Small Molecule Regulators of Ferroptosis.

Ferroptosis is a dedicated mode of cell death involving iron, reactive oxygen species and lipid peroxidation. Involved in processes such as glutathione metabolism, lysosomal iron retention or interference with lipid metabolism, leading either to activation or inhibition of ferroptosis. Given the implications of ferroptosis in diseases such as cancer, aging, Alzheimer and infectious diseases, new molecular mechanisms underlying ferroptosis and small molecules regulators that target those mechanisms have prompted a great deal of interest. Here, we discuss the current scenario of small molecules modulating ferroptosis and critically assess what is known about their mechanisms of action.

Salinomycin Derivatives Kill Breast Cancer Stem Cells by Lysosomal Iron Targeting.

Salinomycin (1) exhibits a large spectrum of biological activities including the capacity to selectively eradicate cancer stem cells (CSC), making it and its derivatives promising candidates for the development of drug leads against CSC. It has been previously shown that salinomycin and its C20-propargylamine derivative (Ironomycin (2)) accumulate in lysosomes and sequester iron in this organelle. Herein, a library of salinomycin derivatives is reported, including products of C20-amination, C1-esterification, C9-oxidation, and C28-dehydration. The biological activity of these compounds is evaluated against transformed human mammary epithelial HMLER CD24low /CD44high cells, a well-established model of breast CSC, and HMLER CD24high /CD44low cells deprived of CSC properties. Unlike other structural alterations, derivative 4, which displays a cyclopropylamine at position C20, showed a strikingly low IC50 value of 23 nm against HMLER CD24low /CD44high cells. This study provides highly selective molecules to target the CSC niche, a potential interesting advance for drug development to prevent cancer resistance.

Deeper insight into protease-sensitive "covalent-assembly" fluorescent probes for practical biosensing applications.

We report a rational and systematic study devoted to the structural optimisation of a novel class of protease-sensitive fluorescent probes that we recently reported (S. Debieu and A. Romieu, Org. Biomol. Chem., 2017, 15, 2575-2584), based on the "covalent-assembly" strategy and using the targeted enzyme penicillin G acylase as a model protease to build a fluorescent pyronin dye by triggering a biocompatible domino cyclisation-aromatisation reaction. The aim is to identify ad hoc probe candidate(s) that might combine fast/reliable fluorogenic "turn-on" response, full stability in complex biological media and ability to release a second molecule of interest (drug or second fluorescent reporter), for applications in disease diagnosis and therapy. We base our strategy on screening a set of active methylene compounds (C-nucleophiles) to convert the parent probe to various pyronin caged precursors bearing Michael acceptor moieties of differing reactivities. In vitro stability and fluorescent enzymatic assays combined with HPLC-fluorescence analyses provide data useful for defining the most appropriate structural features for these fluorogenic scaffolds depending on the specifications inherent to biological application (from biosensing to theranostics) for which they will be used.

2nd PSL Chemical Biology Symposium (2019): At the Crossroads of Chemistry and Biology.

Chemical Biology is the science of designing chemical tools to dissect and manipulate biology at different scales. It provides the fertile ground from which to address important problems of our society, such as human health and environment.

In situ formation of pyronin dyes for fluorescence protease sensing.

We report a reaction-based strategy for the fluorogenic detection of protease activity. Based on the "covalent-assembly" probe design principle recently put forward by the Yang group for detection of Sarin related threats (J. Am. Chem. Soc., 2014, 136, 6594-6597), we have designed two unusual non-fluorescent caged precursors (mixed bis-aryl ethers) which are readily converted into a fluorescent unsymmetrical pyronin dye through a domino cyclisation-aromatisation reaction triggered by penicillin G acylase (PGA) or leucine aminopeptidase (LAP). Fluorescence-based in vitro assays and HPLC-fluorescence/-MS analyses support the claimed activation mechanism whose the further implementation to "smart" imaging agents for the study of protease function in vivo is expected.

Synthesis of N,N-Dialkylamino-nor-Dihydroxanthene-Hemicyanine Fused Near-Infrared Fluorophores and Their First Water-Soluble and/or Bioconjugatable Analogues.

The effective synthesis of extended conjugated N,N-dialkylamino-nor-dihydroxanthene-based fluorophores is described from diversely functionalized salicylic aldehydes. The access to these original fluorescent derivatives proceeded in two steps through a one-pot construction of the unusual nor-dihydroxanthene (nor-DHX) scaffold followed by a diversification step providing a wide variety of nor-DHX-hemicyanine fused dyes emitting in the range of 730-790 nm. The versatility of our approach has enabled a further extension to the late-stage introduction of negatively/positively charged polar groups onto their terminal nitrogen heterocyclic subunit, thereby giving access to the first water-soluble and/or bioconjugatable members of this emerging class of NIR fluorophores. Our water-solubilizing method is easily implementable, and the nor-DHX-hemicyanine skeleton maintains satisfying fluorescence quantum yields (5-20 %) under physiological conditions. Finally, the bioconjugation ability of fluorescent derivatives bearing a free carboxylic acid was demonstrated through the covalent labeling of a model protein, namely, bovine serum albumin.

Dual enzyme-responsive "turn-on" fluorescence sensing systems based on in situ formation of 7-hydroxy-2-iminocoumarin scaffolds.

A new strategy for the simultaneous fluorogenic detection of two distinct enzyme activities namely hydrolase (amidase or esterase) and reductase is described. This innovative biosensing method is based on the powerful "covalent-assembly" principle that involves in situ synthesis of a fluorophore from a non-fluorescent caged precursor and through domino reactions triggered by the two analytes of interest. To establish this approach, penicillin G acylase (PGA) (or pig liver esterase (PLE)) and nitroreductase (NTR) were chosen as model enzymes, and original bis-O-protected 2,4-dihydroxycinnamonitrile derivatives acting as dual-reactive probes readily convertible to highly fluorescent 7-hydroxy-2-iminocoumarin scaffolds upon reacting with the two selected enzymes were synthesised. The two phenolic groups available within the core structure of these probes play a pivotal role in generating iminocoumarin scaffold through an intramolecular cyclisation reaction (hydroxyl group in C-2 position) and in enhancing its push-pull character (hydroxyl group in C-4 position). Their orthogonal and temporary protection with two different enzyme-labile masking groups is the cornerstone in the design of this novel class of fluorogenic "turn-on" probes. Their evaluation using fluorescence-based in vitro assays and HPLC-fluorescence/-MS analyses have enabled us both to demonstrate the claimed activation mechanism (in particular the specific order in which the two enzymes react with the probe) and to highlight the potential utility of these advanced chemical tools in multi-analyte sensing applications.