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Citation: Hamada, M.; Varkoly, K.S.;
Riyadh, O.; Beladi, R.; Munuswamy-
Ramanujam, G.; Rawls, A.; Wilson-
Rawls, J.; Chen, H.; McFadden, G.;
Lucas, A.R. Urokinase-Type
Plasminogen Activator Receptor
(uPAR) in Inflammation and Disease:
A Unique Inflammatory Pathway
Activator. Biomedicines 2024,12, 1167.
https://doi.org/10.3390/
biomedicines12061167
Academic Editors: Ferenc Sipos and
Mafalda Fonseca
Received: 26 March 2024
Revised: 24 April 2024
Accepted: 10 May 2024
Published: 24 May 2024
Copyright: © 2024 by the authors.
Licensee MDPI, Basel, Switzerland.
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4.0/).
biomedicines
Review
Urokinase-Type Plasminogen Activator Receptor (uPAR) in
Inflammation and Disease: A Unique Inflammatory
Pathway Activator
Mostafa Hamada
1
, Kyle Steven Varkoly
2,
*, Omer Riyadh
1
, Roxana Beladi
3
, Ganesh Munuswamy-Ramanujam
4
,
Alan Rawls 5, Jeanne Wilson-Rawls 5, Hao Chen 6, Grant McFadden 7and Alexandra R. Lucas 7,*
1College of Medicine, Kansas City University, 1750 Independence Ave, Kansas City, MO 64106, USA;
mostafa.hamada@kansascity.edu (M.H.); omer.riyadh@kansascity.edu (O.R.)
2Department of Internal Medicine, McLaren Macomb Hospital, Michigan State University College of Human
Medicine, 1000 Harrington St., Mt Clemens, MI 48043, USA
3Department of Neurosurgery, Ascension Providence Hospital, Michigan State University College of Human
Medicine, 16001 W Nine Mile Rd, Southfield, MI 48075, USA; roxanabeladi@gmail.com
4Molecular Biology and Immunobiology Division, Interdisciplinary Institute of Indian System of Medicine,
SRM Institute of Science and Technology, Kattankulathur 603203, India; mrganesh2000@hotmail.com
5School of Life Sciences, Arizona State University, 427 E Tyler Mall, Tempe, AZ 85281, USA;
alan.rawls@asu.edu (A.R.); jeanne.wilson-rawls@asu.edu (J.W.-R.)
6Department of Tumor Center, Lanzhou University Second Hospital, Lanzhou 730030, China;
chenhao3996913@163.com
7Center for Personalized Diagnostics, Biodesign Institute, Arizona State University, 727 E Tyler St.,
Tempe, AZ 85287, USA; grantmcf@asu.edu
*Correspondence: kylevarkoly@gmail.com (K.S.V.); arlucas5@asu.edu (A.R.L.); Tel.: +1-904-859-4401 (K.S.V.);
+1-352-672-2301 (A.R.L.)
Abstract: The urokinase-type plasminogen activator receptor (uPAR) is a unique protease binding
receptor, now recognized as a key regulator of inflammation. Initially, uPA/uPAR was considered
thrombolytic (clot-dissolving); however, recent studies have demonstrated its predominant im-
munomodulatory functions in inflammation and cancer. The uPA/uPAR complex has a multifaceted
central role in both normal physiological and also pathological responses. uPAR is expressed as a
glycophosphatidylinositol (GPI)-linked receptor interacting with vitronectin, integrins, G protein-
coupled receptors, and growth factor receptors within a large lipid raft. Through protein-to-protein
interactions, cell surface uPAR modulates intracellular signaling, altering cellular adhesion and mi-
gration. The uPA/uPAR also modifies extracellular activity, activating plasminogen to form plasmin,
which breaks down fibrin, dissolving clots and activating matrix metalloproteinases that lyse connec-
tive tissue, allowing immune and cancer cell invasion and releasing growth factors. uPAR is now
recognized as a biomarker for inflammatory diseases and cancer; uPAR and soluble uPAR fragments
(suPAR) are increased in viral sepsis (COVID-19), inflammatory bowel disease, and metastasis. Here,
we provide a comprehensive overview of the structure, function, and current studies examining
uPAR and suPAR as diagnostic markers and therapeutic targets. Understanding uPAR is central to
developing diagnostic markers and the ongoing development of antibody, small-molecule, nanogel,
and virus-derived immune-modulating treatments that target uPAR.
Keywords: urokinase-type plasminogen receptor; uPAR;inflammation; cancer; sepsis; virus; inflammatory
bowel disease; Serp-1; serpin
1. Introduction
Hidden strength
Strange power
Pervasive
Biomedicines 2024,12, 1167. https://doi.org/10.3390/biomedicines12061167 https://www.mdpi.com/journal/biomedicines
Biomedicines 2024,12, 1167 2 of 44
Invasive
Universal and unknown
Life to life.
Alexandra Lucas, MD, FRCP(C)—March 2024
1.1. The Thrombolytic Plasminogen Activator (PA) Protease Cascade
The primary physiological role of the urokinase-type plasminogen activator (uPA)
system was first identified as a thrombolytic or clot-dissolving protease cascade. The
binding of uPA to urokinase-type plasminogen activator receptor (uPAR) initiates a cascade
of events, converting plasminogen into plasmin, which then drives fibrinolysis. Through
activation of plasmin the uPA/uPAR complex can also initiate cell activation, tissue remod-
eling, and angiogenesis. Here, we begin with a discussion of the role of uPA in fibrinolysis
and clot breakdown.
The tissue- and urokinase-type plasminogen activators (tPA and uPA, respectively)
are the two serine protease enzymes known to activate plasmin and break down clots.
tPA and uPA have differing cell surface receptors [
1
–
3
]. Both tPA and uPA have been
used as thrombolytic agents for myocardial infarction and other acute thrombotic events
such as strokes (cerebrovascular accident), deep venous thrombosis (DVT), and pulmonary
embolism. In the vascular system, although tPA and uPA both activate plasmin to cleave
fibrin, tPA is the principal thrombolytic or clot-dissolving protease in the circulating blood.
uPA also has clot-dissolving activity but is now reported to function predominantly as an
inflammatory mediator.
Plasminogen is cleaved to form plasmin, which is also a serine protease with a diverse
set of functions, in addition to clot lysis. Plasmin activates pro-matrix metalloproteinases
(MMPs) that drive extracellular matrix degradation and also release and activate latent
tissue growth factors, such as transforming growth factor-beta (TGF-
β
) [
1
,
2
]. Inhibitors
of tPA, uPA, and plasmin function to regulate these pathways. Serine protease inhibitors,
termed serpins, regulate the thrombolytic and thrombotic pathways and have been exten-
sively studied. Mammalian serpins that regulate the uPA, tPA, and plasmin pathways
include plasminogen activator inhibitor-1 (PAI-1, SERPINE1), PAI-2 (SERPINB2) [
1
,
3
], and
other serpins, specifically,
α
2-antiplasmin (SERPINF2) and
α
2-macroglobulin [
3
,
4
]. PAI-1
and PAI-2 are active when binding to the uPA/uPA receptor complex (uPA/uPAR), but
antiplasmin is most active in targeting proteases in the circulation.
The cell-associated uPA/uPAR complex and associated soluble uPAR cleavage prod-
ucts (suPAR) have now been identified as markers for severe immune and inflammatory
diseases, as well as markers for cancer.
1.2. Urokinase-Type Plasminogen Activator (uPA)
The uPA was first identified as a thrombolytic. However, as noted above, this unique
serine protease also has a leading role in the inflammatory response, with a wide range of
physiologically significant functions [5–15].
Historically, the uPA was first identified and reported in 1947 as a thrombolytic by
MacFarlane and Pilling, with the seminal identification of uPA as a protein in urine [
5
].
Approximately five years later, Sobel and colleagues officially christened this protein
“urokinase” [
6
], now used as a thrombolytic. Urokinase, or uPA, is in various bodily
fluids including plasma, seminal fluid, and extracellular matrix (ECM) [
7
]. uPA is also
detected on many immune cells and in many cancers. Thus, uPA, also termed urokinase,
was first studied as a thrombolytic and only later found to have widespread immune
pathway functions.
uPA is synthesized and secreted as a single glycosylated proenzyme or zymogen,
referred to as pro-uPA, which consists of 411 amino acids, with little or no intrinsic enzy-
matic activity. Pro-uPA contains three distinct domains: a growth factor domain, sharing
structural similarities with the epidermal growth factor (GFD); a kringle domain (KD);
and a serine protease domain [
8
,
14
–
17
] (Figure 1). The GFD and KD are situated at the
Biomedicines 2024,12, 1167 3 of 44
N-terminus, whereas the catalytic serine protease domain (amino acids 159–411) is located
at the C-terminus. A linker region connects the N- and C-terminal regions [9].
Biomedicines 2024, 12, x FOR PEER REVIEW 3 of 48
studied as a thrombolytic and only later found to have widespread immune pathway
functions.
uPA is synthesized and secreted as a single glycosylated proenzyme or zymogen,
referred to as pro-uPA, which consists of 411 amino acids, with little or no intrinsic enzy-
matic activity. Pro-uPA contains three distinct domains: a growth factor domain, sharing
structural similarities with the epidermal growth factor (GFD); a kringle domain (KD);
and a serine protease domain [8,14–17] (Figure 1). The GFD and KD are situated at the N-
terminus, whereas the catalytic serine protease domain (amino acids 159–411) is located
at the C-terminus. A linker region connects the N- and C-terminal regions [9].
Once secreted, pro-uPA undergoes cleavage at the peptide bond between Lys158 and
IIe159 to generate the double chain form of uPA. The two-chain form of uPA (tcuPA) is
still linked through a disulfide bond [10]. Thus, a key event in the activation of pro-uPA
is the cleavage of a specific peptide bond within the pro-uPA molecule. tcuPA consists of
a heavy chain (A-chain) and a light chain (B-chain). The structural variations in uPA con-
tribute to the functional versatility of uPA. tcuPA and the single-chain form (scuPA) can
both bind to uPAR; however, tcuPA is 250 times more potent at generating plasmin than
scuPA [11]. A range of different proteases, such as thrombin and elastase, have also been
identified as agents that facilitate cleavage of pro-uPA, resulting in the generation of active
high-molecular-weight uPA [10]. Additionally, various other activating proteins includ-
ing cathepsin B and L, mast cell tryptase, and nerve growth factor-gamma, have been rec-
ognized as capable of activating pro-uPA [12–14]. Among these, plasmin stands out as the
most efficient enzyme for the conversion of pro-uPA into its active counterpart, uPA [10].
Plasmin formed from plasminogen by uPA activation, thus, activates uPA.
Figure 1. Schematic of the enzymatic processing of uPA. uPA is secreted as a 411-amino-acid-long
inactive protein containing growth factor, kringle, and serine protease domains. Activation of uPA
occurs through a proteolytic cleavage between K158 and I159 followed by linkage of the two pep-
tides by a disulfide bond. A second round of proteolytic cleavage between K135 and K136 results in
a catalytically inactive amino terminal fragment and a catalytically active soluble low-molecular-
weight serine protease. uPA—urokinase-type plasminogen activator; uPAR—uPA receptor.
1.3. Urokinase-Type Plasminogen Activator Receptor (uPAR)
The urokinase-type plasminogen receptor (uPAR), also termed CD87 and encoded
by the PLAUR gene, is now recognized as a central immune regulating receptor. uPAR is
a 50–60 kDa glycosylated protein involved in multiple physiological and pathological pro-
cesses [7,17–29]. uPAR is part of the Ly6 (lymphocyte antigen-6)/uPAR family of recep-
tors. Ly6/uPAR encodes proteins containing at least one conserved functional motif,
termed the LY6/uPAR (LU) domain [18,19]. The LY6/uPAR superfamily of protein recep-
tors were first identified in T lymphocytes and comprise over 30 genes found in insects,
fish, amphibians, reptiles, birds, and mammals [18]. uPAR was first discovered by three
researchers simultaneously in 1985: Vassalli, Del Roso, and Blasi [21–23].
Figure 1. Schematic of the enzymatic processing of uPA. uPA is secreted as a 411-amino-acid-long
inactive protein containing growth factor, kringle, and serine protease domains. Activation of uPA
occurs through a proteolytic cleavage between K158 and I159 followed by linkage of the two peptides
by a disulfide bond. A second round of proteolytic cleavage between K135 and K136 results in a
catalytically inactive amino terminal fragment and a catalytically active soluble low-molecular-weight
serine protease. uPA—urokinase-type plasminogen activator; uPAR—uPA receptor.
Once secreted, pro-uPA undergoes cleavage at the peptide bond between Lys158 and
IIe159 to generate the double chain form of uPA. The two-chain form of uPA (tcuPA) is
still linked through a disulfide bond [
10
]. Thus, a key event in the activation of pro-uPA
is the cleavage of a specific peptide bond within the pro-uPA molecule. tcuPA consists
of a heavy chain (A-chain) and a light chain (B-chain). The structural variations in uPA
contribute to the functional versatility of uPA. tcuPA and the single-chain form (scuPA)
can both bind to uPAR; however, tcuPA is 250 times more potent at generating plasmin
than scuPA [
11
]. A range of different proteases, such as thrombin and elastase, have also
been identified as agents that facilitate cleavage of pro-uPA, resulting in the generation
of active high-molecular-weight uPA [
10
]. Additionally, various other activating proteins
including cathepsin B and L, mast cell tryptase, and nerve growth factor-gamma, have
been recognized as capable of activating pro-uPA [
12
–
14
]. Among these, plasmin stands
out as the most efficient enzyme for the conversion of pro-uPA into its active counterpart,
uPA [10]. Plasmin formed from plasminogen by uPA activation, thus, activates uPA.
1.3. Urokinase-Type Plasminogen Activator Receptor (uPAR)
The urokinase-type plasminogen receptor (uPAR), also termed CD87 and encoded by
the PLAUR gene, is now recognized as a central immune regulating receptor. uPAR is a
50–60 kDa glycosylated protein involved in multiple physiological and pathological pro-
cesses [
7
,
17
–
29
]. uPAR is part of the Ly6 (lymphocyte antigen-6)/uPAR family of receptors.
Ly6/uPAR encodes proteins containing at least one conserved functional motif, termed
the LY6/uPAR (LU) domain [
18
,
19
]. The LY6/uPAR superfamily of protein receptors were
first identified in T lymphocytes and comprise over 30 genes found in insects, fish, amphib-
ians, reptiles, birds, and mammals [
18
]. uPAR was first discovered by three researchers
simultaneously in 1985: Vassalli, Del Roso, and Blasi [21–23].
The uPAR is expressed on the surface of endothelial cells, neutrophils, monocytes/mac-
rophages, T cells, and fibroblasts, as well as many cancers, with reported major roles in
inflammatory and cancer cell responses (Figure 2). uPAR can alter both intracellular sig-
naling and cell activation, as well as extracellular adhesion and migration. uPAR binds
Biomedicines 2024,12, 1167 4 of 44
vitronectin and is associated with a large lipid raft of membrane proteins that include inte-
grins, growth factors including transforming growth factor beta (TGF
β
), fibroblast growth
factor (FGF), and vascular endothelial growth factor (VEGF), as well as chemokine receptors
(i.e., CXCR4) and low-density lipoprotein receptor-related protein (LRP). uPA bound to
uPAR (uPA/uPAR), as noted, also activates extracellular-matrix-degrading enzymes, such
as the MMPs, and can activate growth factors [7,17–19].
Biomedicines 2024, 12, x FOR PEER REVIEW 4 of 48
The uPAR is expressed on the surface of endothelial cells, neutrophils, mono-
cytes/macrophages, T cells, and fibroblasts, as well as many cancers, with reported major
roles in inflammatory and cancer cell responses (Figure 2). uPAR can alter both intracel-
lular signaling and cell activation, as well as extracellular adhesion and migration. uPAR
binds vitronectin and is associated with a large lipid raft of membrane proteins that in-
clude integrins, growth factors including transforming growth factor beta (TGFβ), fibro-
blast growth factor (FGF), and vascular endothelial growth factor (VEGF), as well as
chemokine receptors (i.e., CXCR4) and low-density lipoprotein receptor-related protein
(LRP). uPA bound to uPAR (uPA/uPAR), as noted, also activates extracellular-matrix-de-
grading enzymes, such as the MMPs, and can activate growth factors [7,17–19].
uPAR is a single-chain polypeptide, further subdivided into three domains (D1, D2,
and D3), each approximately 90 amino acids [7]. Following cleavage of this single-chain pol-
ypeptide, the D1 domain binds uPA (Figure 2). Although uPAR serves as the attachment
site for uPA to the cell surface, uPAR is not a transmembrane protein, but is instead a gly-
cosylphosphatidylinositol (GPI)-linked receptor (Figure 2). uPAR is covalently bonded to
the GPI, a glycophospholipid in the outer leaflet of the membrane. uPAR is reported to alter
internal cellular activity via interacting with adjacent transmembrane proteins in this large
lipid raft of associated membrane proteins, such as the integrins [7,17,18]. uPAR can also
alter cellular activation when incorporated into a cell, as is seen when uPAR is internalized
after binding to uPA and the plasminogen activator inhibitor-1 (PAI-1).
