Abstract 6045: Interferon signaling induces NAD(H) consumption via PARP9/10/14 and sensitizes pancreatic cancer cells to NAMPT inhibition

Abstract

Abstract Interferons (IFNs) are critical cytokines which, in the tumor microenvironment, have the potential to trigger targetable vulnerabilities. Pancreatic ductal adenocarcinoma (PDAC) has an extensively reprogrammed metabolic network in which nicotinamide adenine dinucleotide (NAD) is required as a co-factor for critical cell processes. NAMPT is the rate-limiting enzyme in the NAD salvage pathway, which is the dominant source of NAD in cancer cells. Findings from clinical trials with NAMPT inhibitors (NAMPTi) suggest the need to identify patient subsets that are highly sensitive to NAD depletion. An NAD assay was used to profile the effect of type I IFN signaling and NAMPTi on NAD levels in a panel of 12 human PDAC models. The effect of this NAD reduction was profiled in vitro with cell proliferation and mitochondrial respiration assays, Incucyte cell imaging, and flow cytometry for DNA damage (pH2A.X assay) and apoptosis (Annexin V assay). After confirmation of the synergy between IFN signaling and NAMPTi, a high throughput screen was performed to identify drugs that are potentiated by reduced NAD levels. Inhibition of PDAC proliferation and induction of DNA damage and apoptosis with the combination treatment was validated in vitro using the same assays previously described. In vivo studies were then performed with orthotopic tumor models (with Luciferase labeling for bioluminescence imaging) and patient-derived xenograft (PDX) models. We found that IFN signaling, present in a subset of PDAC tumors, substantially lowered NAD(H) levels through upregulating the expression of NAD consuming enzymes PARP9, PARP10, and PARP14. IFN and NAMPTi collaboratively reduced NAD levels in PDAC cells, inhibited PDAC cell proliferation and invasion in vitro, and suppressed orthotopic PDAC tumor growth and liver metastasis in vivo. We then identified clinical oncology drugs more potent in PDAC cells with lower NAD levels induced by IFN and NAMPTi, which included DNA-damaging compounds Gemcitabine (GEM) and Paclitaxel (PTX). Mechanistically, the NAD-dependent PARP family members are critical in DNA repair. Their activity can be monitored by measuring PAR abundance, which was substantially reduced by IFNβ and NAMPTi with and without the GEM/PTX combination. NAD(H) reduction and GEM/PTX collaboratively enhanced phosphorylation of CHEK2, a marker of DNA damage response, and triggered PDAC cell apoptosis. All of these findings were rescued by nicotinamide riboside (NR) supplementation (which bypasses the need for rate-limiting NAMPT in NAD synthesis), validating that these findings were due to the effect of on-target NAD(H) reduction. Our findings predict that using PARP9/10/14 as biomarkers will allow the identification of PDAC patients whose tumors have relatively low NAD(H) levels, which we hypothesize would render them vulnerable to combination therapies with NAMPTi and DNA damaging agents. Citation Format: Alexandra M. Moore, Lei Zhou, Shili Xu, Timothy R. Donahue. Interferon signaling induces NAD(H) consumption via PARP9/10/14 and sensitizes pancreatic cancer cells to NAMPT inhibition [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6045.