Cellular senescence in age-related disorders

Abstract

Much of the population is now faced with an enormous burden of age-associated chronic diseases. Recent discoveries in geroscience indicate that healthspan in model organisms such as mice can be manipulated by targeting cellular senescence, a hallmark mechanism of aging, defined as an irreversible proliferative arrest that occurs when cells experience oncogenic or other diverse forms of damage. Senescent cells and their pro-inflammatory secretome have emerged as contributors to age-related tissue dysfunction and morbidity. Cellular senescence has causal roles in mediating osteoporosis, frailty, cardiovascular diseases, osteoarthritis, pulmonary fibrosis, renal diseases, neurodegenerative diseases, hepatic steatosis, and metabolic dysfunction. Therapeutically targeting senescent cells in mice can prevent, delay, or alleviate each of these conditions. Therefore, senotherapeutic approaches, including senolytics and senomorphics, that either selectively eliminate senescent cells or interfere with their ability to promote tissue dysfunction, are gaining momentum as potential realistic strategies to abrogate human senescence to thereby compress morbidity and extend healthspan.

Introduction

Over the past 200 years, the human lifespan in most developed countries has more than doubled, and this trend is projected to continue in the coming decades with further improvements in living conditions and medical care. However, this extension of lifespan is paralleled by increased burden of chronic diseases such as osteoporosis, cardiovascular diseases, frailty/sarcopenia, osteoarthritis, pulmonary fibrosis, renal diseases, neurodegenerative diseases, hepatic steatosis, and metabolic dysfunction, in much of the aging population, thereby reducing the healthspan, i.e., disease-free period of life.⁽¹⁾ These age-related chronic diseases not only account for an enormous socioeconomic burden but are also leading causes of poor quality of life and increased mortality.⁽²⁾ Nevertheless, significant advances in our understanding of the basic fundamental mechanisms that drive aging⁽³⁾ have raised the possibility of implementing biomedical interventions to delay or reverse late-life chronic diseases and co-morbidities of aging as a group, as opposed to the current approach of treating each condition separately.⁽⁴⁾

One such fundamental aging mechanism that has increasingly received much attention over the past decade or so is cellular senescence, which can be defined as the essentially irreversible proliferative arrest that occurs when cells experience oncogenic or other diverse forms of damage.⁽⁵⁾ Examples of cell-damaging insults include DNA lesions (telomere attrition, mutations, radiation), reactive metabolites (reactive oxygen species [ROS], ceramides, high glucose, fatty acids), proteotoxic stress (unfolded protein responses, protein aggregation, upregulated mTOR signaling), damage-associated molecular pattern proteins (DAMPs), and systemic low-grade inflammation.⁽⁶⁾ Cellular senescence favors fitness and reproductive output by preventing malignant cell transformation.⁽⁷⁾ Growth arrest upon activation of the cellular senescence program is achieved by activation of cyclin-dependent kinase inhibitors, including p16^Ink4a and p21^Cip1 (upstream of p53), that trigger cell-cycle arrest to halt cancer cell proliferation.⁽⁸⁾ However, senescent cells accumulate in various tissues throughout the body with natural aging or in accelerated aging conditions and are potential mediators in age-related comorbidities including osteoporosis, frailty, cardiovascular disease, osteoarthritis, pulmonary fibrosis, renal disease, neurodegenerative diseases, hepatic steatosis, and metabolic dysfunction (Figure 1). The overall purpose of this review is to examine the causal roles of cellular senescence in diseases of aging and to assess some of the progress made thus far towards senotherapeutic approaches to extend healthy aging and compress morbidity in preclinical studies.

Figure 1.

Cellular senescence is an emerging risk factor for most serious chronic diseases of natural aging and conditions of accelerated aging. Senescent cells and their senescence-associated secretory phenotype (SASP) have causal roles in mediating osteoporosis, frailty, cardiovascular disease, osteoarthritis, pulmonary fibrosis, renal disease, neurodegenerative diseases, hepatic steatosis, and metabolic dysfunction, among others. There is also evidence suggesting that at least some these diseases feedback to contribute to increased senescent cell accumulation. See text for extended discussion and references.