Figure 2. Schematic of the uPA receptor (uPAR). uPAR has three domains: D1, D2, and D3. D1
binds uPA and D3 is linked by GPI to the outer cell membrane. GPI—glycophosphatidyl inositol;
uPA—urokinase-type plasminogen activator.
The binding of uPA to uPAR is an intricate and tightly regulated molecular interac-
tion with a pivotal role in various physiological and pathological processes. The
uPA/uPAR complex is characterized by high-affinity binding, which is achieved through
specific binding of three consecutive binding domains on both uPA and uPAR. Crystal
structure analysis has provided a detailed uPA/uPAR binding structure. The Ly6/uPAR
family of receptors maintains a three-finger structure. The three-dimensional structure of
uPAR forms a central hydrophobic site deep in its D1 domain core structure. This location
serves as the ligand-binding site for uPA, specifically the β-hairpin structure on the sur-
face of the growth-factor-like domain (GFD) [19,20]. This tight binding near the center of
uPAR enables the external surfaces of uPA to interact with other proteins in the lipid raft
where uPAR sits, including proteins such as vitronectin (Vn). Vn is a multifunctional gly-
coprotein present in both the extracellular matrix and blood and is closely involved in
Figure 2. Schematic of the uPA receptor (uPAR). uPAR has three domains: D1, D2, and D3. D1
binds uPA and D3 is linked by GPI to the outer cell membrane. GPI—glycophosphatidyl inositol;
uPA—urokinase-type plasminogen activator.
uPAR is a single-chain polypeptide, further subdivided into three domains (D1, D2,
and D3), each approximately 90 amino acids [
7
]. Following cleavage of this single-chain
polypeptide, the D1 domain binds uPA (Figure 2). Although uPAR serves as the attachment
site for uPA to the cell surface, uPAR is not a transmembrane protein, but is instead a
glycosylphosphatidylinositol (GPI)-linked receptor (Figure 2). uPAR is covalently bonded
to the GPI, a glycophospholipid in the outer leaflet of the membrane. uPAR is reported
to alter internal cellular activity via interacting with adjacent transmembrane proteins in
this large lipid raft of associated membrane proteins, such as the integrins [
7
,
17
,
18
]. uPAR
can also alter cellular activation when incorporated into a cell, as is seen when uPAR is
internalized after binding to uPA and the plasminogen activator inhibitor-1 (PAI-1).
The binding of uPA to uPAR is an intricate and tightly regulated molecular interaction
with a pivotal role in various physiological and pathological processes. The uPA/uPAR
complex is characterized by high-affinity binding, which is achieved through specific
binding of three consecutive binding domains on both uPA and uPAR. Crystal structure
analysis has provided a detailed uPA/uPAR binding structure. The Ly6/uPAR family
of receptors maintains a three-finger structure. The three-dimensional structure of uPAR
forms a central hydrophobic site deep in its D1 domain core structure. This location serves
as the ligand-binding site for uPA, specifically the
β
-hairpin structure on the surface of the
growth-factor-like domain (GFD) [
19
,
20
]. This tight binding near the center of uPAR enables
the external surfaces of uPA to interact with other proteins in the lipid raft where uPAR sits,
including proteins such as vitronectin (Vn). Vn is a multifunctional glycoprotein present
in both the extracellular matrix and blood and is closely involved in integrin interactions
and cell adhesion. uPAR can interact with uPA, plasminogen, glycosaminoglycans, and
collagen, and when bound to uPA can stabilize interactions with the inhibitor PAI-1, a
mammalian serpin. Such multifaceted binding allows uPAR to regulate other activities
such as proteolysis and cell adhesion, including, for example, formation of actin-rich
lamellipodia on Vn [
24
,
25
]. This molecular model provides a basis for the potential use and
development of small-molecule inhibitors of uPA/uPAR interactions, a growing field, as
discussed in this review.
uPAR, together with Vn, is closely associated with transmembrane integrin proteins
that are associated with cell migration, adhesion, and proliferation. uPAR interacts with
many integrins; however, uPAR has the highest affinity for the fibronectin receptors
α
5
β
1
Biomedicines 2024,12, 1167 5 of 44
and
α
3
β
1 (Figure 3). Other integrins that interact with uPAR include macrophage 1 antigen
(Mac1, a complement receptor also known as CD11b/CD18 or CR3), a leukocyte
α
M
β
2
-integrin receptor that binds fibrinogen and the
α
v
β
5, and the
α
v
β
3 integrins that bind
Vn [
25
]. Through uPAR’s interaction with integrins and other associated receptors including
G protein-coupled receptors (GPCRs) in the uPAR lipid raft, cellular signaling, cellular acti-
vation, and cellular migration are modified. GPCRs include the chemokine receptors that
are central to cellular chemotaxis and migration, linking chemokines bound to glycosamino-
glycans in the cellular matrix to the cell surface-linked GPCRs, forming chemotactic cell
gradients. One such receptor includes uPAR-agonization of the FPRL1/LXA4R GPRC nec-
essary for uPA chemotactic activity, resultantly linking the fibrinolytic and inflammatory
responses in the process [
30
]. Once PAI-1 binds to the uPA/uPAR complex, the inhibited
complex is internalized, further modifying intracellular signaling and activation [24–26].
Biomedicines 2024, 12, x FOR PEER REVIEW 5 of 48
integrin interactions and cell adhesion. uPAR can interact with uPA, plasminogen, gly-
cosaminoglycans, and collagen, and when bound to uPA can stabilize interactions with
the inhibitor PAI-1, a mammalian serpin. Such multifaceted binding allows uPAR to reg-
ulate other activities such as proteolysis and cell adhesion, including, for example, for-
mation of actin-rich lamellipodia on Vn [24,25]. This molecular model provides a basis for
the potential use and development of small-molecule inhibitors of uPA/uPAR interac-
tions, a growing field, as discussed in this review.
uPAR, together with Vn, is closely associated with transmembrane integrin proteins
that are associated with cell migration, adhesion, and proliferation. uPAR interacts with
many integrins; however, uPAR has the highest affinity for the fibronectin receptors α5β1
and α3β1 (Figure 3). Other integrins that interact with uPAR include macrophage 1 anti-
gen (Mac1, a complement receptor also known as CD11b/CD18 or CR3), a leukocyte αMβ2
-integrin receptor that binds fibrinogen and the αvβ5, and the αvβ3 integrins that bind Vn
[25]. Through uPAR’s interaction with integrins and other associated receptors including
G protein-coupled receptors (GPCRs) in the uPAR lipid raft, cellular signaling, cellular
activation, and cellular migration are modified. GPCRs include the chemokine receptors
that are central to cellular chemotaxis and migration, linking chemokines bound to gly-
cosaminoglycans in the cellular matrix to the cell surface-linked GPCRs, forming chemo-
tactic cell gradients. One such receptor includes uPAR-agonization of the FPRL1/LXA4R
GPRC necessary for uPA chemotactic activity, resultantly linking the fibrinolytic and in-
flammatory responses in the process [30]. Once PAI-1 binds to the uPA/uPAR complex,
the inhibited complex is internalized, further modifying intracellular signaling and acti-
vation [24–26].
Through close interactions, the activation of integrins can further modulate cellular
signaling and increase production of pro-inflammatory cytokines such as interleukin-1
beta (IL-1β), IL-6, etc. Additionally, the induction of JAK-STAT by integrins and other
signaling pathways can increase cell-mediated immunity and division (Figure 3).
Figure 3. The urokinase -type plasminogen activator receptor (uPAR) modifies intracellular and
extracellular pathways. uPAR is a GPI-linked membrane protein that modifies intracellular down-
stream pathway responses via protein-to-protein interactions in a large lipid raft of associated mem-
brane proteins. uPAR modifies intracellular activation through interactions with integrins and
GPCRs, as illustrated here. Mammalian serpin PAI-1 binding to the uPA/uPAR complex inhibits
Figure 3. The urokinase-type plasminogen activator receptor (uPAR) modifies intracellular and ex-
tracellular pathways. uPAR is a GPI-linked membrane protein that modifies intracellular downstream
pathway responses via protein-to-protein interactions in a large lipid raft of associated membrane
proteins. uPAR modifies intracellular activation through interactions with integrins and GPCRs, as
illustrated here. Mammalian serpin PAI-1 binding to the uPA/uPAR complex inhibits uPA/uPAR
activity and can also be internalized, further modifying and altering intracellular activity pathways.
uPA/uPAR also modifies extracellular activity via activation of plasminogen to form plasmin, with
subsequent activation of MMPs and growth factors that alter cellular invasion into the extracellular
matrix surrounding adjacent cells. The virus-derived serpin Serp-1 also binds and inhibits uPA and
the uPAR as well as plasmin. MMP—matrix metalloproteinases; GPCR—G protein-coupled receptor;
PAI—plasminogen activator inhibitor; C—complement; Vn—vitronectin; GPI—glycophosphatidyl
inostitol; ECM—extracellular matrix; Serp-1—virus-derived serpin.
Through close interactions, the activation of integrins can further modulate cellular
signaling and increase production of pro-inflammatory cytokines such as interleukin-1 beta
(IL-1
β
), IL-6, etc. Additionally, the induction of JAK-STAT by integrins and other signaling
pathways can increase cell-mediated immunity and division (Figure 3).
To ensure precise control, uPA/uPAR interactions are strictly regulated by various
inhibitors. PAI-1 (SERPINE1 gene) is a serine protease inhibitor, a serpin that is an essential
regulator for both uPA and tPA [
31
], preventing uncontrolled proteolysis and fibrinolysis.
Biomedicines 2024,12, 1167 6 of 44
The balance between uPA, uPAR, and PAI-1 maintains homeostasis. uPA activity is also
diminished by other cellular mechanisms, including endocytosis, when uPA/uPAR is
bound to PAI-1 [32].
For uPAR to effectively bind its ligand with the highest affinity, full involvement of
all three D domains of uPAR is needed. Cleavage of the D1–D2 linker domains by other
proteases irreversibly blocks uPAR’s interaction with other proteins [27,28]. In alternative
pathways, uPAR is converted into a soluble form that is shed from the cell surface. This
soluble uPAR (suPAR) consists of all three domains which still maintain the full capacity to
bind uPA and cannot be cleaved by uPA [27,29].
1.4. Soluble Urokinase-Type Plasminogen Activator Receptor (suPAR)
Enzymatic cleavage of the uPAR GPI anchor leads to the generation of a soluble
variant of uPAR, referred to as suPAR [
29
,
33
]. This cleavage is mediated by various prote-
olytic enzymes, including plasmin and other proteases such as glycerophosphodiesterase
(GDE3). The specific cleavage sites on uPAR may vary, but they are generally located
in the extracellular region of the protein. GDE3 is a membrane-associated GPI-specific
phospholipase C, that releases uPAR from the cell membrane surface. This suPAR fragment
is released into the bloodstream, circulating throughout the body and making it detectable
in plasma samples. suPAR retains the ability to bind to uPA, plasminogen, and other
ligands, contributing to various processes involved in immune response regulation and
inflammation. Increased suPAR levels in the bloodstream are associated with pathological
conditions, including cardiovascular disease, renal disorders, and cancer [31]. In addition
to its proteolytic activity, the binding of uPA to suPAR also triggers the same intracellular
signaling pathways as uPA. suPAR can transduce signals into the cell, influencing processes
such as cell proliferation, migration, and adhesion. This makes the uPA/suPAR interaction
a key player in tumor and cancer progression, both as a marker for disease and potentially
as a mediator of cancer and other diseases [32,34,35].
The interactions between the serpin PAI-1 and suPAR have not been fully defined but
may also lead to irreversible inhibitory actions, as seen with PAI-1 inhibition of uPA. The
exact function of suPAR is an area of ongoing, active research, but it is believed to be in-
volved in innate (inflammatory) immune responses and tissue remodeling, similar to uPAR.
Elevated suPAR levels are reported with increased innate and acquired immune pathway
activation. suPAR in the blood can be detected through the enzyme-linked immunosorbent
assay (ELISA) or other point-of-care immunoassay techniques. This technique has been
studied for patients with SARS-CoV-2 infections and antibiotic administration in patients
with acute SARS-CoV-2 infections (COVID-19) [
36
–
38
]. Additionally, suPAR levels are rela-
tively stable throughout the day, in addition to stability in plasma samples following many
freeze–thaw cycles, but can be elevated in disease [
39
]. suPAR has thus been proposed
as a biomarker for a variety of inflammatory diseases, as well as a potential therapeutic
target [40,41].
2. uPAR as a Marker for Severe Inflammatory Diseases and Cancer
In this section, we will begin with a full review of uPAR as a clinical marker for disease
progression organized according to studies reported for individual organ systems.
The suPAR/uPAR system has been studied as a biomarker and therapeutic target in
a wide variety of diseases, from cardiovascular and hematological diseases to oncology
and chronic inflammatory diseases. suPAR and uPAR have been reported as markers
for systemic inflammatory response syndrome (SIRS) and sepsis, wound healing and
repair, neurological diseases, infectious diseases, cancers and also renal, gastroenterology,
endocrine, pulmonary, and arthritic disorders.
The current studies examining uPAR and suPAR as markers for disease and as poten-
tial targets for new treatments in diseases associated with individual organ systems are
discussed in Section 2and listed in Table 1by individual organ systems and diseases.
Biomedicines 2024,12, 1167 7 of 44
Table 1. uPAR/suPAR as diagnostic markers for clinical pathology.
Disease Role of uPAR/suPAR Target/Outcomes Subjects Citation(s)
Acute Coronary
Syndrome and
Percutaneous Coronary
Intervention
(1) Increased uPAR on circulating
monocytes in patients with ACS
(2) Increased suPAR associated with
increased MI and cardiac mortality
(3) suPAR product of atherogenic cells
(1) uPAR levels are elevated in patients
with ACS and myocardial necrosis.
(2) uPAR predicts future MACE. Human [42–45]
Acute Myocardial
Infarction (AMI)
(1) Deficient uPAR—protection against
myocardial rupture and reduced
myocardial infarct size
(2) Reliable biomarker for mortality in
adults with AMI and cardiac arrest
(1) uPAR-deficient mice protected against
myocardial rupture with reduced infarct
size.
(2) suPAR levels predict 1 year mortality
in AMI and correlate with neurological
outcome and mortality after cardiac
arrest.
Mice, Human [46–54]
Coronary Artery Disease
(CAD) suPAR is a cleavage product of
atherogenic immune cells suPAR and uPAR levels correlate with
extent of CAD. Human [47,55]
Cardiovascular Risk
Stratification suPAR is preditive of CV mortality
(1) suPAR, together with standard clinical
markers, e.g., hs-CRP and NT-proBNP, is
predictive of cardiovascular mortality.
(2) suPAR outperforms hs-CRP for
prognosis of inpatient mortality for
coronary disease.
(3) suPAR independent predictor for
future adverse cardiac events.
Human [56,57]
Viral Myocarditis Remodeling of cardiac tissue with
ventricular dysfunction
uPA-deficient mice with viral myocarditis
protected against cardiac dilatation and
failure. Mice [58]
Chronic Heart Failure suPAR correlates with heart failure suPAR concentration strongly correlates
with mortality in patients with heart
failure. Human [56]
Congenital Heart Block Increased anti-Ro and uPAR associated
with increased scar, collagen deposition,
and heart block
uPAR colocalizes and interacts with Ro60
on apoptotic human fetal cardiomyocytes.
Human [59,60]
Acute Ischemic Stroke uPAR is linked to carotid vascular
pathology
(1) Patients with higher uPAR have
increased risk of ischemic stroke and
carotid atherosclerosis.
(2) uPAR levels in patients with carotid
artery atherosclerosis are reduced with
beta-blocker.
Human [61–63]
Angiogenesis The uPA/uPAR complex is directly
involved in release of pro-angiogenic
growth factors such as FGF-2 and VEGF
(1) Reduced uPAR in endothelial cells
impairs VEGF signaling.
