Resected Non-small Cell Lung Carcinoma Research Articles

A Comprehensive Analysis of p16 Expression, Gene Status, and Promoter Hypermethylation In Surgically Resected Non-small Cell Lung Carcinomas

The role of p16 is gaining importance in non-small cell lung cancer (NSCLC) because of epigenetic therapy options. Further insight into the significance of protein expression, gene status and promoter methylation is needed and has the potential to optimize existing treatment strategies. This population-based study analyzes p16 in 383 surgically resected non-small cell lung carcinomas brought into a standardized tissue microarray platform. Immunohistochemistry and fluorescence in situ hybridization were performed. For selected cases, p16 promoter hypermethylation was assessed by a pyrosequencing assay. Extensive clinical data and a postoperative follow-up period of 15 years enabled detailed correlations. Loss of p16 expression is a common event in NSCLC (232/365, 64%), especially in squamous cell carcinomas (97/115, 84%) in contrast to adenocarcinomas (93/186, 50%). Loss of p16 expression was associated with poorer survival time for the entire cohort and for certain subgroups including men, age younger than 65 years, smokers, early tumor stage, adenocarcinoma, and large-cell carcinoma. Promoter hypermethylation was absent for cases expressing p16 but was commonly observed when (heterozygous) p16 gene deletions were present and in cases negative for p16. Our comprehensive data would be compatible with a two-step process leading to loss of p16 expression in NSCLC. Hypermethylation of the promoter region may represent an early event, followed by heterozygous deletion of the p16 locus. Because of the possibility of detection of hypermethylated gene regions, these data may lead to the identification of specific patient subgroups more likely to benefit from upcoming demethylating treatment strategies.

Predictive clinical outcome of the intratumoral CD66b‐positive neutrophil‐to‐CD8‐positive T‐cell ratio in patients with resectable nonsmall cell lung cancer

The role of the interaction between tumor cells and inflammatory cells in nonsmall cell lung carcinoma (NSCLC) is unclear. In this study, the authors assessed the prognostic impact of intratumoral cluster of differentiation 66b (carcinoembryonic antigen-related cell adhesion molecule 8 [CD66b])-positive neutrophils and of the intratumoral CD66b-positive neutrophil-to-cluster of differentiation 8 (cell surface antigen T8 [CD8])-positive lymphocytes (the CD66b-positive neutrophil-to-CD8-positive lymphocyte ratio [iNTR]) in patients with resectable NSCLC. Expression levels of CD66b and CD8 were evaluated by immunohistochemistry on tissue microarrays consisting of 632 NSCLC specimens from patients who underwent curative surgery. The relation between clinicopathologic variables and patient outcome was assessed. Intratumoral CD66b-positive neutrophils were elevated in 318 patients (50%). In univariate analysis, an increase in CD66b-positive cells was associated with a high cumulative incidence of relapse (CIR) (median CIR, 51 months for low CD66b-positive cell density; 36 months for high CD66b-positive cell density; P = .002) and trended toward worse overall survival (OS) (median OS, 57 months for low CD66b-positive cell density; 54 months for high CD66b-positive cell density; P = .088). The iNTR was elevated in 190 patients (30%). An increased iNTR was strongly associated with both a high CIR (median CIR: 43 months for an iNTR ≤1; 34 months for an iNTR >1; P < .0001) and poor OS (median OS: 60 months for an iNTR ≤1; 46 months for an iNTR >1; P < .0001). In multivariate analysis, independent prognostic factors for a higher CIR were high iNTR (hazard ratio [HR], 0.71; 95% confidence interval [CI], 0.56-0.90; P = .005) and tumor stage >I, (HR, 0.39; 95% CI, 0.30-0.52; P < .0001). Independent prognostic factors for worse OS were a high iNTR (HR, 0.70; 95% CI, 0.54-0.91; P = .007) and tumor stage >I (HR, 0.35; 95% CI, 0.26-0.47; P < .0001). The current results indicated that the iNTR is a novel, independent prognostic factor for a high rate of disease recurrence and poor OS in patients with resectable NSCLC.

