Dynamic Ir(III) Photosensors for the Major Human Drug-Metabolizing Enzyme Cytochrome P450 3A4

Abstract

Probing the activity of cytochrome P450 3A4 (CYP3A4) is critical for monitoring the metabolism of pharmaceuticals and identifying drug–drug interactions. A library of Ir(III) probes that detect occupancy of the CYP3A4 active site were synthesized and characterized. These probes show selectivity for CYP3A4 inhibition, low cellular toxicity, K d values as low as 9 nM, and are highly emissive with lifetimes up to 3.8 μs in cell growth media under aerobic conditions. These long emission lifetimes allow for time-resolved gating to distinguish probe from background autofluorescence from growth media and live cells. X-ray crystallographic analysis revealed structure–activity relationships and the preference or indifference of CYP3A4 toward resolved stereoisomers. Ir(III)-based probes show emission quenching upon CYP3A4 binding, then emission increases following displacement with CYP3A4 inhibitors or substrates. Importantly, the lead probes inhibit the activity of CYP3A4 at concentrations as low as 300 nM in CYP3A4-overexpressing HepG2 cells that accurately mimic human hepatic drug metabolism. Thus, the Ir(III)-based agents show promise as novel chemical tools for monitoring CYP3A4 active site occupancy in a high-throughput manner to gain insight into drug metabolism and drug–drug interactions.

Ruthenium-Based Photoactivated Chemotherapy

“…Like for 11 2+ , the enzyme inhibition properties of 12 2+ in the dark may also explain, at least in part, the significant dark toxicity of this compound toward cancer cells (e.g., EC 50,dark = 23.6 μM in A431 skin cancer cells in normoxia). Although similar unwanted inhibitory properties are observed every now and then with metal complexes, it should be noted that they are more an exception than the rule. In addition, it should be possible to correct them by changing the ruthenium cage or the place where it is installed on the inhibitor, to lower interaction with the targeted protein.…”

The Impact of Inorganic Systems and Photoactive Metal Compounds on Cytochrome P450 Enzymes and Metabolism: From Induction to Inhibition

“…Recently, Kodanko and others developed Ru(II) and Ir(III) based photosensors tethered to pyridine for monitoring the occupancy of the CYP3A4 active site [61,62]. CYP3A4 is a promiscuous enzyme with a large volume active site cavity; the size depends on the nature of the substrate or inhibitor bound within the enzyme, and volumes up to 1400 Å 3 have been reported [63][64][65].…”

“…It was concluded that the hydrophobic and aromatic interactions mediated by the Ir(III) scaffold primarily define the binding ability and inhibitory activity. The optimized CYP3A4 probe (complex 6) possessed a 3-fold higher binding affinity, with a K d value of 24 nM; however, it was cytotoxic against HepG2 cells at the micromolar range (EC 50 = 16 µM) [62]. Unfortunately, the lipophilic efficiency of the new metal-based inhibitors was not discussed in these reports, but it seems likely that the lower charge of the iridium complex was more compatible with the hydrophobic binding site.…”

Cyclometalated Iridium(III) Polypyridine Complexes for Luminescent Detection of Chlorinated Solvents

“…The rich photophysical properties of metal complexes, such as visible range photoexcitation, large Stokes shift, tunable photophysical properties, and relatively longer lifetimes than organic molecules, provide an opportunity to use them as a better fluorescence sensor for various analytes. − Cationic cyclometalated iridium­(III) and heteroleptic ruthenium­(II) polypyridine complexes find extensive applications in chemistry and biology, such as sensors, − OLEDs, , in the fabrication of logic gates, photodynamic therapy, − and cellular imaging probes. − The development of deep red emissive probes is favored in bioimaging and in vivo bioassay because of reduced light scattering, background autofluorescence, and better tissue penetration. − …”