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Additional analyses on IEO and TCGA cohorts

Molecular characteristics of different groups of TCGA lung adenocarcinoma patients

Supplemental Legends Supplemental Table 1.TaqMan Assays Supplemental Table 2. 10-gene model Supplemental Table 3. Cut-offs Supplemental Table 4. Clinical and pathological characteristics of Stage II-III cohort Supplemental Table 7. Causes of death in Training and Validation Sets Supplemental Table 8. Stage I - C1 smoking status Supplemental Table 12A. KEAP1 Mutation in TCGA patients Supplemental Table 12B. KEAP1 copy-number variants in TCGA patients Supplemental Table 12C. KEAP1 promoter methylation in TCGA patients Supplemental Table 13. Mutation, Copy Number Variation and Methylation in C1-patients (TCGA) of KEAP1 and NRF2 genes. Supplemental Table 14. Patient and tumor characteristics of a subgroup of 50 patients from the stage I IEO cohort.

KEAP1 and NFR2 target genes analyses

Purpose: The National Lung Cancer Screening Trial has confirmed that lung cancer mortality can be reduced if tumors are diagnosed early, that is, at stage I. However, a substantial fraction of stage I lung cancer patients still develop metastatic disease within 5 years from surgery. Prognostic biomarkers are therefore needed to identify patients at risk of an adverse outcome, who might benefit from multimodality treatment.Experimental Design: We extensively validated a 10-gene prognostic signature in a cohort of 507 lung adenocarcinoma patients using formalin-fixed paraffin-embedded samples. Furthermore, we performed an integrated analysis of gene expression, methylation, somatic mutations, copy number variations, and proteomic profiles on an independent cohort of 468 patients from The Cancer Genome Atlas (TCGA).Results: Stage I lung cancer patients (N = 351) identified as high-risk by the 10-gene signature displayed a 4-fold increased risk of death [HR = 3.98; 95% confidence interval (CI), 1.73–9.14], with a 3-year overall survival of 84.2% (95% CI, 78.7–89.7) compared with 95.6% (92.4–98.8) in low-risk patients. The analysis of TCGA cohort revealed that the 10-gene signature identifies a subgroup of stage I lung adenocarcinomas displaying distinct molecular characteristics and associated with aggressive behavior and poor outcome.Conclusions: We validated a 10-gene prognostic signature capable of identifying a molecular subtype of stage I lung adenocarcinoma with characteristics remarkably similar to those of advanced lung cancer. We propose that our signature might aid the identification of stage I patients who would benefit from multimodality treatment. Clin Cancer Res; 23(1); 62–72. ©2016 AACR.

IPA analysis of the NFE2L2 modulator

Gene expression analysis of other lung cancer datasets and prognostic signatures

Optimization of the 10-gene signature screening protocol in FFPE samples and effect of long-term storage of FFPE samples on RNA quality

BACKGROUND Lung ischemia/reperfusion (IR) injury contributes to the development of severe complications in patients undergoing transplantation. Mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) exert beneficial actions comparable to those of MSCs without the risks of the cell-based strategy. This research investigated EV effects during IR injury in isolated rat lungs. METHODS An established model of 180-minutes ex vivo lung perfusion (EVLP) was used. At 60 minutes EVs (n = 5) or saline (n = 5) were administered. Parallel experiments used labeled EVs to determine EV biodistribution (n = 4). Perfusate samples were collected to perform gas analysis and to assess the concentration of nitric oxide (NO), hyaluronan (HA), inflammatory mediators, and leukocytes. Lung biopsies were taken at 180 minutes to evaluate HA, adenosine triphosphate (ATP), gene expression, and histology. RESULTS Compared with untreated lungs, EV-treated organs showed decreased vascular resistance and a rise of perfusate NO metabolites. EVs prevented the reduction in pulmonary ATP caused by IR. Increased medium-high-molecular-weight HA was detected in the perfusate and in the lung tissue of the IR + EV group. Significant differences in cell count on perfusate and tissue samples, together with induction of transcription and synthesis of chemokines, suggested EV-dependent modulation of leukocyte recruitment. EVs upregulated genes involved in the resolution of inflammation and oxidative stress. Biodistribution analysis showed that EVs were retained in the lung tissue and internalized within pulmonary cells. CONCLUSIONS This study shows multiple novel EV influences on pulmonary energetics, tissue integrity, and gene expression during IR. The use of cell-free therapies during EVLP could constitute a valuable strategy for reconditioning and repair of injured lungs before transplantation.

Background Bilateral lung neoplasms are becoming more common, but distinguishing two primary lung cancers from metastatic disease can be difficult and only long-term follow-up after treatment can disclose the real behavior of the disease.The present study aimed to identify the appropriate candidates for bilateral surgical resections from the perspective of short-term postoperative complications and long-term outcomes. Methods Two hundred and fifteen consecutive patients undergoing bilateral pulmonary resection for lung cancers over a 20-year period were analyzed. Preoperative patient characteristics were noted, including demographic information, operative details, pathologic information including histology and tumor stage according to the eighth edition of the tumor nodes metastases staging system, and the use of neoadjuvant or adjuvant treatments. Results Patients receiving the second pulmonary resection more than 24 months from the first procedure as well as patients receiving bilateral lobectomies had higher overall 3-, 5-, and 10-year survival rates compared with the others. Conclusion Patients receiving the second resection more than 24 months from the first procedure have the best long-term results irrespective of the type of resection.