Prognostic Value of Epithelial Growth Factor Receptor, Vascular Endothelial Growth Factor, E-Cadherin, and p120 Catenin in Resected Non-Small Cell Lung Carcinoma

Introduction Several markers have been investigated to predict the prognosis of lung cancer. In the present study, the prognostic values of epithelial growth factor receptor (EGFR), vascular endothelial growth factor (VEGF), E-cadherin, and p120 catenin expression were investigated by immunohistochemistry in patients with a surgically resected non-small cell lung carcinoma (NSCLC). Patients and method EGFR, VEGF, E-cadherin, and p120 catenin expression were prospectively determined in resected specimens from patients with NSCLC who had undergone surgery between 2003 and 2007. Patients’ and disease-related general characteristics and survival rate were recorded. Results One hundred seventeen patients with a mean age of 61.3 years were included in the study. After a mean follow-up of 27.5 months, the median survival was determined to be 44.0 months and the 5-year survival was 46.2%. The 5-year survival in negative and positive staining groups were as follows; 32% and 66.7% for EGFR ( P=.02), 37.8%, and 50.7% for VEGF ( P=.5), 41% and 66% for E-cadherin ( P=.19), and 46% and 50% for p120 catenin ( P=.27). The differentiation, N status, stage, and EGFR staining were variables significantly affecting survival ( P=.001, .006, .03, and .02, respectively). In multivariate Cox analysis, the EGFR staining level and N status were variables those significantly affecting survival ( P=.021 and P=.010). Conclusions While negative staining of EGFR was related with poor survival, staining of VEGF, E-cadherin, and p120 catenin were not related with survival in patients with resected NSCLC.

Abstract 2252: ALDH7A1 expression is associated with recurrence in patients with resected non-small cell lung carcinoma

Abstract The aldehyde dehydrogenase (ALDH) enzymes are important for detoxification of endogenous and exogenous aldehydes. Expression of ALDH family members has recently been described as a potential marker for tumor-initiating cancer stem cells in a variety of human malignancies, including lung cancer. We were interested to determine whether expression of ALDH7A1, a member of the ALDH family, has prognostic significance in resected non-small cell lung carcinoma (NSCLC). Tumor specimens were obtained for 107 patients with completely resected stage I through stage III NSCLC from paraffin-embedded tissue microarrays and stained with an antibody specific for ALDH7A1. Staining patterns were graded by a pathologist based on the intensity of staining and the percentage of cells stained. A staining score index was calculated by multiplying intensity score by the percentage area with positive staining. An index of ≥100 was considered positive, while an index &amp;lt;100 was deemed negative, as determined by visually determining a clear positive stain supported by histograms of the range of scores. Staining index was subsequently correlated with clinical data. Positive ALDH7A1 staining was identified in 46 patients, and negative staining was noted in 48 patients, with 13 tumor sections missing. Multivariate analysis demonstrated that NSCLC patients with positive ALDH7A1 expression had decreased overall survival relative to ALDH7A1-negative tumors, although this did not reach significance (hazard ratio 1.42, 95% confidence interval 0.84 to 2.389; p=0.192). However, patients with ALDH7A1-expressing NSCLC tumors had a significantly higher incidence of lung cancer recurrence compared to patients with ALDH7A1-negative tumors (hazard ratio 6.24, 95% confidence interval 2.06 to 18.86; p=0.001). These data indicate that ALDH7A1 staining is present in a substantial number of NSCLC tumors and may be a biomarker predictive for increased incidence of cancer recurrence in patients with surgically resected NSCLC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2252. doi:10.1158/1538-7445.AM2011-2252

Abstract 1610: Predicting NSCLC outcomes based on tumor growth in SCID mice

Abstract Background. Xenograft models of non-small cell lung carcinoma (NSCLC) in immunodeficient mice have played an important role in the study of new chemotherapeutic regimens, including targeted agents. We hypothesize that the success of human tumor implantation in severe combined immune-deficient (SCID) mice will correlate with the observed clinical outcomes in patients with resected NSCLC. Such growth in mice may reflect the aggressiveness of human tumors. Methods. 196 lung cancer specimens from 193 patients who underwent surgical resection of NSCLC from June 1995 to June 2010 were implanted subcutaneously in (SCID) mice. Mice were observed for a period of six months to assess the success of tumor implantation. This was determined by the growth of tumors from 2-3 mm to 10 mm in diameter. Overall and disease free survival in patients were reviewed and were correlated with tumor growth in SCID mice. Results. Median overall and disease free survival in patients with successful tumor implantation in SCID mice was 16.3 months and 4.2 months vs. 38.6 and 20.6 months for patients whose tumor did not grow (logrank p = 0.0039 and 0.0024 respectively.) Half of all patients whose tumor successfully grew in SCID mice recurred or died within twelve months. Conclusions. Success of tumor implantation in SCID correlates with worse clinical outcome in patients with NSCLC. This model can be expanded to evaluate the predicted response of specific tumors to novel chemotherapeutic or molecularly targeted agents. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1610. doi:10.1158/1538-7445.AM2011-1610

