Development and Characterization of Boronate-Masked Prochelators for Peroxide-Triggered Metal Chelation

Chelating agents that can passivate the redox reactivity of transgressing iron have shown promising potential to minimize damage associated with oxidative stress in diseases. Systemic administration of these chelating agents, however, raises safety concerns due to the potential risks of indiscriminate extraction of metals from critical metalloproteins and inhibition of metalloenzymes. To overcome this challenge of unselective metal chelation, we have pursued a prodrug strategy for two extensively studied iron chelators, salicylaldehyde isonicotinic hydrazone (SIH) and deferasirox, where a boronate masking group blocks metal binding in the resulting “prochelator” until peroxide activation release the parent chelator.

Based on the aroylhydrazone chelator SIH, our first prochelator BSIH was previously shown to have favorable therapeutic ratio of low inherent toxicity and profound cytoprotection against oxidative damage in cultured cells. However, it failed to yield a full complement of SIH upon reaction with hydrogen peroxide in aqueous solutions and cell contexts. In the current study, we report that BSIH undergoes rapid hydrolysis equilibrium with its two degradation components isoniazid and Bsal, which ultimately lower the content of intact SIH formation upon peroxide activation. In our search for boronate-masked prochelators with improved hydrolytic stability, a series of BSIH derivatives have been developed by modifying the parent aroylhydrazone framework with various functionalities. Among others, the para-methoxy derivative (p-OMe)BSIH and the meta-, para- double substituted MD-BSIH have shown improved hydrolytic stability, which ultimately maximizes the release of corresponding active chelators upon peroxide activation. Moreover, both prochelators provide profound cytoprotection to ARPE-19 retinal pigment epithelial cells stressed either exogenously with H2O2 addition, or endogenously with paraquat insult.

Deferasirox is a triazole-based chelator used clinically for iron overload, but also is cytotoxic to cells in culture. In order to test whether a prodrug version of deferasirox could minimize its cytotoxicity but retain its protective properties against iron-induced oxidative damage, we present here a prochelator version TIP that contains a self-immolative boronic ester masking group that is removed upon exposure to hydrogen peroxide to release the bis-hydroxyphenyltriazole ligand deferasirox. TIP does not coordinate to Fe3+ or Zn2+ and shows only weak affinity for Cu, in stark contrast to deferasirox, which avidly binds all three metal ions. TIP converts efficiently in vitro upon reaction with hydrogen peroxide to deferasirox. In cell culture, TIP protects retinal pigment epithelial cells from death induced by hydrogen peroxide; however, TIP itself is more cytotoxic than deferasirox in unstressed cells. These results imply that the cytotoxicity of deferasirox may not derive exclusively from its iron withholding properties, which encourages further investigation on identifying potential target proteins responsible for the toxicity of deferasirox.