Establishment, culture and characterization of gemcitabine hydrochloride‐resistant human non‐small cell lung carcinoma cell line derived cancer stem cells.

Due to their high expression profile of multi‐drug resistance genes, cancer stem cells (CSCs) are the main source of drug resistance. The aim of this study was to establish a gemcitabine‐hydrochloride‐resistant (rt) human non‐small cell lung cancer (hNSCLC) cell line and their CSC line to be used as disease models in various cancer studies. In the first phase of study, a gemcitabine hydrochloride‐rt hNSCLC line cells was produced by making them rt through periodic exposure to gemcitabine hydrochloride. This acquired gemcitabine‐hydrochloride‐rt hNSCLC cell line was characterized for resistance. Subsequently, a CSC population with a CD326 + CD133 + CD44+ phenotypes was immunoselectively isolated from gemcitabine hydrochloride‐rt hNSCLCs purified from a single cell by colony forming technology. This rt CSC line was characterized for both resistance and stemness. Rt and non‐rt CSCs were analyzed and compared with each other in terms of immunophenotyping the expression profiles of ALDH1, CD90, ABCG2, CD44 and MDR1, which are CSC specific markers, of demonstrating mitotic capacity with growth curve analysis and of their ability to form tumor spheroids in three different 3D cultures. The results of this study demonstrated for the first time the successful generation of both gemcitabine‐hydrochloride‐rt hNSCLC cells and CSCs derived from gemcitabine‐hydrochloride‐rt hNSCLC cells. It was also shown that isolated and characterized rt CSCs could proliferate and form tumor spheres in vitro using three different 3D in vitro techniques. It was shown that the cell surface markers CD326, CD133 and CD44 can serve as an antibody panel for CSCs. Significance statement: This study aimed to establish gemcitabine‐hydrochloride‐resistant (rt) human non‐small cell lung cancer (hNSCLC) cells and their cancer stem cells (CSCs) for cancer research. Gemcitabine‐rt hNSCLC cells were generated through periodic exposure. A CD326 + CD133 + CD44 + CSC population was isolated from these rt cells. Comparative analysis of rt and non‐rt CSCs revealed significant differences in marker expression, mitotic capacity, and spheroid formation ability. This study demonstrated successful generation of both rt hNSCLC cells and their CSCs, highlighting the potential of CD326, CD133, and CD44 as CSC markers. [ABSTRACT FROM AUTHOR]