The platelet contribution to cancer progression pot

REVIEW ARTICLE
The platelet contribution to cancer progression
N. M. BAMBACE and C. E. HOLMES
Division of Hematology and Oncology, Department of Medicine, University of Vermont, Burlington, VT, USA
To cite this article: Bambace NM, Holmes CE. The platelet contribution to cancer progression. J Thromb Haemost 2011; 9: 237–49.
Summary. Traditionally viewed as major cellular components
in hemostasis and thrombosis, the contribution of platelets to
the progression of cancer is an emerging area of research
interest. Complex interactions between tumor cells and circu-
lating platelets play an important role in cancer growth and
dissemination, and a growing body of evidence supports a role
for physiologic plate let receptors and platelet agonists in can cer
metastases and angiogenesis. Platelets provide a procoagulant
surface f acilitating amplification of cancer-related coagulation,
and can be recruited to shroud tumor cells, thereby shielding
them from immune responses, and facilitate cancer growth and
dissemination. Experimental blockade of key platelet receptors,
such as GP1b/IX/V, GPIIbIIIa and GPVI, has b een s hown t o
attenuate metastases. Platelets are also recognized as dynamic
reservoirs of proangiogenic and anti-angiogenic proteins that
can be manipulated pharmacologically. A bidirectional rela-
tionship between platelets and tumors is also seen, with evidence
of Ôtumor conditioningÕ of platelets. The platelet as a reporter of
malignancy and a targeted delivery system for antican cer
therapy has also been proposed. The development of platelet
inhibitors that influence malignancy progression and clinical
testing of c urrently available a ntiplatelet drugs represen ts a
promising a rea o f t argeted c ancer therapy .
Keywords: angiogenesis, cancer, metastases, platelets, TCIPA.
Introduction
Tumor cells interact with all major components of the

hemostatic system, including platelets. Platelets a nd platelet
activation have been linked to key steps in cancer progression
(summarized in Fig. 1). The contribution of platelets to
malignancy progression has been suggested to be a n organized
process that underlies the pathobiology of cancer growth and
dissemination rather than a simple epiphenomenon of neopla-
sia (reviewed in [1]). Here, we highlight current insights into
how platelets contribute to cancer growth, maintenance and
propagation and identify potential targets and directions for
platelet-directed anticancer therapy in the future.
Platelet structure and function
Often numbering over 3–4 trillion in a n individual patient with
cancer, platelets represent the smallest circulating hematopoi-
etic cells and are anucleate fragments formed from the
cytoplasm o f m egakaryocytes. The platelet membrane consists
of phospholipids and is covered with glycoproteins and
integrins, which are essential for adhesion, aggregation and
activation, the critical s teps in platelet-mediated hemostasis.
Important platelet membrane receptors include Glycoprotein
Ib-IX-V (GPIb-IX-V), Glycoprotein VI (GPVI) and Glyco-
protein IIb-IIIa (GPIIb-IIIa, also as integrin aIIbb3), receptors
that are essential for complete adhesion and aggregation [2,3].
Additional important receptors found on platelet membranes
include the protease-activated receptors (PAR ), PAR-1 and
PAR-4, and the P2 receptors, P2Y
1
and P2Y
12
,which
principally mediate activation a nd aggregation [4]. Platelets

also contain three types of granules: (i) dense granules
containing platelet agonists such as serotonin and ADP that
serve to amplify platelet activation, (ii) a granules containing
proteins that enhance the activation process a nd participate i n
coagulation; and (iii) lysosomal granules containing glycosid-
ases and proteases [5].
Many of the major structural components of platelets and
platelet receptors that contribute to hemostasis have a lso b een
found to relate to malignancy progression (reviewed in
Table 1). For example, in addition to coagulation-related
proteins, platelets also store proteins within the alpha granule
that can regulate angiogenesis and metastases [2,6]. Further,
platelet receptors such as GPIIb/IIIa can mediate platelet
angiogenic protein release in addition to their more t raditional
role in fibrinogen binding. At least one study has f ound
ultrastructural c hanges in platelets from p atients w ith l ung
cancer, including an increase in the number of platelet alpha
granules [7]. Interestingly, these r esearchers also f ound that the
number of alpha granules was associated with survival.
Functionally, platelets are complex cells capable of shape
change, translational protein production, protein and metab-
olite release, cell-cell interactions and paracrine regulation.
Most of these functions relate to the processes of platelet
activation and aggregation that occur following exposure to
Correspondence: Chris E. Holmes, Department of Medicine,
Hematology and Oncology, University of Vermont, Burlington, VT
05401, USA.
Tel.: +1 802 656 0302; fax: +1 802 656 0390.
E-mail:
Journal of Thrombosis and Haemostasis, 9: 237–249 DOI: 10.1111/j.1538-7836.2010.04131.x

