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Introduction: Programmed cell death ligand-1 (PD-L1) expression is a predictive biomarker for the efficacy of anti-programmed cell death receptor-1/PD-L1 antibodies in advanced non-small cell lung cancer (NSCLC). Although several assays have been approved for evaluating PD-L1 expression status, inter-assay discordance has been observed between some assays. The clinical significance of these discrepancies is still unclear., Methods: We retrospectively reviewed treatment-naïve NSCLC patients whose PD-L1 expression was evaluated using both 22C3 and SP142 assays. Among those, efficacy analysis was performed for patients with PD-L1 tumor proportion score (TPS) ≥ 50 % (22C3), who had received first-line pembrolizumab monotherapy. Additionally, transcriptome analysis was conducted in the available tumors with TPS ≥ 50 % to investigate the distinct immune profiles that accompany inter-assay discordance., Results: In total, 611 patients were eligible. Among 198 patients with TPS ≥ 50 %, 91 (46 %) had tumor cell score ≤ 1 (SP142, i.e., inter-assay discrepancy). In the 52 patients who received first-line pembrolizumab monotherapy, treatment efficacy was significantly lower in patients with the discrepancy than that in those without (objective response rate: 18 % vs. 83 %, p < 0.001; median progression-free survival [months]: 3.2 vs. 8.3, p < 0.001). Transcriptome analysis revealed significantly more CD274 splice variants with aberrant 3'-terminal sequences in tumors with the inter-assay discrepancy than in those without., Conclusion: The inter-assay discrepancy in the PD-L1 status of tumor cells between the 22C3 and SP142 assays, reflecting an imbalance in the CD274 splice variants, could be a biomarker for primary resistance against pembrolizumab monotherapy in high PD-L1-expressing NSCLCs., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Tight junction (TJ) protein cingulin (CGN) and transcription factor forkhead box protein O1 (FOXO1) contribute to the development of various cancers. Histone deacetylase (HDAC) inhibitors have a potential therapeutic role for some cancers. HDAC inhibitors affect the expression of both CGN and FOXO1. However, the roles and regulatory mechanisms of CGN and FOXO1 are unknown in non-small cell lung cancer (NSCLC) and normal human lung epithelial (HLE) cells. In the present study, to investigate the effects of CGN and FOXO1 on the malignancy of NSCLC, we used A549 cells as human lung adenocarcinoma and primary human lung epithelial (HLE) cells as normal lung tissues and performed the knockdown of CGN and FOXO1 by siRNAs. Furthermore, to investigate the detailed mechanisms in the antitumor effects of HDAC inhibitors for NSCLC via CGN and FOXO1, A549 cells and HLE cells were treated with the HDAC inhibitors trichostatin A (TSA) and Quisinostat (JNJ-2648158). In A549 cells, the knockdown of CGN increased bicellular TJ protein claudin-2 (CLDN-2) via mitogen-activated protein kinase/adenosine monophosphate-activated protein kinase (MAPK/AMPK) pathways and induced cell migration, while the knockdown of FOXO1 increased claudin-4 (CLDN-4), decreased CGN, and induced cell proliferation. The knockdown of CGN and FOXO1 induced cell metabolism in A549 cells. TSA and Quisinostat increased CGN and tricellular TJ protein angulin-1/lipolysis-stimulated lipoprotein receptor (LSR) in A549. In normal HLE cells, the knockdown of CGN and FOXO1 increased CLDN-4, while HDAC inhibitors increased CGN and CLDN-4. In conclusion, the knockdown of CGN via FOXO1 contributes to the malignancy of NSCLC. Both HDAC inhibitors, TSA and Quisinostat, may have potential for use in therapy for lung adenocarcinoma via changes in the expression of CGN and FOXO1.

Background: The association between duration of smoking abstinence before non-small-cell lung cancer (NSCLC) diagnosis and subsequent survival can influence public health messaging delivered in lung-cancer screening. We aimed to assess whether the duration of smoking abstinence before diagnosis of NSCLC is associated with improved survival., Methods: In this retrospective, pooled analysis of cohort studies, we used 26 cohorts participating in Clinical Outcomes Studies of the International Lung Cancer Consortium (COS-ILCCO) at 23 hospitals. 16 (62%) were from North America, six (23%) were from Europe, three (12%) were from Asia, and one (4%) was from South America. Patients enrolled were diagnosed between June 1, 1983, and Dec 31, 2019. Eligible patients had smoking data before NSCLC diagnosis, epidemiological data at diagnosis (obtained largely from patient questionnaires), and clinical information (retrieved from medical records). Kaplan-Meier curves and multivariable Cox models (ie, adjusted hazard ratios [aHRs]) were generated with individual, harmonised patient data from the consortium database. We estimated overall survival for all causes, measured in years from diagnosis date until the date of the last follow-up or death due to any cause and NSCLC-specific survival., Findings: Of 42 087 patients with NSCLC in the COS-ILCCO database, 21 893 (52·0%) of whom were male and 20 194 (48·0%) of whom were female, we excluded 4474 (10·6%) with missing data. Compared with current smokers (15 036 [40·0%] of 37 613), patients with 1-3 years of smoking abstinence before NSCLC diagnosis (2890 [7·7%]) had an overall survival aHR of 0·92 (95% CI 0·87-0·97), patients with 3-5 years of smoking abstinence (1114 [3·0%]) had an overall survival aHR of 0·90 (0·83-0·97), and patients with more than 5 years of smoking abstinence (10 841 [28·8%]) had an overall survival aHR of 0·90 (0·87-0·93). Improved NSCLC-specific survival was observed in 4301 (44%) of 9727 patients who had quit cigarette smoking and was significant at abstinence durations of more than 5 years (aHR 0·87, 95% CI 0·81-0·93). Results were consistent across age, sex, histology, and disease-stage distributions., Interpretation: In this large, pooled analysis of cohort studies across Asia, Europe, North America, and South America, overall survival was improved in patients with NSCLC whose duration of smoking abstinence before diagnosis was as short as 1 year. These findings suggest that quitting smoking can improve overall survival, even if NSCLC is diagnosed at a later lung-cancer screening visit. These findings also support the implementation of public health smoking cessation strategies at any time., Funding: The Alan B Brown Chair, The Posluns Family Fund, The Lusi Wong Fund, and the Princess Margaret Cancer Foundation., Competing Interests: Declaration of interests We declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY-NC-ND 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Drug resistance limits the efficacy of chemotherapy and targeted cancer treatments, calling for the identification of druggable targets to overcome it. Here we show that the mitochondria-shaping protein Opa1 participates in resistance against the tyrosine kinase inhibitor gefitinib in a lung adenocarcinoma cell line. Respiratory profiling revealed that oxidative metabolism was increased in this gefitinib-resistant lung cancer cell line. Accordingly, resistant cells depended on mitochondrial ATP generation, and their mitochondria were elongated with narrower cristae. In the resistant cells, levels of Opa1 were increased and its genetic or pharmacological inhibition reverted the mitochondrial morphology changes and sensitized them to gefitinib-induced cytochrome c release and apoptosis. In vivo, the size of gefitinib-resistant lung orthotopic tumors was reduced when gefitinib was combined with the specific Opa1 inhibitor MYLS22. The combo gefitinib-MYLS22 treatment increased tumor apoptosis and reduced its proliferation. Thus, the mitochondrial protein Opa1 participates in gefitinib resistance and can be targeted to overcome it., (© 2023. The Author(s).)

Liquid biopsy can identify gene alterations that are associated with resistance to fusion gene-targeted treatments. In this study, we present three cases of advanced non-small cell lung cancer (NSCLC) harboring gene fusions; cell-free DNA (cfDNA) was used to assess the resistance mutations. A patient with MET amplification underwent RET-fusion NSCLC treatment with selpercatinib. A patient with ROS1 G2032R underwent ROS1-fusion NSCLC treatment with crizotinib. A patient who underwent ROS1-fusion NSCLC treatment with crizotinib harbored no somatic mutations. This case series shows that cfDNA analysis can identify potentially actionable genomic alterations, after disease progression, in targeted therapy for fusion genes. TRIAL REGISTRATION: The study was registered in the UMIN Clinical Trial Registry (UMIN 000017581)., Competing Interests: Conflict of Interest Yoshitaka Seki has no conflict of interest. Tatsuya Yoshida received honoraria for lecture fees from AstraZeneca, Chugai, Ono, Eli Lilly, Takeda, and research grants from AMGEN, AstraZeneca, Takeda, Daiichi-Sankyo, Ono, MSD, AbbVie, Novartis, Chugai, Merck Biopharma, Blueprint, and BMS. Takashi Kohno received research grants from Sysmex. Ken Masuda received honoraria for lecture fees from Ono, AstraZeneca, Chugai, BMS, and Healios. Yusuke Okuma received honoraria for lecture fees from AstraZeneca, Eli Lilly, Chugai, and research grants from AbbVie, and Chugai. Yasushi Goto received research grants from AstraZeneca, Chugai and Guardant Health. Hidehito Horinouchi received honoraria for lecture fees from AstraZeneca, MSD, Eli Lilly, Ono, BMS, Chugai, Roche, and Kyowa-Kirin, and research grants from MSD, AbbVie, AstraZeneca, BMS, Ono, Merck Biopharma, Daiichi-Sankyo, Janssen, Genomic Health, Chugai, Roche, and Novartis. Noboru Yamamoto received honoraria for lecture fees from Chugai, and research grant from Astellas, AstraZeneca, Chugai, Eisai, Taiho, BMS, Pfizer, Novartis, Eli Lilly, AbbVie, Daiichi-Sankyo, Bayer, Boehringer Ingelheim, Kyowa-Kirin, Takeda, ONO, Janssen Pharma, MSD, MERCK, GSK, Sumitomo Dainippon Pharma, Chiome Bioscience, Otsuka, Carna Biosciences, Genmab, and Shionogi. Kazuyoshi Kuwano has no conflict of interest. Yuichiro Ohe received honoraria for lecture fees from AstraZeneca, Chugai, and research grants from EPS Holdings, MSD, AstraZeneca, KISSEI PHARMACEUTICAL, BMS, Janssen Pharmaceutical, Takeda, IQVIA Services Japan, CMIC, Eli Lilly, and Medpace Japan., (Copyright © 2022 The Japanese Respiratory Society. Published by Elsevier B.V. All rights reserved.)