Despite increasing clinical adoption, biologic influences of ex vivo lung perfusion (EVLP) remain insufficiently elucidated. The aim of the current study was to investigate biomolecular changes induced by EVLP in rat lungs. EVLP was maintained for 180 min. Hyaluronan, mediators, and cells were assessed in the perfusate. Gene expression, signaling pathways, and ATP content were investigated in lung tissue. EVLP induced the release of medium-high molecular weight hyaluronan and transcription of hyaluronan synthases ( P < 0.001). Increasing concentrations of inflammatory mediators were detected in the perfusate ( P < 0.001). Perfused lungs exhibited a distinctive transcriptional signature compared with organs examined before or after surgery/procurement ( P = 0.003). Up-regulated genes were involved in inflammation and its regulation, apoptosis/survival, heat shock, and oxidative stress response ( q = 0). Down-regulated genes were related to lymphocyte function ( q = 0). The NF-κB, signal transducer and activator of transcription 3, ERK1/2, p38, Akt, and stress-activated protein kinase/JNK signaling pathways were modulated by EVLP ( P < 0.05). Most of these biomolecular changes were examined and confirmed in additional experiments that were performed in lungs procured from donation after cardiocirculatory death after 180 min of warm ischemia. The current study demonstrates that EVLP broadly affects the lung biomolecular phenotype. These findings improve our comprehension of the effects exerted by the procedure and encourage additional research in preclinical models to implement therapeutic interventions.-Lonati, C., Bassani, G. A., Brambilla, D., Leonardi, P., Carlin, A., Faversani, A., Gatti, S., Valenza, F. Influence of ex vivo perfusion on the biomolecular profile of rat lungs.

Objectives While the best approach to malignant pleural mesothelioma has yet to be demonstrated, surgery remains the mainstay of treatment. We analysed a group of candidates for trimodality treatment, aiming to identify prognostic factors guiding patient selection. Methods Between 2003 and 2015, 83 (31.6%) of the 283 patients with malignant pleural mesothelioma were considered for trimodality treatment to perform induction chemotherapy, extrapleural pneumonectomy and adjuvant radiotherapy. All patients underwent cisplatin-based chemotherapy. Radiotherapy was administered at a mean dose of 50.4 Gy. Results Thirty-six patients (43.4%) had 3 cycles of chemotherapy, whereas 21 (25.3%) had more than 3. Progression to chemotherapy was observed in 10.9% (9 of 83) of patients, partial response in 30.1% (25 of 83) and stable disease in 59% (49 of 83). Sixty-three patients underwent extrapleural pneumonectomy. Fifty-five patients (87.3%) had epithelial tumour. Forty-two patients (66.7%) were in pathological Stage 3. Major complications after extrapleural pneumonectomy were observed in 28 patients (44.4%), whereas 30-day postoperative mortality was 11.1% (7/63). Radiotherapy was not administered in 24 patients (38.1%) due to major complications after surgery or patient intolerance. Two patients (3.2%) died within 90 days after the end of radiotherapy. The trimodality treatment was completed in 37 (44.6%) patients. Median overall survival was 35.6 months, with 1- and 3-year overall survival of 82% and 48% for patients who completed the trimodality treatment compared with 32% and 14% for patients who did not undergo radiotherapy. Conclusions Only 45% of patients completed the planned trimodality treatment, and morbidity/mortality remained high. Nonetheless, the patients who completed treatment showed good loco-regional disease control and better overall survival.

Purpose: The National Lung Cancer Screening Trial has confirmed that lung cancer mortality can be reduced if tumors are diagnosed early, that is, at stage I. However, a substantial fraction of stage I lung cancer patients still develop metastatic disease within 5 years from surgery. Prognostic biomarkers are therefore needed to identify patients at risk of an adverse outcome, who might benefit from multimodality treatment. Experimental Design: We extensively validated a 10-gene prognostic signature in a cohort of 507 lung adenocarcinoma patients using formalin-fixed paraffin-embedded samples. Furthermore, we performed an integrated analysis of gene expression, methylation, somatic mutations, copy number variations, and proteomic profiles on an independent cohort of 468 patients from The Cancer Genome Atlas (TCGA). Results: Stage I lung cancer patients (N = 351) identified as high-risk by the 10-gene signature displayed a 4-fold increased risk of death [HR = 3.98; 95% confidence interval (CI), 1.73–9.14], with a 3-year overall survival of 84.2% (95% CI, 78.7–89.7) compared with 95.6% (92.4–98.8) in low-risk patients. The analysis of TCGA cohort revealed that the 10-gene signature identifies a subgroup of stage I lung adenocarcinomas displaying distinct molecular characteristics and associated with aggressive behavior and poor outcome. Conclusions: We validated a 10-gene prognostic signature capable of identifying a molecular subtype of stage I lung adenocarcinoma with characteristics remarkably similar to those of advanced lung cancer. We propose that our signature might aid the identification of stage I patients who would benefit from multimodality treatment. Clin Cancer Res; 23(1); 62–72. ©2016 AACR.

Background: Choriocarcinoma is a rare malignant disease that is usually associated with a gestational event. Lung metastasis with no evident primary origin and choriocarcinoma, which mimics features of non-small-cell lung cancer, might be misdiagnosed as adenocarcinoma or large-cell carcinoma. This is a pivotal clinical concern since the tumor can lead to various symptoms, seriously affecting the quality of life and can escalate rapidly, with a high mortality rate, compared to lung cancer. Methods: We reported a case of a 37-year-old woman with a history of one-year enhancement of beta-human chorionic gonadotropin levels and only a single nodule in the right upper lobe, with no abnormal findings on the gynecological investigation. Then we retrospectively examined all cases treated in the Division of Thoracic Surgery at the European Institute of Oncology in the last twenty years (from 1998 to 2018). Results: This was the first time in our experience that choriocarcinoma presentation was with a single nodule without a gynecological finding. Moreover, the differential diagnosis between lung carcinoma and choriocarcinoma was achieved only after surgical removal. Conclusions: As confirmed by our literature search, precise and expedited differential diagnosis is essential in choriocarcinoma care (both with single or multiple metastases), to successfully remove the tumor and increase the patient&rsquo, s chances of survival.

