Nrf2 and Keap1 Abnormalities in Non–Small Cell Lung Carcinoma and Association with Clinicopathologic Features

Purpose: To understand the role of nuclear factor erythroid-2–related factor 2 (Nrf2) and Kelch-like ECH-associated protein 1 (Keap1) in non–small cell lung cancer (NSCLC), we studied their expression in a large series of tumors with annotated clinicopathologic data, including response to platinum-based adjuvant chemotherapy. Experimental Design: We determined the immunohistochemical expression of nuclear Nrf2 and cytoplasmic Keap1 in 304 NSCLCs and its association with patients' clinicopathologic characteristics, and in 89 tumors from patients who received neoadjuvant (n = 26) or adjuvant platinum-based chemotherapy (n = 63). We evaluated NFE2L2 and KEAP1 mutations in 31 tumor specimens. Results: We detected nuclear Nrf2 expression in 26% of NSCLCs; it was significantly more common in squamous cell carcinomas (38%) than in adenocarcinomas (18%; P < 0.0001). Low or absent Keap1 expression was detected in 56% of NSCLCs; it was significantly more common in adenocarcinomas (62%) than in squamous cell carcinomas (46%; P = 0.0057). In NSCLC, mutations of NFE2L2 and KEAP1 were very uncommon (2 of 29 and 1 of 31 cases, respectively). In multivariate analysis, Nrf2 expression was associated with worse overall survival [P = 0.0139; hazard ratio (HR), 1.75] in NSCLC patients, and low or absent Keap1 expression was associated with worse overall survival (P = 0.0181; HR, 2.09) in squamous cell carcinoma. In univariate analysis, nuclear Nrf2 expression was associated with worse recurrence-free survival in squamous cell carcinoma patients who received adjuvant treatment (P = 0.0410; HR, 3.37). Conclusions: Increased expression of Nrf2 and decreased expression of Keap1 are common abnormalities in NSCLC and are associated with a poor outcome. Nuclear expression of Nrf2 in malignant lung cancer cells may play a role in resistance to platinum-based treatment in squamous cell carcinoma. Clin Cancer Res; 16(14); 3743–53. ©2010 AACR.

[1]  M. Hannink,et al.  PGAM5, a Bcl-XL-interacting Protein, Is a Novel Substrate for the Redox-regulated Keap1-dependent Ubiquitin Ligase Complex* , 2006, Journal of Biological Chemistry.

[2]  P. Muti,et al.  RIPK1/RIPK3 promotes vascular permeability to allow tumor cell extravasation independent of its necroptotic function , 2017, Cell Death & Disease.

[3]  M. Hannink,et al.  Structure of the Keap1:Nrf2 interface provides mechanistic insight into Nrf2 signaling , 2006, The EMBO journal.

[4]  M. McMahon,et al.  NRF2 and KEAP1 mutations: permanent activation of an adaptive response in cancer. , 2009, Trends in biochemical sciences.

[5]  S. Goodman,et al.  RNAi-mediated silencing of nuclear factor erythroid-2-related factor 2 gene expression in non-small cell lung cancer inhibits tumor growth and increases efficacy of chemotherapy. , 2008, Cancer research.

[6]  N. Yoo,et al.  Oncogenic NRF2 mutations in squamous cell carcinomas of oesophagus and skin , 2010, The Journal of pathology.

[7]  M. Ostland,et al.  Mutations in the epidermal growth factor receptor and in KRAS are predictive and prognostic indicators in patients with non-small-cell lung cancer treated with chemotherapy alone and in combination with erlotinib. , 2005, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[8]  H. Hansen,et al.  Lung cancer. , 1990, Cancer chemotherapy and biological response modifiers.

[9]  A. Kong,et al.  Molecular mechanisms of Nrf2‐mediated antioxidant response , 2009, Molecular carcinogenesis.

[10]  Rui Wang,et al.  Hypermethylation of the Keap1 gene in human lung cancer cell lines and lung cancer tissues. , 2008, Biochemical and biophysical research communications.

[11]  N. Colburn,et al.  Targeting Transcription Factors for Cancer Prevention—the Case of Nrf2 , 2008, Cancer Prevention Research.

[12]  E. Wanker,et al.  Nuclear Oncoprotein Prothymosin α Is a Partner of Keap1: Implications for Expression of Oxidative Stress-Protecting Genes , 2005, Molecular and Cellular Biology.

