High TUBB3 expression, an independent prognostic marker in patients with early non-small cell lung cancer treated by preoperative chemotherapy, is regulated by K-Ras signaling pathway

• Published in: Molecular cancer therapeutics 2012-05, Vol.11 (5), p.1203-1213
• Main Authors: Levallet, Guénaëlle, Bergot, Emmanuel, Antoine, Martine, Creveuil, Christian, Santos, Adriana O, Beau-Faller, Michelle, de Fraipont, Florence, Brambilla, Elisabeth, Levallet, Jérôme, Morin, Franck, Westeel, Virginie, Wislez, Marie, Quoix, Elisabeth, Debieuvre, Didier, Dubois, Fatéméh, Rouquette, Isabelle, Pujol, Jean-Louis, Moro-Sibilot, Denis, Camonis, Jacques, Zalcman, Gérard
• Format: Article
• Language: English
• Subjects: Adult; Aged; Antineoplastic Combined Chemotherapy Protocols - therapeutic use; Biomarkers, Tumor - genetics; Biomarkers, Tumor - metabolism; Cancer; Carcinoma, Non-Small-Cell Lung - drug therapy; Carcinoma, Non-Small-Cell Lung - metabolism; Carcinoma, Non-Small-Cell Lung - mortality; Female; Gene Expression Regulation, Neoplastic; Humans; Life Sciences; Lung Neoplasms - drug therapy; Lung Neoplasms - metabolism; Lung Neoplasms - mortality; Male; Middle Aged; Mutation; Neoadjuvant Therapy; Prognosis; Proto-Oncogene Proteins p21(ras) - genetics; Proto-Oncogene Proteins p21(ras) - metabolism; Signal Transduction; Survival Analysis; Treatment Outcome; Tubulin - genetics; Tubulin - metabolism
• ISSN: 1535-7163; 1538-8514
• DOI: 10.1158/1535-7163.MCT-11-0899

We assessed the prognostic and predictive value of β-tubulin III (TUBB3) expression, as determined by immunohistochemistry, in 412 non-small cell lung cancer (NSCLC) specimens from early-stage patients who received neoadjuvant chemotherapy (paclitaxel- or gemcitabine-based) in a phase III trial (IFCT-0002). We also correlated TUBB3 expression with K-Ras and EGF receptor (EGFR) mutations in a subset of 208 cryopreserved specimens. High TUBB3 protein expression was associated with nonsquamous cell carcinomas (P < 0.001) and K-Ras mutation (P < 0.001). The 127 (30.8%) TUBB3-negative patients derived more than 1 year of overall survival advantage, with more than 84 months median overall survival versus 71.7 months for TUBB3-positive patients [HR, 1.58; 95% confidence interval (CI), 1.11-2.25)]. This prognostic value was confirmed in multivariate analysis (adjusted HR for death, 1.51; 95% CI, 1.04-2.21; P = 0.031) with a bootstrapping validation procedure. TUBB3 expression was associated with nonresponse to chemotherapy (adjusted HR, 1.31; 95% CI, 1.01-1.70; P = 0.044) but had no predictive value (taxane vs. gemcitabine). Taking account of these clinical findings, we further investigated TUBB3 expression in isogenic human bronchial cell lines only differing by K-Ras gene status and assessed the effect of K-Ras short interfering RNA (siRNA) mediated depletion, cell hypoxia, or pharmacologic inhibitors of K-Ras downstream effectors, on TUBB3 protein cell content. siRNA K-Ras knockdown, inhibition of RAF/MEK (MAP-ERK kinase) and phosphoinositide 3-kinase (PI3K)/AKT signaling, and hypoxia were shown to downregulate TUBB3 expression in bronchial cells. This study is the first one to identify K-Ras mutations as determinant of TUBB3 expression, a chemoresistance marker. Our in vitro data deserve studies combining standard chemotherapy with anti-MEK or anti-PI3K drugs in patients with TUBB3-overexpressing tumors.