Background In stage III to IVa thymic epithelial tumors (TETs), infiltration of the superior vena cava (SVC) is not rare. The extent of SVC resection depends on the width of the area of neoplastic invasion. Our article aims to evaluate the safety and long-term outcomes of extended thymectomy for TETs with SVC resection compared with advanced-stage TETs patients without SVC resection. Methods Retrospective review of the experience on patients who underwent extended thymectomy for TETs in the last 20 years, according to STROBE (Strengthening the Reporting of Observational Studies in Epidemiology) methodology. Progression-free survival (PFS) and overall survival (OS) were calculated using the Kaplan-Meier method. A backward stepwise Cox regression multivariate analysis was performed to determine factors associated with long-term outcomes. Results A total of 78 patients underwent surgery for advanced-stage TETs (Masaoka-Koga stages III-IVa) from January 1998 to April 2019. Fourteen (17.9%) underwent thymectomy with resection of SVC. Presence of a thymic carcinoma (hazard ratio , 2.26; 95% confidence interval, 1.82-6.18; P = .038) and the SVC resection (hazard ratio, 1.89; 95% confidence interval, 1.11-3.96; P = .041) were adverse prognostic factors at multivariate analysis. The median OS and the PFS of all SVC resected patients were 50 (range, 5-207) months and 31 (range, 5-151) months, respectively. There was no significant difference in OS (P = .28) and PFS (P = .32) between SVC-resected and non–SVC-resected patients. Conclusions SVC resection is a safe and effective procedure to restore the venous system continuity and does not seem to affect survival and disease recurrence. This surgical approach allows radical resection of locally advanced TETs, even after neoadjuvant chemotherapy.

Background This study analyzed the short- and long-term outcomes of robotic-assisted thoracic surgery (RATS) for early stage non-small cell lung cancer (NSCLC). Methods From November 2006 to December 2016, we performed 363 RATS procedures. This study retrospectively reviewed 339 patients who underwent RATS for clinical stages I (n = 318) or II (n = 21) NSCLC. Results Twenty-nine patients underwent segmentectomy, 307 lobectomy, and 3 pneumonectomy. Conversion occurred in 22 patients (6.5%): 15 (4.4%) due to technical issues, 4 (1.2%) for oncological reasons, and 3 (0.9%) for bleeding. The median number of N1 and N2 stations resected was 2 and 3, respectively, and the median number of N1 and N2 lymph nodes resected was 9 and 6, respectively. Median operative time was 192 minutes for lobectomy, 172 minutes for segmentectomy, and 275 minutes for pneumonectomy. Median length of hospital stay was 5 days (2–191). The most common postoperative complication was prolonged air leak (12.1%). Major complications occurred in eight patients (2.4%). The 30-day and 90-day operative mortality was 0% and 0.3%, respectively. Two and 5-year cancer-specific survival rate was 96.1% and 91.5%, respectively. Five-year survival rate was 96.2% for patients who underwent segmentectomy, and 89.1% for lobectomy. All three patients who underwent pneumonectomy were alive at 5 years with no disease. Conclusions Besides the well-known short-term outcomes showing very low morbidity and mortality rates, mediastinal lymph node dissection during RATS adequately assesses lymph node stations detecting occult lymph node metastasis and leading to excellent oncologic results. However, these results await longer follow-up studies.

Background Prolonged air leak is one of the most common complications after lung surgery and the cause of prolonged hospital stay frequently associated with major postoperative morbidity and thus responsible for even higher hospital costs. This case-control study was designed to test the sealing efficacy and safety of Enable-Innoseal TP4 in patients undergoing pulmonary resection for lung cancer. Methods This was a case-control trial enrolling patients with primary or single site metastatic lung cancer scheduled for elective anatomic or nonanatomic pulmonary resection presenting intraoperative grade 1 or 2 air leak at water submersion test; the study group population was then matched 1:1 according to surgical procedure, male/female ratio, preoperative FEV1, and age. Results In the study population, 21 patients (70.0%) presented intraoperative grade 1 air leak and 9 patients grade 2 (30.0%) air leak; after comparison with the control group, we observed a significant shorter time for chest drain removal in the study population ( P = 0.0050), whereas no difference was registered in terms of number of days needing for discharge ( P = 0.0762). Conclusions Enable-Innoseal TP4 was effective in treating limited intraoperative air leaks after pulmonary resection and preventing prolonged postoperative air leaks in patients receiving either anatomic or nonanatomic lung resections. Further randomized double-arm studies are required to confirm the efficacy of Enable-Innoseal TP4 demonstrated by this pilot study.

The aim of this study was to analyse the feasibility and safety of robotic-assisted thymectomy (RoT) in patients with clinically early stage thymoma, investigating clinical and early oncological results. Between 1998 and 2017, we retrospectively reviewed 76 (42.2%) patients who underwent radical thymectomy for clinically early stage thymoma (Masaoka-Koga I and II), identifying all patients who underwent RoT (n = 28) or open thymectomy (OT) with eligibility criteria for robotic surgery (n = 48). Using a propensity-score matched for tumor size (3.9 ± 1.8 cm) and stage (35% stage I, 42% stage IIA, 23% stage IIB), we paired 24 patients who had RoT with 24 patients undergoing OT. RoT was left-sided in 19 (79.2%) patients. None of the patients required conversion to open surgery. OT was via sternotomy in 21 (87.5%) patients and thoracotomy in 3 (12.5%). Mean operating time was shorter in the RoT group (117 ± 40 min) than in the OT (141 ± 46 min) (p = 0.06), even if not statistically significant. Length of stay was significantly shorter in the RoT group (mean 4.0 ± 1.9 days) than in the OT (mean 5.9 ± 1.7 days) (p = 0.0009). No significant difference between the two groups regarding post-operative complications. Five patients died in the OT group after a median follow-up of 6.1 years (only one for recurrence). After a median follow-up of 1.3 years, all patients in the RoT group were alive without disease. RoT is feasible and safe for early stage thymoma with clear advantage compared to OT in term of short term outcomes. A longer follow-up is needed to better evaluate the oncological results.