(2) uPAR-deficent mice have incomplete
angiogenesis.
Human Cells,
Mice [64–66]
Metastatic Cancer
uPAR is involved in extracellular matrix
degradation, tumor angiogenesis, cell
migration and proliferation, apoptosis,
and contributes to multidrug resistance
(MDR) in multiple cancers, specifically,
breast, lung, prostate, head and neck, and
ovarian cancers
Increased uPAR expression is consistently
observed in various metastatic solid
tumor tissues. Humans [67–70]
Breast Cancer
(1) Malignant breast tumors exhibit
slightly higher uPA and PAI-1 mRNA
expression but significantly increased
uPAR mRNA expression compared to
benign tissue.
(2) HER-2 (+) breast cancer has higher
uPAR expression.
Humans [68,69,71]
Lung Cancer
(1) Monoclonal antibodies targeting
uPAR decreased the invasive potential of
lung cancer strain 95D cells in vitro.
(2) uPAR-targeting U11 peptide
conjugated with pH-sensitive
doxorubicin and curcumin has synergistic
anti-tumor effects on lung tumor cells.
In vitro, Mice [72–75]
Prostate Cancer
(1) Inhibition of uPAR via MIR143 in
nanoparticles inhibits tumor growth.
(2) uPAR is highly expressed in
non-homeostatic prostate tissue
Human Cell
Lines [70,76–78]
Head and Neck
Squamous Cell
Carcinoma (HNSCC)
uPAR is associated with poor prognosis
and resistance to anticancer agents. Human Cells,
Clinical Samples [79]
Ovarian Cancer uPAR expression increases with
lysophosphatidic acid (LPA) stimulation
in ovarian cancer.
Human Cell
Lines [80,81]
Prostate Cancer uPAR is highly expressed in prostate
cancer.
Human
Xenograft SCID
mice [82]
Biomedicines 2024,12, 1167 8 of 44
Table 1. Cont.
Disease Role of uPAR/suPAR Target/Outcomes Subjects Citation(s)
Leukemia
uPAR mRNA variants play a specific role
in the progression of AML Transfection of uPAR 3′UTR modulates
pro-tumoral factors and cell adhesion. Human Cell
Lines [83]
Chronic Inflammation
suPAR is a cleavage product that is
elevated in acute and chronic
inflammatory diseases and comorbidities,
specifically, RA, SLE, SIRS, sepsis, HIV,
TB, pancreatitis, hepatitis, liver failure,
diabetes, kidney damage, asthma,
pneumonia, COPD, and smoking
suPAR levels correlate with chronic
inflammation. Human Studies [31,55–57,84]
Rheumatoid Arthritis
(RA)
(1) suPAR levels exhibit a direct
correlation with the number of inflamed
joints.
(2) Elevated suPAR levels are observed in
RA patients.
Humans [85,86]
Systemic Lupus
Erythematous (SLE) suPAR levels are elevated and predict
disease progression. Humans [87]
Systemic Inflammatory
Response Syndrome
(SIRS)
(1) Initial suPAR concentrations
significantly higher in patients who died
within 28 days.
(2) suPAR—good biomarker for
differentiating SIRS from sepsis.
Humans [88]
Bacterial and Viral Sepsis
(1) suPAR is a biomarker for identifying
adult bacterial sepsis.
(2) suPAR outperforms hs-CRP for
hospital mortality prognosis for bacterial
septic shock.
(3) suPAR is superior to other biomarkers
for differentiating septic vs. non-septic
neonates.
Humans [88–91]
HIV
(1) uPAR expression increased on
lymphocytes and monocytes after HIV
infection.
(2) uPAR levels correlate with prognosis
of HIV-1 similar to CD4+ count and viral
load.
Human Studies [92,93]
Tuberculosis (TB)
(1) suPAR levels are elevated in active
tuberculosis.
(2) suPAR is marker for treatment efficacy
and elimination of TB.
Human Studies [94]
Acute Pancreatitis
(1) suPAR differentiates severe acute
pancreatitis (SAP) with high diagnostic
accuracy.
(2) suPAR differentiates SAP from mild
pancreatitis.
(3) uPAR predicts inpatient mortality
from SAP.
(4) suPAR correlates with clinical scores
and lab values.
Humans [95]
Chronic Hepatitis Fibrosis
Progression
(1) suPAR is fair biomarker for liver
fibrosis in chronic HCV.
(2) suPAR distinguishes early and
advanced fibrosis.
(3) suPAR distinguishes cirrhotic and
non-cirrhotic patients.
Humans [96–98]
Acute Decompensated
Liver Failure (ADLF)
(1) suPAR levels > 14.4 ng/mL predict 28
day mortality in patients with ADLF.
(2) Ascitic suPAR levels increased in
bacterial peritonitis.
Humans [99]
Type 1 Diabetes Mellitus
(T1DM)
(1) suPAR is elevated across all patients
with T1DM.
(2) Patients with CVD carried a 2.5 times
higher suPAR level, 2.7 times for patients
with autonomic dysfunction, 3.8 times for
albuminuria, and 2.5 times higher for stiff
arterial walls.
Human [100]
Type 2 Diabetes Mellitus
(T2DM)
(1) Higher baseline suPAR levels showed
a higher risk of microalbuminuria in
patients at risk for T2DM or
patients with diagnosed T2DM.
Human [101]
Acute Kidney Injury
(AKI)
(1) Patients undergoing coronary
angiography, cardiac surgery, or admitted
to ICU within the upper quartile suPAR
levels had increased risk for AKI and
death at 90 days across all cohorts.
(2) Mice given suPAR and contrast had
greater pathologic evidence of AKI.
(3) Monoclonal-uPAR-antibody-treatment
mice had reduced AKI.
Humans, Mice
Overexpressing
uPAR [101,102]
Biomedicines 2024,12, 1167 9 of 44
Table 1. Cont.
Disease Role of uPAR/suPAR Target/Outcomes Subjects Citation(s)
Focal Segmental
Glomerosclerosis (FSGS)
Identified optimal suPAR value for
diagnosis of FSGS was 4.644 ng/mL;
sensitivity and specificity of 0.91 and 0.91;
AUC of 0.946 and may be used to
differentiate FSGS from other glomerular
diseases.
Humans [103]
Asthma
(1) Patients who were readmitted to
hospital due to an acute asthma
exacerbation or died had higher suPAR
and decreased eosinophil on admission.
(2) Patients in the 4th quartile for suPAR
levels or eosinophil counts < 150 cells/uL
had an increase in readmission or
mortality.
Human [104]
Community-Acquired
Pneumonia
(1) suPAR levels were significantly
elevated in patients with CAP and
correlate with Pneumonia Severity Index.
(2) LPS expression increases suPAR levels
in macrophages.
Mice, Human [105]
Ventilator-Associated
Pneumonia
(1) Plasma suPAR levels were increased
on day of diagnosis (AUC 0.77, p= 0.01)
and in deceased patients (AUC 0.79, p<
0.001) in patients diagnosed with VAP.
(2) suPAR significantly increased in
patients with VAP 3 days before
definitive diagnosis.
Human [106,107]
Chronic Obstructive
Pulmonary Disease
(1) Correlation superior for suPAR when
compared to other acute-phase reactants,
including CRP and fibrinogen.
(2) suPAR is a clinically useful biomarker
for early COPD diagnosis: sensitivity and
specificity of 87% and 79%.
(3) suPAR predictor for acute COPD
exacerbation and monitoring treatment
response.
Human [108,109]
Smoking Exposure
(1) suPAR levels significantly elevated in
smokers at 3.2 ng/mL vs. 1.9 ng/dL,
respectively.
(2) Four weeks following cessation,
suPAR levels are comparable to never
smokers when compared to those who
did not stop smoking.
Humans [110]
Neurological Disorders
(1) suPAR levels are elevated in the CSF
in distinct CNS pathologies
(2) uPAR is upregulated in microglial cells
during acute intracerebral LPS exposure
(1) suPAR identified in CSF of patients
with HIV dementia.
(2) suPAR superior specificity when
compared to CRP for discriminating
osteomyelitis and other
neurodegenerative spinal diseases.
Mouse Model,
Humans [63,111–115]
COVID-19 (SARS-CoV-2)
Pneumonia
Altered uPA/uPAR expression associated
with severe COVID-19
(1) uPAR associated with hypoxia and
pneumonia.
(2) COVID-19 patients with low suPAR
have lower mortality.
(3) suPAR guided anakinra treatment
provided survival benefit.
Human Studies,
Mouse [116]
Inflammatory Bowel
Disease (IBD) uPAR maintains the integrity of the
intestinal epithelial barrier
(1) uPAR expression increases during
active-gut-damage IBD.
(2) uPAR suppresses EGFR-modulated
repair and signaling.
Cell Lines,
Mouse [117,118]
Parenchymal Lung Injury
and Repair uPAR associated with lung fibrotic
processes and tissue remodeling suPAR correlates with aggressive
management in pleural effusions. Human, Mouse [119,120]
Ocular Diseases
uPAR linked to inflammatory
neovascular formation in ocular diseases,
specifically, premature retinopathy,
retinitis pigmentosa, and
wet macular degeneration
uPA/uPAR is reported in disease
progression. Mouse Model [121]
Allograft Transplant
Rejection
(1) uPA/uPAR activation correlates with
allograft rejection
(2) uPAR is necessary for TNF-alpha and
C5a signaling, inducing integrin ICAM-1
signaling on allograft endothelial cells for
leukocyte diapedesis
Serp-1 efficacy in aortic allograft models
was blocked in uPAR-deficient allograft
implants. Mice [122–124]
Biomedicines 2024,12, 1167 10 of 44
2.1. Cardiovascular Disease
Cardiovascular disease is the leading cause of death in North America. Previous
research has identified uPAR as a key player in the pathogenesis of cardiovascular disease,
and specifically, atherosclerosis, with strong associations to leukocyte invasion and tissue
remodeling (Table 1) [
125
]. Various cell types, such as smooth muscle cells, macrophages,
and endothelial cells all express uPAR. Such discoveries have led to broad clinical studies
that correlate the levels of suPAR to cardiovascular morbidity and mortality [125,126].
2.1.1. Acute Coronary Syndrome
Acute coronary syndrome (ACS) is a medical emergency caused by thrombosis with
sudden complete or partial blockage of blood flow to the heart muscle. ACS is typically
caused by macrophage invasion and rupture of an atherosclerotic plaque within a coronary
artery, exposing the inner atheromatous plaque, leading to platelet activation and activation
of the clotting cascade, with subsequent formation of a blood clot (thrombosis). The clotting
cascade is a series of serine proteases activated on the platelet surface forming a fibrin
clot. The fibrin clot partially or completely blocks blood flow or can cause intermittent
obstruction [
127
]. When blood flow is blocked this reduces oxygen supply to the heart mus-
cle, termed ischemia. Early diagnosis and prompt treatment, often involving antiplatelet
agents, anticoagulants, and revascularization procedures such as balloon angioplasty and
stent implant, restore blood flow and reduce heart muscle damage, with improved survival.
In flow cytometry studies conducted on 263 angina patients, Zhang et al. found
increased expression of uPAR on circulating monocytes. In those patients, elevated uPAR
levels strongly correlated with other clinically used inflammatory biomarkers and clinical
instability [
42
]. Individuals with coronary artery disease (CAD) and with elevated suPAR
levels had a significantly increased all-cause mortality, including increased cardiovascular
mortality [
42
]. Additional studies in this field have identified suPAR as a pro-inflammatory
biomarker for increased risk for unstable coronary artery disease, but not for stable coronary
atherosclerosis [
43
,
44
]. Increased local production of suPAR in coronary vessels is also
associated with a dysfunctional endothelial cell layer in patients [
45
]. suPAR may allow
identification of patients with a higher risk of unstable plaque and plaque rupture in
coronary artery disease, potentially identifying patients who will benefit from earlier
interventions, such as stent implant or bypass surgery.
2.1.2. Acute Myocardial Infarction
An acute myocardial infarction (AMI), commonly known as a heart attack, occurs when
there is a complete and persistent obstruction of coronary blood flow to the
heart [127,128].
The
role of uPAR in the context of acute myocardial infarction (AMI) has drawn considerable
attention, particularly due to its association with inflammation, leukocyte adhesion, and
invasion. uPA/uPAR research is rapidly evolving in the cardiovascular field. Mouse
uPA-knockout studies have shed light on the role of uPA in unstable plaque rupture and
tissue infarction. One such study observed a significant reduction in myocardial infarct
areas in uPA-deficient (uPA
−/−
) but not uPAR
−/−
mice, when compared to wildtype mice,
indicating a pro-inflammatory and pro-necrotic function for uPA [
46
]. Moreover, uPA defi-
ciency (uPA
−/−
) protected against myocardial rupture, a complication of AMI [
46
]. Thus,
although uPA has thrombolytic, or clot-dissolving, pathways when used as a therapeutic,
in the circulation there is an apparent paradox in that reducing uPA reduces risk of unstable
plaque rupture and thrombosis and ischemic myocardial damage. These studies highlight
the varying functions of the uPA/uPAR complex in fibrinolysis and inflammation, with
a greater role for the uPA/uPAR complex in inflammatory cell responses in unstable and
inflamed atherosclerotic plaque.
In a study examining patients with AMI, uPAR expression was increased, but in stable
angina patients, uPAR was not increased [
47
]. In this study, researchers discovered that
targeting uPAR with monoclonal antibodies blocked monocytic adhesion as well as integrin-
mediated fibrinogen adhesion. This study further established the pro-inflammatory role of
Biomedicines 2024,12, 1167 11 of 44
uPAR in unstable vascular disease and indicates that uPAR is a potential target for prevent-
ing leukocyte adhesion and migration in areas of unstable plaque, arterial thrombosis, and
myocardial injury.
In another study, of 1314 patients presenting to the ED with suspected MI, suPAR
reliably predicted mortality after one year of presentation [
48
]. Further studies have
linked the levels of suPAR six hours post-cardiac arrest to neurological outcomes and
mortality [49].
2.1.3. Percutaneous Coronary Intervention (PCI)
Percutaneous coronary intervention (PCI) involves the insertion of an angioplasty
balloon catheter guided over a wire across an area of atherosclerotic plaque. The angioplasty
balloon is dilated to open an area of stenosis, arterial narrowing or occlusion, to restore
blood flow. Balloon angioplasty restores normal blood flow to areas of myocardium that
are ischemic, with reduced blood and oxygen supply. In most cases a stent is delivered
on the angioplasty balloon to the site of vessel occlusion. PCI in ST-elevation myocardial
infarction (STEMI) is the gold standard of care and has revolutionized the management of
this life-threatening condition, offering rapid and effective restoration of blood flow to the
heart and improving survival.
However, in contrast to the pro-inflammatory role of uPA in the formation of atheroscle-
rotic plaque, where invasive macrophages drive rupture and thrombosis, uPA, or urokinase
(UK), was in fact initially developed as a thrombolytic agent that was used to dissolve
coronary clots during heart attacks. UK (uPA) was used as a thrombolytic to dissolve clots
before coronary stent implants for primary PCI were available or tested for the treatment
of STEMI. In 1995, Mitchell and colleagues published a study on patients that revealed a
decrease in coronary thrombotic stenoses following the local infusion of 150,000 units of
UK administered over a 30 min period [
50
]. In their
in vitro
studies, they demonstrated de-
creased thrombus weight (66% decrease) compared to the control groups with no treatment
(25% decrease) when administering UK by direct infusion with a dispatch catheter.
In vivo
studies further validated the capacity for UK to localize to sites of thrombosis for up to 5 h.
Ultimately, the intracoronary infusion of UK resulted in a reduction in the amount of drug
required and a more localized treatment. A study involving 345 patients presenting with an
acute ST-elevation myocardial infarction (STEMI) caused by a high-grade thrombus were
given intracoronary UK prior to stenting. Researchers found more complete ST-segment
resolution with the UK group compared to saline treatment [
51
]. Additionally, the peak
CK-MB levels were lower in patients treated with UK. The incidence of bleeding with
UK was not significant when compared to untreated controls. A meta-analysis of five
randomized, controlled studies with a total of 761 subjects found that UK intracoronary
injection increased perfusion to the myocardium, improved cardiac function, and reduced
infarction size [52].
In the pursuit of enhancing thrombolytic therapy, researchers have explored innovative
strategies, one of which involves the utilization of urokinase-coated nanoparticles (UK-NPs).
