Senescence: a new weapon for cancer therapy

Senescence: a new weapon for cancertherapyJuan Carlos Acosta and Jesus Gil

Cell Proliferation Group, MRC Clinical Sciences Centre, Imperial College London, Hammersmith Campus, London W12 0NN, UK

Review

Senescence is a stable cell cycle arrest that can beactivated by oncogenic signaling and manifests withchanges in cellular organization and gene expression,such as the induction of a complex secretome. Impor-tantly, senescence limits tumor progression and deter-mines the outcome of conventional anticancertherapies. In recent years, therapeutic approaches suchas p53 reactivation, inhibition of c-MYC in addictedtumors or treatment with cyclin-dependent kinase(CDK) inhibitors have proven effective by invoking asenescence response. The possibility of using prose-nescence therapies for cancer treatment has provokedconsiderable interest. We propose that the senescencesecretome can be a source of novel targets for prose-nescence therapies, as it has tumor suppressiveactions. Overall, tailored prosenescence therapies havethe potential to be used for treating cancer and otherpathologies.

Replicative senescence: an arresting cell stateSenescence is a stable cell cycle arrest induced at the end ofthe cellular lifespan or in response to different stresses.What we refer to today as replicative senescence was firstdescribed by Hayflick and Moorhead [1], who challengedthe existing dogma that normal cells were capable ofunlimited proliferation in culture [2]. Although senescentcells remain arrested even when stimulated by growthfactors, these cells are metabolically active. Senescent cellsalso display senescence-associated b-galactosidase (SA-b-Gal) activity, as a consequence of an increase in lysosomenumber [3], and undergo chromatin remodeling giving riseto the so-called senescence-associated heterochromatin foci(SAHF) [4]. In addition, senescent cells secrete a complexmixture of extracellular matrix and soluble factors, re-ferred to as the senescence-associated secretory phenotype(SASP) or senescence messaging secretome (SMS) [5,6].Recent studies suggest that manifestations of the senes-cent phenotype influence the stable cell cycle arrest char-acteristic of senescent cells [5].

In human cells, replicative senescence is triggered by acombination of two main factors. The first is the activationof a DNA damage response mainly triggered by telomereshortening and uncapping [7]. The second is the derepres-sion of the INK4/ARF (inhibitor of CDK4A/alternativereading frame) locus, which behaves as a sensor for linkingstress detection with activation of key tumor suppressornetworks [8]. The relative contribution of these pathways

Corresponding author: Gil, J. ([email protected]).Keywords: senescence; cancer; therapy; tumor suppressors; secretome; p53; Myc; Ras.

0962-8924/$ – see front matter � 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.tcb.2011.1

to other types of senescence and how they are engaged arevery much dependent on cell type, organism of origin andthe specific stimuli activating senescence.

Oncogene-induced senescence (OIS) as a barrier fortumor progressionExpression of oncogenic RasG12V in normal cells induces aphenotype almost indistinguishable from replicative se-nescence, termed OIS (Figure 1). Besides OIS, agentscausing DNA damage, oxidative stress, chemotherapeuticdrugs or even the process of reprogramming to inducedpluripotent stem cells can also trigger premature senes-cence or stress-induced senescence [9]. In 2005, severalgroups independently identified the presence of cellsundergoing OIS in premalignant mouse and humanlesions and their conspicuous absence in more advancedtumors [10–14]. Prototypic examples of such lesions arenevi, skin lesions of a benign nature that are precursors tomelanomas and that can persist in the skin for years.Human nevi stain positive for SA-b-Gal activity andexpress high levels of p16INK4a, although in a mosaicpattern. Almost all human nevi bear an activated BRAFor NRAS oncogene, but only after they acquire additionalmutations that cancel OIS can they progress to melanoma[12,15].

