Oncogene (2013), 112 & 2013 Macmillan Publishers Limited All rights reserved 0950-9232/13 www.nature.

com/onc

REVIEW

Sorting out functions of sirtuins in cancer

M Roth and WY Chen The sirtuins (SIRT 17) comprise a family of NAD -dependent protein-modifying enzymes with activities in lysine deacetylation, adenosinediphospho(ADP)-ribosylation, and/or deacylation. These enzymes are involved in the cells stress response systems and in regulating gene expression, DNA damage repair, metabolism and survival. Sirtuins have complex roles in both promoting and/or suppressing tumorigenesis. This review presents recent research progress concerning sirtuins and cancer. On one hand, functional loss of sirtuin genes, particularly SIRT1, involved in maintaining genome integrity and DNA repair will promote tumorigenesis because of genomic instability upon their loss. On the other hand, cancer cells tend to require sirtuins for these same processes to allow them to survive, proliferate, repair the otherwise catastrophic genomic events and evolve. The bifurcated roles of SIRT1, and perhaps several other sirtuins, in cancer may be in part a result of the nature of the genes that are involved in the cells genome maintenance systems. The in-depth understanding of sirtuin functions may have signicant implication in designing precise modulation of selective sirtuin members to aid cancer prevention or treatment under dened conditions. Oncogene advance online publication, 22 April 2013; doi:10.1038/onc.2013.120 Keywords: acquired resistance; cancer; genetic instability; longevity; sirtuin; SIRT1

INTRODUCTION Sirtuins are mammalian homologs of yeast silent information regulator 2 (Sir2) encoding a histone deacetylase.1,2 The seven mammalian members of sirtuins (termed SIRT17) share a conserved catalytic core domain (Figure 1). Despite this homology, sirtuins have divergent enzymatic activities with SIRT13 and SIRT7 primarily as lysine deacetylases, SIRT4 primarily as adenosinediphospho (ADP)-ribosyltransferase, SIRT5 as deacetylase and deacylase, and SIRT6 as ADP-ribosyltransferase and deacetylase.15 In addition, primary localization of these proteins in the cell varies signicantly, reecting functional distinction of sirtuin members (see Figure 1). Growing interest in sirtuins largely stems from previous studies of yeast Sir2 showing that in lower organisms, increased Sir2 gene dosage is sufcient to extend lifespan,68 and that Sir2 is at a nexus between caloric restriction, resveratrol or other caloric restriction mimetics and longevity.9,10 More recently, these initial observations have been scrutinized and rened to show a lack of an effect of Sir2 in extending lifespan in lower species as well as in mammals.1113 Further adding to the complex nature of the role of sirtuins in extending lifespan, Kan et al.14 described that transgenic SIRT6 male mice live B1015% longer than their wild-type littermates. The conicting literature on the roles of Sir2 in longevity contributes in part to the confusion of functions of mammalian sirtuins, but may also foretell the complexity of these genes in mammalian cells. Research in the past decade has revealed the perplexed and often controversial roles of sirtuins in promoting versus suppressing cancer. Cancer cells alter normal cellular machineries to promote unabated cell proliferation and maximize their lifespan, but as a consequence of their malignant growth, they cut short the lifespan of the host organism. Hanahan and Weinberg15 have elegantly outlined the fundamental mechanisms of cancer promotion as consisting of sustaining proliferative signaling,

enabling replicative immortality, activating invasion and metastasis and inducing angiogenesis; meanwhile, as normal cells possess innate mechanisms to antagonize cancer-promoting signals, the transformed cells must also overcome tumor suppression mechanisms, namely, by evading growth suppressor signals and resisting cell death. Crucially underlying these hallmarks of cancer is the genetic instability of cancer cells.15 Although cancer biologists tend to classify genes into either tumor promoting or tumor suppressing, only a limited number of genes unambiguously fall into one of these categories, for examples, MYC as an oncogene and retinoblastoma gene RB as a tumor suppressor gene.16 Other genes including sirtuins are less apparent, and the tumor-promoting or -inhibiting properties of the genes may depend on the stages of cancer development and contextual variables such as tissue of origin, the microenvironment and the exact experimental conditions.15 However, rapid progress has been made most recently in the research on mammalian sirtuins and cancer, which may improve our understanding of these elusive genes and will be highlighted in this review.

SIRT1 Biochemical overview SIRT1 shares the greatest homology with yeast Sir2, which was initially characterized as an nicotinamide adenine dinucleotide (NAD) -dependent histone deacetylase2 (so-called class III histone deacetylase that is structurally and biochemically distinct from class I, II and IV histone deacetylases). SIRT1 deacetylates histone H4 lysine 16 (H4K16) as well as histone H3 lysine 9 and 14 (H3K9 and H3K14, respectively).1 Additionally, SIRT1 deacetylates histone H1 lysine 26 (H1K26) and is involved in the deposition of histone variants.17 These modications of histone tails are closely related to gene silencing and heterochromatin formation that may

Department of Cancer Biology, Beckman Research Institute, Duarte, CA, USA. Correspondence: Dr WY Chen, Department of Cancer Biology, Beckman Research Institute, City of Hope, 1500 East Duarte Road, Duarte, CA 91010, USA. E-mail: wechen@coh.org Received 4 January 2013; revised 11 February 2013; accepted 18 February 2013

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Sirtuin protein and catalytic domain 254 SIRT1 40 SIRT2 137 SIRT3 47 SIRT4 51 SIRT5 45 SIRT6 100 SIRT7 314 410 Deacetylase Nucleolus 257 355 301 310 308 314 ART Deacetylase& Deacylase Deacetylase& ART Mitochondria Mitochondria Nucleus 373 399 294 352 495 747 Deacetylase Deacetylase Deacetylase Nucleus Cytosol Mitochondria Enzymatic Activities Primary Localization

Figure 1. Comparison of mammalian sirtuins. The amino-acid length of each mature main form of sirtuins is indicated at the end of each protein, with the conserved catalytic domain illustrated in grey. Principal enzymatic activities and primary cellular localization are shown.