Ó 2011 International Society on Thrombosis and Haemostasis
an in vivo stimulus. In thrombosis formation, thrombin and
collagen contribute substantially, but not exclusively, to
platelet activation in vivo [2]. In malignancy, tumor cells can
activate platelets by direct contact, or via release of mediators
such as ADP, thrombin, thromboxane A2 or tumor-associ-
ated proteinase s [8–11]. The relative importance of each
platelet activator in malignancy is unknown and some data
suggest the mechanism of platelet activation by tumor cells
may be tumor cell specific and, in some cases, mutually
exclusive [12].
Several studies have suggested an increase in platelet
activation in the blood of patients with cancer [13–19]. Both
tumor secretion of activators and direct contact with tumors
have been related to t his platelet activation [ 20–22]. To date, no
differences in platelet receptor function or composition have
been described in patients with a ctive malignancy a s compared
with healthy subjects to explain this increase in platelet
activation.
Early observations on platelets and cancer
Gasic et al., in 1968 [23], fi rst described t he association b etween
platelet number and metastatic cancer potential. This group
found neuraminidase-induced thrombocytopenia was associ-
ated with decreased metastasis of T A3 ascites tumor cells. This
antimetastatic effect was neutralized by infusion of platelet-
rich plasma (PRP). Thrombocytop enia experimentally induced
by a variety of mechanisms has also been associated with a
reduction in the number of m etastases in tumor transplant
models [23,24].
Thrombocytosis is observed i n 10–57% of patients with

cancer, with the number varyin g bas ed o n cancer type [1]. The
relationship between elevated platelet count and malignant
tumors was initially reported b y Reiss et al. in 1872 [25].
Subsequent studies have established this relationship for
common cancers, including colorectal, l ung and breast cancer,
as well as gastric, renal and most urogenital malignancies [26–
32]. Further, for the majority of malignancies, the extent of
platelet count elevation is inversely correlated with survival,
making thrombocytosis a marker of poor prognosis [26–32].
Insights into the mechanisms underlying the initial observa-
tions of thrombocytosis in malignancy h ave been forthcoming
in more recent decades. Sierko & Wojtukiewicz [1] have
recently summarized mechanisms underlying the humoral
interaction between tumor cells, bone marrow e ndothelial cells
(BMEC) and megakaryocytes. An important driver for
thrombocytosis in malignancy is the secretion o f t umor-derived
cytokines such as IL-1, G M-CSF, G-CSF and IL-6, which
stimulate thrombopoiesis through a thrombopoietin-depen -
dent mechanism, influencing largely m egakaryopoietic growth
and differentiation [33–38]. Megakaryocytes have a similar
ability to secrete inflammatory cytokines, which can in turn
influence bone marrow endothelial cells to support mega-
karyocytopoiesis [39,40]. VEGF and b-FGF are also released
by megakaryocytes, a nd influence megakaryocytic m aturation
and transendothelial migration via an autocrine loop [41–43].
Although incompletely elucidated, the interactions between
tumor c ells, megakaryocytes and bone marrow endothelial cells
appear to promote thrombopoiesis, and may influence tumor
angiogenesis.
Tumor cell-induced platelet aggregation, activation and

metastases
Platelets contribute t o critical steps in cancer metastasis,
including facilitating tumor cell m igration, invasion [44–46]
and a rrest within the vasculature [47–49]. In cellular models of
both breast cancer and ovarian cancer, invasiveness has
increased following exposure to platelets [46,50]. In the latter,
both activated platelet membranes and platelet releasate
increased invasion. Platelet contents may be released into the
peritumoral space following platelet activation and enhance
tumor cell e xtravasation and metastases [51–55]. An important
step in metastatic dissemination is the breakd own of v essel
basement membrane. By releasing proteolytic enzymes such as
gelatinase, heparanase and various matrix metalloproteinases
(MMPs), activated platelets can directly degrade s tructural
components, or alternatively, support this p rocess b y a ctivating
other proteinases and/or enabling tumor cells and endothelial
cells to do the same [46,56–58]. Moreover, modulation of
proteolytic activity is a ccomplished by g rowth factors released
Platelet
1
Angio-
genesis
Tumor
microenvironment
Tumor
cell
Tumor
thrombus
4
2

