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Nature Reviews Drug Discovery | Voume 23 | November 2024 | 817–837 817
nature reviews drug discovery https://doi.org/10.1038/s41573-024-01033-z
Review article Check for updates
Therapeutic targeting of
senescent cells in the CNS
Markus Riessland 1,2, Methodios Ximerakis 3, Andrew A. Jarjour 4, Bin Zhang5,6 & Miranda E. Orr 7,8
Abstract
Senescent cells accumulate throughout the body with advanced
age, diseases and chronic conditions. They negatively impact health
and function of multiple systems, including the central nervous
system (CNS). Therapies that target senescent cells, broadly
referred to as senotherapeutics, recently emerged as potentially
important treatment strategies for the CNS. Promising therapeutic
approaches involve clearing senescent cells by disarming their
pro-survival pathways with ‘senolytics’; or dampening their
toxic senescence-associated secretory phenotype (SASP) using
‘senomorphics’. Following the pioneering discovery of rst-generation
senolytics dasatinib and quercetin, dozens of additional therapies
have been identied, and several promising targets are under
investigation. Although potentially transformative, senotherapies
are still in early stages and require thorough testing to ensure reliable
target engagement, specicity, safety and ecacy. The limited
brain penetrance and potential toxic side eects of CNS-acting
senotherapeutics pose challenges for drug development and
translation to the clinic. This Review assesses the potential impact
of senotherapeutics for neurological conditions by summarizing
preclinical evidence, innovative methods for target and biomarker
identication, academic and industry drug development pipelines and
progress in clinical trials.
Sections
Introduction
Mechanisms leading to CNS
cell senescence
CNS senotherapeutics in
development
Challenges in developing CNS
senotherapies
Applying AI to senotherapy
development
Conclusions
1Department of Neurobiology and Behavior, Stony Brook University, Stony Brook, NY, USA. 2Center for Nervous
System Disorders, Stony Brook University, Stony Brook, NY, USA. 3Merck & Co.,Inc., Cambridge, MA, USA. 4MSD
(UK) Ltd, London, UK. 5Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai,
New York, NY, USA. 6Mount Sinai Center for Transformative Disease Modeling, Icahn School of Medicine at Mount
Sinai, New York, NY, USA. 7Department of Internal Medicine, Section on Gerontology and Geriatric Medicine, Wake
Forest University School of Medicine, Winston-Salem, NC, USA. 8Salisbury VA Medical Center, Salisbury, NC, USA.
e-mail: morr@wakehealth.edu
Nature Reviews Drug Discovery | Voume 23 | November 2024 | 817–837 818
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cells and cerebral vasculature cells, may undergo senescence owing to
a range of stressors, such as physical, psychological, viral, oxidative and
proteotoxic insults, DNA damage and protein accumulation (Table1).
In this Review, we describe mechanisms involved in central nervous
system (CNS) cell senescence, and consider how these processes might
be therapeutically targeted. We balance optimistic promise for their
potential, including results from first clinical trials33, with an objective
perspective on the challenges faced and strategies to overcome them.
Mechanisms leading to CNS cell senescence
Cell cycle dysfunction
34,35
, ‘loss of cell identity’
36,37
or lysosomal stress
38
may all represent key steps in the conversion from a healthy brain cell
type into a final senescent state. Neurons, the cellular building blocks
of the nervous system, communicate through electrical signals. They
are necessary for the processing, storage and retrieval of information.
Neurons are vulnerable to somatic mutations39,40 and insoluble protein
accumulation41, which are known drivers of senescence through DNA
damage pathway activation10 and protein aggregation9,19.
Non-neuronal brain cell types, such as glia, immune cells, endothe-
lial cells and vascular smooth muscle cells, have crucial roles in main-
taining the health and function of the nervous system by modulating
neuronal activity, preventing and removing toxins, and nourishing the
tissue. These cells may be especially vulnerable to cell non-autonomous
stressors such as external protein aggregates and cellular by-products.
For example, in AD and other tauopathies, intraneuronal tau aggre-
gates drive neuronal senescence9,19, and tau oligomers released from
neurons trigger astrocyte42 and vascular senescence43. Microglia can
enter senescence after phagocytosing protein aggregates, includ-
ing clearing senescent neurons that contain neurofibrillary tangles
(NFTs) composed of aberrantly phosphorylated tau aggregates44.
Similarly, amyloid-β, another hallmark of AD, can induce senescence
in oligodendrocyte precursor cells
45
and endothelial cells
46
. TNF, a
cytokine secreted by many cell types, including brain microglia and
senescent cells, can induce neuronal senescence, increasing the release
of α-synuclein
47
, a synaptic protein that aberrantly accumulates in
Parkinson disease and other neurodegenerative diseases. A model
depicting senescent cell initiation and propagation across brain cell
types is presented in Fig.1, and the features of senescent cells, including
postmitotic senescent cells, are depicted in Fig.2.
Senescent cells that accumulate within the brain or in periph-
eral tissues negatively impact brain health and function (Table1) .
Senescent cell accumulation in the brain may be driven by comorbidi-
ties that increase the risk of developing cognitive impairment or brain
dysfunction (for example, insulin resistance, obesity, alcohol use
disorder
48–50
), or by physical stress to the nervous system such as trau-
matic brain injury
51
or spinal cord injury
52
. Emerging evidence suggests
that senescence is a contributing and causal factor for chronic pain and
psychological stress-induced mental disorders, primarily through the
release of inflammatory SASP factors
49,50,53,54
. Individuals with a high
bloodSASP index tend to experience more severe depressive episodes
than individuals with low SASP indices
55
. Furthermore, elevated SASP
expression levels have been linked to poorer treatment response and
reduced remission rates in late-life depression
56
. These findings in
humans are supported by mechanistic studies in mice, demonstrating a
causal link between senescence and depressive behaviours49.
In 2018, studies began to report improvements in brain structure
and function by elimination of senescent cells in transgenic mouse mod-
els of neurodegeneration
9,12,57,58
. Mechanistic data indicated that clear-
ing senescent cells improved brain structure, function and memory
Introduction
Cellular senescence was first described in 1961 as a phenomenon
whereby primary cells in culture undergo a finite number of divisions
1
.
This limited replicative potential was believed to be an artefact of cells in
culture driven by telomere erosion2,3. The eventual discovery of molecu-
lar biomarkers enabled the identification of senescent cells in vivo4,5.
However, the inclusion ofcell cycle arrest in the definition of senes-
cence cast doubt on the relevance of senescence in tissues with mostly
non-dividing cells, such as the brain. The first study to report senescent
neurons in mouse brain tissue was transformative to the field6 but was
met with scepticism. Later discoveries of senescent-like neurons
7–10
and astrocytes11,12 in post-mortem tissue from patients with Alzheimer
disease (AD) and Parkinson disease supported the initial findings and
highlighted the potential translational importance of senescent cells
in the brain. These studies, along with the identification of postmitotic
senescent-like cells in other tissues
13
, inspired the field to reconsider
the senescence-defining criteria. Cellular senescence now refers to
an end-stage change in cell fate, orchestrated through simultaneous
and sustained activation of opposing pro-apoptotic and pro-survival
molecular pathways
14
. For the context of this Review, we focus only on
pathological senescence that arises through chronic stress activation,
distinguishing it from physiological senescence, which occurs during
development, tissue regeneration or wound healing15,16.
Similar to other changes in cell fate, senescent cells acquire
distinct morphologies and remain metabolically active17. They
often develop enlarged somas both in vitro
18
and in vivo
9,19
, exhibit
increased lysosomal senescence-associated β-galactosidase (SA-β-gal)
activity20,21, elevated glycolysis22 and lipid metabolism23, and acquire
a secretory phenotype16,24. The molecules they release, known as
thesenescence-messaging secretome (SMS)
24
, or more commonly
as thesenescence-associated secretory phenotype (SASP)16, com-
municate with their neighbouring environment and the immune sys-
tem either to prepare the tissue for disease or to facilitate their own
clearance, respectively. The presence of senescent cells perpetuates
tissue inflammation and destruction through multiple mechanisms
25
.
If senescent cells are not actively removed by the immune system,
their sustained SASP signalling induces cytotoxicity or transforms
healthy cells into senescent cells. Phagocytic cells can also become
senescent after clearing senescent cells, which perpetuates inflam-
mation and pathogenic processes (Fig.1). Throughout the lifespan,
tissues and organs accumulate senescent cells, and their contribution
to disease and dysfunction earned them recognition as a “biological
hallmark of aging”26.
During the transition to senescence, cells undergo significant
changes in chromatin structure, alter their metabolism and morphol-
ogy, and become resistant to apoptosis
14
. The features of senescent cells
depend on the original cell type and the specific stressor involved
27
.
Various markers have been used to identify senescent cells, but a
confident identification requires evidence of multiple overlapping
phenotypes within the same cell, such as changes in morphology, macro-
molecular damage, cell cycle arrest and an inflammatory secretome14,28.
In addition to applying traditional senescence detection methods to
tissues of interest, the advancement in single-cell and single-nucleus
RNA sequencing, high-resolution multiplex spatial profiling techniques
and machine learning (ML) and artificial intelligence (AI) enable the
identification of rare senescent cells in complex tissues
19,29–32
. These
strategies have propelled the discovery of novel biomarkers of senes-
cence. Experimental evidence with new and established biomarkers
indicates that several brain cell types, including neurons, glia, stem
Nature Reviews Drug Discovery | Voume 23 | November 2024 | 817–837 819
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and reduced neuroinflammation and AD-associated neuropathology
(Table2). The clear link between senescent cell accumulation and
organ dysfunction and healthy lifespan
59–62
, along with the potential for
therapeutic interventions, led to a surge of funding to support research
on cellular senescence in ageing and disease, including in the CNS63,64.
The pioneering discovery of compounds that could eliminate
senescent cells, including in the brain, stimulated great interest in drug
development65. Therapeutic approaches under investigation include
methods to clear senescent cells from tissue, to gently revert them back
to their original cell type or to dampen their SASP. The type of senescent
cell and its associated mechanism of toxicity influence the choice of
therapeutic strategy. For instance, drugs designed to clear senescent
cells may be most suitable for replaceable cell types, such as microglia;
whereas non-clearing strategies may be more appropriate to target cells
that are difficult to replace, such as neurons. Research into lifestyle inter-
ventions and other non-pharmacological strategies to reduce senescent
cell burden, including approaches such as mindfulness to treat insomnia
in AD caregivers (NCT03538574), is also active and promising, and may
be effective for some individuals66–68. However, this Review focuses on
the various pharmacological approaches under investigation (Box1).
CNS senotherapeutics in development
To develop effective senotherapeutics for the CNS, strategies need
to incorporate various drug modalities against diverse targets and
• Protein aggregate
• Chronic stress
• Expose ‘eat me’ signals
Negative impact on
multiple cell types
Secrete SASP
Release partially
digested toxic proteins
b Senescent neuron
• Impaired tissue surveillance
e Senescent microglia
• Phagocytic receptor expression
c Activated microglia
• Microglial phagocytosis
• Partially digested
protein aggregates
d Neuronal phagoptosis
Phagocytic
receptor
'Eat me' signals
(PS, etc.)
