Targeting senescent cells: Approaches, opportunities, challenges

Abstract

Cellular senescence is a hallmark of aging, whose onset is linked to a series of both cell and non-cell autonomous processes, leading to several consequences for the organism. To date, several senescence routes have been identified, which play a fundamental role in development, tumor suppression and aging, among other processes. The positive and/or negative effects of senescent cells are directly related to the time that they remain in the organism. Short-term (acute) senescent cells are associated with positive effects; once they have executed their actions, immune cells are recruited to remove them. In contrast, long-term (chronic) senescent cells are associated with disease; they secrete pro-inflammatory and pro-tumorigenic factors in a state known as senescence-associated secretory phenotype (SASP). In recent years, cellular senescence has become the center of attention for the treatment of aging-related diseases. Current therapies are focused on elimination of senescent cell functions in three main ways: i) use of senolytics; ii) inhibition of SASP; and iii) improvement of immune system functions against senescent cells (immunosurveillance). In addition, some anti-cancer therapies are based on the induction of senescence in tumor cells. However, these senescent-like cancer cells must be subsequently cleared to avoid a chronic pro-tumorigenic state. Here is a summary of different scenarios, depending on the therapy used, with a discussion of the pros and cons of each scenario.

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INTRODUCTION
Cellular senescence is a stress response mechanism
induced by different types of insults such as telomere
attrition, DNA damage, and oncogenic mutations,
among others [1]. First described in cultured human
diploid fibroblasts after successive rounds of division
[2], its main hallmarks are irreversible growth arrest,
alterations of cell size and morphology, increased
lysosomal activity, expression of anti-proliferative
proteins, resistance to apoptosis, activation of damage-
sensing signaling routes. Another important
characteristic is the regulated secretion of interleukins
(ILs), inflammatory factors, termed the senescence-
associated secretory phenotype (SASP) [3].
As there is ample evidence placing senescent cells as
one of the causes of age-related dysfunctions, it has
been considered to be one of the hallmarks of aging [4].
It was recently demonstrated that elimination of
senescent cells by genetic or pharmacological
approaches delays the onset of aging-related diseases,
such as cancer, neurodegenerative disorders or cardio-
vascular diseases, among others, showing that the
chronic presence of these cells is not essential [5–7].
Conversely, local injections of senescent cells drive
aging-related diseases [8, 9]. This data, together with
that obtained from tissues of patients with different
diseases and ages, has established causality of senescent
cells in some aging-related pathologies [10, 11].
Current therapies targeting senescent cells are focused
on: i) specific killing of these cells by senolytics; ii)
specific inhibition of the secretory phenotype (anti-
SASP strategy); and iii) improving clearance of
senescent cells by the immune system [12]. In addition,
currently available senescence-inducing therapies for
www.aging-us.com AGING 2019, Vol. 11, No. 24
Review
Targeting senescent cells: approaches, opportunities, challenges
Cayetano von Kobbe
1
1
Centro de Biología Molecular “Severo Ochoa” (CBMSO), Consejo Superior de Investigaciones Científicas (CSIC),
Universidad Autónoma de Madrid, Madrid
28049, Spain
Correspondence
to: Cayetano von Kobbe; email: cvonkobbe@cbm.csic.es
Keywords
: cellular senescence, senolytics, senomorphics, immunosurveillance, anti-aging therapies
Received:
September 27, 2019 Accepted: November 20, 2019 Published: November 30, 2019
Copyright:
von Kobbe. This is an open-access article distributed under the terms of the Creative Commons Attribution
License
(CC
BY 3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author
and
source
are credited.
ABSTRACT
Cellular
senescence is a hallmark of aging, whose onset is linked to a series of both cell and non-cell
autonomous
processes,
leading to several consequences for the organism. To date, several senescence routes have
been
identified,
which play a fundamental role in development, tumor suppression and aging, among other
processes.
The
positive and/or negative effects of senescent cells are directly related to the time that they remain in
the
organism.
