Screening, Synthesis, and Evaluation of Novel Isoflavone Derivatives as Inhibitors of Human Golgi β-Galactosidase

Abstract

The genes GLB1 and GALC encode GLB1 isoform 1 and galactocerebrosidase, respectively, which exhibit β-galactosidase activity in human lysosomes. GLB1 isoform 1 has been reported to play roles in rare lysosomal storage diseases. Further, its β-galactosidase activity is the most widely used biomarker of senescent and aging cells; hence, it is called senescence-associated β-galactosidase. Galactocerebrosidase plays roles in Krabbe disease. We previously reported a novel β-galactosidase activity in the Golgi apparatus of human cells; however, the protein responsible for this activity could not be identified. Inhibitor-derived chemical probes can serve as powerful tools to identify the responsible protein. In this study, we first constructed a cell-based high-throughput screening (HTS) system for Golgi β-galactosidase inhibitors, and then screened inhibitors from two compound libraries using the HTS system, in vitro assay, and cytotoxicity assay. An isoflavone derivative was identified among the final Golgi β-galactosidase inhibitor compound hits. Molecular docking simulations were performed to redesign the isoflavone derivative into a more potent inhibitor, and six designed derivatives were then synthesized. One of the derivatives, ARM07, exhibited potent inhibitory activity against β-galactosidase, with an IC 50 value of 14.8 µM and competitive inhibition with K i value of 13.3 µM. Furthermore, the in vitro and cellular inhibitory activities of ARM07 exceeded those of deoxygalactonojirimycin. ARM07 may contribute to the development of affinity-based chemical probes to identify the protein responsible for the newly discovered Golgi β-galactosidase activity. The therapeutic relevance of ARM07 against lysosomal storage diseases and its effect on senescent cells should be evaluated further.

Drug Repurposing and Lysosomal Storage Disorders: A Trick to Treat

“…Miura and co-workers [ 32 ] described another candidate PC for the same disease. Upon assessing a fluorogenic-based high-throughput screening for Golgi-associated β-galactosidase, they performed a compound screening, selecting an isoflavone derivative.…”

Recent advances in the synthesis of 4H-chromen-4-ones (2012 − 2021)

“…A common approach to obtain a wide variety of 3-substituted 4H-chromen-4-ones is using (E)-3-(dimethylamino)-1-(2-hydroxyaryl) prop-2-en-1-ones, commonly known as o-hydroxyaryl enaminones, as key intermediates, obtained mainly from the reaction of 2 0 -hydroxyacetophenones with DMF-DMA in DMF as solvent. Various 3-iodo-4H-chromen-4-ones arise from one-pot ring-closure and iodination sequence via addition of molecular iodine to o-hydroxyaryl enaminones employing methanol, [242][243][244][245][246][247] chloroform, 248,249 pyridine and chloroform, [250][251][252][253][254] pyridine and dichloromethane 47,255 as solvents, at room temperature. Examples of 3-fluoro-4H-chromen-4-ones were synthesized through the monofluorination of o-hydroxyaryl enaminones with Selectfluor ® as fluorine source in 1,2-DCE at room temperature, 256 in the presence of sodium acetate and butylated hydroxytoluene (BHT)…”