Senescent cells are present in a wide range of premalig-nant lesions. Mouse models have shown that oncogenic K-RasG12V expression associates with senescence in earlystages of lung and pancreatic tumors [14]. In mice harbor-ing an Em-N-Ras transgene [13], most of the animalsdevelop a nonlymphoid neoplasia with prevalent signs ofsenescence. Conversely, the expression of Em-N-Ras inmouse knockouts for p53 or Suv39h1 causes aggressiveT cell lymphomas [13]. Skin papillomas induced by treat-ment with DMBA and TPA bear activating point muta-tions in H-Ras [16] and are also enriched in senescent cells[14], in a way that is dependent upon p38 activation byPRAK [17]. Moreover, in a serrate colon cancer modeldriven by oncogenic K-Ras, hyperplasias with character-istics of senescence, such as elevated p16Ink4a expressionand SA-b-Gal activity, are observed [18,19]. Nevertheless,the ability of oncogenic Ras to induce senescence dependson the genetic context and relative expression. Lowerlevels of Ras in the mammary gland stimulate proliferationand mammary epithelial hyperplasias whereas higherlevels of Ras lead to senescence and Ink4a/Arf upregula-tion [20]. More strikingly, a recent report suggests thatactivation of K-RasG12D in the pancreas does not triggersenescence but in fact suppresses senescence induced by

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Therapy-induced senescence (TIS)

Normal cells

Oncogenic alterations

Aberrant proliferation

Oncogene-induced senescence (OIS)

Fail-safe mechanisms

Apop tosi s

Additional alterations

Cancer cells

Conventional therapies Prosenescence therapies

Apop tosi s

TRENDS in Cell Biology

Figure 1. Role of senescence in cancer progression and therapy. Normal cells accumulate oncogenic alterations that trigger an initial phase of aberrant cell proliferation

giving rise to preneoplastic lesions. Parallel to this aberrant proliferation, cell-intrinsic fail-safe mechanisms such as oncogene-induced senescence (OIS) and apoptosis are

activated. During cancer progression, additional mutations are acquired to override these protective mechanisms, giving rise to full-blown malignancies. Conventional

treatments such as chemo- or radiotherapies act by inducing cell death or senescence (termed therapy-induced senescence, TIS). Currently, prosenescence therapies are

being explored as an alternative or complement to cancer treatment. Normal cells, gray; preneoplastic cells, yellow; senescent cells, blue; apoptotic cells, purple; cancer

cells, red.

Review Trends in Cell Biology April 2012, Vol. 22, No. 4

inflammation through a mechanism involving Twist-me-diated inactivation of p16Ink4a [21]. Other work has sug-gested that K-RasG12D can only trigger senescence when adetoxifying Nrf2-dependent program commonly activatedby oncogenes is disabled [22].

Premalignant lesions caused by oncogenes other thanRas are also enriched in senescent cells. Tissue-specificexpression of an oncogenic BRAFV600E mutant results inlung adenomas or melanocytic nevi with increased num-bers of senescent cells [23–25]. Deregulated expression ofE2F3 in the mouse pituitary gland induces hyperplasiaswith senescent characteristics [10], whereas c-Myc, usuallylinked to apoptosis, triggers senescence in murine lympho-mas through macrophage-mediated TGF-b secretion [26].Recent work taking advantage of Bmi�/� mice hasrevealed that several leukemic fusion proteins also inducesenescence [27].

Inactivation of tumor suppressors also drives senes-cence in premalignant lesions. The prototypic example isthe inactivation of PTEN in the mouse prostate, whichinduces a phenotype resembling prostate intraepithelialneoplasia (PIN) [11]. PIN lesions have characteristics ofsenescence, such as positive staining for SA-b-Gal activity

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and activation of the Arf–p53–p21 axis in mouse cells.Importantly, features of senescence are similarly observedin human PIN [11,28]. As some of the characteristics ofsenescence induced by Pten loss differ from those of senes-cence driven by RAS/RAF, the Pandolfi group has renamedthis type of senescence Pten loss-induced senescence(PICS) [29]. Specifically, PICS can occur independentlyof DNA replication and in the absence of DNA damage[29]. Loss of other tumor suppressors, such as NF1 inneurofibromas [30], the von Hippel–Lindau tumor suppres-sor gene (VHL) in kidney carcinomas [31] or Rb in thyroidcancers [32], also result in premalignant lesions withmarkers of senescence.

