Investigating strategies to modify PARP14 function through macrodomain inhibition

Macrodomains are conserved protein interaction modules that are present in all domains of life and mediate recognition of sequence motifs harbouring adenosine diphosphate ribose (ADPR) modifications. In addition, some of them are able to control the turnover of ADPR signalling through their catalytic activity removing these modifications. Macrodomains are hence implicated in a variety of cellular processes as well as in diseases including cancer and viral pathogenesis. The polyadenosine-diphosphate-ribose polymerase (PARP) family member PARP14 is one of twelve human macrodomain-containing proteins; it contains three macrodomains in addition to its catalytic PARP domain. PARP14 was shown to be involved in several cellular processes linked to cancer development in for example B-cell lymphoma and hepatocellular carcinoma. Therefore PARP14 including its macrodomains has emerged as a potential therapeutic target. However, the lack of specific small molecule inhibitors has hampered domain-specific target validation studies so far. Current approaches focus on inhibitor development for its PARP domain, yet attaining selectivity of these inhibitors over other PARP enzymes has been challenging. The aim of the work described in this thesis was therefore to evaluate the possibility of targeting PARP14 via inhibition of its macrodomains with small molecule inhibitors as an alternative to PARP domain inhibition. These studies revealed that displacing PARP14 from its target sites requires inhibitor development for both its second and third macrodomain. Druggability of both macrodomains has been demonstrated by the highly selective allosteric macrodomain 2 inhibitor GeA-69 and by several fragment hits targeting the ADPR binding site of macrodomain 3. Finally, PARP14 was confirmed to be implicated in DNA repair mechanisms protecting cells against replication stress, suggesting that a dual PARP14 macrodomain inhibitor may provide the possibility to potentiate genotoxic chemotherapy for cancer treatment.