underlie certain biological processes.18,19 Notably, global genomic hypoacetylation at H4K16 is a hallmark of human cancer cells, both cell lines and clinical samples.20 The biological roles of SIRT1, however, are mostly revealed through its deacetylation of a growing number of non-histone substrates that are involved in a wide variety of cellular functions, particularly in metabolic, oxidative/genotoxic and oncogenic stress responses. These substrates can be broadly categorized as: (1) transcriptional factors p53, forkhead transcription factors (FOXO)1, FOXO3a, nuclear factor (NF)-kB, c-MYC, N-MYC, E2F1 and hypoxia-inducible transcription factors (HIF)-1a/HIF-2a, for regulating cell cycle progression and promoting survival under various conditions; (2) DNA repair machinery elements Ku70, RAD51, NBS1, APE1, XPA/C and WRN, for improving DNA damage repair; (3) nuclear receptor, circadian clock and related factors LXR, FXR, ERa, AR, PPARg, PGC1a, CLOCK and PER2, for regulating metabolism; (4) histone-modifying enzymes SUV39H1, p300, TIP60 and PCAF, for regulating gene expression; (5) cell-signaling molecules STAT3, b-catenin and SMAD7, as detailed in previous reviews.21,22 SIRT1, genetic stability and tumor suppression Several studies using mouse models provide evidence that SIRT1 may improve genetic stability and suppress tumor formation (Table 1). SIRT1 is a critical gene for mouse early development. Homozygous deletion of SIRT1 (DExon56 or DExon57) results in perinatal and postnatal lethality in C57BL/6 and 129 strains.23,24 More severe and early embryonic lethality phenotype occurs in another strain with mixed genetic background containing FVB upon SIRT1 deletion (DExon56).25 In C57BL/6-129 background, wild-type and SIRT1 / mouse embryonic stem cells exhibit similar levels of chromosomal abnormality, but SIRT1 / embryonic stem cells are more prone to genomic instability under oxidative stress.26 In mixed FVB background, SIRT1 / early embryos and mouse embryonic broblasts display markedly increased spontaneous gross chromosomal abnormalities accompanied with altered global histone modications.25 The genetic instability in SIRT1 / embryonic cells is attributed to defective DNA damage repair characterized by reduced gH2AX foci formation and reduced recruitment of DNA-damage repair factors to the foci.25,26 In accord with reduced genetic stability upon SIRT1 loss, SIRT1 heterozygous deletion accelerates tumor formation in p53 / knock-out mice.25 Using a conditional SIRT1 overexpression allele that is specically expressed in the cells lining the intestine, Firestein et al.27 showed that SIRT1 overexpression reduces intestinal tumor
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formation in adenomatous polyposis coli (APC)min/ mice. They demonstrated that SIRT1 deacetylates b-catenin and reduces its nuclear presence and transactivation potential. Similarly, Oberdoerffer et al.26 showed that SIRT1 conditional overexpression reduces tumor burden in p53 / mice. In a recent report, Herranz et al.11 undertook a 3-year study of SIRT1 transgenic mice with SIRT1 overexpression under its own promoter, and showed that increased SIRT1 expression by threefold improves healthy mouse aging and reduces spontaneous carcinomas and sarcomas, as well as carcinogeninduced liver cancer incidence. Taken together, in vivo mouse models suggest that SIRT1 may improve genetic stability of normal cells and suppress tumor formation. In addition to genetic stability, SIRT1 may regulate other pathways that contribute to its tumor suppression function. SIRT1 has been shown to inhibit the NF-kB pathway28 that promotes inammation, survival and cancer metastasis.29 NF-kB comprises heterodimers with p65 and p50 subunits. SIRT1 deacetylates the p65/Rel-A subunit of NF-kB and blocks its transactivation of downstream anti-apoptotic target genes, cIAP-2 and BCL-xL.28 Further in line with tumor suppressor function, SIRT1 acts downstream of BReast CAncer 1 (BRCA1) to negatively regulate the anti-apoptotic gene, Survivin, by deacetylating H3K9 on its promoter and thereby repressing its transcription. Thus, BRCA1 ablation, via reduced SIRT1, results in elevated Survivin levels and enhances tumor growth.30 Dysregulation of SIRT1 gene expression and enzymatic activity in cancer Although SIRT1 has tumor suppression function in the mouse studies described above, so far there is no reported functional SIRT1 genetic mutation or deletion, or SIRT1 promoter hypermethylation in human cancer. The Sangers Catalogue of Somatic Mutations in Cancer database registers only 14 uncharacterized point mutations among 4357 unique samples as of December 2012, perhaps as background mutations. In contrast to rare SIRT1 genetic alterations, numerous studies have revealed that human cancer samples display elevated levels of SIRT1 relative to their nontransformed counterparts. These studies span different cancer types including liver,31 breast,32 gastric,33 prostate34,35 and hematopoietic3641 origin. In colorectal cancer, Kabra et al.42 observed reduction of SIRT1 expression, but Nosho et al.43 showed that SIRT1 overexpression correlates with microsatellite instability as well as a CpG island methylator phenotype. Similarly, in mice, overexpression of SIRT1 is reported in leukemia, lymphoma, sarcoma, lung adenocarcinoma and prostate cancer.34,39,44,45 Alteration of SIRT1 gene expression in cancer is mediated by multiple mechanisms affecting transcription, translation and posttranslational modications. At the transcriptional level, tumor suppressor HIC1 directly represses SIRT1 transcription, thereby leading to acetylation and activation of another tumor suppressor p53 (Chen et al.,44 and Figure 2). Loss of HIC1 results in deacetylation of p53 and survival of aged and damaged cells, which may predispose them to cancers.46,47 p53 itself has been shown to directly repress SIRT1 transcription as well.48 Conversely, SIRT1 is upregulated by oncogenic transformation in several settings. Yuan et al.39 have recently shown that transformation of hematopoietic stem/progenitor cells by oncogenic tyrosine kinase BCR-ABL activates SIRT1 transcription in part through STAT5 (signal transducer and activator of transcription 5). STAT5 is downstream of numerous cytokine and growth factor receptorsignaling cascades and is constitutively active in many cancer types.49 STAT5 also acts downstream of BCR-ABL protein in chronic myelogenous leukemia (CML).50,51 STAT5 directly binds the SIRT1 promoter and enhances SIRT1 expression in CML cells39 (see Figure 2). SIRT1 levels are increased stepwise from normal
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Table 1.
Sirtuin SIRT1 Mouse cancer phenotypes with sirtuin gene knockout or overexpression Genotype Constitutive deletion of SIRT1 SIRT1tg Pten / Phenotype SIRT1 knockout inhibited BCR-ABL transformation and CML development SIRT1 transgenic mice had increased incidence of thyroid cancer with lung metastases. SIRT1 expression also increased prostate carcinomas in situ Double heterozygous knock-out mice had increased mammary tumor incidence Intestinal SIRT1 expression reduced intestinal tumors SIRT1 overexpression reduced overall tumor incidence, particularly thymic lymphoma, in p53 / background Transgenic SIRT1 mice had reduced incidence of spontaneous carcinomas and sarcomas, but not lymphoma. SIRT1tg mice had lower incidence of carcinogen-induced liver carcinomas SIRT2 / -female mice developed mammary tumors, whereas -males predominately developed liver cancers and intestinal tumors SIRT3 / -female mice developed mammary tumors with 35% penetrance at about 24 months of age Premature aging and death at postnatal week 3 SIRT6 / mice developed larger intestinal tumors Organ sites of tumors Blood Thyroid and prostate Role of sirtuins Cancer promoting Cancer promoting Reference Yuan et al.39 Herranz et al.75