Chromosomal backgrounds of cancerous mutations still remain elusive. Here, we conduct the phasing analysis of non-small cell lung cancer specimens of 20 Japanese patients. By the combinatory use of short and long read sequencing data, we obtain long phased blocks of 834 kb in N50 length with >99% concordance rate. By analyzing the obtained phasing information, we reveal that several cancer genomes harbor regions in which mutations are unevenly distributed to either of two haplotypes. Large-scale chromosomal rearrangement events, which resemble chromothripsis events but have smaller scales, occur on only one chromosome, and these events account for the observed biased distributions. Interestingly, the events are characteristic of EGFR mutation-positive lung adenocarcinomas. Further integration of long read epigenomic and transcriptomic data reveal that haploid chromosomes are not always at equivalent transcriptomic/epigenomic conditions. Distinct chromosomal backgrounds are responsible for later cancerous aberrations in a haplotype-specific manner., (© 2022. The Author(s).)

Background: Somatic EGFR mutations define a subset of non-small cell lung cancers (NSCLC) that have clinical impact on NSCLC risk and outcome. However, EGFR-mutation-status is often missing in epidemiologic datasets. We developed and tested pragmatic approaches to account for EGFR-mutation-status based on variables commonly included in epidemiologic datasets and evaluated the clinical utility of these approaches., Methods: Through analysis of the International Lung Cancer Consortium (ILCCO) epidemiologic datasets, we developed a regression model for EGFR-status; we then applied a clinical-restriction approach using the optimal cut-point, and a second epidemiologic, multiple imputation approach to ILCCO survival analyses that did and did not account for EGFR-status., Results: Of 35,356 ILCCO patients with NSCLC, EGFR-mutation-status was available in 4,231 patients. A model regressing known EGFR-mutation-status on clinical and demographic variables achieved a concordance index of 0.75 (95% CI, 0.74-0.77) in the training and 0.77 (95% CI, 0.74-0.79) in the testing dataset. At an optimal cut-point of probability-score = 0.335, sensitivity = 69% and specificity = 72.5% for determining EGFR-wildtype status. In both restriction-based and imputation-based regression analyses of the individual roles of BMI on overall survival of patients with NSCLC, similar results were observed between overall and EGFR-mutation-negative cohort analyses of patients of all ancestries. However, our approach identified some differences: EGFR-mutated Asian patients did not incur a survival benefit from being obese, as observed in EGFR-wildtype Asian patients., Conclusions: We introduce a pragmatic method to evaluate the potential impact of EGFR-status on epidemiological analyses of NSCLC., Impact: The proposed method is generalizable in the common occurrence in which EGFR-status data are missing., (©2022 American Association for Cancer Research.)

Comprehensive genomic profiling has been approved for use in patients with advanced solid tumours; however, it is only indicated in advanced solid tumour patients without available standard chemotherapeutic treatment or those who have completed standard treatments in Japan, and there are no available data on the clinical feasibility and utility of comprehensive genomic profiling in treatment-naive patients. This multicentre, single-arm, prospective study aims to evaluate the feasibility and utility of the OncoGuide NCC Oncopanel System in treatment-naive patients with six advanced major malignancies: non-small cell lung cancer, breast cancer, gastric cancer, colon cancer, pancreatic cancer and biliary tract cancer (NCCH1908). This study (study cohort) will be compared with the other prospective observational study (control cohort), which enrols patients not receiving comprehensive genomic profiling prior to initial systemic treatment. A total of 200 patients will be enrolled in the study over 21 months. This study has been registered in the UMIN Clinical Trials Registry (www.umin.ac.jp/ctr/) (UMIN000040743)., Clinical Trial Registration: This study, initiated in June 2020, has been registered in the UMIN Clinical Trials Registry (www.umin.ac.jp/ctr/) (registration number: UMIN000040743). We plan to enrol a total of 200 patients over a period of 21 months., (© The Author(s) 2021. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Epidermal growth factor receptor(EGFR)is a transmembrane receptor tyrosine kinase that plays an important role in regulating the growth and progression of cancer via signalling pathways. This review summarizes the differences in EGFR kinase structural changes, stability of active forms, and phosphorylation inhibitory effects of EGFR tyrosine kinase inhibitors between the exon 19 deletion mutation and the exon 21 L858R point mutation, which are frequently detected EGFR mutations in patients with non-small cell lung cancer.

Introduction: Programmed death-ligand 1 (PD-L1) expression is not a completely reliable predictive marker of the efficacy of anti-programmed cell death protein-1 (PD-1)/PD-L1 therapy in patients with advanced NSCLC. Immune-related tumor microenvironment (TME) is classified into four different types based on the tumor-infiltrating lymphocyte (TIL) status and PD-L1 expression., Methods: We retrospectively reviewed patients with advanced NSCLC treated with anti-PD-1/PD-L1 therapy between 2015 and 2019. We investigated the association between the efficacy of anti-PD-1/PD-L1 therapy, the types of TME based on PD-L1 (clone: 22C3) expression, the density of CD8-positive TILs assessed by immunohistochemistry, and mutational profiles by next-generation sequencing., Results: Overall, 228 patients were included in the analysis. The patients were classified into the following four groups: type I: PD-L1 High (tumor proportion score ≥ 50%)/TIL High (≥85/mm 2 ; n = 73); type II: PD-L1 Low (tumor proportion score < 50%)/TIL Low (<85/mm 2 ; n = 70); type III: PD-L1 High /TIL Low (n = 37); and type IV: PD-L1 Low /TIL High (n = 48). The objective response rate (ORR) and progression-free survival (PFS) of anti-PD-1/PD-L1 therapy clearly differed according to the different TME types (ORR and PFS; type I: 64%, 14.5 mo; type II: 12%, 2.1 mo; type III: 24%, 3.6 mo; type IV; 41%, 10.8 mo). In patients with PD-L1 High tumors, type I tumors had significantly better ORR and PFS than type III tumors (ORR: p < 0.001 and PFS: p < 0.001). The presence of TP53 and KRAS mutation was related to the density of CD8-positive TILs and PD-L1 expression, respectively., Conclusions: Differential types of TME, including PD-L1 expression and TIL status, could accurately predict the efficacy of anti-PD-1/PD-L1 therapy., (Copyright © 2021 International Association for the Study of Lung Cancer. Published by Elsevier Inc. All rights reserved.)

Histone deacetylase (HDAC) inhibitors have a potential therapeutic role for non-small cell lung cancer (NSCLC). However, more preclinical studies of HDAC inhibitors in NSCLC and normal lung epithelial cells are required to evaluate their antitumor activities and mechanisms. The bicellular tight junction molecule claudin-2 (CLDN-2) is highly expressed in lung adenocarcinoma tissues and increase the proliferation of adenocarcinoma cells. Downregulation of the tricellular tight junction molecule angulin-1/LSR induces malignancy via EGF-dependent CLDN-2 and TGF-β-dependent cellular metabolism in human lung adenocarcinoma cells. In the present study, to investigate the detailed mechanisms of the antitumor activities of HDAC inhibitors in lung adenocarcinoma, human lung adenocarcinoma A549 cells and normal lung epithelial cells were treated with the HDAC inibitors Trichostatin A (TSA) and Quisinostat (JNJ-2648158) with or without TGF-β. Both HDAC inhibitors increased anguin-1/LSR, decrease CLDN-2, promoted G1 arrest and prevented the migration of A549 cells. Furthermore, TSA but not Quisinostat with or without TGF-β induced cellular metabolism indicated as the mitochondrial respiration measured using the oxygen consumption rate. In normal human lung epithelial cells, treatment with TSA and Quisinostat increased expression of LSR and CLDN-2 and decreased that of CLDN-1 with or without TGF-β in 2D culture. Quisinostat but not TSA with TGF-β increased CLDN-7 expression in 2D culture. Both HDAC inhibitors prevented disruption of the epithelial barrier measured as the permeability of FD-4 induced by TGF-β in 2.5D culture. TSA and Quisinostat have potential for use in therapy for lung adenocarcinoma via changes in the expression of angulin-1/LSR and CLDN-2.