3
Fig. 1. Platelets are involved in ke y steps of malignancy progression. In in
vitro and in vivo m urine models, a role for platelets has b een d emonstrated
in tumor m etastasis, tumor growth a nd angiogenesis. Our w orking
unders tan ding of t he ro le of pla te let s in ma lig nan cy in vol ves : (i) tum or ce ll-
induced platelet aggregation can occur following tumor cell intravasation
into the v asculature, thereby ÔprotectingÕ or ÔcloakingÕ circulating tumor
cells from p hysical clearance a nd immune s urveillance, (ii) platelets facil-
itate t umor cell arrest within t he vasculature, endothelial cell retraction
and subsequent tissue invasion, (iii) platelets induce endothelial cell pro-
liferation and new blood vessel formation, which are requisi te for tumor-
associated angiogenesis and growth and (iv) platelet-tumor and platelet-
stromal interactions in t he tumor microenvironment depend, in part, on
platelet activation a nd platelet protein release, w hich contribute to the
inflammatory response. Additional platelet-related proteins an d metabo-
lites that facilitate proteolysis and tissue remodelling also e nhance tumor
growth and metastasis (including bony metastases).
238 N. M. Bambace and C. E. Holmes
Ó 2011 International Society on Thrombosis and Haemostasis
by platelets, a topic recently reviewed by Sierko &
Wojtukiewicz [1].
Tumor cells have the ability t o aggregate platelets, a finding
first reported in 1968, and referred to as tumor cell-induced
platelet aggregation ( TCIPA) [23]. It is now recognized that this
aggregation correlates with the metastatic potential of cancer
cells in b oth in vitro and in vivo mod els of experimental
metastasis [59,60]. The mechanisms by which tumor cells
induce platelet aggregation m ay differ by cancer type, but have
in common t he theme o f conferring surviva l advantage. I n
turn, platelets can protect tumor cells in at least two ways: by

coating them and thereby directly shielding t hem from physical
stressors within the vasculature and by p ermitting e vasion from
the i mmune system Õs effector cells. For example, platelets h ave
been shown to protect tumors from NK cells and T NF-a
cytotoxicity [61,62]. Timar et al. [63] have raised the hypothesis
that some malignant ce lls can acquire a platelet-like phenotype,
with expression of similar adhesion molecules and receptors.
This concept of Ôplatelet-mimicry Õ h as been suggested to relate
to the perceived lack of tumor-directed immune surveillance.
Recently, platelet-derived transforming growth factor-b has
been shown to down-regulate the activating immunoreceptor
NKG2D on NK cells and impair NK cell antitumor activity
[64].
Tumor-platelet aggregates have the a bility to d isseminate
and embolize within the pulmonary microvasculature and have
been directly observed to do s o in m urine models [65]. A brief
discussion of several major mechanisms of TCIPA a nd tumor
Table 1 Key p latelet components and their c ontribu tion to hemostasis a nd malignancy
Platelet
component
Principal role in
thrombus formation
Role in malignancy
Reference
In vitro models In vivo models
GPIIb/IIIa
(a
IIbb3
)
Activation allows

fibrinogen binding and
platelet plug
reinforcement
Tumor cell and platelet
interaction (via fibronectin,
fibrinogen and VWF)
demonstrated in numerous cell
lines; inhibition decreases
TCIPA and platelet-mediated
angiogenic growth factor
release
Decreased pulmonary
metastasis following
inhibition of receptor by
antibody and receptor
antagonists
[3,60,71,86–88,90,160,
165,166]
GP Ib-IX-V Binding of von
Willebrand factor;
anchors platelet to
subendothelium
Limited data to suggest role in
TCIPA; conflicting data on
tumor cell-platelet interactions
Pulmonary metastasis
decreased in mice lacking
GPIb but increased when
GPIb functionally
inhibited by monovalent,

monoclonal antibodies
[71,91,92]
GPVI Platelet adhesion to
collagen
Not studied to date 50% reduction in
pulmonary metastases in
GPVI-deficient mice
[93]
P-selectin Mediates platelet-
leukocyte tethering;
triggers leukocyte
activation
Facilitated interaction between
tumor cells and endothelial
cells via sialylated fucosylated
carbohydrates
Deficiency or blockade of
P- selectin inhibits the
formation of melanoma
metastases
[94–100]
P2Y
receptors
ADP-mediated platelet
aggregation
ADP-mediated VEGF release
from platelets; ADP induced
TCIPA
ADP depletion associated
with reduced metastases

