p53-independent apoptosis mediated by tachpyridine, an anti-cancer iron chelator

Iron is involved in essential biochemical reactions ranging from respiratio n to DNA synthesis. Consequently, iron deprivation has been proposed as a s trategy for inhibition of tumor cell growth. We recently described a novel iron chelator, tachypyridine [N,N ' ,N " -tris(2-pyridylmethyl)-cis,cis-1,3 ,5-triaminocyclohexane], and demonstrated that it not only inhibited growth of cultured tumor cells, but was actively cytotoxic. Here we explore the m echanisms underlying tachpyridine cytotoxicity. Using several criteria, inc luding time-lapse video microscopy, DNA staining and TUNEL assays, tachpyri dine was shown to specifically induce apoptotic cell death. Further, unlike numerous cytotoxic chemotherapeutic drugs which induce apoptosis by activa ting p53-dependent pathways, tachpyridine-mediated cell death did not requi re p53 activation. Although immunoblotting revealed rapid accumulation of p 53 following treatment with tachpyridine, p21(WAF1) was not induced. Furthe r, neither cytotoxicity nor apoptosis required p53. p53 null human lung can cer H1299 cells transfected with an ecdysone-inducible p53 exhibited equiva lent sensitivity to tachpyridine in the presence and absence of p53, demons trating the lack of requirement for p53 in an isogenic cell system. Further , time-lapse video microscopy and TUNEL assays demonstrated that both p53 n ull and p53 wildtype cells underwent apoptotic cell death in response to ta chpyridine. In addition, in 55 human cancer cell lines the mean GI(50) of t achpyridine in cells with mutant p53 was virtually identical to the GI(50) in cells with wild-type p53. These results demonstrate that tachpyridine in itiates an apoptotic mode of cell death that does not require functional p5 3. Since over 50% of human tumors contain a functionally defective p53 that reduces sensitivity to commonly used chemotherapeutic agents, such as etop oside and cisplatin, the ability of tachpyridine to induce apoptosis indepe ndently of p53 may offer an advantage in antitumor therapy.