Pu, Xia

Abstract Definitive radiotherapy combined with chemotherapy is associated with enhanced locoregional control and clinical outcomes in non-small cell lung cancer (NSCLC) patients who are not candidates for surgery. However, radiation-induced pneumonitis and esophagitis are common dose-limiting inflammatory conditions that occur in these patients. These events can compromise the effect of the overall treatment and reduce quality of life in these patients. It is not clear what factors contribute to differences in the development of toxicities among patients with same clinical or dosimetric parameters. In this study, we conducted a large scale pathway-based analysis to identify inflammation-related genetic variants associated with radiation-induced pneumonitis or esophagitis. In the initial discovery phase, 11,930 inflammation-related genetic variants were genotyped in 243 stage I-III NSCLC patients who were treated with definitive radiotherapy at MD Anderson Cancer Center. A fast-track validation was performed for the top candidate SNPs in an additional 212 NSCLC cases following the same criteria. A total of 14 SNPs and 12 SNPs were validated for pneumonitis and esophagitis, respectively. Meta-analysis was used to summarize the results from two phases. The most significant SNPs for esophagitis and pneumonitis mapped to chromosome 18p (OR=9.05, 95%CI=2.68-30.59, P-meta=3.90×10−4) and 14q (OR=4.75, 95%CI=2.17-10.41, P-meta =9.61×10−5), respectively. To explore the potential underlying mechanisms, we analyzed radiation sensitivity and eQTL relationships of the identified SNPs and their host genes in 277 ethnically defined human lymphoblastoid cell lines. Among the 24 host genes for the 26 identified SNPs, three genes (DDX58, GZMB, CDC2) showed significant association with radiation responses in these cell lines (P<0.05). Moreover, a significant eQTL relationship was identified between CDC2 expression and the genotype of the identified CDC2 SNP (P=0.022). Interestingly, this CDC2 SNP is predicted to create a new miRNA binding site. Luciferase reporter assays to test the influence of this SNP on miRNA regulation is currently ongoing. Our results provide evidence that inflammation-related genetic variants contribute to the development of radiation-induced pneumonitis and esophagitis. The results also suggest that gene expression changes through altered miRNA binding may be an underlying mechanism for these events. Citation Format: Xia Pu, Liewei Wang, Michelle AT Hildebrandt, Yuanqing Ye, Joe Y. Chang, Charles Lu, Heath D. Skinner, Nifang Niu, Gregory D. Jenkins, Ritsuko Komaki, John D. Minna, Jack A. Roth, Richard M. Weinshilboum, Xifeng Wu. Inflammation-related genetic variants predict radiation-induced toxicity following definitive radiotherapy for non-small cell lung cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3626. doi:10.1158/1538-7445.AM2013-3626

Abstract To identify the genetic factors that influence overall survival in never smokers who have non–small cell lung carcinoma (NSCLC), we conducted a consistency meta-analysis study using genome-wide association approaches for overall survival in 327 never smoker patients with NSCLC from The University of Texas MD Anderson Cancer Center (Houston, TX) and 293 cases from the Mayo Clinic (Rochester, MN). We then conducted a two-pronged validation of the top 25 variants that included additional validation in 1,256 patients with NSCLC from Taiwan and assessment of expression quantitative trait loci (eQTL) and differential expression of genes surrounding the top loci in 70 tumors and matched normal tissues. A total of 94 loci were significant for overall survival in both MD Anderson and Mayo studies in the consistency meta-analysis phase, with the top 25 variants reaching a P value of 10−6. Two variants of these 25 were also significant in the Taiwanese population: rs6901416 [HR, 1.44; 95% confidence interval (CI), 1.01–2.06] and rs10766739 (HR, 1.23; 95%CI, 1.00–1.51). These loci resulted in a reduction of median survival time of at least eight and five months in three populations, respectively. An additional six variants (rs4237904, rs7976914, rs4970833, rs954785, rs485411, and rs10906104) were validated through eQTL analysis that identified significant correlations with expression levels of six genes (LEMD3, TMBIM, ATXN7L2, SHE, ITIH2, and NUDT5, respectively) in normal lung tissue. These genes were also significantly differentially expressed between the tumor and normal lung tissue. These findings identify several novel, candidate prognostic markers for NSCLC in never smokers, with eQTL analysis suggesting a potential biologic mechanism for a subset of these observed associations. Cancer Res; 73(13); 4028–38. ©2013 AACR.

