Nci h716

Normal colon cell line NCM-460 cells and human CRC cell lines CoLo205, DLD-1, HT-29, CoLo320, RKO, NCI-H716, and Caco-2 cells were from ATCC (Manassas, VA, USA). The NCM-460 cells were grown in M3 medium containing 10% FBS (Gibco, Grand Island, NY, USA). All of the CRC cell lines were grown in RPMI1640 supplemented with 10% FBS (Gibco) 37 °C/5% CO₂. For oxaliplatin (L-OHP) treatment, HT-29 and Caco-2 cells were treated with different doses of L-OHP (0, 5, 10, 20 and 40 μg/mL). For irinotecan (CPT-11) treatment, HT-29 and Caco-2 cells were treated with different doses of CPT-11 (0, 5, 10, 20 and 40 μg/mL). To study the protein stability, cycloheximide (CHX, 20 μg/mL) was administered for 0.25, 0.5, 1, 2, and 4 h. To block the ubiquitin–proteasome pathway, the cells were treated with 20 μM MG132. CHX and MG132 were purchased from MedChemExpress. shNC, sh-circ_RNF13-1, sh-circ_RNF13-2, sh-DDX27-1, sh-DDX-27-2, sh-TRIM24, sh-FBXW7, sh-MDM2, sh-TRIM25, sh-NEDD4, sh-TRIM11, sh-RNF180, sh-SYVN1, sh-MIB1, and pcDNA3.1-circ_RNF13 were purchased from GenePharma (Shanghai, China). HT-29 and Caco-2 cells were transfected 0.5 μg shRNA and/or plasmids using Lipofectamine 3000 (Invitrogen, Carlsbad, CA, USA). For double knockdown, 0.5 μg sh-circ_RNF13-1 and/or 0.5 μg sh-FBXW7 were co-transfected into CRC cells. At 48 h post-transfection, cells were harvested for subsequent analysis.

The FGFR2-overexpressing human gastric cell line Snu-16 (no. CRL-5974; American Type Culture Collection, ATCC) and human colorectal cell line NCI-H716 (no. CCL-251; ATCC) were cultured in RPMI-1640 medium (Thermo Fisher Scientific, Waltham, MA, USA). Human osteosarcoma cells U2OS (no. HTB-96; ATCC) were grown in Dulbecco’s modified Eagle’s medium (DMEM) (Invitrogen). All media were supplemented with 10% fetal calf serum (FBS, Invitrogen) and 1% penicillin/streptomycin mix (Gibco). All cells were cultured at 37°C in 5% CO₂ atmosphere and 95% humidity.