[67–71,124,133,163,167]
PAR
receptors
Thrombin mediated
platelet activation
Selective release of angiogenesis
influencing proteins; induces
TCIPA
Promote metastases [11,122,123,168]
Alpha
granules
Storage of proteins that
enhance adhesive
process: fibrinogen,
VWF, MMP-II,
P-selectin, factor V,
PF-4, platelet
activating factor
Uptake and storage of
angiogenic proteins that are
selectively packaged and
released: VEGF, b-FGF
endostatin, angiostatin,
TSP-1; storage and release of
proteolytic enzymes and
metastasis influencing proteins
Maintenance of intra-tumor
vascular integrity
[6,117,120–122,126,141]
Platelet

microparticles
Enhances thrombosis
and secondary
hemostasis
Increased tumor cell
invasiveness, metastasis,
MMP-2 up-regulation and
angiogenesis; increased
leukemia, prostate and
breast cancer invasion/
migration
Increased chemo-
invasiveness and
metastases formation in
lung cancer models
[101–108]
The platelet contribution to cancer progression 239
Ó 2011 International Society on Thrombosis and Haemostasis
cell-induced platelet activation follows as their understanding is
pivotal to the development of s elective agents t argeting the
pharmacologic inhibition of these central pathways (see also
[66] for extensive review).
Adenosine diphosphate (ADP) is contained in platelet dense
granules and is considered a secondary mediator of aggrega-
tion. The major ADP receptors, P2Y
1
and P2Y
12
, are both
involved in platelet aggregation. Stimulation t hrough these

receptors a lso leads to shape change a nd thromboxane A
2
generation by platelets [67]. ADP contributes to TCIPA
induced by various tumor cell lines, including neuroblastoma,
melanoma and breast carcinoma [68,69]. The P2Y
12
receptor
plays a c entral role in platelet activation a nd in TCIPA [ 70,71].
For example, by generating ADP, MCF-7 breast carcinoma
cells activate and aggregate platelets via the P2Y
12
receptor [71].
Thrombin has a multifaceted role in hemostasis and
represents a key link between primary and secondary coagu-
lation responses. Thrombin has also been linked to tumori-
genesis and angiogenesis, with thrombin signaling being a
major contributor to metastatic tumor dissemination [72].
Thrombin has been detected in situ in numerous tumor types,
including small c ell lung cancer, r enal cell, melanoma and
ovarian cancer [73–75]. Tumor-enhancing effects of thrombin
include i nduction of TCIPA, increased tumor-cell adhesiveness,
promigratory and c hemotactic effects, an d up-regulation of
VEGF expression by tumor cells [76–79]. Importantly, t hrom-
bin is also the most potent platelet activator, and exerts its
function via the platelet PAR receptors, PAR-1 and PAR-4.
Secretion of ADP and thrombin by human tumor cells
activates platelets and recruits them to participate in TCIPA
[11]. Following thrombin-mediated platelet activation, up to
300 biologically act ive molecules can be released and deposited
ad lib at sites of vascular i njury, at the s ite of a wound or within

the tumor and tumor vasculature [6].
Cathepsin B, cancer procoagulant factor and the matrix
metalloproteinases (MMPs) are contributors to TCIPA.
Cathepsin B and cancer procoagulant factor can induce
platelet aggregation when released by tumor cells [80,81].
MMPs have demonstrated a similar ability to induce TCIPA in
vitro [82]. MMPs can be released by both platelets and cancer
cells in vivo ( reviewed in [83]). Jurasz and colleagues have
identified enhanced generation of MMP-2 as the potential
cause of human platelet aggregability in the setting of
metastatic prostate cancer [66].
Thromboxane A2 (TXA2) and its receptor (TX) also play
integral roles in platelet-tumor aggregation. It has been shown
that TX mediates platelet aggregation induced by murine and
tumor cell lines [84]. TXA2 can be generated by platelets as a
result of activation induced by other platelet agonists, an
observation that h ighlights the complex a nd interrelated nature
of platelet functional responses in the tumor.
Platelet adhesion receptors also play a critical r ole in t umor-
platelet cross-talk and the process o f h ematogeneous metastasis
(recently r eviewed i n [ 85]). T he r ole of the GPIIb-IIIa receptor
in TCIPA has been established f or decades, and numerous
metastatic models have highlighted the importance of this
receptor in the t umor-platelet i nteraction model [86–89]. A
recent role for the GPIIb/IIIa receptor in the release of
proangiogenic proteins and fibrinogen has also been elucidated
[87,90]. The i nvolvement of the i ntegrin receptor G PIba in
tumor metastasis, on the other hand, has b een more difficult to
define [59,86,91,92]. Recently, the GPVI s urface receptor, a
member of the immunoglobulin superfamily, which p rincipally

