MDM2 mediated nuclear exclusion of p53 attenuates etoposide-induced apoptosis in neuroblastoma cells

The p53 gene in neuroblastoma tumors (NB) is rarely mutated but the protein accumulates in the cytoplasm. Because p53 can mediate the cytotoxic effect s of chemotherapeutic agents, it is important to determine whether accumula tion of p53 in the cytoplasm impairs p53 function. Data presented here indi cate that hyperactive nuclear export of p53 suppresses etoposide-induced ap optosis but does not prevent growth arrest. We compared p53 function in a p air of NB subclones. Our data show etoposide induces complete trans-locatio n of p53 to the nucleus and activation of apoptosis in the neuroblastic NB cell line SH-SY5Y (N-type), which expresses low levels of MDM2. However, in Schwann cell-like SH-EP1 cells (S-type), which have up to 10-fold higher l evels of MDM2, p53 accumulates in the cytoplasm and the cells are extremely resistant to etoposide-induced apoptosis. Notably, when MDM2 expression is inhibited in S-type cells, with a phosphorothioated antisense oligonucleot ide (AS5), then p53 accumulates in the nucleus and the SH-EP1 cells undergo apoptosis. Surprisingly, induction of p21 and G(1)-arrest are not attenuat ed in S-type cells, despite the predominantly cytoplasmic location of p53. Whereas, G(1)-arrest is attenuated in the SH-SY5Y cells, which have high le vels of nuclear p53. Taken together, these findings suggest attenuation of G(1)-arrest is related to the differentiation status of neuroblastomas and occurs downstream of p53 nuclear accumulation. These results demonstrate fo r the first time that hyperactive nuclear export of p53 attenuates chemothe rapy-induced apoptosis in NB cells, and our findings suggest that inhibitor s of MDM2 may enhance the therapeutic efficacy of etoposide by promoting ap optosis rather than trans-differentiation.