The age-associated accumulation of senescent cells is likely due to a combination of factors that include inefficient senescent cell removal by the immune system in old age⁽⁹⁾ and the resistance of senescent cells to apoptosis.⁽¹⁰⁾ Once a cell enters a state of growth arrest or senescence, it can display several hallmark cellular and molecular features that when detected (in combinations) can be used as biomarkers of cellular senescence.⁽¹¹⁾ These hallmark features of senescent cells (see⁽¹¹⁾ for a more detailed review) include, but are not limited to, the following: i) sites of DNA damage within telomeres, termed telomere-associated foci (TAFs);⁽¹²⁾ ii) unraveled satellite heterochromatin DNA, termed senescence-associated distension of satellites (SADS);^(13,14) iii) cytoplasmic chromatin fragments (CCFs);⁽¹⁵⁾ iv) higher CENP-A;^(15,16) v) increased lipofuscin (GL13);⁽¹⁷⁾ vi) reduced lamin B1;⁽¹⁸⁾ and vii) loss of nuclear-mobility group box 1 (HMGB1).⁽¹⁹⁾

Senescent cells can also develop a complex, altered gene expression profile that in addition to higher expression of p16^Ink4a and/or p21^Cip1, is characterized by upregulation of senescent cell anti-apoptotic pathways (SCAPs)⁽⁶⁾ as well as typically the acquisition of a distinctive pro-inflammatory secretome, termed the senescence-associated secretory phenotype (SASP).⁽²⁰⁾ Identification of the first SCAPs was achieved by comparing the expression profile of human senescent versus non-senescent cells followed by confirmation experiments using RNA interference.⁽²¹⁾ This seminal study unveiled multiple SCAP networks, including BCL-2/BCL-X_L, p53/p21/serpines, PI3K/AKT, dependence receptors/tyrosine kinases, and HIF1-α, that were specifically upregulated in human senescent cells, thus paving the way for the identification of first-generation drugs that selectively eliminate senescent cells.⁽²¹⁾ Senescent cells cause degeneration as well as aberrant remodeling and disruption of the normal function of neighboring tissues via secretion of their SASP, which can consist of pro-inflammatory cytokines, chemokines, interleukins, growth factors/regulators, and matrix-degrading proteases.⁽²²⁾ Consistent with these direct detrimental effects of the SASP on neighboring, non-senescent cells, Nelson et al.⁽²³⁾ showed that young (control) fibroblasts co-cultured with senescent fibroblasts have decreased proliferative capacity and diplay increased senescence markers (e.g., SA-β-Gal staining), even days after removal of the senescent cells from culture. These co-culture experiments demonstrate that the SASP of senescent cells can spread the senescence phenotype to neighboring, non-senescent cells and tissues via this phenomenon, which is referred to as the senescence-induced bystander effect.^(23,24) Thus, over prolonged periods the SASP is considered a mechanistic driver by which senescent cells promote aging, tissue dysfunction, and disease.

The biological relevance and causal roles of senescent cells are becoming more evident with the development of transgenic mice that permit the selective elimination or clearance of senescent cells (termed “senolysis”). Such models have established the physiological consequences of senescent cell removal in progeroid and naturally aged mice.^(25,26) It is noteworthy that only a small or low abundance of senescent cells is sufficient to promote tissue dysfunction and multiple aspects of aging. For example, eliminating or clearing even a relatively small proportion of senescent cells (e.g., ~30% ⁽²⁵⁾) using a systemically expressed senescence-associated “suicide” transgene, INK-ATTAC (p16^Ink4a-linked apoptosis through targeted activation of caspase ⁽²⁵⁾), that allows the inducible elimination of p16^Ink4a-expressing senescent cells via administrating a synthetic drug (AP20187), extends healthspan and prevents or alleviates several age-associated co-morbidities in both progeroid and natural chronologically aged mice.^(25–29) Consistent with this, the inducible genetic clearance of p16^Ink4a-expressing senescent cells using p16^Ink4a trimodality reporter (p16–3MR)^(30,31) mice upon administration of ganciclovir (GCV) has also been shown to prevent multiple diseases of aging.⁽³²⁾ These genetic models have inspired interest in the discovery and development of drugs that therapeutically target senescent cells to improve healthspan.