binds collagen, has become a subject of active investigation.
Importantly, a 50% reduction in experimental pulmonary
metastases in GPVI-deficient mice was reported by Jain et al.
[93]. Clinically, patients with GPVI deficiency exhibit a mild
bleeding tendency, suggesting that this receptor could poten-
tially be inhibited without major hemostatic consequence.
Finally, P -selectin is expressed on a ctivated platelets a nd
endothelial cells and has been identified as an important
mediator of the interaction between these cells and the vessel
wall [94]. This f acilitated interaction also applies t o tumor cells
as P-selectin can bind t o different tumor c ell lines through
binding of sialylated fucosylated carbohydrates [95,96]. In a
similar manner, P-selectin appears to facilitate interactions
between t umor cells and t he surrounding endothelium, at least
in the case of melanoma [97]. Deficiency or blockade of P-
selectin has inhibited the formation of metastasis in various
other e xperimental models [97,98]. This effect is most pro-
nounced in mucin-producing cancers [99,100].
Platelet microparticles and malignancy
When platelets are activated or exposed to high shear stress,
they release particles expressing membrane receptors and
cytoplasmic constituents termed platelet microparticles
(PMPs). A growing body of literature s upports the d irect
involvement of PMPs in malignant cell proliferation and
growth. PMPs have the ability to induce chemotaxis of many
hematopoietic cells and increase their adhesive affinity to
fibrinogen [101]. PMPs express multiple proteins and chemo-
kine receptors, which can be transferred to surrounding cell
membranes, including m alignant cells, which then b enefit from
enhanced invasiveness [102–105].

In vitro , P MPs h ave been shown to induce p roliferation
and tube formation of human umbilical vein endothelial cells,
to increase trans-matrigel chemoinvasion of lung cancer cell
lines, and to increase invasiveness of breast cancer cells [105].
In vivo, angiogenesis can be observed in the heart of ischemic
rats when PMPs are injected into myocardium [106]. Injection
of murine Lewis lung cancer cells coated with platelet PMPs
was associated with significantly more metastatic lung disease
[107]. Janowska-Wierczorek et al. [105] have recently demon-
strated that PMPs promote adhesion of tumor cells to
endothelium, induce chemotaxis and chemoinvasion, and up-
regulate MMP production. MMP-2 up-regulation and
increased malignant cell invasiveness have also recently been
reported in prostate cancer [108]. PMPs appear to represent
an important aspect of the functional interaction between
tumors and p latelets and may represent a novel treatment
approach in the future.
240 N. M. Bambace and C. E. Holmes
Ó 2011 International Society on Thrombosis and Haemostasis
The role of platelets in angiogenesis
Evidence supporting the link between platelets and angiogen-
esis has a ccumulated s ince Pined o and F olkman first raise d this
hypothesis [109]. The growth of s olid tumors and formation of
metastases depend on the generation of neovessels, and it is
recognized that tumor cells cannot grow beyond 2–3 mm in
size without a new vascular network [110]. These vessels are
needed not only to sustain and nourish the developing tumor
cells, but also to allow d elivery o f proteases and c ytokines that
permit further invasion, extravasation and dissemination. This
elaborate delivery and transportation system exists secondary

to an altered b alance between angiogenesis stimulators and
inhibitors. These proteins are released by many components of
the tumor microenvironment, including the tumor itself. This
tumor microenvironment is comprised of stromal fibroblasts,
resident macrophages and mast cells, mononuclear cells and
platelets [111–115].
Platelets contain over 30 important angiogenesis regulating
proteins. Platelets are now recognized as the major source of
VEGF (a pro-angiogenic protein) in serum a s t he platelet pool
comprises > 80% of total circulating VEGF in patients with
cancer as well as healthy individuals [116,117]. Of interest is the
observation that i n s ome cancers, plate let-derived VEGF better
predicts tumor progression than serum levels of VEGF [118].
Platelets also c ontain proteins that i nhibit angiogenesis,
including platelet factor-4 (PF-4), TSP-1 and endostatin
[119,120].
Under normal physiologic conditions, platelets have been
suggested to release angiogenic proteins to promote wound
healing. These p ro-angiogenic proteins are later counterbal-
anced by the r elease of angiogenic inhibitors from stro mal cells
andplatelets,tostopuncontrolledgrowthinlaterstagesof
healing in non-malignant wounds [121]. These angiogenic
mediators are packaged into d istinct alpha granule popula-
tions, and selective release based on selective engagement of
platelet receptors has been proposed [122]. Ma and colleagues
first introduced the concept of differential release of platelet
angiogenic proteins, by demonstrating that PAR-1 activation
was associated with VEGF r elease a nd suppression of endost-
atin, while PAR-4 activation, conversely, s timulated endostatin
release and suppressed release of VEGF [123]. These investi-

