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The eighth edition of the American Joint Committee on Cancer (AJCC) staging system for lung cancer adopts new criteria for tumor size, and for determining pTis, pT1a(mi), and pT1a. The latter is based on the size of stromal invasion. It is quite challenging for lung pathologists.All patients who had undergone surgical resection for pulmonary adenocarcinoma (ADC) at Chung Shan Medical University Hospital between January 2014 and April 2018 were reviewed, and restaged according to the eighth AJCC staging system. The clinical characteristics and survival of patients with tumor stage 0 (pTis), I or II were analyzed.In total, 376 patients were analyzed. None of the pTis, pT1a(mi), or pT1a tumors recurred during the follow-up period up to 5 years, but pT1b, pT1c, pT2a, and pT2b tumors all had a few tumor recurrences (p 0.0001). In addition, 95.2%, 100%, and 77.5% of pTis, pT1a(mi), and pT1a tumors, respectively, had tumor sizes ≤1.0 cm by gross examination. All pTis, pT1a(mi), and pT1a tumors exhibited only lepidic, acinar, or papillary patterns histologically.This study demonstrated excellent survival for lung ADC patients with pTis, pT1a(mi), and pT1a tumors when completely excised. To reduce the inconsistencies between pathologists, staging lung ADC with tumors of ≤1 cm in size grossly as pTis, pT1a(mi), or pT1a may not be necessary when the tumors exhibit only lepidic, acinar, or papillary histological patterns. A larger cohort study with sufficient follow-up data is necessary to support this proposal.

Chemotherapy is the treatment of choice for gastric cancer, but the currently available therapeutic drugs have limited efficacy. Studies have suggested that gastric cancer stem cells may play a key role in drug resistance in chemotherapy. Therefore, new agents that selectively target gastric cancer stem cells in gastric tumors are urgently required. Sirtuin-3 (SIRT3) is a deacetylase that regulates mitochondrial metabolic homeostasis to maintain stemness in glioma stem cells. Targeting the mitochondrial protein SIRT3 may provide a novel therapeutic option for gastric cancer treatment. However, the mechanism by which stemness is regulated through SIRT3 inhibition in gastric cancer remains unknown. We evaluated the stemness inhibition ability of the SIRT3 inhibitor 4′-bromo-resveratrol (4-BR), an analog of resveratrol in human gastric cancer cells. Our results suggested that 4-BR inhibited gastric cancer cell stemness through the SIRT3-c-Jun N-terminal kinase pathway and may aid in gastric cancer stem-cell–targeted therapy.

Background: Methods synthesizing multiple data sources without prospective datasets have been proposed for absolute risk model development. This study proposed methods for adapting risk models for another population without prospective cohorts, which would help alleviate the health disparities caused by advances in absolute risk models. To exemplify, we adapted the lung cancer risk model PLCOM2012, well studied in the west, for Taiwan. Methods: Using Taiwanese multiple data sources, we formed an age-matched case–control study of ever-smokers (AMCCSE), estimated the number of ever-smoking lung cancer patients in 2011–2016 (NESLP2011), and synthesized a dataset resembling the population of cancer-free ever-smokers in 2010 regarding the PLCOM2012 risk factors (SPES2010). The AMCCSE was used to estimate the overall calibration slope, and the requirement that NESLP2011 equals the estimated total risk of individuals in SPES2010 was used to handle the calibration-in-the-large problem. Results: The adapted model PLCOT-1 (PLCOT-2) had an AUC of 0.78 (0.75). They had high performance in calibration and clinical usefulness on subgroups of SPES2010 defined by age and smoking experience. Selecting the same number of individuals for low-dose computed tomography screening using PLCOT-1 (PLCOT-2) would have identified approximately 6% (8%) more lung cancers than the US Preventive Services Task Forces 2021 criteria. Smokers having 40+ pack-years had an average PLCOT-1 (PLCOT-2) risk of 3.8% (2.6%). Conclusions: The adapted PLCOT models had high predictive performance. Impact: The PLCOT models could be used to design lung cancer screening programs in Taiwan. The methods could be applicable to other cancer models.

