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The cellular basis of the apparent aggressiveness in lung cancer is poorly understood but likely associated with functional or molecular features of disseminated cancer cells (DCCs). DCCs from epithelial cancers are mostly detected by antibodies directed against histogenetic markers such as cytokeratin or EpCAM. It has been argued that marker-negative metastatic founder cells might escape detection. We therefore used ex vivo sphere formation for functional detection of candidate metastasis founders. We generated cell suspensions from 199 LN samples of 131 lung cancer patients and placed them into non-adherent cell culture. Sphere formation was associated with detection of DCCs using EpCAM immunocytology and with significantly poorer prognosis. The prognostic impact of sphere formation was strongly associated with high numbers of EpCAM-positive DCCs and aberrant genotypes of expanded spheres. We also noted sphere formation in patients with no evidence of lymphatic spread, however such spheres showed infrequent expression of signature genes associated with spheres from EpCAM-positive samples and displayed neither typical lung cancer mutations (KRAS, TP53, ERBB1) nor copy number variations, but might be linked to disease progression >5 years post curative surgery. We conclude that EpCAM identifies relevant disease-driving DCCs, that such cells can be expanded for model generation and that further research is needed to clarify the functional and prognostic role of rare EpCAM-negative sphere forming cells., (© 2023 The Authors. International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.)

Computer tomography-derived skeletal muscle index normalized for height in conjunction with muscle density enables single modality-based sarcopenia assessment that accounts for all diagnostic criteria and cutoff recommendations as per the widely accepted European consensus. Yet, the standard approach to quantify skeletal musculature at the third lumbar vertebra is limited for certain patient groups, such as lung cancer patients who receive chest CT for tumor staging that does not encompass this lumbar level. As an alternative, this retrospective study assessed sarcopenia in lung cancer patients treated with curative intent at the tenth thoracic vertebral level using appropriate cutoffs. We showed that skeletal muscle index and radiation attenuation at level T10 correlate well with those at level L3 (Pearson's R = 0.82 and 0.66, p < 0.001). During a median follow-up period of 55.7 months, sarcopenia was independently associated with worse overall (hazard ratio (HR) = 2.11, 95%-confidence interval (95%-CI) = 1.38-3.23, p < 0.001) and cancer-specific survival (HR = 2.00, 95%-CI = 1.19-3.36, p = 0.009) of lung cancer patients following anatomic resection. This study highlights feasibility to diagnose sarcopenia solely by thoracic CT in accordance with the European consensus recommendations. The straightforward methodology offers easy translation into routine clinical care and potential to improve preoperative risk stratification of lung cancer patients scheduled for surgery., (© 2023. The Author(s).)

In melanoma, immunocytology (IC) after sentinel lymph node disaggregation not only enables better quantification of disseminated cancer cells (DCCs) than routine histopathology (HP) but also provides a unique opportunity to detect, isolate, and analyse these earliest harbingers of metachronous metastasis. Here, we explored lymph node IC in non-small cell lung cancer (NSCLC). For 122 NSCLC patients, 220 lymph nodes (LNs) were split in half and prepared for IC and HP. When both methods were compared, IC identified 22% positive patients as opposed to 4.5% by HP, revealing a much higher sensitivity of IC (p < 0.001). Assessment of all available 2,952 LNs of the same patients by HP uncovered additional patients escaping detection of lymphatic tumour spread by IC alone, consistent with the concept of skip metastasis. A combined lymph node status of IC and complete HP on a larger cohort of patients outperformed all risk factors in multivariable analysis for prognosis (p < 0.001; RR = 2.290; CI 1.407-3.728). Moreover, isolation of DCCs and single-cell molecular characterization revealed that (1) LN-DCCs differ from primary tumours in terms of copy number alterations and selected mutations and (2) critical alterations are acquired during colony formation within LNs. We conclude that LN-IC in NSCLC patients when combined with HP improves diagnostic precision, has the potential to reduce total workload, and facilitates molecular characterization of lymphatically spread cancer cells, which may become key for the selection and development of novel systemic therapies. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland., (© 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.)

Introduction: Detection of disseminated cancer cells (DCC) in bone marrow (BM) of patients with early-stage NSCLC has been associated with poor outcome. However, the phenotype, and hence relevant therapy targets, of DCCs in BM are unknown. We therefore compared a classical pan-Cytokeratin (CK) antibody for DCC detection with an anti-EpCAM antibody that may also detect more stem-like cells and tested whether assay positivity impacts on the survival of NSCLC patients., Materials and Methods: We prospectively collected BM aspirates from 104 non-metastasized NSCLC patients that underwent potentially curative tumor resection from 2011 to 2016 at the Department of Thoracic Surgery of the University Hospital and Hospital Barmherzige Brüder in Regensburg. DCCs were detected by staining with the pan anti-CK antibody A45-B/B3 and the anti-EpCAM antibody HEA-125. We analyzed the association between detection of DCCs and clinicopathological characteristic and patient outcome., Results: CK + and EpCAM + DCCs were detected in 45.2% and 52.9% of patients, respectively. Correlation between the two markers was low and neither of them was associated with sex, age, histology, T or N classification, resection status, grading or smoking habit. No significant association with tumor specific survival (TSS) and progression-free survival (PFS) was observed in patients with CK + DCCs. In contrast, detection of EpCAM + DCCs significantly correlated with reduced PFS (P = 0.017) and TSS (P = 0.017) and remained an independent prognostic variable for PFS and TSS upon multivariate testing (hazard ratio: 7.506 and 3.551, respectively). Detection of EpCAM + DCCs was the only prognostic marker for PFS., Conclusions: EpCAM+, but not CK + DCCs in BM predict reduced PFS and TSS. This finding suggests that EpCAM + DCCs in the BM comprise metastatic founder cells necessitating their in-depth molecular analysis for detection of novel therapy targets., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: Surgical resection of pulmonary metastases leads to prolonged survival if strictly indicated. Usually, thoracotomy with manual palpation of the entire lung with lymph node dissection or sampling is performed. The aim of this study was to evaluate the role of video-assisted thoracoscopic surgery (VATS) in pulmonary metastectomy with curative intent., Methods: In this study, all patients with suspected pulmonary metastasis (n = 483) who visited the Center for Thoracic Surgery in Regensburg, between January 2009 and December 2017 were analysed retrospectively., Results: A total of 251 patients underwent metastectomy with curative intent. VATS was performed in 63 (25.1%) patients, 54 (85.7%) of whom had a solitary metastasis. Wedge resection was the most performed procedure in patients treated with VATS (82.5%, n = 52) and thoracotomy (72.3%, n = 136). Postoperative revisions were necessary in nine patients (4.8%), and one patient died of pulmonary embolism after thoracotomy (0.5%). Patients were discharged significantly faster after VATS than after thoracotomy (p < 0.001). Complete (R0) resection was achieved in 89% of patients. The median recurrence-free survival was 11 months (95% confidence interval 7.9-14.1). During follow-up, eight (12.7%) patients in the VATS group and 42 (22.3%) patients in the thoracotomy group experienced recurrence (p = 0.98). The median overall survival was 61 months (95% confidence interval 46.1-75.9), and there was no significant difference with regard to the surgical method used (p = 0.34)., Conclusions: VATS metastasectomy can be considered in patients with a solitary lung metastasis. An open surgical approach with palpation of the lung showed no advantage in terms of surgical outcome or survival.

