Elevated expression of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM-1) is associated with increased angiogenic potential in non-small-cell lung cancer.

Recent studies have challenged the previously postulated concept of a tumor-suppressive effect of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM-1). A possible angiogenic influence of CEACAM-1 in non-small-cell lung cancer (NSCLC) has not been investigated so far. Therefore, we examined microvessel density (MVD) and CEACAM-1 expression in primary NSCLC and analyzed their possible correlations under consideration of their prognostic effects. Specimens from 82 consecutive patients with completely resected NSCLC were stained immunohistochemically using the monoclonal anti-CEACAM-1 antibody 4D1/C2 and the monoclonal anti-CD31 antibody JC70A. The prognostic relevance of CEACAM-1 expression and MVD was evaluated by univariate Kaplan-Meier and multivariate Cox regression analysis. The median follow-up period was 75 months (range 10-156 months). A high MVD (i.e., > or =31microvessels/400x microscopic field) was observed more frequently in tumors with high CEACAM-1 expression (i.e., >/=66% stained tumor cells) than in tumors with low CEACAM-1 expression (61.8% vs. 33.3%, respectively; p=0.01). In univariate survival analyses, high CEACAM-1 expression and high MVD were associated with development of distant metastasis (p=0.011 and 0.022, respectively) and decreased cancer-related survival (p=0.046 and 0.006, respectively). Multivariate Cox regression analysis demonstrated that the prognostic impact of CEACAM-1 depended on the prognostic influence of MVD, while MVD itself represented an independent prognosticator for unfavorable cancer-related survival (p=0.021; relative risk 2.1; 95% confidence interval, 1.1-4.0). Here we show for the first time that high CEACAM-1 expression is associated with an increased angiogenic activity in NSCLC, and that the prognostic influence of CEACAM-1 might be derived from this association.

Sublobar resections in stage IA non-small cell lung cancer: segmentectomies result in significantly better cancer-related survival than wedge resections.

OBJECTIVE: Sublobar resections spare pulmonary function and offer a method of increasing resection rates in patients with lung cancer and limited functional operability. Previous studies demonstrated an increased local recurrence rate following wedge resections compared to segmentectomies in stage IA non-small cell lung cancer (NSCLC). However, a prognostic impact of this observation has never been shown and is still under debate. Therefore, this study has been performed to analyse the cancer-related survival of sublobar resections in stage IA patients. METHODS: Over a 17-year period 87 patients underwent sublobar complete resection (R0) of stage IA NSCLC via thoracotomy. Sublobar resection was reserved for patients with cardiopulmonary impairment. Wedge resections with selective lymphadenectomy were performed in 31 patients (36%) and segmentectomies with systematic lymphadenectomy in 56 patients (64%). Patient characteristics, functional parameters, tumour specifics and follow-up duration were analysed concerning their distribution between the two groups. Kaplan-Meier curves were compared and possible joint effects between prognostic parameters were analysed by multivariate Cox regression analysis. RESULTS: The median follow-up duration was 45 months. There was no significant difference between the two groups in gender (p=0.11), age (p=0.08), American Society of Anesthesiology physical performance status (ASA)-score (p=0.32), forced expiratory volume in 1s FEV(1) (p=0.08), tumour size (p=0.30), histology (p=0.17), grading (p=0.12), complication rate (p=0.15) and follow-up duration (p=0.29). The mean number of dissected lymph nodes in segmentectomies (12+/-6) was higher than in wedge resections (6+/-3) (p=0.0001). The 5-year survival rate was 63%. There were significantly less locoregional recurrences (p=0.001), an equal distribution of distant metastases (p=0.53) and a better cancer-related survival (p=0.016) following segmentectomies compared to wedge resections. Cox regression analysis showed that the prognostic effect of the resection type was independent from gender, age, ASA-score, respiratory function, tumour size, tumour histology, grading and number of dissected lymph nodes (p=0.04, relative risk 1.16). CONCLUSIONS: Studies investigating survival after sublobar resection of stage IA NSCLC should always distinguish between anatomical segmentectomies and wedge resections. If limited functional operability requires a sublobar resection of stage IA NSCLC, segmentectomy with systematic lymphadenectomy should be preferred.

