Enzymatic and Biological Characterization of Novel Sirtuin Modulators against Cancer.

Vincenzo Carafa,Marzia Di Donato,Antonello Mai,Zhijun Yu,Lucia Altucci,Angelita Poziello,Laura Della Torre,Daniela Tomaselli,Dante Rotili,Alfonso Baldi,Angela Nebbioso,Gabriella Castoria,Pia Giovannelli,Elham Safadeh

doi:10.3390/ijms20225654

Abstract

Sirtuins, a family of nicotinamide adenine dinucleotide (NAD+)-dependent lysine deacetylases, are promising targets for anticancer treatment. Recently, we characterized a novel pan-sirtuin (SIRT) inhibitor, MC2494, displaying antiproliferative effects and able to induce death pathways in several human cancer cell lines and decrease tumor growth in vivo. Based on the chemical scaffold of MC2494, and by applying a structure–activity relationship approach, we developed a small library of derivative compounds and extensively analyzed their enzymatic action at cellular level as well as their ability to induce cell death. We also investigated the effect of MC2494 on regulation of cell cycle progression in different cancer cell lines. Our investigations indicated that chemical substitutions applied to MC2494 scaffold did not confer higher efficacy in terms of biological activity and SIRT1 inhibition, but carbethoxy-containing derivatives showed higher SIRT2 specificity. The carbethoxy derivative of MC2494 and its 2-methyl analog displayed the strongest enzymatic activity. Applied chemical modifications improved the enzymatic selectivity of these SIRT inhibitors. Additionally, the observed activity of MC2494 via cell cycle and apoptotic regulation and inhibition of cell migration supports the potential role of SIRTs as targets in tumorigenesis and makes SIRT-targeting molecules good candidates for novel pharmacological approaches in personalized medicine.

Highlights

It is widely accepted that cancer displays both genomic and epigenomic deregulations [1,2]
To clarify whether these compounds exerted an inhibitory effect against SIRT1 and SIRT2, a fluorescent enzymatic assay of the compounds was performed at a concentration of 5 μL intermediate dilution (50 μM) against SIRT human recombinant enzymes produced in Escherichia coli
Western blot analysis performed for two metalloproteinases (MMP2 and MMP9) involved in cell migration revealed that together with the block of proliferation an accumulation of the metalloproteinase inactive forms occurred (Figure 7E). These findings indicate that SIRT inhibition mediated by MC2494 has a functional role in cancer cell migration

Summary

Introduction

It is widely accepted that cancer displays both genomic and epigenomic deregulations [1,2]. The best-known epigenetic enzymes targeted in cancer therapy are HDACs, which modulate the expression and activity of many proteins deregulated in cancer [3,4,5]. We investigated its anticancer activity by characterizing the type of cell death mechanism activated, and observed a novel kind of tumor-selective apoptosis. This proposed activity of MC2494 corroborates the role of SIRT1 in tumorigenesis and prompted us to further explore the activity of MC2494 and to design new small MC2494-derived molecules able to modulate SIRT activity. We evaluated the action of MC2494 on cell cycle progression and migration, characterizing some of its derivatives at biological and enzymatic levels

Results

Discussion

Conclusion