Kuo an Hsu

R331W Missense Mutation of Oncogene YAP1 Is a Germline Risk Allele for Lung Adenocarcinoma With Medical Actionability

PURPOSE Adenocarcinoma is the most dominant type of lung cancer in never-smoker patients. The risk alleles from genome-wide association studies have small odds ratios and unclear biologic roles. Here we have taken an approach featuring suitable medical actionability to identify alleles with low population frequency but high disease-causing potential. PATIENTS AND METHODS Whole-genome sequencing was performed for a family with an unusually high density of lung adenocarcinoma with available DNA from the affected mother, four affected daughters, and one nonaffected son. Candidate risk alleles were confirmed by matrix-assisted laser desorption ionization time of flight mass spectroscopy. Validation was conducted in an external cohort of 1,135 participants without cancer and 1,312 patients with lung adenocarcinoma. Family follow-ups were performed by genotyping the relatives of the original proband and the relatives of the identified risk-allele carriers. Low-dose computed tomography scans of the chest were evaluated for lung abnormalities. RESULTS YAP1 R331W missense mutation from the original family was identified and validated in the external controls and the cohort with lung adenocarcinoma. The YAP1 mutant-allele carrier frequency was 1.1% in patients with lung adenocarcinoma compared with 0.18% in controls (P = .0095), yielding an odds ratio (adjusted for age, sex, and smoking status) of 5.9. Among the relatives, YAP1-mutant carriers have overwhelmingly higher frequencies of developing lung adenocarcinoma or ground-glass opacity lung lesions than those who do not carry the mutation (10:0 v 1:7; P < .001). YAP1 mutation was shown to increase the colony formation ability and invasion potential of lung cancer cells. CONCLUSION These results implicated YAP1 R331W as an allele predisposed for lung adenocarcinoma with high familial penetrance. Low-dose computed tomography scans may be recommended to this subpopulation, which is at high risk for lung cancer, for personalized prevention and health management.

Clustered Genomic Alterations in Chromosome 7p Dictate Outcomes and Targeted Treatment Responses of Lung Adenocarcinoma With EGFR-Activating Mutations

PURPOSE Although epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have been proven more effective for patients with lung adenocarcinoma with EGFR-activating mutation rather than wild type, the former group still includes approximately 30% nonresponders. The molecular basis of this substantial response heterogeneity is unknown. Our purpose was to seek molecular aberrations contributing to disease progression at the genome-wide level and identify the prognostic signature unique to patients with EGFR-activating mutation. PATIENTS AND METHODS We first investigated the molecular differences between tumors with EGFR-activating mutation and wild-type tumors by conducting high-density array comparative genomic hybridization on a collection of 138 adenocarcinoma tissues. We then used an independent group of 114 patients to validate the clinical relevance of copy-number alterations (CNAs) in predicting overall and disease-free survival. Finally, focusing on 23 patients with EGFR mutation receiving EGFR-TKI treatment, we investigated the association between CNAs and response to EGFR-TKIs. RESULTS We identified chromosome regions with differential CNAs between tumors with EGFR-activating mutation and wild-type tumors and found the aberration sites to cluster highly on chromosome 7p. A cluster of six representative chromosome 7p genes predicted overall and disease-free survival for patients with EGFR-activating mutation but not for those with wild type. Importantly, simultaneous presence of more genes with increased CNAs in this cluster correlated with less favorable response to EGFR-TKIs in patients with EGFR-activating mutation. CONCLUSION Our results shed light on why responses to EGFR-TKIs are heterogeneous among patients with EGFR-activating mutation. They may lead to better patient management in this population.