Objectives: The LURET phase II study evaluated the efficacy and safety of the multikinase inhibitor vandetanib in patients with previously treatedRET-rearranged advanced non-small cell lung cancer (NSCLC). Among the eligible patients included in the primary analysis, the objective response rate met the primary endpoint (53 %, 90 % confidence interval [CI]: 31-74). Here, we report final survival outcomes of the LURET study., Materials and Methods: Nineteen patients with previously treated RET-rearranged advanced NSCLC continuously received 300 mg of oral vandetanib daily. This final analysis provides updated data on progression-free survival (PFS), overall survival (OS) and safety. This study was registered with UMIN-CTR (number UMIN 000010095)., Results: Among the 19 patients in the intention-to-treat population, 42 % had been heavily treated with 3 or more prior chemotherapy regimens. The median PFS was 6.5 months (95 % CI, 3.9-9.3) as determined by an independent radiology review committee. The median OS was 13.5 months (95 % CI, 9.8-28.1) and the overall survival rate at 12 months was 52.6 % (95 % CI 28.7-71.9). The most common adverse events were hypertension (84.2 %), diarrhea (78.9 %), and rash acneiform (63.2 %). Overall, 11 patients (57.9 %) had adverse events leading to a dose reduction, although the safety profile was consistent with that reported in previous studies., Conclusion: Our results indicated that vandetanib enabled a prolonged and clinically meaningful PFS and OS in patients with previously treatedRET-rearranged advanced NSCLC at the updated final analysis. The safety profile was consistent with that reported in previous studies, although most of the patients experienced off-target adverse events besides RET., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Introduction: The relationship between Body-Mass-Index (BMI) and lung cancer prognosis is heterogeneous. We evaluated the impact of sex, smoking and race on the relationship between BMI and overall survival (OS) in non-small-cell-lung-cancer (NSCLC)., Methods: Data from 16 individual ILCCO studies were pooled to assess interactions between BMI and the following factors on OS: self-reported race, smoking status and sex, using Cox models (adjusted hazard ratios; aHR) with interaction terms and adjusted penalized smoothing spline plots in stratified analyses., Results: Among 20,937 NSCLC patients with BMI values, females = 47 %; never-smokers = 14 %; White-patients = 76 %. BMI showed differential survival according to race whereby compared to normal-BMI patients, being underweight was associated with poor survival among white patients (OS, aHR = 1.66) but not among black patients (aHR = 1.06; p interaction  = 0.02). Comparing overweight/obese to normal weight patients, Black NSCLC patients who were overweight/obese also had relatively better OS (p interaction  = 0.06) when compared to White-patients. BMI was least associated with survival in Asian-patients and never-smokers. The outcomes of female ever-smokers at the extremes of BMI were associated with worse outcomes in both the underweight (p interaction <0.001) and obese categories (p interaction  = 0.004) relative to the normal-BMI category, when compared to male ever-smokers., Conclusion: Underweight and obese female ever-smokers were associated with worse outcomes in White-patients. These BMI associations were not observed in Asian-patients and never-smokers. Black-patients had more favorable outcomes in the extremes of BMI when compared to White-patients. Body composition in Black-patients, and NSCLC subtypes more commonly seen in Asian-patients and never-smokers, may account for differences in these BMI-OS relationships., (Copyright © 2020. Published by Elsevier B.V.)

Background: Non-small-cell lung cancer (NSCLC) has been reported to develop in patients with interstitial pneumonia (IP); however, clinical, radiological, and pathological features remain to be elucidated., Methods: We retrieved the records of 120 consecutive NSCLC patients associated with IP who underwent surgery at Toranomon Hospital between June 2011 and May 2017. We classified the patients into three groups according to NSCLC location using high-resolution computed tomography: group A, within a fibrotic shadow and/or at the interface of a fibrotic shadow and normal lung; group B, within emphysematous tissue and/or at the interface of emphysematous tissue and normal lung; and group C, within normal lung. In 64 patients, programmed death ligand-1 (PD-L1) status was assessed with immunohistostaining., Results: Most of the patients (89; 70%) were classified as group A. This group tended to have squamous cell carcinoma with the usual interstitial pneumonia (UIP). These cancers were located mainly in the lower lobes and seven of the eight postoperative acute exacerbations (pAE) of IP developed in this group. NSCLC in the group B were mainly squamous cell carcinomas located in the upper lobes. No patient with PD-L1 negative was classified into group B. None of the patients in group C showed UIP. and most of the cancers were adenocarcinoma. The frequency of epidermal growth factor receptor mutation-positive NSCLC was the highest in this group., Conclusions: The three groups each showed characteristic features in terms of tumor location, histopathology, PD-L1 expression, and frequency of pAEof IP.

Here, we report the application of a long-read sequencer, PromethION, for analyzing human cancer genomes. We first conducted whole-genome sequencing on lung cancer cell lines. We found that it is possible to genotype known cancerous mutations, such as point mutations. We also found that long-read sequencing is particularly useful for precisely identifying and characterizing structural aberrations, such as large deletions, gene fusions, and other chromosomal rearrangements. In addition, we identified several medium-sized structural aberrations consisting of complex combinations of local duplications, inversions, and microdeletions. These complex mutations occurred even in key cancer-related genes, such as STK11 , NF1 , SMARCA4 , and PTEN The biological relevance of those mutations was further revealed by epigenome, transcriptome, and protein analyses of the affected signaling pathways. Such structural aberrations were also found in clinical lung adenocarcinoma specimens. Those structural aberrations were unlikely to be reliably detected by conventional short-read sequencing. Therefore, long-read sequencing may contribute to understanding the molecular etiology of patients for whom causative cancerous mutations remain unknown and therapeutic strategies are elusive., (© 2020 Sakamoto et al.; Published by Cold Spring Harbor Laboratory Press.)

Introduction: The relationships between morbid obesity, changes in body mass index (BMI) before cancer diagnosis, and lung cancer outcomes by histology (SCLC and NSCLC) have not been well studied., Methods: Individual level data analysis was performed on 25,430 patients with NSCLC and 2787 patients with SCLC from 16 studies of the International Lung Cancer Consortium evaluating the association between various BMI variables and lung cancer overall survival, reported as adjusted hazard ratios (aHRs) from Cox proportional hazards models and adjusted penalized smoothing spline plots., Results: Overall survival of NSCLC had putative U-shaped hazard ratio relationships with BMI based on spline plots: being underweight (BMI < 18.5 kg/m 2 ; aHR = 1.56; 95% confidence interval [CI]:1.43-1.70) or morbidly overweight (BMI > 40 kg/m 2 ; aHR = 1.09; 95% CI: 0.95-1.26) at the time of diagnosis was associated with worse stage-specific prognosis, whereas being overweight (25 kg/m 2 ≤ BMI < 30 kg/m 2 ; aHR = 0.89; 95% CI: 0.85-0.95) or obese (30 kg/m 2 ≤ BMI ≤ 40 kg/m 2 ; aHR = 0.86; 95% CI: 0.82-0.91) was associated with improved survival. Although not significant, a similar pattern was seen with SCLC. Compared with an increased or stable BMI from the period between young adulthood until date of diagnosis, a decreased BMI was associated with worse outcomes in NSCLC (aHR = 1.24; 95% CI: 1.2-1.3) and SCLC patients (aHR=1.26 (95% CI: 1.0-1.6). Decreased BMI was consistently associated with worse outcome, across clinicodemographic subsets., Conclusions: Both being underweight or morbidly obese at time of diagnosis is associated with lower stage-specific survival in independent assessments of NSCLC and SCLC patients. In addition, a decrease in BMI at lung cancer diagnosis relative to early adulthood is a consistent marker of poor survival., (Copyright © 2019 International Association for the Study of Lung Cancer. Published by Elsevier Inc. All rights reserved.)

Next generation sequencing (NGS)-based tumor profiling identified an overwhelming number of uncharacterized somatic mutations, also known as variants of unknown significance (VUS). The therapeutic significance of EGFR mutations outside mutational hotspots, consisting of >50 types, in nonsmall cell lung carcinoma (NSCLC) is largely unknown. In fact, our pan-nation screening of NSCLC without hotspot EGFR mutations ( n = 3,779) revealed that the majority (>90%) of cases with rare EGFR mutations, accounting for 5.5% of the cohort subjects, did not receive EGFR-tyrosine kinase inhibitors (TKIs) as a first-line treatment. To tackle this problem, we applied a molecular dynamics simulation-based model to predict the sensitivity of rare EGFR mutants to EGFR-TKIs. The model successfully predicted the diverse in vitro and in vivo sensitivities of exon 20 insertion mutants, including a singleton, to osimertinib, a third-generation EGFR-TKI ( R 2 = 0.72, P = 0.0037). Additionally, our model showed a higher consistency with experimentally obtained sensitivity data than other prediction approaches, indicating its robustness in analyzing complex cancer mutations. Thus, the in silico prediction model will be a powerful tool in precision medicine for NSCLC patients carrying rare EGFR mutations in the clinical setting. Here, we propose an insight to overcome mutation diversity in lung cancer., Competing Interests: The authors declare no conflict of interest., (Copyright © 2019 the Author(s). Published by PNAS.)