Overall, there is very convincing evidence that earlyoncogenic activation gives rise to premalignant lesionsbut can, in parallel, activate senescence. In addition, dis-abling this senescence response is needed for tumor progres-sion. For example, loss of Ink4a/Arf on the serrate colonmodel induces the appearance of adenocarcinomas [18].Similarly, nevi lesions induced by oncogenic BRAF expres-sion take months or even years to spontaneously evolve tomelanomas, but upon inactivation of Cdkn2a or p53, thisprogression is markedly accelerated [24,25]. p53 loss in


Pten�/� PIN lesions also accelerates the appearance ofadvanced metastatic prostate cancer [11]. These examplessuggest that if mutations that disable senescence occur, OIScan be reversed, giving rise to tumors. Although definitiveevidence for the reversibility of senescence in vivo is lacking,acute ablation of Rb or p53 in vitro results in senescencereversal [33,34]. In summary, senescence is not only trig-gered in preneoplastic lesions but also acts as an effectivebarrier that needs to be disabled during tumor progression(Figure 1).

Activation of senescence during conventional cancertherapiesTargeted cancer therapies have been developed in recentdecades and proven effective against specific tumors. Thefirst example was Imatinib, an inhibitor of the BCR–ABLfusion used to treat chronic myeloid leukemia (CML). Sincethen, many other targeted drugs have proven effective,such as Her2 inhibitors in a subset of breast or lung cancers[35], or BRAF inhibitors for the treatment of melanoma[36]. However, even with this collection of targeted drugs,the most widely used treatments for cancer are still chemo-and radiotherapies. The rationale behind their effective-ness is that they cause extensive DNA damage in rapidlydividing cells and thus predominantly target cancer cells.This DNA damage response can induce gross aneuploidies,mitotic catastrophe and apoptosis. Interestingly, senes-cence has also been identified in cancer cells treated withchemotherapeutic drugs or subjected to ionizing radiation(Figure 1) [37,38].

Telomerase inhibition

Terc–/– GRN163L

Cell

Cdk 2–/– CVT-313 Cdk

Myc

PICS

Pten VO-OHpic

Pten+/–

Myc addiction

Myc JQ

(a) (b)

(c) (d)

BET

Figure 2. Prosenescence therapies for cancer treatment. The different types of prosenes

in the therapy are listed at the left of the arrows, and examples of small molecule inh

Targeting cell cycle control. (c) Pten loss-induced senescence (PICS) in Pten+/� tumors

proteins by JQ1 is used as an indirect way to inhibit Myc activity. (e) Reactivation of p

immune system, green and white.

Using the Em-myc model of lymphomagenesis, work hasstarted on the senescent program that contributes to theoutcome of cancer therapy [39]. This senescent response,termed therapy-induced senescence (TIS), relies on en-gagement of the p16INK4a and p53 tumor suppressor net-works. Genetic models in which either of these networks isinactivated show resistance to cancer treatment, directlylinking the ability to mount a robust senescence responsewith the outcome of chemotherapy. It is possible that keyeffectors of TIS are frequently mutated in advanced can-cers. Therefore, resistance to conventional therapies willoften be acquired from the very same mutations drivingtumor progression.

Inducing senescence as an alternative therapy forcancer treatmentOIS has a crucial role in preventing cancer progression, asthere is an active need to bypass OIS for tumors to evolvefrom indolent stages to malignant phases. This, togetherwith the relevance of TIS in the outcome of conventionalcancer therapies, suggests that prosenescence therapiescould be effective in cancer treatment. Therefore, drugsaimed at selectively inducing cellular senescence couldrepresent a promising novel approach for cancer interven-tion. Supporting this, transgenic mice expressing extracopies of key senescent effectors such as p53 or Ink4/Arfhave exhibited extended cancer protection, without un-wanted side effects [40]. In addition, several exampleshave emerged recently of drugs exploiting induction orreinforcement of senescence for cancer treatment. We

cycle control

4–/– PD0332991 Skp2–/– MLN4924

Ras Pten–/–

Myc

1

p53 reactivation

Non-functional p53

p53 Nutlin or PRIMA-1

Senescent secretome

Immune clearance

(e)

Immune system


cence therapies discussed in this review are summarized here. The genes targeted

ibitors used are listed to the right of the arrows. (a) Inhibition of telomerase. (b)

. (d) Targeting Myc addiction. Inhibition of bromodomain and extraterminal (BET)

53. Cancer cells, red; senescent cells, blue; apoptotic cells, purple; cells from the

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review those approaches and their underlying rationale inthis section (summarized in Figure 2).