Short-term (acute) senescent cells are associated with positive effects; once they have executed
their
actions,
immune cells are recruited to remove them. In contrast, long-term (chronic) senescent cells
are
associated
with disease; they secrete pro-inflammatory and pro-tumorigenic factors in a state known
as
senescence
-associated secretory phenotype (SASP). In recent years, cellular senescence has become the center
of
attention
for the treatment of aging-related diseases. Current therapies are focused on elimination of
senescent
cell
functions in three main ways: i) use of senolytics; ii) inhibition of SASP; and iii) improvement of
immune
system
functions against senescent cells (immunosurveillance). In addition, some anti-cancer therapies are
based
on
the induction of senescence in tumor cells. However, these senescent-like cancer cells must be
subsequently
cleared
to avoid a chronic pro-tumorigenic state. Here is a summary of different scenarios, depending on
the
therapy used, with a discussion of the pros and cons of each scenario.

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cancer stop tumor growth while causing accumulation
of senescent cells [13, 14], which subsequently become
a problem for the organism [15].
This review will summarize the hypothetical scenarios
that each anti-cell senescence approach (described
above) could face, either alone or in combination, with
a discussion of open questions that should be kept in
mind when targeting senescent cells.
Triggers of cell senescence
The onset of senescence in healthy tissue occurs in
response to different internal and external stimuli, such
as telomere attrition, DNA damage (alkylating agents,
radiation), oncogene activation, mitochondrial
dysfunction, and spindle, epigenetic, endoplasmic
reticulum (ER) and proteotoxic stress [16–19]. The
type and duration of the stimulus dictates the final
effect on the senescent cells [20]. These cells display
a characteristic phenotype comprising specific
cell/nuclear morphology (increased size, abnormal
shape and nuclear envelope changes), apoptosis
resistance, chromatin redistribution (senescence-
associated heterochromatin foci and senescence-
associated distension of satellites), epigenetic markers
(e.g. H3K9Me3), lipofuscin accumulation, SASP, and
overexpression of proteins such as p53, p16Ink4a,
p21WAF1, Differentiated Embryo Chondrocyte-expressed
gene 1 (DEC1) and senescence-associated β-Gal (SA-β-
Gal) [13, 21–24]. To date there is no universal marker
for senescence, and identification of senescent cells is
based on the combined detection of two or more
phenotypic aspects mentioned above, such as SA-β-Gal,
p16Ink4a or p21WAF1 [10].
One of the characteristic phenotypic hallmarks of cell
senescence is the secretion of a plethora of factors that
affect their environment (SASP), which also serves as
a call for the immune system to recognize and
eliminate the senescent cells [3, 25]. Among the SASP
factors that seem responsible for attraction of immune
cells are CSF (colony stimulating factor 1), CXCL-1
(chemokine C-X-C motif ligand 1), MCP-1 (monocyte
chemoattractant protein 1) and ICAM-1 (intercellular
adhesion molecule 1) [25]. In this scenario of acute or
short-term senescence, the tissue returns to normal
after a regeneration process [17] (Figure 1, steps 1-4).
The regeneration is a fundamental process to avoid
tissue atrophy and dysfunction. In this scenario of
replacement of senescent cells, we should keep in
mind the different capacity of renewal of some
tissues with respect to others, and the exhausted or
damaged state of stem cells that can lead to
functionally compromised differentiated cells or
carcinogenesis [26].
Implication of cell senescence in disease
Acute senescent cells play a direct role in tumor
suppression, efficient wound healing, embryogenesis,
placental formation, and tissue regeneration, among other
processes [17]. At this point, both their onset and primary
effect are positive for the organism [17, 20].
When senescence-inducing stimuli persist and decrease
the ability of the immune system to recognize and
eliminate senescent cells (by either immunosenescence
or immunosuppression), these cells accumulate. The
continual presence of senescent cells negatively affects
their environment, inducing damage, instability or
senescence in other cells through SASP [1, 27]. Over
time, these “secondary” damaged cells can become
either pro-tumorigenic or senescent, which increases the
cellular instability of the tissue, leading to dysfunction
and disease [27] (Figure 1, steps 5 and 6). In this sense,
some SASP factors play a direct role in fibroblast
activation and uncontrolled fibrotic scarring [28].
Chronic senescent cells (also termed “zombie” cells)
have been associated with the onset of several diseases
[1, 10, 13, 17]. In the last few years there have been
extensive studies to elucidate the causative role of
senescence in the onset of different pathologies [17].