SIRT1 / p53 / Villin-Cre SIRT1DSTOP APCmin/ Mx-Cre SIRT1DSTOP p53 / SIRT1tg

Mammary glands Intestine Thymus

Cancer suppressing Cancer suppressing Cancer suppressing Cancer suppressing

Wang RH et al.25 Firestein et al.27 Oberdoerffer et al.26 Herranz et al.11

Liver, and other epithelial and mesenchymal tissues

SIRT2

Constitutive deletion of SIRT2 Constitutive deletion of SIRT3 Constitutive SIRT6 deletion Villin-Cre SIRT6/ APCmin/

Liver and intestine in males and mammary glands in females Mammary glands

Cancer suppressing Cancer suppressing Cancer suppressing

Kim et al.142

SIRT3

Kim et al.164

SIRT6

Intestine

Mostoslavsky et al.192 Sebastian et al.200

Abbreviations: APC, adenomatous polyposis coli; CML, chronic myelogenous leukemia; SIRT, sirtuins; SIRT1tg, SIRT1 transgenic. No tumors have been reported in constitutive SIRT4, SIRT5 and SIRT7 knock-out mice.

hematopoietic cells to chronic-phase CML to accelerated and blast crisis CML.39 In the same setting, Yuan et al.39 also showed that SIRT1 can be activated by KRAS transformation. Recently, several groups have demonstrated that MYC (c-MYC and N-MYC) protooncogenes directly activates SIRT1 transcription.52,54 This occurs despite incomplete agreement of its effect on MYC protein stability: three groups showing that SIRT1 and MYC form a positive feedback loop in which SIRT1 deacetylates MYC and enhances MYC protein stability,52,54,55 whereas another group showing that SIRT1 destabilizes c-MYC protein.53 In addition, the cell cycle and apoptosis regulator E2F1 induces SIRT1 transcription when cancer cells are under genotoxic stress.56 At the message level, SIRT1 expression is increased by stabilizing its messenger RNA (mRNA) in cancer cells. MicroRNAs are small, RNA polymerase II-transcribed genes that generally form clusters and operate by binding to the 30 untranslated regions of genes and thereby negatively regulating mRNA stability and/or translation.57 Several families of microRNAs have been shown to negatively regulate SIRT1, particularly miR-34a.58 The chromosomal region where miR-34a resides is frequently lost in human cancers, while reintroduction of miR-34a into cancer cells triggers apoptosis.59,60 Other miR families including miR-200a that target SIRT1 have been characterized as possessing tumor suppressor function.32,57,61 Conversely, SIRT1 mRNA is stabilized by RNA-binding protein HuR62, which has a role in promoting tumor cell proliferation and survival.6365 SIRT1 protein stability is regulated by phosphorylation, sumoylation and ubiquitination. SIRT1 phosphorylation by cell cycle& 2013 Macmillan Publishers Limited

dependent kinase cyclin B/CDK1 controls cell proliferation.66 The dual specicity tyrosine phosphorylation-regulated kinases DYRK1A and DYRK3 phosphorylate SIRT1 and increase SIRT1 enzymatic activity to promote cell survival.67 SIRT1 phosphorylation by c-Jun N-terminal kinase 2 stabilizes the protein,68 whereas SIRT1 phosphorylation by JNK1 facilitates ubiqitination-mediated degradation.69 Under genotoxic stress, the proapoptotic nuclear desumoylase SENP1 removes SIRT1 sumoylation and reduces its deacetylase activity.70 Finally, SIRT1 enzymatic activity could be regulated during tumorigenesis. Deleted in breast cancer 1 (DBC1) is a tumor suppressor that is lost in a portion of breast cancer patients. DBC1 suppresses SIRT1 activity by direct binding to the SIRT1 catalytic core domain.71,72 In contrast, active regulator of SIRT1 (AROS) increases SIRT1 activity through direct interaction with N-terminus of SIRT1 protein.73 Furthermore, increasing SIRT1 expression is concomitant with activation of the mammalian NAD salvage biosynthesis enzyme nicotinamide phosphoribosyltransferase in several types of cancer cells to provide sufcient NAD for SIRT1 functions, which is important for cancer cell survival and stress response.52,74 SIRT1 promotes cancer genomic instability and cancer evolution Several recent studies provide crucial insight into the roles of SIRT1 dysregulation in cancer. Yuan et al.39 demonstrated that SIRT1 homozygous knockout signicantly inhibits BCR-ABL transformation of mouse bone marrow stem/progenitor cells
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4
BCR-ABL
Normal Cells

SIRT1 Survival with fixed DNA Tumor suppression

STAT5

HIC1

p53

Myc

E2F1

DNA repair with fidelity

STAT5A

STAT5B -1116 -1039 -178 -168 -80 -65

SIRT1 Survival with increased c he mutations mo

Evolve to high grade malignancy

-2235 -1838

Cancer Cells

DNA repair with infidelity

HuR UAG AUG

miRNAs 34a, 200a, etc.

Relapse and drug resistance

AAAA

Figure 3. A model for bifurcated SIRT1 roles in cancer through genome maintenance. SIRT1 promotes genome maintenance in both normal and cancer cells under genotoxic stress and DNA damage. Activation of DNA repair with high delity in normal cells improves genome stability and suppresses tumor formation. Activation of DNA repair with low delity in cancer cells prevents catastrophic genomic events and renders cancer cell survival, but allows cancer cells to accumulate nonfatal lesions and more mutations to evolve toward high-grade malignancy and drug resistance under chemotherapy.