Objective: Tumor-associated macrophages (TAMs) are believed to influence tumor progression and the prognosis of patients. The purpose of this study was to clarify the correlation between the TAM density or location and the clinicopathological features of non-small-cell lung cancer (NSCLC) as well as to explore the prognostic impact of TAMs in NSCLC., Materials and Methods: CD68- and CD204-positive macrophages were detected in tumor islets, tumor stroma and alveolar space in 297 patients with NSCLC using immunochemistry. The clinicopathological and genetic factors surveyed were the disease-free survival, age, gender, smoking status, histological type, disease stage, histological grade, pleural invasion, lymph node metastasis, EGFR gene mutations and ALK rearrangements., Results: There were significantly more CD68-positive macrophages than CD204-positive macrophages in each location of the tumor islets, tumor stroma and alveolar spaces, and they were strongly correlated (P < 0.0001 each). Factors such as male gender, being a smoker, an advanced disease stage and histological grade, positive pleural invasion and node status and wild-type EGFR gene status were significantly correlated with a higher density of CD68- and CD204-positive TAMs in tumor stroma (P < 0.05 each). In contrast, the age of patients was not correlated with CD68- and CD204-positive TAMs (P > 0.05 each). Furthermore, survival analysis revealed that a high number of CD68- and CD204-positive TAMs in tumor stroma, but not in tumor islets or alveolar space, was a significant prognostic factor for the disease-free survival time of NSCLC (P < 0.05, respectively). Moreover, both univariate and multivariate analyses confirmed that higher numbers of CD204-positive TAMs in tumor stroma were an independent worse prognostic predictor for adenocarcinoma., Conclusion: The tumor stroma is the most suitable intratumoral area for the evaluation of TAMs in the setting of the prognostic prediction of NSCLC patients. CD204-positive TAMs are the preferable marker for prognostic prediction in NSCLC, especially in lung adenocarcinoma., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Objective: The aim of the study was to investigate the relationship between germline variations as a prognosis biomarker in patients with advanced Non-Small-Cell-Lung-Cancer (NSCLC) subjected to first-line platinum-based treatment., Materials and Methods: We carried out a two-stage genome-wide-association study in non-small-cell lung cancer patients with platinum-based chemotherapy in an exploratory sample of 181 NSCLC patients from Caucasian origin, followed by a validation on 356 NSCLC patients from the same ancestry (Valencia, Spain)., Results: We identified germline variants in SMYD2 as a prognostic factor for survival in patients with advanced NSCLC receiving chemotherapy. SMYD2 alleles are associated to a decreased overall survival and with a reduced Time to Progression. In addition, enrichment pathway analysis identified 361 variants in 40 genes to be involved in poorer outcome in advanced-stage NSCLC patients., Conclusion: Germline SMYD2 alleles are associated with bad clinical outcome of first-line platinum-based treatment in advanced NSCLC patients. This result supports the role of SMYD2 in the carcinogenic process, and might be used as prognostic signature directing patient stratification and the choice of therapy., Microabstract: A two-Stage Genome wide association study in Caucasian population reveals germline genetic variation in SMYD2 associated to progression disease in first-line platinum-based treatment in advanced NSCLC patients. SMYD2 profiling might have prognostic / predictive value directing choice of therapy and enlighten current knowledge on pathways involved in human carcinogenesis as well in resistance to chemotherapy., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Purpose: miR-451 is a tumor suppressive microRNA with several target genes, including Macrophage migration inhibitory factor (MIF). As little is known about the expression and clinicopathological significance of mir-451 in NSCLC, we performed a clinicopathological study of 370 NSCLC cases to clarify them. Cell biological experiments were also performed on NSCLC cell lines to confirm the tumor-suppressive role of miR-451 and whether or not MIF is targeted by miR-451., Methods: We analyzed 370 NSCLC cases for the miR-451 expression by quantitative real-time polymerase chain reaction and the MIF expression by immunohistochemistry. Eighty-four background lung tissue samples were also evaluated for the miR-451 expression. The clinicopathological and genetic factors surveyed were the disease-free survival, smoking status, histological type, disease stage, EGFR gene mutations and ALK rearrangements. In 286 adenocarcinoma cases, the invasive status (adenocarcinoma in situ, minimally invasive adenocarcinoma and invasive adenocarcinoma) was also evaluated. Five NSCLC cell lines (H23, H441, H522, H1703, and H1975) were cultured and evaluated for their miR-451 and MIF expression. The cell lines with lower miR-451 and higher MIF expressions were then selected and transfected with miR-451-mimic to observe its effects on MIF expression, Akt and Erk status, cell proliferation, and cell migration., Results: The miR-451 expression was down-regulated in cancer tissues compared with background lung tissues (P<0.0001). Factors such as advanced disease stage, positive pleural invasion and nodal status and being a smoker were significantly correlated with a lower expression of miR-451 (P<0.05 each), while EGFR gene mutations and ALK rearrangements were not. In adenocarcinoma, invasive and minimally invasive adenocarcinoma showed lower expression of miR-451 than adenocarcinoma in situ (P<0.0005, respectively). A survival analysis showed that a lower expression of miR-451 was an independent predictor of a poor prognosis for NSCLC (P<0.05). The MIF expression was inversely correlated with the miR-451 expression. Out of 5 NSCLC cell lines examined, H441 and H1975 showed higher MIF and lower miR-451 expressions. After the transfection of miR-451-mimic, the MIF expression and phosphorylated Akt expression of these cell lines was suppressed, as were cell proliferation and cell migration., Conclusion: This clinicopathological study of 370 NSCLC cases and the cell biological studies of NSCLC cell lines clarified the tumor-suppressive role of miR-451 and its prognostic value. We also validated MIF as a target of miR-451 in NSCLC.

Introduction: There are no validated molecular methods that prospectively identify patients with surgically resected lung squamous cell carcinoma (SCC) at high risk for recurrence. By focusing on the expression of genes with known functions in development of lung SCC and prognosis, we sought to develop a robust prognostic classifier of early-stage lung SCC., Methods: The expression of 253 genes selected by literature search was evaluated in microarrays from 107 stage I/II tumors. Associations with survival were evaluated by Cox regression and Kaplan-Meier survival analyses in two independent cohorts of 121 and 91 patients with SCC, respectively. A classifier score based on multivariable Cox regression was derived and examined in six additional publicly available data sets of stage I/II lung SCC expression profiles (n = 358). The prognostic value of this classifier was evaluated in meta-analysis of patients with stage I/II (n = 479) and stage I (n = 326) lung SCC., Results: Dual specificity phosphatase 6 gene (DUSP6) and actinin alpha 4 gene (ACTN4) were associated with prognostic outcome in two independent patient cohorts. Their expression values were utilized to develop a classifier that identified patients with stage I/II lung SCC at high risk for recurrence (hazard ratio [HR] = 4.7, p = 0.018) or cancer-specific mortality (HR = 3.5, p = 0.016). This classifier also identified patients at high risk for recurrence (HR = 2.7, p = 0.008) or death (HR = 2.2, p = 0.001) in publicly available data sets of stage I/II and in meta-analysis of stage I patients., Conclusions: We have established and validated a prognostic classifier to inform clinical management of patients with lung SCC after surgical resection., (Published by Elsevier Inc.)

Background: RET rearrangements are rare oncogenic alterations in non-small-cell lung cancer (NSCLC). Vandetanib is a multitargeted tyrosine kinase inhibitor exhibiting RET kinase activity. We aimed to assess the efficacy and safety of vandetanib in patients with advanced RET-rearranged NSCLC., Methods: In this open-label, multicentre, phase 2 trial (LURET), patients with advanced RET-rearranged NSCLC continuously received 300 mg of oral vandetanib daily. RET-positive patients were screened using a nationwide genomic screening network of about 200 participating institutions. Primary endpoint was the independently assessed objective response in eligible patients. This study is registered with UMIN-CTR, number UMIN000010095., Findings: Between Feb 7, 2013, and March 19, 2015, 1536 patients with EGFR mutation-negative NSCLC were screened, of whom 34 were RET-positive (2%) and 19 were enrolled. Among 17 eligible patients included in primary analysis, nine (53% [95% CI 28-77]) achieved an objective response, which met the primary endpoint. In the intention-to-treat population of all 19 patients treated with vandetanib, nine (47% [95% CI 24-71]) achieved an objective response. At the data cutoff, median progression-free survival was 4·7 months (95% CI 2·8-8·5). The most common grade 3 or 4 adverse events were hypertension (11 [58%]), diarrhoea (two [11%]), rash (three [16%]), dry skin (one [5%]), and QT prolongation (two [11%])., Interpretation: Vandetanib showed clinical antitumour activity and a manageable safety profile in patients with advanced RET-rearranged NSCLC. Our results define RET rearrangement as a new molecular subgroup of NSCLC suitable for targeted therapy., Funding: The Ministry of Health, Labour and Welfare of Japan and the Practical Research for Innovation Cancer Control from the Japan Agency for Medical Research and Development, AMED., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The frequency and clinical profile of patients with stage III non-small cell lung cancer harboring KRAS mutations have not yet been well documented. Here, we analyzed hotspot KRAS mutations using high-resolution melting analyses in tumor specimens from patients who received chemoradiotherapy between January 2001 and December 2010 at the National Cancer Center Hospital. The associations between the presence of KRAS mutations and the response rate, relapse-free survival, first relapse sites, survival post-progression and overall survival were investigated. A total of 274 non-squamous non-small cell lung cancer patients received chemoradiotherapy at our hospital. After excluding 121 patients for whom tumor specimens were not available and 34 patients with EGFR mutations, the remaining 119 patients were included in the analysis. KRAS mutations were found at a frequency of 13%. Patients with KRAS mutations had a shorter median relapse-free survival (6.1 vs 10.9 months) and a lower response rate (63% vs 81%). As for the first relapse site, patients with KRAS mutations had fewer local relapses (8% vs 23%) and more brain metastases (46% vs 12%). After disease progression, patients with KRAS mutations had a significantly shorter median survival post-progression (2.5 vs 7.3 months, P = 0.028) and median overall survival (15.1 vs 29.1 months, P = 0.022). Our results suggested that KRAS mutation could be associated with a reduced efficacy of chemoradiotherapy and a shortened survival time., (© 2015 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.)