[13]  Elisabeth Brambilla,et al.  Pathology and genetics of tumours of the lung , pleura, thymus and heart , 2004 .

[14]  Ronald Simon,et al.  Tissue microarrays in drug discovery , 2003, Nature Reviews Drug Discovery.

[15]  N. Dubrawsky.  Cancer statistics , 1989, CA: a cancer journal for clinicians.

[16]  J. Herman,et al.  Dysfunctional KEAP1–NRF2 Interaction in Non-Small-Cell Lung Cancer , 2006, PLoS medicine.

[17]  Bengt Bergman,et al.  Cisplatin-based adjuvant chemotherapy in patients with completely resected non-small-cell lung cancer. , 2004, The New England journal of medicine.

[18]  A. Jemal,et al.  Cancer Statistics, 2008 , 2008, CA: a cancer journal for clinicians.

[19]  S. Biswal,et al.  Identification of Nrf2-regulated genes induced by the chemopreventive agent sulforaphane by oligonucleotide microarray. , 2002, Cancer research.

[20]  R. Bordoni.  Consensus conference: multimodality management of early- and intermediate-stage non-small cell lung cancer. , 2008, The oncologist.

[21]  K. Jordan-Sciutto,et al.  Expression of the fetal Alz-50 clone 1 protein induces apoptotic cell death. , 2005, Biochemical and biophysical research communications.

[22]  Tsutomu Ohta,et al.  Structural basis for defects of Keap1 activity provoked by its point mutations in lung cancer. , 2006, Molecular cell.

[23]  S. Hirohashi,et al.  Genetic alteration of Keap1 confers constitutive Nrf2 activation and resistance to chemotherapy in gallbladder cancer. , 2008, Gastroenterology.

[24]  Se-Jin Kim,et al.  Suppression of Nrf2-driven heme oxygenase-1 enhances the chemosensitivity of lung cancer A549 cells toward cisplatin. , 2008, Lung cancer.

[25]  S. Hirohashi,et al.  Loss of Keap1 function activates Nrf2 and provides advantages for lung cancer cell growth. , 2008, Cancer research.

[26]  Valeria Panebianco,et al.  Supplementary Figure 2 , 2012 .

[27]  M. Kwak,et al.  Role of the Nrf2-antioxidant system in cytotoxicity mediated by anticancer cisplatin: implication to cancer cell resistance. , 2008, Cancer letters.

[28]  S. Dhakshinamoorthy,et al.  Functional characterization and role of INrf2 in antioxidant response element-mediated expression and antioxidant induction of NAD(P)H:quinone oxidoreductase1 gene , 2001, Oncogene.

[29]  J. D. Engel,et al.  Keap1 represses nuclear activation of antioxidant responsive elements by Nrf2 through binding to the amino-terminal Neh2 domain. , 1999, Genes & development.

[30]  J. Minna,et al.  Clinical and biological features associated with epidermal growth factor receptor gene mutations in lung cancers. , 2006, Journal of the National Cancer Institute.

[31]  H. Stünzi,et al.  Tumours of the lung. , 1974, Bulletin of the World Health Organization.

[32]  Jihnhee Yu,et al.  Elevated Peroxiredoxin 1, but not NF-E2–Related Factor 2, Is an Independent Prognostic Factor for Disease Recurrence and Reduced Survival in Stage I Non–Small Cell Lung Cancer , 2007, Clinical Cancer Research.

[33]  Tsutomu Ohta,et al.  Cancer related mutations in NRF2 impair its recognition by Keap1-Cul3 E3 ligase and promote malignancy , 2008, Proceedings of the National Academy of Sciences.

[34]  D. Hallahan,et al.  Increased expression of nuclear factor E2 p45‐related factor 2 (NRF2) in head and neck squamous cell carcinomas , 2006, Head & neck.

[35]  S. Lippman,et al.  Lung cancer. , 2008, The New England journal of medicine.

[36]  P. Hirvikoski,et al.  Increased BTB-Kelch type substrate adaptor protein immunoreactivity associates with advanced stage and poor differentiation in renal cell carcinoma. , 2009, Oncology reports.

[37]  A. Józkowicz,et al.  Heme oxygenase-1 in tumors: is it a false friend? , 2007, Antioxidants & redox signaling.

[38]  J. Minna,et al.  EGFR tyrosine kinase domain mutations are detected in histologically normal respiratory epithelium in lung cancer patients. , 2005, Cancer research.