Pemetrexed induces both intrinsic and extrinsic apoptosis through ataxia telangiectasia mutated/p53-dependent and -independent signaling pathways

Pemetrexed, a new‐generation antifolate, has demonstrated promising single‐agent activity in front‐ and second‐line treatments of non‐small cell lung cancer. However, the molecular mechanism of pemetrexed‐mediated antitumor activity remains unclear. The current study shows that pemetrexed induced DNA damage and caspase‐2, ‐3, ‐8, and ‐9 activation in A549 cells and that treatment with caspase inhibitors significantly abolished cell death, suggesting a caspase‐dependent apoptotic mechanism. The molecular events of pemetrexed‐mediated apoptosis was associated with the activation of ataxia telangiectasia mutated (ATM)/p53‐dependent and ‐independent signaling pathways, which promoted intrinsic and extrinsic apoptosis by upregulating Bax, PUMA, Fas, DR4, and DR5 and activating the caspase signaling cascade. Supplementation with dTTP allowed normal S‐phase progression and rescued apoptotic death in response to pemetrexed. Overall, our findings reveal that the decrease of thymidylate synthase and the increase of Bax, PUMA, Fas, DR4, and DR5 genes may serve as biomarkers for predicting responsiveness to pemetrexed.

Epidermal Growth Factor Receptor Mutation Status in Stage I Lung Adenocarcinoma with Different Image Patterns

Purpose: Early lung adenocarcinoma may present with ground-glass opacity (GGO) component in computed tomography (CT) scan. Epidermal growth factor receptor (EGFR) mutation had been reported in patients with lung cancer with GGO patterns. Nevertheless, the correlation between clinical characteristics, CT image patterns, and EGFR mutation status was indeterminate. Methods: Patients with stage I lung adenocarcinoma with tumor lesions less than 3 cm were included and classified into pure GGO, part-solid, and solid patterns by CT scan images. All patients had EGFR mutation test from frozen tumors. Available paraffin-embedded archival tissues were microdissected into three different locations similar to CT images with central and peripheral parts of tumor, and adjacent normal part for EGFR mutation tests. Results: Totally, 162 patients were analyzed, 90 women and 72 men, and 128 nonsmokers. The patients included 35 (21.6%) pure GGO, 41 (25.3%) part-solid, and 86 (53.1%) solid lesions. The EGFR mutation rate was 64.2% (n = 104). Analysis of the correlation between CT image patterns and EGFR mutation, the less GGO ratio had more typical mutation, especially L858R (p = 0.037). In 45 microdissected tumors, the central and peripheral parts had the same EGFR mutation status. In adjacent normal parts, 5 of 32 (15.6%) EGFR mutant patients had identical mutation but none in nonmutant patients. Conclusions: In stage I lung adenocarcinoma, typical mutation, especially L858R was detected more frequent in invasive solid pattern and significantly less in pure GGO pattern. EGFR mutation is an early event in the pathogenesis of lung adenocarcinoma and may facilitate the tumor into aggressive behavior.

Identification of Five Driver Gene Mutations in Patients with Treatment-Naïve Lung Adenocarcinoma in Taiwan

Background It is important to select appropriate targeted therapies for subgroups of patients with lung adenocarcinoma who have specific gene alterations. Methods This prospective study was a multicenter project conducted in Taiwan for assessment of lung adenocarcinoma genetic tests. Five oncogenic drivers, including EGFR, KRAS, BRAF, HER2 and EML4-ALK fusion mutations, were tested. EGFR, KRAS, BRAF and HER2 mutations were assessed by MALDI-TOF MS (Cohort 1). EML4-ALK translocation was tested by Ventana method in EGFR-wild type patients (Cohort 2). Results From August 2011 to November 2013, a total of 1772 patients with lung adenocarcinoma were enrolled. In Cohort 1 analysis, EGFR, KRAS, HER2 and BRAF mutations were identified in 987 (55.7%), 93 (5.2%), 36 (2.0%) and 12 (0.7%) patients, respectively. Most of these mutations were mutually exclusive, except for co-mutations in seven patients (3 with EGFR + KRAS, 3 with EGFR + HER2 and 1 with KRAS + BRAF). In Cohort 2 analysis, 29 of 295 EGFR-wild type patients (9.8%) were positive for EML4-ALK translocation. EGFR mutations were more common in female patients and non-smokers and KRAS mutations were more common in male patients and smokers. Gender and smoking status were not correlated significantly with HER2, BRAF and EML4-ALK mutations. EML4-ALK translocation was more common in patients with younger age. Conclusion This was the first study in Taiwan to explore the incidence of five oncogenic drivers in patients with lung adenocarcinoma and the results could be valuable for physicians in consideration of targeted therapy and inclusion of clinical trials.