As an alternative to the genetic senescent cell clearance approaches, and one potentially more applicable to humans, multiple groups have demonstrated in several models of aging and disease that reducing the senescent cell burden using drugs that either selectively eliminate senescent cells (i.e., “senolytics”) or modulate their SASP (i.e., “senomorphics”) prevents, delays, or alleviates chronic diseases in mice. For example, in the first study of its kind, the Kirkland group at Mayo Clinic exploited the dependence of senescent cells on pro-survival, anti-apoptotic pathways to thereby identify that the combination of Dasatinib (D; an FDA-approved tyrosine kinase inhibitor [TKI] in clinical use for treating hematologic disorders^(33,34) plus Quercetin (Q; a natural flavanol present in many fruits and vegetables⁽³⁵⁾) that targets BCL-2 family, HIF-1α, and insulin/IGF-1 networks⁽³⁶⁾), has in vitro senolytic activity by killing senescent cells without having noticeable effects on proliferating or quiescent, differentiated cells.⁽²¹⁾ In addition, several groups have reported that the BCL-2 family (BCL-2, BCL-W, and BCL-XL) inhibitors – navitoclax (ABT-263) and ABT-737, are senolytic in vitro in multiple senescent cell types^(37–39) and in vivo by eliminating senescent cells, thereby preventing progression of morbidity in models of aging and disease. Besides the development of senolytics, yet another approach has been to target the pro-inflammatory SASP of senescent cells using, for example, the senomorphic JAK-STAT signaling pathway inhibitor, ruxolitinib.⁽⁴⁰⁾ Other notable senolytic compounds that have recently been identified include fisetin^(37,41) (a flavonol with a similar structure to Q^(42,43)), piperlongumine,⁽⁴⁴⁾ a FOXO4 peptide that causes p53-dependent apoptosis,⁽⁴⁵⁾ UBX0101,⁽³²⁾ and HSP90 inhibitors.⁽⁴⁶⁾ These and other potential senolytic and senomorphic drugs have been recently reviewed,⁽⁴⁷⁾ and because of the inherently rapid evolving nature of the field, many more novel senotherapeutics are on the horizon. Given the current vast number of emerging senolytic agents that potentially target SCAPs (Table 1), this review will focus primarily on studies that have utilized D+Q or BCL-2 inhibitors (e.g., navitoclax), as these senolytics were among the first-generation compounds identified and, to date, have been most rigorously tested in the context of numerous preclinical models of aging and disease. The predominant focus will be placed on published studies that have utilized a combination of genetic and pharmacologic approaches to target senescent cells.

Table 1.

Compounds exhibiting senolytic activity and their senescent cell anti-apoptotic pathways (SCAPs). Adapted from EBioMedicine 2017;21:21–8.

Compound	SCAP	Reference
Dasatinib (D)*	Dependence receptor / Src kinase / tyrosine kinase	Aging Cell 2015;14:644–28
Quercetin (Q)†	Bcl-2 family, p53 / p21Cip1 / serpine, PI3K / AKT	Aging Cell 2015;14:644–58
Fisetin†	PI3K / AKT	Aging 2017;9:955–63
Navitoclax (ABT-263)‡	Bcl-2 family	Aging Cell 2016;15:428–35; Nat Med 2016;22:78–83; Nat Commun 2016;7:11190
ABT-737‡	Bcl-2 family	Aging Cell 2016;15:428–35; Nat Med 2016;22:78–83; Nat Commun 2016;7:11190
A1331852‡	Bcl-XL	Aging 2017;9:955–63
A1155463‡	Bcl-XL	Aging 2017;9:955–63
UBX0101‡	p53-MDM2 / p21Cip1	Nat Med 2017;23:775–81
Piperlongumine†	P53 / p21Cip1 & Bcl-2 family	Aging 2016;8:2915–26
17-AAG (Tanespimycin)*	HSP-90	Nat Commun 2017;8:422
Geldanamycin*	HSP-90	Nat Commun 2017;8:422
17-DMAG (Alvespimycin)*	HSP-90	Nat Commun 2017;8:422
FOXO4-related peptide‡	p53 / p21Cip1 / serpine & Bcl-2 family	Cell 2017;169:132–47
Ruxolitinib (INCB18424)*	Selective JAK1/2 inhibitor	PNAS 2015;112:E6301–10; Elife 2015;4:e12997

^*FDA approved for other indications;

^†Available as a non-FDA regulated supplement;

^‡Investigational use only.

