears. Concerning surgery, thoracotomy is indicated in tracheal lacerations extending to the main bronchi, whereas the transcervical approach is preferred for intratracheal tears because of its efficacy in reaching and suturing the lesions extending to the carina and for its limited invasiveness.

Grading the neuroendocrine tumors of the lung: an evidence-based proposal.

Lung neuroendocrine tumors are catalogued in four categories by the World Health Organization (WHO 2004) classification. Its reproducibility and prognostic efficacy was disputed. The WHO 2010 classification of digestive neuroendocrine neoplasms is based on Ki67 proliferation assessment and proved prognostically effective. This study aims at comparing these two classifications and at defining a prognostic grading system for lung neuroendocrine tumors. The study included 399 patients who underwent surgery and with at least 1 year follow-up between 1989 and 2011. Data on 21 variables were collected, and performance of grading systems and their components was compared by Cox regression and multivariable analyses. All statistical tests were two-sided. At Cox analysis, WHO 2004 stratified patients into three major groups with statistically significant survival difference (typical carcinoid vs atypical carcinoid (AC), P=0.021; AC vs large-cell/small-cell lung neuroendocrine carcinomas, P<0.001). Optimal discrimination in three groups was observed by Ki67% (Ki67% cutoffs: G1 <4, G2 4-<25, G3 ≥25; G1 vs G2, P=0.021; and G2 vs G3, P≤0.001), mitotic count (G1 ≤2, G2 >2-47, G3 >47; G1 vs G2, P≤0.001; and G2 vs G3, P≤0.001), and presence of necrosis (G1 absent, G2 <10% of sample, G3 >10% of sample; G1 vs G2, P≤0.001; and G2 vs G3, P≤0.001) at uni and multivariable analyses. The combination of these three variables resulted in a simple and effective grading system. A three-tiers grading system based on Ki67 index, mitotic count, and necrosis with cutoffs specifically generated for lung neuroendocrine tumors is prognostically effective and accurate.

Comparison Between Epidermal Growth Factor Receptor (EGFR) Gene Expression in Primary Non-small Cell Lung Cancer (NSCLC) and in Fine-Needle Aspirates from Distant Metastatic Sites

Purpose: Epidermal growth factor receptor (EGFR) gene copy number obtained by fluorescence in situ hybridization (FISH) has been recently found to predict treatment outcome in non-small cell lung cancer (NSCLC) patients receiving EGFR tyrosine kinase inhibitors. However, it is still unknown whether EGFR status differs in metastases compared with primary NSCLC. In all studies FISH have been performed on histologic material. The possibility to perform FISH analysis on cytologic material obtained by fine-needle aspiration from superficial and visceral metastases would allow us to know the real EGFR status avoiding invasive diagnostic procedures. Methods: EGFR gene copy number was analyzed by FISH on fine-needle aspirates obtained from 31 patients with metastatic NSCLC and the results were compared with those obtained on corresponding paraffin histologic sections from the primary tumor. Results: The feasibility of EGFR FISH on cytology was 90% (28 of 31 patients). EGFR FISH was positive in 61% (17 of 28 patients) of the metastases and in 36% (10 of 28 patients) of the primary tumors. Nine of the 28 cases (32%) were EGFR positive on both primary tumor and metastatic site and 10 (36%) were negative on both primary tumor and metastasis. Nine of the 28 cases (32%) showed discordance of primary tumor versus metastasis (McNemar test; p = 0.041). Conclusions: EGFR FISH can be reliably assessed on fine-needle aspirates obtained from NSCLC metastases. We found that EGFR gene copy number is discordant between primary NSCLC and the corresponding distant metastatic sites in a significant proportion of cases. These findings should be considered in future studies designed to elucidate the predictive role of EGFR FISH in NSCLC.