gators subsequently treated rats with established gastric ulcers
with an oral PAR-1 antagonist or vehicle. In this model,
significant healing of ulcers did not occur in the rats treated
with the PAR-1 antagonist [123].
Subsequently, the ADP receptors, P2Y
1
and P2Y
12
, have
been demonstrated to participate in the regulation of angio-
genic protein release, though this pathway of platelet activation
appears to release less VEGF than thrombin-mediated activa-
tion [124]. ADP-mediated platelet ac tivation is associated with
a net increase in the release of VEGF in healthy individuals,
with no effect on endostatin release. T his VEGF r elease can be
abolished by selectively inhibiting the P2Y
12
receptor [124].
The source and mechanism of platelet-derived angiogenesis
proteins remain under a ctive investigation in both h ealthy
individuals and patients with cancer. Recent s tudies have
offered insight. For example, in the circulation, platelets have
been shown to uptake and store proteins that regulate
angiogenesis [1,125,126]. In addition to protein u ptake,
Zaslavsky et al. [120] have recently demonstrated that the
platelet source of TSP-1 is megakaryocyte derived, suggesting
that enhanced production or endocytosis by marrow precursor
cells ma y c ontribute to t he platelet angiogenic p rotein c ontent.
Based o n t he findings that VEGF-A was regulated by Il-6 in a
megakaryoblastic cell line, Salgado et al. [127] bring forward

the h ypothesis that higher VEGF l evels in cancer patients may
partly result from an IL-6 mediated up-regulation of the
expression of VEGF-A in platelet precursors.
In vitro, proangiogenic effects of platelets were observed by
Pipili-Synetos et al. [128], who noted that platelets stimulated
endothelial cell proliferation and growth of capillary-like
structures in Matrigel assays. An additional in vivo model of
angiogenesis showed a reduction of retinal neovascularization
in mice with induction of thrombocytopenia as well as
inhibition of platelet aggregation by a highly specific alpha-
IIbbeta3 receptor antagonist or aspirin [129]. This resulted in a
35–50% reduction of retinal neovascularization, further
supporting the platelet contribution to angiogenesis [129].
Kisucka et al. also examined the role of platelets in four
in vivo animal models of angiogenesis using both a cornea
and Matrigel assay. They report t hat platelet-depleted mice
experienced a significant reduction in corneal neovasculariza-
tion and developed hemorrhage, and postulate that platelets
support angiogenesis through release of growth factors and
platelet-vessel w all i nteractions [130]. Brill has also demon-
strated the role of platelet microparticles in models of
angiogenesis [106].
Importantly, a clear understanding of the contribution of
platelets specifically to tumor-associated angiogenesis remains
under investigation. For example, while platelets enhance
angiogenesis as in the examples above, platelet-endothelial
interactions in tumor microvessels have been found to be
reduced in murine models of tumor angiogenesis [131]. The
platelet as a scavenger of VEGF and therefore a potent anti-
angiogenic cellular component of the tumor microvasculature

couldalsobeconsidered.
A complex and bidirectional relationship between tumor
cells and platelets
There is growing evidence to suggest that the interplay between
platelets and tumors is neither passive n or unidirectional
(Fig. 2). Complex relationships between host, tumor and
platelet within th e cancer patient will need to be carefully
delineated and significant research efforts are required if
antiplatelet therapy is to be used successfully in the clinical
setting. The platelet role in coagulation-mediated cancer
progression, the platelet contribution to the tumor-stromal
interaction and the contribution of platelets to inflammation
and its subsequent role in malignancy progression are just
several examples of these relationships [132]). Shared tumor cell
The platelet contribution to cancer progression 241
Ó 2011 International Society on Thrombosis and Haemostasis
and p latelet agonists and receptors offer both opportu nity and
potential obstacles for d rug targeting. For example, drugs that
inhibit the P2Y receptors on platelets may also interact with
endothelial and cancer cell P2Y receptors and contribute to the
overall impact of the drug [133–135]. The well-delineated role
of thrombin signaling and activation of PARs found on
malignant cells is another example of shared targets between
tumor cells and platelets (reviewed in [136,137].
Some evidence suggests that platelets can be conditioned in
vivo by tumor cells to deliver anti-angiogenic proteins [121,138].
In a murine model, Kerr et al. [138] have r ecently d emonstrated
that platelets preferentially store tumor-derived GM-CSF,
TPO, TNF-a,TGF-bı and especially MCP-1 over host-derived
proteins. An emerging concept in the literature focuses on the