As summarized herein, mounting evidence (mostly in mice) has established that senescent cells represent promising therapeutic targets to delay or improve functional deficits and chronic disorders of aging. For example, senolytic therapy in mouse models of aging and disease has been shown to extend healthspan by preventing or alleviating the following: osteoporosis,⁽²⁷⁾ frailty,⁽⁴⁸⁾ atherosclerosis,^(28,49) hepatic steatosis,⁽⁵⁰⁾ osteoarthritis,⁽³²⁾ idiopathic pulmonary fibrosis,⁽⁵¹⁾ obesity-induced anxiety,⁽⁵²⁾ tau-mediated neurodegenerative disease,^(53,54) and metabolic dysfunction/type 2 diabetes (T2D),^{(21,29,55,56)} as well as several other conditions. In addition, intermittent administration of senolytics has been shown to compress morbidity and extend maximal lifespan in mice even when therapy is initiated late in life.⁽⁴⁸⁾ Thus, based on the evidence summarized herein, targeting cellular senescence appears to be an innovative approach to delay aspects of age-related multi-morbidity and chronic diseases as a group compared to the current approach which treats each condition separately.

Osteoporosis.

To date, multiple groups have found in mice that with aging, at least a subset of cells of different lineages located in the bone marrow and bone itself become senescent.^(57,58) Furthermore, some of these senescent cell populations, including senescent myeloid cells and senescent osteocytes, develop a profound SASP signature in the context of natural chronological aging.⁽⁵⁷⁾ Importantly, similar results were found in bone biopsies from older postmenopausal as compared to younger women,⁽⁵⁷⁾ establishing that senescent cells are present at the time and location of age-related bone loss in humans, as in mice. These findings thus set the stage for determining whether cellular senescence has a causal role in mediating osteoporosis. To examine this question, Farr et al.⁽²⁷⁾ used genetic (INK-ATTAC⁽²⁵⁾), senolytic (D+Q⁽²¹⁾), and senomorphic (JAK1/2 inhibitor⁽⁴⁰⁾) approaches to target senescent cells and assessed the impact of each of these interventions on age-related osteoporosis in old mice. These studies demonstrated that the skeletal effects of intermittent D+Q therapy in old wild-type mice were virtually identical to those observed in the old INK-ATTAC mice treated with AP20187 (a synthetic drug that induces suicide transgene activation⁽²⁵⁾), demonstrating that senescent cell clearance via either genetic or senolytic approaches prevents age-related osteoporosis in mice.⁽²⁷⁾ Similar findings were also observed in old wild-type mice treated with the senomorphic SASP inhibitor, ruxolitinib.⁽²⁷⁾ By histomorphometry, targeting senescent cells using either genetic or senolytic approaches reduced bone resorption, although, despite this change, bone formation was either maintained (trabecular sites) or was higher (cortical sites) following senescent cell clearance.⁽²⁷⁾ Thus, from a translational therapeutic perspective, targeting senescent cells using senolytics or senomorphics may offer advantages over conventional anti-resorptive therapies (reviewed in^(59–61)) as the latter inhibit/eliminate osteoclasts to reduce bone resorption, with a concomitant reduction in bone formation due to coupling between osteoblasts and osteoclasts. By contrast, by reducing the senescent cell burden, senolytics simultaneously suppress/reduce osteoclastic bone resorption with favorable effects on bone formation at both cortical and trabecular skeletal sites, leading to positive “bone balance”.⁽²⁷⁾

In support of these findings, Chandra et al.⁽⁶²⁾ recently showed in young adult mice that focal radiation therapy induces accelerated bone cell senescence, elevated SASP factors in bone, and significant bone loss associated with deficient bone formation, all of which can be at least partially reversed by senoltyic therapy (D+Q). Similarly, Yao et al.⁽⁶³⁾ recently found that the genetic clearance of senescent cells using INK-ATTAC protects against doxorubicin-induced bone loss in young adult mice. Together, these studies have potential translational relavence for the treatment of age-associated osteoporosis as well as radiation- and chemotherapy-induced bone loss.^(27,62,63)

Frailty.