According to Jin and colleagues, UK-NPs sustained uPA activity, exhibiting an enhanced
thrombolytic function over non-conjugated uPA in rabbit models [
53
]. Additional studies
have displayed similar outcomes, affirming the utilization of nanoparticle-conjugated
urokinase as a potential combined therapy for thrombolysis [
53
]. A recombinant form
of pro-uPA (known as Prolyse
®
; developed by Abbott Laboratories) has been developed
for managing thromboembolic disorders. However, this medication has not yet received
approval from the FDA [
54
]. These studies examining the role of UK as a fibrinolytic further
identify the complexity of the uPA/uPAR complex as both a therapeutic thrombolytic as
well as a potential pro-inflammatory mediator and marker for disease progression.
Thus again, the complex interplay between cell surface and plasma uPAR (suPAR)
levels and the progression of coronary artery disease (CAD) with inflammation and plaque
rupture illustrates the multiple roles of uPA and uPAR in vascular disease. The use of
uPA (UK) as a beneficial therapeutic, clot-dissolving thrombolytic appears at variance
Biomedicines 2024,12, 1167 12 of 44
with the increased inflammation of atheroma in vascular disease associated with increased
circulating uPAR and the potential to target the uPA/uPAR complex as a new treatment
paradigm. UK is used as a thrombolytic treatment in acute MI. However, elevation of
plasma suPAR concentration is associated with increased severity of CAD, and is now
considered a predictor of future adverse cardiac events [
55
]. uPAR, thus, has a pivotal role
in atherosclerosis progression and plaque rupture, guiding atherogenic cellular invasion
and proliferation. A rise in uPAR levels mirrors the release of suPAR from inflammatory
cells present at the site of atherosclerosis [
55
]. The regulation of this balance between the
pro-inflammatory functions of uPA/uPAR in driving cardiovascular disease (CVD) and the
therapeutic benefit of uPA as a fibrinolytic remains incompletely defined.
2.1.4. Viral Myocarditis
Viral infection, and specifically Coxsackievirus-B3 (CVB3) infection, can lead to my-
ocardial inflammation and damage, termed viral myocarditis. CVB3 myocarditis is charac-
terized by infiltration of inflammatory cells into the myocardium, resulting in cardiomy-
ocyte damage and cardiac dysfunction (heart failure). With viral infection there can be
direct damage to the myocardium by viral infection as well as damage induced by the
body’s response, a pro-inflammatory response. Both can increase cardiac damage, remodel-
ing, and cardiac dysfunction. There is thus a complex interplay between viral infections, the
host inflammatory response, and cardiac function. In mouse models, uPA-deficient mice
(uPA-genetic-knockout mice) were tested and found to be protected against remodeling of
cardiac tissue after viral infection, demonstrating reduced heart dilatation and failure after
CVB3 viral myocarditis [58,129].
2.1.5. Congenital Heart Block (CHB)
Maternal autoantibodies and the uPA/uPAR system are reported to affect fetal cardiac
health. Autoantibodies can cross the placental barrier, entering fetal circulation and trigger-
ing events that disrupt normal cardiac development, manifesting as structural abnormalities
and affecting the cardiac conduction system. CHB is a rare but serious condition, potentially
necessitating lifelong medical interventions [
130
]. Due to circulating maternal anti-SSA/Ro
and anti-SSB/La, the fetal heart can become susceptible to fibrosis due to a loss of healthy
cardiomyocytes rather than the removal of dead or apoptotic cardiomyocytes. This fibrosis
can cause progressive heart block [
130
,
131
]. In mouse studies, Park et al. demonstrated
that uPAR has a role in phagocytic clearing of damaged cells [
59
]. Flow cytometry of cell
cultures highlighted the localization of uPAR on the surface of apoptotic cardiomyocytes.
This suggested an interplay between anti-Ro binding and increased uPAR gene expression.
As a result, there is increased scar and collagen deposition causing heart block [
60
]. These
observations highlight the importance of continued research into the role of uPA and uPAR
in CHB, with the potential to improve diagnostic and therapeutic strategies, and ultimately
improve the care and outcomes of affected infants and their families.
2.1.6. Chronic Heart Failure
The incidence of heart failure has increased, and heart failure is associated with high
mortality. An accurate prognostic indicator for the detection of heart failure would be of
benefit. As discussed above, suPAR is the cleavage product of membrane-bound uPAR
induced by circulating immune cells, and levels of suPAR mirror increased inflammatory
responses. In prior work on chronic heart failure, suPAR concentration was strongly
correlated to increased mortality in patients [
56
]. This strong correlation between suPAR
levels and mortality highlights the role of uPAR and immune activation as a method
for identifying the risk of heart failure and progression. By monitoring suPAR levels,
clinicians gain a powerful tool to gauge the intensity of the inflammatory response, which
is intrinsically linked to the progression of heart failure.
Biomedicines 2024,12, 1167 13 of 44
2.1.7. Cardiovascular Risk Stratification
In a 10-year prospective cohort study of 1951 apparently healthy patients, it was
discovered that suPAR, together with markers already in clinical use, e.g., hs-CRP and
NT-proBNP, predicted cardiovascular death. In this study, suPAR was suggested as a
prognostic biomarker for cardiovascular death in otherwise healthy patients [57].
2.1.8. Acute Ischemic Stroke
In a large population-based cohort study of 6103 subjects, among the key observations,
a compelling correlation emerged, drawing a direct link between heightened uPAR levels
and the presence of carotid plaques in individuals. This finding is of great clinical relevance,
as carotid plaques serve as harbingers of potential local occlusive and embolic atheroscle-
rotic vascular thromboses, with risk of stroke. This study also went on to uncover a height-
ened risk of ischemic stroke among patients who exhibited increased uPAR levels [
61
,
132
].
This study not only adds to the growing body of evidence linking uPAR to vascular pathol-
ogy, but also underscores its clinical significance as a potential tool for improved risk
assessment and personalized management in the realm of cerebrovascular health.
In studies exploring interventions to modulate suPAR levels, it has been observed
that individuals on extended courses of beta-blocker treatment exhibit decreased suPAR
levels, as demonstrated in carotid endarterectomy biopsy samples [
63
]. This intriguing
link prompts further investigation into the precise mechanisms by which beta-blockers
influence suPAR, with the potential to unveil novel therapeutic strategies and diagnostic
applications [63].
An acute ischemic stroke, termed a cerebrovascular accident (CVA), is a medical
emergency characterized by acute occlusion of cerebral blood vessels causing disabling
neurological deficits. Rapid intervention is necessary to restore blood flow and minimize
long-term brain damage. Early thrombolytic treatment or mechanical interventions to
dissolve or remove blood clots, when given early after the onset of the CVA, improves
outcomes, but with an associated increased risk of hemorrhage. uPA (UK) has been
employed in the treatment of acute ischemic stroke, as it can help dissolve blood clots in
the cerebral vasculature, potentially improving neurological outcomes [61].
The PROACT trial (Prolyse in Acute Cerebral Thromboembolism) is a pivotal trial
in the treatment of acute thromboembolic strokes. This phase II randomized human trial
indicated that the combination of pro-urokinase and heparin resulted in an improved
recanalization efficacy for individuals with acute thromboembolic strokes [
62
]. This break-
through not only suggests a potential therapeutic avenue but also underscores the critical
importance of addressing the intricate mechanisms underlying thromboembolic strokes.
The improved recanalization efficacy observed in the PROACT trial promises to enhance pa-
tient recovery and reduce neurological sequelae [
62
]. Additional clinical trials utilizing uPA
as a thrombolytic agent have demonstrated positive results, consistently demonstrating
enhanced recanalization, improved functional outcomes, and rates of cerebral hemorrhage
that mirror those seen with recombinant tissue plasminogen activator (rtPA) [61].
Here, again the role of suPAR as a diagnostic indicator for unstable disease demon-
strates a correlation and a potential role for uPA and suPAR in disease progression. A
pro-inflammatory role for uPAR in cerebrovascular disease appears superficially to be con-
tradictory to the use of uPA as a thrombolytic. However, these studies indicate correlations,
they do not prove cause. With these various studies, the question arises as to whether
the local inflammatory responses driven by the uPA/uPAR complex represents an early
disease stage or is limited to local events in the vasculature or may represent a secondary
or rebound response. In summary, the use of higher doses of uPA (UK) to dissolve clots has
therapeutic benefit, but may also represent a non-physiological response to the high dose
of uPA, whereas local uPAR activity may represent a physiological, pathological tissue
response. Analyses of local and individual cellular responses to uPA/uPAR activation will
provide new insights into the physiological roles of uPA/uPAR in inflammatory responses
in contrast to the use of uPA (UK) as a thrombolytic.
Biomedicines 2024,12, 1167 14 of 44
2.1.9. Angiogenesis
Angiogenesis is the formation or growth of new blood vessels from existing ones.
Angiogenesis plays a crucial role in normal physiological and also pathological conditions,
such as wound healing, tissue repair, and tumor growth, and is tightly regulated by
a complex interplay of pro-angiogenic and anti-angiogenic factors [
133
]. uPA is directly
involved in the release of pro-angiogenesis growth factors such as FGF-2 and VEGF, playing
a key role in endothelial cell proliferation [134].
uPA binding to uPAR on the surface of vascular smooth muscles induces an intricate
intracellular signaling cascade which ultimately results in increased migration of cells [
64
].
Downstream of uPA/uPAR binding, the activation of the JAK-STAT kinase pathway results
in platelet-derived growth factor receptor-beta (PDGFR-
β
) internalization via the LR11
receptor [65].
Studies exploring angiogenesis in human umbilical vein endothelial cells have also
demonstrated impaired VEGF-mediated signaling with the knockdown of uPAR. This is
further supported by studies in uPAR-deficient mice, where incomplete angiogenesis was
observed [
66
]. Angiogenesis and the growth of blood vessels is necessary to support tumor
growth, supplying blood, oxygen, and nutrients to the tumor. In the next sections, we
will discuss studies linking cancer progression to uPAR (Section 2.2) and subsequently
discuss the role of uPAR in inflammation and disease (Section 2.3). One study investigating
endothelial colony-forming cells with properties of endothelial progenitor cells found that
whole-uPAR localization in caveolae is required for further angiogenic proliferation [135].
2.2. Cancer
The level of suPAR in the blood has been extensively studied as a marker for cancer
progression, metastases, and abnormal immune responses to cancer in patients (Table 1).
Aberrant immune responses are now understood to drive tumor growth and invasion,
both in local tumor- and stromal-associated mononuclear cells and in systemic immune
cell responses. Increases in suPAR have been detected in patients with metastatic can-
cer. Based on these observed increases in uPAR with metastatic cancer, new approaches
that target uPAR have been developed using small molecules, selective anti-uPAR anti-
bodies, uPA-mediated toxin activity, uPAR-targeting nanoparticles designed to transport
chemotherapeutic agents for local delivery to the tumor, and one virus-derived immune-
modulating serpin. Several studies have reported reduced cancer invasion and metastasis
with these new uPAR-targeting reagents. Here, we describe a variety of cancers for which
suPAR and uPAR have been identified as markers for cancer progression and metastasis
and as new therapeutic approaches.
2.2.1. Cancer Metastasis
Recent studies have indicated a pivotal role for uPAR in malignant tumor invasion
and metastasis [
67
]. High uPAR expression is consistently observed in solid tumors.
uPAR is intricately involved in extracellular matrix degradation, tumor angiogenesis,
cell proliferation, and apoptosis, as well as multidrug resistance (MDR) in cancer cells
(Figures 2and 3). Here, we discuss specific malignancies in which associated changes in
uPAR expression have been identified in breast, lung, prostate, head and neck, leukemia,
ovarian, and pancreatic cancers (Table 1).
2.2.2. Breast Cancer
Breast cancer is the most common cancer among females and the incidence is estimated
at 2.3 million new cases globally each year [
68
,
136
]. Recent studies have demonstrated an
increase in the expression of uPA, uPAR, and plasminogen activator inhibitor type-1 (PAI-1)
in breast cancer and bony metastases. Studies have analyzed tissue samples from various
breast carcinomas, metastases, and normal breasts, identifying associated changes in uPAR.
The majority of the tumors analyzed had moderate uPA mRNA levels and variable uPAR
and PAI-1 mRNA levels, primarily localized in epithelial tumor cells. Malignant tumors
Biomedicines 2024,12, 1167 15 of 44
exhibited significantly increased uPAR mRNA expression as well as slightly increased uPA
and PAI-1 mRNA expression on comparison to benign breast tissue [68,69,71].
A specific subset of breast carcinomas expressing human epidermal growth factor
receptor type 2 (HER2-positive) display high uPAR expression. HER2-positive breast
cancer is known for an aggressive and metastatic nature, although treatment with tar-
geted therapies is available and effective with early treatment. A microarray analysis of
advanced breast cancer lesions identified interactions between HER2 and uPAR, along
with a wide array of downstream molecules. The HER2-positive/uPAR-positive subtype
demonstrates increased expression of transcriptional factors, contributing to the aggressive
nature of this tumor. This overexpression of uPAR may suggest a reason for resistance to
treatment in HER2-positive cancers and the potential for uPAR to provide a new thera-
peutic
target [68,69,71].
Antibody drug constructs that target uPAR are in development as
potential treatments for triple-negative breast cancer [71].
2.2.3. Lung Cancer
Bronchogenic lung carcinoma is the leading cause of cancer-related deaths in the
United States. Smoking is the primary cause of lung malignancies, but it is further exac-
erbated by exposure to environmental factors such as asbestos and polycyclic aromatic
hydrocarbons [
137
]. Two large-cell lung carcinoma strains in humans, one highly metastatic
(strain 95D) and one less metastatic (strain 95C), were evaluated for their
in vitro
and
in vivo
invasive and metastatic potentials. Strain 95D exhibited greater invasiveness than strain
95C. The expression levels of uPA and tPA, uPAR, and PAI-1 and PAI-2 were assessed by
RT-PCR and immunohistochemical staining in the high- and low-metastatic strains [
72
].
The high-metastatic strain 95D displayed elevated uPA and uPAR expression and reduced
tPA and PAI-2 levels compared to the low-metastatic strain 95C. PAI-1 expression was
similar in both strains. Monoclonal antibodies targeting uPAR effectively decreased the
invasive potential of strain 95D cells
in vitro
. This suggests that uPAR plays a significant
role in the invasiveness of these lung carcinoma strains [72].
Small-cell lung carcinoma (SCLC) is a subset of highly proliferative lung carcinoma
that presents with rapid growth, early metastasis, and poor prognosis [
73
]. Studies have
also investigated the role of uPAR in six different SCLC cell lines. The findings revealed
that a subpopulation of cells in these SCLC lines expressed uPAR. These uPAR-positive
cells exhibited resistance to multiple drugs, with high clonogenic (survival) activity, and
also co-expressed CD44 and MDR1, which are markers of potential cancer stem cells.
This work suggests that uPAR-positive cells represent a functionally important subset
of chemotherapy-resistant cancer cells in SCLC and could be valuable targets for more
effective therapeutic interventions [74].
The overexpression of uPAR in a range of lung cancer subtypes further suggests a role
for uPAR as a suitable target for chemotherapy. Peptide sequences of the amino-terminal
domain of uPA have recently been proposed as an efficient method to target uPAR in
lung cancer cells. Investigation of the uPAR-targeting U11 peptide conjugated with a
pH-sensitive doxorubicin and curcumin combination has demonstrated a synergistic anti-
tumor effect on cultured non-small-cell lung cancer cells
in vitro
and
in vivo
in a mouse
model [75].
2.2.4. Prostate Cancer
Prostate cancer (PCa) is a heterogeneous disease involving genetic, environmental,
and social influences, commonly affecting men aged 45 to 60 [
138
]. A recent analysis of
uPA, uPAR, and PAI-1 in patients undergoing radical prostatectomy for cancer, identified
increased levels of these proteins. An immunohistochemical analysis of tissue samples from
3121 patients revealed overexpression of uPA, uPAR, and PAI-1 in varying proportions [
70
].
Overexpression of uPAR was associated with aggressive PCa. All three markers were
linked to an increased risk of cancer recurrence as detected on biochemical assay. The
likelihood of recurrence increased with a higher number of overexpressed markers. A
Biomedicines 2024,12, 1167 16 of 44
decision curve analysis indicated that the inclusion of data for uPA, uPAR, and PAI-1
improves clinical decision-making when compared to standard clinical–pathological fea-
tures [
70
]. A follow-up analysis using tissue microarrays and immunohistochemistry found
that overexpression of uPAR was present in more than half of primary PCa tissues and over
90% of lymph node metastases, but not in normal or benign tissues. The overexpression
of uPAR was further associated with higher Gleason scores, indicating a link between
their expression and tumor differentiation, even in patients with favorable pathological
characteristics [76].
