Human SirT2 co-localizes with microtubules in cytoplasm. Thus, knockdown of SirT2 results in tubulin hyperacetylation. SirT2 is also able to bind FOXO3a, reducing its acetylation levels.

SirT3 is localized in mitochondrial inner membrane, and is activated by caloric restriction. SirT4 is also mitochondrial, but no NAD⁺-dependent deacetylase activity has so far been detected for this sirtuin. SirT4 uses NAD⁺ for ADP-ribosylation of glutamate dehydrogenase, resulting in inhibition of insulin secretion and enzymatic activity of β-cells. The third mitochondrial sirtuin, SirT5, is localized in the inter-membrane space and can deacetylate cytochrome c. The last two sirtuins, SirT6 and SirT7, have recently been characterized. SirT6 exerts an NAD⁺-dependent histone H3 K9 deacetylase action able to modulate telomeric chromatin functions (Alcaín and Villalba, 2009). SirT6 has a role in genome stability. In mammalian cells subjected to oxidative stress, SirT6 is recruited on DNA double-strand breaks (DSB) and stimulates repair through both homologous and non-homologous end-joining recombination. Moreover, some results indicate that SirT6 is physically associated with poly [adenosine diphosphate (ADP)-ribose] polymerase 1 (PARP1). The amount of SirT6–PARP1 complexes increase after DNA damage. PARP1 is mono-ADP-ribosylated on lysine residue K521, thus increasing PARP1 activity and enhancing DSB repair under oxidative stress (Mao et al., 2011). SirT7 is a predominantly nucleolar protein associated with active rRNA genes (rDNA), where it interacts with RNA polymerase I and histones (Alcaín and Villalba, 2009). SirT7 does not seem to have any NAD⁺-dependent activity. Other functional characteristics of SirT6 and SirT7 remain to be further elucidated.

SirT1 is responsible for heterochromatin formation by a deacetylation process. Histone deacetylation can in turn facilitate histone methylation, thus enhancing global transcriptional repression. SirT1 has no effect on H3K4 methylation (a marker of open chromatin) and promotes H3K9 tri-methylation through its involvement with the histone methyltransferase Suv39H1 (Liu et al., 2009; Bosch-Presegue and Vaquero, 2011).

SirT2 and SirT3 are responsible for regulation of H4K16 acetylation. Deacetylation occurs specifically during cell-cycle in G2/M transition, in particular when SirT2 is shuttled to the nucleus (Shogren-Knaak et al., 2006). The mechanism of SirT3 is more complex. Although SirT3 is localized in mitochondria, its feeble nuclear expression is probably responsible for deacetylation of H4K16ac and H3K9ac in some settings (McGuinness et al., 2011). The finding that SirT3 knockdown does not lead to global H4K16 hyperacetylation suggests that its action occurs in limited regions of the genome.

SirT1 has been shown to be significantly up-regulated in different types of cancer including acute myeloid leukemia (AML), prostate, colon, and skin cancers.

Given that only SirT1 and SirT2 seem to control levels of H4K16 acetylation in many settings, and that only SirT1 is proven to be altered in certain cancers, H4K16ac loss in cancer has been correlated with SirT1 deregulated activity.

Nevertheless, SirT1 seems to play contradictory roles, both as tumor suppressor or tumor promoter (Deng, 2009; Bosch-Presegue and Vaquero, 2011). The initial evidence that SirT1 acts as tumor promoter derives from its repressive effect on tumor suppressor p53 (Deng, 2009). By interacting physically with p53, SirT1 blocks its functions through deacetylation at the C-terminal K382 residue (Vaziri et al., 2001). Moreover, SirT1 overexpression in cancer represses p53-dependent cell-cycle arrest and apoptosis in response to DNA damage and oxidative stress. DNA damage-induced acetylation of p53 leads to its activation. In response to damage, SirT1 binds to and deacetylates p53, thus reducing its functional and transcriptional activities. Overexpression of SirT1 disrupts p53-dependent pathways, resulting in a significant reduction in the cell’s ability to respond to stress and DNA damage. In contrast, the inhibition of SirT1 potentiates p53-dependent apoptosis (Deng, 2009; Rahman and Islam, 2011). These findings underline the involvement of SirT1 in tumorigenesis, given that SirT1 overexpression may increase the risk of cancer in mammals by inhibiting p53 and potentially other tumor suppressor genes, including FOXO family members, p73, Rb, and several others.

In contrast, several studies have suggested that SirT1 also has a tumor suppressor role. Decreased SirT1 levels have been reported for glioma, bladder, prostate, and ovarian cancers. Furthermore, some studies have suggested that SirT1 overexpression in APC^−/+ mice reduces rather than increases colon cancer formation (Firestein et al., 2008). This action seems to be caused by SirT1 deacetylation of β-catenin, which promotes cytoplasmic localization of the nuclear-localized oncogenic form of β-catenin. In addition, in SirT1-deficient mice, embryos die at middle gestation stages, displaying increased acetylation of H3K9 and H4K16, reduced chromosome condensation and aberrant mitosis (Firestein et al., 2008; Deng, 2009). SirT1 deficiency also causes reduced DNA double-strand break repair and radiation sensitivity (Wang et al., 2008). It has also been suggested that SirT1 plays a role as tumor suppressor by regulating c-Myc. c-Myc binds to and induces SirT1. This interaction results in decreased c-Myc stability, compromising its transformational capability. These data provide strong evidence for a tumor suppressor role of SirT1 in these settings. Importantly, it still seems possible that the expression level and activity of SirT1 may modulate a delicate balance between suppression and promotion of oncogenesis, thus displaying dual dependence on its spatial and temporal distribution and stage of tumorigenesis (Bosch-Presegue and Vaquero, 2011). A potential tumor suppressor role has also been proposed for the other human sirtuins (McGuinness et al., 2011). This hypothesis is supported by several findings such as the reduction of SirT2 in a large number of human brain tumor cell lines, and its involvement in cell-cycle progression. SirT3 is the only mitochondrial sirtuin implicated in tumorigenesis. Its reduction in several cancers leads to an increase in ROS (reactive oxygen species) production, which results in enhanced tumor growth (Kim et al., 2010). SirT5 overexpression has been found in a study of pancreatic cancer (Ouaïssi et al., 2008). Recently, a role for SirT6 and SirT7 in tumorigenesis has also been proposed. SirT6 might be involved as a result of its control of the NFκB pathway and DNA double-strand repair. SirT7, whose expression inversely correlates with the tumorigenic potential in several murine cell lines (Vakhrusheva et al., 2008), displays increased expression levels in breast cancer (Ashraf et al., 2006).