EGFR L858R Mutation and Polymorphisms of Genes Related to Estrogen Biosynthesis and Metabolism in Never-Smoking Female Lung Adenocarcinoma Patients

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 ORs (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.

Characteristics of young lung cancer: Analysis of Taiwan's nationwide lung cancer registry focusing on epidermal growth factor receptor mutation and smoking status

Lung cancer is relatively rare in young patients as the median age at diagnosis is 65–70 years. The main objective of this nationwide study was to investigate the characteristics of young lung cancer in Taiwan, especially the relationships among smoking behavior, epidermal growth factor receptor (EGFR) mutation, and age. The National Taiwan Lung Cancer Registry, a database contain detailed cancer statistics, was analyzed in this study for the period 2011–2012. Young lung cancer was defined as age ≦ 45 years. There were 21,536 lung cancer patients (13,187 men and 8349 women). Among these patients, 1074 (5.0%) were in the younger group, and 20,462 patients (95.0%) were in the older group. Female gender (48.8% versus 38.2%, P < 0.001), never-smokers (47.3% versus 43.8%, P = 0.015), and adenocarcinoma (70.4% versus 58.1%, P < 0.001) were more frequent in the younger group. While the EGFR mutation rate was lower in the younger group (52.5% versus 60.6%, P = 0.001), the primary site of lung cancer and stage distribution were not significantly different. If only adenocarcinoma patients were included in the analysis, female gender, older age, and never-smokers were more likely to have EGFR mutation. In conclusion, lung cancer in young patients (≦ 45 year-old) was associated with unique characteristics, with greater percentages of female patients, adenocarcinoma, and never-smokers and a lower EGFR mutation rate compared with older patients.

Predictive factors for EGFR-tyrosine kinase inhibitor retreatment in patients with EGFR-mutated non-small-cell lung cancer – A multicenter retrospective SEQUENCE study

BACKGROUND Acquired resistance occurs in most non-small cell lung cancer (NSCLC) patients with epidermal growth factor receptor (EGFR) mutations experiencing a response to EGFR-tyrosine kinase inhibitor (TKI) initially. We investigated EGFR-TKI retreatment in patients who had previously received EGFR-TKI followed by chemotherapy. MATERIALS AND METHODS This was a retrospective multicenter study. Patients with locally advanced or metastatic adenocarcinoma or TTF-1 (+) NSCLC, positive EGFR sensitive mutation, and EGFR-TKI reuse after initial EGFR-TKI followed by chemotherapy were enrolled. The objectives were to assess the objective response rate (ORR), disease control rate (DCR), progression-free survival (PFS), and overall survival (OS) of EGFR TKI switched retreatment. RESULTS In total, 205 patients were enrolled, with a median age of 61.8 years (range 31.4-92.9). There was a larger proportion of females (62.9%) than males, and more never-smokers (73.2%) than ever-smokers. In the initial EGFR-TKI administration, 57.6% of patients showed a complete response (CR) or partial response (PR), and 34.6% had stable disease (SD); in the second-line chemotherapy, 13.7% had PR, and 58.0% had SD; in the EGFR-TKI retreatment, 7.3% had PR, and 37.1% had SD. The median PFS of first-line EGFR-TKI was 8.0 months (95% CI 7.3-8.2), and retreatment EGFR-TKI was 4.1 months (95% CI 2.7-4.6). The median OS since the start of the first-line EGFR-TKI therapy was 35.9 months (95% CI 28.8-50.9), and since the start of EGFR-TKI retreatment was 12.6 months (95% CI 10.4-20.9). In the univariable and multivariable regression analysis of factors associated with PFS of EGFR-TKI retreatment, time interval between the two EGFR TKIs equal to or more than 7 months was statistically significant (HR=0.62, 95% CI 0.44-0.86; HR=0.6, 95% CI 0.43-0.86), both p<0.01. Females with exon 21 mutation also showed a significant difference between the two groups (HR=0.51, 95% CI 0.30-0.86; HR=0.52 (0.31-0.88), both p<0.05). CONCLUSIONS EGFR-TKI retreatment was effective in prolonging survival, and was shown to be a worthwhile option for EGFR-mutated NSCLC patients after failure of first-line EGFR-TKI and chemotherapy. The survival benefit was especially pronounced in patients with longer drug holidays from the initial EGFR-TKI and in females with the exon 21 mutation.