PTPRH is a receptor-type protein tyrosine phosphatase thought to be a potential regulator of tumorigenesis. The aim of the present study was to clarify the significance of PTPRH expression and its regulation by DNA methylation in non-small cell lung cancer (NSCLC), especially in lung adenocarcinoma (LADC). PTPRH mRNA expression was examined in 89 NSCLC and corresponding non-cancerous tissues. The correlation between DNA methylation and PTPRH gene expression was investigated in another cohort that consisted of 145 patients with LADC, a major NSCLC subtype. Gene regulation by DNA methylation was assessed using a DNA methylation inhibitor. PTPRH mRNA expression was significantly upregulated in NSCLC. PTPRH DNA methylation was reduced in LADC samples and inversely correlated with mRNA expression. 5-Aza-2'-deoxycytidine treatment of lung cancer cell lines with low PTPRH expression, restored mRNA PTPRH expression levels. Furthermore, low PTPRH methylation was associated with shorter recurrence-free survival (P=1.64x10(-4)) and overall survival (P=5.54x10(-5)). Multivariate analysis revealed that PTPRH DNA methylation was an independent prognostic factor (P=6.88x10(-3)). It was confirmed that PTPRH is overexpressed in NSCLC. Furthermore, we determined that PTPRH is epigenetically regulated by DNA hypomethylation, with prognostic implications for LADC.

Carbon-ion radiotherapy (CIRT) holds promise to treat inoperable locally-advanced non-small cell lung carcinoma (NSCLC), a disease poorly controlled by standard chemoradiotherapy using X-rays. Since CIRT is an extremely limited medical resource, selection of NSCLC patients likely to benefit from it is important; however, biological predictors of response to CIRT are ill-defined. The present study investigated the association between the mutational status of EGFR and KRAS, driver genes frequently mutated in NSCLC, and the relative biological effectiveness (RBE) of carbon-ion beams over X-rays. The assessment of 15 NSCLC lines of different EGFR/KRAS mutational status and that of isogenic NSCLC lines expressing wild-type or mutant EGFR revealed that EGFR-mutant NSCLC cells, but not KRAS-mutant cells, show low RBE. This was attributable to (i) the high X-ray sensitivity of EGFR-mutant cells, since EGFR mutation is associated with a defect in non-homologous end joining, a major pathway for DNA double-strand break (DSB) repair, and (ii) the strong cell-killing effect of carbon-ion beams due to poor repair of carbon-ion beam-induced DSBs regardless of EGFR mutation status. These data highlight the potential of EGFR mutation status as a predictor of response to CIRT, i.e., CIRT may show a high therapeutic index in EGFR mutation-negative NSCLC.

Background: Dysregulation of HER2 signaling pathways results in tumor progression in several types of carcinomas. The aim of the current study is to identify clinicopathological characteristics of HER2-mutated non-small cell lung carcinomas (NSCLCs) in conjunction with HER2 protein expression, gene amplification, and phosphorylation., Patients and Methods: We investigated 1275 patients including 1055 adenocarcinomas (ADCs), 146 squamous cell carcinomas, 2 large cell carcinomas, 8 sarcomatoid carcinomas, and 64 adenosquamous carcinomas. High-resolution melting analysis of HER2 hot spot inflame deletion mutations, chromogenic in situ hybridization for HER2 amplification, and immunostaining of wild-type and phosphorylated HER2 was performed., Results: HER2 mutations were detected in 46 (3.6%) of the NSCLCs, with mutations only present in the ADC. When analyzing ADC cases alone, the incidence of HER2 mutation was increased to 4.3%. All HER2-mutated tumors were negative for other driver gene alterations. HER2 mutation status correlated with never-smoker status and patients with smaller tumor size. HER2 amplifications were also identified in approximately half of the tumors with HER2 mutations. The overall survival rate was not significantly different between patients without and with HER2 mutations. When analyzing only invasive ADCs, HER2 mutation status was an independent factor for an unfavorable outcome. Amongst the 46 patients harboring HER2 mutations, univariate and multivariate analysis revealed that HER2 amplification was an unfavorable prognostic factor, while HER2 phosphorylation was a favorable prognostic factor., Conclusions: HER2 mutations were observed in 3.6% of NSCLCs, particularly in younger patients, those with no history of smoking, and those with small tumors. Amongst the patients with HER2 mutations, HER2 amplification and phosphorylation were independent prognostic factors., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Purpose: To determine the frequency and clinical significance of epidermal growth factor receptor (EGFR) mutations in patients with potentially curable stage III non-small-cell lung cancer (NSCLC) who are eligible for definitive chemoradiotherapy (CRT)., Patients and Methods: Between January 2001 and December 2010, we analyzed the EGFR mutational status in consecutive NSCLC patients who were treated by CRT. The response rate, relapse-free survival, 2-year relapse-free rate, initial relapse sites, and overall survival of the patients were investigated., Results: A total of 528 patients received CRT at our hospital during the study period. Of these, 274 were diagnosed as having nonsquamous NSCLC. Sufficient specimens for mutational analyses could be obtained from 198 of these patients. The proportion of patients with EGFR activating mutations was 17%. In addition to the well-known characteristics of patients carrying EGFR mutations (female, adenocarcinoma, and never/light smoker), the proportion of cases with smaller primary lesions (T1/2) was found to be higher in patients with EGFR mutations than in those with wild-type EGFR. Patients with EGFR mutations showed similar response rate, relapse-free survival, and 2-year relapse-free rates as compared to patients with wild-type EGFR. Local relapses as the site of initial relapse occurred significantly less frequently in patients with EGFR mutation (4% vs 21%; P=.045). Patients with EGFR mutations showed longer local control (adjusted hazard ratio 0.49; P=.043). After disease progression, a majority of the patients with EGFR mutations received EGFR tyrosine kinase inhibitors (62%), and these patients showed longer postprogression survival than those with wild-type EGFR., Conclusions: Our study is the first to show radiosensitive biology of EGFR-mutated tumors in definitive CRT with curative intent. This finding could serve as a credible baseline estimate of EGFR-mutated population in stage III nonsquamous NSCLC., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background and Purpose: Chromatin remodeling through histone modifications, including acetylation, plays an important role in the appropriate response to DNA damage induced by ionizing radiation (IR). Here we investigated the radiosensitizing effect of C646, a selective small molecule inhibitor of p300 histone acetyltransferase, and explored the underlying mechanisms., Materials and Methods: A549, H157 and H460 human non-small cell lung carcinoma (NSCLC) cells, and HFL-III human lung fibroblasts were assessed by clonogenic survival assay. Apoptosis and necrosis were assessed by annexin V staining. Senescence was assessed by Senescence-associated β-galactosidase staining. Mitotic catastrophe was assessed by evaluating nuclear morphology with DAPI staining. Cell cycle profiles were analyzed by flow cytometry. Protein expression was analyzed by immunoblotting., Results: C646 sensitized A549, H460 and H157 cells to IR with a dose enhancement ratio at 10% surviving fraction of 1.4, 1.2 and 1.2, respectively. C646 did not radiosensitize HFL-III cells. In A549 cells, but not in HFL-III cells, C646 (i) enhanced mitotic catastrophe but not apoptosis, necrosis, or senescence after IR; (ii) increased the hyperploid cell population after IR; and (iii) suppressed the phosphorylation of CHK1 after IR., Conclusions: C646 radiosensitizes NSCLC cells by enhancing mitotic catastrophe through the abrogation of G2 checkpoint maintenance., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Background: To elucidate clinicopathological characteristics of non-small-cell lung carcinoma (NSCLC) cases carrying RET rearrangements causing oncogenic fusions to identify responders to therapy with RET tyrosine kinase inhibitors., Methods: We investigated 1874 patients with carcinomas, including 1620 adenocarcinomas (ADCs), 203 squamous cell carcinomas (SCCs), 8 large cell carcinomas, and 43 sarcomatoid carcinomas (SACs). Fluorescence in situ hybridisation (FISH) and/or reverse transcription-PCR (RT-PCR) were performed to detect RET gene rearrangement., Results: In all, 22 cases (1.2%) showed RET rearrangements; all cases were of ADC histology. Of the 22 patients, 19 possessed KIF5B-RET fusion genes, whereas 3 possessed CCDC6-RET fusion genes. The RET-rearranged tumours were significantly more common in younger patients (P=0.038) and tended to occur in patients with no history of smoking (P=0.051). In addition, RET rearrangements were not associated with gender, occupational history (particularly radioactive exposure), tumour size, lymph node status, tumour stage, or patient survival. The predominant growth pattern in RET-rearranged ADCs was lepidic in 6 cases, papillary in 9 cases, acinar in 2 cases, micropapillary in 1 case, and solid in 4 cases. Cells with cytoplasmic mucin production were at least focally present in 12 of the 22 (54.5%) RET-rearranged ADC cases. Among the 21 analysed RET-rearranged tumours, RET immunopositivity was observed in 15 cases (71.4%), and was significantly associated with RET rearrangement (P<0.001)., Conclusions: The RET rearrangements were observed in 1.2% of NSCLCs. All cases of RET rearrangement were ADCs. The RET rearrangements were more likely to be observed in younger patients. Although cytoplasmic mucin production was at least focally present in 54.5% of RET-rearranged ADCs, specific histological features were not detected.

Our knowledge of small cell lung cancer (SCLC) genetics is still very limited, amplification of L-MYC, N-MYC, and C-MYC being some of the well-established gene alterations. Here, we report our discovery of tumor-specific inactivation of the MYC-associated factor X gene, MAX, in SCLC. MAX inactivation is mutually exclusive with alterations of MYC and BRG1, the latter coding for an ATPase of the switch/sucrose nonfermentable (SWI/SNF) complex. We demonstrate that BRG1 regulates the expression of MAX through direct recruitment to the MAX promoter, and that depletion of BRG1 strongly hinders cell growth, specifically in MAX-deficient cells, heralding a synthetic lethal interaction. Furthermore, MAX requires BRG1 to activate neuroendocrine transcriptional programs and to upregulate MYC targets, such as glycolysis-related genes. Finally, inactivation of the MAX dimerization protein, MGA, was also observed in both non-small cell lung cancer and SCLC. Our results provide evidence that an aberrant SWI/SNF-MYC network is essential for lung cancer development.

