NAD⁺ metabolism: a therapeutic target for age-related metabolic disease

Abstract

Nicotinamide adenine dinucleotide (NAD) is a central metabolic cofactor by virtue of its redox capacity, and as such regulates a wealth of metabolic transformations. However, the identification of the longevity protein silent regulator 2 (Sir2), the founding member of the sirtuin protein family, as being NAD⁺-dependent reignited interest in this metabolite. The sirtuins (SIRT1-7 in mammals) utilize NAD⁺ to deacetylate proteins in different subcellular compartments with a variety of functions, but with a strong convergence on optimizing mitochondrial function. Since cellular NAD⁺ levels are limiting for sirtuin activity, boosting its levels is a powerful means to activate sirtuins as a potential therapy for mitochondrial, often age-related, diseases. Indeed, supplying excess precursors, or blocking its utilization by poly(ADP-ribose) polymerase (PARP) enzymes or CD38/CD157, boosts NAD⁺ levels, activates sirtuins and promotes healthy aging. Here, we discuss the current state of knowledge of NAD⁺ metabolism, primarily in relation to sirtuin function. We highlight how NAD⁺ levels change in diverse physiological conditions, and how this can be employed as a pharmacological strategy.

Figure 1. De novo biosynthesis and salvage pathway of NAD⁺

The first step of the NAD⁺ de novo biosynthesis is the conversion of tryptophan into N-formylkynurenine through an enzymatic reaction catalyzed by either indoleamine 2,3-dioxygenase (IDO) or tryptophan 2,3-dioxygenase (TDO). N-Formylkynurenine is then converted in four successive enzymatic reactions into α-Amino-β-carboxymuconate-ε-semialdehyde (ACMS), which can undergo either enzymatic conversion directed to total oxidation or spontaneous cyclization to quinolinic acid. The following step is the formation of nicotinic acid mononucleotide (NAMN) through the quinolinate phosphoribosyltransferase (QPRT) activity. NAMN is then transformed to nicotinic acid adenine dinucleotide (NAAD) by the nicotinamide mononucleotide adenylyltransferase (NMNAT) enzymes. The final step in the biosynthesis of NAD⁺ is the amidation of NAAD by the NAD synthase enzyme. NAD⁺ is also synthesized through the NAD⁺ salvage pathway from its precursors NA, NAM, or NR. From NA, the first step in NAD⁺ synthesis is catalyzed by nicotinic acid phosphoribosyltransferase (NAPT) and leads to the formation of NAMN. Similarly, NAM is converted by nicotinamide phosphoribosyltransferase (NAMPT), forming NMN, which is also the product of phosphorylation of NR by nicotinamide riboside kinase (NRK). Both NAMN and NMN are then converted by NMNAT, after which the NAMN-derived NAAD requires the final amidation through NAD synthase.

Figure 2. Sirtuins and PARPs as competing NAD⁺-consuming enzymes

Sirtuins are NAD⁺-consuming deacetylases, using NAD⁺ to cleave acetyl groups from acetylated lysine residues of target proteins, in a reaction that generates NAM and 2′-O-acetyl-ADP-ribose. PARP family members are also NAD⁺-consuming enzymes. They catalyze a reaction in which multiple ADP-ribose groups are transferred to a mono ADP ribosylated substrate protein, forming long chains and branches of ADP-ribosyl polymers.

Figure 3. NAD⁺ as a keystone for mitochondrial regulation

NAD⁺ is a rate-limiting metabolite for the SIRT1 enzymatic activity. SIRT1 activity can be increased by different types of physiological or experimental interventions that increase NAD⁺ levels, such as caloric restriction, treatment with STACs/resveratrol, enhancement of NAD⁺ biosynthesis through supplementation with precursors (NA, NR, NMN), or through inhibition of NAD⁺-consuming activities, such as the PARPs or CD38. This stimulation of the SIRT1 deacetylation activity leads to an improvement of the mitochondrial adaptation and ultimately to beneficial effects on health and lifespan.

Figure 4. Compounds increasing NAD⁺ levels

Chemical structures of compounds that increase NAD⁺ levels. We distinguish four types of NAD⁺ boosters, including resveratrol, the primary NAD⁺ precursors NA, NMN and NR, the PARP inhibitors PJ34, olaparib and veliparib, and the CD38 inhibitor 1-{[2-(4-phenoxyphenoxy)ethoxy]methyl}-3-(aminocarbonyl)-pyridinium chloride (Dong et al., 2011).

Figure 5. Mechanisms of action of resveratrol

Resveratrol promotes mitochondrial biogenesis and functions through indirect AMPK and SIRT1 activation. First, compelling evidence suggests that the metabolic actions of resveratrol are based on its ability to act as a mitochondrial poison by inhibiting ATP synthase activity (1). The resulting energy stress will in turn activate AMPK, leading to the stimulation of SIRT1 by increasing NAD⁺ levels. SIRT1 will then activate downstream targets through deacetylation, ultimately leading to an improvement of mitochondrial function. Another potential explanation of how resveratrol acts is based on a recent report showing that resveratrol inhibits phosphodiesterase (PDE) 4 activity and induces cAMP signaling resulting in Ca²⁺ release and, ultimately the activation of the CamKKb-AMPK pathway (2).