Department of Chemistry, Wayne State University, 5101 Cass Avenue, Detroit, Michigan 48202, United States.
Department of Chemistry and Biochemistry, The Ohio State University, Columbus, Ohio 43210, United States.
Inorg Chem. 2022 Sep 5;61(35):13673-13677. doi: 10.1021/acs.inorgchem.2c02587. Epub 2022 Aug 22.
Cytochromes P450 (CYPs) are a superfamily of enzymes responsible for biosynthesis and drug metabolism. Monitoring the activity of CYP3A4, the major human drug-metabolizing enzyme, is vital for assessing the metabolism of pharmaceuticals and identifying harmful drug-drug interactions. Existing probes for CYP3A4 are irreversible turn-on substrates that monitor activity at specific time points in end-point assays. To provide a more dynamic approach, we designed, synthesized, and characterized emissive Ir(III) and Ru(II) complexes that allow monitoring of the CYP3A4 active-site occupancy in real time. In the bound state, probe emission is quenched by the active-site heme. Upon displacement from the active site by CYP3A4-specific inhibitors or substrates, these probes show high emission turn-on. Direct probe binding to the CYP3A4 active site was confirmed by X-ray crystallography. The lead Ir(III)-based probe has nanomolar and high selectivity for CYP3A4, efficient cellular uptake, and low toxicity in CYP3A4-overexpressing HepG2 cells.
细胞色素 P450（CYPs）是一个负责生物合成和药物代谢的酶超家族。监测 CYP3A4 的活性，作为主要的人药物代谢酶，对于评估药物代谢和识别有害的药物相互作用至关重要。现有的 CYP3A4 探针是不可逆的“开-关”底物，可在终点测定中监测特定时间点的活性。为了提供更具动态性的方法，我们设计、合成并表征了发光的 Ir(III)和 Ru(II)配合物，可实时监测 CYP3A4 活性部位占有率。在结合状态下，探针的发射被活性部位的血红素猝灭。当被 CYP3A4 特异性抑制剂或底物从活性部位置换出来时，这些探针表现出高的发射开启。通过 X 射线晶体学证实了直接探针与 CYP3A4 活性部位的结合。基于 Ir(III)的先导探针对 CYP3A4 具有纳摩尔级的亲和力和高选择性、有效的细胞摄取能力，并且在 CYP3A4 过表达的 HepG2 细胞中毒性低。
