Abstract 3016: KPT-9274 inhibits cellular NAD and synergizes with NAD depleting enzymes to induce cancer cell death

Abstract

Abstract Nicotinamide adenine dinucleotide (NAD) is an essential metabolite and important cofactor of several biological processes (i.e. metabolism and genomic stability) that undergo significant alterations during malignant transformation. In cancer cells, NAD undergoes quick turnover through the high metabolic demands of rapidly proliferating cells and increased activity of NAD consuming enzymes, such as sirtuin 1 (SIRT1) and poly ADP ribose polymerase 1 (PARP1). NAD can be generated de novo from tryptophan or regenerated by nicotinamide phosphoribosyl transferase (NAMPT) or nicotinate phosphoribosyl transferase 1 (NAPRT1) in NAD salvage pathways. However, cancer cells do not utilize the de novo or NAPRT1 pathways effectively and instead rely on the NAMPT-dependent salvage pathway to generate NAD, making NAD depletion a promising anti-cancer therapy. We have previously described the PAK4 allosteric modulator, KPT-9274, which can also inhibit the enzymatic activity of NAMPT. KPT-9274 and other NAMPT inhibitors rapidly deplete cellular NAD levels, ultimately leading to ATP depletion and cell death. Similarly, several studies have shown that hyper activation of NAD consuming enzymes can lead to apoptosis. The purpose of this study is to determine whether NAD inhibition by KPT-9274 can synergize with NAD depleting enzymes to enhance the cytotoxic effects of KPT-9274 in cancer cells. Methods: Celltiter-Glo was used to measure ATP levels and viability of cells. NAD/NADH-Glo was used to measure total NAD levels in cells. Gene and protein expression was measured using quantitative PCR and western blot analysis, respectively. Protein knockdown was accomplished using RNAi. Results: We have identified an orally bioavailable dual inhibitor of PAK4 and NAMPT, which demonstrated potent anti-cancer activity in a variety of cell lines both in vitro and in vivo. We have identified a mechanistic combination that increase the antitumor activity of KPT-9274, through the activation of NAD consuming enzymes such as SIRT1 and PARP1. Specifically, we found that SRT1720 (an activator of SIRT1) synergizes with KPT-9274 to increase cancer cell death while RNAi of SIRT1 diminishes the efficacy of KPT-9274. In addition, activation of PARP1 by DNA damaging agents (e.g. gemcitabine) significantly enhances the effectiveness of KPT-9274 mediated cell death. Finally, we show that PARP activation through DNA-damaging agents enhances the cytotoxicity and anti-tumor properties of KPT-9274 in xenograft models. Conclusions: Here we report that KPT-9274 synergizes with NAD depleting enzymes to induce cancer cell death in vitro and in vivo. This noteworthy enhancement to the anticancer activity of KPT-9274, together with its previously described PAK4 inhibition, support the continued development of this orally bioavailable small molecule in combination with current therapies. Citation Format: Christian Argueta, Trinayan Kashyap, Margaret Lee, Yosef Landesman, Sharon Shacham, William Senapedis, Erkan Baloglu. KPT-9274 inhibits cellular NAD and synergizes with NAD depleting enzymes to induce cancer cell death. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3016.