Expression of ID2 in NSCLC Figure 3. Survival analysis. A–F: Kaplan—Meier curves established for all NSCLC patients (n = 59) according to disease stage (A), nodal status (B), T status (C), nuclear (D), cytoplasmic (E), and global expression of ID2 (F). G–I: Kaplan—Meier curves obtained for patients with poorly differentiated tumors (n = 18) according to nuclear (G), cytoplasmic (H), and global (I) expression of ID2. Curves were compared with log-rank test. doi:10.1371/journal.pone.0004158.g003 reduce the pro-tumoral effect of ID2. These results were globally in agreement with our study, as we demonstrated a direct relationship between nuclear expression of ID2 and the survival of patients. However, differences regarding ID2 localization may originate in part from differences between the two pathologies, but also from differences in the specificity of the antibodies. Indeed, we Table 2. Mutivariate survival analysis after stratification according to patients with poorly differentiated tumors. Variables T status N status Nuclear expression of ID2 Relative Risk 6.7 2.05 13.7 95% CI 1.1–39.1 0.4–9.5 1.2–161 p value 0.035 0.36 0.034 doi:10.1371/journal.pone.0004158.t002 have used Zymed antibodies as recommended by Perk et al., who warned against the use of poorly specific ID2 antibodies [24]. Altogether, our data suggest that both ID2 expression and intracellular localization must be evaluated in order to determine the prognosis for cancer patients, including those with lung cancer. The intracellular localization of ID2 may play a major role in the control of ID2 function and cell fate regulation. Kurooka et al. determined that the nucleo-cytoplasmic shuttling of ID2 is an active mechanism dependent on the CRM-1 protein, and is inversely correlated to transcriptional repression via the E-box sequence [25]. In addition, Lasorella and Iavarone demonstrated that the interaction between ID2 and the actin-associated protein enigma homolog (ENH) also controls the localization of ID2 [26]. Indeed, downregulation of ENH expression prevents relocalization of ID2 into the cytoplasm in differentiated neuroblastoma cells. This result allows us to speculate on the possible implication of ENH or CRM-1 protein in the dedifferentiation mechanism of NSCLC. Our results indicate that nuclear ID2 is likely to be a PLoS ONE | www.plosone.org 5 January 2009 | Volume 4 | Issue 1 | e4158