Frequency of local recurrence following segmentectomy of stage IA non-small cell lung cancer is influenced by segment localisation and width of resection margins--implications for patient selection for segmentectomy.

OBJECTIVE: Segmentectomy has recently been suggested as alternative to lobectomy for curative treatment of early-stage non-small cell lung cancer (NSCLC). This study was performed to investigate if localisation of the resected segment or width of resection margins influence local recurrence following complete segmentectomy of stage IA NSCLC. METHODS: Between 1987 and 2002, 49 segmentectomies and 150 lobectomies were performed in patients with pT1pN0cM0-NSCLC in our institution. Indications for segmentectomy were a limited pulmonary function or severe comorbidity. The median follow-up duration was 54 months. Local recurrence was distinguished from secondary primary lung cancer and was defined as tumour within the same lung or in the ipsilateral mediastinum. Segment localisation, width of resection margins, tumour size, tumour type, grading and age were analysed concerning their influence on local recurrence. RESULTS: Local recurrence occurred in 16% of patients with segmentectomy and was significantly more frequent than in patients with lobectomy (5%; p=0.005; log-rank test). Segmentectomy in the S1-3 region tended more frequently to local recurrence than segmentectomy in the remaining segments (p=0.08; log-rank test): There was no recurrence following segmentectomy in the S7-10 region (n=6) or of S4-5 (n=5). Recurrence occurred in 7 (23%) out of 30 patients with segmentectomy in the S1-3 region and in 1 (12%) out of 8 patients with S6-segmentectomy. Also, resection margins < or = 1cm tended to be associated with local recurrence (p=0.06; log-rank test). CONCLUSIONS: The frequency of local recurrence following segmentectomy might be influenced by segment localisation and width of resection margins. Segmentectomy within the S1-3 region should be avoided whenever possible.

Inhibition of T cell homing by down-regulation of CD62L and the induction of a Th-2 response as a method to prevent acute allograft rejection in mice.

OBJECTIVE: For a successful immune response, migration of lymphocytes to lymphoid organs and other tissues is a key step, as the initial recognition of foreign antigens and activation of lymphocytes takes place in these organs. CD62L is a homing receptor that mediates entry of naïve T cells to peripheral lymph nodes. Maybe the preventing of T cell homing will change the immune response against allogeneic tissue and suppress rejection. METHODS: We treated different mouse strains with pertussis toxin to manipulate T cell homing and measured the rejection of allografts in terms of allogeneic tumor cells. We transferred pertussis toxin treated or nontreated transgenic T cells into BALB/c wild type mice. The transgenic T cells could be followed ex vivo by specific antibodies. Cytokine production from purified (1x10(5)/ml) T cells after different stimulations in vitro and expression of surface markers on T cells following pertussis toxin treatment by FACS analysis were performed. RESULTS: Pertussis toxin-treated C57BL/6 mice with the MHC class I molecule H-2K(b) could not reject allogeneic tumor cells R1.1, which expressed the MHC class I molecule H-2K(k) and were killed by these cells. This allograft survival could be demonstrated for various allogeneic cells in different mouse strains with different MHC class I expression and emphasizes the general mechanism in these studies. In vivo CD62L expression on T cells was down-regulated by pertussis toxin in normal mice and transgenic mice that produce only one specific T cell, and after the pertussis toxin treatment the mice showed 4-5 times larger spleens compared to untreated mice. In transfer experiments, we demonstrated that CD62L low transgenic T cells could not home to lymph nodes. Furthermore, spleen cells from pertussis toxin-treated mice produced high amounts of the Th-2 cytokine interleukin 4 after stimulation in primary culture. CONCLUSIONS: Our data suggest that the inhibition of T cell homing changes the immune response. Prevention of homing of T cells in combination with the induction of a Th-2 response is a mechanism to prevent specific acute rejection of allogeneic tissue.