Background:Methods synthesizing multiple data sources without prospective datasets have been proposed for absolute risk model development. This study proposed methods for adapting risk models for another population without prospective cohorts, which would help alleviate the health disparities caused by advances in absolute risk models. To exemplify, we adapted the lung cancer risk model PLCOM2012, well studied in the west, for Taiwan.Methods:Using Taiwanese multiple data sources, we formed an age-matched case–control study of ever-smokers (AMCCSE), estimated the number of ever-smoking lung cancer patients in 2011–2016 (NESLP2011), and synthesized a dataset resembling the population of cancer-free ever-smokers in 2010 regarding the PLCOM2012 risk factors (SPES2010). The AMCCSE was used to estimate the overall calibration slope, and the requirement that NESLP2011 equals the estimated total risk of individuals in SPES2010 was used to handle the calibration-in-the-large problem.Results:The adapted model PLCOT-1 (PLCOT-2) had an AUC of 0.78 (0.75). They had high performance in calibration and clinical usefulness on subgroups of SPES2010 defined by age and smoking experience. Selecting the same number of individuals for low-dose computed tomography screening using PLCOT-1 (PLCOT-2) would have identified approximately 6% (8%) more lung cancers than the US Preventive Services Task Forces 2021 criteria. Smokers having 40+ pack-years had an average PLCOT-1 (PLCOT-2) risk of 3.8% (2.6%).Conclusions:The adapted PLCOT models had high predictive performance.Impact:The PLCOT models could be used to design lung cancer screening programs in Taiwan. The methods could be applicable to other cancer models.

Supplementary Data from Recalibrating Risk Prediction Models by Synthesizing Data Sources: Adapting the Lung Cancer PLCO Model for Taiwan

Supplementary Figure from Recalibrating Risk Prediction Models by Synthesizing Data Sources: Adapting the Lung Cancer PLCO Model for Taiwan

The standard treatment for early-stage lung cancer is complete tumor excision by limited resection of the lung. Pre-operative localization is used before video-assisted thoracoscopic surgery (VATS) to improve the accuracy of pulmonary nodule excision. However, lung atelectasis and hypoxia resulting from controlling apnea during the localization procedure may affect the localization accuracy. Pre-procedural pulmonary recruitment may improve respiratory mechanics and oxygenation during localization. In this study, we investigated the potential benefits of pre-localization pulmonary recruitment prior to pulmonary ground-glass nodule localization in a hybrid operating room. We hypothesized that pre-localization pulmonary recruitment would increase localization accuracy, improve oxygenation, and prevent the need for re-inflation during the localization procedure. We retrospectively enrolled patients with multiple pulmonary nodule localizations before surgical intervention in our hybrid operating room. We compared localization accuracies between patients who had undergone pre-procedure pulmonary recruitment and patients who had not. Saturation, re-inflation rate, apnea time, procedure-related pneumothorax, and procedure time were also recorded as secondary outcomes. Patients who had undergone pre-procedure recruitment had better saturation, shorter procedure time, and higher localization accuracy. The pre-procedure pulmonary recruitment maneuver was effective in increasing regional lung ventilation, leading to improved oxygenation and localization accuracy.

