Figure 5. UbcH10-deficient DLD1 cells were more susceptible to ALLN.

(A) ALLN was identified as an effective drug for UbcH10-/- DLD1 cells. Approximately 1 × 10⁴ cells of the indicated cell lines were seeded in 96-well plates and treated with various drugs for 24 hours. The viability of the cells was measured 12 hours later using the CCK8 method. The concentration of ALLN was 10 μg/mL (26 μM). (B) ALLN was effective at decreasing the viability of the UbcH10-/- DLD1 cells at a concentration of 26 μM. Approximately 1 × 10⁴ cells of the indicated cell lines were seeded in 96-well plates and treated with various concentrations of ALLN for 24 hours. The viability of the cells was measured immediately using the CCK8 method. (B) Colony formation assay to determine the sensitivity of the indicated cell lines to ALLN. Approximately 1 × 10³ cells of the indicated cell lines were seeded in 24-well plates and treated with the indicated concentrations of ALLN for 24 hours. The drug was then removed, and the cells were cultured for an additional two weeks. The cell colonies were stained with crystal violet and photographed. (D–F) Xenograft experiments were performed by injecting wild-type DLD1 or UbcH10^-/- DLD1 cells (5 × 10⁶) into the flanks of 6-week-old nude mice to form tumors. The mice were then randomly assigned to the vehicle control group or the ALLN treatment group. The mice in the vehicle control group were intraperitoneally injected with vehicle (saline and ethanol), and the mice in the treatment group were intraperitoneally injected with 10 mg/kg/day of ALLN. The injections were performed once each day for two weeks. The tumor volumes were measured every 3 days. At the end of the experiment, the mice were euthanized, and the tumors were weighed on an electronic balance. The numbers of colonies are expressed as the means ± S.E. from three assays. *P< 0.05, **P< 0.01, ***P< 0.001 compared with controls.