The high prevalence of uPAR in PCa suggests its potential as an effective therapeutic
intervention. Noscapine, an anticancer compound, has shown promise in inhibiting tumor
growth in various types of cancers, but its effectiveness has been limited by bioavailability.
With recent developments in nanoscale delivery systems, researchers have utilized the
human-type amino-terminal fragment of uPA as a natural ligand for uPAR to improve
drug targeting and availability [
77
]. uPAR-targeted nanoparticles significantly enhanced
the intracellular accumulation of noscapine in PCa, leading to a more potent inhibitory
effect compared to free noscapine [
77
]. Further research identified the effects of uPAR
knockdown using small interfering RNA (siRNA) or combined treatment with microRNA
(miRNA) and siRNA. Systemic treatment with tumor suppressor gene MIR143 in polymeric
nanoparticles inhibits tumor growth in mice with subcutaneous PC-3 tumor xenografts.
The nanoparticle-mediated delivery of MIR143 significantly downregulated uPAR protein
levels without affecting mRNA levels, indicating translational inhibition [78].
2.2.5. Head and Neck Cancer
Head and neck cancers are notorious for their late discovery, as most patients initially
present with lymph node metastases [
139
]. In comparison to more common cancers,
head and neck squamous cell carcinoma (HNSCC) has a poorly described mechanism
for invasion and metastasis. Therefore, identification of biological markers may enable
earlier diagnosis and the potential for new treatment targets. The plasminogen activator
system, especially uPA and uPAR, plays a significant role in head and neck squamous cell
carcinoma (HNSCC). The overexpression of uPAR and PAI-1 (SERPINE1 gene) is reported to
contribute to increased tumor cell migration, invasion, and metastasis, ultimately resulting
in a poor prognosis. Both uPAR and PAI-1 are associated with the induction of epithelial-to-
mesenchymal transition, the acquisition of stem cell properties, and resistance to anticancer
agents [79].
2.2.6. Leukemia
Leukemias are hematologic cancers with a wide prevalence in both pediatric and
adult patients. The major sub-classifications of leukemia encompass chronic lymphocytic
leukemia (CLL), acute myeloid leukemia (AML), chronic myeloid leukemia (CML), and
acute lymphoblastic leukemia (ALL) [
140
]. While the primary focus of uPAR analysis has
been on the effect of the translated protein, recent research has indicated that transfection of
uPAR 3
′
UTR in AML tumor cells affects the expression of pro-tumoral factors, cell adhesion,
and migration. These findings demonstrated that uPAR 3
′
UTR-recruited microRNAs can
modulate multiple intracellular functions by targeting various transcripts. Additionally,
variants of uPAR transcripts with 3
′
UTR regions are detected in U937 leukemia cells, with
higher uPAR expression [
83
]. These findings suggest that uPAR mRNA variants play an
important role in the progression of AML.
2.2.7. Ovarian Cancer
Ovarian cancer is a leading cause of death among women with gynecologic cancer.
Similar to HNSCC, as discussed previously, ovarian cancer is typically diagnosed at a
later stage, leading to worse outcomes [
139
,
141
]. A major driver in ovarian cancer (OC) is
lysophosphatidic acid (LPA), which stimulates both cellular migration and proliferation. A
Biomedicines 2024,12, 1167 17 of 44
recent analysis of uPAR expression in ovarian epithelial cancer cells stimulated with LPA
revealed an increase in uPAR aggregation and uPA binding [80].
More recently, the Kazal-type serine protease inhibitor-13 (SPINK13) gene was re-
ported as associated with decreased mortality in OC patients. Analysis of the SPINK13
molecular pathway identified its role in inhibiting the expression of uPA, further emphasiz-
ing the significant role of uPA in OC [
81
]. Similarly, subtypes of ovarian cancer, such as
leptin-induced OC cell invasion, rely heavily on uPA.
2.2.8. Pancreatic Cancer
uPAR expression has been reported to be highly expressed in pancreatic cancer, more
so than many other cancers. Treatment of a xenograft model of pancreatic cancer cells
in SCID mice with the virus-derived immune-modulating serpin, Serp-1, demonstrated
significant reduction in tumor growth along with altered stromal and regulatory T cell
immune responses [
82
]. Serp-1 requires uPAR for normal immune-modulating functions
(Table 2).
Table 2. Analysis of Serp-1 and PEGSerp-1 treatments targeting uPAR.
Therapeutic Inflammatory Disorder Treatment Effect/Outcome Subjects Studied Reference
Serp-1
Atherosclerotic Plaque
Acute Coronary
Syndromes with Stent
Implant
(1) Multicenter phase II Clinical Trial in
USA and Canada. Mechanism extensively
studied. Reduced markers of cardiac
damage; MACE = 0. No neutralizing
antibodies.
(2) Preclinical—reduced intimal
hyperplasia.
(1)
Human clinical trial,
randomized
dose-escalating trial at 7
sites in Canada and US
(2)
Preclinical—rabbits
[142,143]
Serp-1 Angiogenesis Serp-1 reduced angiogenesis in
chorioallantoic membrane. Chicken [144]
Serp-1 Pancreatic Cancer Treated pancreatic xenografts had
decreased growth with altered myeloid
cell responses.
SCID mice (severe combined
immunodeficiency model) [82]
Serp-1 Corneal Abrasion Corneal wound healing was enhanced by
reducing immune cell infiltration, fibrosis,
and neovascularization. Mice [123]
Serp-1 Uveitis Inflammation was decreased with AAV
expression Serp-1. Given intraocularly. Mice [124,145]
PEGSerp-1 Diffuse Alveolar
Hemorrhage
PEGSerp-1 reduced lung hemorrhage and
inflammation—effective when given as a
delayed treatment. Mice [142,143]
Serp-1
Aortic, Renal, and Cardiac
Allograft Acute and
Chronic Transplant
Rejection and Vasculitis
Serp-1 reduces chronic renal and aortic
allograft vascular inflammation.
Serp-1 lost activity in uPAR-KO mouse
aortic allografts.
Mice, rats [146–149]
Serp-1 Lethal Viral Infection Serp-1 reduces vascular inflammation in a
lethal gammaherpes MHV68 infection. Mice [150]
PEGSerp-1 SARS-CoV-2 Infection
PEGSerp-1 reduced immune
coagulopathic lung damage after
SARS-CoV-2 infection and reduced
mortality and vascular inflammation in
gammaherpes-infected mice.
Mice [151]
Serp-1 Hyperlipidemic
Atherosclerotic Plaque
Serp-1 infusion by osmotic pump
significantly reduced carotid plaque after
carotid cuff injury.
Mice—ApoE−/−mice with
carotid cuff injury [152]
Serp-1 Giant Cell Arteritis
Reduced inflammation in human
xenograft giant cell arteritis biopsy
implants in SCID mice with PBMC
infusions.
Mice [153]
Serp-1 Inflammatory Arthritis
(Rheumatoid Arthritis) Serp-1 reduced joint inflammation in
collagen-induced arthritis. Rabbits [154]
PEGSerp-1 Inflammatory Colitis Colon damage was reduced in dextran
sulfate sodium-induced colitis Mice [155]
Biomedicines 2024,12, 1167 18 of 44
Table 2. Cont.
Therapeutic Inflammatory Disorder Treatment Effect/Outcome Subjects Studied Reference
Serp-1 Inflammation in Wound
Healing
Serp-1 given at sites of skin wounds
improved healing rates in mice and
reduced inflammation
Activity blocked by anti-uPAR antibody.
Mice [156]
Serp-1 Spinal Cord Injury Local infusion of Serp-1 after spinal cord
injury in rats reduced inflammation and
neuronal damage. Rats [157]
PEGSerp-1 Duchene Muscular
Dystrophy
PEGSerp-1 reduced inflammation in
mouse diaphragm and skeletal
musculature. Mice [158]
2.3. Inflammation
2.3.1. Chronic Inflammation
Chronic inflammation (CI) is characterized by persistent, low-grade innate immune-
cell activation that significantly impacts an individual’s quality of life and contributes to the
development of disease. Currently, CI lacks well-defined diagnostic criteria, often relying
on markers associated with acute inflammation, despite its long-lasting, chronic nature,
spanning years [
159
]. CI is associated with chronic diseases such as DM, CVD, arthritis,
and renal disease, and even long-term infections such as tuberculosis. The transition to
CI inflammation typically occurs when the body struggles to repair and resolve an initial
acute inflammatory response and associated damage leading to ongoing excess systemic
immune-cell activation and tissue damage. Recent research has investigated the response of
immune cells under inflammatory stimuli and has revealed an increase in suPAR (Table 1).
Blood suPAR demonstrates a strong correlation with inflammation as well as a strong
association with increased circulating immune cells [
55
]. The role of suPAR is further
supported by the fact that suPAR shares common risk factors for age-related diseases as
mentioned in this review, including atherosclerotic coronary and carotid artery disease and
COPD [31,55].
Interestingly, in contrast to CRP, suPAR plasma levels remain unaffected by circadian
fluctuations, with relatively steady expression even during periods of acute stress [
84
]. For
instance, serial measurements demonstrated a mere 15% average increase in suPAR levels,
in stark contrast to the 365% rise in hs-CRP levels in the setting of an MI. Overall, suPAR
was reported to be a better prognostic indicator for adverse outcomes than hs-CRP [55].
Macrophage invasion of atherosclerotic plaque in coronary arteries leads to plaque
rupture and acute thrombosis in MI, as noted above in Section 2.1. Unstable coronary
plaque is, thus, an example of an inflammatory disorder and the uPA/uPAR complex
is associated with activation of matrix metalloproteinases (MMPs) that allow for tissue
breakdown and inflammatory cell invasion into plaque and into the arterial wall. suPAR
appears to outperform hs-CRP in terms of prognosis for hospital mortality for individuals
with coronary artery disease and has been established as an independent predictor for
future adverse cardiac events [
55
–
57
,
84
]. The utilization of suPAR as a diagnostic marker
for CI represents an exciting avenue of research.
2.3.2. Rheumatoid Arthritis
Rheumatoid arthritis (RA) is a complex autoimmune disease associated with various
genetic and environmental factors. RA typically begins unilaterally in a peripheral joint and
subsequently progresses to proximal joints, resulting in cartilage loss and bone erosion [
160
].
RA often has a delay in initial diagnosis. Extensive analysis has revealed that major histo-
compatibility complex, class II, DR beta 1 (HLA-DRB1) carries a segment of five conserved
amino acids in the hypervariable regions strongly associated with RA development.
Earlier investigations on the assessment of circulating suPAR in RA demonstrated ele-
vated suPAR levels in RA patients compared to their healthy counterparts [
85
,
86
,
161
]. These
suPAR levels exhibited a direct correlation with the number of inflamed joints, even among
Biomedicines 2024,12, 1167 19 of 44
patients with limited disease activity [
85
]. In a recent study involving
252 patients
from
a Swedish prospective observational cohort with early RA, serum suPAR was evaluated
using an enzymatic immunoassay at disease onset, as well as after 3 and 36 months [
160
]. In
contrast, tPA is generally downregulated in RA. Based on current research, suPAR detection
has the potential to serve as an adjunctive tool for the screening and monitoring of disease
progression in patients with RA [85,86,160–162].
2.4. Critical Care
2.4.1. SIRS—Systemic Inflammatory Response Syndrome
SIRS (systemic inflammatory response syndrome) is associated with high morbidity
and mortality, with or without an antecedent infectious process (sepsis). A study of
132 patients
investigated the utility of suPAR as a biomarker for SIRS, and the capacity for
suPAR to differentiate SIRS from bacteremia in comparison with other biomarkers. suPAR
was found to have an AUC (area under receiving operator curve) of 0.726 in differentiating
SIRS from sepsis; an acceptable biomarker for differentiation. When combined with other
current clinically used biomarkers, procalcitonin and IL-6, the AUC rose to 0.804, which
is considered a good classification level for differentiating SIRS from sepsis. Interestingly,
initial suPAR concentrations were significantly higher in patients who later died within
28 days,
supporting its utility as a prognosticator for overall inpatient mortality upon initial
presentation [163].
2.4.2. Sepsis
Sepsis is defined as a systemic inflammatory response syndrome (SIRS) in response to
a confirmed infectious process such as a bacteremia or viremia. Sepsis is, thus, considered
a form of SIRS but with an identified source of infection as the culprit. Sepsis is a serious
condition produced by infections with bacteria and also viruses, fungi, and parasites.
In sepsis, there is an extreme immune response causing systemic symptoms of fever,
tachycardia, tachypnea, and shock, with associated high mortality. Sepsis is a major cause
of morbidity and mortality in newborns. A meta-analysis conducted in 1959 patients over
six studies concluded that suPAR had superior specificity for differentiating neonatal sepsis,
by clinical definitions, from non-septic neonates when compared to the commonly used
procalcitonin and CRP levels. Furthermore, the diagnostic odds ratio was 117, indicating
high efficiency and precision. The positive likelihood ratio was 14, indicating that the
suPAR level is 14 times higher in neonates with sepsis than those without. Finally, the
negative likelihood ratio was low at 0.12. This means that if the suPAR level is negative,
the probability of neonatal sepsis is 12%, allowing confidence in ruling neonatal sepsis.
Interestingly, in neonatal patients, and in concordance with the adult patients, the measured
levels of suPAR on the first day helped in predicting mortality in late-onset neonatal
sepsis [88–90]. Finally, suPAR is also useful as a prognostic biomarker in adult sepsis [91].
2.5. Wound Healing and Tissue Repair
Tissue repair is a complex process that entails the orchestrated activation of diverse
intracellular mechanisms that facilitate cellular migration, proliferation, and differentia-
tion [
164
,
165
]. These intricate pathways are subject to a very rigorous regulation, as their
dysregulation can predispose individuals to a wide spectrum of infections and malignan-
cies. The skin’s dermal layers have both normal structural cells but also intrinsic or local
tissue immune cells that can act as a primary response to tissue damage, similar to the
central nervous system. As noted, uPAR plays a pivotal role in cellular migration, with
increased prevalence in injured tissues (Table 1).
uPAR has been identified as playing a major role in both epidermal and dermal
wound healing environments. Recent investigation of overexpressed demogelin 2, an
important regulator of cell survival and proliferation, identified an increased release in
uPAR expression [
166
]. Leveraging the increased activity of the uPAR signaling pathway
in the context of wound healing holds promise for the identification of pharmacological
Biomedicines 2024,12, 1167 20 of 44
interventions that can expedite tissue repair. Recent investigations have unveiled the
significance of spermidine (SPD), an abundant natural polyamine essential for endothelial
cell proliferation and angiogenesis, that strongly promotes the activation of uPAR when
administered both systemically and topically [
167
,
168
]. Initial
in vitro
scratch wound assay
studies in tissue cultures provided evidence of spermidine/uPA/uPAR wound healing-
promoting properties.
Subsequent
in vivo
analyses were conducted in a murine model of skin-wound repair
to determine the effectiveness of topical SPD treatment. Notably, mice treated with SPD
exhibited substantial enhancements in the healing. Examination of the wounds not only re-
vealed heightened activity within the uPA/uPAR signaling pathway but also demonstrated
increased expression of interleukin-6 (IL-6) and tumor necrosis factor (TNF), two pivotal
proteins associated with both inflammation with potential damage as well as with tissue
regeneration [
168
]. Further research is needed to better understand the role of uPA/uPAR
in wound healing as well as identify novel promoters of this pathway.
2.6. Neurology
uPAR has also been reported to have a role in neuronal damage and repair (Table 1).
The CNS also has an intrinsic immune system that includes glial and astrocyte cellular
actions. Through immunohistochemical analysis of necrotic brain lesions and focal cerebral
infarcts, continuous uPAR expression was identified up to four days post-injury, with a
peak at the 12 h mark [111].
In a study that investigated the expression of uPAR in the murine central nervous
system (CNS) during inflammatory responses, suPAR was identified in the CSF in various
diseases or pathologies with increases in the CSF in patients with HIV dementia, a chronic
neuroinflammatory disease [
24
,
111
,
112
]. There is significant upregulation of uPAR at both
mRNA and protein levels in microglial cells during acute intracerebral lipopolysaccharide
(LPS) exposure. Expression of uPAR in neuronal cell lines with subsequent binding to
vitronectin, a protein located predominantly in the ECM, have shown to cause extensive
changes in cellular morphology and actin cytoskeleton formation [24].
uPA expression and activity are prominently increased during chronic neurodegenera-
tion, suggesting potential proteolysis-independent roles for uPAR in acute disease. In both
kainate and LPS challenges, uPAR was significantly upregulated, whereas uPA was not
altered. Further investigations have found a correlation between suPAR levels in the CSF
with the presence of any form of CNS inflammation, e.g., not with specific pathological
conditions, as highlighted by the diffuse staining found on histology [111].