Supplementary Figure 2 PDF file - 132K, (A) The expression levels of MnSOD evaluated by Western blotting were not changed by FoxM1-knockdown in A549 and H1355 cells. (B) The expression levels of MMP1 and MMP2 evaluated by Western blotting decreased in a dose-dependent manner; however, the change of MMP7 and MMP9 expression was not observed in MnSOD-knockdown A549 cells

Supplementary Figure 1 PDF file - 101K, The expression level of E2F1 evaluated by Western blotting was not changed by MnSOD-knockdown in H1355 cells compared with non-specific shRNA control cells

Supplementary Table 1 PDF file - 142K, Relationship of MnSOD IHC, FoxM1, MMP2 and MnSOD mRNA expression with clinical parameters in lung adenocarcinoma patients

Supplementary Table 2 PDF file - 144K, List of primer sequences (5'�|��3') and their reaction conditions used in the present study

Supplementary Data PDF file - 268K,Supplementary Figure 1. The expression level of E2F1 evaluated by Western blotting was not changed by MnSOD-knockdown in H1355 cells compared with non-specific shRNA control cells. Supplementary figure 2. (A) The expression levels of MnSOD evaluated by Western blotting were not changed by FoxM1-knockdown in A549 and H1355 cells. (B) The expression levels of MMP1 and MMP2 evaluated by Western blotting decreased in a dose-dependent manner; however, the change of MMP7 and MMP9 expression was not observed in MnSOD-knockdown A549 cells. Supplementary Table 1. Relationship of MnSOD IHC, FoxM1, MMP2 and MnSOD mRNA expression with clinical parameters in lung adenocarcinoma patients. Supplementary Table 2. List of primer sequences (5'�|��3') and their reaction conditions used in the present study

Manganese superoxide dismutase (MnSOD) is an antioxidant enzyme responsible for the elimination of superoxide radical. The role of MnSOD in tumor progression in different human cancers is still controversial. In the present study, MnSOD expression in lung cancer cells was explored by knockdown or overexpression using transfection of a short hairpin RNA (shRNA) or an expression vector, respectively, to determine whether MnSOD expression mediates lung cancer cell migration, invasion, and oncogenic potential by increasing FoxM1 and MMP2 expression. Western blotting showed that FoxM1 and MMP2 expression was dependent on MnSOD expression, suggesting that FoxM1 could be upregulated by MnSOD. Three FoxM1 promoters were constructed to verify this activation of FoxM1 by MnSOD and to determine the transcription factors responsible. Luciferase reporter and chromatin immunoprecipitation assays indicated that MnSOD overexpression in lung cancer cells promoted binding of E2F1 and Sp1 to their putative FoxM1 promoter-binding sites and activated FoxM1 reporter activity. MnSOD also enhanced the potential for cell migration, invasion, and anchorage-independent colony growth on soft-agar plates, again via upregulation of FoxM1 and MMP2 expression. In patients with lung cancer, evaluation of MnSOD expression in lung tumors by immunohistochemistry indicated a positive correlation between FoxM1 and MMP2 mRNA expressions. Kaplan–Meier and Cox regression analysis revealed a poorer overall survival (OS) and relapse-free survival (RFS) in patients with MnSOD-positive tumors than with MnSOD-negative tumors. We conclude that MnSOD may promote tumor aggressiveness via upregulation of the FoxM1–MMP2 axis, and that MnSOD expression can independently predict survival and relapse in patients with resected lung adenocarcinoma. Mol Cancer Res; 11(3); 261–71. ©2012 AACR.

Supplementary Tables 1-2 from Identification of α-Enolase as an Autoantigen in Lung Cancer: Its Overexpression Is Associated with Clinical Outcomes