AROS

DBC1

254 Ncatalytic core

495 -C

Figure 2. Regulation of SIRT1 gene expression and activities in cancer cells. SIRT1 transcription is activated by Myc, E2F1 or BCR-ABL in part through STAT5, but repressed by HIC1 and p53. The sites for transcriptional factor binding are shown. SIRT1 mRNA is stabilized by HuR but degraded by miR34a and miR200a. SIRT1 activity is enhanced by AROS but inhibited by DBC1.

and development of CML-like myeloid leukemia in a mouse model in the BALB/c strain. In this model, bone marrow cells were harvested from adult wild-type and SIRT1 / mice and transduced by BCR-ABL in vitro, followed by transplantation to lethally irradiated syngenic recipients. The effect of SIRT1 knockout on leukemogenesis is thus not inuenced by SIRT1 loss on mice, but is a result of cell autonomous impact on leukemia cells. Pharmacological inhibition of SIRT1 also inhibits leukemia development to a similar extent as SIRT1 knockout. This study provides the rst genetic evidence for a causal role of dysregulated SIRT1 expression in efcient oncogenic transformation and leukemia progression. In another study, Herranz et al.75 crossed SIRT1 transgenic mice to PTEN / mice, and found that SIRT1 overexpression increased incidence of thyroid carcinomas and their lung metastasis, suggesting that increased SIRT1 gene expression has a direct role in promoting tumor progression. These authors found that SIRT1 enhanced Myc stability, although they did not directly link deacetylation of Myc to protein stability. Furthermore, Herranz et al.75 showed that PTEN / SIRT1 transgenic mice also developed prostate carcinomas in situ, whereas PTEN / or SIRT1 transgenic cohort mice did not, indicating that SIRT1 overexpression may also have a role in promoting tumor initiation. This study provides additional genetic evidence that SIRT1 activation can facilitate tumorigenesis. The above studies ostensibly contrast other mouse genetic studies suggesting roles of SIRT1 in tumor suppression, as described above. The precise mechanisms for this difference are not clear, but recent studies raise the possibility that SIRT1 might differentially regulate genomic stability in normal versus cancer cells (Figure 3). In normal cells, loss of SIRT1 causes defective DNA repair leading to genomic instability. Intriguingly, in cancer cells, SIRT1 knockdown also reduces the efciency of DNA damage
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repair including both non-homologous end joining (NHEJ) and homologous recombination (HR) repair in CML cells76 and osteosarcoma cells.26 It is shown that SIRT1 is recruited to sites of DNA damage, presumably to remodel local chromatin structure to facilitate repair in cancer cells.77,78 Therefore, it is likely that SIRT1 may have a role in genome maintenance in both normal and cancer cells. However, this may lead to complex outcomes in cancer cells. On one hand, the maintenance by SIRT1 may ensure cancer cell survival and proliferation (see more in next section) by avoiding deleterious impact of DNA damage on key oncogenes, given that transformation is accompanied with increased production of reactive oxygen species (ROS) and oxidative DNA damage.79,80 On the other hand, increased DNA repair by SIRT1 could lead to more mutations in cancer cells and further genomic instability for cancer evolution. This is supported by a recent report by Wang et al.76, showing that SIRT1 promotes de novo acquisition of genetic mutations for drug resistance in CML and prostate cancer cells upon therapeutic (by tyrosine kinase inhibitors) and/or genotoxic (by camptothecin) stress. SIRT1 promotion of mutation acquisition is associated with its ability to increase error-prone DNA damage repair in cancer cells, in particular, through deacetylating the key NHEJ repair factor Ku70. Wang et al.76 proposed that the unusual effect of SIRT1 in mutation promotion may be attributed to the compromised DNA repair delity in cancer cells,81,82 and thus new genetic mutations may arise as a consequence of misrepair under stress. Genomic instability is one of the most critical enabling characteristics of cancer. As cancer progresses toward highergrade malignancy, more mutations are acquired, but mechanisms for such increasing mutation acquisition are not well understood.15 The new ndings by Wang et al.76 may suggest a previously underestimated pathway for cancer genomic instability and evolution, that is, to accumulate mutations through enhanced indelity DNA repair, which has important biological signicance for cancer drug resistance. SIRT1 appears to be at the center of such an evolution pathway;83 however, more detailed mechanisms still need to be worked out in the future. SIRT1 promotes cancer hallmark capability Cancer evolves not only with accumulation of genetic alterations, but also with profound epigenetic changes. Cancer cells display global genomic DNA hypomethylation with concurrent hypermethylation at tumor suppressor gene loci. In addition, cancer cells display gross aberrant histone modication patterns relative to normal cells.84 In addition to SIRT1 directly deacetylating
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Functions of sirtuins in cancer M Roth and WY Chen

