(-)-CHANA, a fluorogenic probe for detecting amyloid binding alcohol dehydrogenase HSD10 activity in living cells.

The association of 17ß-hydroxysteroid dehydrogenase 10 (HSD10) with ß-amyloid in the brain is known to contribute to the progression of Alzheimer's disease. Further, it has been shown that the interaction between the purified HSD10 and ß-amyloid inhibits its enzymatic activity. However, to date no system has been developed to enable the study of HSD10 activity in intact living cells. To address this significant shortcoming, we have developed a novel fluorogenic probe, (-)-cyclohexenyl amino naphthalene alcohol [(-)-CHANA], to observe and measure the activity of HSD10 in living cells. The oxidation of (-)-CHANA by HSD10 results in the production and accumulation of a fluorescent product, which can be measured using real-time fluorescence microscopy. This compound permits the measurement of mitochondrial HSD10 activity and its inhibition by both a small molecule HSD10 inhibitor and by ß-amyloid, in living cells. Herein, we define the parameters under which this probe can be used. This compound is likely to prove useful in future investigations aimed at developing therapeutic compounds targeting the HSD10-ß-amyloid association.

Harnessing functional plasticity of enzymes: a fluorogenic probe for imaging 17beta-HSD10 dehydrogenase, an enzyme involved in Alzheimer's and Parkinson's diseases.

In this paper, we describe the development of a fluorogenic substrate for 17beta-hydroxysteroid-dehydrogenase type 10 (17beta-HSD10), which is a multifunctional metabolic enzyme fulfilling several metabolic roles (beta-oxidation of fatty acids, catabolism of isoleucine, and metabolism of steroids). In recent years, it has emerged as an important stress and pathological marker in neurons and glial cells (expression down-regulation in Parkinson's disease, up-regulation and association with beta-amyloid peptide in Alzheimer's disease). Through the iterative molecular design and chemical synthesis described herein, compound 1 was developed, which possesses all required properties for a selective optical reporter substrate: alcohol-ketone optical switching, the ability to function as a good enzyme substrate (expressed in kinetic parameters), cell permeability, and cell retention. Probe 1 provides a blue-to-green/yellow bright switch and enables non-invasive, real-time imaging of 17beta-HSD10 in live human cells. The selectivity of reporter 1 was established by the quantitative correlation of metabolic activity to protein expression in human kidney cell line HEK-293T.