Type Lung Adenocarcinoma Research Articles

Ferroptosis-related gene signature for predicting prognosis and identifying potential therapeutic drug in EGFR wild-type lung adenocarcinoma

Epidermal growth factor receptor wild type lung adenocarcinoma (EGFRWT LUAD) still has limited treatment options and unsatisfactory clinical outcomes. Ferroptosis, as a form of cell death, has been reported to play a dual role in regulating tumor cell survival. In this study, we constructed a 3-ferroptosis-gene signature, FeSig, and verified its accuracy and efficacy in predicting EGFRWT LUAD prognosis at both the RNA and protein levels. Patients with higher FeSig scores were found to have worse clinical outcomes. Additionally, we explored the relationship between FeSig and tumor microenvironment, revealing that enhanced interactions between fibroblasts and tumor cells in FeSighigh patients causing tumor resistance to ferroptosis. To address this challenge, we screened potential drugs from NCI-60 (The US National Cancer Institute 60 human tumour cell line anticancer drug screen) and Connectivity map database, ultimately identifying 6-mercatopurine (6-MP) as a promising candidate. Both in vitro and in vivo experiments demonstrated its efficacy in treating FeSighigh EGFRWT LUAD tumor models. In summary, we develop a novel FeSig for predicting prognosis and guiding drug application.

Computed tomography radiomics study of invasion and instability of lung adenocarcinoma manifesting as ground glass nodule.

Ground-glass nodule (GGN) is the most common manifestation of lung adenocarcinoma on computed tomography (CT). Clinically, the success rate of preoperative diagnosis of GGN by puncture biopsy and other means is still low. The aim of this study is to investigate the clinical and radiomics characteristics of lung adenocarcinoma presenting as GGN on CT images using radiomics analysis methods, establish a radiomics model, and predict the classification of pathological tissue and instability of GGN type lung adenocarcinoma. This study retrospectively collected 249 patients with 298 GGN lesions who were pathologically confirmed of having lung adenocarcinoma. The images were imported into the Siemens scientific research prototype software to outline the region of interest and extract the radiomics features. Logistic model A (a radiomics model to identify the infiltration of lung adenocarcinoma manifesting as GGNs) was established using features after the dimensionality reduction process. The receiver operating characteristic (ROC) curve of the model on training set and the verification set was drawn, and the area under the curve (AUC) was calculated. Second, a total of 112 lesions were selected from 298 lesions originating from CT images of at least two occasions, and the time between the first CT and the preoperative CT was defined as not less than 90 days. The mass doubling time (MDT) of all lesions was calculated. According to the different MDT diagnostic thresholds instability was predicted. Finally, their AUCs were calculated and compared. There were statistically significant differences in age and lesion location distribution between the "noninvasive" lesion group and the invasive lesion group (P<0.05), but there were no statistically significant differences in sex (P>0.05). Model A had an AUC of 0.89, sensitivity of 0.75, and specificity of 0.86 in the training set and an AUC of 0.87, sensitivity of 0.63, and specificity of 0.90 in the validation set. There was no significant difference statistically in MDT between "noninvasive" lesions and invasive lesions (P>0.05). The AUCs of radiomics models B1, B2 and B3 were 0.89, 0.80, and 0.81, respectively; the sensitivities were 0.71, 0.54, and 0.76, respectively; the specificities were 0.83, 0.77, and 0.60, respectively; and the accuracies were 0.78, 0.65, and 0.69, respectively. There were statistically significant differences in age and location of lesions between the "noninvasive" lesion group and the invasive lesion group. The radiomics model can predict the invasiveness of lung adenocarcinoma manifesting as GGNs. There was no significant difference in MDT between "noninvasive" lesions and invasive lesions. The radiomics model can predict the instability of lung adenocarcinoma manifesting as GGN. When the threshold of MDT was set at 813 days, the model had higher specificity, accuracy, and diagnostic efficiency.

Retrospective analysis of change in frequency of STK11 mutation in lung adenocarcinomas over a 10-year period.