Various kinds of inflammation are predicted to prompt release of suPAR from mi-
croglial cells membranes. For instance, the CSF of patients with human immunodeficiency
virus (HIV)-associated dementia had elevated levels of suPAR, yet specific demyelinating
pathologies such as multiple sclerosis or Guillian–Barre syndrome did not show altered
suPAR levels [
112
]. The determination of resident versus reactive resident microglia and
invasive systemic inflammatory macrophage and lymphocytes that express suPAR as a
response to neuroinflammation remains under investigation.
PAI-I is upregulated in a variety of neurodegenerative states including Parkinson’s
disease and Alzheimer’s disease (AD). Plasmin cleaves and degrades
α
-synuclein, and
α
-synuclein upregulates PAI-1. It has been proposed that an excess of PAI-1 in the brains
of PD patients prevents plasmin-induced clearance of
α
-synuclein aggregates, which is
corroborated by the fact that elevated PAI-1 levels are associated with worse outcomes
for PD patients. Finally, PAI-1 is downregulated, with associated upregulation of tPA
and uPA, during exercise in patients with AD [
169
]. As suPAR is more bioavailable than
uPAR, it is reasonable to hypothesize that suPAR may contribute to plasmin-associated
alpha-synuclein disease states, providing a biomarker.
In the assessment of stroke risk, suPAR levels were elevated in patients with symp-
tomatic carotid artery plaques in a study on 162 patients, supporting the use of suPAR as a
biomarker for unstable carotid artery atherosclerosis at risk of thrombotic occlusion [
114
].
Biomedicines 2024,12, 1167 21 of 44
Additionally, suPAR was found to be superior to CRP in specificity for discrimination
between vertebral osteomyelitis and other neurodegenerative spinal diseases in a 36-person
study [114].
2.7. Infectious Disease
uPAR has been associated with infectious disease, specifically, HIV, SARS-CoV-2, and
tuberculosis (Table 2).
2.7.1. HIV
In addition to the studies examining bacterial sepsis in critical care units, increased
uPAR expression has also been detected in other infections. Speth and colleagues were the
first to identify increased levels of uPAR expression on the surface of lymphocytes and
monocytes with HIV infection [92]. Based on these findings, another group of researchers
found a strong correlation between prognosis of HIV-1 and blood suPAR levels, similar to
the prognostic value of the CD4+ count and viral load, and thus, survivability [93].
2.7.2. COVID-19 Pneumonia
SARS-CoV-2 infection is the coronavirus infection that caused the recent coronavirus
pandemic (COVID-19). CoV-2 causes acute lung injury and acute respiratory distress
syndrome, stemming from severe inflammatory pneumonia and associated immune and
thrombotic vascular damage. It has been proposed that dysregulation in the uPA/uPAR
system may induce some of the CoV-mediated inflammatory damage. suPAR levels have
been investigated in patients with COVID-19, with suPAR levels reported as a marker
for increased risk for intensive care admission. The Ly-6/uPAR family of receptors have
been proposed as sites activated by the SARS-CoV-2 S protein, increasing cellular infection
by the virus. uPA/uPAR may also provide a potential therapeutic target for treating the
dysregulated immune and thrombotic responses in COVID-19 [170,171].
The SAVE-MORE (suPAR-Guided Anakinra Treatment for Validation of the Risk and
Early Management of Severe Respiratory Failure by COVID-19) trial examined 606 patients
randomized to a phase 3 controlled trial. Initially, 1060 patients were enrolled; however,
those with suPAR levels less than 6 ng/mL were excluded. Based upon suPAR stratification,
patients treated with anakinra experienced a survival benefit with a 3.9% 28-day mortality,
markedly increased when compared to the 8.7% mortality at 28 days in control subjects.
Thus, suPAR has shown efficacy in the improvement of patient selection criteria as a useful
biomarker in patient stratification to assist patients with COVID-19 infections treated with
anakinra [116].
Of those who were excluded due to not reaching threshold suPAR levels
(i.e., <6 ng/mL),
a
post hoc analysis revealed that only 2.9% of those patients progressed to respiratory failure
or death, yet again highlighting the importance of suPAR as an early inpatient biomarker
and major predictor of impending intrahospital morbidity and mortality [116].
In a mouse-adapted SARS-CoV-2 (MA30) model, PEGSerp-1 treatment reduced weight
loss, clinical symptoms, inflammation, and damage to lung and vascular tissue
(Table 2) [151].
Of particular interest, tissue uPAR detected by IHC was reduced by PEGSerp-1, but PCR
(polymerase chain reaction) analysis indicated increased uPAR gene expression.
2.7.3. Tuberculosis
In a community study, individuals with an active infection of Mycobacterium tuberculosis
had significantly increased levels of suPAR using ELISA detection assays [
94
]. After an
8-month treatment period, suPAR levels decreased to levels of tuberculosis (TB)-negative
subjects. Therefore, suPAR may provide a biomarker for treatment efficacy in TB.
Biomedicines 2024,12, 1167 22 of 44
2.8. Nephrology
2.8.1. Acute Kidney Injury (AKI)
suPAR was elevated in 3827 patients undergoing coronary angiography, 250 patients
undergoing cardiac surgery, and upon admission in 692 critically ill ICU patients. Those
within the upper quartile for suPAR levels had an increased risk of acute kidney injury
(AKI) and death at 90 days across all three cohorts. Mice studies performed by the in-
vestigators revealed that mice overexpressing suPAR and mice given suPAR when given
contrast for imaging had greater pathologic evidence of AKI, energetic demand, and mi-
tochondrial superoxide generation than wildtype mice. Pre-treatment in these mice with
a monoclonal uPAR antibody reduced AKI in mice overexpressing uPAR, normalizing
oxidative stress [102].
2.8.2. Focal Segmental Glomerulosclerosis
Focal segmental glomerulosclerosis (FSGS) causes roughly 20% of all glomerular dis-
ease and represents a major cause of end-stage renal disease (ESRD) requiring hemodialysis.
Two-thirds of all patients with FSGS will have elevated suPAR levels (using 3.0 as the cutoff),
which is significantly higher than all other glomerular diseases. Additionally, higher suPAR
levels were predictive for FSGS recurrence following kidney transplantation.
54 patients
with FSGS were found to have elevated suPAR levels when compared to patients with
glomerulonephritis and healthy control subjects. The optimal cut-off value for diagnosis
of FSGS using suPAR was found to be 4.644 ng/mL, with a sensitivity and specificity
of 0.91 and AUC of 0.946, providing an excellent diagnostic test for FSGS. The authors
supported suPAR as a biomarker for a variety of purposes, some of which will require
more randomized controlled trials for individualized immunosuppressive therapy in FSGS
recurrence post-transplant, ESRD prediction, and diagnosis of other glomerular diseases
when kidney biopsy is not possible. SuPAR may be most useful when combined with other
established biomarkers, such as anti-PLA2R antibodies [103].
2.9. Gastroenterology
uPAR has also been examined as a marker for severe gastrointestinal disorders such
as pancreatitis, hepatitis, and inflammatory bowel disease (Table 1).
2.9.1. Acute Pancreatitis
Acute pancreatitis is inflammation of the pancreas leading to increased capillary
permeability. Increased capillary leak is considered an antecedent to non-septic SIRS
with potential to cause organ failure and multi-organ distress syndrome. The severity of
pancreatitis ranges from mild pancreatitis, causing local complications without signs of
organ failure; to moderate, showing signs of organ failure recovering within 48 h; and
finally, to severe, leading to organ failure persisting past 48 h [
172
]. Given its potential
for morbidity and mortality, predicting the type of pancreatitis is important for planning
treatment modalities.
Researchers conducted a study on 225 hospitalized patients with acute pancreatitis.
Of those, 75 had severe acute pancreatitis (SAP), 75 had moderate–severe acute pancreatitis
(MSAP), and 75 had mild acute pancreatitis (MAP). Another 75 healthy patients served as
controls. Serum samples of suPAR were taken 24 h after admission. suPAR could signifi-
cantly differentiate SAP from healthy controls with high diagnostic accuracy
(AUC = 0.920).
SAP was statistically significant, differentiating MAP with an AUC of 0.855 and MSAP
with an AUC of 0.684. suPAR correlated with clinically used lab values and clinical scores
including Ranson’s score and CRP. Finally, suPAR was able to predict inpatient mortality
from SAP with an AUC of 0.806. The study concluded that suPAR could be used as a
potential biomarker for inflammation, severity, and inpatient mortality in SAP patients [
95
].
Biomedicines 2024,12, 1167 23 of 44
2.9.2. Chronic Hepatitis and Fibrosis Progression
Twelve potential biomarkers were evaluated in twenty-one cirrhotic patients positive
for chronic hepatitis C (HCV) genotype I versus twenty-one healthy controls, assessing
for progression of liver fibrosis as evidenced by transient elastography. suPAR was found
to have an AUC of 0.78, a fair diagnostic biomarker for progression of liver fibrosis in
chronic HCV patients [
95
]. A later study by another research group corroborated these
findings. Researchers examined 146 chronic infections in HCV patients in two cohorts.
Mean suPAR levels were not elevated in the earlier-stage fibrosis F1 and F2 stages, but
were significantly increased in the F3 and F4 stages, when compared to healthy subjects.
The AUC in distinguishing F1/F2 from F3/F4 was 0.774, and the AUC in distinguishing
non-cirrhotic (F1, F2, and F3) from cirrhotic patients (F4) was 0.791. Serum suPAR levels
also strongly correlated with noninvasive clinically used biomarkers of fibrosis in the
aspartate transaminase-to-platelets ratio index score (r = 0.52) and transient elastography
imaging studies (r = 0.44; all pvalues < 0.0001) [96].
The gold standard for monitoring liver progression is invasive liver biopsy for cirrhosis
staging in hepatitis B cirrhosis. To this end, a biomarker is needed to assess liver cirrhosis
progression. A total of 76 patients and 21 healthy controls were recruited to assess various
biomarkers for liver cirrhosis progression. Clinically used biomarkers including mean
platelet volume or aspartate aminotransferase-to-platelet ratio index scores failed to achieve
statistical significance. However, suPAR and IL-10 were significantly higher in those
patients with severe fibrosis versus mild fibrosis [
97
]. The finding of elevated suPAR levels
was corroborated in another study including 105 cirrhotic patients when compared to
19 liver-healthy controls [98].
2.9.3. Acute Decompensated Liver Failure
No biomarker yet exists which separates the severity of decompensated cirrhosis
from bacterial infections common in the patient population. In a single-center study of
162 patients
with decompensated liver cirrhosis who underwent diagnostic paracentesis at
a tertiary care center in Aachen, Germany, suPAR levels were increased in patients with de-
compensated cirrhosis and correlated with the severity of liver dysfunction and systemic in-
flammation, but were not indicative of bacterial infection.
Serum suPAR levels > 14.4 ng/mL
predicted 28-day mortality in these patients. Ascitic fluid suPAR levels were elevated
during episodes of spontaneous bacterial peritonitis, but were not elevated during episodes
involving bacterial translocation into the ascitic fluid. The reasoning by the authors, un-
covered in
in vitro
experiments, was that monocytes (and to a lesser extent neutrophils)
secreted suPAR following TLR activation, leading to rapid uPAR cleavage and upregula-
tion [99,173].
2.9.4. Inflammatory Bowel Disease
Inflammatory bowel disease (IBD), including ulcerative colitis (UC) and Crohn’s
disease, is characterized by recurrent chronic inflammation of the intestinal tract. The
significant rise in IBD cases in the 21st century has been strongly associated with the
rapid industrialization of countries [
114
]. Despite ongoing research, the exact cause of IBD
remains a mystery, posing challenges to diagnosis and treatment given its multifactorial
nature. A recent comprehensive transcriptomic meta-analysis of public IBD datasets has
identified uPAR as a potential key factor in IBD. Mouse models of IBD have increases
in uPAR expression during epithelial cell breakdown. Pharmacological blockade and
knockdown of uPAR in the same mouse model demonstrated a protective effect against
cytokine-induced mucosal barrier breakdown [174].
The role of uPAR in IBD suggests a potential novel therapeutic target. The breakdown
of the intestinal epithelial barrier is a crucial factor in the development of inflammatory
bowel disease (IBD), causing inflammation, damage, and loss of crypt architecture. A
potential avenue for innovative IBD therapeutics lies in targeting specific ligand–receptor
pairs in IBD mucosa, which may play a role in maintaining the integrity of the intestinal
Biomedicines 2024,12, 1167 24 of 44
epithelial barrier. The uPA–uPAR complex was presented as a therapeutic target for
IBD based upon meta-analysis indicating that there is upregulation of urokinase-type
plasminogen receptor–ligand genes in damaged mucosa. Interestingly, the same genes
were expressed less during barrier formation. This coordinated upregulation of uPA–uPAR
in UC and CD biopsies is hypothesized to be linked to cytokine-induced damage to the
epithelial barrier [117].
Human intestinal epithelial cell lines with knockout of uPA and uPAR genes have
shown increased protective barrier formation, further indicating the dysregulatory role of
uPA–uPAR in IBD. In primary-organoid-derived cell monolayers, researchers were able to
demonstrate improved barrier function with small-molecule inhibitors, peptide antagonists,
and neutralizing antibodies targeting the uPA–uPAR complex. The uPA protease inhibitor
did not show the same significant epithelial barrier protection effect. These findings stress
the importance of targeting intracellular receptor signaling to attenuate IBD, while also
avoiding potential unwanted side effects of uPA inhibition. This paper also highlights
the lesser importance of inhibiting ECM formation, which is modified by uPA-uPAR. To
gain enhanced insights into the therapeutic effects
in vivo
, it will be necessary to use a
monoclonal antibody and/or a highly effective small-molecule inhibitor to hinder the
interaction between uPA and uPAR [117].
A dextran sulfate sodium (DSS) colitis model was tested in either wildtype or uPAR-
knockout mice. Mice lacking uPAR exhibited significant protection, evident in the restora-
tion of colon mucosa architecture, a reduction in inflammatory infiltrate, and improved
surface integrity. Notably, in wildtype mice treated with DSS, a disruption of cellular tight
junctions was observed, and this effect was markedly diminished in the knockout (KO)
mice, indicating a safeguarding of the epithelial barrier
in vivo
by uPAR deficiency. The
improved colon morphology and reduced epithelial damage in uPAR-KO mice indicated a
protective role of uPAR inhibition against DSS-induced colitis [117].
Since uPAR is a GPI-anchored receptor, it has the capacity to interact with various
membrane and intracellular pathways. Through co-immunoprecipitation, Cheng and col-
leagues found that uPAR–epidermal growth factor receptor (EGFR) binding is undisturbed
following barrier damage. In contrast, the association of uPAR with integrin subunits was
lost following the cytokine-induced damage [117].
Repair signals from EGFR were also increased in uPAR-deficient cells during bar-
rier breakdown. The signaling pathway of EGFR plays a crucial role in regulating ep-
ithelial functions, including maintenance of cell junctions, cell survival, and secretion of
mucin [
118
]. Since EGFR-uPAR binding is unchanged during IBD pathogenesis, this finding
could support the key role of uPAR in suppressing EGFR-modulated repair mechanisms
and signaling. Upon cytokine challenge, tight junctions were disrupted with cell death
increased, indicating dysfunction in EGFR. In uPAR-knockout cells, an overall increase in
cell survivability and decreased junction damage was observed, in addition to enhanced
activity of 45 different kinases/phosphoproteins associated with EGFR [118].
2.10. Endocrinology
2.10.1. Type I Diabetes
suPAR levels were assessed in 667 patients with type I diabetes (T1DM) versus
51 nondiabetic
control patients in a single-center cross-sectional study, stratified into as-
sociated complications of diabetes. suPAR was elevated across all cohorts with T1DM.
suPAR levels were more elevated in T1DM patients with cardiovascular complications.
Patients with cardiovascular disease were observed to have a 2.5 times higher suPAR
level, autonomic dysfunction carried a 2.7 times greater suPAR level, 3.8 times elevated
for albuminuria, and finally 2.5 times higher for patients determined to have stiff arterial
walls [100].