Purpose: Although existence of humoral immunity has been previously shown in malignant pleural effusions, only a limited number of immunogenic tumor-associated antigens (TAA) have been identified and associated with lung cancer. In this study, we intended to identify more TAAs in pleural effusion–derived tumor cells.Experimental Design: Using morphologically normal lung tissues as a control lysate in Western blotting analyses, 54 tumor samples were screened with autologous effusion antibodies. Biochemical purification and mass spectrometric identification of TAAs were done using established effusion tumor cell lines as antigen sources. We identified a p48 antigen as α-enolase (ENO1). Semiquantitative immunohistochemistry was used to evaluate expression status of ENO1 in the tissue samples of 80 patients with non–small cell lung cancer (NSCLC) and then correlated with clinical variables.Results: Using ENO1-specifc antiserum, up-regulation of ENO1 expression in effusion tumor cells from 11 of 17 patients was clearly observed compared with human normal lung primary epithelial and non-cancer-associated effusion cells. Immunohistochemical studies consistently showed high level of ENO1 expression in all the tumors we have examined thus far. Log-rank and Cox's analyses of ENO1 expression status revealed that its expression level in primary tumors was a key factor contributing to overall- and progression-free survivals of patients (P < 0.05). The same result was also obtained in the early stage of NSCLC patients, showing that tumors expressing relatively higher ENO1 level were tightly correlated with poorer survival outcomes.Conclusions: Our data strongly support a prognostic role of ENO1 in determining tumor malignancy of patients with NSCLC.

Supplementary Figure Legends - PDF file 57K, Supplementary Figure Legends

Supplementary Figure 1 - PDF file 36K, Doubling time of IL-10-knockdown TL1 cells and IL-10-overexpressing TL4 cells and the error bars represent the S.D. of triplicate results

Supplementary Figure 4 from Identification of α-Enolase as an Autoantigen in Lung Cancer: Its Overexpression Is Associated with Clinical Outcomes

Supplementary Fig. 1. PAK1 confers erlotinib resistance in lung adenocarcinoma cells regardless of EGFR mutation Supplementary Fig. 2. The optimal concentration of gefitinib was obtained from the dose-dependent experiments Supplementary Fig. 3. PAK1 transcription is regulated by the PI3K/AKT/C/EBP-β /miR-145 cascades and PAK1 targeted by miR-145 may confer PAK1-mediated TKI resistance in EGFR-wild-type cells Supplementary Fig. 4. PAK1 modulates Mcl-1 protein stability via phosphorylation of Mcl-1 at Ser-159. Supplementary Fig. 5. The association of PAK1 and E-cadherin mRNA expression with overall survival (OS) and relapse free survival (RFS) in lung adenocarcinoma patients. Supplementary Table 1. Relationships of PAK1 and E-cadherin mRNA levels with clinico-pathological parameters, and the association between PAK1 and E-caherin mRNA levels in lung adenocarcinoma patients. Supplementary Table 2. The association of PAK1 and E-cadherin mRNA level with the response to TKI therapy in lung adenocarcinoma patients. Supplementary Table 3. Cox regression analysis for the prognostic value of PAK1 and E-cadherin mRNA levels on OS and RFS in lung adenocarcinoma patients.

Supplementary Figure 2 from Identification of α-Enolase as an Autoantigen in Lung Cancer: Its Overexpression Is Associated with Clinical Outcomes

Supplementary Figure 2 - PDF file 65K, Kaplan-Meier analysis was used to assess the influence of IL-10 and CIP2A on overall survival in (A) all study population, (B) patients with HPV 16/18 E6 positive tumor, and (C) patients with HPV 16/18 E6 negative tumor

PDF file - 430K

Supplementary Figures S1-S4; Supplementary Tables S1-S3.