8032 Background: Previous studies have linked the presence of inactivating mutations in the tumor suppressor gene STK11 with reduced efficacy of immune checkpoint inhibitor (ICI) treatments in lung adenocarcinoma (LA). We queried whether there has been a change in the STK11 mutation frequency over the last 10-year period that could potentially reflect the emergence of additional acquired resistance mutations. Methods: Over a 10-year period from 2013 through 2022, 109,763 clinically advanced LA underwent hybrid capture based comprehensive genomic profiling (CGP) to assess all classes of genomic alterations (GA) at a single reference laboratory (Foundation Medicine). MSI-high status and tumor mutational burden (TMB) were determined from the sequencing data. PD-L1 status was measured by immunohistochemistry (IHC) using the Dako 22C3 kit with TPS scoring. Statistical comparisons utilized the 2-tailed Chi Square method with the Yates’ correction. Results: The frequency of STK11 inactivating GA in all cases of LA significantly increased during the study period from 2013 to 2022 (16.1% vs 20.1%; p=0.0005). This increase appeared to be predominantly in KRAS wild type LA (10.1% vs 15.5%; p&lt;0.0001). The STK11 GA frequency slightly declined in the KRAS mutated LA over the 10-year period (29.9% vs o 27.0%, P=0.25). During the same time period, the KEAP1 GA frequency increased from 12.6% to 14.9% (p= 0.076) and the frequency of combined STK11 and KEAP1 GA increased from 4.8% to 8.0% (p=.0006). The frequencies of MSI-high status, TMB and PD-L1 expression did not significantly change over the observation period. Conclusions: The continuously increasing approved indications for ICI-based treatments for both early stage and clinically advanced LA appears to be accompanied by a significant increase in the frequency of inactivating STK11 GA predominantly in the KRAS mutation negative LA population. This inactivation is associated with an attenuated response rate and progression-free survival in this population. Given the development of novel therapies focused on potential restoration of STK11 function by HDAC/CoREST inhibition, further study of the STK11 GA distribution in LA appears warranted. [Table: see text]

Expression patterns of HNF4α, TTF-1, and SMARCA4 in lung adenocarcinomas: impacts on clinicopathological and genetic features.

HNF4α expression and SMARCA4 loss were thought to be features of non-terminal respiratory unit (TRU)-type lung adenocarcinomas, but their relationships remained unclear. HNF4α-positive cases among 241 lung adenocarcinomas were stratified based on TTF-1 and SMARCA4 expressions, histological subtypes, and driver mutations. Immunohistochemical analysis was performed using xenograft tumors of lung adenocarcinoma cell lines with high HNF4A expression. HNF4α-positive adenocarcinomas(n = 33) were divided into two groups: the variant group(15 mucinous, 2 enteric, and 1 colloid), where SMARCA4 was retained in all cases, and the conventional non-mucinous group(6 papillary, 5 solid, and 4 acinar), where SMARCA4 was lost in 3/15 cases(20%). All variant cases were negative for TTF-1 and showed wild-type EGFR and frequent KRAS mutations(10/18, 56%). The non-mucinous group was further divided into two groups: TRU-type(n = 7), which was positive for TTF-1 and showed predominantly papillary histology(6/7, 86%) and EGFR mutations(3/7, 43%), and non-TRU-type(n = 8), which was negative for TTF-1, showed frequent loss of SMARCA4(2/8, 25%) and predominantly solid histology(4/8, 50%), and never harbored EGFR mutations. Survival analysis of 230 cases based on histological grading and HNF4α expression revealed that HNF4α-positive poorly differentiated (grade 3) adenocarcinoma showed the worst prognosis. Among 39 cell lines, A549 showed the highest level of HNF4A, immunohistochemically HNF4α expression positive and SMARCA4 lost, and exhibited non-mucinous, high-grade morphology in xenograft tumors. HNF4α-positive non-mucinous adenocarcinomas included TRU-type and non-TRU-type cases; the latter tended to exhibit the high-grade phenotype with frequent loss of SMARCA4, and A549 was a representative cell line.

A rare case of synchronous thymoma type AB and lung adenocarcinoma in a non-smoker middle-aged woman

A rare case of synchronous thymoma type AB and lung adenocarcinoma in a non-smoker middle-aged woman

Survival benefit in EGFR-wild and ALK negative NSCLC patients who participate in clinical trials compared to standard-of-care: Propensity-matched analysis

