Operationalizing DNMT1-Targeting to Treat Chemorefractory Pancreatic Cancer

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Pancreatic ductal adenocarcinoma (PDAC) is a recalcitrant, deadly problem. Most PDAC patients receive aggressive multi-agent chemotherapy but median overall survivals remain at <1 year. A well-established basis for limited chemotherapy response is mutation and deletion of the master regulator of apoptosis p53 in most PDAC. Thus, alternative treatment modalities that do not rely on p53 are needed. The key epigenetic regulator DNA methyltransferase 1 (DNMT1) is a scientifically validated molecular target to cytoreduce chemo-refractory malignancies including PDAC, since it effects cancer cell cycling exits by p53-independent epithelialization. DNMT1 can be inhibited/ depleted by the pyrimidine nucleoside analog pro-drugs decitabine (Dec) or 5- azacytidine (5Aza), approved to treat myeloid malignancies. However, results from several completed clinical trials using Dec, 5Aza, or analogs to treat PDAC and other solid tumors have disappointed. A reason for this was suggested by correlative studies that revealed failure of Dec to elicit DNMT1-inhibiting pharmacodynamic effect in >75% of solid cancer tissues. We explored reasons for this further, in both pre-clinical and clinical studies we found inherent and adaptive configurations of pyrimidine metabolism in PDAC cells that forestall Dec or 5Aza processing into DNMT1-depleting nucleotide. Fortunately, there are practical modifications to therapy that can counter these mechanisms of resistance to DNMT1-targeting by Dec/5Aza. Specifically, we hypothesize that resistance to Dec, 5Aza emerges from adaptive responses of pyrimidine metabolism that can be anticipated and exploited using alternative schedules, and by combination with non-toxic modifiers of pyrimidine metabolism. Aim 1: Evaluate schedules of Dec and 5Aza administration designed to exploit peaks/troughs in pyrimidine metabolism adaptation and increase S-phase dependent DNMT1-depletion. Our prelim data indicates that scheduling Dec/5Aza administration to exploit reactive peaks and troughs in key pyrimidine metabolism enzymes, and frequent, distributed administration to increase overlap between malignant cell S-phase entries and treatment exposures, adds substantial benefit in vivo – we will extend this data to guide mechanistically-rational, readily implementable schedule improvements to clinical therapy. Aim 2: Enhance non-cytotoxic DNMT1-targeting by combining Dec and 5Aza with non-cytotoxic modifiers of pyrimidine metabolism. We have found that auto-upregulation of cytidine deaminase (CDA) that rapidly catabolizes Dec/5Aza, and of de novo pyrimidine synthesis that competes with Dec/5Aza for incorporation into DNA, contributes critically to resistance. We will therefore combine Dec/5Aza with clinical non-cytotoxic inhibitors of catabolic and de novo pyrimidine metabolism pathways, to identify combinations that increase DNMT1-targeting, response rates and durations. The treatment concepts we evaluate here are practical for clinical translation, e.g., we have developed into the clinic oral forms of CDA-inhibitor+Dec/5Aza that are readily combined with clinical inhibitors of de novo pyrimidine synthesis, to rapidly translate pre-clinical proof-of-principle into clinical therapy.