Uptake of (201)Tl into primary cell cultures from human thyroid tissue is multiplied by TSH.

UNLABELLED: According to current guidelines, (201)Tl scintigraphy aiming at the detection of iodine-negative metastases of differentiated thyroid carcinoma is usually performed during hormone replacement in thyroid-stimulating hormone (TSH)-suppressive doses. The aim of this study was to determine the effect of TSH on thyroid (201)Tl uptake in vitro. METHODS: Adherent monolayers of human thyroid tissue (1.965-3.000 million cells) were cultured after mechanical disintegration and enzymatic digestion by neutral protease. The samples were derived from patients undergoing surgical treatment of nodular goiter. Studies were performed on paranodular tissue components, exclusively. Cells originating from identical tissue samples were dispensed into matched-pair cultures and incubated in parallel by a TSH-free medium and by a medium containing 10 IU/L bovine TSH. Thyroglobulin (Tg) accumulation was calculated after repetitive measurements of Tg concentrations by radioimmunoassay. Uptake studies were initiated by adding fresh medium containing (201)Tl (8.2-91.0 kBq). After 1 h of incubation, the media were removed and the cells were detached by trypsin and collected by centrifugation. Uptake was measured by a gamma-counter, and cellular uptake values were calculated as percentages of total activity normalized to 1 million cells. The statistical significance of differences in Tg release and (201)Tl uptake was corroborated in a generalized estimating equations analysis taking the variability of unbalanced replicate measurements into account. RESULTS: Cells cultured in the presence of TSH displayed a 2-fold release of Tg (12.949 ng/h/million cells vs. 6.049 ng/h/million cells, P = 0.001) and triplicate (201)Tl uptake (0.718%/million cells vs. 0.249%/million cells, P = 0.0002). CONCLUSION: (201)Tl uptake in human thyroid cells is significantly increased by TSH. These data suggest that withdrawal of thyroid hormone substitution has the potential to improve the sensitivity of (201)Tl scintigraphy for detecting thyroid remnants or cancer recurrences. This suggestion should be further investigated in patients and in cell cultures from thyroid carcinomas.

The impact of surgery and mild hyperthermia on tumor response and angioneogenesis of malignant melanoma in a rat perfusion model.

BACKGROUND: The aim of this experimental study was to determine the effect of mild hyperthermia on tumor response and angioneogenesis in an isolated limb perfusion model with a human melanoma xenograft. METHODS: A human melanoma xenograft was implanted into the hindlimbs of 30 athymic nude rats. The animals were randomized into five groups: group I: control, group II: sham group, group III: external hyperthermia with a tissue temperature of 41.5 degrees C for 30 minutes without ILP, group IV: normothermic ILP (tissue temperature 37 degrees C for 30 minutes, group V: hyperthermic ILP (tissue temperature 41.5 degrees C for 30 minutes). Tumor response was evaluated by tumor size determination and immunohistochemical analysis 6 weeks postoperatively. Tissue sections were investigated for expression of CD34 and basic fibroblast growth factor (bFGF). RESULTS: Average tumor volumes of the controls (I) increased from 105 mm3 to 1388 mm3. In the sham operated group (II) tumor volumes were significantly larger than in group I. Tumor volumes in group IV were significantly smaller than in group I and lowest in group V. There were no significant differences in size between group I and group III after six weeks. In group III and IV each, 5 animals showed tumor progression and one had a partial tumor response. In group V only 2 animals showed tumor progression. Immunhistochemical analysis of the tissue sections demonstrated that angioneogenesis was more pronounced in group II than in group I and less pronounced in group IV and V compared with group I. CONCLUSIONS: Our results suggest that even a surgical manipulation such as a skin incision promotes tumor growth, probably by induction of growth factors like bFGF. External hyperthermia of 41.5 degrees C tissue temperature for 30 minutes only has no impact on tumor growth and angioneogenesis in vivo.