These studies were mainly based on: i) detection of
senescent cells in tissues/organs from patients or animal
models; or ii) improvement in tissue/organ functions
upon removal of senescent cells in mice, by either
genetic or pharmacological interventions. This is a list
of some age-related diseases where cellular senescence
seems to play an important role:
Cancer
Aging is the main cause of cancer [29], and the
presence of senescent cells in aged tissues or xenograft
models correlates with the incidence of cancer [30,
31]. Their specific removal led to a delay in tumor
formation and reduced metastasis [6]. It is also
important to note that both senolytics and senomorphics
are currently being used in clinical trials for the
treatment of numerous types of cancer, such as leukeia,
lung cancer, melanoma and glioblastoma, among others
[16].
Neurodegenerative disorders
Senescent cell accumulation has been detected at sites of
brain pathology [7, 32, 33]. The presence of senescent
astrocytes correlates with the onset of pathologies such as
Parkinson’s and Alzheimer’s disease [34]. Interestingly,
Tau protein induces cellular senescence in neurons, and
specific clearance of senescent astrocytes and microglia,

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reduced Tau-containing neurofibrillary tangle, neuron loss
and ventricular enlargement [7, 8]. Moreover, it has been
proposed a role of senescent cells in multiple sclerosis
[33].
Cardiovascular disease
Senescent cells play a key role in atherosclerosis, and
their specific removal reduced progression of the
disease [35]. Moreover, senescent macrophages seem to
contribute to coronary heart disease, and cell senescence
in the aorta increases vascular stiffness [13].
Osteoarthritis
This degenerative disease causes the joints to become
painful and stiff, and accumulation of senescent cells
correlates with its progression [36]. In mouse models,
local injections of these cells induce an osteoarthritis-
like condition [9], whereas their clearance improves
health by attenuating development of post-traumatic
osteoarthritis [37].
Type 2 diabetes
Aging is the main cause of type 2 diabetes, and there is
association between disease progression and detection
of senescent markers. Senescent β-cells affect glucose
homeostasis, although further work is needed to
elucidate the exact role of senescence [20, 38, 39].
Kidney-related diseases
Diseases such as glomerulosclerosis and nephropathies
are associated with an increase of senescent cells [10].
Remarkably, when these cells were removed by genetic
approaches, kidney functions improved [6].
Idiopathic pulmonary fibrosis (IPF)
This chronic lung disease results in scarring, affecting
primarily older adults. Tissues from IPF patients
display some phenotypical characteristics of senescent
cells, and when these cells were removed by
senolytics, pulmonary functions improved [104].
Cachexia
In this disease adipocyte differentiation is disrupted by
senescent cells, causing weight loss, muscle wasting
and loss of body fat, leading to metabolic dysfunction
and loss of adaptive thermogenic capacity [10]. When
senescent cells were removed, tissue homeostasis
recovered [6, 75].
Figure 1. The onset of cellular senescence in normal tissue takes place in response to different stimuli (1). Some SASP factors are involved in
immune cell recruitment, which act in the clearance of the senescent cells (2). Then, to restore the normal tissue, a regeneration process is
necessary (3, 4). When a combination of persistent damage and immune system decay occurs, senescent cells accumulate, creating a pro-

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inflammatory and pro-tumorigenic environment and fibrotic tissue. Over time, this leads to disease, such as cancer progression, insulin
resistance, osteoarthritis, atherosclerosis, and brain pathologies, among others (5, 6).

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Cataracts
Characterized by opacity of the lens of the eye [109],
the lens capsules from patients suffering cataracts show
accumulation of senescent human lens epithelial cells
[105]. Removal of these cells by genetic approaches
decreased the incidence of cataracts in old mice [6].
Liver diseases
The presence of senescent cells correlates with the onset
of liver fibrosis, cirrhosis and non-alcoholic fatty liver
disease. Elimination of these cells reduced liver fat
accumulation [10, 106].
Metabolic syndrome
A collection of metabolic disorders such as increased
blood pressure, high blood sugar, excess body fat
(around the waist) and abnormal cholesterol levels.
Endothelial cell senescence is involved in systemic
metabolic dysfunction and glucose intolerance [13,
107].
Erectile dysfunction
The presence of senescent cells is directly related to
endothelial dysfunction. SASP factors seem mediate
this effect, and importantly, removal of senescent cells
led to improvement of erectile function in mice [40].
Altogether, this data highlights the importance of
targeting these cells in order to delay or cure different
diseases.
