The type of DNA damage response after Decitabine treatment depends on the level of DNMT activity

Abstract

Decitabine (AzadC) and Azacytidine (AzaC) are considered as epigenetic drugs that trap DNA-methyltransferases (DNMTs), resulting in DNA-hypomethylation and -damage through DNMT-DNA crosslinks as well as RNA-damage for AzaC. They are applied against myeloid cancers and have promising features for therapy of solid tumours. However, important aspects of their mode of action remain unknown, which highly limits their clinical potential. Using a combinatorial approach, we reveal that under low DNMT-activity, application of AzaC results in only limited phenotypic changes, whereas AzadC displays substantial toxicity, even independent from DNMT-crosslinking. When DNMT-activity is high, toxicity and cellular response to both compounds are dramatically increased, but do not primarily depend on DNA-hypomethylation or RNA-associated processes. By applying spatial proteomics, we show that AzadC induces a strictly DNMT-dependent multifaceted DNA-damage response based on chromatin-recruitment of various repair-associated proteins, with Rad50 as a central player. The choice of DNA-repair pathway herby depends on the severity of the AzadC-induced DNA lesions, which is determined by DNMT-activity. While under moderate DNMT-activity, mismatch (MMR), base-excision (BER) and Fanconi anemia-dependent DNA-repair combined with homologous recombination are activated in response to AzadC, high DNMT-activity and therefore immense replication stress, induce activation of MMR and BER followed by non-homologous and alternative end-joining.