platelet as a reporter of malignancy. For e xample, both platelet
associated PF-4 and TSP-1 have been associated with early
cancer growth and been proposed as biomarker s of early tumor
progression [120,139].
Platelet granule proteins not only promote growth o f tumor
vessels, but prevent tumor hemorrhage, presumably by main-
taining the integrity of the existing tumor vascular supply
[140,141]. Though t he precise mechanism underlying this
phenomenon has not been fully elucidated, t his appears t o
occur i ndependently from thrombus formation. The prevention
of tumor hemorrhage by platelets has more recently b een found
to relate, i n part, to their ability to modulate vascular damage
by tumor-infiltrating leukocytes [142]; an observation that
further illustrates the complex tumor-stromal interaction,
including the ability of platelet to influenc e inflammatory
responses [140,143–145]. These observations suggest that the
mechanism underlying the maintenance of neoplastic v essels by
platelets m ay b e distinct from that used fo r maintenance of host
vessels, rendering pharmacologic inhibition of the former
plausible. Selective platelet storage and release of stimulatory,
inhibitory and regulatory proteins represents a novel concep-
tual framework to be explored in the understanding of tumor
angiogenesis.
Antiplatelet therapy in the treatment of cancer
In 1989 and 1993, Dr Leo Zacharski and colleagues, writing
for the Scientific and Standardization Committee of the
International Society on Thrombosis and Haemostasis Sub-
committee on Hemostasis and Malignancy, published an
update of clinical trials using antiplatelet therapy and antico-
agulants i n c ancer [146,147]. A t the time, over 2 0 studies, most

of them pilot studies with 50 patients or fewer, were reported
in the literat ure using an ant iplatelet drug in the treatment (not
prevention) of cancer. T he majority of thes e studies focused o n
the drug dipyridamole in non-randomized studies, which
reported variable response rates. An a nalogue of dipyridamole
(RA-233, mopidamol) has a lso been studied in prospective
randomized studies, with no survival benefit demonstrated in
small cell and ovarian cancer but an approxim ate 1 00-day
improvement in s urvival in non-small-cell lung cancer patients
[148–150].
The r emaining prospective studies using antiplatelet therapy
focused on the use of aspirin in renal cell and small cell lung
cancer and showed no effect [151]. Aspirin use has been most
extensively studied in colorectal and breast cancer, with
demonstrated efficacy in the colorectal cancer prevention
setting [152]. Aspirin-mediated inhibition of platelet aggrega-
tion is well documented, and recently aspirin has also been
shown to attenuate platelet protein release [153]. In vivo data
suggesting a possible inhibitory role in the formation of
metastasis were initially reported by Gasic et al. [154], who
observed metastatic inhibition of MCA6 ascites sarcoma cells
in mice, in the presence of a spirin. In a more recent publication,
aspirin but not indomethaci n suppressed the formation o f lung
metastasis in a metastatic hepato-cellular murine model [155].
Antimetastatic effects of aspirin, however, have not been seen
consistently in all laboratory models.
Fig. 2. A d iagramatic representation of th e multiple bidirectional inter-
actions between pla telets and tum or cells. P latelets and tumor cells express
many of the s ame receptors, illustrating the concept of Ôplatelet mimicryÕ.
These receptors, such as GPIb and GPIIbIIIa, may participate in T CIPA

by promoting a rrest of tumor cells i n the vasculature and b y promoting
interactions with bridging proteins such as VWF, fibronectin and fibrin-
ogen. Tissue factor expression is u p-regulated in tumor cells, leading to
thrombin generation. Tumor cells also have the ability t o directly secrete
platelet agonists such as ADP and thrombin, which activate platelet s in the
tumor microenvironment. In turn, activated platelets secrete g rowth
factors and prot einases that can re gulate tumor growth a nd invasion.
Activated plate lets shed m icroparticles, which facilitate cell i nvasion and
angiogenesis. ADP, a denosine diphosphate; PAR, proteinase-activated
receptors; P2Y, P2Y receptors; GPIb, Glycoprotein I b; GPIIbIIIa,
Glycoprotein IIbIIIa ; VWF, V on Willebrand f actor; Lpa, lyso phos-
phatidic acid; Txa2, thromboxane; PMP, plate let microparticle.
242 N. M. Bambace and C. E. Holmes
Ó 2011 International Society on Thrombosis and Haemostasis
Clinical data evaluating the impact of aspirin therapy on
cancer survival have begun to emerge. Fontaine et al. [156]
have rece ntly reported p re liminary d ata suggesting t hat aspirin
used in combination w ith the surgical treatment o f non-small-
cell lung cancer is assoc iated with increased survival. Similarly,
in a prospective observational study of women d iagnosed with
breast cancer, as reported in the NursesÕ Health Study, aspirin
use w as associated with decreased risk of breast cancer
recurrence and death [157]. Add itionally, a spirin use was
found to decrease the proangiogenic effects of tamoxifen in
patients with breast cancer [116]. Importantly, the clinical
benefits of t he drug are likely to also relate to its anti-
inflammatory effects.
A review of selected published human clinical studies using
antiplatelet th erapy in the treatment of c ancer is found in
Table 2. T his t able does not contain data related to the role o f