STRATEGIES TO SUPPRESS SENESCENT
CELLS
Senolytics
An option to eliminate the negative effects of chronic
senescent cells is to kill them specifically, using
compounds called senolytics (Figure 2), which target
pathways activated in senescent cells [16]. The list of
these senolytic tool compounds is extensive and
continuously growing. In Table 1 are shown the
noteworthy ones. Chronic/periodic administration of
senolytics kills senescent cells that are generated in the
tissues, and the immune system is responsible for
clearing apoptotic bodies for subsequent regeneration
with new cells (Figure 2, steps 1-3). Senolytics target
key proteins mainly involved in apoptosis, such as Bcl-
2, Bcl-XL, p53, p21, PI3K, AKT, FOXO4 and p53. See
Table 1 for references.
Although senolytics are supposed to be specific for
senescent cells, there are always unwanted damage/side
Figure 2. Treatment with senolytics to specifically kill senescent cells (1). Over time, these apoptotic bodies will be cleared by the immune
system (2). Finally, a regenerative process will lead to normal tissue functions (3). Normal cells could be affected by either the lack of
specificity of the senolytics or chronic treatment, leading to tissue dysfunction (4).

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Table 1. List of senolytics and their targets.
Senolytic
Target/function
References
Apoptosis
Dasatinib (D)
Inhibitor EFNB*-dependent suppression of apoptosis
[51]
Quercetin (Q)
PI3K/AKT, BCL-2, p53, p21, Serpine
[51]
ABT 737
BCL-W and BCL-XL inhibitor
[52]
ABT 263 (Navitoclax; UBX0101)
BCL-2, BCL-XL and BCL-W inhibitors
[37, 53, 54]
A1331852, A1155463
BCL-XL
[55]
Fisetin
PI3K/AKT and ROS
[55]
FOXO4-related peptide (DRI)
Inhibitor of FOXO4-p53 interaction
[44]
Delivery options**
Gal-encapsulated cytotoxics
SA-β-Gal
[42]
*AKT; protein kinase B. BCL; B-cell lymphoma. EFNB; ephrin ligand B. FOXO; forkhead box proteins O. PI3K;
phosphatidylinositol 3-kinase. ROS; reactive oxygen species.
**It helps improve senolysis by directed targeting.
effects since the administration is not directed [41]
(Figure 2, step 4). In this regard, a new strategy has
been recently described to specifically target senescent
cells in mice, using nanocapsules containing toxins (or
senolytics) [42]. The outer layer of these nanocapsules
are composed of substrates for enzymes that are
overexpressed in senescent cells. In this way, the toxin
(senolytic) will only be released inside senescent cells,
killing them [42]. Thus, these nanocapsules are a
vehicle to specifically deliver any type of senolytic into
senescent cells in mice. The specificity of the delivery is
important in non-targeted senolytics (natural product
derivatives with less defined biological activities), such
as quercetin and fisetin.
Though there have been numerous reports showing the
benefits of senolytics, it is important to highlight the
recently described effects of dasatinib + quercetin (D + Q)
treatment on lifespan in old animals [43]. Transplant of
senescent cells into healthy mice caused physical
dysfunction, which was reversed by oral administration of
D + Q [43]. Also, clearance of senescent neurons
improved neurological functions in transgenic mice
mimicking Tau aggregation-dependent neurodegenerative
disease [8]. It is also important to note that the treatment
with the peptide FOXO4-DRI restored renal functions in
both old (normal) mice and mice with accelerated aging
[44]. As indicated above, some senolytics are currently
being used in clinical trials for treating different diseases
[16]. In this sense it is important to mention that MDM2
inhibitors, targeting p53, are also in clinical phases as
anti-cancer therapies [45].
Remaining questions
There is reasonable doubt about the fate of the dead
senescent cells, especially when the immune system of
the patient is depressed (by either immunosenescence or
immunosuppression). The accumulation of these
apoptotic bodies may have undesired side effects (i.e.
further release pro-inflammatory factors in an already-
damaged tissue) [10]. Also, as indicated before, the
possible side effects of periodic/chronic treatments
should not be ignored. In fact, toxic effects after systemic
administration of BCL family inhibitors have been
described in patients, such as thrombocytopenia and
neutropenia [41]. It would be desirable that treatments
with senolytics are as sporadic as possible, without
affecting efficacy. Lastly, and as indicated above, the
regeneration process is an important issue to be analyzed
in the tissues where senescence clearance has taken
place.