Inhibiting telomerase activity

One of the hallmarks of tumors is their ability to proliferatebeyond the normal replicative lifespan [41,42]. A key factorinfluencing this limit is telomeric length. Eroded or unpro-tected telomeres resulting from the progressive shorteningcaused by cellular division can cause chromosomal fusionsand activate a DNA damage signaling cascade [7]. Tobypass this limitation, most tumors acquire telomeraseactivity (around 90%). Consistent with these observations,telomerase-deficient mice engineered by deletion of Tercshow reduced cancer susceptibility [43], with the onlynotable exception being p53�/�/Terc�/� mice, in whichcancer incidence increases [44]. Therefore, the inhibition oftelomerase activity has been proposed as a valid anticancerapproach. Different strategies have been pursued to inhibittelomerase activity in tumors [45], with small moleculeenzyme inhibitors (such as GRN163L) being the mostpromising so far. However, given that inhibition of telome-rase can also cause gross aneuploidies or apoptosis, therelative contribution of senescence induction when inhibit-ing telomerase in tumors is unclear.

Modulation of CDK activities

Despite the deregulated cell cycle of cancer cells, drugsinhibiting CDKs have had limited success as cancer treat-ments in general clinical trials [46,47]. However, recentstudies suggest that targeting specific cell cycle-dependentkinases or CDK inhibitors (CDKI) in the appropriate ge-netic context can result in synthetic lethal interactionspromoting a tumor-specific prosenescence response withtherapeutic benefit [48–50].

One of those studies showed that inhibiting CDK2 inc-Myc-driven tumorigenesis induces senescence. Two dif-ferent small molecule inhibitors that inhibit Cdk2 (CVT-313 and CVT-2584) caused Myc-dependent senescence inMEFs, IMR-90 primary human fibroblasts and even inU937 cancer cells [48]. Similarly, targeting Cdk4 unveileda synthetic lethal interaction with oncogenic K-RasG12V

that has therapeutic potential in lung adenocarcinomas[50]. Genetic inactivation of Cdk4 induces senescence of K-RasG12V-driven lung adenomas and results in tumor re-gression and senescence in more advanced adenocarcino-mas. Further proving the therapeutic potential of thisinteraction, treatment with a selective Cdk4 inhibitor(PD0332991) partially prevented or slowed the appearanceof non-small cell lung tumors driven by oncogenic K-RasG12V [50].

Targeting cell cycle control in the context of Pten lossalso induces a prosenescence response with therapeuticsignificance. Genetic inhibition of Skp2 synergizes withPten loss in mouse lymphoma, sarcoma, prostate andadrenal cancer to induce senescence and suppress tumori-genesis [49]. Skp2 regulates expression of the CDKI p27,which is a key target downstream of Skp2 in this syntheticlethal interaction. Experiments in mouse models sug-gested that targeting Skp2 could trigger senescence intumors driven by Pten inactivation [49]. To target Skp2activity, the authors used MLN4924, an inhibitor of

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neddylation. Cullin, a protein associated with Skp2, isneddylated and treatment with MLN4924 results inSkp2 inhibition, preventing the formation of tumors in aPC3 human prostate cancer cell xenograft model by induc-ing senescence that was independent of p53.

Activation of a PICS response

Another strategy to mount a senescent response in Ptenheterozygous tumors was proposed recently [29]. It isbased in inhibiting Pten activity in Pten+/� tumors. Al-though inhibiting a tumor suppressor such as Pten iscounterintuitive and could potentially be dangerous,proof-of-principle experiments indicate that it can be ther-apeutically useful. Previous work demonstrated the con-trasting effect of deleting one or both alleles of Pten.Treatment with VO-OHpic, a Pten inhibitor, induces se-nescence specifically and differentially in Pten +/� tumorswith no deleterious effect on surrounding Pten +/+ cells.This senescence response is referred as Pten loss-inducedsenescence (PICS). As Pten overexpression or PI3K inhi-bition can also cause senescence [30], it seems that thePten/PI3K pathway must be very finely tuned and smallalterations in either direction could have unwanted con-sequences. Whether this could be a limitation hamperingthe therapeutic use of PICS remains to be established, butthis highlights how the effectiveness of prosenescencetherapies is highly dependent on genetic context.