Carinal resection (CR) is defined as the resection of the trachea-bronchial bifurcation with or without lung resection. It is an uncommon challenging surgery performed in case of NSCLC, primitive airway tumors or benign lesions invading the carina. A well-organized team is essential to manage patients undergoing CR and it must involve several specialists experienced in diagnosis, operative treatment and postoperative care. Before and during surgery a strict cooperation between surgeon and anesthesiologist is essential; cross-field ventilation is generally used to maintain the adequate gas exchange during surgical airway reconstruction, but also high frequency jet ventilation (HFJV) or extracorporeal membrane oxygenation (ECMO) could be valid alternative options when cross-field ventilation is not feasible. Right-sided lesions requiring a CR with pulmonary resection are better approached through an ipsilateral thoracotomy (IV intercostal space), whereas tumor involving the carina as well as the left main bronchus and requiring a left carinal pneumonectomy could be treated using a left thoracotomy with subaortic dissection (only for very limited tracheal resection due to a bad exposure of the trachea after moving the aortic arch). Instead, CR without pulmonary resection and left carinal pneumonectomy are better approached through a median sternotomy. Reconstruction of the airway could be performed in different ways according to the extension of the resection and to the surgeon experience with the sole purpose to obtain a tension-free anastomosis to reduce as much as possible the possible post-operative complications due to impairment healing of the suture. Based on tumor histology and the pathological staging the patient should be referred to the oncologist and/or radiotherapist for the further medical treatments. Improved patient selection, anesthetic management, surgical technique and better postoperative management are essential in such a challenging surgery to have the lowest possible rate of postoperative morbidity and mortality.

Background The role of extended pulmonary resection for lung metastases is still unclear, and little information is available in the literature. This study was performed to analyze the outcomes and prognostic factors of patients who underwent extended resections for pulmonary metastases. Methods From 1998 to 2013, 1027 patients underwent lung metastasectomy procedures. Twenty-nine patients had extended pulmonary resections: three resections of the chest wall, one azygos, one diaphragm, four vascular resections/reconstructions, six sleeve resections, and 14 pneumonectomies. Results Extended resection was performed for metastatic disease mainly from epithelial (62.1%) and sarcomatous (20.7%) tumors. Complete resection was obtained in all patients. Thirty-day operative morbidity and mortality rates were 38% (11 of 29) and 0%, respectively. Only one patient had a major complication due to a bronchopleural fistula. Mean hospital stay was 6.3 days. After a median follow-up of 27 months, 16 patients (55%) had died. At univariate analysis, survival was determined by primary tumor histology ( p = 0.03); the number of metastases, nodal status, disease-free interval or extension of surgery (pneumonectomy vs. lobar resection) were not related to survival probably due to the low number of patients. Overall survival after a complete extended metastasectomy was 66% at 2 years, 42% at 5 years, and 36% at 10 years. Conclusions Extended resections performed during pulmonary metastasectomies are associated with low mortality and morbidity rates and an acceptable long-term survival when performed in selected patients susceptible to complete resection.

Background and Objective. EBUS-TBNA has revolutionized the diagnostic approach to thoracic diseases from a surgical to minimally invasive procedure. In non small-cell lung cancer (NCSLC) patients, EBUS-TBNA is able to dictate the consecutive therapy both for early and advanced stages, providing pathological diagnosis, mediastinal staging, and even adequate specimens for molecular analysis. This study reports on the ability of EBUS-TBNA to make different diagnoses and dictates the consecutive therapy in a large cohort of patients presenting different thoracic diseases. Methods. All procedures performed from January 2012 to September 2016 were reviewed. Five groups of patients were created according to the main indications for the procedure. Group 1: lung cancer staging; Group 2: pathological diagnosis in advanced stage lung cancer; Group 3: lymphadenopathy in previous malignancies; Group 4: pulmonary lesions; Group 5: unknown origin lymphadenopathy. In each group, the diagnostic yield of the procedure was analysed. Non malignant diagnosis at EBUS-TBNA was confirmed by a surgical procedure or clinical and radiological follow-up. Results. 1891 patients were included in the analysis. Sensitivity, negative predictive value, and diagnostic accuracy in each group were 90.7%, 79.4%, and 93.1% in Group 1; 98.5%, 50%, and 98.5% in Group 2; 92.4%, 85.1%, and 94.7% in Group 3; 90.9%, 51.0%, and 91.7% in Group 4; and 25%, 83.3%, and 84.2% in Group 5. Overall sensitivity, negative predictive value, and accuracy were 91.7%, 78.5%, and 93.6%, respectively. Conclusions. EBUS-TBNA is the best approach for invasive mediastinal investigation, confirming its strategic role and high accuracy in thoracic oncology.

Mediastinal lymph node enlargement is common in the follow-up of patients with previously treated malignancies. The aim of this study is to assess the role of endobronchial ultrasound (EBUS) transbronchial needle aspiration (TBNA) for cyto-histological evaluation of positron emission tomography with 18fluorodeoxyglucose (PET) positive mediastinal and hilar lymph nodes developed in patients with previous malignancies. All EBUS-TBNA cases performed from January 2012 to May 2016 were retrospective reviewed. Results of EBUS-TBNA in patients with mediastinal and/or hilar lymphadenopathies were analysed. Non-malignant cytopathologies were confirmed with surgical procedures or clinical and radiological follow-up. Among 1780 patients, 176 were included in the analysis. 103 of these (58.5%) had a diagnosis of tumour recurrence whereas 73 (41.5%) had a different diagnosis: 63 (35.8%) had a non-neoplastic diagnosis and 8 patients (4.6%) had a different cell type malignancy. Samples were false-negative in 5 (2.8%) out of 176 patients. The overall sensitivity, specificity, negative predicted value and diagnostic accuracy were 95.7% (95% CI 90.2–98.6%), 100% (95% CI 94.0–100%), 92.3% (95% CI 83.2–96.7%) and 97.2% (95% CI 93.5–98.8%), respectively. EBUS-TBNA demonstrated a pathological diagnosis different from the previous tumour in a large percentage of patients, confirming its strategic role in the management of patients with previously treated malignancies., EBUS-TBNA changes the management of treated cancer patients http://ow.ly/vTnh30fBFaE