Study Aim: The 8th edition of the TNM classification combined with the latest update of the S3-guideline (by AWMF/Scientific Medical Societies in Germany) on prevention, diagnosis, therapy and follow-up of lung cancer led to several changes in staging and treatment of lung cancer. The aim of this study was to identify differences in the distribution of patients due to changes from the 7th to the 8th edition that affected staging. The influence on surgical therapy will be discussed by using the recommendations of the latest S3 guideline., Methods: Prospective analysis of all primary cases at two thoracic surgical centres in the year 2016 and follow-up in March 2019. Comparison of the 7th edition of tumour classification for lung cancer with the 8th edition, focused on changes in tumour staging and its effects on the appropriate surgical therapy according to the latest S3 guideline., Results: A total of 432 primary cases comprised the study population. According to the 8th edition, 82 patients (7th edition: n = 85) in stage I, 43 (n = 49) patients in stage II, 100 (n = 91) patients in stage III and 207 (n = 207) patients are assigned to stage IV. 81 changes (18.7%) were detected (77 upgrades vs. 4 downgrades). 63 patients (14.6%) exhibited a different graduation within the stages. 18 patients (4.1%) were classified in different tumour stages. As a result, fewer patients (n = 12; 2.8%) should have surgery according to the latest S3 guidelines. 290 patients (67.1%) were classified to new subgroups (IA1-3, IIIC and IVA/B). Two-year survival was significantly higher in IVA (25.2%) vs. IVB (13.0%) patients (p < 0.05)., Conclusion: The 8th edition of the TNM-classification affords a higher level of differentiation. In this study, the new TNM classification led to a shift in the distribution, with a tendency to increase the tumour stage. This is mainly caused by changes in the T-descriptor and stage grouping. As a result, fewer patients in stage I - IIIA should have surgery according to the latest S3 guidelines. A significantly higher two-year survival rate was detected in stage IVA (M1a and M1b) compared to IVB and justifies the new differentiation due to the metastatic pattern., Competing Interests: Die Autorinnen/Autoren geben an, dass kein Interessenkonflikt besteht., (Thieme. All rights reserved.)

Background: The Federal Joint Committee (G‑BA) is currently discussing the introduction of new minimum volume regulations (MVR) in Germany. The present study examined the current opinions of active thoracic surgeons regarding minimum volumes (MV) for the surgical treatment of lung cancer., Methods: The participating centers for the online survey were identified on the basis of the thoracic surgery departments in the 2017 hospital directory (Federal Statistical Office), lung cancer centers (German Cancer Society), certified centers of excellence for thoracic surgery (German Society for Thoracic Surgery), hospitals with a focus on lung surgery and German university hospitals. They were asked about the potential effects of MVR on the quality of results and quality of care, economic aspects and the structure of care. Furthermore, a recommendation for MV was requested and possible provisions for exemption were evaluated., Results: A total of 145 hospitals (response rate 85%) with 454 thoracic surgeons (response rate 54%) were surveyed. The results showed a high degree of approval for MV to improve the quality of results and 78.4% of the surgeons surveyed expected it to result in centralization of surgical care, although this would not lead to a deterioration in care according to 70.1% of the participants. Approximately 46.1% of the participants expected care to become more economical and 83.3% supported the introduction of an MVR, with the average recommended MV being 67 anatomical lung resections per center per year., Conclusion: An MVR for the surgical treatment of lung cancer met with a high degree of approval among active thoracic surgeons. The MV that was called for (n = 67) was slightly below the prerequisite for primary surgical cases at a certified lung cancer center.

Background: The lungs are the second most common organ site for metastases in patients with colorectal cancer (CRC). Lymph node metastasis of CRC represents a prognostic factor for survival., Objective: The present study investigated the influence of CRC lymph node metastasis on lung metastasis, in particular thoracic lymph node metastasis., Material and Methods: A retrospective analysis of 88 patients (n = 56 male) with curative resection of lung metastases of CRC was performed. Primary endpoint: influence of lymph node status of CRC on lung metastases. Secondary endpoints: disease-free survival and overall survival. Statistical evaluation was carried out with SPSS., Results: In 48 patients a positive lymph node status of CRC and in 9 patients an N+ status of lung metastases were determined. The lymph node status of the CRC significantly affected the incidence of synchronous metastases (p = 0.03), disease-free interval until formation of metachronous lung metastases (p = 0.012) and the overall survival of patients with CRC (p = 0.048). The 5‑year survival rate for CRC patients with lung metastases was 48.7% after pulmonary metastasectomy. Thoracic lymph node involvement also significantly affected survival (p = 0.001)., Conclusion: Screening for pulmonary metastases should be included in the staging and follow-up of all patients with CRC, especially in patients with a positive lymph node status of the CRC.