Background: Recently, driver tyrosine kinase gene mutations have been detected in malignant tumors, including lung tumors. Notwithstanding their attractiveness as targets for molecular therapy, limited information is available regarding BRAF-mutated lung carcinomas., Materials and Methods: BRAF mutation status was determined in 2001 surgically resected nonsmall-cell lung cancer (NSCLC) cases using high-resolution melting analysis (HRMA) followed by Sanger sequencing and/or deep sequencing using next generation sequencer., Results: BRAF mutations were detected in 26 (1.3%) of 2001 NSCLC cases (25 adenocarcinomas and 1 squamous cell carcinoma). In the 26 cases, 13 mutation genotypes were identified, including V600E (8 of 26; 30.8%), G469A (6 of 26; 23.1%), K601E (4 of 26; 15.4%), and other residual mutations (1 of 26; 0.04%). Of the 13 genotypes, 4 genotypes (G464E, G596R, A598T, and G606R) had not been previously reported in lung cancer. The overall survival rate was not significantly different between patients with wild-type BRAF and those with V600E or non-V600E BRAF mutations (P = 0.49 and P = 0.15, respectively). Histomorphological analysis revealed that focal clear cell changes were present in 75% of V600E-mutated tumors. All V600E BRAF-mutated tumors were negative for other driver gene alterations including epidermal growth factor receptor (EGFR) and KRAS mutations and the anaplastic lymphoma kinase gene translocation, whereas five tumors with non-V600E BRAF mutations (four G469A and one G464E/G466R) showed concomitant EGFR mutations., Conclusion: The frequency of BRAF mutations in lung cancer was low in an Asian cohort. Furthermore, BRAF mutation status lacked prognostic significance in this patient population.

The recent discovery of fusion oncokinases in a subset of non-small cell lung carcinomas (NSCLCs) is of considerable clinical interest, since NSCLCs that express such fusion oncokinases are reportedly sensitive to kinase inhibitors. To better understand the role of recently identified ROS1 and RET fusion oncokinases in pulmonary carcinogenesis, we examined 114 NSCLCs for SLC34A2-ROS1, EZR-ROS1, CD74-ROS1 and KIF5B-RET fusion transcripts using RT-polymerase chain reaction and subsequent sequencing analyses. Although the expression of SLC34A2-ROS1, EZR-ROS1, or KIF5B-RET fusion transcripts was not detected in any of the cases, the expression of CD74-ROS1 fusion transcripts was detected in one (0.9%) of the 114 NSCLCs. The fusion occurred between exon 6 of CD74 and exon 34 of ROS1 and was an in-frame alteration. The mutation was detected in a woman without a history of smoking. Histologically, the carcinoma was an adenocarcinoma with a predominant acinar pattern; notably, a mucinous cribriform pattern and a solid signet-ring cell pattern were also observed in part of the adenocarcinoma. ROS1 protein overexpression was immunohistochemically detected in a cancer-specific manner in both the primary cancer and the lymph node metastatic cancer. No somatic mutations were detected in the mutation cluster regions of the KRAS, EGFR, BRAF and PIK3CA genes and the entire coding region of p53 in the carcinoma, and the expression of ALK fusion was negative. The above results suggest that CD74-ROS1 fusion is involved in the carcinogenesis of a subset of NSCLCs and may contribute to the elucidation of the characteristics of ROS1 fusion-positive NSCLC in the future.

The occurrence of inactivating mutations in SWI/SNF chromatin-remodeling genes in common cancers has attracted a great deal of interest. However, mechanistic strategies to target tumor cells carrying such mutations are yet to be developed. This study proposes a synthetic-lethality therapy for treating cancers deficient in the SWI/SNF catalytic (ATPase) subunit, BRG1/SMARCA4. The strategy relies upon inhibition of BRM/SMARCA2, another catalytic SWI/SNF subunit with a BRG1-related activity. Immunohistochemical analysis of a cohort of non-small-cell lung carcinomas (NSCLC) indicated that 15.5% (16 of 103) of the cohort, corresponding to preferentially undifferentiated tumors, was deficient in BRG1 expression. All BRG1-deficient cases were negative for alterations in known therapeutic target genes, for example, EGFR and DDR2 gene mutations, ALK gene fusions, or FGFR1 gene amplifications. RNA interference (RNAi)-mediated silencing of BRM suppressed the growth of BRG1-deficient cancer cells relative to BRG1-proficient cancer cells, inducing senescence via activation of p21/CDKN1A. This growth suppression was reversed by transduction of wild-type but not ATPase-deficient BRG1. In support of these in vitro results, a conditional RNAi study conducted in vivo revealed that BRM depletion suppressed the growth of BRG1-deficient tumor xenografts. Our results offer a rationale to develop BRM-ATPase inhibitors as a strategy to treat BRG1/SMARCA4-deficient cancers, including NSCLCs that lack mutations in presently known therapeutic target genes., (©2013 AACR.)

Recent discovery of ROS1 gene fusion in a subset of lung cancers has raised clinical interest, because ROS1 fusion-positive cancers are reportedly sensitive to kinase inhibitors. To better understand these tumors, we examined 799 surgically resected non-small cell lung cancers by reverse transcriptase polymerase chain reaction and identified 15 tumors harboring ROS1 fusion transcripts (2.5% of adenocarcinomas). The most frequent fusion partner was CD74 followed by EZR. The affected patients were often younger nonsmoking female individuals, and they had overall survival rates similar to those of the ROS1 fusion-negative cancer patients. All the ROS1 fusion-positive tumors were adenocarcinomas except 1, which was an adenosquamous carcinoma. Histologic examination identified an at least focal presence of either solid growth with signet-ring cells or cribriform architecture with abundant extracellular mucus in 53% of the cases. These 2 patterns are reportedly also characteristic of anaplastic lymphoma kinase (ALK)-rearranged lung cancers, and our data suggest a phenotypic resemblance between the ROS1-rearranged and ALK-rearranged tumors. All tumors except 1 were immunoreactive to thyroid transcription factor-1. Fluorescence in situ hybridization using ROS1 break-apart probes revealed positive rearrangement signals in 23% to 93% of the tumor cells in ROS1 fusion-positive cancers, which were readily distinguished using a 15% cutoff value from 50 ROS1 fusion-negative tumors tested, which showed 0% to 6% rearrangement signals. However, this perfect test performance was achieved only when isolated 3' signals were included along with classic split signals in the definition of rearrangement positivity. Fluorescence in situ hybridization signal patterns were unrelated to 5' fusion partner genes. All ROS1 fusion-positive tumors lacked alteration of EGFR, KRAS, HER2, ALK, and RET genes.

Genetic rearrangement of the ROS1 receptor tyrosine kinase was recently identified as a distinct molecular signature for human non-small cell lung cancer (NSCLC). However, direct evidence of lung carcinogenesis induced by ROS1 fusion genes remains to be verified. The present study shows that EZR-ROS1 plays an essential role in the oncogenesis of NSCLC harboring the fusion gene. EZR-ROS1 was identified in four female patients of lung adenocarcinoma. Three of them were never smokers. Interstitial deletion of 6q22-q25 resulted in gene fusion. Expression of the fusion kinase in NIH3T3 cells induced anchorage-independent growth in vitro, and subcutaneous tumors in nude mice. This transforming ability was attributable to its kinase activity. The ALK/MET/ROS1 kinase inhibitor, crizotinib, suppressed fusion-induced anchorage-independent growth of NIH3T3 cells. Most importantly, established transgenic mouse lines specifically expressing EZR-ROS1 in lung alveolar epithelial cells developed multiple adenocarcinoma nodules in both lungs at an early age. These data suggest that the EZR-ROS1 is a pivotal oncogene in human NSCLC, and that this animal model could be valuable for exploring therapeutic agents against ROS1-rearranged lung cancer.

Introduction: The hepatocyte growth factor/MET pathway has been shown to cause tumor progression in several types of carcinomas. The aim of this study was to examine the correlations between c-MET/phospho-MET expression as well as MET gene copy number alterations and overall survival (OS) in non-small cell lung carcinomas (NSCLCs)., Methods: We analyzed 906 NSCLCs including 704 adenocarcinomas (ADCs), 150 squamous cell carcinomas (SCCs), 43 sarcomatoid carcinomas, and 9 large cell carcinomas. The mutational status of epidermal growth factor receptor and K-ras and anaplastic lymphoma kinase rearrangements were retrospectively examined. We performed immunohistochemistry to detect c-MET/phospho-MET expression and MET gene copy number using bright-field in situ hybridization (BISH)., Results: c-MET/phospho-MET expression and MET BISH positivity were observed in 22.2%, 5.6%, and 10.9% of NSCLCs, respectively; they were more prevalent in ADCs (27.3%, 6.9%, and 11.5%, respectively) and sarcomatoid carcinomas (20.9%, 9.3%, and 36.6%, respectively) than in SCCs and large cell carcinomas. Among ADCs, poorly differentiated cases exhibited c-MET expression and MET BISH positivity more commonly than well-differentiated ones. An analysis of all patients revealed that c-MET/phospho-MET expression and MET BISH positivity were not correlated with OS. However, when SCC cases were excluded, both univariate (p=0.019) and multivariate (p=0.020) analyses revealed a significant correlation between MET BISH positivity and OS., Conclusions: c-MET/phospho-MET expression and MET BISH positivity differed according to histological type. Among ADCs, c-MET expression and MET BISH positivity were more common in poorly differentiated cases. MET BISH positivity was an independent prognostic factor in nonsquamous NSCLCs.