Prognostic gene-expression signature of carcinoma-associated fibroblasts in non-small cell lung cancer

The tumor microenvironment strongly influences cancer development, progression, and metastasis. The role of carcinoma-associated fibroblasts (CAFs) in these processes and their clinical impact has not been studied systematically in non-small cell lung carcinoma (NSCLC). We established primary cultures of CAFs and matched normal fibroblasts (NFs) from 15 resected NSCLC. We demonstrate that CAFs have greater ability than NFs to enhance the tumorigenicity of lung cancer cell lines. Microarray gene-expression analysis of the 15 matched CAF and NF cell lines identified 46 differentially expressed genes, encoding for proteins that are significantly enriched for extracellular proteins regulated by the TGF-β signaling pathway. We have identified a subset of 11 genes (13 probe sets) that formed a prognostic gene-expression signature, which was validated in multiple independent NSCLC microarray datasets. Functional annotation using protein-protein interaction analyses of these and published cancer stroma-associated gene-expression changes revealed prominent involvement of the focal adhesion and MAPK signaling pathways. Fourteen (30%) of the 46 genes also were differentially expressed in laser-capture-microdissected corresponding primary tumor stroma compared with the matched normal lung. Six of these 14 genes could be induced by TGF-β1 in NF. The results establish the prognostic impact of CAF-associated gene-expression changes in NSCLC patients.

Detection of circulating tumor cells as a prognostic factor in patients undergoing radical surgery for non‐small‐cell lung carcinoma: comparison of the efficacy of the CellSearch Assay™ and the isolation by size of epithelial tumor cell method

Comparison of the efficacy of different enrichment methods for detection of circulating tumor cells (CTCs) before radical surgery is lacking in non-small-cell lung carcinoma (NSCLC) patients. Detection and enumeration of CTCs in 210 consecutive patients undergoing radical surgery for NSCLC were evaluated with the CellSearch Assay™ (CS), using the CellSearch Epithelial Cell Kit, and by the isolation by size of epithelial tumor (ISET) method, using double immunolabeling with anti-cytokeratin and anti-vimentin antibodies. CTCs were detected in 144 of 210 (69%) patients using CS and/or ISET and in 104 of 210 (50%) and 82 of 210 (39%) patients using ISET and CS, respectively. Using ISET, 23 of 210 (11%) patients had vimentin-positive cells with cytological criteria of malignancy. Disease-free survival (DFS) was worse for patients with CTCs compared to patients without CTCs detected by CS alone (p < 0.0001; log rank = 30.59) or by ISET alone (p < 0.0001; log rank = 33.07). The presence of CTCs detected by both CS and ISET correlated even better with shorter DFS at a univariate (p < 0.0001; log rank = 42.15) and multivariate level (HR, 1.235; 95% CI, 1.056-1.482; p < 0.001). CS and ISET are complementary methods for detection of CTCs in preoperative radical surgery for NSCLC. CTC detection in resectable NSCLC patients using CS and/or ISET could be a prognostic biomarker of great interest and may open up new avenues into improved therapeutic strategies for lung carcinoma patients.

Preoperative Circulating Tumor Cell Detection Using the Isolation by Size of Epithelial Tumor Cell Method for Patients with Lung Cancer Is a New Prognostic Biomarker

Pathologic TNM staging is currently the best prognostic factor for non-small cell lung carcinoma (NSCLC). However, even in early-stage NSCLC, the recurrence rates after surgery range from 25% to 50%. The preoperative detection of circulating tumor cells (CTC) could be useful to tailor new therapeutic strategies in NSCLC. We assessed the presence of CTC in NSCLC patients undergoing surgery, using cytologic analyses, after their isolation by size of epithelial tumor cells (ISET method). The presence and the number of CTCs were considered and correlated with clinicopathologic parameters including patient follow-up. Of the 247 blood samples tested, 208 samples were from patients with resectable NSCLC and 39 from healthy subjects. The mean follow-up was 24 months. An image of detected cells with presumably nonhematologic features [initially defined as "circulating nonhematologic cells" (CNHC)] was recorded. The presence of CNHC was assessed blindly and independently by 10 cytopathologists, using cytologic criteria of malignancy on stained filters. The count of detected CNHCs was made for each filter. One hundred two of 208 (49%) patients showed CNHCs corresponding to CNHC with malignant cytopathologic features in 76 of 208 (36%) cases. CNHCs were not detected in the control group. A level of 50 or more CNHCs corresponding to the third quartile was associated with shorter overall and disease-free-survival, independently of disease staging, and with a high risk of recurrence and death in early-stage I + II-resectable NSCLC. A high percentage of NSCLC patients show preoperative detection of CNHC by the ISET method. The presence and level of 50 or more CNHCs are associated with worse survival of patients with resectable NSCLC.