Protein aggregate (tau, Aβ, α-synuclein,
FUS, Htt, PrP, SOD1, TDP43)
Secrete
neuronal SASP
Aberrant
neuronal
activity
Inlammatory
molecules DNA
damage
Metabolic
dysfunction
Oxidative
stress
Toxic
proteins
a Healthy neuron
Negative impact on
multiple cell types
Fig. 1 | Senescence initiation and spread across brain cells. a, Age and
disease-associated stressors that drive inflammation, reactive oxygen species,
toxic protein accumulation, metabolic dysregulation or DNA damage may
cause senescence in many cell types, including neurons. b, Senescent neurons
exhibit altered excitability and/or activity, display ‘eat me’ signals and secrete
deleterious molecules (the senescence-associated secretory phenotype
(SASP)) that negatively impact neuronal, vascular and glial cells. They may also
contain aggregate-prone, neurotoxic proteins that they transmit to other cells
as neuronal SASP9,19. c, Activated microglia expressing phagocytic receptors
recognize neuronal phagoptosis ‘eat me’ signals. d, Activated microglia engulf
senescent neurons166 and their content, including difficult-to-digest protein
aggregates, which may cause microglial senescence. e, Senescent microglia
exhibit reduced phagocytic and surveillance function44. They release partially
digested, neurotoxic fragments of protein aggregates and SASP factors
that cause astrocytes42, microglia44,57, vasculature43 and oligodendrocyte
precursor cells45 to become senescent. Aβ, amyloid-β; Htt, huntingtin; PrP,
prion protein;PS,phosphatidylserine; SOD1,superoxide dismutase 1;
TDP43,TAR DNA-binding protein 43.
Nature Reviews Drug Discovery | Voume 23 | November 2024 | 817–837 820
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senescent cell types. This section describes therapeutic strategies
that range from inhibition of specific senescence-associated sig-
nalling pathways, such as those that control SASP, to activation of
apoptosis-inducing pathways. Several molecules have been identified
as potential senotherapeutic targets (Table 2 and Fig.3). Small-molecule
approaches are the most straightforward option for targeting senes-
cent cells in the CNS owing to their potential for oral bioavailability
and relatively favourable prof ile regarding CNS penetration across
the blood–brain barrier (BBB). Candidate molecular processes can
be targeted with senolytics, which destroy senescent cells, or with
senomorphics, which modulate senescent cells (Box1). We also explore
innovative approaches such as targeting proteins specific to senescent
cells for degradation. Although not all the targets and approaches dis-
cussed in the following paragraphs have been directly tested in the CNS,
they describe basic principles for tackling senescent cells and lay the
groundwork for the future development of CNS-targeted applications.
Senolytics
Senolytics are a class of small-molecule therapies designed to selectively
eliminate senescent cells. The term ‘senolytics’ originates from combin -
ing ‘senescence’ and ‘lytic,’ indicating the literal destruction of senescent
cells. These cells are inherently damaged with upregulated apoptotic
pathways. Their long-term survival relies on concurrent upregulation
of pro-survival pathways inhibiting full engagement of apoptosis. Many
senolytics function by deactivating these survival pathways, referred
to as senescent cell anti-apoptotic pathways (SCAPs),allowing senes-
cent cells to undergo apoptosis (Table 2 and Fig.3). The therapeutic
druggability of senescence survival pathways is explored fur ther below.
PI3K and Src kinase inhibition. Phosphatidylinositol 3-kinases
(PI3Ks) are lipid kinases that have been shown to take crucial roles in
the regulation of cell cycle, apoptosis, DNA repair, angiogenesis, cel-
lular metabolism and cellular senescence
69
. Dysregulation of the PI3K
pathway is observed in one-third of human tumours, which has led
to the development of PI3K inhibitors as cancer therapeutics70. Simi-
larly, the Src family kinases are involved in regulation of important
cellular functions such as cell proliferation, differentiation, apoptosis,
migration and metabolism
71
. Early activation of Src has been implicated
in shifting the cell fate from apoptosis to senescence72.
A significant advancement in the field of senolytic agents was the
discovery of a combinatorial therapeutic approach involving dasat-
inib and quercetin. Dasatinib, a Src tyrosine kinase inhibitor with FDA
Table 1 | Evidence of cellular senescence in ageing and CNS conditions
Condition Astrocytes Microglia Neurons Other CNS cells
Alzheimer
disease p16 (refs. 11,251), p53 (ref. 251)p16 (ref. 202) morphology, DNA damage,
HMGB1 (ref. 252); protein panel(ref.251)Transcriptome(refs.9,19,208,253)
p19+ NFTs, karyomegaly,
lipofuscin(ref.19)
OPCs(ref.45)
Endothelial cells(ref.200)
Ageing p16 (refs. 11,254); p21,
SA-β-gal254; loss of HMGB1,
SASP, p16 (ref. 42)
p16 (ref. 255); protein panel(ref.251)p21 (ref. 6); Gdf11 knockout
(ref.104)OPCs(ref.255)
VSMCs(ref.31)
Neuroblasts(ref.256)
Amyotrophic
lateral sclerosis SA-β-gal(ref.257), DNA
damage(ref.257)
p16 (refs. 251,258–260),
p53 (ref. 251)
p21 (ref. 258); γH2AX(ref.257)
p16 (refs. 259,260), loss of lamin B1
(ref. 259)p16 (ref. 259); p21 (ref. 258) –
Chronic pain Telomere erosion, p53
(ref. 53)Telomere erosion, p53 (ref. 53)
SA-β-gala (ref. 261)
Telomere erosion, p53 (ref. 53) –
Parkinson
disease Loss of lamin B1 (ref. 12) – p21 and loss of SATB1 (ref. 10) –
Stroke SA-β-gal(ref.262),
p16 (refs. 262,263);
SASP(ref.263)
p16, p21, SASP(ref.264)p16, p21, SASP(ref.264)Endothelial cells(ref.263)
Tauopathy FTLD: HMGB1, SASP, p16
(ref. 42)SA-β-gal, SASP, MMP3 (ref. 44); p16 (ref. 57)PSP: Cdkn2a, DNA damage,
karyomegaly(ref.9)–
Traumatic brain
injury or blast
exposure
Cyclin D1, PCNA, p16,
p21, SA-β-gal(ref.265);
γH2AX, loss of lamin B1,
SASP(ref.266)
Cyclin D1 (ref. 265); PCNA(ref.265);
p16 (refs. 265,267); p21 (refs. 265,267);
SA-β-gal(ref.265); Bcl-2 (ref. 267),
lipofuscin(ref.267) and γH2AX(ref.267)
DNA damage(ref.51);
p16 (refs. 51,265);
p21 (refs. 51,265)
Cyclin D1, PCNA,
SA-β-gal (ref.265)
Ependymal cells(ref.266)
Oligodendrocytes(ref.266)
Other conditions Obesity(ref.49) – Alcohol use disorder(ref.48);
Glaucoma(ref.268)
Insulin resistance(ref.50)
Spinal cord injury(ref.52)
SARS-CoV-2 (ref. 269)
Glioblastoma
(refs.203,270,271)
MS: NPCs(ref.272)
CNS, central nervous system; FTLD, frontotemporal lobar degeneration; MS, multiple sclerosis; NFT, neuroibrillary tangle; NPC, neural precursor cells; OPC, oligodendrocyte precursor cell;
p16, p16INK4a; p19, p19INK4d; p21, p21CIP1; PSP, progressive supranuclear palsy; SA-β-gal, senescence-associated β-galactosidase; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2;
SASP, senescence-associated secretory phenotype; VSMC, vascular smooth muscle cell. Evidence of senescence using bulk tissues without cell speciicity not included. aCell type inferred
from in vitro model, but not conirmed in vivo.
Nature Reviews Drug Discovery | Voume 23 | November 2024 | 817–837 821
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approval for leukaemia treatment, was combined with quercetin,
a naturally occurring flavonoid that inhibits PI3K65. Although dasatinib
had already demonstrated its ability to induce apoptosis in cancer cells,
it initially showed no significant effects on senescent cell clearance
when used alone. Similarly, quercetin, a relatively nonspecific PI3K
inhibitor known for its anti-inflammatory properties and broad kinase
inhibitory profile, demonstrated only modest senolytic potential as
a monotherapy. However, the combination of dasatinib with querce-
tin synergistically influenced SCAPs and demonstrated significant
senolytic potential65.
This combination of dasatinib and quercetin has since proved
effective in eradicating senescent cells and improving health out-
comes in mouse models of tau-associated neurodegeneration9,
AD-associated amyloid-β accumulation45, frailty65, osteoporosis65,
vasomotor dysregulation65,73, hepatic steatosis74, insulin resistance75,
pulmonary fibrosis76, adipose tissue inflammation77, intervertebral
disc degeneration78 and skeletal muscle debility79. The success of
dasatinib–quercetin has stimulated considerable interest in combat-
ting age-related CNS conditions in human trials (Table3). Although the
side effects of long-term use of dasatinib–quercetin remain unknown,
to date all the clinical trials have reported favourable safety outcomes.
Chronic complications of dasatinib treatment in cancer therapy include
recurrent pleural effusions in up to 37% of patients, as well as pos-
sible side effects affecting platelet function, leading to a bleeding
tendency
80
. To minimize potential severe side effects such as these,
dasatinib–quercetin senolysis treatment is administered intermit-
tently. Despite dasatinib’s short elimination half-life of less than 11h,
and quercetin’s poor bioavailbility81, early data from clinical trials
indicate that dasatinib–quercetin is potent enough to significantly
reduce the senescent cell burden in humans using this ‘hit-and-run’
regimen82, which simultaneously improves the safety prof ile.
The first-in-human trial of dasatinib–quercetin demonstrated
safety in a clinical population of idiopathic pulmonary fibrosis
83
,
as did the first phase I trial in older adults with mild cognitive
impairment (MCI) or early AD33. The MCI/early AD open-label study
enrolled five participants to assess the safety of an intermittent
dasatinib–quercetin treatment protocol and determine BBB pen-
etrance of the compounds. Dasatinib and quercetin were adminis-
tered orally for two consecutive days followed by a 2-week break,
repeated for six cycles. Drug levels were measured in plasma and
cerebrospinal fluid (CSF) under fasting conditions at baseline, before
drug administration and 80–150min after the final dose of study drug.
