Chemical Modification of Auranofin Yields a New Family of Anticancer Drug Candidates: The Gold(I) Phosphite Analogues

Abstract

A panel of four novel gold(I) complexes, inspired by the clinically established gold drug auranofin (1-Thio-β-D-glucopyranosatotriethylphosphine gold-2,3,4,6-tetraacetate), was prepared and characterized. All these compounds feature the replacement of the triethylphosphine ligand of the parent compound auranofin with a trimethylphosphite ligand. The linear coordination around the gold(I) center is completed by Cl^-, Br^-, I^- or by the thioglucose tetraacetate ligand (SAtg). The in-solution behavior of these gold compounds as well as their interactions with some representative model proteins were comparatively analyzed through ³¹PNMR and ESI-MS measurements. Notably, all panel compounds turned out to be stable in aqueous media, but significant differences with respect to auranofin were disclosed in their interactions with a few leading proteins. In addition, the cytotoxic effects produced by the panel compounds toward A2780, A2780R and SKOV-3 ovarian cancer cells were quantitated and found to be in the low micromolar range, since the IC₅₀ of all compounds was found to be between 1 μM and 10 μM. Notably, these novel gold complexes showed large and similar inhibition capabilities towards the key enzyme thioredoxin reductase, again comparable to those of auranofin. The implications of these results for the discovery of new and effective gold-based anticancer agents are discussed.

Keywords: anticancer compounds; auranofin; metal-based drugs; phosphite compounds.

Figure 1

Auranofin structure.

Figure 2

The investigated compounds.

Figure 3

(A) Deconvoluted ESI-MS mass spectra of human serum albumin (HSA) 10⁻⁶ M in 20 mM ammonium acetate solution at pH 6.8, incubated at 37 °C for 5 min with (B) AuP(OCH₃)₃Cl, (C) AuP(OCH₃)₃Br, (D) AuP(OCH₃)₃I, (E) AuP(OCH₃)₃SAtg solution in a 1:3 protein-to-gold ratio; 0.1% v/v of formic acid was added just before injection.

Figure 4

(A) Deconvoluted ESI-MS mass spectra of human carbonic anhydrase I (hCA I) 10⁻⁶ M in 20 mM ammonium acetate solution at pH 6.8 and incubated at 37 °C for 24 h with (B) AuP(OCH₃)₃Cl, (C) AuP(OCH₃)₃Br, (D) AuP(OCH₃)₃I, (E) AuP(OCH₃)₃SAtg solution in a 1:1 protein-to-gold ratio.

Figure 5

Analysis of gold content in A2780 cells following treatment; the gold content is indicated as a ratio between µg of gold and total protein amount. Histograms report the mean values ± SD.

Figure 6

Caspase-3 activity shown by fluorescence-activated cell sorting analysis using FAM FLICA in A2780 cells treated for 72 h with gold compounds concentration corresponding to their 72 h exposure IC₅₀-dose. Histograms report the mean values ± SD. P2 represents the percentage of the cell population with activated caspase-3 (P2 population). The statistical analysis was carried out using one-way ANOVA test followed by Tuckey’s multiple comparisons test using Graphpad Prism v 6.0 (* p < 0.05, ** p < 0.01, *** p < 0.001).

Figure 7

TrxR enzyme inhibition assay was performed after 24 h of treatment using a commercial thioredoxin reductase assay kit, whereby histograms report the residual TrxR enzyme activity in gold-treated cells expressed in percentage with respect to control cells. The analysis was performed in triplicate. The statistical analysis was carried out using one-way ANOVA test followed by Tukey’s multiple comparisons test using Graphpad Prism software v 6.0 (** p < 0.01, **** p < 0.0001).