Lung cancer is the first cause of cancer death in men and the second in woman. The first therapeutic line of action against lung adenocarcinoma is surgery. However only 15% of cases are operable, thus chemotherapy is considered. Cisplatin-based combination chemotherapy regimens are currently used as a front-line therapy in the treatment of non-small cell lung cancer. Unfortunately, such treatment often leads to chemoresistance and hence therapy failure. Thus, it is urgent to investigate the molecular mechanisms involved in this resistance. Cancer cells are known to exhibit a high rate of proliferation, migration ability and resistance to apoptosis. These processes are proved to be controlled by calcium and calcium-permeable ion channels. Store Operated Calcium Channels (SOCs) represent a major calcium entry pathway in non-excitable cells and are shown to be implicated in drug resistance in different types of cancer. Recently, we showed that Orai3 is overexpressed in lung adenocarcinoma and is correlated to high tumor grade and controls cell proliferation via the Akt pathway (Ay et al., 2013). We also showed that Orai3 constitutes a predictive marker of metastasis and survival in resectable lung adenocarcinoma (Benzerdjeb et al., 2016). Furthermore, Orai3 overexpression confers resistance to chemotherapeutic drugs via the pro-survival PI3K/Sgk-1/Sek-1 pathway in breast cancer cells (Hasna et al., 2018). Hence, we aimed to investigate the Orai3 involvement in resistance to chemotherapy in lung adenocarcinoma cell lines. Experiments were conducted on two lung adenocarcinoma cell lines, H23 and A549.The expression of the SOC actors was studied using RT-qPCR and confirmed by Western-Blot. The inhibition of the expression of the SOC actors was done via transfection using (siOrai1, siOrai3 and siStim1) vs. siControl (electroporation, AMAXA). Cellular viability and mortality were assessed (48 hours or 4 days after treatment with Cisplatin) using MTT and Trypan blue methods. Calcium imaging experiments were also conducted to measure the functional effect of the treatment obtained at the level of the channels (quantification of the SOC entry). We found that Cisplatin treatment increased Orai3 expression, SOC entry, and favours cell survival in both cell lines. However, in contrast to H23 cells where Orai3 constitutes a native SOC channel, in A549 cell line, Orai3 doesn’t contribute to SOC entry but upon the treatment with Cisplatin, it becomes a SOC channel. Moreover, we found that the long term treatment with Cisplatin leads to morphological change of A549 cells along with increased stem cell marker, expression (Nanog) suggesting a stem cell enrichment process. In conclusion, we demonstrated that Orai3 channel becomes overexpressed during drug treatment in lung adenocarcinoma cell lines and contributes to Cisplatin resistance and the ability of conferring resistance is related to the SOC activity of the channel.