Biomedicines 2024,12, 1167 25 of 44
2.10.2. Type II Diabetes
SuPAR levels, as in T1DM, are also elevated in patients with T2DM. One study
investigated suPAR as an early biomarker for diabetic nephropathy, potentially providing
an earlier marker than the use of the clinically approved microalbuminuria. Researchers
investigated baseline suPAR levels with incidental microalbuminuria in a prospective
longitudinal cohort study of 258 patients at risk of T2DM. Another cohort was studied
looking for association with albuminuria at later stages of T2DM in a cross-sectional cohort
with diagnosed T2DM. A higher baseline suPAR level was associated with a higher risk
of microalbuminuria in subjects at risk for T2DM for the higher quartile of subjects when
compared to the lower quartile of subjects. Patients with new-onset microalbuminuria
were found to be at increased risk of prediabetes. suPAR levels were consistently elevated
in patients with microalbuminuria in a separate cohort of patients with already-diagnosed
T2DM. Finally, elevated baseline suPAR concentrations were independently associated
with new-onset microalbuminuria in subjects at risk for T2DM. Thus, suPAR may present
earlier than microalbuminuria in patients at risk of T2DM [101].
2.11. Pulmonology
Lung disease has also been associated with altered uPAR activity (Table 1).
2.11.1. Asthma
Asthma is responsible for a substantial number of hospital readmissions, and at
present no prognostic biomarker exists for asthma patients. To this end, serum suPAR
and eosinophils were taken upon admission from 1341 patients admitted with an acute
exacerbation of asthma. The 365-day readmission and all-cause mortality were assessed.
Patients who were either readmitted or died had higher suPAR levels and decreased
eosinophil levels upon admission. Patients in the 4th quartile for suPAR levels or eosinophil
counts < 150 cells/uL had the highest odds for readmission or mortality. The investigators
for this study stated that the concurrent use of suPAR with eosinophils as a biomarker
allows for precision in asthma prognostication for patients at higher risk of adverse events
or lower disease control [104].
2.11.2. Ventilator-Associated Pneumonia
Ventilator-associated pneumonia (VAP) remains a challenge for timely diagnosis
for appropriate treatment. In an observational, prospective, multicenter cohort study of
24 patients
with VAP compared with 19 control patients, suPAR levels were found to be
significantly elevated in VAP patients three days before definitive VAP diagnosis, albeit
poorly, with an AUC of 0.68. The AUCs on the day of diagnosis and in deceased patients
were even higher, at 0.77 and 0.79, respectively; when combined with already clinically
used C-reactive protein, procalcitonin, and the Clinical Pulmonary Infection Score, VAP
prediction and specificity increased. Thus, on its own, suPAR had a fair diagnostic accuracy,
but as an adjunct biomarker suPAR further assisted in diagnosis [
106
]. Furthermore, in a
homogeneous Greecian cohort of 180 patients with concurrent VAP and sepsis, suPAR levels
greater than a 10.5 ng/mL cut-off had 80% specificity and 77.6% positive predictive value to
discriminate between severe sepsis and sepsis. suPAR levels greater than
12.9 ng/mL
had
80% specificity and 76.1% positive predictive value for prognosis of unfavorable outcome.
Finally, in these critically ill patients with sepsis and VAP, suPAR was identified as an
independent factor associated with unfavorable outcome, as identified using stepwise Cox
regression analysis [107].
2.11.3. Community-Acquired Pneumonia
Community-acquired pneumonia (CAP) is the most common infectious disease carry-
ing high mortality. In a study investigating suPAR levels in 75 patients with CAP versus
67 healthy controls, suPAR levels were significantly elevated in patients with CAP, and
correlated with the clinically used Pneumonia Severity Index scores. LPS was found to
Biomedicines 2024,12, 1167 26 of 44
induce suPAR expression in macrophages in a mouse model of CAP. The authors concluded
that plasma suPAR levels may provide a biomarker for CAP severity, and may potentially
serve as a therapeutic target [105].
2.11.4. Chronic Obstructive Pulmonary Disease (COPD)
Chronic obstructive pulmonary disease (COPD) is a chronic inflammatory disease
causing hospital readmission and mortality through repeated exacerbation. Commonly
used acute-phase reactants, biomarkers such as CRP, are increased during these acute
exacerbations. suPAR levels were measured in 43 patients with an acute exacerbation of
COPD and compared to 30 healthy controls on day one of admission and seven days after
treatment. All acute-phase reactants studied, including suPAR, and the clinically used CRP
and fibrinogen, were markedly elevated. The AUC was superior for suPAR when compared
to the other acute-phase reactants. The researchers concluded that suPAR can be used as a
predictor for acute COPD exacerbations and in monitoring response to treatments [
108
]. A
later meta-analysis of 11 studies involving 4520 patients confirmed these findings, with the
additional finding that suPAR serves as a clinically useful biomarker for early diagnosis,
with a sensitivity and specificity of 87% and 79%, respectively, and an AUC of 84%. These
researchers also concluded that suPAR can be used to distinguish acute exacerbations from
stable COPD, and also can be used to guide clinical response to treatment [109].
2.11.5. Pleural and Parenchymal Acute Lung Injury and Repair
In the context of lung injury, uPAR has been associated with fibrotic processes and
tissue remodeling. The uPA/uPAR complex activates plasmin and subsequently MMPs and
other proteolytic enzymes, influencing the balance between tissue repair and fibrosis [
175
].
A correlation between the pleural fluid level of suPAR and the predictive requirement
for aggressive management has been reported in patients with parapneumonic pleural
effusions [
120
]. Clinical studies of 93 patients with pleural effusions found that levels of
suPAR in the pleural fluid were significantly elevated, by even more than pH, glucose, and
lactate dehydrogenase (LDH) [
119
]. In the same study, monitoring suPAR levels was the
most accurate indicator for chest tube insertion. Additional clinical trials, involving more
patients at multiple centers, are required to address this need for accurate biomarkers in
clinical settings.
In mouse studies, diminished uPAR expression attenuated lung injury associated
with hypoxia, and additionally, decreased lung parenchymal destruction [
176
]. Simi-
larly, such anti-inflammatory effects limited the containment of pneumonia, which led to
poorer outcomes.
The uPA/UPAR system has also been implicated in pulmonary fibrosis [
177
]. Shetty
and colleagues reported that lung samples of patients with idiopathic pulmonary fibrosis
(IPF) have increased uPAR expression compared to healthy patients. The same authors
hypothesized that post-transcriptional regulation of uPAR plays a partial role in the devel-
opment of lung fibrosis [178,179].
2.11.6. Smoking Exposure
In a randomized controlled cohort study of 48 smokers vs. 46 never smokers, suPAR
levels were found to be significantly elevated at 3.2 ng/mL vs. 1.9 ng/dL, respectively.
Smokers were randomized to smoking cessation with nicotine patches alone. Four weeks
following cessation, suPAR levels were comparable to never smokers. Individuals with
the highest levels of smoking at the time of cessation maintained the highest levels at
four weeks following cessation, pointing towards the utility of suPAR levels in CI. The
researchers concluded that suPAR may be useful as a biomarker for personalization of
smoking cessation by identifying those at risk and those who may benefit the most from
cessation. However, future studies investigating the longitudinal effects are still needed to
truly assess clinical utility [110].
Biomedicines 2024,12, 1167 27 of 44
2.12. Rheumatology
Rheumatoid arthritis and lupus are associated with altered uPAR activity (Table 1).
2.12.1. Rheumatoid Arthritis
As mentioned in the inflammation subsection, suPAR levels in patients with rheuma-
toid arthritis are elevated when compared to healthy cohorts. suPAR levels correlated with
joint involvement. These findings indicate that suPAR provides a useful biomarker for
the diagnosis and management of progression of RA [
85
,
86
,
160
–
162
]. Potential treatments
utilizing this biomarker are described in Section 3.
2.12.2. Systemic Lupus Erythematosus (SLE)
SuPAR was proposed by rheumatology experts as a clinically useful biomarker for
systemic lupus erythematous (SLE) as early as 2015 [
180
]. suPAR levels have been demon-
strated to reflect accrued damage in patients with SLE. In a later study, researchers in-
vestigated suPAR levels in 344 patients with SLE who met the 1997 American College
of Rheumatology classification criteria and compared these levels with organ damage
as assessed by the SLICC/ACR Damage Index (SDI), a clinically used index of overall
organ damage attributable to SLE. suPAR levels were elevated in patients with SLE with
progressive damage when compared to those with no detected damage, in particular with
those carrying an SDI of two or greater. Furthermore, in an optimized logistic regression
analysis predicting damage attributable to SLE suPAR was identified as a useful biomarker
for predicting disease progression, together with baseline disease activity (as identified
by used SLEDAI-2K), age, and non-Caucasian ethnicity, with an AUC of 0.77, on the high
side of a fair prognostication for organ damage accrual during the first five years of SLE
disease [87].
2.12.3. Systemic Sclerosis
Systemic sclerosis (SSc) is a multisystem autoimmune disease characterized by internal
organ and dermal fibrosis, vasculopathy, and widespread immune system dysregulation.
RA is likened to SSc in that they are both autoimmune diseases involving both the fibri-
nolytic and uPAR systems. SSc differs in that SSc is pathologically anti-angiogenic, whereas
RA is pathologically pro-angiogenic [
181
,
182
]. suPAR levels were found to be elevated
in patients with SSc versus controls, and those with pulmonary fibrosis had higher lev-
els [
183
,
184
]. Mechanistically, the anti-angiogenetic properties of SSc in these endothelial
cell types is chiefly due to uPAR diminishment and the resultant loss of the beta2 integrin-
facilitated connection of uPAR with the actin cytoskeleton [
185
]. This cleavage is due to
overproduction of matrix metalloproteinases in SSc cleaving and activating the uPAR,
reducing angiogenesis in the process [186].
3. Development of New Therapeutics—uPAR as a Therapeutic Target
In Section 3, we will discuss uPAR as a potential therapeutic target for treating severe
inflammatory diseases, with a focus on the virus-derived immune-modulating proteins.
Small-molecule-, antibody-, and protein-based uPA and uPAR inhibitors have been
investigated and several have been briefly discussed above in the individual sections
(Table 1),
illustrating the efficacy of drugs that modify uPAR expression in disease. uPAR
has been used for targeted chemotherapy through a variety of mediums including mono-
clonal antibodies and nanogels for treating cancers. A virus-derived immune-modulating
protein, Serp-1 is a protein that binds uPA and uPAR and has been tested in a wide ar-
ray of animal models with demonstrated efficacy in reducing inflammation and disease
progression (Table 2) [
187
,
188
]. Serp-1 activity is lost in uPAR-knockout mouse models of
aortic transplantation [
146
,
147
] and with anti-uPAR antibody treatments in mouse wound
healing studies [
189
]. In-depth reviews have been compiled for many of these new ap-
proaches, and thus, we have provided a broader overview. Given the recent prior reviews
on uPAR-modifying treatments [
189
–
194
], we have provided an overview of these ap-
Biomedicines 2024,12, 1167 28 of 44
proaches together with a more in-depth review of the virus-derived immune-modulating
serpin, Serp-1 (Table 2).
The data examining uPAR blockade as a treatment further supports a central role
for uPA/uPAR in disease progression and provides newer therapeutic targets for treating
disease.
3.1. Oncology
Given uPAR’s diminished expression in healthy homeostatic tissues when compared
to cancers, the receptor has been targeted as a viable therapeutic option [
26
]. Due to
its role in plasminogen activation and tissue remodeling, allowing cellular invasion and
altering immune responses, uPAR provides an avenue for cancer invasion and metastasis.
The uPA/uPAR complex increases MMP proteolytic activity and dissolution of ECM
barriers, along with enhancing stromal angiogenesis in the tumor microenvironment (TME).
uPA/uPAR at the leading edge of immune cells may also enhance tumor-associated stromal
cells, further increasing cancer growth and/or invasion. Many
in vitro
and
in vivo
studies
have proven impaired tumor progression, metastasis, and invasion when the proteolytic
function of uPA/uPAR is either impaired or inhibited [16,24,195,196].
The negative prognostic value with uPAR stromal expression in multiple cancer types,
including colon, breast, and pancreatic cancers, clearly highlights the therapeutic potential
of aiming at stromal TME as an adjuvant anti-angiogenic, anticancer treatment [
191
–
194
].
uPAR’s role in the tumor stromal microenvironment, overexpression in non-homeostatic
tissue, high expression in aggressive cancer subtypes of poor prognosis, along with the lack
of obvious cancer phenotypes when uPAR is deficient, all suggest uPAR as a candidate in
anti-tumor cytotoxic therapy. Targeting strategies using uPAR have been employed.
uPAR targeting is accomplished using uPA-derived peptides, monoclonal antibodies,
and high-affinity receptor-binding fragments of uPA (containing the GFD). As mentioned
previously, uPAR plays an important role in malignancy (tumor invasion) and metastasis
of breast cancer [
66
–
68
,
135
,
136
]. Thus, the therapeutic potential of this system has been
explored. A fully human antibody called 2G10 that effectively blocks uPA/uPAR interac-
tions and has shown promise in treating aggressive triple-negative breast cancer (TNBC) in
mouse models has recently been developed [71].
These uPAR-targeting ligands provide scaffolds for targeted delivery of cytotoxic
drugs, including traditionally used anticancer agents, stromal-targeting oncolytic viruses,
cytotoxic products, clinically used radioisotopes, photosensitizers, chimeric antigen recep-
tor (CAR) T cells, and immunostimulators [
16
]. These approaches increase tumor specificity
and improve intra-tumoral delivery to the tumor stromal microenvironment. Albeit it is in
its early research stages, the utility of uPAR targeting may provide effective targeted drug
invasion through the dense stroma, a chief mechanism behind anti-chemotherapeutic drug
resistance. Few medications exist that can overcome drug resistance [197,198].
uPAR-targeting radionuclide therapy, PET-probes based on the high-affinity peptide
AE105, have been synthesized and tested preclinically in human xenograft mouse studies
and in two clinical studies evaluating uPAR PET studies in breast, bladder, and prostate
cancer patients, with positive results. Although further preclinical validation and toxicity
studies are required, the availability of human anti-uPAR-targeting constructs and their
successful use as adjuvant imaging agents strongly support clinical translation to determine
the therapeutic and prognostic utility of uPAR in the management of aggressive tumors,
such as ovarian, prostate, and breast cancers. These studies have implications for future
use as an imaging or treatment adjuvant. The full cytotoxic effect on human tumor lesions
in xenograft mouse models is likely understated as it leaves the host stromal compart-
ment essentially unharmed, where uPAR operates in the TME [
19
,
195
,
199
–
204
]. This is
further corroborated by the fact that when the radioimmunotherapy is combined with
a stromal-targeting recombinant anti-uPAR antibody, complete tumor regression occurs
with triple-negative breast cancer (TNBC) human tumor xenografts in a metastatic mouse
model [196,205].
Biomedicines 2024,12, 1167 29 of 44
Recombinant immuno- and ligand-targeted fusion toxins (IT and LT) is another ap-
proach to cytotoxic therapy designed to target uPAR. The cell-binding domain of the
toxin/ligand is complexed with the tumor-targeting vehicles, providing a desired binding
specificity [
206
]. First described utilizing diptheria cytotoxin A, the catalytic potency of the
toxin moiety is enhanced, allowing a small number of molecules to be effectively delivered
to the cytosol to kill cancer cells [
207
,
208
]. Internalization of the toxin is, however, required,
unlike the radionuclide therapies. In preclinical studies, utilizing mono- and bi-specific
activating transcription factor (ATF)-fusion recombinant diptheria toxins that target uPAR
has been successful in treating human to mouse xenograft glioblastoma multiforme (GBM)
and non-small-cell lung cancer models [208–213].
Utilizing the same LT therapy with Pseudomonas aeruginosa toxin in GBM subcuta-
neous and intracranial xenograft models has also proven beneficial [
213
]. Mouse models
investigating the recombinant toxin’s use in head and neck squamous cell carcinoma have
found similar success [
213
]. The therapy was deemed safe and effective in targeting sarco-
mas in preclinical studies, but future clinical studies will be required [
214
]. A toxin from the
plant bacteria saporin has been studied in human bladder and triple-negative breast cancer
cell lines and in bladder cancer SC xenograft models with success [
215
,
216
]. The repeated
success amongst different tumor subtypes in a variety of different toxin studies has now
validated the ability to target uPAR expression in the TME once stromal penetrance occurs,
utilizing the toxin-aided vehicles as the means of internalization.
