Design and discovery of 3-aryl-5-substituted-isoquinolin-1-ones as potent tankyrase inhibitors

The ADP-ribosyltranferase diphtheria toxin-like (ARTD) or poly-ADP-ribose polymerase (PARP) protein superfamily comprises 17 proteins that contain a common catalytic domain. Those that are catalytically active use β-NAD+ as an essential co-factor to transfer poly- or mono-ADP-ribose units onto protein substrates. This post-translational modification is best characterised for PARP1 (ARTD1) and PARP2 (ARTD2) substrates, which play an important role in the DNA damage response. The PARP1/2 inhibitor olaparib has now been approved for use in the treatment of ovarian cancer, as it is able to selectively target tumour cells with either a BRCA1 or BRCA2 tumour suppressor gene defect. PARP1/2 inhibitors such as olaparib 1 and veliparib 2 exploit the nicotinamidyl pharmacophore present in β-NAD+ 3. Tankyrase 1 and 2 (TNKS/ARTD5 & TNKS2/ARTD6) are PARP proteins which are involved in a range of cellular functions including telomere maintenance, control of the mitotic checkpoint and WNT signalling, as well as the genetic disorder Cherubism.

The role of TNKS/TNKS2 in WNT signalling has highlighted the potential of tankyrase inhibitors as anti-cancer agents, as this pathway has been associated with the development of many tumour types, including colorectal cancers, where loss-of-function mutations in the tumour suppressor gene APC4 cause constitutive WNT signalling driving a tumourigenic phenotype. APC acts as a molecular scaffold, or hub, for the assembly of the ‘destruction complex’, a key component of canonical WNT signalling; this complex, which includes GSK3β and AXIN1/2, sequesters cytosolic β-catenin (a known transcriptional co-activator). The subsequent phosphorylation of β-catenin, by GSK3β, promotes its degradation by the proteasome, leading to the inactivation of WNT signalling.

Conversely, loss of normal destruction complex function, which can occur when APC is mutated, leads to an enhanced level of nuclear, non-phosphorylated, ‘active’ β-catenin which then drives the transcription of WNT target genes such as MYC. TNKS and TNKS2 normally PARylate two components of the destruction complex, AXIN1 and AXIN2, thereby promoting their ubiquitylation and proteosomal degradation; events that minimise the total amount of active β-catenin. Inhibition of TNKS/TNKS2 minimises AXIN degradation, stabilises the destruction complex and suppresses WNT signalling. As constitutive WNT signalling can often be oncogenic, chemical inhibitors of tankyrase activity have been proposed as potential anti-cancer agents.

Here we describe the design and discovery of a series of novel 3-aryl-5-substituted isoquinolin-1-one compounds that are potent TNKS/TNKS2 inhibitors. The optimised compounds presented in this study were characterised by their biochemical potency (estimated using an in vitro poly-ADP-ribosylation assay), their ability to inhibit cellular WNT signalling (quantified using a transcriptional reporter assay) and their ability to inhibit growth of WNT-dependent, APC-mutant (APCmut) colorectal tumour cells.

Results and discussion

In silico and biochemical screening for TNKS inhibitors

In order to initiate the discovery of small molecule TNKS/TNKS2 inhibitors, we used an in silico small molecule screen to identify a set of commercially available compounds that contained the core (nicotinamide-like) aryl-CONH unit found in PARP superfamily inhibitors such as olaparib 1 (AstraZeneca/KuDOS) and veliparib 2 (AbbVie). One thousand and sixty nine compounds were identified using this approach, which were then assessed using a cell-free biochemical assay, where the ability to inhibit PARylation of a histone pseudo-substrate by recombinant TNKS was quantified. This analysis produced 59 hits with >75% inhibition of TNKS activity at a concentration of 10 μM. Consequently, several robust chemical series were identified after dose response validation assays (data not shown) of which one, based on a dihydroisoquinolin-1-one (DHIQ) scaffold, was selected for further optimisation and derivatisation (for an example from the DHIQ series, compound 4).




Richard J. R. Elliott, Ashley Jarvis, Mohan B. Rajasekaran, Alan Ashworth*, Christopher J. Lord*. Med. Chem. Commun.,2015, 6,1687–1692


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