SASP inhibitors (or senomorphics)
Another strategy to inhibit the functions of senescent cells
is through the specific silencing of SASP [16, 46], the
complex mixture of soluble factors such as cytokines,
chemokines, growth factors, proteases and angiogenic
factors that mediates the paracrine and autocrine functions
of senescent cells [3, 25] (Figure 3). The qualitative and
quantitative composition of this secretome is different
depending on the cell type and the senescence-inducing
stimulus, and becomes fully active a few days after the
persistent stimulus [3, 47, 48]. Senomorphics inhibit
SASP functions by targeting pathways such as p38
mitogen-activated protein kinase (MAPK), NF-κB, IL-1α,
mTOR and PI3K/AKT (Table 2), which act at the level of
transcription, translation or mRNA stabilization [21].
Alternatively, inhibition may be achieved by specific
neutralizing antibodies against individual SASP factors
(protein function inhibition), as is the case for IL-1α, IL-8
and IL-6.

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As IL-1α plays a direct role in SASP regulation,
targeting either the receptor (IL-1αR) or the ligand (IL-
1α) leads to decreased global SASP expression, with
special emphasis on oncogene-induced senescence
(OIS) [49, 50].
Importantly, the MABp1 antibody (neutralizing anti-
human IL-1α monoclonal) has proven efficient in
clinical trials against type 2 diabetes, sarcopenia and
inflammation [56–58], diseases in which senescent cells
play an important causative role [10].
IL-8 is a member of the CXC motif chemokine
upregulated in SASP, and is associated with some types
of cancer [50]. ABX-IL-8 is a humanized monoclonal
antibody against IL-8 that acts as an antagonist,
impairing IL-8 signaling. Treatment with ABX-IL-8
attenuates the growth of some cancer xenografts
models [59].
IL-6 is a pleiotropic cytokine also upregulated in SASP
that is involved in tumor proliferation, invasion and
immunosuppression. Specific inhibition of IL-6 by a
neutralizing monoclonal antibody (Mab-IL-6.8)
completely abolished JAK/STAT signaling [50, 77] and
relieved symptoms of arthritis in a primate model
(Olokizumab) [78]. Arthritis has also been causally
associated with the presence of senescent cells [37].
Finally, SASP-silenced/attenuated senescent cells should
be recognized by the immune system for subsequent
clearance and regeneration (Figure 3, steps 2 and 3).
Remaining questions
One doubt about this strategy is how SASP-
silenced/attenuated senescent cells would be cleared.
Given that some SASP factors are involved in the
recruitment of immune cells, SASP inhibition could
make senescent cells effectively “invisible” to the
immune system, therefore remaining chronically within
the tissue. In fact, two senomorphics (apigenin and
kaempferol) showed inhibition in cultured cells of SASP
components involved in immune cell recruitment, such as
CXCL-1 and CSF [65]. What would the influence of
SASP-silenced senescent cells be in the tissue? Perhaps
instead of being dysfunctional, the tissue would be non-
functional.
Likewise, as senomorphics require chronic/continuous
treatment, a major problem of these types of SASP
inhibitors is the lack of specificity for senescent cells.
Perhaps inhibition of individual SASP components by
neutralizing antibodies (as described above) would
minimize the potential side effects. As indicated for
senolytics, it would be desirable if over time, the
treatments with senomorphics were as sporadic as
possible without affecting efficacy.
Figure 3. Treatment with senomorphics to inhibit SASP factors in senescent cells (1). Over time, these cells will be removed by immune cells
(2). Finally, a regenerative process will lead to normal tissue functions (3). In aged or immunosuppressed individuals, this strategy would lead
to an accumulation of SASP-silenced/attenuated senescent cells (4).

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Table 2. List of senomorphics and their targets.
SASP inhibitor
Target/function*
References
SB 203580
p38 MAPK** inhibitor
([60] Reviewed by [12])
UR-135756, BIRB 796
p38 MAPK inhibitor
[61]
Resveratrol
NF-ƙB inhibitor (IĸB-kinase inhibitor), AMPK and
SIRT1 activator, others
[62–64]
Apigenin, Wogonin, Kaempferol
NF-ƙB inhibitors (IĸB-zeta)
[65]
Metformin
Inhibition of IKK/NF-ƙB, mitochondrial electron
tranport, mitochondrial GPDH, and KDM6A/UTX,
AMPK activator, others
[66–70]
Cortisol/corticosterone
IL-1α/NF-ƙB pathway inhibitors
[71]
NDGA
ROS (free radical scavenger)
[72]
Rapamycin
mTOR inhibitor, membrane-bound IL-1A translation
inhibition, prelamin A, 53BP1
[73] [74] [110]
Ruxolitinib
Inhibition of JAK1/2 and ROCK
[75, 76]
*For many of the SASP inhibitors listed there have been described several targets.