This paper reports on the characteristics, treatment modalities, and outcomes of patients with superior sulcus tumors who underwent surgery over a period of 15 years in 1 institution. Clinical records of 94 consecutive patients operated on by the same surgical team for non–small cell lung cancer between July 1998 and December 2013 were retrospectively reviewed. All patients received lung and chest wall en bloc resection. Forty-eight (51%) received induction treatments. Surgery was an anterior approach in 46 patients (48.9%), Paulson incision in 35 (37.2%), and a combined approach in 13 (13.8%). Lung resections were 78 lobectomies (83%), 3 were pneumonectomies (3.2%), 6 were bronchoplastic reconstructions (6.4%), and 7 were wedge resections (7.4%). Nodal dissection was systematic in 96% of patients. The median number of resected ribs was 2 (1-5), chest wall residual defect was reconstructed in 42 patients (44.7%), and 21 patients had an associated vascular resection (22.3%). Resection was radical in 85 patients (90.4%). Overall 90-day mortality was 9.6%. After a median follow-up of 1.9 years, 5-year and 10-year overall survival rates were 35% and 23%, respectively. A lower 5-year survival was observed in patients with nodal disease (48% in N0 vs 18% in N+; P < 0.0001), incomplete resection (21% for incomplete vs 37% for complete resection; P = 0.15), and anteriorly located tumor (anterior vs posterior: 26% vs 50%; P = 0.05). Pancoast tumor is a severe condition, but long-term survival may be achieved in selected cases. Nodal involvement, completeness of resection, and vascular invasion are the most important prognostic factors, and induction treatment may play a role.

Background: Owing to the expected poor long-term outcomes and high postoperative morbidity and mortality, patients with stage IIIA-N2 tumors candidate to pneumonectomy (PN) are usually excluded from surgery. This study aims to analyze the outcome of patients who underwent PN to prove its safety and feasibility. Patients and Methods: We retrospectively analyzed data from 233 patients who underwent PN for N2 non–small-cell lung cancer (NSCLC) between 1998 and 2015. Eighty-five patients were occult N2 disease (group 1), whereas 148 patients underwent induction therapy (IT) for stage IIIA-N2 (group 2). Results: Overall morbidity, postoperative mortality, and 90-day mortality rates were 46.8%, 2.6%, and 8.6%, respectively. The 2 groups (group 1 vs. 2) had similar postoperative and 90-day mortality rates: 2.4% versus 2.7% (P = 1.00), and 9.4% versus 8.1% (P =.81), respectively. The incidence of major morbidity was higher and statistically significant in group 2 compared with group 1: 23% versus 12.9% (P =.1). Postoperative bronchopleural fistula occurred in 4.7% (4/85) of patients with occult N2 (group 1) and in 10.1% (15/148) of patients undergoing IT (group 2) (P = .10). Median overall survival (OS) was 2.2 years, with a 3 and 5-year OS of 43.4% and 31.6%, respectively. Disease-free survival (DFS) was 1.5 years, with 3 and 5-year DFS of 41.6% and 32%, respectively; no difference in OS and DFS between the 2 groups was found. Conclusions: Considering the acceptable morbidity and mortality rate and the long-term survival, PN should not be excluded for selected patients with stage IIIA-N2 NSCLC as a matter of principle. This study aims to analyze the outcome of patients with stage IIIA-N2 non–small-cell lung cancer who underwent pneumonectomy to prove its safety and feasibility, in particular after induction chemotherapy. Based on the acceptable morbidity and mortality rate and the long-term survival, pneumonectomy should not be excluded for selected patients as a matter of principle.

There is currently little empirical evidence regarding how patients' attachment patterns manifest in individual psychotherapy. This study compared the in-session discourse of patients classified secure, dismissing, and preoccupied on the Adult Attachment Interview (AAI). Rather than focusing on content or form alone, this study analyzed how patients' discourse elicits and maintains emotional proximity with the therapist. The AAI was administered to 56 patients prior to treatment and one session for each patient was rated with the Patient Attachment Coding System (PACS) by four independent raters, blind to patients' AAI classification. Significant differences were found in the discourse of patients with different attachment patterns. Namely, secure and preoccupied patients showed more contact-seeking behavior than dismissing patients, who avoided emotional proximity more, while preoccupied patients resisted therapists' help more than did secure and dismissing patients. These results suggest that the different attachment patterns may have distinctive manifestations in the psychotherapy process that can be tracked by external observers.

Introduction: Mediastinal lymph node enlargement is a common finding in the follow-up of patients with previous radically treated malignancies. PET is routinely used and highs the suspicion of recurrence but histological confirmation is mandatory to plan the correct therapy. Aims and objectives: The aim of this study is to evaluate the role of EBUS-TBNA for histological evaluation in patients with radically treated previous malignancies who developed during the follow-up a PET positive mediastinal lymph node. Methods: Prospected collected data from December 2011 until March 2014 of patients that underwent EBUS-TBNA. Results: One-hundre-eithteen patients out of 859 patients meet the inclusion criteria. Fifty percent of patients (n=59) had the same diagnosis of the previous oncological disease. 46.6% of patients (n=55) had a different diagnosis from the previous tumor and 37.3% of them (n=44) had a benign condition. 9.3% of patients had a different malignancy. In 3.4% of patients (n=4) the sample was false negative. Overall sensitivity, specificity, negative predicted value and diagnostic accuracy were respectively: 94.6% (95% CI: 86.7% to 98.5%), 100% (95% CI: 92.0% to 100%), 91.7% (95% CI: 80.0% to 97.7%), 96.6% (95% CI: 91.6% to 99.1%). Conclusion: Pathological confirmation of PET positive mediastinal lymph nodes in patients with a previously treated malignancy is mandatory to plan a correct treatment. EBUS-TBNA demonstrated a different pathological diagnosis in a high percentage of patients showing a high sensitivity, diagnostic accuracy and negative predicted values confirming its strategic role in the mediastinal investigation in thoracic oncology.