ObjectivesAdvanced non-small cell lung cancer patients harboring EGFR mutation or ALK fusion have achieved significant survival benefit with targeted agents. In contrast, EGFR-wild type and ALK negative lung adenocarcinoma still have poor survival outcome. This study assessed the impact of participating in clinical trials on clinical outcomes in patients with EGFR-wild-type and ALK-negative lung adenocarcinoma. Materials and methodsThis study included patients with advanced EGFR-wild-type and ALK-negative lung adenocarcinoma who received systemic treatment between March 2017 and June 2022. We compared clinical outcomes between patients who participated in clinical trials and those treated with standard-of-care (SOC) using propensity score matching (PSM). ResultsOverall, 1,686 patients with EGFR-wild-type and ALK-negative advanced lung adenocarcinoma were included in the final analysis. Of these, 1,380 (81.9 %) received SOC only and 306 (18.1 %) patients were enrolled in at least one clinical trial during their cancer journey. After PSM (1:1), 612 patients were matched to the SOC (n = 306) and clinical trial (n = 306) groups. Among those who participated in clinical trials, 27.8 % and 72.2 % were included in clinical trials involving targeted therapy and immunotherapy respectively. In the clinical trial group, more patients received targeted therapy (31.7 % vs. 5.5 %, p < 0.001) and immunotherapy (88.6 % vs. 62.8 %, p < 0.001) compared to the SOC group. The median overall survival was 17.1 months (95 % confidence interval [CI], 13.2–21.4) in the SOC group and 27.3 months (95 % CI, 22.1–32.4) in the clinical trial group (hazard ratio = 0.71, [95 % CI, 0.58–0.88, P = 0.002]). ConclusionsThis study demonstrated that participating in clinical trials resulted in a survival benefit that reduced the risk of death by 29.6% compared to receiving SOC in EGFR-wild-type and ALK-negative lung adenocarcinoma.

EP07.04-11 Clinicopathological Features and Postoperative Prognosis of Bubble-like Appearance Type Lung Adenocarcinoma

EP07.04-11 Clinicopathological Features and Postoperative Prognosis of Bubble-like Appearance Type Lung Adenocarcinoma

Clinical trial versus standard of care in EGFR-wild type and ALK- negative lung adenocarcinoma: A propensity-matched analysis.

e21116 Background: Clinical trials have been a vital role for survival improvement in recurrent or metastatic lung cancer. Recent advances in targeted therapy for EGFR-mutant or ALK-positive lung adenocarcinoma dramatically increased survival. However, EGFR-wild type and ALK negative lung adenocarcinoma still had relatively poor survival outcome. Clinical trial of targeted therapy for rare driver mutation or immunotherapy was introduced in 2015. Moreover, next-generation sequencing was reimbursed in 2017 in Korea. This study evaluated the benefit of clinical trials for EGFR-wild type and ALK negative lung adenocarcinoma in recent 5 years. Methods: This study included patients with recurrent or metastatic lung adenocarcinoma who received systemic treatment between March 2017 and June 2022 at Samsung Medical Center. We compared overall survival in group with clinical trial and the standard of care (SOC) by using propensity score matching (PSM) (1:1). We used cox proportional hazard models to calculate the hazard ratio (HR) and 95% confidence interval while accounting for clustering among matched pairs. Results: Of total, 1,686 EGFR-wild type and ALK negative lung adenocarcinoma were included for final analysis. Among them, 1,380 received SOC only and 306 patients enrolled at least one clinical trial. In a group of clinical trial, 41.5%, 29.1%, and 23,2% of patients were enrolled for first-, second-, and third-line treatment, respectively. To achieve the balance between the SOC group and the clinical trial group, 8 baseline characteristics (age, sex, smoking history, ECOG performance status, brain metastasis, liver metastasis, bone metastasis, and rare driver mutation) were included. After PSM, 612 patients were matched to group of the SOC (n = 306) and the group of clinical trial (n = 306). Among the clinical trial, 27.8% was a clinical trial of targeted therapy and 72.2% was a clinical trial of immunotherapy or others. Median follow-up duration was 32.8 months (range, 5.0–64.4). Median overall survival was 17.1 months (95% CI, 13.2–21.4) for SOC and 27.3 months (95% CI, 22.1–32.4) for group of clinical trial (HR = 0.71 (95% CI, 0.58–0.88, P = 0.002). Conclusions: This study showed the performance of clinical trials for patients with EGFR-wild type and ALK-negative lung adenocarcinoma at tertiary hospital. After PSM, clinical trials showed survival benefits that reduced the risk of death by 29% in patients with EGFR-wild type and ALK-negative lung adenocarcinoma.

Revealing distinct immune landscape in EGFR mutant and EGFR wild type lung adenocarcinoma by using single-cell RNA sequencing.

