The Heat Shock Protein 90-Targeting Drug Cisplatin Selectively Inhibits Steroid Receptor Activation

Abstract

Cisplatin is an antineoplastic drug that binds to DNA, thereby inhibiting cell division and tumor growth. Cisplatin may also disrupt the function of some proteins, including heat shock protein 90 (Hsp90). We report that cisplatin dose-dependently inhibited transcriptional activity of the androgen receptor and the glucocorticoid receptor (GR) in transient reporter assays. A truncated, hormone-independent GR was only partially inhibited at significantly higher doses of cisplatin. Cisplatin treatment of neuroblastoma cells led to an immediate inhibition of hormone binding by GR, followed by proteasome-dependent degradation of the receptor. Other Hsp90-regulated proteins, i.e. the phosphokinases raf-1, lck, and c-src, were not affected, indicating a specific functional interference of cisplatin with the steroid receptors GR and androgen receptor. Cisplatin did not elicit a stress response, in contrast to geldanamycin. Immunoprecipitation revealed that cisplatin disrupts binding of GR to Hsp90. Moreover, cisplatin-treated Hsp90 was unable to associate with untreated ligand binding domain of GR. Reticulocyte lysate was able to restore hormone binding of GR in vitro, but not when the lysate was pretreated with geldanamycin. Our data reveal that cisplatin influences steroid receptors also independently of its DNA-mediated effects and, thus, suggest a novel modes of action for this cytostatic drug.

Androgen receptor activity is inhibited in response to genotoxic agents in a p53-independent manner

“…The ligand-binding ability of exogenously expressed GR was shown to be significantly inhibited by cisplatin, which led to the hypothesis that cisplatin-induced inhibition of Hsp90 only affects a fraction of Hsp90 client proteins, for example, steroid receptors (Rosenhagen et al, 2003). Our ligand-binding assay performed in LNCaP cells with endogenous expression of AR showed that cisplatin only caused a slight reduction in the ligand-binding activity of this receptor.…”

“…It should be noted that our cell fractionation experiment (Figure 3c) did not reveal a subcellular redistribution of Ser81 phosphorylated AR at early time points in response to genotoxic stress (data not shown). The activity of exogenously expressed AR and glucocorticoid receptor (GR) in neuroblastoma cells has been demonstrated to be inhibited in response to cisplatin, and for the GR, this was linked to the cisplatin-induced inhibition of the molecular chaperone Hsp90 (Rosenhagen et al, 2003). As the AR is an Hsp90 client protein (Georget et al, 2002), it is possible that loss of AR activity and loss of protein expression at later time points, in response to cisplatin, could be related to a cisplatin-induced inhibition of Hsp90 in LNCaP cells.…”

Dicoumarol down-regulates human PTTG1/Securin mRNA expression through inhibition of Hsp90

“…Overall, effects of dicoumarol, 17AAG, and novobiocin were somewhat variable when compared with each other. However, differences in inhibitory effect profiles are common among Hsp90 inhibitors and may reflect differences in mechanisms of action on the Hsp90 molecule (33).…”

HSP90 regulates cell survival via inositol hexakisphosphate kinase-2

“…6c). Half-maximal inhibition of binding occurs with Ϸ100 M NB and 1 M CP, comparable to their apoptotic potencies (36,37).…”

“…However, CP and NB bind to the C terminus of HSP90 both in vitro and in vivo (35)(36)(37). CP binds HSP90, decreasing the transcriptional activity of androgen and glucocorticoid receptors although not affecting other HSPregulated proteins, such as the phosphokinases Raf-1, Lck, and c-Src (37).…”