Objectives: Preoperative computed tomography (CT) scans for transcatheter aortic valve implantation (TAVI) are used routinely. In elderly high-risk patients, incidental radiographic findings are frequently reported. The aim of this study was to investigate the impact of auxiliary findings on the patients' mid-term survival, which might affect the treatment strategy., Methods: Between March 2011 and April 2016, all radiological reports of contrast-enhanced CT scans of 976 patients scheduled for TAVI were analysed retrospectively for incidental thoracic findings including solitary pulmonary nodules (SPN) and thoracic lymphadenopathy (LAP). The minimum follow-up period was 1 year after TAVI., Results: The median age of all patients was 79 years; 51.9% (n = 507) were women. Approximately 37% (n = 361) of patients showed 1 of the determined findings. An SPN ≥5 mm was diagnosed in 16.4% (n = 160) of patients. Four of them developed lung cancer and 2 nodules were identified as metastases during follow-up. In addition, 12% (n = 117) of the patients had thoracic LAP. Whereas SPN had no significant effect on the overall survival rate, evidence of LAP turned out to be a statistically significant factor regarding 4-year survival (P = 0.001; hazard ratio 1.66; 95% confidence internal 1.19-2.31)., Conclusions: SPN ≥5 mm were detected in 16.4% of patients scheduled for TAVI. Nevertheless, the incidence of lung cancer was low and the effect on survival in this high-risk group of patients was statistically not significant. In contrast, thoracic LAP had a significant negative effect on survival. It needs to be proven if the outcome of this cohort can be enhanced by further diagnostics and therapy., (© The Author(s) 2018. Published by Oxford University Press on behalf of the European Association for Cardio-Thoracic Surgery. All rights reserved.)

Introduction: Adjuvant platinum-based chemotherapy is standard treatment for surgically resected stage II to IIIA NSCLC, but the relapse rate is high. The preferentially expressed antigen of melanoma (PRAME) tumor antigen is expressed in two-thirds of NSCLC and offers an attractive target for antigen-specific immunization. A phase I dose escalation study assessed the safety and immunogenicity of a PRAME immunotherapeutic consisting of recombinant PRAME plus proprietary immunostimulant AS15 in patients with surgically resected NSCLC (NCT01159964)., Methods: Patients with PRAME-positive resected stage IB to IIIA NSCLC were enrolled in three consecutive cohorts to receive up to 13 injections of PRAME immunotherapeutic (recombinant PRAME protein dose of 20 μg, 100 μg, or 500 μg, with a fixed dose of AS15). Adverse events, predefined dose-limiting toxicity, and the anti-PRAME humoral response (measured by enzyme-linked immunosorbent assay) were coprimary end points. Anti-PRAME cellular responses were assessed., Results: A total of 60 patients were treated (18 received 20 μg of PRAME, 18 received 100 μg of PRAME, and 24 received 500 μg of PRAME). No dose-limiting toxicity was reported. Adverse events considered by the investigator to be causally related to treatment were grade 1 or 2, and most were injection site reactions or fever. All patients had detectable anti-PRAME antibodies after four immunizations. The percentages of patients with PRAME-specific CD4-positive T cells were higher at the dose of 500 μg compared with lower doses. No predefined CD8-positive T-cell responses were detected., Conclusion: The PRAME immunotherapeutic had an acceptable safety profile. All patients had anti-PRAME humoral responses that were not dose related, and 80% of those treated at the highest dose showed a cellular immune response. The dose of 500 μg was selected. However, further development was stopped after negative results with a similar immunotherapeutic in patients with NSCLC., (Copyright © 2016 International Association for the Study of Lung Cancer. Published by Elsevier Inc. All rights reserved.)

Background: Pulmonary metastases occur in 10-20% of patients with colorectal cancer and significantly influence long-term survival. In this study, the immunological architecture of colorectal lung in comparison to liver metastases and its impact on patient survival were examined., Methods: Specimens of patients with colorectal lung and liver metastases were stained for HE, CD4, CD8, CD20, CD68 and CD45RO. Besides histomorphological evaluation, immunohistochemical stainings were analyzed for the respective cell numbers separately for tumor area, infiltrative margin and distant lung or liver stroma. These findings were correlated with clinical data and patient outcome., Results: In colorectal lung (n = 69) in comparison to liver (n = 222) metastases, the immunological focus is located in the tumor region. A high CD4+ cell infiltration of this area is associated with prolonged survival of patients after resection of colorectal lung metastases [103 ± 33 (high) vs. 37 ± 6 months (low); p = 0.0246]. Patients who were treated with preoperative chemotherapy did not show differences in immune infiltrates compared to chemotherapy-naïve patients., Conclusion: Colorectal lung and liver metastases showed a distinct immunological architecture. A dense cell infiltration of colorectal lung metastases by CD4+ cells was related to prolonged patient survival. Preoperative chemotherapy did not influence cellular immune infiltrates., (© 2016 S. Karger AG, Basel.)

A 64-year-old man was diagnosed with complex empyema after a second course of palliative chemotherapy for metastatic lung cancer. Because of the poor general condition of the patient, the decision was made to proceed with vacuum-assisted closure (VAC) therapy of the empyema without Eloesser or Clagett open-window thoracostomy (OWT). Installation and changing of the VAC sponge were performed using the ALEXIS Wound Protector/Retractor (Applied Medical, Rancho Santa Margarita, CA), a flexible polymer membrane tube. After 10 days of VAC treatment, the pleural cavity was sterile and was closed with single stitches. Chemotherapy was resumed 1 week later., (Copyright © 2012 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

The plasma fluorescence related to the standard fluorescence of advanced glycation end products (AGEs) is a simple measurable blood parameter for distinct diseases but its importance in human cancer, including non-small cell lung carcinoma (NSCLC), is unknown. Plasma samples of 70 NSCLC patients who underwent resection surgery of the tumor were analyzed for the distinct AGE-related fluorescence at 370 nm excitation/440 nm emission. In a retrospective study, we tested the prognostic relevance of this AGE-related plasma fluorescence. The effect of circulating AGEs on the NSCLC growth was studied experimentally in vitro and in vivo. NSCLC patients with high (> median) AGE-related plasma fluorescence were characterized by a later reoccurrence of the tumor after curative surgery and a higher survival rate compared with patients with low plasma fluorescence (25% versus 47% 5-y survival, P = 0.011). Treating NSCLC cell spheroids with patients' plasma showed an inverse correlation between the growth of spheroids in vitro and the individual AGE-related fluorescence of each plasma sample. To confirm the impact of circulating AGEs on the NSCLC progression, we studied the NSCLC growth in mice whose circulating AGE level was elevated by AGE-rich diet. In vivo tumorigenicity assays demonstrated that mice with higher levels of circulating AGEs developed smaller tumors than mice with normal AGE levels. The AGE-related plasma fluorescence has prognostic relevance for NSCLC patients in whom the tumor growth-inhibiting effect of circulating AGEs might play a critical role.