e21008 Background: Non-small cell lung cancer (NSCLC) with EGFR mutations displays heterogeneity in the extent of intratumoral immunocytes infiltration compared to those with EGFR wild types, which may explain their variable responsiveness to PD-1 blockade. Here, we performed single-cell RNA sequencing (scRNA-seq) to unveil the immunescape and heterogeneity of lung adenocarcinoma (LUAD) between EGFR mutant (EGFR-mut) patients and wild type (EGFR-wt) patients. Methods: Eleven EGFR-mut and 7 EGFR-wt NSCLC tumor samples were analyzed. Immune cells (CD45+) were enriched through fluorescence-activated cell sorting (FACS) and subjected to single-cell RNA sequencing (scRNA-seq) using the 10X Genomics platform. Sequencing reads were normalized and analyzed using the R/Seurat package. Cellular components of each sample were determined based on known marker genes. Wilcox test was used to compare promotion by subtype composition and immune cell infiltrates. Results: Different immune cell compositions were observed between EGFR-mut and EGFR-wt LUAD tumor samples. We found EGFR-wt tumors are marked by an enrichment of CCL15+ tumor-associated macrophages(TAMs), which display pro-inflammatory properties and high expressions of TNF-a-signaling genes. In contrast, EGFR-mut tumors exhibit enrichment of SPARCL1+TAMs, which are involved in epithelial-mesenchymal transition (EMT) and angiogenesis. EGFR-wt tumors exhibited enrichment of CD8-Teff-MX1 and Gamma delta T cells, both of which displayed higher cytotoxic and naive features. Moreover, the high expression of Gamma delta T corresponds with a better clinical prognosis. In contrast, CD8-Teff-XCL1 and NKT-FGFBP2 were found enriched in EGFR-mut tumors, both of which contributed to higher exhaustion and decreased cytotoxic function. Additionally, NK-IL32 was found enriched in EGFR-mut tumors, showing a downregulated T cell receptor signaling and stronger exhausted features. The high expression of this subcluster correlated with poor outcomes in LUAD patients harboring EGFR mutation. Conclusions: Our study provided a deeper understanding of the immune landscape of EGFR-mut and EGFR-wt LUAD and gained better insights into the immunological mechanisms involved in the response to ICIs. The results of our study could be instructive for the development of future immunotherapy strategies for NSCLC patients harboring EGFR mutations.

Various impacts of driver mutations on the PD-L1 expression of NSCLC.

We aimed to evaluate whether different driver mutations have varying impacts on the programmed cell death-ligand 1 (PD-L1) expression of non-small cell lung cancer (NSCLC), and whether the prognostic roles of PD-L1 amongst our patients were divergent. This was a single-institute study that included patients with NSCLC. Six driver mutations, PD-L1 status, and the outcomes of treatment were assessed. A total of 1,001 NSCLC patients were included for analysis. Overall, the PD-L1 positive (TPS ≥ 1%) and strong positive (TPS ≥ 50%) rates were 52.2% and 17.3%, respectively. As compared with wild type lung adenocarcinoma, EGFR-mutant and HER2-mutant patients had similarly low PD-L1 and strong PD-L1 positive rates. BRAF-mutant patients had numerically higher PD-L1 and strong PD-L1 positive rates. Patients with fusion mutation (ALK and ROS1) (aOR 2.32 [95% CI 1.10–4.88], P = 0.027 and 2.33 [95% CI 1.11–4.89], P = 0.026), KRAS mutation (aOR 2.58 [95% CI 1.16–5.75], P = 0.020 and 2.44 [95% CI 1.11–5.35], P = 0.026), and non-adenocarcinoma histology (aOR 2.73 [95% CI 1.72–4.34], P < 0.001 and 1.93 [95% CI 1.13–3.30], P = 0.016) all had significantly higher PD-L1 and strong PD-L1 positive rates. A trend towards longer survival was noted in ROS-1 rearranged and KRAS-mutant patients with strong PD-L1 expression who had received crizotinib and chemotherapy, respectively. In conclusion, individual driver mutations had various impacts on the PD-L1 expression of NSCLC patients. The prognostic role of PD-L1 may also be divergent amongst patients harboring different driver mutations.

Impact of LncRNA GAS5 Genetic Variants and the Epidermal Growth Factor Receptor Phenotypes on the Clinicopathological Characteristics of Lung Adenocarcinoma Patients.

The aim of the current study was to evaluate the combined effect of the single nucleotide polymorphism (SNP) in long non-coding RNA growth arrest-specific 5 (GAS5) and the phenotypes of epidermal growth factor receptor (EGFR) on the clinicopathological characteristics of lung adenocarcinoma. The present study examined the relationship between the GAS5 single-nucleotide polymorphisms (SNPs; rs145204276 Ins/Del, rs55829688 T/C) and the clinicopathological factors in 539 lung adenocarcinoma patients with or without EGFR mutations. We found that the genotype distributions of the two GAS5 SNPs between different EGFR genotypes were similar after adjusting for age, gender and smoking history. The GAS5 SNP rs145204276 Ins/Del + Del/Del illustrated a higher distribution with an advanced tumor stage (p = 0.030), larger tumor T status (p = 0.019), positive lymph node status (p = 0.014) and distal metastases (p = 0.011) in the EGFR wild type group. In the subgroup analysis of the EGFR wild type population, the presence of GAS5 SNP rs145204276 Ins/Del + Del/Del was correlated to an advanced tumor stage (p = 0.014) and distal metastases (p = 0.020) in non-smokers. In conclusion, these data indicate that the GAS5 SNP rs145204276 variant may help predict tumor stage, lymph node metastasis and distal metastases in patients with EGFR wild type lung adenocarcinoma.

