Organometallic Iridium(III) anticancer complexes with new mechanisms of action: NCI-60 screening, mitochondrial targeting, and apoptosis

Abstract

Platinum complexes related to cisplatin, cis-[PtCl2(NH3)2], are successful anticancer drugs; however, other transition metal complexes offer potential for combating cisplatin resistance, decreasing side effects, and widening the spectrum of activity. Organometallic half-sandwich iridium (Ir(III)) complexes [Ir(Cp(x))(XY)Cl](+/0) (Cp(x) = biphenyltetramethylcyclopentadienyl and XY = phenanthroline (1), bipyridine (2), or phenylpyridine (3)) all hydrolyze rapidly, forming monofunctional G adducts on DNA with additional intercalation of the phenyl substituents on the Cp(x) ring. In comparison, highly potent complex 4 (Cp(x) = phenyltetramethylcyclopentadienyl and XY = N,N-dimethylphenylazopyridine) does not hydrolyze. All show higher potency toward A2780 human ovarian cancer cells compared to cisplatin, with 1, 3, and 4 also demonstrating higher potency in the National Cancer Institute (NCI) NCI-60 cell-line screen. Use of the NCI COMPARE algorithm (which predicts mechanisms of action (MoAs) for emerging anticancer compounds by correlating NCI-60 patterns of sensitivity) shows that the MoA of these Ir(III) complexes has no correlation to cisplatin (or oxaliplatin), with 3 and 4 emerging as particularly novel compounds. Those findings by COMPARE were experimentally probed by transmission electron microscopy (TEM) of A2780 cells exposed to 1, showing mitochondrial swelling and activation of apoptosis after 24 h. Significant changes in mitochondrial membrane polarization were detected by flow cytometry, and the potency of the complexes was enhanced ca. 5× by co-administration with a low concentration (5 μM) of the γ-glutamyl cysteine synthetase inhibitor L-buthionine sulfoximine (L-BSO). These studies reveal potential polypharmacology of organometallic Ir(III) complexes, with MoA and cell selectivity governed by structural changes in the chelating ligands.

Figure 1

Ir^III complexes used in this work.

Figure 2

Heat map showing the log₁₀ GI₅₀ values for each iridium complex in the NCI-60 screen, where high intensity (red) cells indicate high activity and low intensity (blue) cells indicate low activity.

Figure 3

Mechanistic insights derived through use of the COMPARE algorithm. Bar chart summarizing the top 100 correlations, with r > 0.5, and known MoA for each of the four complexes. Correlations were segregated into six classes: oxidative stress, mitosis inhibitors, protein synthesis inhibitors, topoisomerase inhibitors, DNA antimetabolites, and DNA-interacting agents.

Figure 4

Detection of apoptosis in A2780 ovarian cancer cells after treatment with 1. (A–D) Control cells showing typical heterochromatin (H) and euchromatin (E) distributions, mitochondria (M), nuclear pores (NP), and rough endoplasmic reticulum (ER). (E–H) Cells exposed to 1 μM 1 for 24 h, showing swollen mitochondria (V), nuclear vacuoles (NV), and membrane blebbing (B). (I–L) Cells exposed to 5 μM 1 for 24 h, showing abnormal chromatin distributions and further vacuole formations.

Figure 5

Bar graph showing the mitochondrial membrane polarization, determined by a reduction in red (FL2) fluorescence, by complexes 1–4 against that of a negative and positive control (5 μM carbonyl cyanide m-chlorophenylhydrazone). p-values were calculated after a Welch t test with the negative control data. Each value is the mean taken from 3 replicates, with error bars for the standard deviation.

Figure 6

Enhancement of cytotoxicity by co-incubation of A2780 human ovarian cancer cells with complexes 2–4 and 5 μM L-BSO. Bars show IC₅₀ values with and without L-BSO, with the fold increase in activity above each bar. Each value is a mean of 3 replicates, with error bars for the standard deviation.