NAMPT inhibition impairs mTOR-dependent regulation of translation in melanomaNAMPT inhibition impairs mTOR-dependent regulation of translation in melanoma.

The frequent emergence of drug resistance in melanoma remains a challenge. This phenomenon relies on the rewiring of multiple processes, including cancer metabolism, epigenetics, translation process, and interactions with the tumor microenvironment that are only partially understood. Activation of NAD metabolism through its rate-limiting biosynthetic enzyme nicotinamide phosphoribosyltransferase (NAMPT) has been identified as key driver of targeted therapy resistance and melanoma progression. Another major player in this context is the mammalian target of rapamycin (mTOR) pathway, which plays key roles in the regulation of melanoma cell anabolic functions, energy metabolism and protein translation. An interplay between NAD/NAMPT and mTOR signaling axes in regulating translation was revealed some years ago in a leukemic model, however no further studies fully addressed this functional connection. We showed that NAMPT inhibitors (FK866, OT-82) induce a translational arrest in BRAF-mutated human melanoma cell lines sensitive and resistant to BRAF inhibitors. The molecular mechanism involved the activation of 5' AMP-activated protein kinase or (AMPK), the inhibition of mTOR/4EPB1 and an increased phosphorylation of the initiation factor EIF2A. All these events lead to protein synthesis arrest, measured directly using Click-it chemistry based on the incorporation of an aminoacid analog (AHA). Analysis using TCGA database confirmed the correlation between NAMPT and mTOR pathways/translation. In addition, we performed NAMPT immunoprecipitation following mass-spectrometry in cellular extract to identify NAMPT-interacting proteins. Data showed major enrichment of NAMPT-interacting proteins involved in RNA processing, translation, nuclear protein and metabolic processes. The interaction between NAMPT and initiation/elongation factors will be further investigated, however these data suggest potential a direct impact of NAD/NAMPT axis in translational reprogramming in melanoma.