EGFR exon 19 in-frame deletion and polymorphisms of DNA repair genes in never-smoking female lung adenocarcinoma patients

We explored potential associations between genetic polymorphisms in genes related to DNA repair and detoxification metabolism and epidermal growth factor receptor (EGFR) mutations in a cohort of 410 never‐smoking patients with lung adenocarcinoma. Multivariate‐adjusted odds ratios (aORs) and corresponding 95% confidence intervals (CI) of EGFR mutation status in association with the genotypes of DNA repair and detoxification metabolism genes were evaluated using logistic regression analysis. We found an association between in‐frame deletion in EGFR exon 19 and a single nucleotide polymorphism (SNP) rs1800566C/T located in NQO1 (aOR, 2.2 with 95% CI, 1.0–4.8) in female never‐smokers. The SNP rs744154C/G in ERCC4 was also associated with the EGFR exon 19 in‐frame deletion both in never‐smokers (aOR, 1.7 with 95% CI, 1.0–3.0) and female never‐smokers (aOR, 1.9 with 95% CI, 1.0–3.6). Although the association was marginally significant in multivariate logistic regression analysis, the A/A genotype of rs1047840 in EXO1 was associated with a 7.6‐fold increase in the occurrence of the EGFR exon 19 in‐frame deletion in female never‐smokers. Moreover, risk alleles in NQO1, ERCC4 and EXO1 were associated with an increasing aOR of the EGFR exon 19 in‐frame deletion both in never‐smokers (p = 0.007 for trend) and female never‐smokers (p = 0.002 for trend). Our findings suggest that the in‐frame deletion in EGFR exon 19 is associated with polymorphisms in DNA repair and detoxification metabolism genes in never‐smoking lung adenocarcinoma patients, especially in females.

Divergent epidermal growth factor receptor mutation patterns between smokers and non-smokers with lung adenocarcinoma

INTRODUCTION Smoking status is an important determinant of the prevalence of epidermal growth factor receptor (EGFR) mutations in lung cancer patients. However, it is unclear whether smoking status could also influence the spectrum of EGFR mutations. METHODS We enrolled patients with lung adenocarcinoma from three medical centers in Taiwan. EGFR mutations were assessed by Sanger direct sequencing. The objective of this study was to evaluate the influence of smoking status on both the frequency and patterns of EGFR mutations. RESULTS From 2001 to 2013, a total of 1175 patients with lung adenocarcinoma were enrolled for EGFR mutation analysis. The overall EGFR mutation rate was 59.6%, which was significantly higher in females than males (69.1% vs. 49.8%) and in non-smokers than current/former smokers (73.8% vs. 29.8%) (both P<0.001). Among patients harboring EGFR mutations, smokers expressed L858R mutation less frequently (35.2% vs. 50.2%, P=0.005) and exon 19 deletions more frequently (52.8% vs 38.8%, P=0.008) than non-smokers. Smokers and non-smokers also had divergent exon 19 deletions subtypes (Del E746-A750 82.5% vs. 57.6%, respectively, P<0.001). Among subgroup patients harboring the L858R mutation, smokers were associated with a higher rate of complex mutations than non-smokers (34.2% vs. 8.4%, P<0.001). CONCLUSIONS Our results suggested that smoking status could influence not only the frequency but also the spectrum of EGFR mutations. These findings provide a clue for further investigation of EGFR mutagenesis.