Purpose: To assess whether polymorphisms of genes related to estrogen biosynthesis and metabolism are associated with EGFR mutations.Experimental Design: We studied 617 patients with lung adenocarcinoma, including 302 never-smoking women. On the basis of multiple candidate genes approach, the effects of polymorphisms of CYP17, CYP19A1, ERα, and COMT in association with the occurrence of EGFR mutations were evaluated using logistic regression analysis.Results: In female never-smokers, significant associations with EGFR L858R mutation were found for the tetranucleotide (TTTA)n repeats in CYP19A1 (odds ratio, 2.6; 95%CI, 1.2–5.7 for 1 or 2 alleles with (TTTA)n repeats >7 compared with both alleles with (TTTA)n repeats ≤7), and the rs2234693 in ERα (OR, 2.1; 95% CI, 1.1–4.0 for C/T and C/C genotypes compared with T/T genotype). The C/C genotype (vs. T/T genotype) of ERα was significantly associated with EGFR L858R mutation (OR, 3.0; 95% CI, 1.1–8.1), in-frame deletion (OR, 2.9; 95% CI, 1.1–7.6) and other mutations (OR, 4.3; 95% CI, 1.3–14.0). The genotype of COMT rs4680 was significantly associated with EGFR L858R mutation in female and male never-smokers showing OR's (95% CI) of 1.8 (1.0–3.2) and 3.6 (1.1–11.3), respectively, for genotypes G/A and G/G compared with genotype A/A. The number of risk alleles of CYP17, CYP19A1, ERα, and COMT was associated with an increasing OR of EGFR L858R mutation in female never-smokers (P = 0.0002 for trend).Conclusions: The L858R mutation of EGFR is associated with polymorphisms of genes related to estrogen biosynthesis and metabolism in never-smoking female lung adenocarcinoma patients. Clin Cancer Res; 17(8); 2149–58. ©2011 AACR.

Supplementary Figures 3 and 4 - PDF file 262K, S3: Exogenous IL-10 regulates tumor migration capability of lung cancer cells; S4: The migration and invasion ability of TL1 cells was decreased in IL-10R knock down stable clone

Supplementary Figure Legends 1-4 from Identification of α-Enolase as an Autoantigen in Lung Cancer: Its Overexpression Is Associated with Clinical Outcomes

Supplementary Methods; Supplementary Table S1.

Purpose: Interleukin-10 (IL-10) determines virus persistent infection and promotes viral-associated tumor progression via tumor immune escape. However, the role of IL-10 in tumor progression and prognosis in lung adenocarcinoma remains controversial.Experimental Design: To investigate how IL-10 is regulated by HPV E6, IL-10 promoter was constructed to understand which transcriptional factor could be responsible for its transcription. To verify which molecule could be responsible for IL-10–mediated soft agar growth and invasion capability, PCR array and mechanistic strategies were conducted. IL-10 and CIP2A mRNA levels in lung tumors from patients with lung cancer were determined by real-time reverse transcription PCR. The prognostic value of both molecules on survival was estimated by Cox regression model.Results: Mechanistic studies showed that IL-10 protein and mRNA expression was decreased in E6 knockdown TL1 cells and increased in E6- overexpressing TL4 cells. In addition, IL-10 transcription was predominantly regulated by E6-mediated phosphorylation of cAMP response element-binding protein (CREB) and C/enhancer-binding protein β (C/EBPβ) via phosphoinositide 3-kinase (PI3K) signaling pathway. IL-10–mediated tumor aggressiveness in vitro and in vivo occurs through increased CIP2A expression via PI3K signaling pathway. Among patients, IL-10 mRNA expression in lung tumors was positively correlated with CIP2A mRNA expression. Cox-regression analysis showed that IL-10 and CIP2A mRNA levels may independently predict survival in patients with lung adenocarcinoma, especially in patients with E6-positive tumors.Conclusion: IL-10 production from lung tumors and immune cells promotes lung adenocarcinoma aggressiveness and patients with poor survival. We thus suggest that PI3K inhibitor combined with chemotherapy may potentially enhance tumor regression and improve patients’ outcome and life quality. Clin Cancer Res; 19(15); 4092–103. ©2013 AACR.

