A supramolecular indicator displacement assay for acetyl amantadine, a proxy biomarker for spermidine/spermine N1-acetyltransferase (SSAT) activity

Abstract

Acetyl amantadine (AcAm) is produced from amantadine (Am) in vivo upon catalysis by spermidine/spermine N1-acetyl transferase (SSAT). SSAT is a biomarker for multiple aggressive cancers, and the analysis of AcAm in urine has been promoted as a proxy measure for the early detection of cancer. We report here the development and optimization of cucurbit[7]uril–dye pair based indicator displacement assay (IDA) for the detection of AcAm in solution. In deionized water, using Rhodamine B as the dye, the limit of detection of AcAm was 0.087 μM with a linear response range from 0 to 1 μM. Using berberine as the dye, the limit of detection was 0.077 μM with the same range of linear response. Our efforts and difficulties in translating this assay to function in human urine are also described. We achieve a partial response of the berberine IDA to the presence of AcAm in urine that has undergone a simple PD-10 desalting step.