It is intriguing that many cell proliferation genes inversely correlate to the levels of CUDC-907 Nkx2-1 in NSCLC. This inverse correlation may explain the poorer prognosis of patients with NSCLC with low levels of NKX2-1. To determine if the reverse correlation is due to repression by direct NKX2-1 binding, ChIP analyses may be performed in human tumor tissues or tumor cell lines. Alternatively molecular analyses of the promoters of these genes in cell lines may provide information about the repression of these genes by Nkx2-1 binding. These experiments will be the focus of our future studies. Adenocarcinomas sub-classification based on gene expression profiling was proposed to improve prediction of malignant potential and prognosis. The associations identified in our studies may contribute to the molecular classification of these tumors and clarify NSCLC heterogeneity, holding great potential to increase the understanding of this disease. Our findings point to potential molecular mechanisms by which Nkx2-1 may differentially regulate transcriptional activity. First, inverse correlation in expression level of NKX2-1 and targets in tumors, and of Nkx2-1 and c-Met in MLE15 cells suggests a more widespread role of Nkx2-1 in transcriptional repression. This effect could be by direct binding or, alternatively, by recruitment or activation of transcriptional repressors by Nkx2-1 to downregulate particular genes. Nkx2-1 has been mostly linked to transcriptional activation in lung and other organs, although neuropilin-2 and RAGE have been reported to be downregulated by direct binding of Nkx2-1 to a cis-element in their promoters strongly supporting Nkx2-1 repressor activity. Second, there are target genes bound by Nkx2-1 at both developmental time points, whereas others are bound only at E11.5 or E19.5. Interactions with alternative co-factors differentially expressed at each time point might result in differential affinity and binding to alternative targets. Different isoforms and/or modifications of Nkx2-1 proteins by phosphorylation, acetylation or oxidation may affect affinity for particular ciselements or interactions to different co-factors at each time point. Identification of different forms of Nkx2-1 protein at E11.5 and E19.5 will be necessary to fully understand the different targets in alternative cell contexts. The specificity of Nkx2-1 binding has also been linked to promoter structure. Differences in Tubulin Acetylation Inducer chromatin modifications surrounding these cis-elements in different cell contexts could affect affinity of Nkx2-1 proteins. For example binding of Nkx2-1 to the Sftpb promoter is prevented by DNA methylation of the Sftpb promoter in non-expressing tissues, such as thyroid.