Tankyrases: structure, function and therapeutic implications in cancer

Abstract

Several cellular signaling pathways are regulated by ADP-ribosylation, a posttranslational modification catalyzed by members of the ARTD superfamily. Tankyrases are distinguishable from the rest of this family by their unique domain organization, notably the sterile alpha motif responsible for oligomerization and ankyrin repeats mediating protein-protein interactions. Tankyrases are involved in various cellular functions, such as telomere homeostasis, Wnt/β-catenin signaling, glucose metabolism, and cell cycle progression. In these processes, Tankyrases regulate the interactions and stability of target proteins by poly (ADP-ribosyl)ation. Modified proteins are subsequently recognized by the E3 ubiquitin ligase RNF146, poly-ubiquitinated and predominantly guided to 26S proteasomal degradation. Several small molecule inhibitors have been described for Tankyrases; they compete with the co-substrate NAD(+) for binding to the ARTD catalytic domain. The recent, highly potent and selective inhibitors possess several properties of lead compounds and can be used for proof-of-concept studies in cancer and other Tankyrase linked diseases.

Fig. (1)

Domain organization of human ARTD enzymes. Evolutionary relation according to Citarelli and co-workers [40].

Fig. (2)

Covalent protein modification through ADP-ribosylation as catalyzed by ARTDs and removal of ADP-ribosylation. ADP-ribosylation reactions are shown with solid arrows, while the removal of the modifications is shown as dashed arrows. ARTDs catalyze the ADP-ribosylation onto Asp, Glu, and Lys residues (aa) of the acceptor proteins. PARG/ARH3 hydrolyze the O-glycosidic ribose-ribose 1"-2' bond [51, 52], while MDO1/2 remove the proximal modification [53]. TARG catalyzes the removal of PAR but also a single ADPr modification [54]. The catalytic activity responsible for removing the branching from the PAR chain has not been identified. RNF146 poly-ubiquitinates PARsylated proteins, which may subsequently be targeted for degradation in the 26S proteasome [55]. PARG, poly(ADP-ribose) glycohydrolase; ARH, ADP-ribosylhydrolase; TARG, terminal ADP-ribose protein glycohydrolase, MDO, macro domain; RNF146, E3 ubiquitin-protein ligase RNF146.

Fig. (3)

Structure and catalytic sites of Tankyrases. A) The donor and acceptor NAD+ binding sites of TNKS1 (PDB ID 2RF5). The nicotinamide (NI) and adenosine (ADE) subsites are labeled. N-terminus marks the approximate position of the SAM domain which is connected to the catalytic domain with a linker of 18 residues. B) Superposition of TNKS1 (purple) and TNKS2 (aquamarine) (PDB ID 3KR7) showing the HYE conserved triad and the zinc binding site. C) Superposition of TNKS2 and ARTD1 (purple) (PDB ID 3GJW). The regulatory domain (ARD) of ARTD1 is missing in Tankyrases. D) Binding of EB-47 to tankyrase 2 (PDB ID 4BJ9). E) Binding of NAD+ to Diphtheria toxin (PDB ID 1TOX). The disordered D-loop is shown as a dashed line. F) Differences of the acceptor sites of ARTD1 (PDB ID 1A26) and TNKS2 (PDB ID 4HYF). The ADP moiety of an NAD+ analog bound to the ARTD1 is shown. For branching reaction ADP should rotate 180 degrees (from green to blue area), which is blocked in TNKS by acceptor loops.

Fig. (4)

The binding of canonical TNKS inhibitors to the donor NAD+ binding site. A) Binding of XAV939 to TNKS2 nicotinamide subsite (PDB ID 3KR8). B) Binding of IWR-1 to the adenosine subsite of TNKS2 (PDB ID 3UA9).

Fig. (5)

In vitro selectivity profile of selected TNKS inhibitors. No inhibitors have so far been profiled using activity assays with ARTD8, ARTD9, ARTD13, ARTD15, or ARTD16. NI subsite, nicotinamide subsite; ADE subsite, adenosine subsite. IC50 values have been gathered from the literature [15, 23, 68, 157, 159-168].

Fig. (6)

The binding of selected inhibitors to TNKS1/TNKS2. Inhibitors binding to the nicotinamide subsite: A) CMP8 (PDB ID 4BUD), B) CMP11 (PDB ID 4IUE), and C) the general phramacophore of the inhibitors binding to this site. Hydrogen bond interactions of the compounds are shown in dashed lines. Hydrophobic features are shown as spheres and π-π stacking features are black discs. Inhibitors binding mainly to the adenosine subsite are shown in D) G007- LK (PDB ID 4HYF) and E) WIKI4 (PDB ID 4BFP), together with F) showing the common pharmacophoric features at this site. Compounds spanning both sites G) CMP4b (PDB ID 4I9I) and H) NVP-TNKS656-analog (PDB ID 4LI8) are shown as well as I) the combined pharmacophoric features of the dual binders.