Exploiting tumor addiction to c-Myc

Prototypic oncogenes such as RAS or MYC have profoundeffects on cell cycle control, cell growth and cell metabolism– all the criteria necessary to cause a strong, exploitableaddiction in tumors [51,52]. Murine models have shownthat tumors can become addicted to c-Myc and suggestedc-Myc inhibition as a valid target in cancer therapy. Takingadvantage of tet-dependent c-Myc expression, it wasreported that switching off c-Myc caused senescence inlymphomas, osteosarcomas or hepatocellular carcinomas(HCCs) [53]. The addiction of tumors to c-Myc expressionwas also shown by taking advantage of a Myc mutantknown as OmoMyc [54]. OmoMyc can homodimerize orheterodimerize with wild-type Myc or Max to form com-plexes with low DNA binding efficiency. Therefore,OmoMyc behaves as a dominant-negative mutant by se-questering and inhibiting c-Myc. Although in early stageK-Ras-induced lung adenomas the tumor suppressive re-sponse observed after c-Myc inactivation is mediated byapoptosis induction, in advanced lung adenocarcinomasOmoMyc caused decreased proliferation accompanied bysenescence [55]. In addition, inactivation of c-Myc in lym-phomas caused senescence and tumor regression in amanner dependent upon CD4+ T lymphocytes. This sug-gests that beyond cell-intrinsic effects, the tumor microen-vironment, and in particular the immune system, canmediate prosenescent tumor suppression [56].

Despite the validity of exploiting tumor addiction to c-Myc for therapy, an important obstacle remains in identi-fying effective inhibitors for clinical use. In this regard,three recent studies identified inhibition of bromodomainand extraterminal (BET) proteins by small molecules suchas JQ1 or I-BET151 as a way to affect c-Myc function

Box 1. The senescence-associated secretory phenotype (SASP)

Senescent cells secrete a complex mixture of secreted and extracellular

factors usually referred to as the SASP. It has been also called

senescence messaging secretome (SMS). The production by senescent

cells of secreted factors was noted more than 20 years ago with the

identification of individual factors promoting changes in the extra-

cellular matrix and influencing the cellular microenvironment, such as

a-(I) procollagen, fibronectin, collagenase, stromelysin and gelatinase-

TIMP-2 [74–78]. Gene expression profiles confirmed that senescent

cells secrete factors related to wound healing, inflammatory response

or cytokine and chemokine signaling [14,79–81]. This upregulation of

secreted proteins is common to cells undergoing replicative senes-

cence or OIS and observed in cells of epithelial, fibroblast or endothelial

origin [80]. The SASP is present not only in human cells but also in

mouse cells undergoing senescence when cultured in 3% oxygen [6].

Antibody arrays have been used for characterizing the senescent

secretome [28,82], confirming its complex nature. The senescence

secretome includes extracellular proteases (urokinase-type plasmino-

gen activator, uPA), tPA, matrix components (MMP), growth factors,

proinflammatory cytokines (IL-6, IL-1a, IL-1b) or chemokines (IL-8,

GROa, MCP-1). Transcription factors such as CEBPb and NF-kB act as

master regulators of the senescence secretome [28,81,83,84].

The effects exerted by the senescence secretome on the micro-

environment and surrounding cells are diverse and can often be

opposing. Senescent fibroblasts can promote tumorigenesis of

transformed epithelial cells through a combination of cell–cell

interaction and soluble factors [85]. Senescent fibroblasts can also

contribute to the protumorigenic behavior of neighboring epithelial

cancer cells and can, for example, affect the differentiation of

epithelial cells [86], promote angiogenesis through VEGF produc-

tion [87], induce epithelial-to-mesenchymal transition or promote

migration of epithelial cancerous cells [82]. However, the senes-

cence secretome can have tumor suppressive effects. Some factors

of the SASP, such as PAI-1, IGFBP7, IL-6 or IL-8, have a role in

establishing or reinforcing the cell cycle arrest characteristic of

senescence [5]. These factors engage the senescence machinery by

several different mechanisms: IL-6 induces p15INK4b [81]; IL-8

increases ROS production and causes DNA damage [28]; PAI-1

increases GSK3b activity and disrupts the nuclear localization of

cyclin D1 [88]. In addition, factors of the senescence secretome can

signal to the immune system to clear senescent cells [62], therefore

also contributing to tumor suppression in a non-cell-autonomous

way.