Serum collected in a time-course mode during the pregnancy of a group of heifers was analyzed by 2-DE under various experimental conditions to optimize resolution of all protein spots. Changes in the levels of some components were detected during the last phase of pregnancy and early postpartum. These included a decrease of alpha2-HS-glycoprotein, an increase of alpha1-antichymotrypsin and, with a much larger and more abrupt variation, of orosomucoid and haptoglobin. These findings associate the weeks preceding calving with an acute-phase reaction. Analysis of individual animal's sera by 1-DE demonstrated a higher level of orosomucoid in the sera of cows developing postpartum endometritis during the 2 wk after calving (i.e., in the course of the infection) but a lower level during the 2 wk before calving. This observation could represent an important tool for the prepartum detection of animals prone to develop postpartum endometritis and lead to a more accurate peripartum management of those animals.

A laboratory-prepared total protein extract (TPE) and a lupin protein isolate (LPI-E) produced in a pilot plant were submitted to a detailed two-dimensional (2DE) proteomic investigation. Recent findings have indicated that in an established rodent model of hyperlipidemia, moderate daily intakes of LPI-Es lead to a reduction of total and low-density lipoprotein cholesterol levels, and the knowledge of the actual composition of the protein sample used in that study is at the basis of further structure/action investigations. The experimental results indicate that the semi-industrial procedure used for the production of LPI-E damages only marginally the proteins. It does, however, cleave some disulfide bridges and induce mild proteolysis, as confirmed by the higher number of resolved protein spots in the low Mr and acidic pI region of the 2DE map. Out of 72 spots submitted to mass spectrometry and compared with available protein databases, 42 correspond to fragments of beta-conglutin, the 7S globulin of lupin, spanning between positions 37 and 495 of the protein sequence. Using the bioinformatic tool BlastP, these peptides were compared to the alpha'-subunit of beta-conglycinin, the 7S globulin of soybean, this being the most active hypocholesterolemic component of soybean protein, as shown by in vitro and in vivo experiments. At least 18 peptides derived from beta-conglutin, having a percentage identity higher than 50% and a similarity percentage higher than 70% vs the alpha'-subunit of beta-conglycinin, are likely candidates to be the biologically active components of lupin protein.

Ex vivo lung perfusion (EVLP) is a promising procedure for evaluation, reconditioning, and treatment of marginal lungs before transplantation. Small animal models can contribute to improve clinical development of this technique and represent a substantial platform for bio-molecular investigations. However, to accomplish this purpose, EVLP models must sustain a prolonged reperfusion without pharmacological interventions. Currently available protocols only partly satisfy this need. The aim of the present research was accomplishment and optimization of a reproducible model for a protracted rat EVLP in the absence of anti-inflammatory treatment. A 180 min, uninjured and untreated perfusion was achieved through a stepwise implementation of the protocol. Flow rate, temperature, and tidal volume were gradually increased during the initial reperfusion phase to reduce hemodynamic and oxidative stress. Low flow rate combined with open atrium and protective ventilation strategy were applied to prevent lung damage. The videos enclosed show management of the most critical technical steps. The stability and reproducibility of the present procedure were confirmed by lung function evaluation and edema assessment. The meticulous description of the protocol provided in this paper can enable other laboratories to reproduce it effortlessly, supporting research in the EVLP field.

Patients with stage IIIA-ipsilateral mediastinal lymph node involvement (N2) non-small cell lung cancer (NSCLC) represent a heterogeneous group with different clinical presentation. The aim of this study was to analyze a series of patients with "potentially resectable" stage IIIA-pathologically proven N2 (pN2) NSCLC undergoing induction chemotherapy followed by surgery to evaluate their long-term outcomes and to identify prognostic factors. Out of 287 patients who underwent induction chemotherapy for NSCLC with ipsilateral mediastinal lymph node involvement pathologically proven, we retrospectively evaluated 141 (49%) patients with no clinical evidence of progression after induction chemotherapy and candidates for surgery. Most of them (73%) underwent at least 3 cycles of cisplatin-based chemotherapy. We used the Kaplan-Meier method to plot survival and the log-rank test to assess the survival difference between groups. Multivariable analysis was performed using Cox proportional hazards regression. A total of 15 (10.6%) patients underwent explorative thoracotomy; 126 patients underwent surgical anatomical resection after a median 27 days (range: 21-30) from the last cycle of chemotherapy. A total of 113 (89.7%) patients had a radical resection. A total of 22 (17.5%) patients had a complete pathologic lymph node downstaging (pN0), and 8 (6.3%) patients had a complete pathological response (pT0N0). The median overall survival was 24 months, with a 5-year overall survival of 30%. At multivariable analysis, downstaging and number of cycles of chemotherapy were independent prognostic factors (P = 0.006); downstaging benefit was mostly because of complete pathological response (hazards ratio = 0.23, 95% CI: 0.07-0.76). In conclusion, more than 3 cycles of chemotherapy and pathological downstaging could significantly improve 5-year survival in selected patients with "potentially resectable" pathologically proven N2 disease.

http://hdl.handle.net/20.500.11768/96439 Background: During the period that randomized clinical trials were establishing the role of adjuvant therapy in tumors larger than 5 cm without lymph-node invasion, which shifted from stage IB (6th TNM) to stage II (7th TNM), we derived the rate of shifted patients in our series and analyzed the relationship between specific patient- and tumor-characteristics, and clinical outcome, to identify putative prognostic factors. Methods: We retrospectively collected data (age, sex, smoking status, type of surgery grading, and histological type) from 467 patients who underwent radical surgery for primary 6th TNM-T2N0 non-small cell lung cancer patients between 1998 and 2009 at our institute. Categorical variables were cross-tabulated by tumor staging according to the 7th TNM edition, and they were tested both for association with stage and survival. Results: One hundred and eighteen patients shifted to stage II, mainly older patients and patients with a sarcomatoid or a poorly differentiated carcinoma. Median overall survival time was significantly different across stages. Among the factors investigated, only the tumor dimension resulted in being statistically significant in multivariate analysis. Conclusions: Nearly a quarter of patients shifted from stage I (6th TNM) to stage II (7th TNM), raising a major need for information on the effects of adjuvant chemotherapy in this group of patients. Our findings suggest that randomized clinical trials aimed at addressing this topic should consider only tumor dimensions as principal selection criteria.

