5-Fluorouracil-induced RNA stress engages a TRAIL-DISC-dependent apoptosis axis facilitated by p53

Abstract

Despite recent advances in targeted therapeutics, administration of 5-fluorouracil (5-FU) remains a common clinical strategy for post-surgical treatment of solid tumors. Although it has been proposed that RNA metabolism is disturbed by 5-FU treatment, the key cytotoxic response is believed to be enzymatic inhibition of thymidylate synthase resulting in nucleotide pool disproportions. An operating p53 tumor suppressor signaling network is in many cases essential for the efficiency of chemotherapy, and malfunctions within this system remain a clinical obstacle. Since the fate of chemotherapy-insensitive tumor cells is rarely described, we performed a comparative analysis of 5-FU toxicity in p53-deficient cells and conclude that p53 acts as a facilitator rather than a gatekeeper of cell death. Although p53 can act as a regulator of several cellular stress responses, no rerouting of cell death mode was observed in absence of the tumor suppressor. Thus, the final death outcome of 5-FU-treated p53−/− cells is demonstrated to be caspase-dependent, but due to a slow pace, accumulation of mitochondrial reactive oxygen species contributes to necrotic characteristics. The oligomerization status of the p53 target gene DR5 is determined as a significant limiting factor for the initiation of caspase activity in an intracellular TRAIL-dependent manner. Using several experimental approaches, we further conclude that RNA- rather than DNA-related stress follows by caspase activation irrespectively of p53 status. A distinct 5-FU-induced stress mechanism is thereby functionally connected to a successive and discrete cell death signaling pathway. Finally, we provide evidence that silencing of PARP-1 function may be an approach to specifically target p53-deficient cells in 5-FU combinatorial treatment strategies. Together, our results disclose details of impaired cell death signaling engaged as a consequence of 5-FU chemotherapy. Obtained data will contribute to the comprehension of factors restraining 5-FU efficiency, and by excluding DNA as the main stress target in some cell types they propose alternatives to currently used and suggested synergistic treatment regimens.

Detection of Superoxide Alterations Induced by 5-Fluorouracil on HeLa Cells with a Cell-Based Biosensor

“…The determination of dynamic radical changes in cellular microenvironment with non-invasive approaches is an essential mean for assessing the effectiveness of anticancer agents on cancer treatment [45][46][47][48]. As mentioned above, 5-FU's chemotherapeutic properties are attributed to thymidylate synthase (TS) activity inhibition, to mitochondrial ROS generation and the caspases cascade activation [49]. Our findings demonstrate that 5-FU differentially affects superoxide production and caspase-3 activation when applied in cytotoxic concentrations against HeLa cells.…”

Loss of Uracil DNA Glycosylase Selectively Resensitizes p53-Mutant and -Deficient Cells to 5-FdU

“…Recently, several studies have observed that the cytotoxicity of 5-FU is more dependent on its incorporation into RNA than its inhibition of TS, diminishing its effect on DNA (21,(34)(35)(36)(37). In addition, activation of p53 following 5-FU exposure has been identified as working through RNA mechanisms (43,44). Because UDG recognizes only DNA lesions, it is not surprising that depletion of UDG does not significantly alter cellular responses to agents that primarily affect RNA function.…”

Aberrant DR5 transport through disruption of lysosomal function suggests a novel mechanism for receptor activation

“…Moreover, retinoids and interferons have been demonstrated to stimulate cell death by means of TRAIL [ 12 ]. Similarly, our recent work emphasized that 5-Fluorouracil (5-FU) can induce apoptosis following the activation of DRs via an intracellular TRAIL-stimulated mechanism [ 13 ]. In the present study, initiated with the aim to analyze crosstalk between apoptosis and autophagy, we demonstrate that cell death generated by 5-FU requires cytoplasmic transport of DR5, which in turn seems to be tightly associated with free cholesterol.…”