Successful therapy of the non-small cell lung carcinoma (NSCLC) depends on its early detection, and non-invasive detection methods are preferred. As plasma proteins are modified by N-linked glycosylation, we tested the importance of the N-glycan profile in diagnosing and prognosticating NSCLC. We analysed desialylated plasma samples from 75 NSCLC patients, and 71 healthy individuals by the high-throughput DNA sequencer-based carbohydrate analytical profiling technique. We detected alterations in the levels of several N-glycans in NSCLC patients. Total α-1,6-core fucosylated biantennaries (NGA2F, NG1A2F, NA2F) and total bisecting α-1,6-core fucosylated biantennaries (NGA2FB, NA2FB) were reduced in NSCLC patients, whereas the branching α-1,3-fucosylated triantennary N-glycan (NA3FB) was increased. Best diagnostic accuracy was identified for NG1A2F. NSCLC patients with TNM stage I stage did not show further differences, but patients with higher stages did (TNM II to IV). Those patients additionally had a reduced level in the α-1,6-core fucosylated structure NA2F with parallel increase in the non-fucosylated structure NA2. In this regard, NSCLC patients with a relatively low amount of NA2 per NA2F had a better three-year survival than patients with high amount. NSCLC patients show an altered N-glycan profile of plasma proteins that may be regarded as a supportive tool for cancer diagnosis.

Background: Both diabetes mellitus (DM) and non-small cell lung carcinoma (NSCLC) are age-related diseases and therefore occur simultaneously. DM and, in particular, diabetic nephropathy are associated with an increased modification of the extracellular tissue matrix with advanced glycation end-products (AGEs)., Objective: As AGE-modified matrix impairs the invasive migration of lung carcinoma cells, our study aimed at the effect of DM on the NSCLC-related outcome., Methods: In a retrospective clinical study, we analyzed the time-dependent survival of NSCLC patients with DM compared to patients without DM, who had been treated for cancer by resection surgery. The age-matched study population (51-80 years) included 55 DM patients and 111 non-DM patients., Results: Comparative analyses between DM and non-DM patients did not find significant differences for NSCLC histology, tumor stage and stay in the hospital. Analyses of the mid-term (20-month) survival showed that the DM group had a significantly higher proportion of NSCLC patients being alive 20 months after tumor resection than the non-DM group (76 vs. 59%, p = 0.048). The DM-related increase in the mid-term survival rate was particularly observed for patients without lymph node metastasis (88%) and those patients also suffering from nephropathy (90%). In contrast to the mid-term survival, DM had no effect on the long-term (60-month) survival of NSCLC patients (35 vs. 32% in the non-DM group)., Conclusion: Our study indicates that DM-associated changes mediate beneficial effects in terms of NSCLC progression, in which AGE modifications might play a critical role., (Copyright © 2011 S. Karger AG, Basel.)

The aim of study was to investigate the pattern of mediastinal lymph node metastases in patients with colorectal cancer metastasis. Twenty-four pulmonary metastasectomies with mediastinal lymphadenectomies were performed on 19 patients (14 unilateral and five bilateral operations). The metastases were centrally localised in eight cases; the primary tumour was colon cancer in 15 patients and rectal cancer in nine cases. The number and the localisation of metastases were recorded, as the clinico-pathological data of the primary tumours. The results were compared with the pattern of metastases in mediastinal lymph nodes. The data were subjected to statistical processing with the chi(2)-test and Mann-Whitney test. Mediastinal lymph node metastases were confirmed in eight cases (33.3%). The proportion of positive lymph nodes was significantly higher for central metastases (62.5% vs. 18.8%, P=0.032). When the pathological stage of the primary tumour was more advanced, the proportion of lymph node metastases displayed a statistically not significant increase. The pattern of lymph node metastases did not correlate with the localisation of the lung metastases, disease-free interval and the diameter of the greatest pulmonary metastasis. The frequency of lymph node metastasis is relatively high, therefore, mediastinal lymphadenectomy during the resection of colorectal cancer metastases is necessary., (2010 Published by European Association for Cardio-Thoracic Surgery. All rights reserved.)

The omicron-acyltransferase porcupine contributes to secretion and function of Wnt signaling molecules, which stimulate the expression of various cancer-related genes. Porcupine is also involved in the Wnt-induced cell signaling via beta-catenin in non-small cell lung carcinoma (NSCLC) cells. Herein, we report that the expression level of porcupine in human NSCLC tissues (n=89) positively correlates with the expression of several genes coding for cancer-related molecules such as beta-catenin, hypoxia-inducible factor-1alpha and jun B. However, the mRNA expression of porcupine was not generally increased in NSCLC compared to normal lung tissues. In NSCLC tissues we also found a positive correlation between the expression level of porcupine and the calcium-binding protein S100P, which contributes to initiation and invasion of cancer cells. Subsequent studies showed that the DNA hypomethylation with 5-aza-2'deoxycytidine increased the mRNA expression of S100P in NSCLC cells but did not alter that of porcupine. Silencing the expression of porcupine with small interfering (si)RNA reduced the expression of S100P in NSCLC cells, whereas silencing the expression of S100P with siRNA did not effect the level of Porcupine expression. In conclusion, besides DNA methylation processes, porcupine might regulate the expression of some cancer-related molecules including S100P in human NSCLC.