After treatment, both dasatinib and quercetin were detectable in the
plasma of all participants at higher levels than baseline. Dasatinib was
also detected in the CSF in 80% of the participants after treatment;
however, quercetin was not. The lack of quercetin detection could
be attributed to its low bioavailability, levels being below detection
sensitivity, the timing of CSF collection not capturing quercetin
Change in
membrane
potential
Axonal/dendritic retraction
SASP
DNA
damage ↓ Lamin B1
↑ Nuclear size
Morphological changes
Lysosomal
dysfunction
Mitochondrial
dysfunction
↑ p21/p16/p19
expression
Fig. 2 | Cellular phenotypes of postmitotic senescent
neurons. Cellular phenotypes of postmitotic
senescent neurons include morphological changes,
lysosomal and mitochondrial dysfunction, DNA
damage, increased nuclear size, decreased expression
of lamin B1, increased expression of p21, p16 and p19,
changes in membrane potential and a senescence-
associated secretory phenotype (SASP). The black
arrows indicate axonal and dendritic retraction,
which is a feature of postmitotic senescent neurons
that does not occur in non-neuronal cell types.
Processes highlighted in blue are being explored as
senotherapeutic targets across senescent cell types.
Increased β-galactosidase, lipofuscin accumulation,
proliferation arrest and telomere attrition are
senescence phenotypes that may occur in replicative
or stress-induced senescence but are not specific
markers for postmitotic senescent neurons and are not
shown.p16: p16INK4a; p19: p19INK4d; p21:p21CIP1.
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Table 2 | Selected senotherapeutic molecular targets under preclinical investigation for CNS conditions
Drug target Compounds (examples) BBB penetrance Condition/indication (CNS-speciic
only) Mechanism
SRC kinase
plus PI3K Dasatinib–quercetin(ref.65)Dasatinib: yes(ref.273)
Quercetin: yes/weak
(refs.274,275)
Parkinson disease mice(ref.12)
Overtraining-induced deicits in
learning and memory(ref.276)
Age-related cognitive
decline(ref.277)
Tauopathy mice(ref.9)
AD amyloid mice(ref.45)
Senolysis: activation of apoptosis in
senescent cells
mTOR Rapamycin(ref.148)Yes/weak(ref.278)Brain health/ageing/
memory(refs.279,280)Blocks NRF2-mediated senescence
induction and reduces NF-κB-mediated
SASP gene expression
HSP90 Geldanamycin(ref.101)
Tanespimycin(ref.101)
Alvespimycin(ref.101)
Ansamycin(ref.101)
Resorcinol(ref.101)
Purine and pyrimidine-like
N-terminal HSP90
inhibitors(ref.101)
No(ref.281)
Yes(ref.282)
Yes(ref.282)
ND
ND
ND
ND
ND
ND
ND
ND
ND
Autophagy-mediated senolysis
Seno-antigens:
surface
markers of
senescent cells
uPAR targeted by CAR-T
cells(ref.169)
Antibody-based approach for
DPP4 (ref. 170)
GPNMB as immunization
antigen(ref.171)
Yes(ref.283)
No(ref.284)
No(ref.284)
ND
ND
ND
Phagocytosis-mediated clearance of
senescent cells
Na+K+-ATPase Proscillaridin A(ref.103)
Ouabain(refs.103,105)
Bufalin(refs.103,105)
Cinobufagin(ref.103)
Peruvoside(ref.103)
Digitoxin(refs.103,105)
Convallatoxin(ref.103)
Digoxin(refs.103,105)
ND
Yes(ref.285)
Yes(ref.286)
ND
ND
Weak(ref.287)
Yes(ref.288)
Weak(ref.287)
ND
TBI(ref.289)
ND
ND
ND
ND
ND
Dementia(ref.290)
Senolysis by modulating plasma
membrane potential
BCL-2 protein
family ABT-737 (ref. 107)
ABT-263 (navitoclax)
(refs.61,90,109)
A-1331852 (ref. 90)
A-1155463 (ref. 90)
UBX1325 (ref. 291)
No/weak(ref.292)
No/weak(ref.292)
Yes(ref.293)
ND
ND
COVID-19 neuropathology (ref.294)
BBB disruption(ref.295)
AD(ref.57)
COVID-19 neuropathology(ref.294)
Learning and memory in age(ref.296)
ND
ND
ND
Senolysis: deactivation of
anti-apoptotic pathway/activation of
apoptosis in senescent cells
PUMA/NOXA PCC1 upregulates
PUMA/NOXA(ref.139)ND ND Mitochondria-mediated senolysis
USP7 P5091 (ref. 121)
P22077 (ref. 121)
Modiied(ref.297)
Yes(ref.298)
Tau phosphorylation (AD
model)(ref.299)
ND
p53-mediated senolysis (elevated p53
expression)
MDM2 Nutlin3a(ref.300) and UBX1325
(ref. 223) attenuate SASP
UBX0101 (ref. 301)
RG-7112 (ref. 302)
No/weak(ref.303),
unclear
ND
Yes(ref.304)
ND
ND
ND
ND
p53-mediated senolysis (elevated p53
expression)
p53–FOXO4
interaction FOXO4-DRI (cell-penetrating
peptide that disrupts interaction
of p53 with FOXO4)(ref.120)
ND ND p53-mediated senolysis (p53 cytosolic
translocation)
HDAC Panobinostat(ref.129)Yes(ref.305)ND BCL-XL and H3-mediated senolysis
NF-κB BAY11-7082 (ref. 306)ND Aggregation of amyloid-β protein and
memory deicits(ref.307)Reduced NF-κB activity to decrease
SASP gene expression
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owing to its short half-life, or quercetin not penetrating the BBB. Two
additional open-label phase I trials testing dasatinib–quercetin in AD
(NCT04785300 and NCT05422885) will be important for providing
confirmatory safety data, and the ongoing phase II SToMP-AD study
(NCT04685590 (ref. 84)) will help to establish efficacy and timing
of senescent cell re-emergence (Table3). An interventional trial has
also been initiated using dasatinib–quercetin in treatment-resistant
depression, given the observed association between SASP and psy-
chiatric symptoms
55,56
. Collectively, the data generated in these early
trials are crucial for informing safety, dosing strategies and outcome
measures that will guide future trials85.
Other nonspecific PI3K-inhibiting flavonoids have been investi-
gated for their senolytic properties. Luteolin and curcumin have shown
weak senolytic activity at a dose at which quercetin was ineffective 86.
Fisetin, a natural compound structurally related to quercetin, is found
in various fruits, vegetables and teas, has gained considerable atten-
tion in the area of senolytic therapies
87,88
. Fisetin exhibits a diverse
pharmacological profile including antioxidant, anti-inflammatory,
antimicrobial as well as neuroprotective properties87–89. In addition
to PI3K, fisetin acts on multiple signalling pathways that have pivotal
roles in the senescence process, including BCL-2, PI3K–AKT, p53 and
NF-κB. This multifaceted action has led to the recognition of fisetin
as a senolytic compound, a discovery that emerged from systematic
screening studies involving flavonoid polyphenols and senescent
human and murine fibroblasts86,90. Notably, f isetin has demonstrated
the ability to restore tissue homeostasis, mitigate age-related patholo-
gies and extend lifespan in mice
86
. In relevance to neurodegenera-
tion, fisetin has been shown to modulate inflammatory pathways and
preserve cognitive function in AD transgenic mouse models91. It also
enhances memory and induces hippocampal long-term potentiation
in rats, suggesting its positive action in the brain and ability to ame-
liorate AD-related symptoms in vivo
92
. Fisetin is currently undergoing
clinical testing in approximately 20 clinical trials targeting a range of
age-related diseases, including MCI (NCT02741804)93.
Of note, like quercetin, fisetin has low oral bioavailability owing
to factors such as low aqueous solubility, high lipophilicity and high
first-pass metabolism
94
. To improve the bioavailability of these drugs,
strategies may include careful modification of their chemical struc-
tures to enhance their pharmacokinetic profiles while maintaining
their efficacy95, or using novel drug delivery systems96,97.
Drug target Compounds (examples) BBB penetrance Condition/indication (CNS-speciic
only) Mechanism
BRD4 ARV825 (ref. 132)Modiied(ref.308)ND PROTAC, senolysis mediated by
elevated DNA damage
IKK EF24 (curcumin
analogue)(ref.309)ND ND Reduced NF-κB activity to decrease
SASP gene expression
ATM KU-55933 suppressed
senescence and SASP(ref.143)No(ref.310)Neuroprotection against
hydrogen peroxide-induced cell
damage(ref.311)
Reduced NF-κB activity to decrease
SASP gene expression
OXR1 Piperlongumine(ref.135)Yes(ref.312)Age-related cognitive
impairment(ref.313)Induces degradation of OXR1 to induce
senolysis
Lysosome Cytotoxic drugs encapsulated
with galacto-oligosaccharides
(ref.114) e.g., Nav-Gal (ref.116),
5FURGal (ref.314), SSK1 (ref. 315)
ND ND Cytotoxic drugs are released in the
senescent lysosomes, causing senolysis
SASP SR12343 (ref. 154)
SB203580 (ref. 158)
UR-13756 (ref. 159)
BIRB 796 (ref. 159)
MW01-18-150SRM(ref.316)
Ruxolitinib (suppressor of C/EBPβ
activity)(ref.161)
Yes(ref.155)
ND
ND
ND
Yes(ref.317)
Yes(ref.318)
Improved dystonia and motor
function(ref.154)
ND
ND
ND
Neuroinlammation, psychiatric
and cognitive behaviour
(AD)(refs.316,319)
Learning and memory(ref.320)
TBI(ref.321)
Blocks NF-κB–IKK; reduces
NF-κB-mediated SASP gene expression
p38 inhibition; reduces NF-κB-mediated
SASP gene expression
Reduced C/EBPβ activity leading to
lower expression of SASP genes
BCL-XLPZ15227 (ref. 113)ND ND ABT-263-based BCL-XL PROTAC;
senolysis
cGAS Aspirin Yes(ref.322)Reduced SASP and senescence in
brain organoids(ref.182)Reduced SASP gene expression by
inhibition of COX2
STING H‐151 Yes(ref.183)Ameliorated SASP in aged
mice(ref.183) and reduced
senescence in brain
organoids(ref.182)
Reduced SASP gene expression by
reducing NF-κB activity
AD, Alzheimer disease; ATM, ataxia–telangiectasia mutated; BBB, blood–brain barrier; CAR, chimeric antigen receptor; CNS, central nervous system; COVID-19, coronavirus disease 2019;
COX2, cyclooxygenase 2; DPP4, dipeptidyl peptidase 4; FOXO4, forkhead box protein O4; GPNMB, glycoprotein nonmetastatic melanoma protein B; HDAC, histone deacetylase; HSP90,
heat shock protein 90; IKK, IκB kinase; mTOR, mechanistic target of rapamycin; ND, not determined; OXR1, oxidation resistance gene 1; PCC1, procyanidin C1; PI3K, phosphatidylinositol
3-kinase; PROTAC, proteolysis-targeting chimera; SASP; senescence-associated secretory phenotype; TBI, traumatic brain injury; uPAR, urokinase-type plasminogen activator receptor; USP7,
ubiquitin-speciic peptidase 7.
