Reduction of etoposide induced cell killing by hyperthermia can occur without changes in etoposide transport or DNA topoisomerase II activity

Abstract

We investigated the modification of etoposide (i.e. VP-16)-induced cell killing by hyperthermia in a radioresistant human melanoma (Sk-Mel-3) and a human normal (AG1522) cell line. VP-16, a DNA topo II poison, was given as a 1 h exposure at variable doses up to 35 microM; hyperthermia was given either before or following VP-16 treatment. Hyperthermic treatment comprised one of the following: 41 degrees C for 8 h, 42 degrees C for 2 h or 45 degrees C for 15 min. Hyperthermia preceding VP-16 treatment reduced the cytotoxicity of the latter; the reduction of VP-16 cytotoxicity was directly proportional to the severity of the hyperthermic treatment. For a particular combination of hyperthermic dose and VP-16 concentration, generally similar responses were seen in both cell lines. There were no effects on VP-16 cytotoxicity when both Sk-Mel-3 and AG1522 cells were heated at 41 degrees C for 8 h following treatment with VP-16. However, heating both cell lines at 45 degrees C for 15 min following VP-16 treatment again reduced the amount of cytotoxicity associated with VP-16. In addition, we found that a preceding exposure to 45 degrees C, 15 min heating did not affect either cellular accumulation or efflux of [3H]VP-16 in both cell lines. This suggested that the reduction in VP-16 cytotoxicity observed under those conditions was not due to a modification of VP-16 transport. We found no differences between the catalytic activities of topo II extracted from nuclei of Sk-Mel-3 and AG1522 cells that were either heated at 45 degrees C for 15 min or that were not subjected to such treatment. These results therefore suggested that the substantial reduction of cytotoxicity seen when 45 degrees C, 15 min heating preceded VP-16 treatment was also not due to an effect on topo II catalytic activity. Our results therefore demonstrate that hyperthermia, given either before or after VP-16, can actually reduce the amount of VP-16 cytotoxicity and that this can occur without any overt changes in VP-16 accumulation and efflux or in topo II catalytic activity.

High Temperature Drives Topoisomerase Mediated Chromosomal Break Repair Pathway Choice

“…Notably, our viability data and the hyperthermia conditions used here are consistent with the published reports showing that hyperthermia protects cells from the cytotoxic effect of topoisomerase inhibitors [ 33 , 34 , 35 , 36 ]. The possibility that hyperthermia reduced the formation of TOPcc was excluded since multiple independent reports have shown that hyperthermia did not reduce the activity of TOP1, TOP2 or the level of TOP1cc and TOP2cc [ 33 , 35 , 36 ]. On the contrary, it has been suggested that hyperthermia induces the formation of stable TOP1cc during early S-phase [ 61 ].…”

“…In contrast to radiation and several chemotherapeutic agents, hyperthermia failed to sensitize cells to topoisomerase (TOP) inhibitors and instead exerted a protective effect. This paradigm remains unexplained [ 33 , 34 , 35 , 36 ]. Topoisomerases remove topological stress by cleaving and resealing DNA strands.…”

Etoposide toxicity on human neuroblastoma cells in vitro is enhanced by preceeding hyperthermia

“…Etoposide differs from almost all other cytostatics in that thermal enhancement of its effect has not been observed [2,4]. Tumour cells were even found to be protected from etoposide by heat [3,5].…”

“…Enhancement of the cytotoxicity of anticancer drugs has been shown in various in vitro and in vivo models, particularly for alkylants [1]. For etoposide, this effect could never be demonstrated [2]. Some data show even a protective effect of heat against etoposide toxicity [3].…”

In vitro test-system for chemo- and thermosensitivity: an analysis of survival fractions and cell-cycle distributions in human Ewing's sarcomas as a modelfor tumors in Pediatric Oncology

“…The fact that cDDP can suppress the accumulation of heat shock protein HSP-72 as reported by Matsumoto et al indicate that higher treatment efficacy under hyperthermic conditions can also be explained by a drug-related inhibition of cell protection [21]. 13,15,22,24]. These conflicting data show that various tumor cell specific parameters are of great importance for treatment response to hyperthermia.…”