5 histone substrates to affect the epigenetic state, SIRT1 can deacetylate DNA methyltransferase 1 and can either enhance or hinder its methyltransferase activity,85 thus indirectly affecting global or local DNA methylation patterns. SIRT1 is a component of the polycomb repressor complex (PRC), which is involved in silencing genes during normal development. In cancer cells, PRC is also involved in silencing tumor suppressor genes.86,87 The catalytic subunit of the PRC II complex is EZH2, which is responsible for trimethylation of H3K2788 and itself a noted protooncogene frequently mutated in human cancers.89 SIRT1 directly complexes with EZH2 and other components of the PRC complex and is an integral part of the PRCs silencing functions.90 Kuzmichev et al.45 identied a SIRT1-containing PRC complex, termed PRC4, which is specically found in transformed cells and embryonic stem cells. Inhibition of SIRT1 has been shown to reactivate silenced tumor suppressor genes.91 In addition, SIRT1 interacts and deacetylates the SUV39H1 methyltransferase, promoting histone H3 methylation and fostering heterochromatin formation92 and repressing ribosomal RNA transcription to protect cells from energy deprivationdependent apoptosis.93 The dysregulated epigenome by SIRT1 in cancer cells may act in concert with genetic alterations to facilitate cancer evolution and reach the biological endpoint hallmarks.15 This section will discuss the cancer hallmarks that are affected by SIRT1. Cancer hallmark 1: resisting cell death. SIRT1 regulates multiple cell death pathways. SIRT1 deacetylates p53 at lysine 382, reducing its transcriptional activity and leading to a blockade of p53-dependent apoptosis in response to DNA damage signals.94,95 Similar patterns are observed for p73, which is part of the p53 family.96 Leukemia stem cells are typically refractory to chemotherapy. Recently, Li et al.40 have demonstrated that inhibiting SIRT1 in CD34 CML stem/progenitor cells results in enhanced p53 acetylation, transcriptional activity and apoptosis of these cells, and sensitizes them to the BCR-ABL inhibitor, imatinib, both in vitro and in vivo. This effect is dependent on p53, as cells with p53 knockdown fail to respond to SIRT1 inhibition. This study reveals a novel drug resistance mechanism mediated by SIRT1 in leukemia stem cells. The intrinsic apoptotic pathway is tightly orchestrated by the BCL-2 family, which contains pro- and anti-apoptotic members, of which Bax is the prototypical pro-apoptotic gene.97 Cancer cells can resist apoptosis by sequestering Bax from mitochondria. This is accomplished through the non-canonical action of Ku70.98 SIRT1 regulates Bax sequestration through deacetylation of Ku70, thereby increasing Ku70 interaction with Bax and blocking Bax translocation to mitochondria.99 By modulating Ku70, SIRT1 inhibition induces apoptosis in leukemia cells lacking p53 in vitro and suppresses growth of tumor xenograft in vivo.39 The transcriptional regulator BCL6 is also subjected to deacetylation by SIRT1.100 BCL6 is expressed in the germinal center within lymph nodes and represses genes that are involved in cell cycle regulation, apoptosis and differentiation.101 Translocations involving BCL6 are a frequent event in B-cell lymphomas.102,103 Deacetylation by SIRT1 promotes BCL6 transcriptional repression activity and resists cell apoptosis.100 Heltweg et al.104 demonstrate that a chemical inhibitor of SIRT1, cambinol, has anti-lymphoma properties in vitro and in vivo in part by preventing the deacetylation and activation of BCL6. As described above, SIRT1 can promote cancer cell death resistance by facilitating acquisition of genetic mutations.76 In addition, SIRT1 promotes chemotherapy resistance by enhancing efux of drugs. Chu et al.105 report that SIRT1 is elevated in drugresistant cell lines as well as in patient samples. SIRT1 increases the expression of multidrug resistance 1 in cancer cells, whereas inhibition of SIRT1 sensitizes these cancer cells to anticancer
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drugs. Another group reported that a novel SIRT1 inhibitor, Amurensin G, reduces expression of multidrug resistance 1 and sensitizes otherwise resistant cells to cytotoxic drugs.106 Cancer hallmark 2: sustaining proliferation signaling. As discussed above, SIRT1 is activated by MYC and forms a positive feedback loop to stabilize MYC protein in most studies. Stabilized MYC further enables cell proliferative signaling and stimulates cell growth. This likely explains why Burkitts lymphoma, which is initiated by c-MYC transformation, is most sensitive to the SIRT1 inhibitor cambinol used as a single agent.104 SIRT1 is also shown to regulate WNT signaling,107 and WNT activation is associated with transcriptional activation of c-MYC in high-grade malignancy of colorectal cancer in the serrated route.108 Cancer hallmark 3: evading growth suppressors. As described above, SIRT1 deacetylates and inactivates p53, a negative cell cycle regulator, to bypass growth arrest. Another key tumor suppressor family that is deacetylated by SIRT1 is the FOXO.109112 FOXO transcription factors are involved in cell cycle control113 as well as antioxidant and DNA-damage repair pathways.114 Motta et al.110 showed that SIRT1 deacetylation of FOXO3a suppresses FOXO3-mediated cell apoptosis induction, in parallel to p53 inhibition. Wang et al.115 showed that deacetylation of FOXO3 by SIRT1 or SIRT2 results in its ubiquitination and subsequent degradation, thereby facilitating cell cycle progression and contributing to the promotion of cancer. Cancer hallmark 4: inducing angiogenesis. It is known that SIRT1 controls endothelial angiogenic functions through deacetylating FOXO1 and notch1 intracellular domain during vascular growth.116,117 SIRT1 enhances tumor angiogenesis through negatively modulating d-like ligand 4 (DLL4)/notch signaling in Lewis lung carcinoma xenograft-derived vascular endothelial cells.118 SIRT1 also activates endothelial nitric oxide synthase by deacetylation to enhance nitric oxide production and improve vascular function.119 Cancer hallmark 5: activating invasion and metastasis. Epithelialto-mesenchymal transition (EMT) is an essential process to promote cancer invasion and metastasis. It has been shown that SIRT1 is activated during EMT-like transformation of mammary epithelial cells, in part due to epigenetic silencing of miR-200a, which negatively regulates SIRT1.32 SIRT1 is also a positive regulator of EMT and metastasis of prostate cancer.120 Overexpression of SIRT1 induces EMT through transcription factor ZEB1 in prostate cancer cells; SIRT1 knockdown restores cell-cell adhesion and reverses EMT of prostate cancer cells in vitro and in vivo.120 Cancer hallmark 6: deregulating cellular energetics and tumor microenvironment. The HIF-1 and HIF-2 are activated in cancer cells because of chronically low oxygen tension in the tumor bulk. HIF-1 activates numerous genes that promote angiogenesis, survival and glucose uptake, all necessary for tumor growth. HIF-1 is emerging as a major player in metastasis, chemo- and radiotherapy resistance.121 HIF-1a is the regulatory subunit of HIF-1 and is subjected to post-translational modication.122,123 Lim et al.124 showed that SIRT1 can deacetylate HIF-1a and repress its biological function in vitro and in tumor xenografts. However, such effect of SIRT1 on HIF-1a has been recently disputed by Laemmle et al.,125 showing that SIRT1 stabilizes HIF-1a under hypoxia and SIRT1 inhibition impairs hypoxic response of hepatocellular carcinoma cells. The study by Laemmle et al.,125 is in line with a previous report demonstrating that SIRT1 helps cells survive against hypoxic environment by activating HIF-2a.126
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6 Summary of SIRT1 and cancer All the data so far suggest that SIRT1 has both pro- and anticancer roles. We propose that SIRT1 may act as a genome caretaker in normal cells, and therefore suppress tumorigenesis. However, upon oncogenic events, tumor cells co-opt SIRT1-regulated cellular pathways to promote unabated proliferation, progression, resisting death signals and genetic/epigenetic evolution. Such dual roles in tumorigenesis are not unprecedented. For examples, telomerase reverse transcriptase (TERT) can both hinder and promote tumor formation;16 Transforming growth factor-b can be antiproliferative, but is redirected toward promoting EMT and high-grade malignancy at the later stages of tumorigenesis;127 aberrant DNA hypermethylation by DNA methyltransferase 1 promotes tumorigenesis by silencing tumor suppressor genes, but DNA methyltransferase 1 deletion or expression of a hypomorphic allele in mice results in tumorigenesis.128,129 As such, the precise role of SIRT1 in tumorigenesis may be dependent on cellular and molecular contexts. SIRT2 Biochemical overview SIRT2 shares certain biochemical parallels with SIRT1. SIRT2 has thus far been characterized primarily as an NAD -dependent protein deacetylase. Although being mostly cytoplasmic, SIRT2 can translocate to the nucleus where it deacetylates H4K16 during mitosis.130,131 SIRT2 has also been shown to deacetylate H3K56.132 Much fewer non-histone substrates have thus far been identied for SIRT2. a-tubulin is the rst known SIRT2 substrate,133 but the exact biological role of tubulin acetylation/deacetylation remains elusive. Sharing substrates with SIRT1, SIRT2 also deacetylates FOXO1,134,135 FOXO3115,136 and p53137139 for regulating autophagy, apoptosis and differentiation. Most recently, SIRT2 is shown to regulate programmed necrosis by targeting receptorinteracting proteins.140 Mitotic regulation is another biological function frequently ascribed to SIRT2, as detailed in a previous review.141 Kim et al.142 described that SIRT2 regulates mitosis by deacetylating components of the anaphase-promoting complex/ cyclosome ubiquitin ligase machinery to enhance its activity, directing degradation of aurora kinases as cells exit mitosis. In the dietary obesity setting, the intra-adipose tissue hypoxia and activation of HIF-1a results in transcriptional repression of SIRT2, leading to increased acetylation of PGC-1a, and affecting b-oxidation and mitochondrial gene expression.143 SIRT2 and cancer Analogous to the bifurcated results with SIRT1 in cancer, SIRT2 has both roles in tumor suppression and promotion. SIRT2 is reported to be a tumor suppressor in gliomas144,145 and melanomas,146 with chromosomal loss at the SIRT2 locus and point mutations within the catalytic domain that abrogates its enzymatic activity. SIRT2 expression is reduced in certain cancers,142,145,147 and gene silencing by histone deacetylation is associated with SIRT2 downregulation in glioma cells.148 However, forced SIRT2 expression only moderately reduces proliferation of glioma cells.149 The direct support of SIRT2s tumor suppressor role is provided by Kim et al.,142 showing that SIRT2 knock-out mice develop tumors in various organs due to abnormal chromosomal segregation and aneuploidy caused by increased expression of mitotic regulators including aurora kinases upon SIRT2 knockout. On the other hand, it is observed that SIRT2 may promote oncogenic phenotypes. SIRT2 is increased in acute myeloid leukemia cells compared with normal bone marrow cells, and SIRT2 inhibition causes apoptosis of acute myeloid leukemia cells in vitro.150 Hou et al.151 showed that SIRT2 expression positively correlates with cortactin in high-grade prostate cancer samples and predicts poor prognosis. Knockdown of SIRT2 induces p53