Table 2 (continued) | Selected senotherapeutic molecular targets under preclinical investigation for CNS conditions
Nature Reviews Drug Discovery | Voume 23 | November 2024 | 817–837 824
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HSP90 inhibition. The essential family of heat shock protein 90
(HSP90) is involved in multiple cellular processes and regulatory path-
ways such as apoptosis, cell cycle control, cell viability, protein folding
and degradation, and various signalling cascades98. Dysregulation
of HSP90 is implicated in diverse pathologies, including cancer and
neurodegenerative diseases such as Parkinson disease
99
and AD
100
. In a
senescence assay-based screen of compounds that regulate autophagy,
inhibitors of the HSP90 chaperone were identified to have senolytic
activity in mouse and human cells, including fibroblasts, mesenchy-
mal stem cells and umbilical vein endothelial cells101. Various HSP90
inhibitors showed dose-dependent senolytic potential, and inhibition
of HSP90 delayed the onset of age-related symptoms, extended health
span and reduced p16INK4a levels in a mouse model of a human progeroid
(premature ageing) syndrome101.
Na+K+-ATPase inhibitors. The Na+K+-ATPase pump is a crucial trans-
membrane protein that actively transports sodium and potassium
ions against their respective concentration gradients to maintain the
cellular osmotic balance and membrane potential
102
. Recent studies
have reported that senescent cells possess a marginally depolarized
plasma membrane and elevated concentrations of H+ compared with
control cells
103
. A similar aberrant membrane potential phenotype has
been described in senescent neurons10,37,104.
A high-throughput approach was used to determine the in vitro
and in vivo senolytic potential of cardiac glycosides, which inhibitthe
Na
+
K
+
-ATPase
103
. In vivo treatment with cardiac glycosides induced
the clearance of senescent tumour cells and of senescent fibroblasts
in a mouse model of lung fibrosis
103
. The authors acknowledge that the
therapeutic range for cardiac glycosides such as digoxin is relatively
narrow. However, they also emphasize that digoxin has been success-
fully used for many years, and that monoclonal antibodies capable of
reversing potential drug overdoses have been developed103. Another
cardiac glycoside, ouabain, was shown to selectively eliminate cells
undergoing oncogene-induced senescence in a mouse model of liver
cancer
105
. Both studies suggest that the senolytic effects of cardiac gly-
cosides are due to on-target inhibition of the Na
+
K
+
-ATPase, but further
research is required to explore the molecular senolytic properties of
these compounds, including for application to the CNS.
BCL-2 protein family inhibitors. The BCL-2 family has a key role in
regulation of apoptosis and has broader functions. These include
neuronal activity, autophagy, calcium regulation, mitochondrial
dynamics and energy production, as well as various other processes
essential for maintenance of the health and functionality of cells
106
.
The BCL-2 family includes both inhibitors and promoters of cell death,
collectively orchestrating the intrinsic apoptosis pathway, with mito-
chondria having a pivotal role. Senescent cells use an anti-apoptotic
programme driven by members of the BCL-2 family, including BCL-2,
BCL-XL and BCL-W, to persist within tissues and contribute to chronic
inflammation107.
Inhibition of the BCL-2 pathway can induce apoptosis in some,
but not all, senescent cell types61,107. This discover y led to the identi-
fication of BCL-2 family inhibitors as senolytic compounds, including
ABT-737, ABT-263 (navitoclax), A-1331852 and A-1155463. ABT-737,
an inhibitor of BCL-W, BCL-X
L
and BCL-2, was the first in this class
shown to specifically induce apoptosis in senescent cells through
a BCL family-dependent mechanism107. In mice, ABT-737 effectively
removed senescent cells in the lungs and epidermis, resulting in
increased proliferation of hair-follicle stem cells
107
. However, ABT-737
has limitations such as poor oral bioavailability and low aqueous solu-
bility, leading to the development of an orally bioavailable derivative,
ABT-263, as a pan-BCL inhibitor
108
. In addition to its in vitro senolytic
Box 1 | Senescent cell-targeting strategies
Senotherapeutic drug strategies broadly fall into two categories,
senolytics and senomorphics. Senolytics clear senescent cells from
the tissue, whereas senomorphics allow senescent cells to remain
but mitigate their pathogenic properties. Non-pharmacological
strategies are also gaining attention, but are beyond the scope of this
Review.
Senolytics
•Senescence-associated pathway inhibition
-SRC kinase and phosphatidylinositol 3-kinase inhibitors
-Heat shock protein 90 inhibitors
-Na+K+-ATPase inhibitors
-BCL-2 protein family inhibitors
-Ubiquitin-speciic peptidase 7 (USP7), MDM2 and p53–forkhead
box protein O4 (FOXO4) interaction inhibitors
-Histone deacetylase inhibitors
-BRD4 inhibitors
•Harnessing the immune system
-Glycoprotein nonmetastatic melanoma protein B vaccine
-Chimeric antigen receptor-T therapy (urokinase-type
plasminogen activator receptor (uPAR))
•Prodrugs
-Galactose‐modiied duocarmycin prodrugs
•Proteolysis-targeting chimeras
-PZ15227
-ARV825
Senomorphics
•Senescence-associated secretory phenotype modulation
-Rapamycin
-Metformin
-NF-κB inhibitors
-p38 inhibitors
-JAK–STAT inhibitors
-Ataxia–telangiectasia mutated (ATM) inhibitors
•Cell reprogramming
-CDGSH iron–sulfur domain 2
-G-quadruplex structures
-DNA methylation
-Mitochondrial decay
Non-pharmacological senotherapeutics
•Caloric restriction
•Intermittent fasting
•Exercise
Nature Reviews Drug Discovery | Voume 23 | November 2024 | 817–837 825
Review article
activity, ABT-263 has been shown to rejuvenate aged haematopoietic
stem cells and effectively clear senescent cells in irradiated or aged
mice
109
. Nonetheless, notable side effects, including thrombocy-
topenia and transient thrombocytopathy, have been reported for
ABT-263 (ref. 110).
Inhibiting BCL-2 family members can pose challenges in diseases
of the CNS. Treatment with ABT-737 has been found to be toxic to stem
cell-derived dopaminergic neurons in vitro
10
, likely due to the reliance
of neuronal development and survival on BCL-XL activity111. ABT-263
does not easily penetrate the BBB, hindering its ability to reach the brain
Apoptosis
Cytochrome c
Mitochondrion
Proteasome
BC L-XL
BCL- 2
BCL-W
NOXA
PUMA
ATM
MDM2
OXR1
PDK1 PI3K
STING
JAK
Ras
STAT3
STAT3
COX2
C/EBPβ
JAK
cGAS
USP7
HSP90
cGAMP
AKT
AKT
Bak
Bax
Bad
HDAC BRD4
FOXO4
p53 NF-κB
IκB
IKK
NF-κB
P
P P
P
P
Rb
SASP
Na+/K+
ATPase
mTORC1
mTORC2
uPAR, DPP4, GPNMB
RTKs, Integrins etc.
PIP3PIP2
SRC
P
P
P
E3
Ub
Ub
Ub
Ub
Lysosome
Cardiac glycosides
BC L-XL PROTAC
Immune-based approaches
SB203580
Quercetin
Rapamycin
HSP90 inhibitors
Piperlongumine
SB203580
Rapamycin
FOXO4-DRI
PCC1
P5091
UBX0101
Dasatinib
EF24
KU-55933
H-151
Aspirin
Gal-encapsulated
drugs
β-Galactosidase
Panobinostat
BCL-2 family
inhibitors
BETd
IL-6
IL-6
IL-6
SR12343
р38МАРК SB20358
Apoptosome
Fig. 3 | Potential molecular drug targets for CNS senotherapeutics. Senescent
cells feature upregulated pro-survival pathways and a senescence-associated
secretory phenotype (SASP). Several molecular players in these pathways
are under investigation as potential targets for senotherapeutic strategies.
Drug-targeting approaches to activate (red arrows), inhibit (red inhibitory lines)
or promote (dashed arrows) protein transport within the cell can result in cell
clearance (senolytics) or modulate the SASP (senomorphics). A combination of
dasatinib and quercetin, which target Src tyrosine kinase signalling, were the f irst
generation of potential senolytics to be discovered. Other promising approaches
include targeted inhibition of the pro-survival BCL-2-related pathways, or NF-κB
signalling to ameliorate the expression of SASP genes. Selected compounds
are shown as examples (red boxes); see Table2 for further details. Note
that galacto‐oligosaccharide (gal)-encapsulated drugs, which target raised
β-galactosidase levels in the lysosomes of senescent cells, might not be suitable
for central nervous system (CNS) applications owing to high β-galactosidase
expression in non-senescent neurons. COX2, cyclooxygenase 2; DPP4, dipeptidyl
peptidase 4; GPNMB, glycoprotein nonmetastatic melanoma protein B; HDAC,
histone deacetylase; HSP90, heat shock protein 90; OXR1, oxidation resistance
gene 1; RTK, receptor tyrosine kinase; Ub, ubiquitin; uPAR, urokinase-type
plasminogen activator receptor; USP7, ubiquitin-specific peptidase 7.
Nature Reviews Drug Discovery | Voume 23 | November 2024 | 817–837 826
Review article
parenchyma
112
. Nevertheless, studies in tau transgenic mice indicate that
ABT-263 reduces senescent cells in the brain and improves behaviour57.
Despite their demonstrated senolytic effects, the clinical translation
of BCL-2 family inhibitors to CNS disorders is collectively constrained
by potential side effects, off-target effects on platelets and limited CNS
penetrance. Although systemic removal of senescent cells might posi-
tively impact the brain regardless of BBB penetrance (discussed below),
future research efforts are still needed to develop more specific, safe and
brain-penetrant BCL-2 family inhibitors for use as CNS senolytic therapies.
More recently, an ABT-263-based BCL-X
L
proteolysis-targeting
chimera (PROTAC) senolytic has been developed to target BCL-X
L
to
the cereblon E3 ligase for degradation113. Additionally, to selectively
target senescent cells, researchers leveraged the elevated activity
of the lysosomal β-galactosidase that is common in senescent cells,
developing a drug delivery system based on encapsulating drugs
with galacto‐oligosaccharides114,115. In a mouse model involving
chemotherapy-induced senescence, gal-encapsulated cytotoxic drugs
effectively targeted palbociclib -induced senescent tumour cells, lead-
ing to improved regression of tumour xenografts. In another mouse
model of pulmonary fibrosis, gal-encapsulated cytotoxic drugs or
ABT-263 specifically targeted senescent cells, resulting in reduced
collagen deposition and restored pulmonary function. Notably, the
gal-encapsulation approach mitigated the toxic side effects associated
with senolytic drugs
114
. Furthermore, Nav-Gal, engineered by linking
the BCL-2 family inhibitor ABT-263 to an acetylated galactose group,
selectively eliminated senescent cells both in vitro and in vivo, demon-
strating reduced platelet toxicity and ameliorating the common issue
of thrombocytopenia associated with ABT-263 treatment
116
. Although
this approach is promising, targeting β-galactosidase for CNS condi-
tions may not be appropriate as many non-senescent neuronal cell
populations express high levels of β-galactosidase activity9,117 (Fig.2),
as discussed further below.