Objective: The introduction of modern staging systems such as computed tomography (CT) and positron emission tomography/CT (PET/CT) with fluorodeoxyglucose ([ 18 F]FDG) has increased the detection of small peripheral lung cancers at an early stage. We analyzed the behavior of pathological T1 non-small-cell lung cancer (NSCLC) to identify criteria predictive of nodal involvement, and the role of cancer size in lymph node metastases. Methods: We retrospectively analyzed 219 patients with pathological T1 NSCLC. All patients were staged by high-resolution CTand PETas stage I, and underwentanatomical resectionand radical lymphadenectomy. Our data were collected based on pathological nodulesize (0— 10 mm; 11—20 mm; and 21—30 mm); morphological features of lung nodule and FDG uptake of the tumor measured by standardized uptake value (SUV). Results:A total of 190 patients (87%) were pN0, 14 (6%) pN1, and 15 (7%) pN2. No nodal involvement was observed in any of the 62 patients with nodule size less than 10 mm, in 20 out of 120 patients (17%) with nodule size 11—20 mm, and in nine out of 37 tumors (28%) 21—30 mm in size (p = 0.0007). All 55 patients with nodule SUV < 2.0 and all 26 non-solid lesions were pN0 (respectively, p = 0.0001 and p = 0.03). All nodal metastases occurred amongthe group of 132patients with size larger than 10 mm and SUV higher than 2.0 with a 22% rate of nodalinvolvement of (29 patients) (p < 0.0001). Conclusions: The low probability of lymph node involvement in NSCLC

Objective After pneumonectomy, quality of life may be impaired in a proportion of patients because of the presence of symptoms causing severe limitations in daily activities. This is a prospective study on patients who have undergone pneumonectomy for cancer, assessing quality of life modifications 6 months after surgery. Methods Beginning in August 2006, candidates for pneumonectomy had their quality of life assessed by the European Organization for Research and Treatment of Cancer questionnaire (QLQ-C30 + LC13) preoperatively and at 1, 3, and 6 months after surgery. Poor quality of life at 6 months was defined as global health values 10% or more below baseline values. The impact of several clinical variables was tested to discover predictors of poor postoperative quality of life. Results Forty-one of the 50 patients enrolled in the study had a complete quality of life follow-up by January 2008, representing the population of the study. Six months after pneumonectomy, global health showed a minimal impairment in the whole population (baseline 60.4 ± 26.5, at 6 months 56.3 ± 24.2, P = .15). Ten patients (24.4%) were identified as having poor quality of life at 6 months. Age of 70 years or more was identified as a significant risk factor for poor 6-month quality of life using multivariate analysis (odds ratio, 1.13; 95% confidence interval, 1–1.26). The baseline global health score was the strongest predictor of postoperative global health quality of life (odds ratio, 0.16; 95% confidence interval, 0.02–0.46; P = .0086). Conclusion The overall quality of life after pneumonectomy was impaired in 25% of surviving patients at 6 months after surgery; thus, this aspect of recovery should be routinely discussed with patients before pneumonectomy. Patients aged 70 years or more and those with low preoperative quality of life seem to be at risk for unsatisfactory quality of life after surgery.

Background We have assessed the survival rate of patients with non–small cell lung cancer and synchronous hematogenous solitary metastasis identified with complete staging workup, including total body [ 18 F]fluorodeoxyglucose positron emission tomography scan, and treated with a multidisciplinary approach. Methods We examined the database of all patients who underwent surgery for primary non–small cell lung cancer in our institute. The criteria required for inclusion in this analysis were diagnosis of non–small cell lung cancer with synchronous hematogenous solitary metastasis by staging workup with total body computed tomography scan and brain magnetic resonance if indicated, total body positron emission tomography scan, radical surgery for the primary tumors, local treatment of the solitary metastasis, and systemic chemotherapy administration. Results We analyzed the data from 1,509 patients treated from January 2000 to December 2005: 10 patients (0.7%) satisfied the selection criteria. The median overall survival was 26 months, and the median time to progression was 20 months; 6 patients were alive at the time of analysis, with a median follow-up of 30 months. Four patients were tumor progression–free after 9, 18, 23, and 32 months from the start of their treatment. Conclusions The presentation of non–small cell lung cancer with a synchronous hematogenous solitary metastasis identified by [ 18 F]fluorodeoxyglucose positron emission tomography containing complete staging workup is extremely rare. This subset of patients can achieve long-term survival after a multidisciplinary treatment approach.

Two-dimensional electrophoresis is usually run on fully reduced samples. Under these conditions even covalently bound oligomers are dissociated and individual polypeptide chains may be fully unfolded by both, urea and SDS, which maximizes the number of resolved components and allows their pI and M(r) to be most accurately evaluated. However, various electrophoretic protocols for protein structure investigation require a combination of steps under varying redox conditions. We review here some of the applications of these procedures. We also present some original data about a few related samples -- serum from four species: Homo sapiens, Mus musculus, Rattus norvegicus, Bos taurus -- which we run under fully unreduced and fully reduced conditions as well as with reduction between first and second dimension. We demonstrate that in many cases the unreduced proteins migrate with a better resolution than reduced proteins, mostly in the crowded 'alpha-globulin' area of pI 4.5-6 and M(r) 50-70 kDa.