Purpose: Patient survival after resection of non-small cell lung cancer (NSCLC) strongly correlated with the occurrence of distant metastasis. Cathepsins are members of the lysosomal cysteine proteases family and can support the metastatic process by degrading the extracellular matrix. The purpose of this study was to identify members of the Cathepsin family that correlate with recurrence-free and overall survival of NSCLC patients., Patients and Methods: The expression of 13 Cathepsins was examined using DNA-microarray technology in tumor tissues of 89 surgically treated NSCLC patients. All NSCLC samples were classified according to median Cathepsin expression value into either a high or a low expression group. All Cathepsin expression groups were subjected to clinical prognostic analyses regarding survival and local as well as distant recurrences., Results: Patients with high Cathepsin C tumor expression showed higher tumor recurrence rate compared to patients with low Cathepsin C expression (p = 0.02). The tumor expression of Cathepsins K and B significantly correlated with recurrence-free and overall survival as determined by multivariate analysis. A high expression of Cathepsin B or K was associated with a considerable reduction of recurrence-free as well as overall survival. NSCLC patients with a high expression of both Cathepsin B and K had a significantly (p = 0.001) poorer outcome (5-year survival rate: 13%) than patients with low expression of both genes (5-year survival rate: 75%)., Conclusions: The combined expression level of Cathepsins B and K identifies high-risk NSCLC patients. A selection of gene expression panels is theoretically superior to established clinical and pathological criteria.

The efficient treatment of lung carcinomas with chemotherapeutics still poses a challenge for anti-cancer therapy. Since stromal cells of the tumor may alter the responsiveness of tumor cells to chemotherapeutics, we studied the impact of lung fibroblasts (WI-38) on the chemotherapy-induced death of non-small cell lung carcinoma cells (H358). Conditioned medium from WI-38 fibroblasts impaired the H358 cell death induced by paclitaxel but not by cisplatin. Comparable results were observed when culturing H358 cells in conditioned medium from primary tumor fibroblasts or co-culturing H358 cells with fibroblasts. This anti-apoptotic effect induced by paracrine signaling from fibroblasts was associated with less necrosis (membrane leakage, mitochondrial dysfunction) and apoptosis (outer membrane phosphatidylserine exposure, pycnotic nuclei, nuclear translocation of the apoptosis-inducing factor from mitochondria, caspase-9 and -3/-7 activation) in response to paclitaxel but not cisplatin. Additionally, we demonstrated that WI-38 fibroblasts mediate activation of both extracellular signal-regulated kinases (Erk) 1/2 and Akt kinase in H358 cells. Subsequent application of specific inhibitors revealed that the paclitaxel-induced cell death is highly impaired by active Erk1/2 and Akt, whereas the cisplatin-induced cell death is independent of both kinases. Pro-apoptotic Bcl-2 family proteins such as Bim cannot explain the differential impact of fibroblasts on the H358 cell death induced by paclitaxel compared with cisplatin. Our data support the preferential use of the cisplatin-based lung carcinoma therapy, because the cell death-inducing efficiency of cisplatin is not impaired by stromal fibroblasts.

Lung adenocarcinoma is characterised by an upregulation of S100P, which mediates its function intracellularly but also extracellularly. Recent studies suggest that extracellular S100P contributes to tumour development following interaction with the receptor for advanced glycation end-products (RAGE). As RAGE is highly downregulated in lung cancer, one might speculate that S100P supports tumorigenesis via other pathways. Here, we showed that S100P primarily localises in the cytoplasmic and nuclear region as determined for lung adenocarcinoma specimens (immunohistochemistry) and H358 lung adenocarcinoma cells stably overexpressing S100P (GFP-S100P; fluorescence microscopy). S100P overexpression in H358 cells induced a more frequent formation of tumour colonies in vitro (soft agar assay). However, the S100P-overexpressing cell colonies showed a smaller colony size. This observation was supported by proliferation assays demonstrating a reduced proliferation per increasing cell density of the H358 cells overexpressing S100P. Migration of S100P-overexpressing H358 cells was diminished (transwell migration and in vitro wound scratch assays). Inhibiting DNA methylation with 5-aza-2'deoxycytidine enhanced the mRNA expression of S100P, whereas high S100P levels following overexpression suppressed the mRNA expression of endogenous S100P. In this regard, an upregulation of S100P in lung adenocarcinomas was only determined for early/T1 stage but not more advanced/T2 stage tumours compared with normal lung tissues. Thus, S100P induction may be considered an important step in the initial stage of lung adenocarcinomas, whereas its downregulation in advanced stages seems to be important for tumour progression in which DNA methylation and/or feedback transcription processes play a critical role.

Objective: The incidence of second primary malignancies seems to be increasing. The aim of this study was to investigate the incidence, treatment and outcome for patients with second primary lung cancer (SPLC)., Methods: Between January 1996 and December 2005, 163 patients with SPLC, occurring after an extrapulmonary malignancy, were recruited by the Tumor Center of Halle (Saale), which represents a region of nearly 1.0 million inhabitants in Germany. The SPLCs were treated under curative aim (n=59), with palliative intend (n=76) or best supportive care (n=28)., Results: The incidence of SPLC was 1.6 per 100,000 inhabitants. The localization of the first tumor differed depending on the sex of the patients. The actuarial 5-year survival rate of all patients was 12.7% (median survival time 11.4 months). Univariate analysis revealed treatment strategy as a prognostic factor (p=0.0001). Patients with SPLC having undergone curative treatment turned out to have the best prognosis (median survival: 31.0 months). The Cox proportional hazards model demonstrated that only TNM-staging system was a multivariate and significant independent prognostic predictor for overall survival. The method of surgery, standard lung resection (e.g. lobectomy) versus limited resection had no considerable influence on overall survival (p=0.22), respectively recurrence-free survival (p=0.55)., Conclusions: In cases of operability, standard resection must be the method of choice, because of its best survival rates. The results support the demand of an exact and short-term oncological care system to detect early stages of SPLC for patients operated upon for tumors at different sites.

