The potential for cobalt(III) complexes in medicine, as chaperones of bioactive ligands, and to target tumours through bioreductive activation, has been examined over the past 20 years. Despite this, chemical properties such as reduction potential and carrier ligands required for optimal tumour targeting and drug delivery have not been optimised. Here we review the chemistry of cobalt(III) drug design, and recent developments in the understanding of the cellular fate of these drugs.

Inorganic complexes are versatile platforms for the development of potent and selective pharmaceutical agents. Cobalt possesses a diverse array of properties that can be manipulated to yield promising drug candidates. Investigations into the mechanism of cobalt therapeutic agents can provide valuable insight into the physicochemical properties that can be harnessed for drug development. This review presents examples of bioactive cobalt complexes with special attention to their mechanisms of action. Specifically, cobalt complexes that elicit biological effects through protein inhibition, modification of drug activity, and bioreductive activation are discussed. Insights gained from these examples reveal features of cobalt that can be rationally tuned to produce therapeutics with high specificity and improved efficacy for the biomolecule or pathway of interest.

We report a potential means of selectively delivering matrix metalloproteinase (MMP) inhibitors to target tumour sites by use of a bioreductively activated Co(III) carrier system. The carrier, comprising a Co(III) complex of the tripodal ligand tris(methylpyridyl)amine (tpa), was investigated with the antimetastatic MMP inhibitor marimastat (mmstH(2)). The X-ray crystal structure of [Co(mmst)(tpa)]ClO(4) x 4H(2)O was determined and two-dimensional NMR revealed the existence of two isomeric forms of the complex in solution. Electrochemical analysis showed that the reduction potential of the complex is suitable for it to be bioreductively activated at hypoxic tumour sites. In vitro assays confirmed the stability of the prodrug in solution prior to reduction and revealed very low cytotoxicity against A2780 cells. In vivo testing in mice showed a higher level of tumour-growth inhibition by the complex than by free marimastat. Both free marimastat and and its Co(III) complex increased metastasis in the model used, with the complex significantly more active.

The rational design of prodrugs for selective accumulation and activation in tumor microenvironments is one of the most promising strategies for minimizing the toxicity of anticancer drugs. Manipulation of the charge of the prodrug represents a potential mechanism to selectively deliver the prodrug to the acidic tumor microenvironment. Here we present delivery of a fluorescent coumarin using a cobalt(III) chaperone to target hypoxic regions, and charged ligands for pH selectivity. Protonation or deprotonation of the complexes over a physiologically relevant pH range resulted in pH dependent accumulation of the fluorophore in colon cancer cells. Furthermore, in a spheroid solid tumor model, the anionic complexes exhibited preferential release of the fluorophore in the acidic/hypoxic region. By fine-tuning the physicochemical properties of the cobalt-chaperone moiety, we have demonstrated selective drug release in the acidic and hypoxic tumor microenvironment.

The penetration of anthraquinones and their platinum complexes into cancer cell spheroids reveals that they model well the distribution of such compounds in solid tumours and that the proportion of the compound that accumulates deep in the spheroid is inversely related to the rate of cellular uptake which is affected by the charge of the compound.

The development of complexes that allow the monitoring of the release and distribution of fluorescent models of anticancer drugs initially bound to cobalt(III) moieties is reported. Strong quenching of fluorescence upon ligation to cobalt(III) was observed for both the carboxylate- and the hydroximate-bound fluorophores as was the partial return of fluorescence following addition of ascorbate and cysteine. The extent of the increase in the fluorescence intensity observed following addition of these potential reductants is indicative of the fluorophore being displaced from the complex by the action of ascorbate or cysteine, by ligand exchange. The cellular distribution of the fluorescence revealed that coordination to cobalt can dramatically alter the subcellular distribution of a bound fluorophore. This work shows that fluorescence can be an effective means of monitoring these agents in cells, and of determining their sites of activation. The results also reveal that the cytotoxicity of such agents correlates with their uptake and distribution patterns and that these are influenced by the types of ligands attached to the complex.

The transition from a non-invasive to an invasive phenotype is an essential step in tumor metastasis. The Snail family of transcription factors (TFs) is known to play a significant role in this transition. These TFs are zinc fingers that bind to the CAGGTG Ebox consensus sequence. CoIII-Ebox is a cobalt(III) complex attached to an Ebox oligonucleotide that confers specificity towards Snail TFs. CoIII-Ebox has been shown to inhibit Snail-mediated embryonic neural crest development in Xenopus laevis, but its efficacy in inhibiting Snail-induced cancer cell invasiveness has not been explored. Here, we describe the efficacy of CoIII-Ebox in inhibiting the invasive aspects of heregulin β1(HRG)-treated breast cancer cells. CoIII-Ebox was found to inhibit the capacity of Snail to repress target genes after HRG induction. Snail inhibition by CoIII-Ebox reduced the invasive propensity of cells in 2D and 3D, thereby demonstrating promise in inhibiting metastasis.

A peptide tagged at the N-terminus with FITC, at the C-terminus with coumarin-343, and incorporating a sequence selectively cleaved by the matrix metalloproteinase, MMP-2, was synthesized to investigate the effect of peptide cleavage on both cellular accumulation and distribution in cancer cell spheroids. The peptide was shown by HPLC and mass spectroscopy to be cleaved in the presence of MMP-2 at the expected site. The cellular and spheroid distribution of each of the fragments was monitored using confocal fluorescence microscopy. The intact peptide had minimal accumulation in 2D-cultured DLD-1 cells that do not express MMP-2 in these conditions. Following addition of serum containing MMP-2 to the cell media, the cleaved C-terminal fragment was seen to enter the cells, while the N-terminal fragment remained extracellular, evidently blocked by the presence of the FITC group. 3D culture of DLD-1 cells as spheroids resulted in measurable MMP-2 activity. Different distribution patterns of the two fluorophores were seen in spheroids treated with the intact peptide, consistent with cleavage occurring. Different rates of accumulation of each of the fragments were observed within the spheroid over time, which is attributed to the extent of accumulation and sequestration of the fragments by cells residing in the periphery of the spheroids. The outcomes suggest that tumor-associated enzymes have the potential to modify the distribution of peptides and peptide fragments in solid tumors by modifying the cellular uptake of those peptides.