Cellular Consequences of Copper Complexes Used To Catalyze Bioorthogonal Click Reactions

• Published in: Journal of the American Chemical Society 2011-11, Vol.133 (44), p.17993-18001
• Main Authors: Kennedy, David C, McKay, Craig S, Legault, Marc C. B, Danielson, Dana C, Blake, Jessie A, Pegoraro, Adrian F, Stolow, Albert, Mester, Zoltan, Pezacki, John Paul
• Format: Article
• Language: English
• Subjects: Catalysis; Cell Survival - drug effects; Cells, Cultured; Copper - chemistry; Fluorescent Dyes - chemistry; Fluorescent Dyes - metabolism; Fluorescent Dyes - pharmacology; HEK293 Cells; HeLa Cells; Humans; Microscopy, Fluorescence; Mitochondria - drug effects; Mitochondria - metabolism; Molecular Structure; Organometallic Compounds - chemistry; Organometallic Compounds - metabolism; Organometallic Compounds - pharmacology; Structure-Activity Relationship
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/ja2083027

Copper toxicity is a critical issue in the development of copper-based catalysts for copper(I)-catalyzed azide–alkyne cycloaddition (CuAAC) reactions for applications in living systems. The effects and related toxicity of copper on mammalian cells are dependent on the ligand environment. Copper complexes can be highly toxic, can induce changes in cellular metabolism, and can be rapidly taken up by cells, all of which can affect their ability to function as catalysts for CuAAC in living systems. Herein, we have evaluated the effects of a number of copper complexes that are typically used to catalyze CuAAC reactions on four human cell lines by measuring mitochondrial activity based on the metabolism of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) to study toxicity, inductively coupled plasma mass spectrometry to study cellular uptake, and coherent anti-Stokes Raman scattering (CARS) microscopy to study effects on lipid metabolism. We find that ligand environment around copper influences all three parameters. Interestingly, for the Cu(II)–bis-l-histidine complex (Cu(his)2), cellular uptake and metabolic changes are observed with no toxicity after 72 h at micromolar concentrations. Furthermore, we show that under conditions where other copper complexes kill human hepatoma cells, Cu(I)–l-histidine is an effective catalyst for CuAAC labeling of live cells following metabolic incorporation of an alkyne-labeled sugar (Ac4ManNAl) into glycosylated proteins expressed on the cell surface. This result suggests that Cu(his)2 or derivatives thereof have potential for in vivo applications where toxicity as well as catalytic activity are critical factors for successful bioconjugation reactions.