Emissive Alkylated Guanine Analogs as Probes for Monitoring O⁶-Alkylguanine-DNA-transferase Activity

Human O6-alkylguanine-DNA-transferase (hAGT) is a repair protein that provides protection from mutagenic events caused by O6-alkylguanine lesions. As this stoichiometric activity is tissue-specific, indicative of tumor status, and correlated to chemotherapeutic success, tracking the activity of hAGT could prove to be informative for disease diagnosis and therapy. Herein, we explore two families of emissive O6-methyl- and O6-benzylguanine analogs based on our previously described thGN and tzGN, thieno- and isothiazolo-guanine surrogates, respectively, as potential reporters. We establish that O6-BnthGN and O6-BntzGN provide a spectral window to optically monitor hAGT activity, can be used as substrates for the widely used SNAP-Tag delivery system, and are sufficiently bright to be visualized in mammalian cells using fluorescence microscopy.