**53BP1; p53 binding protein 1. AMPK; AMP-activated protein kinase. IKK; IĸB kinase. JAK; Janus kinase. KDM6A/UTX; lysine
demethylase 6A. MAPK; mitogen-activated protein kinase. mTOR; mammalian target of rapamycin. NDGA;
nordihydroguaiaretic acid. NF-ƙB; nuclear factor kappa light chain enhancer of activated B cells. ROS; reactive oxygen species.
Improving immune system function
A third strategy to target senescent cells is to strengthen
the immune system for efficient recognition and
elimination of these cells, a process termed immuno-
surveillance (Figure 4, steps 1-3). The role of the
immune system in the elimination of senescent cells is
fundamental, and a decline in immune function is
associated with an increase in the number of senescent
cells and finally, disease (Figure 4, step 4) [12, 20, 79,
80].
In this regard, there are two strategies: i) improving the
specific anti-senescent cell functions; and ii) general
enhancement of immune functions (to avoid senescence
of immune cells involved in recognition of senescent
cells).
Anti-senescent cell functions have been described in NK
cells, macrophages and CD4+ T cells [20, 81]. Since these
functions take place through membrane receptors, one
option is to increase the binding affinity of the involved
receptors. In this sense, the use of chimeric antigen
receptor (CAR) T cells to target specific senescent-related
molecules would be an attractive approach. This strategy
is currently showing extraordinary results as anti-cancer
therapy [82]. Alternatively, specifically increasing the
surface expression of these receptors in senescent cells
could be attempted. NK cells recognize the CD58/ICAM1
receptor present in senescent cells [83]. In the case of
macrophages this recognition is not clear, and may occur
through modified membrane receptors in senescent cells
(glycans, lipids or vimentin), recognized by receptors
present in macrophages such as CD36, IgM, SIRPα, and
leptins. For T cells this process would be mainly mediated
by TCRs [84].
Another possibility is to reduce the number of senescent
immune cells, perhaps by depletion using specific
antibodies recognizing surface markers of senescence,
and in this way “rejuvenate” the immune system [84].
In this sense the recent identification of a targetable
senescent cell surface marker supports this strategy
[85].
NK and T cell functions decrease in older individuals.
The constitutive activation of the nutrient-sensing
component adenosine 5´-monophosphate-activated
protein kinase (AMPK) seems to play a central role in
this process [86]. Thus, an alternative approach to
increase functions of these immune cells is to target
AMPK functions, as the p38 MAPK inhibitor does [87].
Another approach would be to inhibit the killer cell
lectin-like receptor G1 (KLRG1, or CD57 in humans),
which increases on NK and T cells of older individuals.
Activation of KLRG1 in NK cells is associated with
activation of AMPK (via protein stabilization), which in
turn would inhibit cell functions. In the case of CD8+ T
cells, this mechanism may involve other inhibitory
receptors, such as programmed death 1 (PD-1) and
cytotoxic T lymphocyte antigen 4 (CTLA-4) [86].
The down-regulation of the CD28 receptor is a hallmark
of human CD8+ T cell senescence. Interestingly these

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senescent T cells have been found not only in old
individuals (aging process), but also associated to
diseases such as cancer and arthrosis [83], which are
aging-related diseases where senescent cells seem to
play a causative role, as discussed above.
This fact reinforces the idea of a pivotal role of immune
cells by delaying the onset of diseases related to the
accumulation of chronic senescent cells. In this regard, a
recent article shows that mice lacking the main cytotoxic
functions of NK and T cells (perforin pathway),
accelerates both senescent cell burden and aging [80].
