Figure S1. 5CHQ suppresses etoposide-induced apoptosis in wt-p53-expressing cells; Figure S2. Effect of 5CHQ on p53's DNA-binding to CDKN1A and BBC3 promoter oligonucleotides; Figure S3. 5CHQ by itself activates p53-mediated transcription; Figure S4. Effect of 5CHQ on cell cycle distribution; Figure S5. MTD study of 5CHQ; Figure S6. The technique of SBI to avoid BM aplasia; Figure S7. Additional TBI experiments; Figure S8. Sixty-day survival test of C57BL/6 mouse after 20 Gy-SBI; Figure S9. Immunoblotting also revealed the transactivation-modulating activity of 5CHQ; Figure S10. Dose response of 5CHQ on etoposide-induced apoptosis in FM3A cells; Figure S11. 5CHQ relieves the decrease of the stem cell marker Lgr5 mRNA expression after SBI.
ARTICLE ABSTRACT
Inhibiting p53-dependent apoptosis by inhibitors of p53 is an effective strategy for preventing radiation-induced damage in hematopoietic lineages, while p53 and p21 also play radioprotective roles in the gastrointestinal epithelium. We previously identified some zinc(II) chelators, including 8-quinolinol derivatives, that suppress apoptosis in attempts to discover compounds that target the zinc-binding site in p53. We found that 5-chloro-8-quinolinol (5CHQ) has a unique p53-modulating activity that shifts its transactivation from proapoptotic to protective responses, including enhancing p21 induction and suppressing PUMA induction. This p53-modulating activity also influenced p53 and p53-target gene expression in unirradiated cells without inducing DNA damage. The specificity of 5CHQ for p53 and p21 was demonstrated by silencing the expression of each protein. These effects seem to be attributable to the sequence-specific alteration of p53 DNA-binding, as evaluated by chromatin immunoprecipitation and electrophoretic mobility shift assays. In addition, 5-chloro-8-methoxyquinoline itself had no antiapoptotic activity, indicating that the hydroxyl group at the 8-position is required for its antiapoptotic activity. We applied this remarkable agonistic activity to protecting the hematopoietic and gastrointestinal system in mouse irradiation models. The dose reduction factors of 5CHQ in total-body and abdominally irradiated mice were about 1.2 and 1.3, respectively. 5CHQ effectively protected mouse epithelial stem cells from a lethal dose of abdominal irradiation. Furthermore, the specificity of 5CHQ for p53 in reducing the lethality induced by abdominal irradiation was revealed in Trp53-KO mice. These results indicate that the pharmacologic upregulation of radioprotective p53 target genes is an effective strategy for addressing the gastrointestinal syndrome. Mol Cancer Ther; 17(2); 432–42. ©2017 AACR.See all articles in this MCT Focus section, “Developmental Therapeutics in Radiation Oncology.”