It is well known that overexpression of Sir2 (or its orthologs) can extend organism lifespan in a wide range of lower eukaryotes (Bosch-Presegue and Vaquero, 2011). Sir2 function is often correlated to caloric restriction. The link between the role of sirtuins, caloric restriction, and longevity was first described in S. cerevisiae. In yeast, caloric restriction leads to increased replicative lifespan. Lifespan extension has not been observed in yeast lacking Sir2 (Lin et al., 2000). Currently, the role of sirtuins in the regulation of mammalian lifespan is still unclear. However, considering that sirtuins are an evolutionary conserved family of proteins, it is fair to speculate on their role in the modulation of aging-related processes in higher organisms (Westphal et al., 2007; Brooks and How, 2009). In humans, the aging process is associated with telomere erosion. SirT1 and SirT6 have a role in both the maintenance of telomeres and their function, as well as the aging process. Recent studies have demonstrated that reduction or removal of SirT6 results in telomere dysfunction and end-to-end chromosomal fusions. Symptoms displayed in the absence of SirT6 are similar to those caused by a premature aging disease, known as Werner’s syndrome. To date, little is known about other sirtuins, but no evidence suggests their involvement with telomere function, formation, and stability (Westphal et al., 2007; McGuinness et al., 2011). Recently, SirT1 has been shown to promote replicative senescence in response to chronic oxidative stress, by upregulation of the p19^ARF senescence regulator, which in turn positively regulates p53 via inhibition of MDM2, a protein mediating p53 degradation (Chua et al., 2005). This effect seems to be in contrast to Sir2 function in yeast, which extends replicative lifespan.

Splitomicin and its derivatives were discovered in a screening for inhibitors of yeast Sir2. While splitomicin did not act very efficiently against human sirtuins, a series of its analogs with a different orientation of the β-phenyl group were developed and characterized for their activity on SirT2 (Bedalv et al., 2001; Neugebauer et al., 2008). Sirtinol is mainly an inhibitor of the yeast Sir2 and human SirT2 (Grozinger et al., 2001). Its inhibitory effect on growth has been described in human breast, lung, and prostate cancer cells, though using a high micromolar range.

Another potent and selective inhibitor of SirT2 is AGK2 with IC₅₀ of 3.5μM. It was demonstrated that the inhibition of SirT2 protects against dopaminergic cell death in a Drosophila model of Parkinson’s disease (Outeiro et al., 2007).

Suramin is a potent inhibitor of many sirtuins: SirT1 with IC₅₀ of 297nM, SirT2 with IC₅₀ of 1.15μM, and SirT5 with IC₅₀ of 22μM. However, its reported neurotoxicity limits therapeutic use (Schuetz et al., 2007; Trapp et al., 2007).

Cambinol and salermide (Heltweg et al., 2006; Lara et al., 2009) are reported as inhibitors of SirT1 and SirT2. Cambinol shows the most potent activity against Burkitt lymphoma cell lines by a mechanism involving BCL6 acetylation. Salermide induces apoptosis in cancer cells in a p53-independent manner. The indole derivative, EX-527, is a cell-permeable, selective inhibitor of SirT1 (IC₅₀=98nM). Treatment with EX-527 dramatically increases acetylation at K382 of p53 after DNA damage induction in primary human mammary epithelial cells and several cell lines (Solomon et al., 2006). The direct interaction between SirT1 and p53 highlights potential SirT1 involvement in p53 functions, including differentiation, DNA repair, and aging. Currently, EX-527 is in phase 1 clinical trials for the treatment of Huntington’s disease. In preclinical studies this inhibitor was shown to reduce neuronal death.

The only SirT inhibitors that have been tested in cancer mice models are cambinol, tenovins, and UVI5008. Tenovins, a family of small-molecule inhibitors, are able to inhibit SirT1 at single-digit micromolar concentration and prevent tumor growth in vivo in a p53-dependent manner as single agents. Tenovins have a potential therapeutic interest for the development of new drugs that act against tumor cells overexpressing SirT1 (Lain et al., 2008).

UVI5008 has recently been reported as a novel epigenetic modifier able to inhibit simultaneously histone deacetylases, sirtuins, and DNA methyltransferases. This compound induces cancer cell-selective death in several human models. Importantly, UVI5008 action is independent of p53, Bcl-2-modifying factor (BMF), and TNF-related apoptosis-inducing ligand (TRAIL), affecting the growth of tumor cells deficient or mutated for these factors. This finding potentiates its application in therapy by overcoming potential drug resistance limits. The simultaneous modulation of three classes of epigenetic enzymes offers a promising strategy for applying a single drug exerting multiple-interventions against cancer (Nebbioso et al., 2011).