Some current anti-cancer therapies are based on
immunotherapy, that stimulates the immune system to
recognize and kill disease-associated cells based on
differences in the expression of antigens between
pathogenic and normal cells [88]. Immunotherapy is
currently used not only for different types of cancer, but
also for infectious diseases, Alzheimer’s disease, and
even some types of addictions [89, 90]. Senescent cells
display a characteristic phenotype, which make them
suitable targets for this strategy. Cell and antibody
mediated responses are possible approaches, however,
the specificity of senescent antigens would be the
bottleneck to avoid undesirable side effects [108].
Remaining questions
Improving immune system functions to target senescent
cells could be difficult in scenarios such as immuno-
senescence (in older individuals or patients suffering
from premature aging of the immune system [91]) or
immunosuppression (i.e. patients treated with
corticosteroids or radiation, in cases of organ transplant,
autoimmune disease or cancer). CAR-based strategies
and immune system “rejuvenation” would be
personalized treatments, and thus very time consuming
and expensive. These strategies would rely on specific
(universal) senescence receptors, and a limiting factor
when detecting cell senescence is the lack of universal
markers [13]. Although novel technologies are making
detection of senescent cells in tissues more reliable [92,
93], the use of a combination of different biomarkers is
still necessary for confirmation. Thus, personalized
treatment targeting at least 2 senescence markers would
increase the challenge and difficulty of the process.
Moreover, the described connection between NK and T
cell activation and nutrient-sensing machinery suggests
that dietary interventions could be a promising approach
to maintain a healthy immune system in older
individuals, and thus the ability to efficiently clear
senescent cells. The up-regulation of CD28 (by forced
Figure 4. Improving immune system functions to efficiently remove senescent cells (1). A robust immune system targets senescent cells,
leading to their removal (2). Then a regenerative process will maintain normal tissue functions (3). In situations where the immune system
decays (e.g. immunosenescence or immunodepression), there will be an accumulation of senescent cells, increasing instability in the
tissue/organ (4).

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expression of either the receptor itself or other receptor
related to T cell activation) could be another attractive
approach to delay the senescence process in CD8+ T
cells. Last, but not least, it is important to keep in mind
that a general stimulation of the components of the
immune system might also induce autoimmune diseases
or may also promote some hematopoietic malignancies
[94, 95].
Targeting senescent cancer cells
A way to stop cancer progression is to induce
senescence in tumor cells (TIS; therapy-induced
senescence), through treatments targeting key pathways
activated in highly proliferative cells. These treatments
include DNA damage inducers (e.g. mitoxantrone,
doxorubicin, γ-radiation), and inhibitors of Aurora
kinase A (i.e. MLN8054, alisertib) and CDK4/6
(abemaciclib, palbociclib, ribociclib), among others [14,
96–98]. While stopping tumor growth, TIS becomes a
problem for the organism in the long-term, as cancer
survivors have a higher incidence of age-related
diseases linked to senescence, including cardiovascular
disease, neurodegeneration, sarcopenia and secondary
neoplasia [19]. Cancer cells that escape from TIS (or
“senescence-like” cancer cells) display some features,
such as polyploidy, stemness and aggressiveness. It has
been calculated that only 1 in 106 of senescent cancer
cells escape from TIS. Although it seems to be a rare
event, it occurs [99, 100].
At this point, it is conceivable to imagine a tissue that is
already damaged, not only by tumor cells but also a mix
of pre-tumorigenic and senescent cells, together with
fibrosis and SASP (Figure 5). The newly senescent cells
(from the tumor; TIS) would increase the level of SASP
in the tissue, leading to: i) growth of new tumors (or
sprouts of the former); ii) senescence induction in
neighboring cells; as well as iii) an increase in fibrotic
tissue. This scenario would lead to an exacerbation of
the pathology that was described in the starting point
(step 3).
One solution to this situation would be to combine TIS
(effective therapy to stop the growth of the tumor that is
already present) with one or more of the three anti-
senescent strategies presented above (senolytics,
senomorphics and improved immune function) (Figure
5). Then clearance and tissue renewal processes will be
necessary to restore tissue functions (Figure 5, step 4).
Figure 5. Inducing senescence in tumor cells will lead to an accumulation of senescence burden (1). The pro-inflammatory and pro-
tumorigenic environment (more SASP factors) leads to exacerbation of the pathology (e.g. cancer relapse, fibrosis, inflammation) (2, 3). By
targeting senescent cells with a combination of the approaches currently used, a better final scenario is possible (4). Fibrotic scarring may be
treated by other means, or cured over time.

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Table 3. Comparison of the therapies presented in this review.