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accumulation and promotes apoptosis of cancer cells.138,139 Recently, Liu et al.152 showed that SIRT2 is activated by N-MYC in neuroblastoma cells and by c-MYC in pancreatic cancer cells. Activated SIRT2 stabilizes N-MYC and c-MYC protein and promotes cell proliferation through downregulating ubiquitin-protein ligase NEDD4. Liu et al.152 also found that SIRT2 increases Aurora A expression in these cells, in contrast to the nding by Kim et al.142 In this regard, it is worthy to note that Aurora A has bifurcated roles in cancer with either gene overexpression or knockout promoting tumorigenesis.153,154 More studies are needed to further delineate the precise roles of SIRT2 in cancer. SIRT3 Biochemical overview SIRT3 possesses NAD -dependent deacetylase as well as ADPribosyltransferase activity. SIRT3 is the major mitochondrial deacetylase with a broad range of substrates.155 Although principally localized to mitochondria, SIRT3 can be detected in the nucleus as well.156 There are two forms of SIRT3, and processing of the N-terminus results in a smaller protein that localizes to mitochondria.157,158 Within the nucleus, SIRT3 deacetylates H4K16 and H3K9 to repress transcription.156 In mitochondria, SIRT3 regulates metabolism, energy homeostasis, thermogenesis and mitochondrial biogenesis. For a more thorough review on SIRT3 and metabolic regulation, we refer the readers to other reviews.159,160 SIRT3 increases fatty acid b-oxidation by deacetylating, and therefore enhancing the activity of long-chain acyl CoA dehydrogenase.161 The catabolism of fatty acids by boxidation yields acetyl CoA, which gets further metabolized by the Krebs cycle. SIRT3 regulates several components of the Krebs cycle and electron transport chain to promote adenosine triphosphate production.160,162 This is punctuated by the phenotype seen in SIRT3 knock-out mice that show reduced cellular adenosine triphosphate levels as well as increased ROS.162 Uncontrolled ROS in the cell results in DNA and protein damage, aging and cancer.163 SIRT3 and cancer SIRT3 acts like a tumor suppressor as evidenced by increased mammary tumor formation in SIRT3 knock-out mice.164 One major tumor suppression mechanism of SIRT3 is through modulation of ROS. As noted above, SIRT3 / cells show increased ROS due to faulty electron transport.162,164,165 Another means by which SIRT3 regulates ROS is through the activation of antioxidant enzymes, such as superoxide dismutase that is a major mitochondrial detoxifying enzyme.166,167 Further, Someya et al.168 revealed that SIRT3 lowers ROS by deacetylation and activation of mitochondrial isocitrate dehydrogenase 2, which in turn increases NADPH levels and the abundance of active glutathione, a major cellular antioxidant. Deacetylation and activation of FOXO3a by SIRT3 also promotes its activation of antioxidant enzymes.169 Yet another mechanism whereby SIRT3 reduces ROS, and therefore tumorigenesis is in its capacity to destabilize HIF-1a.165,170 Because of SIRT3s effect in regulating oxidative phosphorylation and HIF-1, SIRT3 / cells may be further prone to cancer by shifting its metabolism toward aerobic glycolysis, a phenomenon termed the Warburg effect, which is thematic of most cancer cells.171,172 The studies with SIRT3 knock-out mice lend credence to the notion that SIRT3 acts to protect against tumorigenesis. However, it is unclear why SIRT3 knockout only increases mouse mammary tumors, given that these pathways are not mammary gland-specic. In human cancers, Kim et al.164 showed that SIRT3 protein and mRNA are reduced in breast cancer. Consistently, Bell et al.165 showed that knockdown of SIRT3 in human cancer cells increases tumors size and reduces latency when injected into mice. Conversely, forced expression of SIRT3 blocks proliferation and reduces tumor xenografts. Finley et al.170 found that about 20% of
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7 all human cancer samples and 40% of breast and ovarian cancer samples contain deletions of SIRT3. Recently, a novel tumor suppressor function of SIRT3 was uncovered in its ability to deacetylate the proto-oncogene, Skp2.173 Skp2 is overexpressed in a wide range of cancers, and its expression levels foretell a poor clinical prognosis. Skp2 functions biochemically as an E3 ubiquitin ligase and is responsible for targeting numerous tumor suppressors such as p21 and p27 for proteasome-mediated degradation.174 Deacetylation of Skp2 by SIRT3 leads to its nuclear import where Skp2 is excluded from targeting E-cadherin. In contrast, acetylated Skp2 gets exported to the cytoplasm where it ubiquitinates E-cadherin for proteasome-mediated degradation.173 Reduced E-cadherin is observed in many cancers and is a feature of EMT and cancer metastases.175 Together, these studies indicate that SIRT3 suppresses tumorigenesis. Yet, as with SIRT1 and SIRT2, there are opposing studies that propose a tumor-promoting role for SIRT3. Alhazzazi et al.176 showed that SIRT3 levels are elevated in oral squamous carcinoma cell lines and patient samples, and that knockdown of SIRT3 in OSCC lines reduces cell viability and proliferation as well as tumors in xenografts. Ectopic SIRT3 expression reverses p53-induced cell cycle arrest and senescence, which would provide a mechanism for a tumor-promoting role of SIRT3.177 Like SIRT1, SIRT3 can deacetylate Ku70 and thereby facilitate its interaction with Bax to mitigate Bax-mediated apoptosis.178 These studies, albeit few and contradictory, provide some evidence that SIRT3 may have a cellprotective role under stress and may give cancer cells a growth advantage. SIRT4 SIRT4 is a mitochondrial ADP-ribosyltransferase without recognized deacetylase activity. Its primary function thus far uncovered is in regulation of metabolic function. In vitro, SIRT4 can ADP-ribosylate histones.179 SIRT4 ADP-ribosylates glutamate dehydrogenase and inhibits its catalytic activity. Glutamate dehydrogenase catalyzes the conversion of glutamate to a-ketoglutarate.179,180 Yang et al.181 reported that SIRT3 and SIRT4 are required for cell viability in response to genotoxic stress along with nicotinamide phosphoribosyltransferase to maintain NAD levels within mitochondria. To date, there are no systematic studies evaluating the role of SIRT4 in cancer, although Bradbury et al.41 revealed that SIRT4 levels are consistently down in acute myeloid leukemia samples. Speculatively, as SIRT4 regulates a-ketoglutarate production from glutamate, we wonder what role SIRT4 may have on a-ketoglutarate-dependent enzymes that have been shown to be involved in tumorigenesis, such as the TET enzymes and jmjC histone demethylases.182,183 Metabolic dysfunction leading to abnormal epigenetic regulation and ultimately tumorigenesis has a precedent, as is seen with isocitrate dehydrogenase mutations in several types of cancers.184,185 SIRT5 Mitochondrial SIRT5 is perhaps the most unusual of sirtuins, as it possess NAD -dependent deacetylase as well as deacylase (demalonylase and desuccinylase) activities.4 The best studied substrate of SIRT5 is carbamoyl phosphate synthetase 1 for the rate-limiting step in urea synthesis. Several groups have demonstrated that SIRT5 can deacetylate, demalonylate and desuccinylate carbamoyl phosphate synthetase 1.4,186,187 Deacetylation of carbamoyl phosphate synthetase 1 activates its enzymatic activity, which promotes the production of urea and clearance of excessive ammonia from the cell. The biological signicance of demolonylation and desuccinylation of carbamoyl phosphate synthetase 1 by SIRT5 remains unknown.188 To date, there are no studies evaluating the role of SIRT5 in tumorigenesis.
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SIRT6 Biochemical overview SIRT6 is a chromatin-bound NAD -dependent deacetylase and ADP-ribosyltransferase.189 Its major histone substrates are H3K9 and H3K56.190,191 Michisita et al.190 showed that SIRT6 associates with telomeres and deacetylates H3K9 to maintain proper telomeric chromatin, and disruption of telomere functions likely accounts for the premature aging phenotype observed in SIRT6 / mice.192 SIRT6 has an integral role in DNA repair pathways including BER,192 HR193,194 and NHEJ.195,196 SIRT6 can ADP-ribosylate PARP1, leading to its activation and promotion of double-strand break repair.196 Intriguingly, overexpression of SIRT6, but not SIRT1, increases NHEJ and HR repair efciency in nonmalignant cells.196 Kawahara et al.197 demonstrated that SIRT6 is recruited with NF-kB to deacetylate H3K9 and silence its target genes. SIRT6 is also a corepressor of HIF1 and represses expression of the genes involved in energy metabolism by deacetylating H3K9.198