Prognostic Significance of ERCC1 Expression in Resected Non Small Cell Lung Carcinoma

The effects of DNA repair pathway on survival were assessed by measuring the tumoral excision repair cross complementation 1 (ERCC1) expression in patients with resected non-small cell lung carcinoma (NSCLC). We aimed to determine the prognostic and predictive significance of ERCC1 in patients with completely resected NSCLC. Immunohistochemistry (IHC) was used to assess the expression of ERCC1 in resected lung tumor samples obtained from 98 patients untreated without pre- or post-operative chemotherapy and/or radiotherapy. The median H score was used as a cut-off for ERCC1 IHC. Univariate and multivariate analyses were performed for factors influencing patient survival. The 5-year survival rates of patients for ERCC1 positive expression and ERCC1 negative expression were 76% and 49%, respectively; this difference was statistically significant (p = 0.004). Subsequent multivariate analysis suggested that ERCC1 expression (adjusted hazard ratio for death, 0.38; 95% CI, 0.18 to 0.78; p = 0.008) and pathological stage (2.2; 95% CI, 1.09 to 4.5; p = 0.027) were both independent prognostic factors. The level of ERCC1 expression in tumors a strong predictor of survival in resected NSCLC patients untreated without pre- or post-operative chemotherapy and/or radiotherapy.

MicroRNA Expression and Clinical Outcomes in Patients Treated with Adjuvant Chemotherapy after Complete Resection of Non–Small Cell Lung Carcinoma

This study determined whether expression levels of a panel of biologically relevant microRNAs can be used as prognostic or predictive biomarkers in patients who participated in the International Adjuvant Lung Cancer Trial (IALT), the largest randomized study conducted to date of adjuvant chemotherapy in patients with radically resected non-small cell lung carcinoma (NSCLC). Expression of miR-21, miR-29b, miR-34a/b/c, miR-155, and let-7a was determined by quantitative real-time PCR in formalin-fixed paraffin-embedded tumor specimens from 639 IALT patients. The prognostic and predictive values of microRNA expression for survival were studied using a Cox model, which included every factor used in the stratified randomization, clinicopathologic prognostic factors, and other factors statistically related to microRNA expression. Investigation of the expression pattern of microRNAs in situ was performed. We also analyzed the association of TP53 mutation status and miR-34a/b/c expression, epidermal growth factor receptor and KRAS mutation status, and miR-21 and Let-7a expression. Finally, the association of p16 and miR-29b expression was assessed. Overall, no significant association was found between any of the tested microRNAs and survival, with the exception of miR-21 for which a deleterious prognostic effect of lowered expression was suggested. Otherwise, no single or combinatorial microRNA expression profile predicted response to adjuvant cisplatin-based chemotherapy. Together, our results indicate that the microRNA expression patterns examined were neither predictive nor prognostic in a large patient cohort with radically resected NSCLC, randomized to receive adjuvant cisplatin-based chemotherapy versus follow-up only.

Primary Tumor Xenografts of Human Lung Adeno and Squamous Cell Carcinoma Express Distinct Proteomic Signatures

