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

Abstract

Iron is involved in essential biochemical reactions ranging from respiration to DNA synthesis. Consequently, iron deprivation has been proposed as a strategy 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 mechanisms underlying tachpyridine cytotoxicity. Using several criteria, including time-lapse video microscopy, DNA staining and TUNEL assays, tachpyridine was shown to specifically induce apoptotic cell death. Further, unlike numerous cytotoxic chemotherapeutic drugs which induce apoptosis by activating p53-dependent pathways, tachpyridine-mediated cell death did not require p53 activation. Although immunoblotting revealed rapid accumulation of p53 following treatment with tachpyridine, p21(WAF1) was not induced. Further, neither cytotoxicity nor apoptosis required p53. p53 null human lung cancer H1299 cells transfected with an ecdysone-inducible p53 exhibited equivalent sensitivity to tachpyridine in the presence and absence of p53, demonstrating the lack of requirement for p53 in an isogenic cell system. Further, time-lapse video microscopy and TUNEL assays demonstrated that both p53 null and p53 wild-type cells underwent apoptotic cell death in response to tachpyridine. In addition, in 55 human cancer cell lines the mean GI(50) of tachpyridine in cells with mutant p53 was virtually identical to the GI(50) in cells with wild-type p53. These results demonstrate that tachpyridine initiates an apoptotic mode of cell death that does not require functional p53. Since over 50% of human tumors contain a functionally defective p53 that reduces sensitivity to commonly used chemotherapeutic agents, such as etoposide and cisplatin, the ability of tachpyridine to induce apoptosis independently of p53 may offer an advantage in anti-tumor therapy.