[57,58]. This indirect inhibition of c-Myc is a promisingtherapeutic strategy for acute myeloid leukemia (AML)[57], mixed-lineage leukemia (MLL) [59] and multiplemyeloma (MM) [58], as shown using murine models andprimary patient samples. In particular, in MM, treatmentwith JQ1 results in cell cycle arrest and senescence induc-tion, showing that disrupting addiction to c-Myc can en-gage a senescent response [58].

Reactivation of the tumor suppressive function of p53

As tumors inactivate the p16/Rb and p53 networks tobypass OIS and thrive, the reactivation of these tumorsuppressors could conversely restore senescence and halttumor progression. Although ineffective in tumors sustain-ing p53 deletions, drugs that enhance p53 function throughMDM2 inhibition (such as nutlin) or that can restore thewild-type activity of p53 mutants (such as PRIMA-1) arecurrently being tested [60]. The validity of this strategy hasbeen shown in mouse models in which p53 expression canbe reactivated. Using a Cre-loxP-based transgene, restora-tion of p53 activity in tumors was shown to cause theirregression. Lymphoma regression is caused by massiveapoptosis induction, whereas in sarcomas cell cycle arrestwith features of senescence is observed [61]. Using a tet-inducible shRNA to conditionally regulate endogenous p53levels in a mosaic mouse model of HCC driven by H-RasG12V expression, even a brief reactivation of p53 wasshown to result in tumor regression, accompanied by se-nescence but negligent apoptosis [62]. Interestingly, theactivation of senescence in cancer cells was associated witha strong SASP. Secretion of proinflammatory cytokines bysenescent cells initiated an innate immune response re-sponsible for tumor clearance [62].

The senescence secretome as a novel target forprosenescence therapiesSenescent cells secrete a complex mixture of extracellularproteins and soluble factors that are referred to asthe SMS, for their ability to signal and influence theirsurrounding environment, or the SASP (Box 1). The

senescence secretome exerts diverse and opposite effectsover the microenvironment and on neighboring cells [6].Although initial interest in studying the SASP was focusedon its protumorigenic potential, such as promoting growth,migration or angiogenesis, more recently, its tumor sup-pressive properties have also been highlighted. Compo-nents of the senescence secretome reinforce or implementstable cell cycle arrest and contribute to tumor suppressionby signaling to and recruiting the immune system [5](Figure 3).

Secreted factors and their receptors are prototypic drug-gable molecules. Indeed, most biotherapeutics used in theclinic are soluble factors or antibodies targeting such factorsor their receptors. Therefore, it is feasible to manipulatecomponents of the senescence secretome for prosenescencetherapies and many of the necessary reagents have alreadybeen developed. Members of the senescence secretome con-tributing to senescence arrest include PAI1, IL-8, IL-6,IGFBP7 and TGF-b [5]. However, some of these factorsdisplay opposing effects; they can be tumor suppressive orprotumorigenic depending on the context. This is the case forproinflammatory cytokines such as IL-6, IL-8 and GROa,which display multiple protumorigenic activities. As a re-sult, IL-6 and IL-8 are both required mediators of Ras-driven tumorigenesis [63,64]. What switches the senescencesecretome from tumor suppressive to protumorigenic is notwell understood, although it is thought that the geneticcontext of the target cells (e.g. whether they have intactRb and p53 pathways) is a factor.

Despite increased knowledge on SASP composition andthe existence of recombinant proteins, small molecules andantibodies to target these factors, there are no currentexamples of prosenescence cancer therapies that exploitcomponents of the senescence secretome, at either theexperimental or the preclinical stage. However, precedentssuggest that such an approach could succeed. Some of theprosenescence therapies described previously, such as re-activation of p53 in Ras-driven HCC [62] or c-Myc lympho-mas treated with chemotherapeutic drugs [26], rely onsecreted factors to drive senescence and activate innate