Nonsmall cell lung carcinoma (NSCLC) accounts for 80% of lung cancers. The most prevalent subtypes of NSCLC are adenocarcinoma (ADC) and squamous cell carcinoma (SCC), which combined account for approximately 90%. Ten resected NSCLC patient tumors (5 ADC and 5 SCC) were directly introduced into severely immune deficient (NOD-SCID) mice, and the resulting xenograft tumors were analyzed by standard histology and immunohistochemistry (IHC) and by proteomics profiling. Mass spectrometry (MS) methods involving 1- and 2-dimensional LC-MS/MS, and multiplexed selective reaction monitoring (SRM, or MRM), were applied to identify and quantify the xenograft proteomes. Hierarchical clustering of protein profiles distinguished between the ADC and SCC subtypes. The differential expression of 178 proteins, including a comprehensive panel of intermediate filament keratin proteins, was found to constitute a distinctive proteomic signature associated with the NSCLC subtypes. Epidermal growth factor receptor (EGFR) was expressed in ADC and SCC xenografts, and EGFR network activation was assessed by phosphotyrosine profiling by Western blot analysis and SRM measurement of EGFR levels, and mutation analysis. A multiplexed SRM/MRM method provided relative quantification of several keratin proteins, EGFR and plakophilin-1 in single LC-MS/MS runs. The protein quantifications by SRM and MS/MS spectral counting were associated with superior dynamic range and reproducibility but were otherwise consistent with orthogonal methods including IHC and Western immuno blotting. These findings illustrate the potential to develop a comprehensive MS-based platform in oncologic pathology for better classification and potentially treatment of NSCLC patients.

Abstract 374: VEGFR2/KDR polymorphisms and gene copy gain with outcome in non-small cell lung cancer

Abstract Background. The VEGF receptor 2 (VEGFR2) is the predominant mediator of VEGF-stimulated endothelial cell function. Recently, VEGFR2/KDR copy number gain and mutation, and VEGF copy number gain, have been described in lung adenocarcinoma tumor specimens. To better characterize the molecular changes of these two genes in non-small cell lung carcinoma (NSCLC), we investigated their abnormalities in NSCLC tumor tissue specimens and correlated with patients clinico-pathological characteristics. Methods. We extracted DNA from microdissected tissue obtained from 200 surgically resected NSCLCs. KDR single nucleotide polymorphism (SNP) 889G/A (rs2305948), SNP 1416A/T (rs1870377) and SNP −37A/G (rs2219471) were genotyped by PCR-based sequencing. KDR and VEGF copy number were examined by quantitative (q)-PCR. Protein expressions of VEGF and VEGFR2, and CD34 for microvascular density (MVD) analysis were studied by immunohistochemistry. SNP genotypes, genes copy number and protein expression were correlated with NSCLC clinico-pathological features, including overall survival (OS) and recurrence-free survival (RFS). Results. KDR 1416 AT/TT genotypes had significantly improved OS (HR = 0.56, 95% CI 0.33 to 0.96, P = 0.035) compared with KDR 1416 AA in all NSCLC patients in the multivariate analysis with adjustment of histology and tumor stage. In lung adenocarcinomas, KDR 1416 AT/TT genotypes and KDR −37AG/GG were associated with a favorable OS (HR = 0.43, 95% CI 0.20 to 0.92, P = 0.029; HR = 0.47, 95% CI 0.23 to 0.96, P = 0.039, respectively). Strikingly, KDR −37 AG/GG genotypes predicted a superior OS compared with KDR −37 AA genotype in the NSCLC patients treated with adjuvant therapy (HR = 2.45, 95% CI 1.06 to 5.66, P = 0.036). No genotype in KDR SNPs was associated with RFS in NSCLC patients. Gene copy gain of KDR and VEGF were detected respectively in 34/91 (37.4%) and 2/91 (2.2%) NSCLC tumors. Gene copy gain of KDR was associated with a poor overall survival in NSCLC patients (HR= 2.96, 95% CI 1.41 to 6.24, P = 0.004). Furthermore, tumors with gene copy gain of KDR showed significantly higher cytoplasmic (P = 0.013) and membrane (P = 0.009) VEGFR2 protein expression, lower cytoplasmic VEGF expression (P = 0.044), and higher MVD (P = 0.018) and larger vessel areas (P = 0.033) compared with tumors lacking KDR gene copy gain. Conclusion. Our findings suggest an association between KDR SNP genotypes and survival in NSCLC patients receiving adjuvant therapy. KDR copy number gain was frequently identified in NSCLC and was associated with worse survival in NSCLC patients. (Supported by grant US DoD W81XWH-07-1-0306). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 374.