A subset (1% to 5%) of non-small-cell lung carcinomas harbors the EML4-ALK fusion gene. Data from previous studies on the histomorphology of ALK-rearranged lung cancer are inconsistent, and the specific histologic parameters that characterize this subset and how accurately such parameters predict underlying ALK abnormality remain uncertain. To answer these questions, we performed a comprehensive histologic analysis of 54 surgically resected, extensively sampled ALK-rearranged lung carcinomas and compared them with 100 consecutive resections of ALK-wild-type lung cancers. All 54 cases showed at least a focal adenocarcinoma component, and 3 and 2 cases had additional squamous and sarcomatoid differentiation, respectively. Solid or acinar growth pattern, cribriform structure, presence of mucous cells (signet-ring cells or goblet cells), abundant extracellular mucus, lack of lepidic growth, and lack of significant nuclear pleomorphism were more common in ALK-positive cancers. Two recognizable constellations of findings, a solid signet-ring cell pattern and a mucinous cribriform pattern, were present at least focally in the majority (78%) of ALK-positive tumors, but were rare (1%) in ALK-negative tumors. Multivariate analysis showed that a combination of these 2 patterns was the most powerful histologic indicator of ALK rearrangement. Characteristic histologies were present both in primary sites and in metastases. Thus, histologic findings may help to identify cases for ALK testing. However, none of the histologic parameters were completely sensitive or specific to ALK rearrangement, and histomorphology should not replace confirmatory molecular or immunohistochemical studies. ALK-positive cancers commonly showed coexpression of thyroid transcription factor-1 and p63, and its significance is currently unclear.

Purpose: There is increasing evidence that altered microRNA expression is associated with tumor progression and survival in cancer patients. We tested if the expression of specific microRNAs was associated with prognosis and disease progression in early-stage lung adenocarcinoma., Experimental Design: The expression of miR-21, miR-17, and miR-155 was measured by quantitative RT-PCR in tissues from 317 non-small cell lung cancer (NSCLC) patients that originated from Maryland, Norway, and Japan. Kaplan-Meier and Cox regression analysis evaluated associations of microRNA expression with cancer-specific mortality and disease-free survival., Results: Elevated miR-21 (HR 2.06, 1.13-3.75), miR-17 (HR 2.00, 1.10-3.61), and miR-155 (HR 2.37, 1.27-4.42) was associated with worse cancer-specific mortality in the Maryland cohort. These were evaluated in two additional cohorts and only miR-21 was associated with worse cancer-specific mortality in the Norwegian cohort (HR 2.78, 1.22-6.31) and worse relapse-free survival in the Japanese cohort (HR 2.82, 1.57-5.07). More advanced stage tumors expressed significantly higher levels of miR-21 compared with TNM stage I tumors. TNM stage I patients were evaluated separately and high levels of miR-21 was associated with worse cancer-specific mortality (HR 2.16, 1.11-4.21) and relapse-free survival (3.40, 1.57-7.36) independent of other clinical factors., Conclusions: This is the first study to report that increased miR-21 expression is associated with disease progression and survival in stage I lung cancer. This suggests that expression of miR-21 may contribute to lung carcinogenesis and serve as a therapeutic target or early-stage prognostic biomarker for lung adenocarcinoma.

Purpose: To identify polymorphisms in DNA repair genes that affect responses to platinum-based doublet chemotherapy in patients with non-small-cell lung cancer (NSCLC)., Patients and Methods: In total, 640 patients with NSCLC who received platinum-based doublet chemotherapy in the National Cancer Center Hospital in Japan from 2000 to 2008 and whose responses were evaluated by Response Evaluation Criteria in Solid Tumors (RECIST) participated in a study of the association between response and genotypes for 30 single nucleotide polymorphisms (SNPs) in 27 DNA repair genes. Candidate SNPs were selected in a discovery set of 201 patients, and their associations were validated in an independent set of 439 patients by prespecified P value criteria., Results: Homozygotes for the minor allele TP53-72Pro of the Arg72Pro SNP in the TP53 gene showed a better response rate (54.3%) than those for the major allele TP53-72Arg (29.1%; P = 4.4 × 10(-5)) irrespective of therapeutic regimens, and minor allele homozygotes had significantly longer progression-free and overall survivals than major allele homozygotes (hazard ratio [HR], 0.85; 95% CI, 0.74 to 0.98; P = .020; and HR, 0.86; 95% CI, 0.74 to 0.99; P = .039). Minor allele carriers for SNP Lys940Arg in the poly (ADP-ribose) polymerase 1 (PARP1) gene showed a better response rate to the paclitaxel regimen (45.8%) than to the gemcitabine regimen (10.5%; P for interaction = .019)., Conclusion: Polymorphisms in the TP53 and PARP1 genes are involved in inter-individual differences in the response to platinum-based doublet chemotherapy in patients with NSCLC.

Components of the SWI/SNF chromatin-remodeling complex, such as INI1, are inactivated in human cancer and, thus, act as tumor suppressors. Here we screened for mutations the entire coding sequence of BRG1 (SMARCA4), which encodes the ATPase of the complex, in 59 lung cancer cell lines of the most common histopathological types. Mutations were detected in 24% of the cancer cell lines, many of them in cells commonly used for lung cancer research. All mutations were homozygous and most predicted truncated proteins. The alterations were significantly more frequent in the non-small-cell lung cancer (NSCLC) type (13/37, 35%) as compared to the small-cell lung cancer (SCLC) type (1/19, 5%) (P<0.05; Fisher's Exact test) and BRG1 was the fourth most frequently altered gene in NSCLC cell lines. BRG1 mutations coexisted with mutations/deletions at KRAS, LKB1, NRAS, P16, and P53. However, alterations at BRG1 always occurred in the absence of MYC amplification, suggesting a common role in lung cancer development. In conclusion, our data strongly support that BRG1 is a bona fide tumor suppressor and a major factor in lung tumorigenesis.

Mutations of the epidermal growth factor receptor gene (EGFR) have been reported to be present in a considerable fraction of lung adenocarcinomas showing dramatic response to EGFR tyrosine kinase inhibitors. To clarify pathogenic significance of the mutations for the development of lung adenocarcinoma, we investigated stage I lung adenocarcinomas for the mutations. First, 107 cases of macrodissected stage I adenocarcinomas were examined for mutations in exons 18-21 of the EGFR gene. EGFR mutations were detected in 36 of the 107 cases (34%). In particular, among the stage I cases, the mutations were detected in 17 of 42 small-sized adenocarcinomas (

Genome scanning at a 1-Mb resolution was undertaken in 29 lung cancer cell lines to clarify the distribution of homozygous (i.e., both allele) deletions along lung cancer genomes, using a high-resolution single nucleotide polymorphism array. Eighteen regions, including two known tumor suppressor loci, CDKN2A at 9p21 and FHIT at 3p14, were found homozygously deleted. Frequencies of deletions at the 18 regions were evaluated by genomic polymerase chain reaction in 78 lung cancer cell lines. Seven regions, 2q24, 3p14, 5q11, 9p21, 9p23, 11q14, and 21q21, were homozygously deleted in two or more cell lines. The CDKN2A locus at 9p21 was most frequently deleted (20/78, 26%), and the deletions were detected exclusively in non-small-cell lung carcinomas (NSCLCs). The PTPRD (protein tyrosine phosphatase receptor type D) locus at 9p23 was the second-most frequently deleted (8/78, 10%), and the deletions were detected in both small-cell lung carcinomas (SCLC) and NSCLC. In addition, the 9p24 region was deleted in a NSCLC. In total, 24 (31%) cell lines carried at least one deletion on chromosome arm 9p, while deletions on the remaining chromosome arms were observed at most in four (5%) cell lines. Deletions at 9p24, 9p23, and 9p21 were not contiguous with one another, and preferential co-occurrence or mutual exclusiveness for the deletions at these three loci was not observed. Thus, it was indicated that 9p is the most frequent target of homozygous deletions in lung cancer, suggesting that the arm contains multiple lung tumor suppressor genes and/or genomic features fragile during lung carcinogenesis., ((c) 2005 Wiley-Liss, Inc.)

Background: The detection rate of small nodules in the peripheral lung area is increasing due to the widespread use of CT scanning. However, the radiological and pathological characteristics of very small tumors have not been fully investigated., Methods: We evaluated 44 lung tumors with the size of 1cm or less in diameter resected from 38 patients (19 men and 19 women, with an average of 62 years) from 1997 through 2001. The clinical records, the findings of high-resolution CT (HRCT) and histopathological features of resected specimens were analyzed. Adenocarcinoma was histologically further subclassified into types A to F according to the Noguchi's classification., Results: Lobectomy was performed in 20 patients, wedge resection in 15 and segmentectomy in 3, respectively. Thirty-two tumors were adenocarcinomas, 4 were squamous cell carcinomas, and eight were atypical adenomatous hyperplasia (AAH), respectively. All carcinoma cases were proved to be stage IA. In adenocarcinoma, type A was detected in 12 tumors, type B in 13, type C in 1, type D in 2, type E in 1, and type F in 3, respectively. Most of AAH and type A showed pure ground-glass attenuation on HRCT scan, whereas types B to F as well as squamous cell carcinoma frequently had malignant CT signs such as lobulation and convergence of peripheral vessels. Lymphatic or vascular invasion was observed in two adenocarcinomas (types D and F) and two squamous cell carcinomas, and HRCT scan of these four tumors showed soft-tissue attenuation occupying more than two-thirds of each nodule. All patients are currently alive without signs of recurrence after a mean follow-up period of 35.5 months., Conclusion: Types A and B of adenocarcinoma were the most common histologic types among lung tumors with the size of 1cm or less in diameter. Limited lung resection appears to be an adequate for such small lung tumors in which soft-tissue attenuation consists of less than two-thirds of the nodule on HRCT.