Clinical Characteristics and Risk Factors Analysis for Visceral Pleural Invasion in Mixed Ground-glass Nodular Lung Adenocarcinoma

背景与目的目前国内肺癌仍是发病率和致死率最高的恶性肿瘤，肺腺癌是其最常见的亚型，影像学中表现为混合性磨玻璃结节(mixed ground glass nodule, mGGN)的肺癌逐渐增多。脏层胸膜侵犯(visceral pleural invasion, VPI)是影响mGGN型肺腺癌预后的重要因素。本研究旨在探索分析mGGN型肺腺癌发生VPI的危险因素。方法回顾性分析2016年11月-2019年11月南京医科大学第一附属医院收治的接受根治性手术的128例mGGN型肺腺癌患者的临床资料，包括影像、病理和生物学特征，其中男性40例、女性88例，年龄30岁-81(60.3±9.3)岁。采用单因素卡方检验与多因素Logistic回归分析调查mGGN型肺腺癌发生VPI的各项危险因素。结果符合纳入标准的128例mGGN型肺腺癌患者中57例被诊断有VPI发生。性别、实性成分最大径、实性成分比例(consolidation tumor ratio, CTR)、毛刺征、肺部疾病史、高血压家族史、肿瘤-胸膜空间位置分型(relation of lesion to pleura, RLP)、支气管与结节走行关系在VPI(+)组和VPI(-)组间的差异均有统计学意义(P < 0.05)。Logistic多因素回归分析发现RLP(OR=3.529, 95%CI: 1.430-8.713, P=0.006)和支气管与结节走行关系(OR=3.993, 95%CI: 1.517-10.51, P=0.005)是VPI发生的独立危险因素(P < 0.05)。结论临床诊治应综合上述参数评估mGGN型肺腺癌发生VPI的可能性。RLP和支气管与结节走行关系异常作为VPI的独立危险因素，对判断mGGN型肺腺癌发生VPI具有一定的指导意义。

Surgical Extent for Ground Glass Nodules.

As diagnoses of small ground glass nodule (GGN)-type lung adenocarcinoma are increasing due to the increasing frequency of computed tomography (CT) screening, surgical treatment for GGN-type lung adenocarcinoma has rapidly become more common. However, the appropriate surgical extent for these lesions remains unclear; therefore, several retrospective studies have been published and prospectively randomized controlled trials are being undertaken. This article takes a closer look at each clinical study. Convincing evidence must be published on 2 issues for sublobar resection to be accepted as a standard surgical option for GGN lung adenocarcinoma. In the absence of such evidence, it is better to perform lobar resection as long as the patient has sufficient lung function. The first issue is the definition of a sufficient resection margin, and the second is whether lymph node metastasis is conclusively ruled out before surgery. An additional issue is the need for an accurate calculation of the total size and solid size on CT. Given the results of clinical studies so far, wedge resection or segmentectomy shows a good prognosis for GGNs with a total size of 2 cm or less. Therefore, sublobar resection will play a key role even in patients who can tolerate lobectomy.

Abstract 2094: Correaltions between genome-wide DNA methylation profiles and genomic driver aberrations during multistage lung adenocaricinogenesis