Subtyping of Undifferentiated Non-small Cell Carcinomas in Bronchial Biopsy Specimens

The emergence of treatments for non-small cell lung carcinoma (NSCLC) with differential efficacy and toxicity between subtypes has highlighted the importance of specific pathologic NSCLC subtyping. Most NSCLCs are inoperable, and pathologic diagnosis is made only on small tissue samples that are prone to diagnostic inaccuracy. In a substantial proportion of cases, standard morphology cannot specifically subtype the tumor, necessitating a diagnosis of NSCLC-not otherwise specified (NOS). Histochemical staining for mucin and immunohistochemical (IHC) identification of NSCLC subtype-associated markers could help predict the final subtype of resected NSCLCs diagnosed as NSCLC-NOS on preoperative bronchial biopsy samples. Paraffin sections of 44 bronchial biopsy samples diagnosed as NSCLC-NOS were stained for mucin (Alcian blue/periodic acid Schiff) and thyroid transcription factor 1 by IHC-(markers of adenocarcinoma), and for S100A7, cytokeratin 5/6, high molecular weight cytokeratins, and p63 proteins-markers of squamous cell carcinoma. A predictive staining panel was derived from statistical analysis after comparing staining profiles with the final postsurgical NSCLC subtype. This panel was prospectively applied to 82 small biopsy samples containing NSCLC. True NSCLC subtype of undifferentiated NSCLC samples was best predicted using Alcian blue/periodic acid Schiff plus p63 and thyroid transcription factor 1 IHC, allowing specific subtyping in 73% of NSCLC-NOS cases with 86% accuracy. When applied prospectively, this staining panel showed 100% concordance with specific NSCLC morphologic subtyping in small biopsies. This approach can facilitate treatment selection by accurately predicting the subtype in undifferentiated NSCLC biopsies, reducing to 7% the proportion of cases without a definite or probable histologic subtype.

Role of Loss of O6-Methylguanine DNA Methyltransferase (MGMT) Expression in Non-Small Cell Lung Carcinomas (NSCLCs): with Reference to the Relationship with p53 Overexpression

Functional inactivation of the O⁶-methylguanine-DNA methyltransferase (MGMT) gene has been demonstrated as loss of MGMT protein and suggested that it plays an important role in primary human neoplasia, including lung cancer. It has also been reported to be associated with the G : C-->A : T transition mutation in the p53 gene of lung cancer. The aims of this study were to investigate the role of MGMT expression loss and its prognostic significance in non-small cell lung carcinomas (NSCLCs), and its correlation with p53 overexpression as well as influence on patient survival. 112 surgically resected NSCLC specimens were reviewed by medical records for their clinicopathologic variables. Their tissue microarray blocks were immunostained with anti-human MGMT and p53 primary antibodies. Correlation between MGMT loss and the clinicopathologic prognostic factors, including p53 overexpression and the single or combined actions of MGMT loss and p53 overexpression on patient survival were statistically analyzed by SPSS15.0. Reduced or absent MGMT expression was found in 48 of 112 NSCLCs (43%), and significantly associated with nodal metastasis and squamous or undifferentiated cell types. Loss of MGMT expression was correlated with p53 overexpression in adenocarcinomas, but not in overall NSCLCs. Its solitary or combined actions with p53 overexpression did not have influence on patient survival. Loss of MGMT expression is a relatively common event in NSCLCs and significantly associated with nodal metastasis and p53 overexpression, suggesting that it may play a major role in pulmonary carcinogenesis, and also in disease progression of NSCLCs.

The variation of prognostic significance of Maximum Standardized Uptake Value of [18F]-fluoro-2-deoxy-glucose positron emission tomography in different histological subtypes and pathological stages of surgically resected Non-Small Cell Lung Carcinoma

Even if the prognostic role of SUVmax of 18-FDG-PET has been largely investigated, many issues regarding its relationship with pathologic staging and histological subtypes still remain controversial. This retrospective study investigated the prognostic significance of SUVmax in 119 completely resected, pathologically proven NSCLC. The SUVmax values resulted significantly related to histological subtypes ( p < 0.001), histological grading ( p < 0.001), and pathologic stage ( p < 0.001). The optimal cut-off value of SUVmax to predict prognosis in the whole series was 6.7 ( p = 0.029). 2-Year disease-specific survival (DSS) was 91% for SUVmax ≤6.7 and 55% for SUVmax >6.7 ( p < 0.001). SUVmax still remain a significant predictor of survival in Stage IB (2-year DSS of 100% for SUVmax ≤6.7; 51% for SUVmax >6.7, p = 0.016). The optimal cut-off values of SUVmax to predict prognosis were 5 for adenocarcinoma ( p = 0.027) and 10.7 for other non-adenocarcinoma NSCLC subtypes ( p = 0.010). These histologic-specific cut-offs resulted significantly related to survival when stratified for stage: 2-year DSS for Stage IB adenocarcinoma were 100% for SUV ≤ 5 and 40% for SUVmax >5 ( p = 0.051); 2-year DSS for Stage IB non-adenocarcinoma were 83% for SUVmax ≤10.7 and 26% for SUVmax >10.7 ( p = 0.018). Adenocarcinomas showed significantly lower survival results respect to other NSCLC for intermediate SUVmax level (range 5.5–11.3) ( p = 0.021). High SUVmax resulted an independent negative prognostic factor at multivariate analysis (HR of 15.7, 95% CI of 2.50–98.44, p = 0.003). In conclusion, SUVmax represents a significant prognostic factor in surgically resected NSCLC but a great variability between different histological subtypes, even when adjusted for stage, is present and could be considered when planning future trials on prognostic role of FDG uptake.

