Sw1463

All cell lines, including HPAC, MIA Paca-2, Capan-1, NCI-H2122, NCI-H2030, SW837, SW1463 and 293 T cells, were purchased from ATCC and authenticated on their own cell lines. Pa01c, Pa02c, Pa03c, Pa14c, and Pa16c were kind gifts from Dr. Channing Der at UNC-CH and whole-exome sequenced^{72 (link)}. All lines were used for <6 months after receipt or resuscitation from cryopreservation. All cell lines were cultured in DMEM supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin. Mycoplasma testing was performed semiannually using a MycoSEQ Detection kit (Applied Biosystems).

Human colorectal cancer cell lines, including HCT116, HT29, RKO, DLD-1, SW403, SW1463, DiFi, and NCI-H508, were purchased from the ATCC and cultured in McCoy's 5A modified media. SNU-407 was purchased from AddexBio and cultured in RPMI1640 (Invitrogen). NCM356D non-transformed colonic epithelial cell line was purchased from INCELL and cultured in M3 media (INCELL). DR5-knockout (KO), PUMA-KO, and p53-KO HCT116 cells were described previously (20 (link), 27 (link)). Cells were authenticated by genotyping and analysis of protein expression by Western blotting, and routinely checked for Mycoplasma contamination by PCR. All cell lines were maintained at 37°C and 5% CO₂ atmosphere. Cell culture media were supplemented with 10% defined FBS, 100 U/mL penicillin, and 100 μg/mL streptomycin. For drug treatment, cells were plated in 12-well plates at 20%–30% density 24 hours before treatment. DMSO stocks of NEO2734, CPI-637, I-BET151, OTX015, 5-FU, oxaliplatin, and SN-38 were prepared and diluted in cell culture media before adding to cells. All drugs were purchased from commercial sources, except for NEO2734 and CPI-637 which were provided by NEOMED Therapeutics 1, Inc.

The use of Transfection and Cell Culture Leibovitz's 15 (L-15) culture media (ATCC) supplemented with 10% fetal bovine serum (FBS; ATCC) was used to cultivate the human rectal mucosa epithelial cell line (PriCells) and two types of human READ cell lines (SW837 and SW1463; ATCC). Logarithmic-phase cells were collected for further study. Genechem (Shanghai, China) provided both the pcDNA3.1 vector targeting SYNGAP1 and the empty vector. These plasmids were used in a Lipofectamine 3000 (Invitrogen) transfection to introduce them into the cells.

Human CRC Cell lines including SW480, HT-29, SW1463 and HCT116, as well as FHCs (fetal human cells), were obtained from ATCC. SW480 and SW1463 cells were seeded in ATCC-formulated Leibovitz’s L-15 medium. HT-29 and HCT116 cells were cultured in ATCC-formulated McCoy’s 5a medium modified. FHCs were maintained in DMEM:F12 medium. All the cell lines were incubated under 37 °C with 5% CO₂ in air atmosphere, and routinely subcultured every 3 days.