Neuroendocrine tumors of the bronchopulmonary system (typical and atypical carcinoid tumors): current strategies in diagnosis and treatment. Conclusions of an expert meeting February 2011 in Weimar, Germany.

Neuroendocrine tumors (NETs; syn. carcinoid tumors) are highly or moderately differentiated neoplasms. They comprise a large variety of rare and heterogeneous tumors with an estimated incidence of 3-5/100,000/year. They can arise in virtually every internal organ, but mainly occur in the gastroenteropancreatic and bronchopulmonary systems. Around 25% of the NETs are localized in the bronchopulmonary system. Approximately 2% of all lung tumors are NETs. According to the World Health Organization (WHO) classification of lung tumors, bronchopulmonary NETs are subdivided into typical carcinoids (TCs) and atypical carcinoids (ACs). The parameter with the highest impact on NET behavior and prognosis is the histological classification and staging according to the tumor/node/metastasis (TNM) system. The diagnosis of NETs is established by histological examination and the immunohistochemical detection of general neuroendocrine markers, such as chromogranin A (CgA) and synaptophysin. Serum markers and the use of functional imaging techniques are important additive tools to establish the diagnosis of a NET. The only curative option for lung NETs is complete surgical resection. Beyond that, the currently available interdisciplinary therapeutic options are local ablation, biotherapy (somatostatin analogues), or chemotherapy. New therapeutic options such as peptide receptor radionuclide therapy (PRRT) and molecularly targeted therapies achieve promising results and are under further evaluation. This report is a consensus summary of the interdisciplinary symposium 'Neuroendocrine Tumors of the Lung and of the Gastroenteropancreatic System (GEP NET) - Expert Dialogue' held on February 25-26, 2011 in Weimar, Germany. At this conference, a panel of 23 German experts shared their knowledge and exchanged their thoughts about research, diagnosis, and clinical management of NETs, whereby special attention was paid to NETs of the respiratory tract.

Stromal CD4/CD25 positive T-cells are a strong and independent prognostic factor in non-small cell lung cancer patients, especially with adenocarcinomas.

Within the concert of immune reactions against tumour cells cytotoxic and regulatory T-cells are of utmost importance. Several studies revealed contradictory results on this issue. We therefore focused on functional expression patterns and localization of tumour-infiltrating T-lymphocytes in non-small cell lung cancer (NSCLC) and their impact on patient's survival. 232 curatively operated NSCLC patients were included. After histological reevaluation and construction of tissue-multi-arrays immunohistochemical doublestains for CD3/CD8 and CD4/CD25 were performed to evaluate the total number of T-cells and their subsets of cytotoxic and activated T-cells. Additionally, the localization of the lymphocytes was included in the analysis. Hereby, T-cells within the tumour stroma were regarded as stromal, those among cancer cells as intraepithelial. The number of lymphocytes differed significantly between the histological subtypes being most prominent in large cell carcinomas. Survival analysis showed that high numbers of stromal T-lymphocytes are of beneficial prognostic influence in NSCLC patients. This also proved to be an independent prognostic factor in adenocarcinomas. Thus, in a large and well characterized cohort of NSCLC this is the first study to determine the prognostic value of stromal T-lymphocytes, as these are an independent prognosticator in NSCLC especially in adenocarcinomas whereas intraepithelial T-cells are not.

Impact of chest tube clearance on postoperative morbidity after thoracotomy: results of a prospective, randomised trial.

