Sirna duplexes targeting

All NSCLC cell lines, were obtained from the U.S. National Cancer Institute (NCI; MTA no. 2702–09). Cells were incubated at 37°C and maintained at 5% CO₂. H23, H226, IMR-90 cell was grown in DMEM/HIGH GLUCOSE medium (SH30243.01, Hyclone, Logan, UT, USA) containing 10% FBS. Lung primary cell was airway epithelial cell basal medium (PCS-300-030, ATCC, Manassas, VA, USA) with the bronchial epithelial cell growth kit (PCS-300-040, ATCC, Masassas, VA, USA). NSCLC cells were grown in RPMI 1640 medium (SH30027.01, HyClone, Logan, UT, USA) containing 10% fetal bovine serum (FBS) (SH30070.03HI, HyClone, Logan, UT, USA), penicillin, and streptomycin. siRNA duplexes targeting human ALDH1L1 (sc-78373), DHFR (sc-37078), GOT2 (sc-60052), and MDH2 (sc-89622) (Santa Cruz Biotechnology, Santa Cruz, CA, USA) were introduced into cells using Lipofectamine^® 3000 (L3000015, Invitrogen, Carlsbad, CA, USA) according to the manufacturer's instructions. As negative controls, cells were incubated with Lipofectamine^® 3000 alone and a negative siRNA (sc-37007, sc-44230) (Santa Cruz). For ALDH overexpression, p3x FLAG-CMV-ALDH isoform constructs individually expressing ALDH1L1 and DHFR were produced by Cosmogenetech (Seoul, KOREA). Each cDNA sequence of ALDH1L1 and DHFR was obtained from NCBI. The plasmids were transfected into cells using Lipofectamine^® 3000 according to the manufacturer's instructions.