Combining drugs that bypass p53 to treat <i>TP53</i> -mutated leukemias

Acute myeloid leukemias (AML) containing TP53 (p53) mutations are routinely treated with decitabine or 5-azacytidine, which deplete DNA methyltransferase 1 (DNMT1; ie, hypomethylating agents [HMA]). Unfortunately, resistance/relapse, characterized by preserved DNMT1, is rapid. HMA are pyrimidine analogs, and to deplete DNMT1, must compete with endogenous pyrimidines. These were substantially increased in HMA-resistant vs parental AML cells, together with upregulation of CAD (carbamoyl-phosphate-synthetase-2/aspartate transcarbamylase/dihydroorotase) that rate limits de novo pyrimidine synthesis. Moreover, TP53-mutated AML appeared primed for such resistance, with higher baseline CAD. Pyrimidine synthesis can be depowered with the B-cell lymphoma 2 (BCL2) inhibitor venetoclax to release BCL-2-associated X protein (BAX) to depolarize mitochondrial membranes. However, BAX, a p53 target gene, was substantially less expressed in TP53-mutated vs wild-type TP53 cells, and venetoclax impacts were correspondingly limited. Alternatively, pyrimidine synthesis can be inhibited directly at dihydroorotate dehydrogenase (DHODH) using the clinical drug teriflunomide. Contrasting with venetoclax, teriflunomide decreased pyrimidine levels several-fold, restored DNMT1 depletion, and cytoreduced HMA-resistant TP53-mutated AML cells via p53/apoptosis-independent terminal-differentiation. This noncytotoxic pathway preserved viability and proliferation of normal hematopoietic stem/progenitor cells (NHSPCs). Inhibiting pyrimidine synthesis triggered compensatory pyrimidine salvage, such that schedules for teriflunomide combination with HMA, which are taken up by salvage, mattered. In mice with TP53-mutated AML, teriflunomide scheduled the day before HMA was more efficacious than same-day or day-after schedules. Chronic teriflunomide exposure paradoxically increased pyrimidines via sustained compensatory salvage, conferring resistance rather than sensitivity to HMA. In sum, DNMT1 and DHODH targeting, administered by timed, intermittent (metronomic) schedules, can circumvent genetic resistance caused by TP53 mutations and adaptive resistance caused by metabolic homeostasis, without cytotoxicity to HSPCs.