Antibodies targeting uPAR utilizing the 2G10 antibody have also been designed with
preclinical success. Clinical studies are still needed, but these may have future therapeutic
implications in aggressive triple-negative breast cancer, for which current medical therapies
are limited with minimal efficacy due to the lack of molecular targets [
196
,
205
]. Following
blockage of the uPAR with antisense oligonucleotides, a uPAR expression was strongly
reduced against murine prostate cancer bony metastases [
217
]. This very therapy also
worked successfully in preclinical studies, reducing the invasion of human cartilage in
synovial fibroblasts in vitro [218].
uPAR-targeting drug-loaded iron oxide nanoparticles (NPs) also have the capacity
to direct therapy to target sites when complexed with guiding moieties, synergistically
amplifying anti-tumor chemotherapeutic response. Preclinical trials have shown promise
in treating human breast, prostate, and lung cancer cell lines. The chemotherapeutic
drugs involved include doxorubicin with curcumin, gemcitabine, cisplatin, noscapine, and
paclitaxel [75,77,219–225].
While the direct therapeutic uPAR-targeting antibodies have had success, immune-
checkpoint blockade and adoptive cell therapy via chimeric antigen receptor (CAR) T cells
have recently emerged as a breakthrough in the treatment of malignant tumors including
ovarian, glioblastoma, and melanoma in preclinical studies. uPAR targeting was reported
to selectively induce immune-mediated clearance of non-homeostatic uPAR-positive cancer
cells via antibody-recruiting small molecules (ARMs) in a metastatic glioblastoma mouse
model [226,227].
Oncolytic virotherapy is a new and rapidly growing field in cancer therapeutics, with
proven late-stage clinical studies. Presently, one FDA-approved oncolytic virus exists, a
herpesvirus [
228
]. However, virotherapy has taken advantage of the oncolytic measles virus,
where success was seen in MV targeting the stromal in breast and colon human-mouse
xenograft preclinical studies [229–233].
The virus-derived serpin Serp-1 binds uPA and uPAR in human macrophage cell
lines. One study in severe combined immunodeficient (SCID) mice demonstrated that
Serp-1 produced from myxomyavirus reduced pancreatic xenograft growth with altered
myeloid immune cell responses (Figure 3, Table 2) [
81
]. Myxoma virus is being studied as
an oncolytic virus for treating cancers. Angiogenesis, the growth of new blood vessels, also
increases tumor growth. In very early work, chorioallantoic membrane angiogenesis was
reduced by Serp-1 in an angiogenic model [144].
Biomedicines 2024,12, 1167 30 of 44
Despite these many promising studies—targeting uPAR as a new approach to cancer
treatment has demonstrated great potential—this area remains in development.
3.2. Inflammatory Bowel Disease
uPA inhibitors have been recently investigated as potential new therapeutics for IBD.
One study detected a dramatic reduction in colitis in mice with experimental colitis when
treated with uPAR inhibitors.
The authors in the study suggested uPA as a therapeutic target for patients with UC.
A modified, PEGylated Serp-1 protein, PEGSerp-1, has also been examined recently in
acute severe and chronic DSS-induced colitis models, demonstrating improved survival
and reduced colon inflammation and damage with loss of crypt architecture (Table 2) [
155
].
3.3. Anti-Inflammatory Anti-Angiogenic Immunomodulation
While originally investigated for its anti-metastatic potential, UPARANT (UPAR anti-
body) has been researched as an anti-inflammatory drug to treat angiogenic inflammatory
disease states in patients with ocular pathologies and complications of diabetes [
234
].
UPARANT was found to reduce VEGF-induced angiogenesis caused by oxygen-induced
retinopathy in a dose-dependent fashion by diminishing VEGF, VEGF-receptor 2, and tran-
scription factors regulating VEGF expression [
235
]. In a mouse study modeling choroidal
neovascularization, intravitreal injection of UPARANT reduced leakage from the choroid
and choroidal neovascularization area [
235
,
236
]. In mouse edema and peritonitis models,
intraperitoneal injection of UPARANT reduced pro-inflammatory enzymes [121].
In a mouse diabetic nephropathy model, injection of UPARANT subcutaneously re-
stored vascular membrane integrity, making the vascular membrane less permeable [237].
Subcutaneous injection of UPARANT reduced Mueller cell gliosis, upregulated inflam-
matory markers, and had anti-apoptotic effects, all of which improved retinal function,
in a retinitis pigmentosa mouse model [
238
]. In a mouse model of rubeosis iridis with
proliferative retinopathy, local intravitreal injection of UPARANT was found to be superior
to anti-VEGF treatment. Systemic subcutaneous injection also reduced neovasculariza-
tion in this study [
239
]. All these findings are highly suggestive for UPARANT being a
co-occurring anti-inflammatory and anti-angiogenic immune-modulating therapeutic.
Purified Serp-1 protein is a highly effective anti-inflammatory and immunomodulator,
as has been proven in a wide variety of animal models and a clinical trial [
188
,
240
]. Serp-
1 carries out its potent effects through binding to uPA, tPA, plasmin, thrombin, factor
Xa and complement; Serp-1 requires uPAR for this potent immunomodulatory and anti-
inflammatory activity. Upon binding, Serp-1 modulates monocyte and T cell cellular
migration through uPAR-linked integrins and actin-binding proteins in Serp-1 cellular
responses. Serp-1 interacts with uPAR to reduce macrophage activation, adhesion, and
invasion. Serp-1 similarly diminished monocyte and T cell migration across endothelial
layers and ascitic fluid both in vitro and in vivo [188].
Serp-1 increased expression of filamin B, an actin-binding protein and decreased
beta-integrin (CD18) expression. These molecular functions were dependent upon uPAR,
as demonstrated by immunoprecipitation analysis. Blocking Serp-1 induced changes in
filamin B expression through small inhibitory RNA (siRNA)-reduced Serp-1-mediated
inhibition of monocyte adhesion and diapedesis. In mouse models of aortic allograft
transplantation, uPAR-deficient (uPAR-KO) donor allografts inhibited Serp-1-mediated
reductions of aortic transplant vasculitis (Table 2). Antibodies to uPAR reduced the efficacy
of Serp-1-mediated wound healing. The capacity of Serp-1 to reduce inflammation and as-
sociated diseases is, thus, closely associated with interactions with the uPA/uPAR complex,
with Serp-1 treatment downregulating beta-integrin and increasing filamin B expression in
human macrophage cell cultures. These findings further support the uPA/uPAR complex
as a new therapeutic target [188,240].
Visual impairment due to photoreceptor degeneration in inherited eye diseases is
chiefly due to inflammation, without angiopathy. uPA/uPAR dysfunction has been re-
Biomedicines 2024,12, 1167 31 of 44
ported as a key player in a variety of eye diseases including retinitis pigmentosa and
retinopathy of prematurity; emerging as a new inflammatory pathway in a variety of ocular
diseases, a pathway that differs from the classic pro-angiogenic pathway. Through the
counteraction of neovessel formation and microvascular dysfunction, uPAR modulates the
inflammatory response as a possible treatment for a variety of eye diseases of neovascularity,
including diabetic retinopathy, wet macular degeneration, retinopathy of prematurity, and
retinitis pigmentosa. Finally, uPAR modulates the inflammatory cascade during rod-cell
degeneration in retinitis pigmentosa. Retinitis pigmentosa is an eye disease with associated
low uPA/uPAR levels. The uPA/uPAR complex may thus provide a possible therapeutic
target through anti-inflammatory activity and reduction of oxidative stress [241].
In prior work, Serp-1 reduced alkali-induced corneal damage in a mouse model [
233
].
In a separate study, AAV (adeno-associated virus) expression of Serp-1 also reduced uveitis
in vitro, corroborated with mouse models [124,145].
3.4. Diffuse Alveolar Hemorrhage in SLE
Diffuse alveolar hemorrhage (DAH) is a fatal complication of SLE with mortality
of up to 50–80%. No proven effective treatments presently exist for DAH. In a mouse
DAH model, treatment with PEGSerp-1 protein as well as the unmodified Serp-1 protein
markedly reduced DAH, with associated reductions in uPAR distribution and reduced
macrophage alveolar invasion (Table 2) [
142
,
143
]. In a collagen-induced arthritis model,
early prophylactic treatment also reduced joint inflammation [154].
Future studies are needed to determine clinical significance; however, this finding
again highlights the importance of uPAR in the processes of this potentially fatal autoim-
mune disease [142,143].
3.5. Acute Transplant Rejection
uPAR is linked to immune responses across the body including, but not limited to,
the lungs and kidneys. Under conditions of oxidative stress and hypoxia the uPA/uPAR
pathway is upregulated. Growing evidence has revealed that ischemia-reperfusion injury
induces immune cell activation in both acute and chronic allograft rejection. Human biopsy
studies have detected uPA/uPAR activation correlating with allograft rejection. The pathol-
ogy contributing to acute allograft rejection is in part due to transplant vascular disease and
occlusion and subsequent ischemia-induced cell damage and apoptosis. Recipient and also
resident renal allograft leukocytes are reported to infiltrate the transplant organ. This is be-
lieved to be mediated in part through increased uPA/uPAR expression and dysregulation.
uPAR has been found to be necessary for TNF-alpha and C5a signaling, inducing integrin
ICAM-1 signaling on allograft endothelial cells, a crucial step in leukocyte diapedesis [
242
].
In prior work Serp-1 treatment improved outcomes in aortic allografts, renal allografts,
heterotopic cardiac allografts, and rat-mouse cardiac xenografts (Table 2) [
148
,
149
]. As
noted above, Serp-1 efficacy in aortic allograft models was blocked in uPAR-deficient
allograft implants, demonstrating that Serp-1 efficacy is in part dependent upon uPAR.
3.6. Atherosclerotic Plaque Stabilization
Serp-1 has demonstrated preclinical efficacy in treating inflammatory atherosclerotic
diseases in both preclinical and clinical studies (Table 2). In studies on hyperlipidemic rats
and rabbits, continuous infusions of Serp-1, and later Serp-2, were found to reduce carotid
cuff compression in ApoE null hyperlipidemic mouse models, stabilizing the carotid plaque
at the affected sites [152].
In a randomized, blinded, dose-escalating phase 2A clinical trial at seven sites in the
US and Canada, Tardif et al. demonstrated that Serp-1 given at the time of coronary stent
implant for unstable coronary syndromes significantly reduced troponin and creatinine
kinase MB (CK-MB) levels at the higher dose of 15
µ
g/kg. Serp-1 treatment reduced
clinical markers of myocardial damage with no significant increases in adverse effects. Of
interest, there were minimal if any detectable neutralizing antibodies to treatment with this
Biomedicines 2024,12, 1167 32 of 44
virus-derived serpin protein [
243
]. The doses used were microgram/kilogram doses so
the protein is highly active. All treatments with proteins and even antibodies can lead to
antibody development, but the lack of neutralizing antibodies in response to this foreign
protein is certainly promising for potential repeat treatments.
3.7. Inflammatory Vascular Disease
Giant cell arteritis is an inflammatory vascular disease with significant risks for sudden
loss of vision (blindness) if left untreated, as well as other vascular complications including
strokes, cardiomyopathy, and aneurysms. Serp-1 treatment successfully reduced inflamma-
tion in human giant cell arteritis xenograft implants in severe combined immunodeficient
(SCID) mice [
153
] in the presence of peripheral human blood mononuclear cell infusions.
The study of these temporal artery biopsy implants was blinded to GCA diagnosis. Gamma-
herpesvirus infection (MHV68) is also a model for inflammatory vasculitis [
150
]. Serp-1
treatment significantly improved survival and reduced vascular and lung inflammation in
mouse models of MHV68 infection.
3.8. Inflammatory Arthritis
RA is the most common inflammatory rheumatologic disease in the world. In a
collagen-induced arthritis model, rats administered with Serp-1 at 50
µ
g/kg via intravenous
(IV) injections had reduced clinical arthritis, with reduced joint swelling when it was given
at the time of inducing the disease and reduced bony erosions on radiographs, with overall
improved clinical status [154].
3.9. Severe Acute Respiratory Distress Syndrome Due to SARS-CoV-2
Severe acute respiratory distress syndrome is a lethal sequalae of increased lung per-
meability, usually due to an infectious insult. In a mouse-adapted MA-30 SARS-CoV-2
model, SARS-CoV-2 treated with PEGSerp-1 altered uPAR gene expression and detectable
protein on IHC analysis. Further treatment with PEGSerp-1 reduced lung and arterial in-
flammation and associated damage in the MA-30 SARS-CoV-2-infected C57Bl/6 mice [
151
].
As noted above, in prior work using an MHV68 mouse herpes model, causing an otherwise
lethal infection, Serp-1 improved survival and reduced lung and vessel inflammation [
149
].
3.10. Inflammation in Wound Healing
Factors involving clotting, hemorrhaging, and inflammation help dictate early wound
healing. In a mouse study, Serp-1 treatment in a chitosan–collagen hydrogel accelerated
wound healing. This wound healing acceleration was blocked by the urokinase-type
plasminogen activator (uPAR) antibody [156].
3.11. Neuromuscular Disorders, Duchene Muscular Dystrophy, and Spinal Cord Injury
Similar studies examined local infusions of Serp-1 as well as an implant of a chitosan–
collagen hydrogel in a rat model of spinal cord injury. This treatment was compared to local
dexamethasone treatments. Serp-1 improved motor function and reduced inflammation
in this model without associated toxicity [
157
]. Mice deficient in dystrophin (Dmd) were
administered PEGSerp-1 intraperitoneally for four weeks beginning at four weeks of age.
These treatments reduced muscular inflammation and diaphragm fibrosis [158].
4. Discussion/Conclusions
In conclusion, within the realm of clinical applications, the uPA/uPAR complex is
now proven to be a very promising biomarker for inflammatory disease and cancer. Note-
worthy is the potential prognostic value in diverse clinical scenarios, specifically in cancer
metastasis and severe inflammatory diseases such as viral infections and inflammatory
bowel disease. A nuanced understanding of uPA/uPAR interactions, as presented in this
review, is indispensable for the formulation of targeted therapies and interventions across
various clinical domains.
Biomedicines 2024,12, 1167 33 of 44
The known benefits of using higher doses of uPA as a thrombolytic or clot-dissolving
agent for acute thrombotic events, heart attacks, and strokes, where there is improved
survival, appears to be at variance with the potential for damage from inflammation
associated with the uPA/uPAR complex. However, the elevated uPA/uPAR complex
response may have greater impact as part of an early inflammatory response. This early
uPA/uPAR response may be initiated as an early healing response which, when excessive,
then leads to more aggressive and damaging immune and inflammatory responses. The
wide range of potential actions of uPA and suPAR may also provide differing responses to
differing pathologies and/or, on a simpler level, differing cellular responses in differing
organs. Further in-depth analyses of the local effects of the uPA/uPAR response in differing
organs and cells may identify individual cell responses. An in-depth analysis will be of
great interest and may provide new insights into these unique immune modifiers. The
uPAR complex has a very wide range of functions in the inflammatory response, in addition
to acting as a fibrinolytic. There are likely many new discoveries to be made in this field,
both using uPAR as a marker for disease and as a new therapeutic target for treating
diseases where treatments are limited.
Therapies targeting the uPA/uPAR complex from small molecules and antibod-
ies, chemotherapy designed to target uPAR in cancer cells, and virus-derived immune-
modulating protein such as the virus-derived serpin Serp-1 are under investigation and
provide new approaches to treatment. The ongoing research and potential future direc-
tions, as discussed here, encompass the development of uPA/uPAR as diagnostic and
prognostic tools and underscore a dynamic landscape of possibilities. In summation, this
comprehensive review is intended to illuminate the diverse roles of uPA/uPAR and their
clinical significance across multiple medical fields, setting the stage for novel insights and
therapeutic avenues.
Author Contributions: Concept, research, literature review, manuscript preparation, revisions—
M.H., K.S.V., O.R., R.B., G.M.-R., A.R., J.W.-R., H.C., G.M. and A.R.L. Figures and tables research and
preparation, review, and revisions—K.S.V., M.H., O.R., A.R., J.W.-R. and A.R.L. All authors have read
and agreed to the published version of the manuscript.
Funding: This research received no external funding.
Data Availability Statement: No new data were created or analyzed in this study. Data sharing is
not applicable to this article.
Conflicts of Interest: Lucas is the founding scientist for Serpass Biologics, however this is a new
Biotech startup and has not provided funding for the research reported here.
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