USP7, MDM2 and p53–FOXO4 interaction inhibitors. Modulators of
the p53 pathway have gained attention as potential senolytic agents
owing to the role of p53 in controlling senescence and apoptosis, par-
ticularly through its regulation of BCL-2 family proteins
118,119
. One note-
worthy example is forkhead box protein O4 (FOXO4), an anti-apoptotic
transcription factor that is upregulated in some senescent cell types.
FOXO4 prevents cell death by binding to and sequestering p53 within
the nucleus
120
. To target this interaction, a senolytic peptide named
FOXO4--Retro-Inverso (FOXO4-DRI) was developed. FOXO4-DRI
disrupts the binding between FOXO4 and p53, which allows p53 to
translocate to the cytosol and induce apoptosis in senescent cells.
FOXO4-DRI has demonstrated selective removal of senescent cells and
has shown promise in alleviating age-related symptoms across organ
systems in animal models120.
p53 protein levels are tightly regulated through ubiquitylation
by the E3 ubiquitin ligase murine double minute 2 (MDM2). Inhibitors
that disrupt the MDM2–p53 interaction, such as UBX0101 and RG7112
(RO5045337), as well as inhibitors of the de-ubiquitinating enzyme
ubiquitin-specific peptidase 7 (USP7), serve to increase and stabilize
p53 levels. RG7112, UBX0101 and USP7 inhibitors, such as P5091 and
P22077, cleared various peripheral senescent cells by upregulating
p53 (ref. 121).
Moving into clinical applications, however, has faced challenges.
Although UBX0101 has been shown to induce apoptosis of senescent
synoviocytes, and a phase I study (NCT03513016) suggested that it has
clinically meaningful effects on pain and function in patients with knee
osteoarthritis, preliminary reports from the follow-up phase II trial
of UBX0101 for osteoarthritis suggest that it has not demonstrated
significant efficacy compared with placebo122. The authors speculated
that possible reasons for this could include the dosing route, patient
and/or investigator expectedness, and gender dimorphism in pain
reporting. Manipulating p53 levels in senescence regulation requires
careful consideration, as p53 can have both promoting and inhibitory
effects depending on cell type and stress levels. Adding complexity
to the therapeutic approach, p53 activity can induce p21-dependent
senescence. Additionally, some p53-targeting agents are peptide
based, which may limit their applicability to address brain senescence.
Table 3 | Selected senotherapeutics under clinical investigation for central nervous system conditions
Compounds Targets Indication Sponsor Study phase/type NCT identiier
Dasatinib–quercetin
combination therapy Primary target
Dasatinib: SRC kinase(ref.65)
Quercetin: PI3K(ref.65)
MCI with slow gait speed Hebrew Senior Life Phase I/II open label NCT05422885
Other drug targets
Dasatinib: ABL kinases(ref.323)
(ABL1, BCR–ABL); receptor
tyrosine kinases(ref.324) (EGFR,
EPHA2, DDR1); non-receptor
tyrosine kinases(ref.324)
(FRK, BRK, ACK), p90RSK (ref. 324)
Quercetin: suppression
of COX2, NF-κB and AP-1
pathways(ref.325), nonspeciic,
multiple targets(ref.326)
MCI/early AD University of Texas He alth
Science Center at San Antonio Phase I/II POC open
label NCT04063124
MCI/early AD Wake Forest University Health
Sciences Phase II RCT NCT04685590
MCI/early AD Mayo Clinic Phase I/II open label NCT04785300
Older adults with
depression or
schizophrenia
Washington University School
of Medicine Phase II RCT NCT05838560
UBX1325 BCL-XLDME or neovascular AMD Unity Biotechnology Phase I open label NCT04537884
Wet AMD Unity Biotechnology Phase II RCT NCT05275205
DME Unity Biotechnology Phase IIb NCT06011798
Non-pharmacological trials or those using compounds with senotherapeutic potential (e.g., polyphenols, rapamycin, etc.) but not in a senescence indication are not included. AD, Alzheimer
disease; AMD, age-related macular degeneration; DME, diabetic macular oedema; MCI, mild cognitive impairment; PI3K, phosphatidylinositol 3-kinase; POC, proof-of-concept; RCT,
randomized controlled trial.
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As mentioned above, senescent cells have been reported in Parkinson
disease post-mortem human brain tissue and model organisms
10,12,47,123
.
Elevated p53 levels and activity were observed in the brains of patients
with Parkinson disease, as well as in Parkinson disease animal and cel-
lular models, mostly correlating with DNA damage124. p53-dependent
brain senescence has also been reported in Huntington disease and
AD
124
, suggesting that p53 may be an especially appealing target for
CNS senescence. Recently, it was reported that a ketogenic diet induced
p53-dependent cellular senescence in multiple tissues, including the
brain
125
. These findings may have serious implications for humans
on the ketogenic diet; however, we note that immunoblotting with
an antibody to β-galactosidase was the primary readout, which may
reflect changes in lysosomal function rather than senescence. Further
research is needed to unravel the complicated molecular mechanisms
by which p53 regulates senescence, with potential implications for
clinical applications.
HDAC inhibitors. Histone deacetylases (HDACs) modulate gene expres-
sion by removing acetyl groups at the lysine residues of histones and
non-histone proteins. They impact many cellular functions including
differentiation, proliferation and survival126. Senescent cells show
changes in H3 acetylation and BCL-X
L
expression
107,127,128
. In vitro, the
FDA-approved HDAC inhibitor panobinostat cleared cancer cells with
chemotherapy-induced senescence. This activity was associated with
decreased BCL-XL expression and increased H3 acetylation129.
BRD4 inhibitors. Bromodomain and extra-terminal domain (BET)
family proteins act as transcriptional regulators
130
. The family member
BRD4 is well-characterized for its role in transcriptional elongation
131
.
In a high-throughput screen of compounds alongside bio-functional
analysis, the BET family protein degrader (BETd) was identified as
senolytic
132
. The authors showed that inhibition of the BET family pro-
tein BRD4, either with chemical inhibitors or gene-specific RNA inter-
ference, induced senolysis. The hetero-bifunctional PROTAC ARV825
proved particularly effective, as it recruits BET family proteins to the E3
ubiquitin ligase cereblon, leading to their degradation
133
. The senolytic
effects of ARV825 were mediated by attenuating non-homologous end
joining (NHEJ) and enhancing autophagic gene expression. Specifically,
the authors noted that ARV825 induces senolysis by exacerbating DNA
double-strand breaks through targeting at least two independent
mechanisms of the NHEJ machinery in senescent cells: first, inhibition
of gene expression of XRCC4, which encodes a protein required for NHEJ
repair; and secondly, blockade of the recruitment of 53BP1, an adaptor
protein necessary for the assembly and activation of the DNA repair
machinery, to double-strand break sites
132
. Treatment with ARV825 was
sufficient to eliminate senescent cells by targeting BRD4, as confirmed
by small interfering RNA (siRNA)-based gene-specific experiments,
which showed that ARV825 reduced levels of BRD3 and BRD4, but not
BRD2. Furthermore, the compound showed senolytic activity in the
livers of obese mice and cleared senescent hepatic stellate cells, as well
as chemotherapy-induced senescent cells in mice132.
OXR1 inhibitors. Oxidation resistance gene 1 (OXR1) acts as a sensor
and regulator of cellular oxidative stress. It is involved in the tran-
scriptional networks needed to detoxify reactive oxygen species and
modulate cell cycle and apoptosis
134
and is upregulated in senescent
human fibroblasts
135
. Interestingly, OXR1 is the target of the previously
identified senolytic piperlongumine, a natural compound found in long
pepper
135,136
. Piperlongumine was identified by screening a library of
rationally selected compounds for their senolytic activity in human
senescent fibroblasts136. The initial study reported that piperlongumine
kills senescent fibroblasts without the induction of reactive oxygen
species; however, it was later demonstrated that piperlongumine binds
to OXR1, leading to its degradation via the ubiquitin–proteasome sys-
tem specifically within senescent cells. The reduction of OXR1 induces
apoptosis exclusively in senescent cells, likely by increasing their
vulnerability to oxidative stress
135
. In a recent computational study
investigating natural senotherapeutics to find candidates that could
replace dasatinib, based on their similarity in gene expression effects,
piperlongumine was identified as the highest-potential substitute,
suggesting that further development of the compound is warranted
137
.
PUMA/NOXA activators. PUMA and NOXA are pro-apoptotic genes
that are functionally repressed by BCL-2 family molecules in senescent
cells138. Through screening natural products, procyanidin C1 (PCC1),
a polyphenolic component of grape seed extract, was identified as
being capable of ameliorating the SASP at low concentrations and
exhibiting senolytic effects at higher concentrations
139
. It was hypoth-
esized that PCC1-induced apoptosis in senescent cells is partially medi-
ated by NOXA and PUMA, accompanied by increased reactive oxygen
species production and mitochondrial dysfunction139. Additionally,
PCC1 ameliorated physical dysfunction and prolonged survival in mice.
Senomorphics
Senomorphics, also known as senostatics, mitigate the toxicity of senes-
cent cells without removing them from the tissue, with the ultimate
goal of retaining the cells’ initial physiological functions (Table 2 and
Fig.3). This section focuses on the molecular pathways targeted in
senomorphic drug development.
ATM inhibitors. The ataxia–telangiectasia mutated (ATM) kinase is
crucial for DNA damage response signalling and is known to activate
NF-κB in response to stress. NF-κB is a transcription factor involved
in the regulation of the immune response, inflammation, cell sur-
vival, cellular senescence and organismal ageing140. ATM kinase medi-
ates the equilibrium between senescence and apoptosis. Active ATM
promotes autophagy, with a specific emphasis on sustaining the
lysosome–mitochondrion connection, boosting senescence while
restraining apoptosis141. Disruptions in autophagy mechanisms,
however, have been demonstrated to elevate DNA damage levels,
thereby promoting the development of cancer and neurodegenera-
tive diseases
142
. Activated ATM has been reported in senescent cells,
and genetic and pharmacological (using KU-55933) inhibition of ATM
resulted in a reduction in NF-κB activity, markers of senescence and
the SASP in cells as well as in mouse models of ageing
143
. This treat-
ment also extended the health span ofprogeroid mice, indicating that
ameliorating ATM-mediated NF-κB activity can slow the progression
of ageing143.
mTOR inhibitors. The PI3K-related kinase mechanistic target of rapa-
mycin (mTOR) regulates numerous cellular pathways including apop-
tosis, growth and autophagy
144,145
. mTOR inhibition has emerged as
an avenue to improve longevity and ameliorate the impact of cellular
senescence. The mTOR inhibitor rapamycin, an FDA-approved immu-
nosuppressive drug, has repeatedly proved its ability to extend lifespan
in various organisms, ranging from yeast to mice
146,147
. Rapamycin pro-
motes autophagy, a cellular process that has a crucial role in recycling
damaged components and maintaining cellular health. Although not a
Nature Reviews Drug Discovery | Voume 23 | November 2024 | 817–837 828
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senolytic, rapamycin has gained attention for its potential to suppress
the SASP
148
. Although rapamycin holds promise for extending lifespan
and curbing the SASP, clinical applications have encountered chal-
lenges, including metabolic dysregulation; impaired wound healing,
a process in which senescent cells appear to have a beneficial role149;
and the development of hyperlipidaemia150. Despite these challenges,
the positive outcomes observed in various animal models of ageing
and neurodegeneration have propelled rapamycin into clinical trials to
assess its potential in treatment of AD (NCT04200911, NCT04629495),
although it is not known whether senescence-associated outcomes
will be assessed.
