Apoptosis Protease Activator Protein-1 Expression Is Dispensable for Response of Human Melanoma Cells to Distinct Proapoptotic Agents

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Abstract
Loss of expression of the apoptosis protease activator protein-1 (APAF-1) in human melanoma is thought to promote resistance to programmed cell death by preventing caspase-9 activation. However, the role of the APAF-1–dependent pathway in apoptosis activated by cellular stress and/or DNA damage has been recently questioned. We investigated APAF-1 expression in a large panel of human melanomas and assessed cellular response to several proapoptotic agents in tumors expressing or lacking APAF-1 protein. In two melanomas with wild-type p53 but with differential expression of APAF-1, treatment with camptothecin, celecoxib, or an nitric oxide synthase inhibitor (1400W) significantly modulated expression of 36 of 96 genes in an apoptosis-specific cDNA macroarray, but APAF-1 mRNA levels were not induced (in APAF-1− cells) nor up-regulated (in APAF-1+ cells), a finding confirmed at the protein level. Treatment with cisplatin, camptothecin, etoposide, betulinic acid, celecoxib, 1400W, and staurosporine promoted enzymatic activity not only of caspases -2, -8, and -3 but also of caspase-9 in both APAF-1+ and APAF-1− tumor cells. Moreover, drug-induced caspase-9 enzymatic activity could be not only partially but significantly reduced by caspase-2, -3, and -8 –specific inhibitors in both APAF-1+ and APAF-1− tumor cells. In response to 1 to 100 μmol/L of cisplatin, camptothecin, or celecoxib, APAF-1+ melanomas (n = 12) did not show significantly increased levels of apoptosis compared with APAF-1− tumors (n = 7), with the exception of enhanced apoptosis in response to a very high dose (100 μmol/L) of etoposide. These results suggest that the response of human melanoma cells to different proapoptotic agents may be independent of their APAF-1 phenotype.