The Prognostic Impact of Main Bronchial Lymph Node Involvement in Non-Small Cell Lung Carcinoma: Suggestions for a Modification of the Staging System

The therapeutic strategies for non-small cell lung carcinoma (NSCLC) with N1 and N2 disease differ remarkably. Debate exists about the definition of the borderline between N1 and N2 stations. This study evaluated the prognostic effect of N1 disease, especially focused on the significance of the main bronchial node (No. 10) vs N2 disease. The records of 1601 patients who underwent complete pulmonary resection for NSCLC were reviewed to examine the clinical features of lymph nodal involvement. There were 1086 patients (67.8%) with pN0 disease, 202 (12.6%) with pN1, and 274 (17.1%) with pN2 disease; overall 5-year survival rates were 74.7%, 56.1% and 28.9%, respectively (p < 0.001). Overall 5-year survival rates were 60.2% in hilar N1 and 49.6% in intralobar N1. Overall 5-year survival rates were 58.6% in N1 without node 10 and 35.1% in N1 with node 10. A significant difference was observed between N0 and N1 without node 10 (p < 0.001), and N1 without node 10 and N1 with node 10 (p = 0.033); however, the difference between N1 with node 10 and N2 was not significant. The status of node 10 involvement was an independent prognostic factor of pN1 patients, as well as age and gender. Patients with node 10-positive N1 disease have an unfavorable prognosis, and the disease behaves like N2 disease. The definition of clear borderline between N1 and N2 is mandatory to achieve a uniform classification map. This study offers further information for clinical and therapeutic purposes.

The prognostic value of intraepithelial and stromal innate immune system cells in non‐small cell lung carcinoma

The major value of prognostic markers in potentially curable non-small cell lung carcinoma (NSCLC) should be to guide therapy after surgical resection. The prognostic significance of tumour-infiltrating macrophages, their growth factor, macrophage colony-stimulating factor (M-CSF), and its receptor, colony-stimulating factor-1 receptor (CSF-1R), as well as natural killer cells and dendritic cells, is controversial. The aim of this study was to elucidate the prognostic significance of these markers in the epithelial and stromal compartments of NSCLC. Tissue microarrays from 335 resected NSCLC, stage I-IIIA were constructed from duplicate cores of epithelial and stromal areas. Immunohistochemistry was used to evaluate epithelial and stromal areas for CD68, M-CSF, CSF-1R, CD56 and CD1a. On univariate analysis, increasing numbers of stromal CD1a+ (P = 0.011) and CD56+ cells (P = 0.014) correlated significantly with improved disease-specific survival (DSS). On multivariate analysis, stromal CD56+ cells were an independent prognostic factor for DSS (hazard ratio = 2.3, confidence interval = 1.1, 5.0, P = 0.031). High density of stromal CD56+ cells is an independent factor associated with improved prognosis in resected NSCLC, suggesting that these cells mediate an antitumour immune response in the tumour stroma.

Clinical relevance of neuroendocrine differentiation in non-small cell lung cancer assessed by immunohistochemistry: a retrospective study on 405 surgically resected cases

Neuroendocrine differentiation in non-small cell lung cancer is a common feature, which has caused contradictory conclusions concerning survival estimates and responsiveness to therapy. Aiming to clarify this conflict, we analyzed neuroendocrine differentiation by immunohistochemistry in 405 surgically resected non-small cell lung carcinomas using standardized tissue microarray platform and the currently recommended antibody panel consisting of chromogranin-A, synaptophysin, and neural-cell adhesion molecule. Diagnostic criteria provided by the World Health Organization were applied. Histological subtypes were primarily reclassified according to current guidelines, assisted by auxiliary immunohistochemistry. Extensive clinical data was acquired, enabling detailed clinicopathological correlation. Importantly, neuroendocrine differentiation assessed by immunohistochemistry showed no significant relation to overall survival estimates, which remained unaffected by histological subtype, neuroendocrine marker type, adjuvant therapy, and recurring disease. The only exception was a small group consisting of three large cell carcinomas, each expressing all three neuroendocrine markers and demonstrating decreased survival. In conclusion, additional immunohistochemical detection of neuroendocrine differentiation in non-small cell lung cancer is presently not of prognostic importance and does not justify a distinct consideration.