Abstract The aim of this study was to clarify correlations between epigenomic and genomic alterations during multistage lung adenocarcinogenesis. Single-CpG resolution genome-wide DNA methylation analysis with the Infinium HumanMethylation27 BeadChip was performed using 162 paired samples of non-cancerous lung tissue (N) and corresponding tumorous tissue (T) from patients with lung adenocarcinomas. Correlations between DNA methylation data on the one hand and clinicopathological parameters and genomic driver mutations, i.e. mutations of EGFR, KRAS, BRAF, and HER2 and fusion involving ALK, RET, and ROS1, were examined. In N samples, which were considered to be at precancerous stages, DNA methylation levels in 12,629 probes were significantly correlated with recurrence-free survival. DNA methylation profiles at the precancerous N stages may determine the prognostic outcome. Principal component analysis revealed that distinct DNA methylation profiles at the precancerous N stage tended not to induce specific genomic driver mutations. Most of the genes showing significant DNA methylation alterations during transition from N to T were shared by two or more driver aberration groups, indicating that, in general, each driver mutation does not induce a specific DNA methylation profile. Commonly shared DNA methylation alterations which could potentially result in expression abnormalities were enriched among transcriptional factors. In contrast, only 67 genes showed EGFR mutation-specific or “pan-negative”-specific DNA methylation alterations. Among the 67 genes, most of the DNA methylation alterations were correlated with clinicopathological parameters reflecting tumor aggressiveness. We further focused on the ZNF132 gene, for which DNA hypermethylation had been observed only in the “pan-negative”-type lung adenocarcinomas and was significantly correlated with their vascular invasion. 5-aza-2'-deoxycytidine treatment restored the expression levels of ZNF132 mRNA in lung adenocarcinoma cell lines, indicating that ZNF132 had been silenced due to DNA hypermethylation. Knockdown of ZNF132 using siRNA transfection led to increased cell migration ability, rather than increased cell growth or reduced apoptosis. We concluded that DNA hypermethylation of the ZNF132 gene participates in the clinicopathological aggressiveness of “pan-negative”-type lung adenocarcinomas. In addition, DNA methylation alterations at the precancerous stage may determine tumor aggressiveness, and such alterations that accumulate after driver mutation may additionally modify clinicopathological features through alterations of gene expression. Citation Format: Kenichi Hamada, Ying Tian, Mao Fujimoto, Takahashi Yoriko, Takashi Kohno, Koji Tsuta, Shun-ichi Watanabe, Teruhiko Yoshida, Hisao Asamura, Yae Kanai, Eri Arai. Correaltions between genome-wide DNA methylation profiles and genomic driver aberrations during multistage lung adenocaricinogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2094.

DNA hypermethylation of the ZNF132 gene participates in the clinicopathological aggressiveness of 'pan-negative'-type lung adenocarcinomas.

Although some previous studies have examined epigenomic alterations in lung adenocarcinomas, correlations between epigenomic events and genomic driver mutations have not been fully elucidated. Single-CpG resolution genome-wide DNA methylation analysis with the Infinium HumanMethylation27 BeadChip was performed using 162 paired samples of adjacent normal lung tissue (N) and the corresponding tumorous tissue (T) from patients with lung adenocarcinomas. Correlations between DNA methylation data on the one hand and clinicopathological parameters and genomic driver mutations, i.e. mutations of EGFR, KRAS, BRAF and HER2 and fusions involving ALK, RET and ROS1, were examined. DNA methylation levels in 12 629 probes from N samples were significantly correlated with recurrence-free survival. Principal component analysis revealed that distinct DNA methylation profiles at the precancerous N stage tended not to induce specific genomic driver aberrations. Most of the genes showing significant DNA methylation alterations during transition from N to T were shared by two or more driver aberration groups. After small interfering RNA knockdown of ZNF132, which showed DNA hypermethylation only in the pan-negative group and was correlated with vascular invasion, the proliferation, apoptosis and migration of cancer cell lines were examined. ZNF132 knockdown led to increased cell migration ability, rather than increased cell growth or reduced apoptosis. We concluded that DNA hypermethylation of the ZNF132 gene participates in the clinicopathological aggressiveness of ‘pan-negative’ lung adenocarcinomas. In addition, DNA methylation alterations at the precancerous stage may determine tumor aggressiveness, and such alterations that accumulate after driver mutation may additionally modify clinicopathological features through alterations of gene expression.

Differences in molecular epidemiology of lung cancer among ethnicities (Asian vs. Caucasian).

BackgroundDifferences in carcinogenesis and therapeutic efficacy according to ethnicity have been reported for lung cancer, and understanding differences in genetic mutation profiles among ethnicities is important for interpreting the results of clinical trials, preventing carcinogenesis, and individualizing treatment. However, no studies have focused on differences in mutation profiles among different ethnicities using large-scale genomic analysis data with detailed information on smoking history, the main cause of lung cancer.MethodsTo clarify the differences in genetic mutation profiles between Caucasian and Japanese subjects, we compared data from The Cancer Genome Atlas, which mainly included Caucasians, with results from the Japan Molecular Epidemiology for lung cancer study, which is an epidemiological study only involving Japanese subjects. We divided the participants into four groups according to smoking status and performed comparative analysis by tissue type (lung adenocarcinoma and squamous cell lung cancer).ResultsIn patients with lung adenocarcinoma, the frequency of EGFR mutations was lower in Caucasian subjects than in Japanese subjects (14.6% vs. 51.1%), whereas the frequencies of mutations in other genes, namely KRAS (32.9% vs. 9.3%), TP53 (45.2% vs. 20.7%), BRAF (9.6% vs. 1.3%), PIK3CA (5.9% vs. 2.6%), KEAP1 (17.8% vs. 0.5%), NF1 (10.9% vs. 0.5%), STK11 (17.8% vs. 0.7%), RBM10 (8.7% vs. 0.1%), and MET (7.8% vs. 0.1%), were higher in Caucasian subjects. Among patients with squamous cell carcinoma, TP53 (81.2% vs. 49.1%), PIK3CA (14.5% vs. 6.8%), KEAP1 (12.7% vs. 0.9%), and NFE2L2 mutations (15.8% vs. 13.6%) were more common in Caucasian subjects.ConclusionsEthnicity is an important and complex characteristic that must be recognized and considered, even in the era of precision medicine. We should collaborate to share data for different ethnicities and incorporate them into clinical practice and the design of global clinical studies. Carefully designed molecular epidemiological studies focusing on ethnic differences are warranted.

