Abstract C150: Depletion of cellular NAD using the NAMPT inhibitor GNE-617 leads to oncosis blister cell death.

Abstract

Abstract Nicotinamide adenine dinucleotide (NAD) is a critical cellular metabolite and the key enzyme in the NAD salvage pathway is nicotinamide phosphoribosyl transferase (NAMPT). Here we describe GNE-617, a novel orally bioavailable NAMPT inhibitor, and show that this compound promotes rapid depletion of NAD both in cells and in xenograft tumor models. Moreover, GNE-617 rapidly induced tumor regressions in colorectal (HCT-116), fibrosarcoma (HT-1080), prostate (PC3) and pancreatic (MiaPaca2) xenograft models. Despite increasing interest in exploiting cancer cell metabolism as a therapeutic strategy, there is still a poor understanding of cell death mechanisms following disruption of key metabolic processes. Here we show that depletion of NAD in cells induces a series of cellular events that leads to oncosis-Blister Cell Death (oncosis-BCD). In all cell lines examined, NAD was depleted >95% within 25-39 hours, which was rapidly followed by a loss of ATP (>95% depleted by 41-76 hours). Depletion of NAD correlates with loss of cell motility and a reduction in mitotic index, which may be explained by reduced activity of NAD-dependent protein deacetylases. In support of this, we find that α-tubulin-K40 is hyper-acetylation, a modification that is known to reduce tubulin dynamics and cell motility. Additionally, there is no evidence of histone H3-K9 deacetylation, which normally occurs as cells enter mitosis. Using a combination of approaches including live cell imaging and electron microscopy we found that cell lines with a more rapid depletion of ATP display signs of necrosis and oncosis-BCD, while cell lines with a slower depletion of ATP show signs of autophagy, apoptosis, necrosis and oncosis-BCD. In all cases however, oncosis-BCD is the predominant form of cell death and is characterized by plasma membrane swelling to form large organelle-free blisters. Finally, we conclude that cell death is due to loss of plasma membrane ion-homeostasis associated with ATP depletion. In conclusion, our data shows that there is an ordered progression of events that occurs in cancer cells following NAD depletion and reveals that oncosis-BCD is the predominant form of cell death in response to NAD depletion. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C150. Citation Format: Thomas O'Brien, Christopher Del Nagro, Yang Xiao, Jason Oeh, Linda Rangell, Mike Reichelt, Xiaorong Liang, Bianca M. Liederer, Peter S. Dragovich, Deepak Sampath. Depletion of cellular NAD using the NAMPT inhibitor GNE-617 leads to oncosis blister cell death. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C150.