Lung metastasis resection of adenoid cystic carcinoma of salivary glands

BACKGROUND Adenoid cystic carcinoma is a rare tumour originating from the exocrine mucous glands, known for its high propensity for distant metastases. The value of lung metastasis resection from adenoid cystic carcinoma of salivary glands origin is evaluated. METHODS A retrospective study was conducted on patients undergoing surgery for primary adenoid cystic carcinoma of the salivary glands between 1982 and 2006. Patients were excluded who had primary tumour macroscopic incomplete resection or were lost at follow-up. From a database of 50 eligible patients, 27 were identified as having presented a tumour recurrence during follow-up; in 20 it was first diagnosed in the form of distant metastases, and in 7 in the form of loco-regional recurrence. Nine patients who presented isolated lung recurrence underwent complete lung metastasectomy. Demographic data, pathologic characteristics and operative and postoperative record were reviewed, as well as updated survival. RESULTS Twenty-six men and 24 women with a median age of 57 years (range 33-79) underwent radical surgery for adenoid cystic carcinoma during the study period. In 20 patients, at a median free interval time of 3 years (range 1-12), a distant metastasis relapse was observed. Nine patients with a median free interval time of 5 years (range 1-12) underwent lung metastasectomy: five had single metastasis resection, one multiple mono-pulmonary and three multiple and bilateral. In six of these patients a new disease recurrence was noted: four patients underwent further lung metastasectomy, but in all of them progression of the disease was observed. Mean survival of the population as a whole resulted as being 16 years (SE=1.4) with an actuarial survival of 77% at 5 years, 66% at 10 years and 56% at 15 years. Mean survival of patients having presented with distant metastases resulted as being 11 years (SE=2.2). Mean survival after appearance of distant metastases resulted as being 72 months (SE=15.8) in the 9 patients treated by metastasectomy, and 62 months (SE=15.1) in the 11 who did not have metastasis resection. CONCLUSIONS Patients with adenoid cystic carcinoma could be frequently encountered with disease recurrence confined to the lung. The impact of complete lung metastasis resection on the course of the disease, however, is yet to be determined.

Dose-dependent effect of FHIT-inducible expression in Calu-1 lung cancer cell line

Abnormalities in the expression of the tumour suppressor fragile histidine triad (FHIT) gene have been reported in a variety of human tumours, including lung cancer and restoration of its expression in cancer cell lines resulted in the inhibition of proliferation and apoptosis induction. Most of the studies that have assigned a proapoptotic role to the FHIT gene were performed in adenoviral-FHIT-transduced cancer cells expressing high levels of the Fhit protein. The present work was the first study designed to investigate the effects of FHIT gene replacement in a human FHIT-negative non-small-cell lung cancer (NSCLC) cell line (Calu-1) by using a hormone-inducible expression system that allows tight modulation of the transgene expression. Through this approach, we demonstrated that a prolonged induction was required to accumulate the Fhit protein at levels adequate to promote a significant decrease of cell proliferation. Analysis of cell-cycle phase distribution showed an accumulation of cells in the G0/G1 phase and a concomitant decrease in the S phase. Moreover, an upregulation of p21waf1 transcript was found, which could account for the alteration of the cycling properties of the cells. The growth-inhibitory effects observed were not associated with apoptosis appearance, and although in these conditions the Fhit protein content was higher than in normal bronchial human epithelial cells (NHBE), it was still significantly lower than the level capable of inducing apoptosis in Calu-1 cells after adenoviral-mediated FHIT gene transfer. These results indicate that the tumour suppressor properties of Fhit are strictly related to its expression level and show that the Fhit protein has a dose-dependent antiproliferative effect on the Fhit-negative Calu-1 lung cancer cell line.

Plasma and EBC microRNAs as early biomarkers of non-small-cell lung cancer

Abstract Lung cancer is a major cause of death in Western countries. Current screening methods are invasive and still lead to a high percentage of false positives. There is, therefore, a need to find biomarkers that increase the probability of detecting lung cancer early. MicroRNAs (miRNAs) are stable molecules in blood plasma and exhaled breath condensate (EBC). We quantified miRNA-21 and miRNA-486 expression from plasma and EBC samples from patients with a diagnosis of non-small-cell lung cancer (NSCLC) and controls. miRNA-21 was significantly higher in plasma and in EBC of the NSCLC patients and miRNA-486 was significantly lower. This difference indicates a significantly improved diagnostic value, and suggests that these miRNAs could be clinically used as a first-line screening test in high-risk subjects.

Reliability of quantitative computed tomography to predict postoperative lung function in patients with chronic obstructive pulmonary disease having a lobectomy.