SIRT6 and cancer Forced expression of SIRT6 induces apoptosis of cancer cells but not in nontransformed cells.199 SIRT6-mediated apoptosis of cancer cells requires its ADP-ribosyltransferase, but not deacetylase, activity, as well as the intact ataxia telangiectasia mutated (ATM) and p53 pathways.199 Mice with constitutive SIRT6 knockout display genetic instability and die of a premature aging syndrome B25 days postnatally.192 Using a conditional gene knock-out strategy, Sebastian et al.200 very recently demonstrated a role of SIRT6 in tumor suppression. These authors revealed that loss of SIRT6 is sufcient to transform immortalized mouse embryonic broblasts. Conditional knockout of SIRT6 in the intestine of APCmin/ mice results in increased tumor incidence, in part by derepression of Myc activity and activation of aerobic glycolysis. They further showed that SIRT6 is frequently deleted and its expression is signicantly reduced in human cancer samples. Together with SIRT6s roles in genome maintenance, the data so far suggest that SIRT6 may act as a tumor suppressor.

SIRT7 Biochemical overview SIRT7 is primarily a deacetylase and is localized to the nucleolus as well as the nucleus.3 Within nucleoli, SIRT7 regulates ribosomal DNA gene expression in part by activating RNA polymerase I.201 SIRT7 deacetylates H3K18, specically, to repress transcription.202 SIRT7 is also shown to deacetylate p53 in vitro and in vivo, and SIRT7 ablation increases p53-mediated apoptosis.203

SIRT7 and cancer Several groups showed that SIRT7 is elevated in human cancers of breast,204 thyroid205 and liver206 origin. Ford et al.201 demonstrated that knockdown of SIRT7 inhibits cell growth and induces apoptosis. Barber et al.202 showed that knockdown of SIRT7 in cancer cell lines reduces cell growth both in vitro and in vivo, and demonstrated a novel oncogenic mechanism of SIRT7 by deacetylating H3K18. Interestingly, many different cancers display global hypoacetylation of H3K18, which is associated with a poor prognosis in prostate cancers.207,208 Viral oncogenes can stimulate hypoacetylation of H3K18.209,210 Consistently, Barber et al.202 showed that knockdown of SIRT7 abolishes adenoviral E1A-mediated proliferation and transformation of broblasts. These studies point to a growth advantage that SIRT7 renders cancer cells. Further in vivo studies should be undertaken to evaluate roles of SIRT7 in cancer in the future.
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Functions of sirtuins in cancer M Roth and WY Chen