Concurrent somatic alterations of TP53 and RB1 in Chinese lung adenocarcinoma with or without EGFR mutations.

e21526 Background: Endothelial growth factor receptor (EGFR)-mutant lung adenocarcinoma displays diverse responses to tyrosine kinase inhibitor therapy. Concurrent somatic alterations represent different biological substantial proportion of patients with EGFR mutations and impacts their prognosis. Methods: We conducted this retrospective study to clarify the comprehensive concurrent genetic alterations of TP53 and RB1 and tumor mutational burden (TMB) in 712 EGFR-mutant or EGFR-wild type lung adenocarcinoma by next generation sequencing-based gene panel tests. Results: EGFR was the most frequently mutated gene altered in 58.0% (413/712) of lung adenocarcinoma, followed by TP53 altered in 56.7%, KRAS altered in 13.3%, and RBM10 altered in 11.2% of all patients. Concurrent genetic alteration of TP53 and RB1 is more likely to be found in EGFR-mutant patients than in EGFR-wild type patients, with a frequency of 11.4% (45/413) and 4.3% (13/299), respectively (p = 0.003). However, the frequency of TP53 and RB1 concurrent alteration was similar in lung cancer with sensitive EGFR mutation compared to those with non-sensitive mutation (10.5% versus 11.6%). TP53 mutation was most frequently found in patients with RB1 mutation (58/61), irrespective of EGFR mutational status. Furthermore, significant difference was found regarding median TMB in patients with mutant EGFR compared to those with non-mutant EGFR (3.8 versus 4.3; p &lt; 0.001). Median TMB was higher for patients with TP53 and RB1 concurrent alteration than those without concurrent alteration in all patients (5.4 versus 4.3, p = 0.018). Conclusions: Our data showed that high prevalence of concurrent somatic alterations of TP53 and RB1 genes among lung adenocarcinoma patients with EGFR mutations, which might help understand several key biological processes and develop potential therapeutic strategies.

The Anticancer Mechanism of Bile Acid Receptor TGR5 in Mouse Models of Lung Cancer

In order to explore the anticancer mechanism of bile acid receptor TGR5 in lung cancer mouse models, healthy C57BL/6J mice of the wild-type and the TGR5 gene knockout type and human lung adenocarcinoma cell line A549 were selected as test system to construct animal models. The mouse models were used to observe the inhibitory effects of bile acid receptor TGR5 on lung cancer and related indicators, inflammatory mediators and mRNA expression levels. Through comparisons between the genes of wild-type mice and TGR5 knockout mice, it was found that the expression levels of IP-10, IL-1β, IFN-γ, CCL4, CXCL2, and CCL3 were significantly higher in the TGR5 knockout model group than those in the wild-type control group. Analysis of the effects of TGR5 on the proliferation of lung adenocarcinoma cells, A549, revealed that the RGT5 agonist could effectively inhibit proliferation of lung cancer cells. Analysis of the effects of TGR5 on apoptosis of lung adenocarcinoma cells A549, indicated that after adding DY240 or INT777 agonist, the level of cell apoptosis was significantly increased compared to the control group. Analysis of the effects of TGR5 on NF-κB signalling pathway showed that inactivating TGR5 could effectively inhibit the expression and phosphorylation of inflammatory factors related to signalling pathways in lung cancer cells. Therefore, it was found that the intervention with TGR5 agonist could effectively inhibit the proliferation of lung cancer cells, which achieved the expected results. Despite the deficiencies in the research process, it has provided a certain basis and ideas for subsequent research on exploring the treatments of lung cancer more accurately. Thus, the study is of great significance.