Prognostic Impact of Fibroblast Growth Factor 2 in Non-small Cell Lung Cancer: Coexpression with VEGFR-3 and PDGF-B Predicts Poor Survival

Fibroblast growth factor 2 (FGF2; basic fibroblast growth factor, b-FGF) and its main receptor FGFR-1 are important in both hemangiogenesis and lymphangiogenesis. Murine studies have indicated a close interplay between both FGF2 and platelet-derived growth factor-B (PDGF-B) as well as FGF2 and vascular endothelial growth factor-3 (VEGFR-3). This study investigates the prognostic impact of FGF2 and FGFR-1 in tumor cells and tumor stroma of resected non-small cell lung carcinomas (NSCLC) and explores the importance of their coexpression with VEGFR-3 or PDGF-B. Tumor tissue samples from 335 resected patients with stage I to IIIA NSCLC were obtained and tissue microarrays were constructed from duplicate cores of tumor cells and tumor-related stroma from each specimen. Immunohistochemistry was used to evaluate the expression of the molecular markers FGF2, FGFR-1, VEGFR-3, and PDGF-B. In univariate analyses, high tumor cell FGF2 expression (p = 0.015) was a negative prognostic indicator for disease-specific survival. In tumor stroma, high FGF2 (p = 0.024) expression correlated with good prognosis. In multivariate analyses, high expression of FGF2 in tumor cells (p = 0.038) was an independent negative prognostic factor whereas increased FGF2 in stroma (p = 0.015) was a positive prognosticator. Tumor cell coexpressions of FGF2/VEGFR-3 (p < 0.001) and FGFR-1/PDGF-B (p = 0.002) were significant indicators of poor prognosis. Expression of FGF2 in tumor cells is an independent negative prognostic factor, and the coexpressions of FGF2/VEGFR-3 and FGFR-1/PDGF-B are strongly associated with poor survival in NSCLC patients.

Analysis of Survival in 400 Surgically Resected Non-small Cell Lung Carcinomas: Towards a Redefinition of the T Factor

The tumor, node, metastasis (TNM) system has been recognized internationally as the standard for staging disease extension, but despite the improvements of the 1997/2002 international staging system, there may be marked differences in postoperative 5-year survival rates within each stage. There is controversy about the impact of tumor size itself as a variable unrelated to stage.The objective of this study was to analyze the influence of tumor size on the survival in patients with surgically resected non-small cell lung carcinoma (NSCLC). Between August 1985 and January 2006, 400 patients underwent pulmonary resection with a curative intention for non-small cell lung carcinoma. Patients were excluded if they had received neoadjuvant chemotherapy. The clinicopathological records of each patient were examined for prognostic factors such as age, sex, right or left side cancer, histology, tumor location, tumor size, clinical nodal stage number, and distribution of metastatic nodes. Operative mortality was 2.2% for lobectomy and 18% for pneumonectomy (p < 0.05). Adenocarcinoma was the most common type (n = 245, 61.2%). Surgery was considered a complete resection in 341 patients (85.2%). When only patients without neoplastic hilar or mediastinal metastases (pN0) were included, the difference in survival was significantly different in terms of tumor size (log rank 28.46, p < 0.0001). Univariate analysis for the group of pN0 patients showed survival was not significantly affected by age, sex, side, or adenocarcinoma histology. In the multivariate analysis, tumor size and the T factor were found to have maintained its independent prognostic effects on overall survival. Among patients with pN0 tumors smaller that 15 mm in diameter, 5-year survival was 95% whereas patients with tumors bigger than 16 mm in diameter had a 5-year survival of 65% (p < 0.0001). In conclusion, our data suggest that tumors over 15 mm are associated with shorter 5-year survival in all TNM stages. Current TNM categories are not sufficiently discriminatory and the T factor requires to be reevaluated in further revisions of the TNM classification.