The INK4a/ARF locus on human chromosome band 9p21 carries two tumor suppressor genes, TP14ARF and TP16INK4a, and both are frequently inactivated in nonsmall cell lung carcinoma (NSCLC. TP14ARF and TP16INK4a play important roles in the TP53 and RB tumor suppressor pathways, respectively. To elucidate the genetic and epigenetic status of the TP14ARF and TP16INK4a genes in NSCLC, we comprehensively analyzed mutations, homozygous deletions, methylations in the CpG regions, and expression of the TP14ARF and TP16INK4a genes in 31 NSCLC cell lines. TP16INK4a (84%) was inactivated more frequently than TP14ARF (55%). Moreover, p16INK4a was inactivated in all 17 cell lines with TP14ARF inactivation. Three cell lines with base substitutions in exon 2 resulted in missense mutations of TP16INK4a but silent mutations of TP14ARF. There was a case of mutation in exon 1alpha unique to TP16INK4a, but not a mutation in exon 1beta unique to TP14ARF. The TP16INK4a gene was methylated in 6 cell lines, but the TP14ARF gene was not methylated in any cell line. Unlike a mutually exclusive relationship for inactivation between TP16INK4a and RB, TP14ARF and TP53 did not show such a relationship (P = 0.61, Fisher exact test). Thus, the present results indicate the TP16INK4a gene to be the primary target of INK4a/ARF locus alterations. Transient TP14ARF expression induced G1 arrest in the cells with wild-type TP53, but not in the cells with mutated TP53. Thus, the pathogenetic and biologic significance of TP14ARF inactivation is different between NSCLC cells with wild-type TP53 and those with mutated TP53.

Lung adenocarcinoma has replaced squamous cell lung carcinoma as the most frequent histological subtype in lung cancers. However, genetic factors that affect cancer susceptibility are much less understood in adenocarcinoma than in squamous cell carcinoma. In this study, polymorphisms in five genes involved in the metabolism of carcinogens or in the repair of damaged DNA in lung cells, NQO1-Pro187Ser, GSTT1-positive/null, GSTM1-positive/null, CYP1A1-Ile462Val, and OGG1-Ser326Cys, were examined for association with lung adenocarcinoma risk in a case-control study of 198 patients and 152 control subjects. The NQO1 and GSTT1 polymorphisms were associated with lung adenocarcinoma risk with adjusted odds ratio of 2.15 for the NQO1-Pro/Pro genotype versus the Ser/Ser genotype and adjusted odds ratio of 1.61 for the GSTT1-null genotype versus the positive genotype, respectively. Furthermore, individuals with the combined genotype of NQO1-Pro/Pro and GSTT1-null showed greater risk compared with those of NQO1-Ser/Ser and GSTT1-positive. In contrast, significant association was not observed for the GSTM1, CYP1A1, and OGG1 polymorphisms with lung adenocarcinoma risk, although several studies have shown their implication in the risk for squamous cell lung carcinoma. The result indicates that the NQO1-Pro/Pro and GSTT1-null genotypes are risk factors for lung adenocarcinoma development, and that the genetic factors for susceptibility to adenocarcinoma are different from those to squamous cell carcinoma. The enhanced risk of the NQO1-Pro/Pro genotype combined with the GSTT1-null genotype was more evident in smokers than in nonsmokers. Therefore, carcinogens in tobacco smoke, which are activated by NQO1 and detoxified by GSTT1, could have a role in lung adenocarcinoma development.

Background: The costs of videothoracoscopic procedures for patients with lung carcinoma were compared with those of patients who underwent open thoracotomy in Japan., Methods: The cost of surgical treatment in 1997 and 1998 for patients with resectable primary or metastatic lung carcinoma was analyzed from itemized statements of hospital charges. For patients with lung carcinoma who are in relatively poor health, the authors chose videothoracoscopic lobectomy or partial resection of the lung instead of an open thoracotomy. For patients with solitary pulmonary metastasis, the authors routinely performed thoracoscopic partial resection of the lung., Results: Among 102 patients who were selected for the analysis, 79 patients had primary lung carcinoma, and 23 had metastatic lung carcinoma. Sixty-six open thoracotomies and 36 thoracoscopic surgeries were performed. The mean hospital charge for the entire group of patients was $11,348 (U.S.). The total charges accrued in the operating room amounted to 63% of the hospital charges. Hospitalization was significantly shorter in the group of patients who underwent videothoracoscopy (17.3 days) compared with the group of patients who underwent open thoracotomy (23.8 days). The charges for laboratory examinations, anesthesia, disposable equipment, and hospitalization were significantly higher in patients who underwent open thoracotomy compared with the patients who underwent videothoracoscopy. There were no statistically significant differences in the charges for medication or surgical fees between the two groups., Conclusions: The hospital charges for patients undergoing videothoracoscopic surgery were lower than the charges for patients undergoing open thoracotomy at the authors' hospital. The lower cost was attributable mainly to the less invasive nature of thoracoscopic surgical procedures, which also may be beneficial for decreasing the risk of postsurgical complications in patients in who are in relatively poor health.

To identify genes differentially expressed between small cell lung carcinoma (SCLC) cells and non-SCLC cells, mRNA differential display was applied to 3 SCLC cell lines and 6 non-SCLC cell lines. The LAMB3 gene was identified as being expressed only in non-SCLC cells and not in SCLC cells. The LAMB3 gene encodes the laminin beta3 chain, which is a unique component of laminin-5. Laminin-5 is a heterotrimer protein consisting of the alpha3, beta3, and gamma2 chains, and another unique component of laminin-5 is the gamma2 chain encoded by the LAMC2 gene. RT-PCR analysis of the LAMB3 and LAMC2 genes in 45 lung cancer cell lines revealed that both the LAMB3 and LAMC2 genes were co-expressed in 21 of 32 non-SCLC cell lines (66%) but only in one of 13 SCLC cell lines (8%). Coexpression of the LAMB3 and LAMC2 genes was also observed in all 4 cases of primary non-SCLC cells examined but not in the corresponding non-cancerous lung cells. Since alpha6beta4 integrin, the specific laminin-5 binding receptor, is known to be expressed only in non-SCLC cells and not in SCLC cells, it was indicated that laminin-5 is a critical microenvironmental factor for the growth of non-SCLC cells but not of SCLC cells. The differences in the expression of integrins and laminins would be critical factors to distinguish SCLC and non-SCLC cells, and such differences might be associated with the unique biological properties of SCLC cells, including metastatic potential and drug sensitivity., (Copyright 2000 Academic Press.)

Chromosome 18q is frequently deleted in lung cancers, and a common region of 18q deletions was mapped to chromosome 18q21. Since the DCC candidate tumor suppressor gene has been mapped in this region, mutation and expression of the DCC gene were examined in 46 lung cancer cell lines, consisting of 14 small cell lung carcinomas (SCLCs) and 32 non-small cell lung carcinomas (NSCLCs), to elucidate the pathogenetic significance of DCC alterations in human lung carcinogenesis. A heterozygous missense mutation was detected in a NSCLC cell line, Ma26, while homozygous deletion was not detected in any of the cell lines. The DCC gene was expressed in 11 (24%) of the 46 cell lines, and the incidence of DCC expression was significantly higher in SCLCs (7/14, 50%) than in NSCLCs (4/32, 13%) (P = .01, Fisher's exact test). Therefore, genetic alterations of DCC are infrequent; however, the levels of DCC expression vary among lung cancer cells, in particular, between SCLCs and NSCLCs. The present result does not implicate DCC as a specific mutational target of 18q deletions in human lung cancer; however, it suggests that DCC is a potential target of inactivation by genetic defects including intron or promoter mutations and/or epigenetic alterations. The present result also suggests that DCC expression is associated with some properties of SCLCs, such as a neuroendocrine (NE) feature.

To date, the characteristics of p53 gene mutations in lung cancer have been extensively investigated. However, current estimates of p53 alterations are inaccurate, since most investigators have limited their analyses to exons 5 to 8 of the p53 gene. We examined 52 lung carcinoma cell lines and 106 primary non-small cell lung carcinomas (NSCLC) for mutations in the entire coding region of the p53 gene, from exons 2 to 11. High resolution single strand conformation polymorphism analysis was performed using a modified electrophoretic apparatus with a high concentration gel (14%) and accurate temperature control. The prevalence of mutations was high (more than 80%) in both small cell lung carcinoma (SCLC) (15 of 18) and NSCLC cell lines (28 of 34), and 9 of 45 mutations (20%) were detected outside the region of exons 5 to 8. The frequency of the mutations in primary NSCLC was 48% (51 of 106) and was significantly different (p=0.01) between adenocarcinoma (39%) and squamous cell carcinoma (67%). A-->G transitions (14%, 6 of 43 cases) as well as G-->T transversions (26%, 11 of 43 cases) were frequently detected with significant strand bias in smoking patients, suggesting that carcinogens causing these mutations are involved in smoking associated lung carcinogenesis.