P65BTK is a novel potential actionable target in KRAS-mutated/EGFR-wild type lung adenocarcinoma

BackgroundLung cancer is still the main cause of cancer death worldwide despite the availability of targeted therapies and immune-checkpoint inhibitors combined with chemotherapy. Cancer cell heterogeneity and primary or acquired resistance mechanisms cause the elusive behaviour of this cancer and new biomarkers and active drugs are urgently needed to overcome these limitations. p65BTK, a novel isoform of the Bruton Tyrosine Kinase may represent a new actionable target in non-small cell lung cancer (NSCLC).Methodsp65BTK expression was evaluated by immunohistochemistry in 382 NSCLC patients with complete clinico-pathological records including smoking habit, ALK and EGFR status, and in metastatic lymph nodes of 30 NSCLC patients. NSCLC cell lines mutated for p53 and/or a component of the RAS/MAPK pathway and primary lung cancer-derived cells from Kras/Trp53 null mice were used as a preclinical model. The effects of p65BTK inhibition by BTK Tyrosine Kinase Inhibitors (TKIs) (Ibrutinib, AVL-292, RN486) and first-generation EGFR-TKIs (Gefitinib, Erlotinib) on cell viability were evaluated by MTT. The effects of BTK-TKIs on cell growth and clonogenicity were assessed by crystal violet and colony assays, respectively. Cell toxicity assays were performed to study the effect of the combination of non-toxic concentrations of BTK-TKIs with EGFR-TKIs and standard-of-care (SOC) chemotherapy (Cisplatin, Gemcitabine, Pemetrexed).Resultsp65BTK was significantly over-expressed in EGFR-wild type (wt) adenocarcinomas (AdC) from non-smoker patients and its expression was also preserved at the metastatic site. p65BTK was also over-expressed in cell lines mutated for KRAS or for a component of the RAS/MAPK pathway and in tumors from Kras/Trp53 null mice. BTK-TKIs were more effective than EGFR-TKIs in decreasing cancer cell viability and significantly impaired cell proliferation and clonogenicity. Moreover, non-toxic doses of BTK-TKIs re-sensitized drug-resistant NSCLC cell lines to both target- and SOC therapy, independently from EGFR/KRAS status.Conclusionsp65BTK results as an emerging actionable target in non-smoking EGFR-wt AdC, also at advanced stages of disease. Notably, these patients are not eligible for EGFR-TKIs-based therapy due to a lack of EGFR mutation. The combination of BTK-TKIs with EGFR-TKIs is cytotoxic for EGFR-wt/KRAS-mutant/p53-null tumors and BTK-TKIs re-sensitizes drug-resistant NSCLC to SOC chemotherapy. Therefore, our data suggest that adding BTK-TKIs to SOC chemotherapy and EGFR-targeted therapy may open new avenues for clinical trials in currently untreatable NSCLC.

ALK and ROS1 rearrangements, coexistence and treatment in epidermal growth factor receptor-wild type lung adenocarcinoma: a multicenter study of 732 cases.

Anaplastic lymphoma kinase (ALK) and c-ros oncogene 1 (ROS1) rearrangements represent two most frequent fusion targets in lung adenocarcinoma. Our study was intended to explore the clinicopathological characteristics, coexistence and treatment of ALK/ROS1-rearranged patients of lung adenocarcinoma without epidermal growth factor receptor (EGFR) mutation. Patients with wild-type EGFR mutation were screened for ALK/ROS1 at four domestic hospitals. ALK/ROS1 rearrangements were detected by reverse transcription-polymerase chain reaction (RT-PCR). Progression-free survival (PFS) curve was plotted with the Kaplan-Meier method. Among 732 eligible cases, ALK and ROS1 rearrangements were detected in 89 (12.2%) and 32 (4.4%) patients respectively. One patient harbored coexisting ALK/ROS1 fusion. Both ALK and ROS1-positive phenotypes were predominantly detected in younger non-smokers. More ALK/ROS1-rearranged patients were correlated with the expressions of TTF1, napsin A and solid predominant adenocarcinoma subtype. Thirty-three ALK and six ROS1 rearrangement patients received crizotinib treatment at an advanced stage. The median PFS was 9.5 months for ALK-positive patients and it was not attained in ROS1-rearranged counterparts. The frequency of ALK and ROS1 rearrangements is elevated in EGFR-wild-type patients and the phenomenon of coexisting ALK/ROS1 has remained extremely rare. The rearrangements of ALK/ROS1 are correlated with age, smoking status, expressions of TTF1 & napsin A and solid predominant adenocarcinoma subtype.