p21WAF1 Prevents Down-modulation of the Apoptotic Inhibitor Protein c-IAP1 and Inhibits Leukemic Apoptosis

Abstract

Background: Prior studies indicate that leukemias expressing high levels of the p21WAF1 cell cycle inhibitor have a poorer prognosis than p21WAF1-negative leukemias. Although p21WAF1 is up-regulated by p53 in the setting of DNA damage, the prognostic significance of p21WAF1 is independent of p53 status. The molecular basis of the negative prognostic effect of p21WAF1 remains obscure, but it is believed to result from decreased apoptosis of p21WAF1-expressing leukemias. Materials and Methods: We studied the effects of p21WAF1 on apoptosis of K562 leukemic cells, which lack wild-type p53 and do not express endogenous p21WAF1. An inducible p21WAF1 system was used and the effect of p21WAF1 induction on susceptibility to etoposide-mediated apoptosis was measured. Results: p21WAF1 decreased apoptotic death of K562 leukemic cells in response to etoposide. Analysis of intermediaries in the apoptotic pathway indicated that p21WAF1 had no effect on cytochrome c release or cleavage of procaspase-3. In contrast, p21WAF1 was protective against cleavage of caspase targets poly(ADP-ribose)polymerase (PARP), retinoblastoma protein (Rb), and lamin. The expression of the inhibitor of apoptosis protein c-IAP1, which inhibited the function of executioner caspases 3 and 7, was studied. c-IAP1 protein expression was found to be present in a majority of leukemic blasts from untreated patients, but absent in normal differentiating myeloid progenitor cells. In K562 cells, treatment with etoposide in the absence of p21WAF1 induction resulted in post-transcriptional down-modulation of c-IAP1 levels. c-IAP1 loss involved proteasomal, rather than caspase, degradation pathways. Expression of p21WAF1 sustained c-IAP1 protein levels in the presence of etoposide. Conclusions: Etoposide-mediated apoptosis involves down-modulation of the anti-apoptotic protein c-IAP1. Our findings support the hypothesis that p21WAF1 contributes to leukemic chemoresistance by stabilizing c-IAP1 levels in the presence of chemotherapy.