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Reinforce growth arrest

Prosenescence therapies

Senescent secretome

Immune clearance

Cancer cells

Cancer cells

Immune system

Cell growthCell differentiation

AngiogenesisEMT

Migration

Secretom e manipulation


Figure 3. The senescence secretome in therapy: roles, possibilities and problems. Therapies that result in senescence induction (prosenescence therapies) stop tumor

progression through cell-intrinsic mechanisms and also induce the production of a senescence secretome. This senescence secretome has tumor suppressive effects

(shown in red), such as its contribution to reinforce growth arrest and signaling to the immune system for clearance, but can also exert protumorigenic actions in cancer

cells (shown in green). By manipulating the secretome, targeting individual components or using it in tumors or preneoplastic lesions of specific genetic composition,

we aim to enhance their tumor suppressive effects, minimizing its protumorigenic actions. Cancer cells, red; senescent cells, blue; cells from the immune system, green

and white.


immunity. In addition, factors secreted by senescent cellsare able to not only invoke innate immunity to clear tumorsduring prosenescence therapies [62] but also initiate animmune surveillance of premalignant senescent cells thatrelies on an adaptive immune response [65].

The best example of components of the senescencesecretome used in a prosenescence therapy comes notfrom a model of cancer but from studying the fibroticresponse associated with wound healing [66]. The extra-cellular matrix protein CCN1 (also known as CYR61)restricts fibrosis induction during wound healing.CCN1 is secreted by senescent cells and binds to integrina6b1 in target cells. This binding activates the RAC1–

NOX1 pathway and results in the production of reactiveoxygen species (ROS) and senescence induction [66]. Theauthors observed that senescent fibroblasts preferential-ly accumulate in granulation tissues of healing woundsand express antifibrotic genes. However, mice expressinga CCN1 mutant that cannot bind to integrin a6b1 exhibitexacerbated fibrosis and do not accumulate senescentfibroblasts. Interestingly, topical application of recombi-nant CCN1 on cutaneous wounds reduced fibrosis, in-creased senescence and improved healing [66]. Theseresults suggest that components of the senescence secre-tome could be successfully used for prosenescence thera-pies (Figure 3).

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Concluding remarksEver since the identification of senescence as a stablegrowth arrest with potential implications in aging, itsphysiological relevance has expanded as we realize thatstresses such as oncogenes, ionizing radiation or exposureto chemotherapeutic drugs also activate senescence. Theroles of senescence in suppressing progression of prema-lignant lesions and as a determinant of the outcome ofcancer therapies have promoted the idea that enhancingsenescence (prosenescence therapies) could be an alterna-tive or a complement to conventional anticancer treat-ments. As such, proof-of-principle studies using mousemodels have shown the therapeutic potential of engagingsenescence. In parallel, pioneering studies such as thosesummarized in this review have shown the feasibility ofprosenescence therapies. Indeed, several compounds thatcould be used in prosenescence therapies (summarized inTable 1) are currently undergoing clinical trials.

In recent years, the list of pathologies to which senes-cence is associated has increased well beyond cancer.Senescence limits fibrotic responses during progressionto cirrhosis or in cutaneous wound healing [66,67]. In-creased incidence of senescence has been shown in diabe-tes, atherosclerotic plaques and multiple age-relateddiseases. Moreover, recent genome-wide association stud-ies (GWAS) have shown that multiple single-nucleotide

Table 1. Molecules with possible applications in prosenescence therapies

Strategy Drug Efficacy Limitations References

Inhibition of

telomerase activity

e.g. GRN163L General, most tumors

reactivate telomerase

Possible side effects in normal tissue

homeostasis. Inhibition of telomerase

increases cancer incidence in p53-null

background

[45]

Inhibition of CDK2 CVT-313, CVT-2584 MYC-driven tumors Limited success in trials with

CDK inhibitors so far

[48]

Inhibition of CDK4 PD0332991 K-RasG12V-driven tumors Limited success in trials with

CDK inhibitors so far

[50]

Inhibition of Skp2/

neddylation

MLN4924 PTEN-null tumors. Works

independently of p53 status

Unknown side effects of global inhibition

of neddylation

[49]

PTEN inhibition VO-OHpic PTEN+/� tumors Balance of PTEN activity is delicate.

Potentially dangerous to inhibit a

tumor suppressor such as PTEN

[29]

Inhibition of BET family

proteins

JQ1, I-BET151 Tumors ‘addicted’ to MYC.