Early and correct diagnosis of non-small cell lung carcinoma (NSCLC) is essential for the choice of an appropriate anti-cancer therapy. Besides the histopathological diagnosis, molecular profiling by detection of the tumour-associated gene expression might play an upcoming role. As proteins of the S100 gene family show a distinct cell type-specific expression profile, our study focused on the relevance of the S100 family for identification and classification of NSCLCs. Among the S100 members, we identified the expression of S100A1, S100A2, S100A4, S100A6, S100A9 and S100P in human lung carcinoma cells (H358(p53-), A549(p53+)) or NSCLC tissues. Distinct S100 members are increased in NSCLCs compared with control lung specimens depending on the histopathological subtype. In particular, S100A2 was upregulated in squamous cell carcinomas, whereas S100P was mainly increased in adenocarcinomas. The upregulation of either S100A2 or S100P was detected in early but less in advanced tumour stages and not at all in NSCLC patients who had received neoadjuvant chemotherapy. In conclusion, our study indicates an important role of the S100A2-S100P expression profile for molecular diagnosis of NSCLCs at early and, therefore, prognostically more favourable tumour stage. As the S100A2-S100P profile also allows the histopathological classification, it might significantly support the conventional tumour diagnostics.

Improvements in detection, treatment and prognosis for patients with non-small cell lung cancer (NSCLC) depend on the molecular understanding of tumor development and progression. Using Affymetrix GeneChips comprising 59,620 elements, we determined the gene expression profiles of 89 NSCLC and 15 normal lung samples. We found 187 (0.3%) genes, which are at least 2-fold overexpressed and 157 (0.3%) genes 2-fold less expressed in NSCLC compared with normal lung. Cell cycle regulation, cell adhesion and nucleotide metabolism were the major biological processes connected to a large proportion of genes up-regulated in NSCLC. Down-regulated genes were frequently involved in metabolic/catabolic processes and signal transduction. The expression of specific genes revealed reliable differentiation of tumor from normal lung tissues, as well as the classification of both NSCLC subtypes squamous cell carcinoma and adenocarcinoma. In this context, collagens (COL7, 17) and cytokeratins (CK6, 15, 17) are preferentially induced in squamous cell carcinoma, whereas several transcription factors (TTF1, DAT1, TF-2) are exclusively elevated in adenocarcinomas. Some gene products involved in the metastatic process [matrixmetalloproteinase 12 (MMP-12) and urokinase plasminogen activator alpha (uPA)] were found as prognostic markers for the recurrence free interval and survival. Particularly, the simultaneous use of the MMP-12 and uPA expression predicted relapse-free and global survival of the patients. Screening of NSCLC with a genome-wide array revealed diagnostic, prognostic and potential therapeutic targets that might be suitable for an individual risk profile by tumor specific arrays.

Lung cancer is still considered the leading cause of cancer-related deaths in industrialized countries. Despite improvements in cancer diagnosis and therapy, the present 5-year survival rate of 14% is only slightly higher than the survival rate of 8% in the early 1960s. At present, a survival plateau has been reached by the current management of lung cancer, including surgery, chemo- and radiotherapy. Although prevention and early diagnosis are undoubtedly important, more innovative treatment approaches and new agents that affect alternate mechanisms in tumor cells are needed. Since molecular targets of tumorigenesis and metastasis are currently the focus of lung cancer research, this review will summarize the most important molecular approaches and clinical trials of novel therapeutics.

Expression profiling by gene microarray techniques have been developed to predict malignant tissue but there are no experiences with the application of antibody arrays to identify malignancy-related proteins. Because altered protein patterns might also better interpret biological processes, we applied tumour samples from 12 patients with squamous cell lung carcinoma and individual lung tissue controls to antibody arrays spotted with 378 distinct monoclonal antibodies. Array analysis defined 20 proteins with higher and nine with lower abundance in lung tumours. Comparison with gene microarray data revealed that 31% of the differentially regulated proteins correlate with altered mRNA expression in squamous cell lung carcinomas, including PEX1, MKK7 and HDAC3 for up-regulated proteins. The histone deacetylase (HDAC) 3 was investigated in detail by immunoblot analysis showing that HDAC3 is indeed elevated in 92% of tumours (n=22/24; P<0.001). Thus, antibody microarrays can be useful for detection of some target proteins related to lung cancer.

Objective: Pulmonary metastasectomy as well as immunotherapy have reproducible, albeit limited efficacy in advanced renal cell carcinoma (RCC). We examined whether metastasectomy improved overall survival compared with results of immunotherapy., Methods: Between 1975 and 2003, 64 patients (41 men, 23 women) underwent pulmonary resection of metastatic RCC. Only patients who met the criteria for potentially curative operation, that means, control of primary tumor, ability to resect metastatic disease and no other extrapulmonary metastases, were included., Results: The overall 5-year survival was 33.4% (median survival: 39.2 months). A significant longer survival was observed using multivariate analysis in patients with complete pulmonary resection (R0), with a 5-year survival of 39.9% and a median survival of 46.6 months in correlation to patients with incomplete resection (5-year survival 0%, median survival 13.3 months). In multivariate analysis patients with synchronous metastases had a significant worse prognosis in correlation to patients with metachronous metastases. The 5-year survival of curative resected patients with metachronous metastases was 43.7% versus 0% for synchronous metastases, respectively. In patients with solitary metastasis and R0 resection, we observed a 5-year survival of 49%, whereas the rate was 23% in patients with more than a single metastasis. When establishing prognostic groups as suggested by the International Registry based on the risk factors disease-free interval, number of metastasis and complete resection the group with the best prognosis showed a 5-year survival of 52% (median survival 75.2 months)., Conclusion: Metastasectomy nowadays is the best treatment option in cases with technical resectable metastases with as much as possible good prognostic factors (metachronous metastases with long DFI, number up to 6 metastases).