Background This study assessed perioperative outcome and long-term survival in a large series of patients with malignant pleural mesothelioma who underwent extrapleural pneumonectomy (EPP) to identify prognostic factors allowing better patient selection. Methods We retrospectively collected data from nine referral centers for thoracic surgery in Italy. Perioperative outcome and survival data were available for 518 malignant pleural mesothelioma patients (84.4% with epithelial tumors, 68.0% with pathologic stage 3 disease) who underwent EPP with intention-to-treat (R0/R1) between 2000 and 2010. Induction chemotherapy was administered in 271 patients (52.3%) and adjuvant therapy in 373 patients (72.0%), including radiotherapy in 213 patients (41.1%), adjuvant chemotherapy in 43 patients (8.3%), and both in 117 patients (22.6%). Results In all, 136 patients (26.3%) had major complications after EPP, and 36 (6.9%) died within 90 days after surgery. The median overall survival was 18 months, with a 1-, 2-, and 3-year overall survival of 65%, 41%, and 27%, respectively. At multivariable analysis adjusted for age and disease stage, male sex (hazard ratio [HR] 1.47, 95% confidence interval [CI]: 1.12 to 1.92), nonepithelial histology (HR 1.96, 95% CI: 1.48 to 2.58), and trimodality treatment using induction chemotherapy (HR 0.61, 95% CI: 0.43 to 0.85) were significantly associated with survival. Development of a major complication also significantly worsened outcome (HR 1.85, 95% CI: 1.37 to 2.50). Conclusions The success of EPP in the context of a multimodality treatment depends on a series of patient characteristics. Female patients, patients with epithelial tumors, and patients who received induction chemotherapy will best benefit from EPP. © 2014 by The Society of Thoracic Surgeons Published by Elsevier Inc.

Background Bilobectomy for lung cancer is considered a high-risk procedure for the increased postoperative complication rate and the negative impact on survival. We analyzed the safety and the oncologic results of this procedure. Methods We retrospectively reviewed patients who underwent bilobectomy for lung cancer between October 1998 and August 2009. Age, gender, bilobectomy type and indication, complications, pathology, stage, and survival were analyzed. Results Bilobectomy was performed on 146 patients (101 men; mean age, 62 years). There were 77 upper-middle and 69 middle-lower bilobectomies. Indications were tumor extending across the fissure in 27 (18.5%) patients, endobronchial tumor in 39 (26.7%), extrinsic tumor or nodal invasion of bronchus intermedius in 66 (45.2%), and vascular invasion in 14 (9.6%). An extended resection was performed in 24 patients (16.4%). Induction therapy was performed in 43 patients (29.4%). Thirty-day mortality was 1.4% (n = 2). Overall morbidity was 47.2%. Mean chest tube persistence was 7 days (range, 6 to 46 days). Overall 5-year survival was 58%. Significance differences in survival were observed among different stages (stage I, 70%; stage II, 55%; stage III, 40%; p = 0.0003) and the N status (N0, 69%; N1, 56%; N2, 40%; p = 0.0005). Extended procedure ( p = 0.0003) and superior bilobectomy ( p = 0.0008) adversely influenced survival. Multivariate analysis demonstrated that an extended resection ( p = 0.01), an advanced N disease ( p = 0.02), and an upper-mild lobectomy ( p = 0.02) adversely affected prognosis. Conclusions Bilobectomy is associated with a low mortality and an increased morbidity. Survival relates to disease stage and N factor. Optimal prognosis is obtained in patients with lower-middle lobectomy without extension of the resection.

Background: To address whether systematic lymph node dissection is always necessary in early lung cancer, we identified factors predicting nodal involvement in a screening series and applied them to nonscreening-detected cancers. Methods: In the 97 patients with clinical T1-2N0M0 lung cancer (10 mm and maxSUV ≤2.0 (p = 0.002 versus the other 62 cases). In the nonscreening series, 42 of 43 cases with negative PET-CT (usually maxSUV 10 mm were pN+ (p = 0.001 versus the 43 cases). Conclusions: This limited experience suggests that in early-stage clinically N0 lung cancers with maxSUV

In this case report we studied alterations in mitochondrial proteins in a patient suffering from recurrent profound muscle weakness, associated with ethylmalonic-adipic aciduria, who had benefited from high dose of riboflavin treatment. Morphological and biochemical alterations included muscle lipid accumulation, low muscle carnitine content, reduction in fatty acid beta-oxidation and reduced activity of complexes I and II of the respiratory chain. Riboflavin therapy partially or totally reversed these symptoms and increased the level of muscle flavin adenine dinucleotide, suggesting that aberrant flavin cofactor metabolism accounted for the disease. Proteomic investigation of muscle mitochondria revealed decrease or absence of several flavoenzymes, enzymes related to flavin cofactor-dependent mitochondrial pathways and mitochondrial or mitochondria-associated calcium-binding proteins. All these deficiencies were completely rescued after riboflavin treatment. This study indicates for the first time a profound involvement of riboflavin/flavin cofactors in modulating the level of a number of functionally coordinated polypeptides involved in fatty acyl-CoA and amino acid metabolism, extending the number of enzymatic pathways altered in riboflavin-responsive multiple acyl-CoA dehydrogenase deficiency.

We present reference maps of the mouse serum proteome (run under reducing and non-reducing conditions), from control animals, from mice injected with lipopolysaccharide (LPS) to induce systemic inflammation, and from mice transgenic for human apolipoproteins A-I and A-II. Seventy-seven spots/spot chains from the reducing gels were identified by HPLC MS/MS, representing 28 distinct proteins, including a species-specific protease inhibitor, contrapsin, and high levels of carboxylesterase. The concentrations of acute-phase reactants were monitored for 96 h after LPS challenge. The greatest changes (four-fold 48 h after LPS administration) were observed for haptoglobin and hemopexin. Orosomucoid/alpha(1)-acid glycoprotein and apolipoprotein A-I increased steadily, to 50-60% above baseline at 96 h from stimulation. In mice transgenic for human apolipoprotein A-I the levels of expression of orosomucoid/alpha(1)-acid glycoprotein, alpha(1)-macroglobulin, esterase, kininogen and contrapsin were altered compared to knockout mice lacking apolipoprotein A-I. In contrast, except for the presence of apolipoprotein A-II, no statistically significant difference was observed in mice transgenic for human apolipoprotein A-II.