Cellular protein synthesis inhibition assay

The inhibitory effect of rGel on cellular protein synthesis was determined according to a previous report (Shin et al. 2016). Briefly, HCT116 and HCT-8 cell were seeded into 12-well plates at a density of 1 × 10⁵ cells per well and allowed to attach to the bottom of plates overnight in RPMI-1640 medium-containing 10% FBS at 37 °C. Then, the cells were treated with various concentrations (10⁻⁹, 10⁻⁸, 10⁻⁷, 10⁻⁶, and 10⁻⁵ M) of rGel for 48 or 72 h. The medium was removed and cells were washed twice with PBS, and 1% triton X-100 was added to lyse cells. Cell lysates were harvested and centrifuged at 12,000 rpm for 15 min. The protein content in the supernatant was determined by BCA protein assay and the relative cellular protein levels were calculated as the ratio of protein contents in treated groups to those of control groups.