Therapy
Pros
Cons
Senolytics
High specificity
- Targeted drugs
Sporadic treatments
- Depending on compound efficacy
Low specificity
- Non-targeted compounds
Side effects
- BCL family inhibitors
Increase in apoptotic bodies
Chronic treatments
- Depending on compound efficacy
Senomorphics
High specificity
- Targeting individual SASP
components
Sporadic treatments
- Depending on compound efficacy
Low specificity
- Targeting central pathways
Chronic treatments
- Depending on compound efficacy
Lack of senescent cell clearance?
Side effects
Improving immune
system
High specificity
- Personalized treatments
- Immunotherapy
Sporadic treatments
Dietary interventions
Time consuming and expensive
- Personalized treatments
Low specificity
- General activation
Side effects
- Autoimmunity?
- Hematopoietic malignancies?
Chronic treatments
Patients affected by immunosuppression and/or
immunosenescence
TIS
High specificity
- Specific targets
Sporadic treatments
Stops tumor growth
Possibility to combine with other therapies
Low specificity
- General damage (chemo-radiotherapies)
Side effects
- New tumors
- Fibrosis
Chronic treatments
Increasing senescence burden
Remaining questions
Importantly these therapies would rely on the state of the
patient´s immune system, and many patients have been
affected by treatments they have received previously
(immunosuppression), or by age (immunosenescence). In
this sense, it is likely that in some cases it would only be
necessary to inhibit SASP and not specifically induce
death of the senescent cells, to avoid depending on the
immune system for removal of apoptotic bodies.
And what about fibrosis? Fibrotic scarring can resolve
over time, being replaced by new tissue. However, if
this process is not completed (e.g. older people), the
normal function of key organs can be compromised.
Thus, alternative therapies should be kept in mind to
treat senescence-associated fibrosis [101].
CONCLUDING REMARKS
Targeting senescent cells has become an alternative
therapy for treating different aging-related diseases.
This therapy can be approached on three levels: i)
specific killing of these cells; ii) inhibition of their
secretory phenotype, therefore making them less
efficient; and iii) improving our immune system for
elimination of senescent cells.
The use of senolytics and senomorphics are showing
promising results, although is still too early to draw
conclusions. It is necessary to improve the specificity of
these compounds, as well as optimize the treatment (i.e.
dosage) to avoid unwanted effects. In this sense, progress
has been made on the specific delivery of drugs into
senescent cells by using nanocapsules. This elegant
approach may overcome the problem of specificity of
senolytic tool compounds when administrated in a chronic
manner [42]. Importantly, senolytics and senomorphics
are found in natural compounds, showing new
(nutraceutical) approaches to treat aging-related diseases,
although in a non-targeted way [102].
The “transformation” of normal cells into senescent
ones is accomplished by a multitude of internal and
external stressors in different physiological situations.

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Cancer cells can become senescent as well after
different therapies, though the new tumor-induced
senescent cells (TIS) generated are harmful in the long
term. In this scenario, the three options presented here
to either eliminate or “silence” the senescent cells are
important to combat TIS. The combination of these pro-
and anti-senescence approaches (TIS + senolytics
and/or senomorphics and/or improved immune system),
will play an important role in the cure of some types of
cancer [98].
In future clinical trials focused on eliminating senescent
cells, it will be important to determine when to initiate
the treatments (age of the patients), the schedule
(continuous, periodic and/or sporadic), as well as the
specific markers to determine the efficacy of the therapy
(see Table 3 for comparison of the therapies presented
in this review). Clinical trials should be supported by
robust preclinical results obtained in proper animal
models.
Senescent cells are the cause of several age-related
diseases, which account for a high percentage of all
causes of death worldwide and an expansion of
morbidity. Likewise, it is estimated that by the year
2045, the number of people older than 60 will surpass,
for the first time in history, the number of people under
the age of 15 [103]. Thus, the approaches presented in
this review highlight the urgent need for new therapies
to delay or cure age/senescence-related diseases.
ACKNOWLEDGMENTS
I am grateful to Adrián V. and Victoria Colombo for
their productive discussions and support. The
professional editing service NB Revisions was used for
technical editing of the manuscript prior to submission.
CONFLICTS OF INTEREST
The author declares that he is co-founder of SenCell
Therapeutics.
FUNDING
Consejo Superior de Investigaciones Científicas (CSIC).
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