Decitabine- and 5-azacytidine resistance emerges from adaptive responses of the pyrimidine metabolism network

Abstract

Mechanisms-of-resistance to decitabine and 5-azacytidine, mainstay treatments for myeloid malignancies, require investigation and countermeasures. Both are nucleoside analog pro-drugs processed by pyrimidine metabolism into a deoxynucleotide analog that depletes the key epigenetic regulator DNA methyltranseferase 1 (DNMT1). Here, upon serial analyses of DNMT1 levels in patients' bone marrows on-therapy, we found DNMT1 was not depleted at relapse. Showing why, bone marrows at relapse exhibited shifts in expression of key pyrimidine metabolism enzymes in directions adverse to pro-drug activation. Further investigation revealed the origin of these shifts. Pyrimidine metabolism is a network that senses and regulates deoxynucleotide amounts. Deoxynucleotide amounts were disturbed by single exposures to decitabine or 5azacytidine, via off-target depletion of thymidylate synthase and ribonucleotide reductase respectively. Compensating pyrimidine metabolism shifts peaked 72-96 h later. Continuous pro-drug exposures stabilized these adaptive metabolic responses to thereby prevent DNMT1-depletion and permit exponential leukemia out-growth as soon as day 40. The consistency of the acute metabolic responses enabled exploitation: simple treatment modifications in xenotransplant models of chemorefractory leukemia extended noncytotoxic DNMT1-depletion and leukemia control by several months. In sum, resistance to decitabine and 5-azacytidine originates from adaptive responses of the pyrimidine metabolism network; these responses can be anticipated and thus exploited.

Resistance to venetoclax and hypomethylating agents in acute myeloid leukemia

“…There is evidence to support a dual mechanism of action for these agents: (1) a direct cytotoxic effect at higher doses, where the formation of covalent DNMT-DNA adducts leads to DNA damage; and (2) DNA hypomethylation and epigenetic modulation at lower doses with subsequent cell differentiation and tumor suppression [36,37] . Proposed mechanisms of auto-resistance to hypomethylating agents [35] . A: mechanisms related to changes in nucleoside metabolism: (1) decitabine inhibits thymidylate synthase (TYMS).…”

“…HMAs are pyrimidine analogs of the nucleoside cytidine that showed promising cytostatic activity at higher doses in the 1960s [ 34 ] . In both azacitidine and decitabine, a nitrogen atom replaces a carbon atom in position 5 of the pyrimidine ring [ Figure 1 ], but the sugar moiety is deoxyribose in decitabine and ribose in azacitidine [ 35 ] . There is evidence to support a dual mechanism of action for these agents: (1) a direct cytotoxic effect at higher doses, where the formation of covalent DNMT-DNA adducts leads to DNA damage; and (2) DNA hypomethylation and epigenetic modulation at lower doses with subsequent cell differentiation and tumor suppression [ 36 , 37 ] .…”

“…Emerging evidence suggests that metabolism of pyrimidines, including HMAs, reflects an adaptive network of enzymes that detect nucleotide levels and respond in a compensatory manner [ Figure 1 ] [ 35 ] . For example, DCK protein is diminished in MDS samples with decitabine resistance and upregulated in azacitidine-resistant cells.…”

“…For example, DCK protein is diminished in MDS samples with decitabine resistance and upregulated in azacitidine-resistant cells. Conversely, UCK2 protein is upregulated in decitabine-resistant cells and diminished in azacitidine-resistant cells [ 35 ] . These observations raise the possibility of tailoring therapy based on DCK and UCK levels or diminishing HMA resistance by alternating HMAs [ 35 , 97 ] .…”

“…The enzyme carbamoyl-phosphate synthetase (CAD) facilitates the synthesis of dCTP, which competes with aza-dCTP, for incorporation into DNA. Notably, CAD has been found to be upregulated in both cell lines and clinical MDS samples that are resistant to HMAs, although this has not been a universal finding [ 35 , 98 , 99 ] .…”