Potentially useful in AML,

MM and MLL

Identification of adequate tumors

to be targeted

[57–59]

MDM2 inhibition e.g. Nutlin, RITA Tumors with wild-type p53 Seems to have a good therapeutic window,

but could select for p53 mutations

[60]

Mutant p53 reactivation e.g. PRIMA-1, MIRA-1 Tumors with mutant p53 No clear mechanism of action or not known

if it will be effective in different mutants

[60]

Treatment with CCN1 Recombinant CCN1 Tested in restricting fibrosis

in wound healing

Efficacy as cancer treatment unknown [66]


polymorphisms (SNPs) at non-coding regions of the INK4/ARF locus, which encodes for key senescence effectors, areassociated with Alzheimer’s disease or susceptibility toatherosclerotic vascular disease (ASVD) [68,69]. A mousemodel in which the corresponding syngenic region wasdeleted showed decreased expression of the Ink4/Arf locusand suggested that ASVD is associated with diminishedsenescence [70]. Although current evidence falls short ofsuggesting that re-establishment of senescence could bebeneficial for ASVD, perhaps prosenescence therapiescould also be useful in treating pathologies other thancancer. However, as a note of caution we should mentionthat increased senescence in vascular cells has been linkedto atherosclerosis [71].

We envision that novel prosenescence therapies will bedeveloped in different ways: through improved knowledgeof the molecular pathways controlling senescence; by spe-cifically exploiting cell culture systems to screen for senes-cence regulators; or through the discovery of drugseffective for cancer treatment without a priori knowledgethat they will have prosenescence effects. In particular,given the current knowledge on senescence, we suggestthat components of the senescence secretome and drugstargeting the epigenetic control of senescence should beexplored to find additional prosenescence therapies.

Exploiting senescence as an anticancer therapy can beless intuitive than inducing apoptosis, as one results in cellcycle arrest whereas the other effectively eliminates theaffected cells. Although there are clear examples (such asnevi) in which senescent cells remain ‘halted’ for years anddecades in vivo without active clearance, it is becomingevident that, in other cases, senescent cells signal throughtheir secretome to be actively eliminated by the immunesystem. Therefore, a possible advantage of prosenescencetherapies is that they could work twofold, by intrinsicallystopping cell proliferation and eventually by recruiting theimmune system to cause clearance of premalignant cells orregression of tumors (Figure 3). A recent study takes

advantage of an ingenious mouse model to prove thateliminating senescent cells can improve age-related phe-notypes [72]. Therefore, an alternative to prosenescencetherapies could be to enhance the elimination of cellsundergoing OIS. However, we first need to determinewhether ablating senescent cells could be beneficial fortumor suppression and how this could be achieved.

A word of caution must be added as senescence regu-lators can often behave as double-edged swords, havingopposite effects on cell proliferation and tumor growthdepending on genetic context. Factors of the senescencesecretome such as IL-6 and IL-8 can be either tumorsuppressive or protumorigenic, slight changes in Ptenactivity can make a difference between tumor growth orsenescence induction, and oncogenic Ras itself eitherbehaves as a potent oncogene or halts proliferationdepending on expression levels, genetic context and otherfactors. Therefore, prosenescence therapy must be appliedvery specifically, and will require stratification of patientsby tumor type and genotyping. An additional challengeremains in monitoring the engagement of senescence aftertreatment. Although efforts are being taken to establish asenescence index, similar to the apoptotic or proliferationindexes [73], there is a need to identify more reliableand robust markers of senescence and, ideally, methodsto detect senescence compatible with advanced in vivoimaging.

Overall, we are just starting to see how prosenescencetherapies are joining the arsenal of advanced weaponryavailable for the fight against cancer, and we anticipatethat, spearheaded by improved knowledge on the molecu-lar basis of senescence and the ability to monitor it, novelstrategies relying on senescence induction will reach theclinic as potential cancer therapies in the coming years.

AcknowledgmentsCore support from the Medical Research Council (MRC) and grants fromMRC Technology, Cancer Research UK and the Association for

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International Cancer Research fund the research in J. Gil’s laboratory. J.Gil is also supported by the EMBO Young Investigator Programme.

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Senescence: a new weapon for cancer therapy