Several studies have aimed to establish whether genetic or pharmacological removal of senescent cells in mice improves physical function and prevents aspects of frailty, which is a multifaceted geriatric disorder characterized by lower physical activity, decreased muscle strength and endurance, and the inability to appropriately respond to stress.⁽⁴⁰⁾ In BubR1 progeroid mouse background, clearance of p16^Ink4a-expressing cells using INK-ATTAC improved physical function, however, parallel findings using this approach were not necessarily found in naturally aged INK-ATTAC mice (treated with AP20187 from 12- to 18-months).⁽²⁶⁾ Xu et al.⁽⁴⁰⁾ reported that continuous administration of the senomorphic JAK1/2 inhibitor, ruxolitinib, to old wild-type mice for ten weeks alleviated inflammation in fat tissue as well as systemically in the circulation, and that blocking the SASP with ruxolitinib enhanced physical function and improved multiple aspects of frailty, such as rearing (decreased rearing is a measure of frailty in mice⁽⁶⁴⁾) as well as ambulation counts, hanging endurance, and grip strength. In subsequent studies, Xu et al.⁽⁴⁸⁾ first aimed to determine the extent to which cellular senescence is causative in directly driving frailty by transplanting relatively small (e.g., less than 1.0 × 10⁶) numbers of senescent cells into young mice, which not only caused long-term persistent physical dysfunction but also resulted in the spread of cellular senescence to distant tissues within the host animal. Importantly, these phenotypes resembling accelerated aging were rescued by intermittently treating these young mice “aged” by senescent cell transplantation with senolytics, D+Q.⁽⁴⁸⁾ Furthermore, intermittent D+Q administration to naturally aged, old wild-type mice over four months resulted in senescent cell clearance and improved muscle performance/function (e.g., maximal treadmill speed, treadmill endurance, hanging endurance, and grip strength) and significantly extended lifespan as late-life post-treatment survival was increased by 36%.⁽⁴⁸⁾ Collectively, these studies demonstrate that targeting cellular senescence using multiple approaches in old mice and mouse models of accelerated aging can enhance healthspan by improving physical function and reversing aspects of frailty, which contributes to lifespan extension. Furthermore, these studies potentially implicate cellular senescence as a key driver of the diminished response to anabolic skeletal muscle interventions, including both pharmacologic and exercise, which older individuals experience with advancing age.

Cardiovascular Disease.

Cellular senescence was hypothesized years ago to have causal roles in mediating cardiovascular disease⁽⁶⁵⁾ as senescent cells were known to accumulate in endothelial and vascular smooth muscle cells with atherosclerosis.^(66,67) Nevertheless, whether and how senescent cells directly contribute to the development of cardiovascular disease and the extent to which clearance of senescent cells reverses these effects remained unclear until recently. In the first study to report the effects of senolytic therapy on cardiovascular functional outcomes, a single dose of D+Q administered to old mice was followed by significant improvements in cardiac ejection fraction and carotid vascular reactivity five days later.⁽²¹⁾ In subsequent studies, Roos et al.⁽²⁸⁾ found that a longer duration (3-months) of intermittent D+Q administration resulted in senescent cell clearance in the aorta medial layer of old and hypercholesterolemic mice that was associated with improvements in vasomotor function as reflected by increased vascular smooth muscle contractile function of the carotid arteries and increased levels of NADPH oxidase-related protein in the intima of the aorta. Parallel findings were observed by genetic clearance of senescent cells using INK-ATTAC in old mice with normal cholesterol levels.⁽²⁸⁾ Using the low-density lipoprotein receptor-deficient (Ldlr^−/−) mouse model of atherosclerosis, Childs et al.⁽⁴⁹⁾ demonstrated that macrophages displaying features of senescence accumulate in the subendothelial space of the aorta during early atherogenesis and drive aberrant pathology via their SASP. They then found in Ldlr^−/− mice with more advanced plaques that eliminating senescent cells using genetic (INK-ATTAC and p16–3MR) or pharmacological (with the senolytic, navitoclax) approaches blocks plaque growth and aberrant remodeling processes.⁽⁴⁹⁾ With regards to heart tissue, recent work by Anderson et al.⁽⁶⁸⁾ showed that with aging, both human and murine post-mitotic cardiomyocytes acquire a senescence-like phenotype characterized particularly by DNA damage at telomere regions. Moreover, senescent cell clearance using either INK-ATTAC or navitoclax in old mice alleviated key aspects of cardiac aging, such as myocardial fibrosis and hypertrophy.⁽⁶⁸⁾ Collectively, these studies establish that senescent cells are key drivers of vasomotor dysfunction, atherosclerosis, and myocardial dysfunction at various stages of disease pathogenesis. Therefore, senescent cells may represent promising therapeutic targets for disease management in patients with cardiovascular disease.