NF-κB inhibition and SASP modulation. The transcription factor
NF-κB has a dual role, acting as both a guardian against and a potential
contributor to neuroinflammation and the accumulation of senescent
cells in the brain151. It mediates neuroinflammation by enhancing the
expression of pro-inflammatory molecules, many of which are part of
the SASP, such as inducible NO synthase (iNOS), cyclooxygenase 2, TNF
and interleukins (for example, IL-1 and IL-6). Under normal conditions,
the activity of NF-κB is restrained by inhibitors known as IκB proteins.
However, in response to stress or inflammation, IκB inhibitors are
phosphorylated by the IκB kinase (IKK) complex, unleashing NF-κB
and triggering a cascade of immune responses152. Studies have shown
that enhanced NF-κB activity can lead to neuroinflammation and an
increased burden of senescent cells in the brain151. Conversely, inhibi-
tion of the binding between NF-κB and IKK, using a peptide inhibitor,
ameliorated senescence in progeroid mice1 53. Building on this, research-
ers developed a small molecule, SR12343, which disrupts the interac-
tion between NF-κB and IKK. This molecule significantly reduced the
release of SASP factors, alleviated senescence and improved markers
of senescence in mice154. Importantly, SR12343 can traverse the BBB,
making it a compelling candidate for evaluation in the context of neu-
rodegenerative conditions
155
. In a recent study using SR12343, it was
found to lower markers of cellular senescence and inflammation in the
liver, skeletal muscle and blood, which were elevated in response to
chemotherapy. Additionally, it was reported that SR12343 effectively
counteracted chemotherapy-induced skeletal muscle wasting and
dysfunction156.
The p38 pathways regulates the transcriptional activity of NF-κB1 57.
Consequently, inhibiting the p38 pathway has emerged as a promis-
ing avenue to address the detrimental effects of the SA SP and reduce
age-related inflammation, particularly in the context of neurodegener-
ative conditions. The p38 inhibitor SB203580 has been shown to reduce
the expression of SASP factors in human senescent cells158. Similarly,
next-generation p38 inhibitors, such as UR-13756 and BIRB 796, have
demonstrated promising results in limiting the expression of IL-6.
By reducing the p38-dependent transcriptional activity of NF-κB on
pro-inflammatory genes, these inhibitors mitigate SASP-related effects,
marking them as potential therapeutic candidates for age-related
inflammatory conditions159.
The regulation of the SASP is complex. Recent findings indicate
that the immune gene transcriptional effector C/EBPβ cooperates with
NF-κB to activate SASP genes
160
. JAK–STAT signalling is involved in the
regulation of C/EBPβ and vice versa. Inhibition of the JAK–STAT signal -
ling pathway by ruxolitinib suppresses the transcriptional activity of
C/EBPβ. This suppression led to a reduction in systemic inflammation,
attributed to SASP repression, and an improvement in the overall fit-
ness of aged (24-month-old) mice
161
. These findings underscore the
potential of modulating stress kinase p38 as a senomorphic approach
to combat neuroinflammation, a common feature of neurodegen-
erative disorders. To tackle neuroinflammation in neurodegenerative
diseases, a brain-permeable and orally available p38 inhibitor, MW1 50
(also known as MW01-18-150SRM), is currently in phase II trials for AD
(NCT05194163).
Although strategies to reduce NF-κB activity hold promise in the
realm of senomorphic therapies, inhibition may lead to side effects
owing to NF-κB involvement in diverse biological processes. An alter-
native approach to reducing inflammation, perhaps with a better
safety profile, involves the use of the nonsteroidal anti-inflammatory
drug, ibuprofen. This strategy has been shown to mitigate senescent
cell accumulation, neuroinflammation and cognitive dysfunction in
mice
151
. However, clinical trials using ibuprofen as monotherapy for AD
have not significantly impacted disease progression or outcomes
162,163
,
but perhaps could be a useful combination therapy adjunct.
Harnessing the immune system
A healthy immune system actively clears senescent cells and prevents
their accumulation. Therapeutic approaches to boost senescent cell
clearance by the immune system are under investigation. These include
enhancing the recognition of senescent cells by the immune system,
promoting immune cell activity or modulating immune responses
to improve the clearance of senescent cells from tissues. Generally,
modulating the immune system for senolysis represents a promising
approach.
The body’s natural defence against senescent cells involves
immune surveillance mechanisms, with immune cells such
as macrophages, T cells, natural killer cells (NK cells)164,165 and
microglia in the brain44,166 tasked with removing these cells. However,
age-relatedimmunosenescence and the ability of senescent cells to
evade immune detection — for example, by expressing immunosup-
pressive molecules such as PDL1 and PDL2, by producing tolerogenic
major histocompatibility complex class I variants and by secreting
factors that attract immunosuppressive cells
167
— can lead to their
accumulation in tissues168.
Notably, some senescent cells upregulate specific surface markers
that are being explored as therapeutic targets. Urokinase-type plasmi-
nogen activator receptor (uPAR) has been shown to be upregulated in
several senescent cells, including mouse lung adenocarcinoma cells
induced to senesce by MEK and CDK4/6 inhibition, oncogene-induced
senescence in mouse hepatocytes and culture-induced senescence
in mouse hepatic stellate cells
169
. Mass spectrometry analyses com-
paring proliferating human fibroblasts with those undergoing rep-
licative senescence revealed an increase in the surface molecule
dipeptidyl peptidase 4 (DPP4; also known as CD26) in senescent cells
170
.
Transcriptome data from senescent vascular endothelial cells indicated
an enrichment of the transmembrane glycoprotein nonmetastatic
melanoma protein B (GPNMB)
171
. Collectively, uPAR, DPP4 and GPNMB
were identified as ‘seno-antigens’ and targeted for immune-based
interventions as described below.
To bolster immune clearance of senescent cells, chimeric anti-
gen receptor-T (CAR-T) cells have been used. In this approach, T cells
from patients are genetically engineered to target and eliminate
cells expressing specific antigens. Utilizing uPAR as a target antigen,
CAR-T cells have been shown to effectively clear senescent cells, miti-
gating various pathologies in mouse models of senescent lung adeno-
carcinoma and liver fibrosis
169
. Similarly, antibody-based therapies
targeting the senescent cell surface protein DPP4 have demonstrated
efficacy in eliminating senescent cells in vitro170.
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A vaccine-like protocol aimed at senolysis has emerged as another
possible approach to mitigate senescent cell burden. Immunizing
mice against GPNMB, which is elevated in senescent vascular endothe-
lial cells and in samples from patients with atherosclerosis, reduced
GPNMB-positive cells, improved metabolic parameters in mice fed a
high-fat diet, reduced atherosclerotic plaque burden in Apoe-knockout
mice, improved age-related phenotypes and ultimately extended the
lifespan of progeroid mice
171
. Developing immune- and vaccine-based
approaches to eliminate senescent cells is intriguing, but identifying
senescence-specific antigens is crucial to avoid potential autoimmunity
against non-senescent cells. To date, none of the senescence-associated
surface markers seems truly exclusive for senescent cells, as GPNMB is
expressed across tissue and cell types and has diverse roles in inflam-
mation and disease172,173; DPP4 is known to be ubiquitously expressed
and involved in immunoregulatory actions174; and uPAR is present on
many immune cells and is detected in various bodily fluids including
plasma and seminal fluid, and in extracellular matrix175.
In addition to the challenge of identifying markers exclusively
expressed on senescent cells, targeting CNS senescence through
immune modulation faces other challenges. Immune cell traffick-
ing into the CNS is tightly regulated by the BBB, permitting entry
only for specific immune cell subsets176. Additionally, immune-based
approaches trigger an inflammatory response, which could be prob-
lematic in the context of neurodegenerative diseases, where neuro-
inflammation is present. Therefore, while harnessing the immune
system holds great potential for senescent cell clearance, the impact
of on-target, off-tissue effects and unintended immune responses must
be carefully evaluated to minimize potential harm.
Future perspective
Many ‘first-generation’ senolytic drugs such as quercetin, fisetin and
piperlongumine, lack a single precise target and instead affect multiple
pathways that are implicated in cellular ageing. These broad-spectrum
senolytics have the capacity to impact multiple senescence pathways
simultaneously177; however, they present a challenge in predicting
efficacy owing to the considerable heterogeneity between senescent
cell populations. Drug discovery programmes based on identifying
molecules that are exclusively upregulated in senescent cells offer
opportunities for more precisely targeted senolytic therapeutics.
Other exciting senotherapeutic approaches, currently in preclinical
stages, include cellular reprogramming
178
; targeting mitochondrial
function
179–181
; targeting the cGAS–STING pathway
182,183
; and various
genome-targeting approaches including G-quadruplex structures18 4,
DNA methylation
185
and retrotransposon activity
186
. These diverse and
innovative approaches reflect a growing enthusiasm for senotherapeu-
tics and underscore the need for continued research to fully realize
their potential for CNS conditions.
Challenges in developing CNS senotherapies
Senotherapeutic drug development faces several challenges, some
of which are common to CNS drug development more broadly and
contribute to the long development timelines of such therapeutics
187
.
Here, we consider challenges that are particularly important in the
development of CNS senotherapeutic drugs and discuss strategies
to address them.
Senescence-specific targeting
The need to achieve selectivity for therapeutic targets and specific cells,
while sparing healthy cells, to achieve a favourable safety profile is a
common challenge for drug discovery. Senotherapeutics that impact
cellular mechanisms that are vital for neuronal viability and func-
tion, such as Na+K+-ATPase inhibitors188, BCL-2 inhibitors189 and NF-κB
inhibitors
190
, pose a particularly substantial risk. Understanding cellular
senescence in various disease states and identifying relevant cell types
are crucial to identify and select optimal senotherapeutic targets
191
.
Drug development, and most senescent cell identification, occurs in
laboratory models. Discerning the differences between human dis-
ease and experimental models is a crucial step in safely and effectively
translating senotherapies to the clinic. We anticipate that technological
advances in single-cell and spatialmulti-omics, along with advanced
computational and AI approaches capable of analysing the vast data-
sets, will enable the identification of tractable and specific targets for
senotherapeutic interventions28.
