Activity of NF-kappaB transcription factor is usually associated with stimulation of the proliferation potential of cancer cells. On the contrary, our preliminary data suggest that sustained activation of NF-kapp has an opposite effect on certain cell lines. Our research is aimed to dissect mechanisms behind the pro- and anti-proliferative effects of NF-kappaB in non-small cell lung carcinoma (NSCLC) cell lines. Methods: To avoid pleiotropic effects of extracellular stimuli, like TNF or EGF growth factors, we established NSCLC cell lines H1299 and H23 with constant overexpression of the RELA gene, which encodes the primary activation subunit of NF-kappaB. Expression analysis of target genes with qRT-PCR confirmed that H1299/RelA and H23/RelA cell lines maintained enhanced transcriptional activity of the exogenous RelA/p65. The rate of Proliferation, colony formation capacity, cell cycle, apoptosis rate, cellular senescence and expression of genes controlling the cell cycle were investigated. Results: We found that RELA overexpression suppressed proliferation of the H1299 cell culture. The RELA expression level and NF-kappaB transcriptional activity demonstrated inverse correlation with the proliferation rate. Similar results were obtained with the colony formation assay. The cells with high RELA expression had reduced ability to form individual colonies. We than tested a direct involvement of RelA/p65 in the cell cycle control or induction of apoptosis but neither cell cycle nor apoptosis rate were affected. Similarly, the expression analysis of genes involved in regulation of cell cycle and apoptosis (p21/WAF1, PUMA, BAX) did not reveal any difference. However, the SA-b-gal staining indicated that H1299/RelA had higher rate of cellular senescence comparing to the parental cell line. To test whether the negative impact of NF-kappaB activation is common for other NSCLC cell lines, we analyzed effects of RELA overexpression in another NSCLC cells, H23. In contrast to H1299 cells, sustained NF-kappaB activation in H23 cells led to an enhanced proliferation rate. Further investigations of possible mechanisms behind the differences in NF-kappaB activation between the H1299 and H23 cell lines are in the progress. Conclusions: We demonstrated that constant NF-kappaB activation might either suppress or stimulate proliferation of different NSCLC cells. The effect does not involve cell cycle control or apoptosis but may implicate cellular senescence. Considering that H1299 and H23 cells lines differ in p53 status, further investigation of NF-kappaB may reveal novel aspects of NF-kappaB/interplay in NSCLC. Funding: The work was supported by the Russian Government Program for the Recruitment of the leading scientists into the Russian Institutions of Higher Education 14.W03.31.0029.