8 CONCLUDING REMARKS Studies of sirtuins are rapidly growing in the eld of cancer and other diseases. Sirtuins are generally involved in stress response, DNA damage repair and metabolism. SIRT1, SIRT2 and SIRT3 appear to have both roles in tumor inhibition and promotion. Despite the controversy, it appears that SIRT1 has a consistent role in mediating cancer cell survival, in particular for drug resistance, and that SIRT6 has more pronounced roles in tumor suppression. Currently, both sirtuin activators and inhibitors have been developed or under development, and they may be suited for distinct therapeutic purposes.211 For cancer treatment, inhibitors like tenovin-6212 and cambinol,104 which targets both SIRT1 and SIRT2, have shown antitumor effects in vivo. However, sirtuin modulators are in general not specic and potent enough. Given the complex roles of each sirtuin, future effort is needed to develop more selective sirtuin modulators for human use to treat cancer or other diseases. We expect to see continued rapid growth in sirtuin research in coming years, which will help us better understand functions of this family of proteins for the good of human health. CONFLICT OF INTEREST
The authors declare no conict of interest. 17 Vaquero A, Scher M, Lee D, Erdjument-Bromage H, Tempst P, Reinberg D. Human SirT1 interacts with histone H1 and promotes formation of facultative heterochromatin. Mol Cell 2004; 16: 93105. 18 Vaquero A. The conserved role of sirtuins in chromatin regulation. Int J Dev Biol 2009; 53: 303322. 19 Guarente L. Sir2 links chromatin silencing, metabolism, and aging. Genes Dev 2000; 14: 10211026. 20 Fraga MF, Ballestar E, Villar-Garea A, Boix-Chornet M, Espada J, Schotta G et al. Loss of acetylation at Lys16 and trimethylation at Lys20 of histone H4 is a common hallmark of human cancer. Nat Genet 2005; 37: 391400. 21 Houtkooper RH, Pirinen E, Auwerx J. Sirtuins as regulators of metabolism and healthspan. Nat Rev Mol Cell Biol 2012; 13: 225238. 22 Saunders LR, Verdin E. Sirtuins: critical regulators at the crossroads between cancer and aging. Oncogene 2007; 26: 54895504. 23 McBurney MW, Yang X, Jardine K, Hixon M, Boekelheide K, Webb JR et al. The mammalian SIR2alpha protein has a role in embryogenesis and gametogenesis. Mol Cell Biol 2003; 23: 3854. 24 Cheng HL, Mostoslavsky R, Saito S, Manis JP, Gu Y, Patel P et al. Developmental defects and p53 hyperacetylation in Sir2 homolog (SIRT1)-decient mice. Proc Natl Acad Sci USA 2003; 100: 1079410799. 25 Wang RH, Sengupta K, Li C, Kim HS, Cao L, Xiao C et al. Impaired DNA damage response, genome instability, and tumorigenesis in SIRT1 mutant mice. Cancer Cell 2008; 14: 312323. 26 Oberdoerffer P, Michan S, McVay M, Mostoslavsky R, Vann J, Park SK et al. SIRT1 redistribution on chromatin promotes genomic stability but alters gene expression during aging. Cell 2008; 135: 907918. 27 Firestein R, Blander G, Michan S, Oberdoerffer P, Ogino S, Campbell J et al. The SIRT1 deacetylase suppresses intestinal tumorigenesis and colon cancer growth. PLoS One 2008; 3: e2020. 28 Yeung F, Hoberg JE, Ramsey CS, Keller MD, Jones DR, Frye RA et al. Modulation of NF-kappaB-dependent transcription and cell survival by the SIRT1 deacetylase. EMBO J 2004; 23: p 23692380. 29 Wang S, Liu Z, Wang L, Zhang X. NF-kappaB signaling pathway, inammation and colorectal cancer. Cell Mol Immunol 2009; 6: 327334. 30 Wang RH, Zheng Y, Kim HS, Xu X, Cao L, Luhasen T et al. Interplay among BRCA1, SIRT1, and Survivin during BRCA1-associated tumorigenesis. Mol Cell 2008; 32: 1120. 31 Jang KY, Noh SJ, Lehwald N, Tao GZ, Bellovin DI, Park HS et al. SIRT1 and c-Myc promote liver tumor cell survival and predict poor survival of human hepatocellular carcinomas. PLoS One 2012; 7: e45119. 32 Eades G, Yao Y, Yang M, Zhang Y, Chumsri S, Zhou Q. miR-200a regulates SIRT1 expression and epithelial to mesenchymal transition (EMT)-like transformation in mammary epithelial cells. J Biol Chem 2011; 286: 2599226002. 33 Cha EJ, Noh SJ, Kwon KS, Kim CY, Park BH, Park HS et al. Expression of DBC1 and SIRT1 is associated with poor prognosis of gastric carcinoma. Clin Cancer Res 2009; 15: 44534459. 34 Huffman DM, Grizzle WE, Bamman MM, Kim JS, Eltoum IA, Elgavish A et al. SIRT1 is signicantly elevated in mouse and human prostate cancer. Cancer Res 2007; 67: 66126618. 35 Jung-Hynes B, Nihal M, Zhong W, Ahmad N. Role of sirtuin histone deacetylase SIRT1 in prostate cancer. A target for prostate cancer management via its inhibition? J Biol Chem 2009; 284: 38233832. 36 Kozako T, Aikawa A, Shoji T, Fujimoto T, Yoshimitsu M, Shirasawa S et al. High expression of the longevity gene product SIRT1 and apoptosis induction by sirtinol in adult T-cell leukemia cells. Int J Cancer 2012; 131: 20442055. 37 Wang JC, Kafeel MI, Avezbakiyev B, Chen C, Sun Y, Rathnasabapathy C et al. Histone deacetylase in chronic lymphocytic leukemia. Oncology 2011; 81: 325329. 38 Jang KY, Hwang SH, Kwon KS, Kim KR, Choi HN, Lee NR et al. SIRT1 expression is associated with poor prognosis of diffuse large B-cell lymphoma. Am J Surg Pathol 2008; 32: 15231531. 39 Yuan H, Wang Z, Li L, Zhang H, Modi H, Horne D et al. Activation of stress response gene SIRT1 by BCR-ABL promotes leukemogenesis. Blood 2012; 119: 19041914. 40 Li L, Wang L, Li L, Wang Z, Ho Y, McDonald T et al. Activation of p53 by SIRT1 inhibition enhances elimination of CML leukemia stem cells in combination with imatinib. Cancer Cell 2012; 21: 266281. 41 Bradbury CA, Khanim FL, Hayden R, Bunce CM, White DA, Drayson MT et al. Histone deacetylases in acute myeloid leukaemia show a distinctive pattern of expression that changes selectively in response to deacetylase inhibitors. Leukemia 2005; 19: 17511759. 42 Kabra N, Li Z, Chen L, Li B, Zhang X, Wang C et al. SirT1 is an inhibitor of proliferation and tumor formation in colon cancer. J Biol Chem 2009; 284: 1821018217.

ACKNOWLEDGEMENTS
We thank the research support from the National Cancer Institute of the National Institutes of Health under award number R01 CA143421, and the State of California Tobacco Related Disease Research Program (TRDRP) award 20XT-0121 to WYC. The contents are solely the responsibility of the authors and do not represent the ofcial views of the National Institutes of Health.

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