Objective: To verify the reliability of quantitative computed tomography (CT) to estimate the postoperative lung function in patients with mild to severe chronic obstructive pulmonary disease (COPD) who underwent a lobectomy. Methods: Nine COPD patients with lung cancer having a lung lobectomy with preoperative CT were enrolled. By applying a density mask technique and a specific equation, predicted postoperative forced expiratory volume in 1 second (FEV1) and vital capacity (VC) were calculated. Predicted values were correlated with postoperative measured values. Results: Estimated FEV1 and VC were always significantly lower than the corresponding postoperative values; however, CT-estimated postresection FEV1 values were better than the postresection VC values (biases between estimated and measured values were −0.14 and −0.536 L, respectively, according to the Bland-Altman method). Quantitative CT predicted postoperative FEV1 (r = 0.97, P < 0.001) and VC (r = 0.93, P < 0.001) well in all patients, however. Conclusions: Quantitative CT may be an alternative tool to perfusion scan to predict postresection lung function, even in patients with borderline pulmonary function undergoing a lobectomy.

Experimental Tracheal Transplantation Using a Cryopreserved Aortic Allograft

Background: The tracheal reconstruction after wide resections remains a critical surgical problem. Our aim was to replace trachea with a tissue easy to vascularize, which allows a simple reconstruction and does not require an immunosuppressive regimen. Materials and Methods: A segment of cryopreserved aorta was used in order to verify its adequacy as tracheal substitute. In phase 1, the thoracic aorta of 10 rabbits was excised, obtaining 20 segments that were cryopreserved. Ten segments were implanted in the omentum of 10 rabbits that were sacrificed on postoperative days 7, 14 and 21, and the grafts were examined histologically. In phase 2, a segment of cryopreserved aorta arranged with a silicone prosthesis was transplanted in 10 rabbits and wrapped with omentum. The animals were sacrificed on postoperative days 7, 14 and 21. Results: In phase 1, the neovascularization of the grafts was present after 7 days, and after 14 days the fibroblasts invaded the lumen of the aorta. In phase 2, 8 rabbits survived and the histologic examination after 7, 14 and 21 days showed neovascularization, the absence of rejection and the proliferation of fibroblasts inside the lumen of the aorta; this growth has been restrained by an endoluminal prosthesis. Conclusions: Our study demonstrated that replacing the trachea with cryopreserved aorta is technically feasible and does not evoke immunologic reactions. It requires, however, a silicone tube inside the allograft to limit the colonization of fibroblasts.

Exercise capacity assessment in patients undergoing lung resection

BACKGROUND The value is examined of preoperative functional assessment, including exercise capacity measurement by a cycloergometric maximal exercise test, in the prediction of postoperative cardio-pulmonary complication after lobar resection. METHODS In a prospective study over a 3-year period, all patients who were candidates for lung resection underwent preoperative functional evaluation by means of resting pulmonary function tests, measurement of the lung diffusing capacity for carbon monoxide and cardio-pulmonary exercise test. Patients who had had pneumonectomy or less than anatomical segmentectomy were excluded. The study population consisted of 73 patients. The postoperative morbidity and mortality record was collected. RESULTS Sixty-four patients underwent lobectomy, five bilobectomy and four segmentectomy. Indication for surgery was NSCLC in 71 cases. Two postoperative deaths were recorded (2.7%). A pulmonary (n=19) and/or cardiac (n=17) complication was scored in 30 patients (41%). Mean preoperative FEV(1) and VO(2)max of patients who developed pulmonary complications were significantly lower (p=0.013 and p=0.043 respectively) than those of patients without pulmonary complications. Logistic regression analysis found FEV(1) to be an independent factor in pulmonary complication (p=0.002). With regard to pulmonary complication occurrence, the receiver operating characteristic curve showed an area of 0.69 with VO(2)max expressed in ml/kg min and of 0.62 when VO(2)max was expressed as a percentage of the predicted value. The widest point of the curve was found at a VO(2)max value of 18.7 ml/kg min. Six out of the 14 patients (43%) with a preoperative VO(2)max equal to or lower than 15 ml/kg min had a pulmonary complication. No functional preoperative identifiers were found for the 16 patients who presented with postoperative new onset atrial fibrillation. The mean preoperative value of carbon monoxide lung diffusing capacity was significantly lower (p=0.037) in the 30 patients who had postoperative cardio-pulmonary complications than in the complication-free population. CONCLUSIONS Preoperative exercise capacity assessment helps in stratifying patients at risk for postoperative pulmonary complication. However, it does not appear to be an independent prognostic factor for postoperative outcome.