Abstract Purpose: Lung cancer in never-smokers (LCINS) is increasingly recognized as a distinct disease from that in ever-smokers owing to substantial differences in etiology, clinical characteristics, and prognosis. Therefore, we aimed to identify prognostic markers specific for LCINS. Experimental Design: First, 11,930 single-nucleotide polymorphisms (SNP) in 904 inflammation-related genes were genotyped, and their associations with overall survival in 411 patients with LCINS at MD Anderson Cancer Center were analyzed. Next, validation of the top 27 SNPs in 311 patients with LCINS at Mayo Clinic was conducted. Results: Three SNPs (IL17RA:rs879576, BMP8A:rs698141, and STY:rs290229) were validated (P < 0.05), and two SNPs (CD74:rs1056400 and CD38:rs10805347) reached borderline significance (P = 0.08) in the Mayo Clinic population. We validated a survival-tree created in the MD Anderson population exploring gene–gene interactions in the Mayo Clinic population. This survival-tree stratified patients into subsets with significantly different risks of death: patients with the rs1056400_GG/rs698141_GA + AA genotype had significantly higher risk of death in both MD Anderson (HR:2.32, 95%CI: 1.58–3.41) and Mayo (HR:1.97, 95%CI: 1.11–3.50) populations compared with those with the rs1056400_GG/rs698141_GG or rs1056400_GA + AA genotype. We evaluated these five SNPs in 996 ever-smokers from MD Anderson and found no significant associations. Conclusions: Our study provides strong evidence that inflammation-related genetic variations can affect clinical outcomes in LCINS, which may lead to significant biologic insight into these outcomes. Clin Cancer Res; 18(21); 5983–91. ©2012 AACR.

Abstract Lung cancer in never-smokers is increasingly recognized as distinct from that in ever-smokers owing to substantial differences in etiology, clinical characteristics, and prognosis. Therefore, we aimed to identify prognostic markers specific for lung cancer in never-smokers in a two-stage study. First, 11,930 single-nucleotide polymorphisms (SNPs) in 904 inflammation-related genes were genotyped, and their associations with overall survival in 411 never-smokers with lung cancer at The University of Texas MD Anderson Cancer Center were analyzed. Next, fast-track validation of the top 37 SNPs in 311 never-smokers with lung cancer at Mayo Clinic was performed. Three SNPs (interleukin 17 receptor A:rs879576, bone morphogenetic protein 8A:rs698141, and spleen tyrosine kinase:rs290229) were validated (p<0.05), and two more SNPs (CD74:rs1056400 and CD38:rs10805347) were borderline significant (p=0.08) in the Mayo Clinic population. In combined analysis of the MD Anderson and Mayo Clinic patients, IL17RA:rs879576 resulted in a 40% reduction in the risk of death (p=4.1 × 10−5 [p=0.61, heterogeneity test]). We successfully validated a survival tree created in MD Anderson population based on gene-gene interactions in the Mayo Clinic population. This survival tree stratified patients into subsets with significantly different risks of death: patients at MD Anderson with the rs1056400_GG/rs698141_GA+AA genotype had a 2.32-fold higher risk of death (p=1.72 × 10−5) and 9-month shorter median survival time (MST) (p=4.5 × 10−4, log-rank test) than did those with the rs1056400_GG/rs698141_GG or rs1056400_GA+AA genotype in the discovery phase. Also, the patients at Mayo Clinic with the former genotype had an approximately twofold higher risk of death (p=0.02) and a strikingly shorter (by 26 months) MST (p=0.029, log-rank test) than did those with the latter genotype in the validation phase. We evaluated these five SNPs in 996 ever-smokers and found no significant associations. To our knowledge, this is the first study to systematically evaluate the associations of genetic variations in major inflammation-related genes with outcome of lung cancer in never-smokers. It provides strong evidence that inflammation-related genetic variations can affect clinical outcomes in these patients, which may lead to significant biological insight into these outcomes. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2929. doi:1538-7445.AM2012-2929

AbstractLung cancer in lifetime never smokers is distinct from that in smokers, but the role of separate or overlapping carcinogenic pathways has not been explored. We therefore evaluated a comprehensive panel of 11,737 single-nucleotide polymorphisms (SNP) in inflammatory-pathway genes in a discovery phase (451 lung cancer cases, 508 controls from Texas). SNPs that were significant were evaluated in a second external population (303 cases, 311 controls from the Mayo Clinic). An intronic SNP in the ACVR1B gene, rs12809597, was replicated with significance and restricted to those reporting adult exposure to environmental tobacco smoke. Another promising candidate was an SNP in NR4A1, although the replication OR did not achieve statistical significance. ACVR1B belongs to the TGFR-β superfamily, contributing to resolution of inflammation and initiation of airway remodeling. An inflammatory microenvironment (second-hand smoking, asthma, or hay fever) is necessary for risk from these gene variants to be expressed. These findings require further replication, followed by targeted resequencing, and functional validation.Significance: Beyond passive smoking and family history of lung cancer, little is known about the etiology of lung cancer in lifetime never smokers that accounts for about 15% of all lung cancers in the United States. Our two-stage candidate pathway approach examined a targeted panel of inflammation genes and has identified novel structural variants that appear to contribute to risk in patients who report prior exposure to sidestream smoking. Cancer Discovery; 1(5): 420–9. ©2011 AACR.This article is highlighted in the In This Issue feature, p. 367