Targeting NAD metabolism in cancer therapy

Abstract

Nicotinamide adenine dinucleotide (NAD) is fundamental to electron movement and metabolism in all living organisms. Tumor cells exhibit NAD metabolic dependencies that may be amenable to targeting with available agents. Two primary means by which cells synthesize NAD+ are driven by nicotinic acid phosphoribosyltransferase (NAPRT) and nicotinamide phosphoribosyltransferase (NAMPT). Multiple potent and selective NAMPT inhibitors (NAMPTi’s; GMX-1778, FK866) have been developed and demonstrate robust anti-tumor activity in pre-clinical models in vitro and in vivo. First, we tested the hypothesis that specific inhibition of NAMPT would deplete NAD(P)(H) and selectively induce metabolic oxidative stress in non-small cell lung cancer (NSCLC) cells, and that this effect would be augmented by radiation, chemotherapies, and pharmacological ascorbate (P-AscH-). We found that physiologically-relevant doses of NAMPTi’s induce NAD(P)(H) depletion and clonogenic cell death that is prevented by nicotinic acid (NA) supplementation. Furthermore, NAMPTi’s induce metabolic oxidative stress characterized by glutathione depletion and/or oxidation, increased mitochondrial pro-oxidant production and significant sensitization to agents that modulate steady-state levels of hydroperoxides. In addition, NAMPTi’s sensitize tumor cells, but not normal human fibroblasts (NHFs) to temozolomide (TMZ) and cisplatin and serve as effective radiosensitizers. Second, we characterized NAPRT expression as a biomarker for NAMPTi sensitivity utilizing publicly available data from the Cancer Dependency Map (DepMap). We found that rhabdomyosarcoma (RMS) and glioma cell lines exhibit high degrees of NAPRT promoter methylation accompanied by low NAPRT mRNA and protein levels relative to other cancer types. Clonogenic survival assays showed that NAPRT-silenced RMS cells are highly sensitive to GMX-1778 (GMX) and that this toxicity is not rescued by NA-supplementation, confirming that functional NAPRT is not expressed by these cells. Finally, we found that GMX exhibited potent sensitization toward TMZ and P-AscH- in NAPRT-silenced sarcoma cell lines. Overall, these results suggest that NAMPTi’s may serve as promising adjuvants for cancer therapy and designate RMS and gliomas as potential targets for NAMPTi’s.