OBJECTIVE: In many centres of thoracic surgery, milking of chest tubes is performed to prevent them from blocking. The usefulness of chest tube clearance is discussed controversially. Therefore, we investigated the impact of postoperative chest tube milking on postoperative outcome in a prospective, randomised trial. METHODS: Within a period of 11 months, 145 patients undergoing pulmonary resection through thoracotomy were included in the study. Two chest tubes each (silicone drainage, Redax, Mirandola, Italy) were placed in all patients (ventral tube 21Ch and dorsal tube 24Ch). Milking was applied to both chest tubes for 1 min every 2h within the first 48 h postoperatively and continuous suction of -20 cm H(2)O was maintained for 48 h. Duration of chest tube drainage, quantity and quality of effusion or air leakage, co-morbidity, length of hospital stay and 30-day postoperative morbidity and mortality were analysed. Furthermore, outcome was measured by assessment of chest radiographs at the time of discharge from hospital. RESULTS: Randomisation resulted in milking of chest tubes of 73 patients and in observation of chest tubes without any manipulation in 72 patients. Twenty-one patients had to be excluded from further analysis due to violation from the study protocol (n=9), necessity of replacement of a chest tubes (n=9) and re-operation for bleeding (n=3). The 30-day mortality rate was 1.4% in each group and the 30-day morbidity was 49.3% in the milking group and 52.8% in the observation group. Milking of chest tubes was not associated with a lower postoperative mortality or morbidity (p=0.99 and p=0.67, respectively; chi-square test). We observed a significant increase of postoperative pleural effusion drainage in the milking group 48 h after surgery (p=0.004; unpaired t-test). No correlation was seen between milking of chest tubes and the duration of chest tube drainage, quality of effusion, air leakage or length of hospitalisation. CONCLUSIONS: We showed for the first time that postoperative chest tube milking is associated with a significant increase of pleural fluid drainage. Postoperative morbidity and mortality was not improved and therefore chest tube milking cannot be recommended as a routine postoperative procedure.

The role of a pseudocapsula in thymic epithelial tumors: outcome and correlation with established prognostic parameters. Results of a 20-year single centre retrospective analysis.

BACKGROUND: Treatment of thymoma is often based on observation of only a few patients. Surgical resection is considered to be the most important step. Role of a pseudocapsula for surgery, its clinical significance and outcome compared with established prognostic parameters is discussed which has not been reported so far. METHODS: 84 patients with thymoma underwent resection and analysis was carried out for clinical features, prognostic factors and long-term survival. RESULTS: Fifteen patients were classified in WHO subgroup A, 21 in AB, 29 in B and 19 patients in C. Forty two patients were classified in Masaoka stage I, 19 stage II, 9 stage III and 14 stage IV. Encapsulated thymoma was seen in 40, incomplete or missing capsula in 44 patients. In 71 complete resections, local recurrence was 5%. 5-year survival was 88.1%. Thymomas with pseudocapsula showed a significant better survival (94.9% vs. 61.1%, respectively) (p = 0.001) and was correlated with the absence of nodal or distant metastasis (p = 0.04 and 0.001, respectively). Presence of pseudocapsula as well as the Masaoka and WHO classification, and R-status were of prognostic significance. R-status and Masaoka stage appeared to be of independent prognostic significance in multivariate analysis. CONCLUSION: Intraoperative presence of an encapsulated tumor is a good technical marker for the surgeon to evaluate resectability and estimate prognosis. Although the presence of a capsula is of strong significance in the univariate analysis, it failed in the multivariate analysis due to its correlation with clinical Masaoka stage. Masaoka stage has a stronger relevance than WHO classification to determinate long-term outcome.

Characterization of gene expression profiles of T cells during anti-tumor response.