New drug modalities in development could help to increase speci-
ficity for therapeutic targeting. For example, PROTACs, as mentioned
earlier, are an innovative mechanism that capitalizes on the cell’s
ubiquitin–proteasome system
192
. They hold promise for specificity,
reduced side effects and the ability to hit targets previously deemed
‘undruggable’. However, their size presents a significant hurdle to
optimization of CNS delivery, necessitating research efforts to improve
their design for such applications. Beyond modalities that directly
engage the target proteins, others such as gene therapy and RNA-based
therapies offer versatile approaches for gene silencing or functional
restoration and are in early stages of development193.
BBB penetrance
Favourable properties of molecules that target the CNS include small
size, hydrophobicity optimal for penetrating the BBB without becom-
ing trapped in the lipid bilayer or losing blood solubility, and low affinity
for the P-glycoprotein transporter
194
. Peptides and antibodies offer
higher specificity compared with small molecules, but they have low
BBB penetrance and face challenges related to stability, solubility and
incompatibility with oral administration. Advances in engineering
antibodies and designing peptides that are more resistant to degrada-
tion, along with new delivery systems and BBB-crossing strategies, such
as tagging with molecules that allow uptake via receptor-mediated
transcytosis (for example, TfR
195
and CD98hc
196
) are making them more
viable for CNS applications
197
, although this is a complex and ongoing
challenge. Even when drugs penetrate the brain, determining the cor-
rect dosage is challenging because drug metabolism in the brain can
differ from that in other organs198,199.
Notably, the cells that comprise the BBB are susceptible to becom-
ing senescent. Vascular smooth muscle cells were identified as the
most prominent senescent cell type across 50 healthy human tissues
31
.
A separate study identified a significant increase in senescence gene
expression in vascular cells isolated from post-mortem human brains
with advanced Braak stages of AD compared with control cases
200
.
These findings suggest that senotherapies may benefit the brain by
targeting vascular cells, potentially without the need for BBB pen-
etration or direct brain exposure. Although BBB penetrance is a key
consideration for CNS drug development, there is some evidence
that clearing senescent cells in peripheral tissues may benefit brain
function. For example, ABT-263, which does not easily penetrate the
BBB, has shown benefits to the CNS following whole-body irradiation
201
in mouse models that develop AD neuropathology
57,202
and in glio-
blastoma models203,204. Although the senescence-inducing insults
(for example, radiation, toxic tau and amyloid-β, or cancer, respec-
tively) may have allowed ABT-263 to penetrate the BBB, it is also possible
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that the benefits were an indirect result of clearance of senescent
peripheral cells, which have been shown to impact senescent cell
accumulation in the brain
205
. Thus, senolytic therapies may improve
brain function by clearing senescent immune cells and cells of brain
vasculature without a need to fully penetrate the parenchyma.
Although the ‘hit-and-run’ strategy of intermittent dosing has
been used to minimize potential adverse events owing to prolonged
systemic levels of senolytics, an alternative approach is local deliv-
ery. This strategy was used in a recent open-label phase I trial for
UBX1325 (foselutoclax), a senolytic small-molecule inhibitor of BCL-X
L
,
in patients with advanced diabetic macular oedema (DME)
206
. In the
study, UBX1325 was administered via intravitreal injection, keeping
plasma levels of UBX1325 and its active parent molecule below the lower
limit of quantification and minimizing adverse events linked to sys-
temic BCL-X
L
inhibition. Visual acuity improvements were obser ved in
six of eight patients at 12 weeks, and in five of eight patients at 24 weeks.
The favourable safety profile supports the ongoing phase IIb study
efforts (Table3).
Risks of neuronal senolysis
The question of whether postmitotic neurons that have entered a
senescence-like state should be eliminated requires careful consid-
eration. The therapeutic elimination of neurons is not unprecedented,
with the best-known example of this being the surgical removal of small
volumes of brain tissue to treat drug-resistant epilepsy
207
. Evidence
to support the clearance of senescent neurons includes preclinical
studies in tauopathy mice that demonstrate benefits from the clear-
ance of NFT-bearing neurons
9
. Neurons with NFTs maintain the ability
to respond to inputs and remain active in vivo208 but contribute to
pathogenic tau spread in an activity-dependent manner209. Similarly,
senescent neurons maintain electrical activity, but with altered func-
tion. For example, inducing a senescence phenotype in excitatory neu-
rons by deleting Gdf11 causes neuronal hyperexcitability with reduced
synaptic inputs in vivo
104
. A similar hyperexcitable phenotype with
decreased synaptic protein expression was observed in an in vitro
model of neuronal senescence generated by transfecting iNeurons with
CDKN2A208. Removing SATB1 in human dopaminergic neurons drives
a senescence-like phenotype in vitro, whereby the cells maintain
spontaneous pacemaking activity but show significant differences in
maintenance of response to positive current injections
10
. Regulated
neuronal activity is crucial for proper brain function, warranting a
better understanding of the consequences of altered excitability in
senescent neurons.
Senescent cells induce paracrine senescence in neighbouring cells
through their SASP. Neuronal SASP may contain toxic aggregate-prone
proteins47, which could facilitate senescence spread between dis-
tant regions particularly via well-connected, anatomically linked
neurons210. A link between intraneuronal tau19,98 and α-synuclein
deposition
123
and senescence has been established. Given that neu-
rons increase the release of α-synuclein after becoming senescent47,
it is tempting to speculate that neuronal senescence is a key process
mediating the transfer of aggregate-prone proteins between neurons
and across circuits. Senescent neurons also spread senescence to
non-neuronal cells including to astrocytes through traditional SASP
mechanisms208 and to microglia through phagoptosis44,166. Specifically,
neurons that contain NFTs expose phosphatidylserine on their cell
membranes, which signals microglial phagocytosis
166
. Upon engulfing
the NFT-bearing neuron, the microglia become senescent, resulting
in their decreased tissue surveillance and their release of partially
digested NFT toxic fragments into their environment, which ampli-
fies pathogenesis44 (Fig.1). In summary, the connectivity of senescent
neurons within complex neuronal circuits and their close interaction
with glia cells may allow them to propagate neuropathology to mul-
tiple brain cell types, which provides rationale in favour of their clear-
ance. However, in some neurodegenerative models associated with
aberrant microglial activity, positive outcomes have been seen from
retaining damaged neurons by interrupting phagoptosis211–214. These
studies highlight the nuanced, and important, choice of senotherapy
approach (senolytic versus senomorphic) depending on context and
condition, especially when targeting cells and tissues with limited
regenerative capacity.
Measuring CNS target engagement
Target engagement, which measures the interaction of a compound with
its intended molecular target within a living system, provides insights
into the drug’s pharmacological profile, mechanism of action and poten-
tial efficacy. Evaluation of precise target engagement in preclinical
senolytic studies often involves histological analysis, as well as protein
and gene expression measurements on mouse brain tissue9,45,57. These
methods offer detailed insights into the cellular and tissue distribution
of senolytic effects. However, assessment of target engagement in liv-
ing humans requires in vivo imaging strategies or surrogate measures
in accessible biofluids.
MRI provides information on the effects of a drug on brain mor-
phology and physiology
215
and has been used to assess the impact
of senolytics in a tauopathy mouse model
9
. Although MRI cannot
provide direct evidence for the action of senotherapeuticson
pro-survival or SASPpathways within senescent cells, it allows
for the longitudinal tracking of disease progression in the brain,
providing a surrogate marker of drug effects in the target tissue.
Positron emission tomography (PET) tracks the accumulation
of radiolabelled drugs in the body216, with ongoing development of
senescence-specific PET tracers focusing on labelling lipofuscin
217
or β-galactosidase218, which appear when mitotically competent
cells enter senescence. However, these tracers may have limited
applicability to cerebral tissues as lipofuscin deposition and elevated
β-galactosidase activity can also be detected in healthy CNS tissue
throughout the lifespan9,117,219 (Fig. 2).
Other strategies for identifying surrogate markers of target
engagement include characterization of the surface proteome of
senescent cells to discover seno-antigen-specific proteins unique to
these cell populations
220,221
, which can then be visualized using clinical
imaging techniques, such as immuno-PET222. A recent report suggests
that senescent cells produce uniqueoxylipins that are released upon
their death, raising the possibility of developing senolysis-specific
biomarkers223. Although the identification of PET tracers specific
to brain senescent cells could represent a significant advance, it
remains unclear whether the small percentage (2%) of sporadically
dispersed senescent cells19 would produce a strong enough signal to
be detectable in PET imaging.
The limitations with imaging senescent cells in the brain have
increased the reliance on biofluid analytes as surrogate markers of
target engagement. Mouse studies suggest that senescent T cell levels
in blood could serve as a potential surrogate for senescent burden in
solid tissues, including the brain
205
. In this regard, p16
INK4a
(p16) mRNA
expression in peripheral blood T lymphocytes has emerged as a quan-
tifiable measure of total senescent cell load
224
and is being used in
clinical trials
62,225,226
, including an ongoing phase II trial for AD
84
. Other
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Review article
commonly used surrogate markers for senescent cells include cir-
culating SASP factors
227
such as cytokines, chemokines, proteases
and growth factors82,83. Similar to measuring a SASP index, as men
-
tioned earlier, the first phase I clinical trial to test senolytics in older
adults with MCI or early AD, used a SASP cytokine panel to evaluate
senescence-associated markers of inflammation in the blood and
CSF33. However, the challenge with using SASP factors is that they can
be secreted for reasons other than cellular senescence, complicating
the interpretation of outcomes. Ongoing efforts to understand how the
secretedmilieu of senescent cells varies by cell type or context will help
to clarify the utility of these promising biomarkers
228,229
. Moreover,
results from placebo-controlled trials, such as the phase II SToMP-AD
study, are needed to interpret how SASP readouts relate to senolytic
effects84.
Methodological advances with omics technologies are paving the
way for a deeper understanding of molecular changes associated with
CNS diseases230. These approaches may allow for the identification
of rare and distinct senescent cell populations and omics changes in
response to senolytic therapies. For example, whereas traditional SASP
protein markers did not change significantly in the first phase I trial for
AD33, lipidomics and transcriptomics analyses revealed analytes that
may be more sensitive to senolytic treatment85. While we are cautious
not to overinterpret the results from the open-label trial, especially
given the small sample size, the approach underscores the ongoing
efforts to identify biomarkers specific to senescent cells and their
clearance.
Applying AI to senotherapy development
The complex and heterogeneous phenotypes of senescent cells
requires innovative approaches in developing effective senothera
-
peutics. Advances in experimental platforms and omics technologies,
coupled with AI and ML, enable the extraction of relevant information
for chemical activity and functional genomics in drug discovery231.
