Abstract 1088: Advancing non-cytotoxic DNMT1-targeting to treat chemorefractory pancreatic cancer
Authors: Tohme Rita , Enane Francis , Schuerger Caroline , Gu Xiaorong , Fishel Melissa , Pink John , Lindner Daniel , Sohal Davendra , Saunthararajah Yogen 
Published: July 2021

The key epigenetic regulator DNA methyltransferase 1 (DNMT1) is a scientifically validated target in p53-null chemorefractory cancers like pancreatic ductal adenocarcinoma (PDAC) since DNMT1-depletion effects cancer cell cycle exits by p53-independent epithelialization. DNMT1 can be depleted by the pyrimidine nucleoside analog pro-drugs decitabine (Dec) or 5-azacytidine (5Aza). However, PDAC clinical trials with Dec/5Aza disappointed. In pre-clinical and clinical analyses, we found resistance was caused by configurations of pyrimidine metabolism in PDAC cells that forestall Dec or 5Aza processing into DNMT1-depleting nucleotide: high expression of cytidine deaminase (CDA) that rapidly catabolizes Dec/5Aza; and suppression of deoxycytidine kinase (DCK) and uridine kinase 2 (UCK2) that rate limit Dec/5Aza pro-drug processing respectively. Accordingly, combination of Dec with a CDA clinical inhibitor, tetrahydrouridine (THU), enabled DNMT1-depletion and PDAC cytoreduction in vitro and in Dec/gemcitabine-refractory PDAC pre-clinical in vivo models. We then conducted a pilot clinical trial in 13 patients with chemorefractory PDAC given oral THU ~10 mg/kg/day combined with decitabine ~0.2 mg/kg/day, for 5 consecutive days, then twice weekly. This Phase 2 was based on several PK/PD studies in human subjects showing potent non-cytotoxic DNMT1-targeting in myeloid cells. Yet again, there were no meaningful clinical responses in the patients. A reason for this was a surprising lack of neutropenia, the most sensitive indicator of systemic DNMT1-targeting. Upon measuring plasma CDA enzyme activity, we found a &gt;10-fold increase in patients with metastatic vs resectable PDAC. Thus, CDA activity is increased not only locally but also systemically in metastatic PDAC, suggesting a need for higher THU doses. We have also observed DCK downregulation, necessary for Dec/gemcitabine uptake and processing, as a cause of PDAC resistance to Dec/gemcitabine. To counter this mechanism, we discovered that 5Aza upregulates DCK as an adaptive response to 5Aza-mediated decrease in dCTP, while Dec upregulates UCK2 (that mediates 5Aza uptake) as an adaptive response to Dec mediated reductions in dTTP. Thus, we alternated Dec with 5Aza in an in vivo model of gemcitabine-resistant PDAC, to exploit their mutual cross-priming, together with THU to inhibit CDA: median vehicle control tumor measurements 972 mm3(range 726-1267.5); median THU-Dec/THU-5Aza 16 mm3 (range 0-87.5); P&lt;0.00001). A non-cytotoxic, epithelial-differentiation based mechanism was confirmed by significant increases in pancreatic epithelial markers while apoptosis markers were unchanged. In sum, metabolism-based resistance to Dec/5Aza can be countered by clinically relevant modifications to treatment, such as alternating doses of THU/Dec and THU/5Aza, for non-cytotoxic p53-independent therapy, a modality distinct from chemoradiation.

Citation Format: Rita Tohme, Francis Enane, Caroline Schuerger, Xiaorong Gu, Melissa Fishel, John Pink, Daniel Lindner, Davendra Sohal, Yogen Saunthararajah. Advancing non-cytotoxic DNMT1-targeting to treat chemorefractory pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1088.