BACKGROUND AND AIMS: T cells of tumor-bearing mice or cancer patients exhibit an immune dysfunction, enabling the tumor to escape immune surveillance. METHODS: The experiments are based on EL4 thymoma cells that were transfected with costimulatory ligands B7-1, B7-2, or both at the same time. We used oligonucleotide-based DNA chip microarrays to characterize the genomic expression profile of peripheral T cells according to their anti-tumor immune response in vivo. These murine T cells were also characterized by ELISA, FACS analysis, and co-stimulatory assays. RESULTS: Using commonly established methods, such as FACS analysis or the analysis of the cytokine profile by ELISA, it was not possible to determine functional differences in the in vivo activity of T lymphocytes against tumor cells. EL4 tumor cells induced multiple anti-tumor immune responses in vivo depending on their B7 expression. We successfully used microarray analysis to identify genes that were differentially expressed in the dysfunctional T cells, which were unable to reject tumors in vivo. Although Th1 and Th2 cytokine expression was not affected, we observed differential expression of genes involved in the regulation of an innate immune response. CONCLUSION: Our results provide evidence that the anti-tumor response can be identified by the "gene profile" of T cells. Genomic scale analysis offers the opportunity to identify subtle changes in gene expression in T cells reflecting a distinct biological behavior in vivo.

Common denominator genes that distinguish colorectal carcinoma from normal mucosa.

PURPOSE: Microarray technology has been used by a growing number of investigators and several studies have been published that list hundreds of genes differentially expressed by colorectal carcinoma (CRC) and normal mucosa (MC). On the basis of our own and other investigators' microarray data, our goal was to identify a common denominator gene cluster distinguishing CRC from MC. METHODS: Thirty GeneChips (HG-U133A, Affymetrix) were hybridized, 20 with RNA of CRC stages I-IV (UICC) and 10 with MC. Expression signals showing at least a 4-fold difference between CRC and MC (p<0.01) were identified as differentially expressed. In addition, in our integrative data analysis approach only those genes whose expression was altered simultaneously in at least 2 of 5 recently published studies were subjected to an unsupervised hierarchical cluster analysis. RESULTS: We detected 168 up- and 283 down-regulated genes in CRC relative to MC. Twenty-three genes were filtered from the five articles reviewed. An unsupervised hierarchical cluster analysis of these 23 genes confirmed the high specificity of these genes to differentiate between CRC and MC in our microarray data. CONCLUSIONS: Colorectal cancer and mucosa could be clearly separated by 23 genes selected for being differentially expressed more than once in a recent literature review. These genes represent a common denominator gene cluster that can be used to distinguish colorectal MC from CRC.

Tissue preparation for gene expression profiling of colorectal carcinoma: three alternatives to laser microdissection with preamplification.

Colorectal-carcinoma specimens are heterogeneous and include areas of nonmalignant mucosal and connective tissue. For those study designs in which laser microdissection and RNA preamplification are impracticable, the optimal yield of genuine cancer RNA is a key factor in gene-expression analysis. In this study we compared alternative methods of tissue purification. Three contiguous 0.5-cm(3) samples taken from an advanced primary adenocarcinoma of the sigmoid colon were processed immediately after surgery with the use of the following methods: (1) cryotomy after manual dissection (CMD), (2) microscopically assisted manual dissection (MAMD), and (3) tumor-cell isolation with the use of Ber-EP4 antibodies and Dynabeads (Dynal Biotech GmbH, Hamburg, Germany; technique abbreviated as DB). We generated gene-expression profiles with the use of GeneChip technology (Affymetrix, Santa Clara, Calif) and recorded preparation times, costs, and RNA quantity and quality. CMD took 60 minutes, MAMD 180 minutes, and DB 90 minutes to isolate 22, 8, and 23 microg of RNA, respectively. Expenses for materials amounted to 41, 23, and 91 US dollars for CMD, MAMD, and DB, respectively. The 3'/5' ratio, as determined with the GeneChips, for GAPDH/beta-actin was 1.01:1.03 for CMD, 1.13:1.28 for MAMD, 1.43:1.68 for DB, K-ras, APC, smad 2, transforming growth factor-beta, and p53 were marked as present in all cases, with the exception of APC, which was graded as marginal on DB. The correlation values of gene-expression profiles were 91% (CMD/DB), 93% (CMD/MAMD), and 97% (DB/MAMD). All 3 methods provided enough RNA, of sufficient quality, for gene-expression microarray analysis in colorectal carcinoma. Cross-methodologic analyses of array data should not be performed uncritically.