AI refers to the simulation of human intelligence in machines for
performing tasks, making decisions and adapting by learning from
their experiences. ML, a specific AI technique, involves training
computer algorithms to learn from data and take actions based
on the learned knowledge231. Predictive modelling using ML algo-
rithms expedites decision-making
232
, identifies potential drug can-
didates with a higher likelihood of success, and reduces costs and
development time233,234.
In this section, we briefly review AI and ML applications in key steps
of senotherapeutic development, including identification of senescent
cells, identification of senescence targets, assessment of druggability
and virtual screening compounds, and highlight major challenges
in these areas. Overall, the application of AI and ML in senescence
research is still in its early stages but holds potential to accelerate drug
development in this area.
Identifying senescent cells
Senotherapeutic drug development requires accurate identification
of senescent cells. ML approaches can be used to classify and charac-
terize senescent cells using morphological and omics (transcriptom-
ics, proteomics and methylomics) data
235
. For instance, ML methods
such as multivariate regression analysis
236,237
and k-means clustering
238
have been used to identify senescent cells using bulk transcriptomic
data. In multivariate regression analysis, the ‘known’ senescent genes
were treated as variates. For single-cell RNA-sequencing data, Teo
et al.
32
used pseudotime trajectory analysis to identify senescent cell
subpopulations. This is an especially powerful approach as it captures
cells and cell functions at various stages of senescence progression.
Traditional methodologies focus primarily on the end point senes-
cent state at which lysosomal function and cytoskeletal abnormali-
ties are enriched. However, trajector y analyses suggest that cellular
respiration and active transportation are transiently elevated in the
middle trajectories, which could be interesting targets to explore as
their modulation could potentially prevent cells from entering the
terminal senescence state.
Others are investigating morphology-based approaches to iden-
tify and classify senescent cells. A convolutional neural network (CNN)
system, Deep Learning-Based Senescence Scoring System by Morphol-
ogy (Deep-SeSMo), was trained on phase-contrast images of human
umbilical vein endothelial cells treated with senescence-inducing
agents239. Thousands of single cells were given a senescence probabil-
ity score, predicted as senescent or control, and then compared with
predetermined answers. Weighted, automatic and iterative optimiza-
tion was used to improve the accuracy of the system, resulting in an
accuracy of 0.93 and area under the curve of the receiver operating
characteristic of 0.98.
Glossary
Cell cycle arrest
The interruption or halting of the cell
cycle at speciic checkpoints to prevent
the completion of cell division.
Immunosenescence
The ageing-related decline in eicacy
of the adaptive and innate immune
systems, leading to increased
susceptibility to infections, reduced
vaccine eicacy, less eicient clearing
of senescent cells and a higher
incidence of age-related diseases.
Oxylipin
A large class of bioactive lipid
metabolites derived from the oxidation
of polyunsaturated fatty acids.
Phagoptosis
A form of cell death characterized
by the removal of living cells by
phagocytic cells, such as macrophages
and microglia.
Progeroid mice
Genetically engineered or naturally
occurring mouse models that exhibit
accelerated ageing processes similar
to those seen in human progeroid
syndromes.
PROTACsenolytics
Proteolysis-targeting chimeras that
enable the targeted degradation of
speciic pro-survival proteins within
senescent cells.
SASP index
A quantitative composite score of
cellular senescence that measures the
expression of senescence-associated
secretory phenotype (SASP)
components using techniques such
as quantitative PCR, enzyme-linked
immunosorbent assay or mass
spectrometry.
Senescence-associated
secretory phenotype
(SASP). The complex molecular milieu
secreted by senescent cells aects
neighbouring cells through extracellular
matrix remodelling, chronic
inlammation, attracting immune cells
and altering function. SASP factors
include cytokines, chemokines, growth
factors, proteases, bioactive lipids,
exosomes and free non-coding
nucleic acids.
Senescence-messaging
secretome
(SMS). Secreted proteins that contribute
to the induction and maintenance of
senescence; later updated to relect
a greater variety of molecules and
renamed the senescence-associated
secretory phenotype.
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Review article
Although these approaches hold great potential, limitations
include a lack of rigorous wet-lab experiments to confirm the infor-
matic predictions. For example, studies with cell culture validation data
commonly rely on SA-β-gal as a gold-standard marker for senescence,
which may be misleading, especially for brain cells.
Predicting novel senescence-associated genes
Intersecting cancer driver genes and ageing-associated genes iden-
tified dual-purpose targets for the treatment of cancer and ageing
and revealed that senescence pathways were upregulated in one
subgroup
240
. A comprehensive transcriptomic network analysis of
human tissue-specific RNA-sequencing data from the Genotype-Tissue
Expression Project (GTEx) identified co-expressed gene modules
enriched for senescence-associated genes31. This approach revealed
51 senescence-associated genes conserved across all human tissues
and identified regulators centred around CDKN1A (p21)31. Although p21
is not a novel senescence-associated molecule, the study represents
a first effort in systematically identifying senescence pathways and
regulators across multiple human tissues31.
Predicting novel senotherapeutic compounds
Research teams are also using ML approaches to discover novel
senolytics239,241. A dataset with 58 known senolytics and 2,465
non-senolytic drugs, each characterized by 200 molecular and struc-
tural properties
241
, was used to train SVM and Random Forest-based
classifiers to predict new senolytics. The trained classifiers were then
used to screen diverse chemical libraries and identified several poten-
tial new senolytic compounds. Three compounds, all plant-derived
molecules (oleandrin, periplocin and ginkgetin), showed evidence
of senolysis using in vitro models. Mechanistically, oleandrin is a
cardiac glycoside that inhibits Na
+
K
+
-ATPase; periplocin, also a gly-
coside, induces apoptosis through AMPK–mTOR or ERK signalling;
while ginkgetin, a naturally occurring biflavonoid, ac ts on multiple
pathways crucial to senescence, including apoptosis induction,
cell cycle arrest and JAK–STAT and MAPK signalling.
A separate approach used a morphology-based CNN model,
Deep-SeSMo, to calculate senescence scores in a drug screen of
80 compounds tested for their senolytic ability. The readout was per-
centage of SA-β-gal-positive cells, without presenting data on total cell
counts, making it unclear whether senolysis occurred. One possibility is
that these experiments may have identified senomorphics that impact
lysosomal function and SA-β-gal activity, which is consistent with the
senotherapeutic agents chosen to train the system, metformin and
nicotinamide mononucleotide, both of which act as senomorphics,
not senolytics (Box1).
These early studies showcase the power of AI and/or ML to rapidly
identify multiple potential senotherapeutics, increasing the need
for validation in appropriate brain cell culture systems and in vivo
models to understand the true predictive potential of these innovative
models.
Predicting druggability
Although identifying and selecting a target is a crucial initial step in
combatting senescence, the target’s druggability remains a decisive
factor in determining the viability of potential therapeutic interven-
tions. Druggability refers to the likelihood that a specific biological
target can be effectively modulated by a drug-like molecule
242,243
. This
aspect is particularly important when exploring potential therapeutic
interventions for senescence owing to the complex pathways involved
and the diverse nature of cellular senescent responses. For a target
to be considered druggable, it should be accessible to the intended
drug, whether that is a small molecule or a larger biological agent. The
probability of successfully drugging a given target is greatly improved
if its structure can be elucidated using methods such as NMR, X-ray
crystallography or cryo-electron microscopy
179
, as it allows for the
use of structure-based drug design. This approach, which uses com-
puter algorithms to identify potential binding pockets in the relevant
domains of the target protein and virtually screen them for chemical
matter that is likely to engage with these regions before commencing
experimental work, greatly cuts down on the number of compounds
needed in phenotypic screening approaches relative to the conven-
tional ‘brute-force’ approach, in which large chemical libraries are
screened244. Notably, recent advancements in AI systems, such as
AlphaFold245 have the potential to revolutionize protein structure
prediction by reducing reliance on laborious experimental methods,
thereby significantly broadening the scope of available targets for
drug design explorations.
Predicting off-target effects
Using systems biology approaches
246,247
, including network model-
ling and integration of multi-omics data, can provide comprehensive
insights into the global effects of senescence-targeting drugs beyond
the intended target. These tools can facilitate the identification of
potential off-target effects, discover novel pathways impacted by the
drug or drug combinations and optimize therapeutic efficacy while
minimizing adverse effects.
Conclusions
Over the past 5 years, research on cellular senescence in the brain
has evolved from phenomenology to clinical trials. Overcoming
challenges in CNS drug delivery, deepening our understanding of
senescence-associated mechanisms and appropriate targets, and
discerning when to use senolytics versus senomorphics are essential
to advance these strategies. Similarly, refinement of terminology in
the field to reflect specific types of senescent cell (for example, repli-
cative, physiological, stress-induced and postmitotic senescent), will
enable more precise drug development as each may require different
treatment strategies including choice of senotherapeutic, route of
administration, dose and scheduling protocol.
Developing dementia has become the most feared health concern
among adults worldwide
248–250
. As the global population continues
to live longer, the prevalence of dementia is expected to rise signifi-
cantly, creating an urgent need for effective therapeutic strategies.
Interdisciplinary approaches, AI and ML advancements and a compre-
hensive view of CNS senotherapeutics as described in Box1 can pave
the way for innovative treatments in age-related neurodegenerative
disorders. Currently, more than 20 senolytic trials are under way,
with four focusing on CNS conditions. Simply stated, the true poten-
tial benefits of these interventions are yet to be determined. While
acknowledging the challenges involved with advancing this new class
of treatments to the CNS, we maintain optimism and enthusiasm for
the future of senotherapy development and its application to nervous
system health. Addressing these challenges not only holds promise
for improving the quality of life for ageing individuals but also has
the potential to alleviate the broader societal and economic impacts
of an ageing population.
Published online: 30 September 2024
Nature Reviews Drug Discovery | Voume 23 | November 2024 | 817–837 833
Review article
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Acknowledgements
M.R. is supported by the Thomas Hartman Foundation and the National Institute of
Neurological Disorders and Stroke (R01NS124735-01A1). M.E .O. is supported by the
Alzheimer’s Drug Discovery Foundation (GC-201908-2019443), Cure Alzheimer’s
Fund, Hevolution/American Federation for Aging Research, National Institute on Aging
(R01AG068293, R01AG065839, U54AG079754, R24AG073199), National Institute of
Neurological Disorders and Stroke (R21NS125171), Rainwater Charitable Foundation and US
Department of Veterans Aairs (I01BX005717).
Author contributions
The authors contributed equally to all aspects of the article.
Competing interests
M.X. is an employee of Merck Sharp & Dohme LLC, a subsidiary of Merck & Co., Inc., Rahway,
NJ, USA. A.A.J. is an employee of Merck Sharp & Dohme (UK) Limited. M.E.O. has a patent
pending, ‘Detecting and treating conditions associated with neuronal senescence’.
Additional information
Peer review information Nature Reviews Drug Discovery thanks Mark Mattson, Yi Zhu and
Richard Faragher for their contribution to the peer review of this work.
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