Osteoarthritis.

Aging and the accumulation of senescent cells in cartilage have also been implicated in the development and progression of osteoarthritis.^(69,70) To begin to investigate the potential causal roles of cellular senescence in mediating osteoarthritis, Xu et al.⁽⁷¹⁾ transplanted luciferase-labeled senescent cells isolated from ear cartilage into the knee joint space of young adult wild-type mice, which after three months resulted in several features suggestive of osteoarthritis, including leg pain, impaired mobility, and both radiographic as well as histological changes that were consistent with the development of osteoarthritis. By contrast, transplanting non-senescent cells has lesser effects, thus establishing that senescent cells are causative in osteoarthritis pathogenesis.⁽⁷¹⁾ These effects are presumably due to detrimental SASP effects on articular cartilage as Diekman et al.⁽⁷²⁾ recently found that, although p16^Ink4a expression increases in cartilage of both mice and humans with advancing age, the role of senescent cells locally is not due to the loss of chondrocyte replicative function as somatic deletion or inactivation of p16^Ink4a in chondrocytes of adult mutant mice did not alter osteoarthritis progression after trauma invoked by destabilization of the medial meniscus or with natural aging. Further support for the direct causal role of cellular senescence in mediating osteoarthritis comes from a study by Jeon et al.⁽³²⁾ demonstrating in p16–3MR mice that following trauma-induced anterior cruciate ligament transection senescent cells accumulate locally in both articular cartilage and the synovium of the knee joint space of mice. Moreover, in osteoarthritis murine models of trauma or aging, the selective clearance of senescent cells using either genetic or intra-articular injection of a senolytic (UBX0101) mitigated pain, prevented the progression of post-traumatic osteoarthritis and enhanced cartilage development.⁽³²⁾ Therefore, cellular senescence may represent a viable therapeutic target for the treatment of osteoarthritis.

Pulmonary Fibrosis.

Wound healing and the ability to resolve tissue fibrosis in response to damage is significantly diminished in older animals and humans. This is also evident in fibrotic lung diseases, such as idiopathic pulmonary fibrosis (IPF), which are associated with aging and a dramatic accumulation of senescent cells.⁽⁷³⁾ Cellular senescence has been implicated as a central mechanism in IPF pathogenesis and in limiting the disease.⁽⁷³⁾ Indeed, Schafer et al.⁽⁵¹⁾ showed that while acutely generated senescent cells facilitate repair, longer-lasting (i.e., chronic) senescent cells increasingly accumulate in the alveolar epithelium with disease progression, resulting in DNA damage, inflammation, tissue matrix remodeling, and lung fibrosis. They then showed using the bleomycin-injury IPF mouse model, that senescent cell clearance by either INK-ATTAC or senolytics (D+Q) improves pulmonary and physical function as reflected by increased exercise capacity (during a graded treadmill exercise test), although with the caveat that it does not alter lung fibrosis.⁽⁵¹⁾ Nevertheless, these findings establish that targeted elimination of senescent cells in IPF improves lung function and physical function.⁽⁵¹⁾ Furthermore, these findings are also consistent with the first-in-human pilot clinical trial of D+Q that provided initial evidence that senolytic therapy may improve physical function in humans with IPF; however, future placebo-controlled trials are needed to more rigorously test the effects of senolytics in IPF patients.⁽⁷⁴⁾

Renal Disease.

