DNA damage response to the Mdm2 inhibitor nutlin-3.
Mdm2 inhibitors represent a promising class of p53 activating compounds that may be useful in cancer treatment and prevention. However, the consequences of pharmacological p53 activation are not entirely clear. We observed that Nutlin-3 triggered a DNA damage response in azoxymethane-induced mouse AJ02-NM(0) colon cancer cells, characterized by the phosphorylation of H2AX (at Ser-139) and p53 (at Ser-15). The DNA damage response was highest in cells showing robust p53 stabilization, it could be triggered by the active but not the inactive Nutlin-3 enantiomer, and it was also activated by another pharmacological Mdm2 inhibitor (Caylin-1). Quantification of gamma H2AX-positive cells following Nutlin-3 exposure showed that approximately 17% of cells in late S and G2/M were mounting a DNA damage response (compared to a approximately 50% response to 5-fluorouracil). Nutlin-3 treatment caused the formation of double-strand DNA strand breaks, promoted the formation of micronuclei, accentuated strand breakage induced by doxorubicin and sensitized the mouse colon cancer cells to DNA break-inducing topoisomerase II inhibitors. Although the HCT116 colon cancer cells did not mount a significant DNA damage response following Nutlin-3 treatment, Nutlin-3 enhanced the DNA damage response to the nucleotide synthesis inhibitor hydroxyurea in a p53-dependent manner. Finally, p21 deletion also sensitized HCT116 cells to the Nutlin-3-induced DNA damage response, suggesting that cell cycle checkpoint abnormalities may promote this response. We propose that p53 activation by Mdm2 inhibitors can result in the slowing of double-stranded DNA repair. Although this effect may suppress illegitimate homologous recombination repair, it may also increase the risk of clastogenic events.
Department of Molecular & Cell Biology U3125, University of Connecticut, Storrs, CT 06269, USA.
Rajeev
R
Verma
Marc J
MJ
Rigatti
Glenn S
GS
Belinsky
Cassandra A
CA
Godman
Charles
C
Giardina
D000818:Animals; D004249:DNA Damage / Q000187:drug effects* / Q000502:physiology; D045325:HCT116 Cells; D006801:Humans; D007093:Imidazoles / Q000633:toxicity*; D051379:Mice; D010879:Piperazines / Q000633:toxicity*; D051736:Proto-Oncogene Proteins c-mdm2 / Q000037:antagonists & inhibitors* / Q000502:physiology; D013237:Stereoisomerism; D014407:Tumor Cells, Cultured
D007093:Imidazoles; D010879:Piperazines; C482205:nutlin 3; C497207:Mdm2 protein, mouse; D051736:Proto-Oncogene Proteins c-mdm2