Effects of Cer-RUB on the protein expression of p53 and p53-responsive genes. A, Western blots and B, Protein expression levels. A2780 and OVCAR-3 cells were pretreated with Cer-RUB nanomicelles (1 μM), Cer-BSA complex (1 μM) or in vehicle for 4 h, and then co-exposed to CDDP (1 μM) during the last 48 h. Equal amounts of total extracted proteins (50 μg/lane) were used for immunoblotting. #, p<0.001 compared to A2780 cells in vehicle; **, p<0.001 compared to corresponding cells in vehicle. C, Effects of Short-term treatment on phosphorylated p53 (Ser15) levels. Mice bearing OVCAR-3 tumors were treated with either NBD Cer-BSA (1 mg/kg, i.p.) or NBD Cer-RUB (1 mg/kg, i.p.) for 6 h. *, p<0.001 compared with mice treated with NBD Cer-BSA. D, Phosphorylated p53 (Ser15) levels in tumors with prolonged treatments. Mice bearing OVCAR-3 tumors were treated with either CDDP (1 mg/kg, i.p. once every 6-days) or Cer-RUB (1 mg/kg, i.p., once every 3-days) alone or in combination for 24 days. *, p<0.001 compared to CDDP; **, p<0.001 compared to Cer-RUB nanomicelles. E, pp53 (Ser15) and p21 in colons of transgenic mice. The genotypes of mice used in treatments were verified by extracted tail DNA with PCR. Cer-RUB (1 mg/kg, i.p., twice in 6 days) was administered to wild-type (WT) or HTZ p53 R172H/+ transgenic mice. #, p<0.001 compared to WT mice; *, p<0.001 compared to corresponding mice treated with saline. F, Immunohistochemistry of colon of p53 R172H/+ mice (x 200). Green FL, Cer-Alexa Fluor®488-conjugated antibodies; red FL, p21-Alexa Fluor®555-conjugated antibodies. Nuclei were counterstained with DAPI (blue).