Primary pulmonary rhabdomyosarcoma is a rare entity and the histological differential diagnosis can be difficult. We report on a 43-year old female patient, smoker (25 pack-years), in whom a large solitary brain metastasis was diagnosed and enucleated. Histological examination revealed a typical small cell carcinoma and histological examination of biopsies obtained from a tumor in the left upper lobe of the lung was compatible with a small cell carcinoma. Despite chemotherapy there was a progressive tumor growth. Bronchial biopsies again showed a small cell tumor, although immunohistochemistry proved it to be a pleomorphic rhabdomyosarcoma. Due to the progressive tumor growth with necrosis and superinfection and a lack of further metastases lobectomy of the left upper lobe was performed, complicated by postoperative pleural empyema, limiting the possibilities of adjuvant therapy. Early relapse occurred with pleural, pulmonary, chest wall and spinal metastases. Laminectomy and extirpation of the spinal metastases, local radiotherapy and chemotherapy with iphosphamide and doxorubicine led to partial remission and clinical improvement for few months only. The patient died from metastatic primary rhabdomyosarcoma of the lung. This rare tumor mimicked small cell lung cancer. Appraisal of the atypical clinical course and a close dialogue between pathologists and clinicians enabled the correct diagnosis.

In this study, microarray analysis was used to identify tumour-related genes that were down regulated in lung carcinoma. The promoter sequences of the identified genes were analysed for methylation patterns. In lung cancer cell lines, CpG island methylation was frequently detected for TIMP4 (64%), SOX18 (73%), EGF-like domain 7 (56%), CD105 (71%), SEMA2 (55%), RASSF1A (71%), p16 (56%) SLIT2 (100%) and TIMP3 (29%). Methylation was however rarely observed in cell lines for SLIT3 (18%) and DLC1 (18%). In primary lung tumours, methylation of TIMP4 (94%), SOX18 (100%), EGF-like domain 7 (100%), CD105 (69%), SEMA2 (93%), DLC1 (61%), RASSF1A (44%), p16 (47%), SLIT2 (100%) and TIMP3 (13%) was also detected. Methylation of several CpG islands was frequently found in normal lung tissue of cancer patients and this may have been attributed to epigenetic field defect and/or infiltrating tumour cells. Interestingly, inactivation of RASSF1A and p16 correlated well with an extended smoking habit (P=0.02), and exposure to asbestos (P=0.017) or squamous cell carcinoma (P=0.011), respectively. These results have identified genes whose aberrant promoter methylation could play a crucial role in the malignancy of lung carcinoma.

The receptor for advanced glycation end-products (RAGE) is a transmembrane receptor of the immunoglobulin superfamily. Several ligands binding to RAGE have been identified, including amphoterin. Experimental studies have given rise to the discussion that RAGE and its interaction with amphoterin contribute to tumour growth and metastasis. However, none of the studies considered a differential transcription profile in cancer that might change the interpretation of the study results when comparing RAGE in tumours with histologically normal tissues. Here we show that RAGE is strongly reduced at the mRNA and even more so at the protein level in non-small cell lung carcinomas compared with normal lung tissues. Down-regulation of RAGE correlates with higher tumour (TNM) stages but does not depend on the histological subtypes, squamous cell lung carcinoma and adenocarcinoma. Subsequent overexpression of full-length human RAGE in lung cancer cells (NCI-H358) showed diminished tumour growth under some conditions. While proliferation of RAGE-expressing cells was less than that of cells expressing the cytoplasmic domain deletion mutant DeltacytoRAGE or mock-transfected NCI-H358 in monolayer cultures, RAGE cells also formed smaller tumours in spheroid cultures and in vivo in athymic mice compared with DeltacytoRAGE cells. Moreover, we observed a more epithelial growth of RAGE-expressing, but also of DeltacytoRAGE-expressing, cells on collagen layers, whereas mock NCI-H358 cells kept their tumour morphology. This observation was supported by immunofluorescence analyses demonstrating that RAGE preferentially localizes at intercellular contact sites, independent of expression of the cytoplasmic domain. Thus, down-regulation of RAGE may be considered as a critical step in tissue reorganization and the formation of lung tumours.

Purpose: Non-small cell lung cancer (NSCLC) is a very common and aggressive malignancy. Survival after resection of tumor is especially determined by the occurrence of distant metastasis. Matrix metalloproteinases (MMP) support this metastatic process by degradation of the extracellular matrix., Experimental Design: We used DNA microarray technology to examine the expression of 22 MMPs in 89 surgically treated NSCLC patients. Validation of microarray results was done using reverse transcription-PCR and immunohistology., Results: MMP-1, MMP-9, and MMP-12 expression was significantly increased in tumors versus corresponding lung tissues. MMP-12 expression significantly correlated with local recurrence and metastatic disease. Multivariate Cox regression analysis revealed MMP-12 expression as an independent prognostic factor for tumor relapse-free interval. Gene expression analysis of 158 healthy tissues from 32 different organs revealed no MMP-12 expression in these organs and immunohistology identified MMP-12 protein in NSCLC only in tumor cells., Conclusions: MMP-12 might be not only a prognostic marker, but also a valuable therapeutic target.

Simple tools for discrimination of lung tissues can be useful in a fast machine-aided diagnosis, for example, by tumor-specific microarrays. We demonstrate that an easy ratio technique, based on the expression levels of only two genes differentially expressed in lung tumor and normal lung samples, allows discrimination of normal and neoplastic lung with a sensitivity of 100% and specificity of 90.5%. DNA microarray analysis of 99 lung tumor samples and 15 normal lung tissues revealed that receptor for advanced glycation end products (RAGE) mRNA is reduced fourfold (p = 7.8 x 10(-11)) and cyclin-B2 mRNA is upregulated twofold (p = 5.9 x 10(-18)) in lung carcinoma compared with normal lung. The microarray-calculated expression ratio of RAGE to cyclin-B2 was used in polymerase chain reaction analysis of 84 independent blinded samples to discriminate tumor and corresponding normal lung tissues. In 94.7% of the samples this quotient correctly distinguished non-small cell lung cancer from normal lung tissue, suggesting the RAGE/cyclin-B2 quotient as a potential means for diagnosis of lung cancer.

