Benzoxazolone Carboxamides: Potent and Systemically Active Inhibitors of Intracellular Acid Ceramidase

Abstract

The ceramides are a family of bioactive lipid-derived messengers involved in the control of cellular senescence, inflammation, and apoptosis. Ceramide hydrolysis by acid ceramidase (AC) stops the biological activity of these substances and influences survival and function of normal and neoplastic cells. Because of its central role in the ceramide metabolism, AC may offer a novel molecular target in disorders with dysfunctional ceramide-mediated signaling. Here, a class of benzoxazolone carboxamides is identified as the first potent and systemically active inhibitors of AC. Prototype members of this class inhibit AC with low nanomolar potency by covalent binding to the catalytic cysteine. Their metabolic stability and high in vivo efficacy suggest that these compounds may be used as probes to investigate the roles of ceramide in health and disease, and that this scaffold may represent a promising starting point for the development of novel therapeutic agents.

Regulation of hepatic insulin signaling and glucose homeostasis by sphingosine kinase 2

“…To segregate the impacts of ceramides and sphingosine, we treated cells with fumonisin b1 and ARN14974. Fumonisin b1 is a specific inhibitor of ceramide synthase, which reduces ceramides but increases sphingosine levels in cells ( 27 ), while ARN14974 is a novel inhibitor of acid ceramidase (ASAH1), which explicitly blocks ceramide conversion to sphingosine and thus reduces sphingosine content ( 28 ). Fumonisin b1 up to 50 μM failed to reverse Akt activation in SphK2 knockdown cells ( Fig.…”

“…Sphingosine, the central substrate of SphK2 in hepatocytes, is mainly produced via ASAH1-mediated hydrolysis of ceramides ( 41 ). By blocking this pathway, the ceramidase inhibitor ARN14974 is known to elevate the ceramide level and decrease the sphingosine level in the cell ( 28 ). Interestingly, both ARN14974 and siRNA-mediated knockdown of ASAH1 significantly rescued insulin sensitivity in SphK2-deficient hepatocytes ( Fig.…”

Inhibitors of Ceramidases

“…This effect was accompanied by an increase in the intracellular levels of (d18:1/16:0) ceramides and a corresponding decrease in sphingosine levels. More interestingly, this compound was found to inhibit aCDase activity in live mice resulting in a significant increase in (d18:1/14:0, d18:1/ 16:0, and d18:1/18:0) ceramides along with a decrease in sphingosine levels (Pizzirani et al, 2015).…”

The emergence of acid ceramidase as a therapeutic target for acute myeloid leukemia

“…Alone, the compound was only mildly toxic at micromolar concentrations, but viability of proliferative melanoma cells was drastically reduced when combined with chemotherapeutics 45 . Benzoxazolone carboxamide prototypes have also been studied as potent acid ceramidase inhibitors 74, 75 . Additionally, small peptide targeting of AC with cystatin SA lead to reduced AC activity and increased ceramide accumulation 76 .…”