Purpose: EGFR mutation as a biomarker has documented that EGFR-mutant patients will derive clinical benefit from tyrosine kinase inhibitor (TKI) treatment. Unfortunately, most patients show TKI resistance and tumor recurrence after therapy. Therefore, we expected that an adjuvant biomarker other than EGFR mutation is needed for predicting TKI resistance.Experimental Design: Molecular manipulations were performed to verify whether TKI resistance mediated by p21-activated kinase (PAK1) could be through increasing Mcl-1 protein stability via the PI3K/AKT/C/EBP-β/miR-145 cascade. Xenograft mouse models were used to confirm the mechanistic action of PAK1 on TKI resistance. Forty-six tumor tissues from patients with lung adenocarcinoma who received TKI therapy were collected to evaluate PAK1 and E-cadherin mRNA expressions by real-time PCR. The association of PAK1 and E-cadherin mRNA expressions with tumor response to TKI treatment and outcomes was evaluated.Results: We demonstrate that PAK1 confers TKI resistance in EGFR-mutant cells as well as in EGFR–wild-type cells. Mechanistically, the positive feedback loop of PAK1/PI3K/AKT/C/EBP-β/miR-145 cascades persistently activates the PI3K/AKT signaling pathway to protect Mcl-1 degradation by Fbw7, which results, in turn, in TKI resistance and cell invasion via epithelial-to-mesenchymal transition due to a decrease in E-cadherin expression. The mechanism underlying the cell model is further confirmed in xenograft tumors. Among patients, high-PAK1 or low–E-cadherin tumors more commonly exhibited an unfavorable response to TKI and poorer outcome compared with low-PAK1 or low–E-cadherin tumors.Conclusions: The combination of TKI with AKT inhibitor might confer TKI sensitivity and in turn improve outcomes in patients with lung adenocarcinoma who harbored high PAK1 mRNA–expressing tumors. Clin Cancer Res; 22(21); 5370–82. ©2016 AACR.

Purpose:Fas ligand (FasL) −844T/C polymorphism (rs763110) has a demonstrated association with lung cancer risk. FasL −844CC with higher FasL expression has been suggested to contribute to tumor progression via immune escape. However, the impact of FasL −844T/C polymorphism on the clinical outcome of non–small cell lung cancer (NSCLC) remains to be identified.Experimental Design: A total of 385 adjacent normal lung tissues from patients with NSCLC were collected to determine FasL −844T/C polymorphism by PCR-based restriction fragment length polymorphism. FasL mRNA and protein expression in lung tumors were evaluated by real-time PCR and immunohistochemistry. The prognostic value of FasL −844T/C polymorphism on survival and relapse was determined by Kaplan–Meier analysis and Cox proportional hazards models.Results: The FasL −844CC genotype had higher prevalence in those with advanced tumors than in those with early tumors (P = 0.008). In addition, patients with the FasL −844CC genotype were more prone to tumor relapse than those with the FasL −844TT+TC genotype (62.1% vs. 37.9%, P = 0.001). Multivariate Cox regression analysis showed that patients with the FasL −844CC genotype had poorer survival in terms of overall survival (OS) and relapse-free survival (RFS) than those with the FasL −844TT+TC genotype (24.1 vs. 42.8 months for OS, HR = 1.455, P = 0.004; 15.4 vs. 31.4 months for RFS, HR = 1.710, P < 0.001).Conclusions:FasL −844T/C polymorphism may predict survival and relapse in NSCLC. We suggest that FasL may be a molecular target for immunotherapeutic interventions to improve the clinical outcome of patients with NSCLC. This finding should be validated by another investigative group. Clin Cancer Res; 17(18); 5991–9. ©2011 AACR.

Supplementary Figure 1 from Identification of α-Enolase as an Autoantigen in Lung Cancer: Its Overexpression Is Associated with Clinical Outcomes

Supplementary Figure 3 from Identification of α-Enolase as an Autoantigen in Lung Cancer: Its Overexpression Is Associated with Clinical Outcomes

Supplementary Figures 1-2 from Human Papillomavirus 16/18 E6 Oncoprotein Is Expressed in Lung Cancer and Related with p53 Inactivation

Supplementary Methods, Figure 1, Tables 1-9 from Paxillin Predicts Survival and Relapse in Non–Small Cell Lung Cancer by MicroRNA-218 Targeting