Senescent cells increasingly amass in the kidneys with natural aging and are associated with the age-related decline in renal function in rodents and humans.^(75,76) Selective clearance of senescent cells starting at one year of age was shown to significantly extended the median lifespan and prevent glomerulosclerosis as well as other aspects of kidney dysfunction in natural, chronologically aged INK-ATTAC mice.⁽²⁶⁾ These findings suggest that the removal of senescent cells with senolytics may alleviate renal dysfunction in older individuals or patients with kidney disease. While current evidence from preclinical studies supporting this premise is somewhat scarce, Baar et al.⁽⁴⁵⁾ found that administration of a novel senolytic (i.e., a modified FOXO4-p53 interfering peptide) restored kidney function in a mouse model of accelerated aging (Xpd^TTD/TTD mice) and naturally aged wild-type mice. With regards to human translation, three consecutive days of D+Q therapy in an open-label Phase 1 pilot study was recently shown to decrease the senescent cell burden in adipose tissue, skin, and blood of patients with diabetic kidney disease,⁽⁷⁷⁾ which sets the stage for additional clinical testing of D+Q in larger randomized controlled trials.

Neurodegenerative Diseases.

The risk for developing the majority of chronic neurodegenerative diseases, such as Parkinson’s and Alzheimer’s diseases, increases dramatically with advancing age. Numerous recent studies have identified cells with senescence-like features in the setting of both neurodegenerative diseases and in various regions of the brain with natural aging,^(78–81) thus raising the possibility that senescent neurons drive cognitive dysfunction. Tau protein deposition and accumulation (i.e., tau toxicity) is a common hallmark feature among neurodegenerative diseases, including Alzheimer’s disease. Recently, Musi et al.⁽⁵³⁾ utilized multiple models of Alzheimer’s disease to show that tau-containing neurofibrillary tangle-containing neurons display a senescent-like phenotype. Interestingly, in a late-life mouse model with established tau toxicity, intermittent senolytic (D+Q) therapy alleviated neurofibrillary tangle-associated pathology and significantly improved brain structure and function as reflected by reduced cortical brain atrophy and increased cerebral blood flow.⁽⁵³⁾ These findings are consistent with a study by Bussian et al.⁽⁵⁴⁾ that examined the roles of cellular senescence in another tau toxicity mouse model of neurodegeneration (MAPTP^301SPS19 mice). This study demonstrated that neuronal expression of mutant tau causes senescence of astrocytes and microglia and that preventing the accumulation of these senescent cells (using INK-ATTAC or senolytic therapy [ABT-263]) can alleviate the cognitive decline and neurodegeneration that APTP^301SPS19 mice normally experience.⁽⁵⁴⁾ Taken together, these findings establish that cellular senescence represents a mechanism that contributes to tau toxicity in the context of neurodegenerative diseases, and that targeting senescent cells may have benefits even when a particular disease has progressed to symptomatic stages, which has potential broad clinical implications for patients with various different types of brain diseases that share tau-dependent pathology.

Hepatic Steatosis.

The incidence of excessive accumulation of fat in the liver (hepatic steatosis or non-alcoholic fatty liver disease [NAFLD]) is more common in older individuals, suggesting that cellular senescence may have a role in mediating NAFLD. To rigorously examine this question, Ogrodnik et al.⁽⁵⁰⁾ used multiple approaches. First, they demonstrated in mice that as compared to ad libitum feeding, dietary restriction protected against fat deposition and the accumulation of senescent cells in the liver. They next generated mutant mice with liver-specific deletion of the DNA repair gene Xpg (i.e., Alb-Xpg mice), which displayed increased hepatocyte-specific senescence and higher liver fat accumulation earlier in adulthood. These findings were complemented by studies in old mice in which senescent cells were cleared either genetically (using INK-ATTAC) or with senolytics (D+Q); both approaches alleviated hepatic steatosis.⁽⁵⁰⁾ This study thus establishes that cellular senescence mediates hepatic steatosis and that senescent cell clearance may represent a novel approach to treat NAFLD.