Purpose: Apoptotic cell death contributes to the regulation of tumour regression but can be prevented by proteins of the IAP family. Although survivin can be identified as tumour-specific gene product, the role of other members of the IAP family is mainly unclear in non-small cell lung cancer (NSCLC). Therefore, we hypothesise that hIAP-1, hIAP-2, and XIAP are associated with lung carcinogenesis, too., Methods: To define IAP expression levels, lung tumour samples from 34 NSCLC patients with adenocarcinoma (16) and squamous cell carcinoma (18) were included. Analyses were performed by standardised RT-PCR and immunoblotting. Paired non-tumour lung tissues served as controls. All tumour samples showed a strong survivin mRNA up-regulation compared with non-tumour controls., Results: Investigations of the XIAP mRNA expression revealed an overall increase in lung carcinoma (median: 1,083 vs 605 rel. U; P =0.02). In contrast, hIAP-2 mRNA was nearly identical in all tumour and control samples. Furthermore, we identified an elevated hIAP-1 mRNA expression especially in patients with adenocarcinoma (median: 8.58 vs 3.44 rel. U; P <0.01). Using the median increase of 50% determined in all tumours as a cut-off point, 11/16 patients with adenocarcinoma but only 6/18 with squamous cell carcinoma showed an elevation in hIAP-1 mRNA. This hIAP-1 up-regulation could be mainly observed in low TMN adenocarcinomas (7/9 in TMN I, median increase: +289% vs 2/5 in TMN III-IV, +44.6%). An enhanced mRNA expression of hIAP-1 and XIAP in respective tumours could be confirmed on protein level by immunoblot analyses., Conclusions: Our results indicate an involvement of the anti-apoptotic XIAP in pathogenesis of NSCLC, while hIAP-1 preferentially seems to play an important role in low-stage adenocarcinoma.

Objectives: This retrospective study evaluates the probability of survival in patients who had undergone resection for non-small cell lung cancer (NSCLC) and in whom residual disease at the resection margins was found., Methods: During a period of 6 years, 596 patients with NSCLC were operated upon with curative intention. Residual disease at the resection margin was divided into microscopic (R1) and macroscopic (R2)., Results: Twenty-six patients (4.4%) showed R1 and 12 (2%) R2 residual disease. An extrabronchial (thoracic wall, vessels) R1 situation was found in five patients and a bronchial R1 infiltration in 21 cases. The bronchial resection margin was subject to peribronchial infiltration in most cases (16/21). A total of 17/21 (65%) patients with bronchial infiltration had N2 disease. Thirty day lethality was 3.8% in the R1 group. Fifteen patients had postoperative irradiation. The 5-year survival rate for patients with R1 resection was 14%. The differences in survival between patients with extrabronchial vs. bronchial infiltration and N0/N1 vs. N2 were significant using univariate analysis. Adjuvant radiation did not result (especially in N2 disease) in a survival benefit. Among 12 patients with macroscopic residual disease (R2), 3/12 (25%) died within the first 30 days after the operation, and none of the R2 patients survived the first year after the operation., Conclusions: Patients with an R1 situation have a survival rate of 14% comparable to curative resected patients (RO) in stage III. Adjuvant radiation had no clear effect on survival. Patients with macroscopic tumor (R2) should receive palliative treatment after the operation depending on their condition.

Background: The aim of our study was to examine the significance of tumour DNA-content and proliferation in lung cancer., Methods: The DNA content and S-phase fraction (SPF) was determined by flow cytometry in 125 resected tumours of patients with non-small cell lung cancer. In 40 cases we compared the SPF with immunohistochemical staining of the Ki-67 protein using MIB-1 antibody., Results: DNA aneuploidy was detected in 84.8% (106/125). Cell cycle analysis for the determination of proliferation activity was only possible in 69 (55.2%) cases. An SPF of 0-8% as a sign of low proliferation was found in 27 specimens. In advanced tumours at stage III and IV the proportion of tumours with SPF 9-16% was significantly (p<0.05) increased as compared to tumours at stage I and II. There was a significant correlation (p=0.012, ascent: 0.045) between SPF and MIB-1. Patients with aneuploid tumours had a relative risk of 1.4 to die earlier than patients with diploid tumours. Patients with SPF of 9-16% in the tumour tended to decreased survival (5-year survival rate: 29%) in correlation to patients with a percentage of SPF 0-8% (5-year survival rate: 38%, p=0.5). These differences were significant (p=0.048) in patients with adenocarcinomas only. In the multivariate COX-regression model age (p=0.03) and stage (p=0.0001) were significant prognostic factors, ploidy state (p=0.33) was of no prognostic significance., Conclusions: Flow cytometry seems to be a useful method for understanding the clinical behaviour of lung cancer. Especially the SPF in adenocarcinomas may be used as a prognostic indicator.

Between 1975-1996, 39 patients underwent resection of pulmonary metastases from renal cell carcinoma. Multivariate analysis (COX model) for survival of preoperative risk factors showed that time of diagnosis (syn-/metachronous) of the metastases (p = 0.05) and the number of metastases (p = 0.01) were of prognostic significance. It is concluded that metastasectomy in patients with not more than six metachronous metastases after renal cell carcinoma has significant benefit and in cases of synchronous metastases or more than 6 pulmonary metastases indication for resection should be restricted.

The adenosquamous carcinoma is a rare combined tumour of non-small cell lung cancer (NSCLC). The survival prognosis of surgically treated patients with adenosquamous carcinoma and patients with squamous cell carcinoma, large cell carcinoma or adenocarcinoma were compared during a study. Two hundred and seventyfive patients who had been treated surgically because of primary lung cancer in the Department of Thoracic-Surgery at the Martin-Luther-University Halle-Wittenberg between 1980 and 1989 were evaluated. The five year survival study of 172 patients who underwent resection because of squamous cell carcinoma was 45%, the one of patients with adenocarcinoma (n = 84) was 27%. 26% was the five year survival rate of the patients (n = 9) with large cell carcinoma. Of 13 patients (4%) with adenosquamous carcinoma none survived five years after surgical treatment. The two year survival rate was 28%. The presented results demonstrate the poor survival prognosis of patients suffering from adenosquamous carcinoma and ask for an adjuvant therapy.