Metabolic Dysfunction.

Aging and obesity are the leading predictors of T2D, metabolic syndrome, and their related complications, yet available treatments for these conditions are suboptimal.⁽⁸²⁾ Mounting evidence, however, now establishes that aging, obesity, and T2D are each associated with the accumulation of senescent cells in adipose and other tissues of rodents and humans.^{(55,56,83–85)} With aging, senescent cells can accumulate, for example, in fat tissue where they have been hypothesized to cause insulin resistance in metabolic tissues, inhibit adipogenesis, and release factors that attract immune cells to exacerbate metabolic dysfunction.⁽⁸⁴⁾ Targeting cellular senescence in old mice either genetically (using INK-ATTAC) or with a JAK1/2 SASP inhibitor (ruxolitinib) alleviates multiple aspects of age-related metabolic dysfunction including adipose tissue inflammation and progenitor cell dysfunction.⁽²⁹⁾ In mice, prolonged high-fat diet-induced obesity causes the accelerated or premature accumulation of senescent cells earlier in life in adipose and other tissues,^(83–85) resulting in obese young adult mice that are “aged” by their premature cellular senescence and metabolic dysfunction. A recent study by Palmer et al.⁽⁵⁵⁾ used genetic (INK-ATTAC and p16–3MR) as well as senolytic (D+Q) interventions to demonstrate that clearing senescent cells systemically in young adult obese mice (given high-fat diet) alleviates metabolic dysfunction by improving glucose homeostasis and insulin sensitivity, blunting the SASP and its mediators, and promoting healthy adipogenesis. It is noteworthy that exercise alone is also sufficient to prevent the premature accumulation of senescent cells, attenuate the SASP, and reduce aspects of systemic metabolic dysfunction caused by high-fat diet feeding in young adult mice.⁽⁸⁵⁾ These findings point to the unique possibility of combining modifiable lifestyle choices with senolytic therapy to improve healthspan. Insulin resistance in association with T2D has also been shown to cause the premature acceleration of pancreatic β-cell senescence, which in mice and humans with T2D display abnormal insulin secretion, downregulated β-cell genes, and increased expression of SASP factors.⁽⁵⁶⁾ This study induced insulin resistance in mice both pharmacologically (using S961) or physiologically (by high-fat diet), and showed in both situations that systemic senescent cell clearance by either INK-ATTAC or senolytics (i.e., both navitoclax and D+Q) can improve glucose homeostasis and β-cell function while simultaneously reducing senescence and SASP markers.⁽⁵⁶⁾ Collectively, these preclinical findings establish that targeting senescent cells can alleviate age- and obesity-related metabolic dysfunction, and point to evidence-based therapeutic approaches of targeting cellular senescence to prevent and perhaps alleviate T2D and its complications.

Conclusions

In summary, much of the population is now faced with an enormous burden of age-related chronic diseases and co-morbidities. However, recent discoveries in geroscience indicate that healthspan in model organisms such as mice can be manipulated by targeting fundamental hallmark mechanisms of aging, such as cellular senescence. Indeed, senescent cells and their pro-inflammatory secretome have emerged as significant contributors to age-related tissue dysfunction and morbidity. For example, studies in mice demonstrate that cellular senescence has causal roles in mediating osteoporosis, frailty, cardiovascular disease, osteoarthritis, pulmonary fibrosis, renal disease, neurodegenerative diseases, neuropsychiatric disorders, hepatic steatosis, and metabolic dysfunction. Moreover, therapeutically targeting senescent cells in mice can prevent, delay, or alleviate each of these conditions. Therefore, senotherapeutic approaches, including senolytics and senomorphics, that either selectively eliminate senescent cells or interfere with their ability to promote tissue dysfunction, are gaining momentum as potential realistic strategies to abrogate human senescence to thereby compress morbidity and extend healthspan.