antiplatelets (such as aspirin) in cancer prevention or the
potential antiplatelet (p-selectin inhibition) e ffect of heparin
(the latter recently reviewed in [85]). Additional p ilot studies of
antiplatelet drugs alone or in combination with additional
chemotherapy have been reviewed by Hejna et al. [158]. The
table h ighlights the paucity of clinical trial data using currently
available antiplatelet agents. Importantly, while we have
recently reported on t he use o f aspirin therapy i n women with
breast cancer receiving tamoxifen therapy [116,159], there is a
paucity of data to support t he combination of antiplatelet
therapy with existing tumor-targeted therapy.
Despite the limited number of prospective randomized
trials, the laboratory data using antiplatelet therapy continue
to accumulate. Early laboratory studies focused on p rosta-
cyclin and p rostacyclin analogues, which have been previ-
ously reviewed [158]. In addition, blockade of the GPIIb/IIIa
receptor using t he monoclonal a nitbody 10E5, an inhibitor
of human platelet GPIIb/IIIa, decreased lung colonization of
cancer cells [160]. A challenging aspect of the administration
of GPIIb/IIIa antagonists in the clinical setting has been the
need for intravenous administration of these agents, which
are now widely used in high-risk acute coronary syndromes.
Recently, however, an oral inhibitor of GPIIb/IIIa, XV454,
has h alted experimental metastasis formation in a murine
Table 2 Clinical out comes associated with the u se of platelet inhibitors in patients with canc er. Limited clinical da ta are a va ilable on t he impact of plat elet
inhibitors on clinical o utcomes in patients diagnosed wit h cancer. Murine model data are r eviewed in the text and in Table 1
Platelet inhibitor
or modulator
Mechanism of platelet
inhibition

Type of cancer(s)
studied Protocol designs Observations in clinical studies Reference
Aspirin Inhibits platelet thromboxane
production and platelet
aggregation (anti-neoplastic
effects of this drug are also
anticipated to rely on COX-2
tissue and tumor inhibition)
Colon cancer Double blind No difference in overall survival [169]
SCLC Unblinded,
randomized
No effect on survival [170]
Renal cell
carcinoma
Prospective
randomized
No significant response or
effect on survival
[151]
Breast cancer Prospective
observational
study
Decreased recurrence and
mortality from breast cancer
[157]
NSCLC
(early stage)
Retrospective
analysis
Increased survival

post-resection
[171]
Prostate
cancer
Retrospective
analysis
Improved PSA control in
patients undergoing radiation
[172]
Benoral (aspirin-
acetaminophen
conjugate)
Breast cancer Double blind No significant response or
improved survival
[173]
Clopidogrel P2Y12 receptor antagonist;
inhibits platelet aggregation
induced by ADP
Prostate cancer Retrospective
analysis
Improved PSA control in
patients undergoing radiation
[172]
RA-233
(Mopidamole)
Dipyridamole derivative;
increase in platelet cyclic
AMP; decreased platelet
aggregation
Colon cancer Double blind No significant response [149]

NSCLC
(early stage)
Double blind Improvement in survival in
limited stage/resected disease;
no effect in disseminated
disease
[149,150]
SCLC Prospective
randomized
No significant response [150]
Ovarian cancer Prospective
randomized
trial
No effect on survival or
recurrence
[148]
Dipyridamole Increase in platelet cyclic
AMP; decreased platelet
aggregation
Colon cancer
(advanced)
Prospective
randomized
trial
No impact on survival or
response
[174]
NSCLC
(advanced stage)
Prospective

non-randomized
No significant response
compared with historical
controls
[175]
The platelet contribution to cancer progression 243
Ó 2011 International Society on Thrombosis and Haemostasis
model o f lung c ancer [87]. I ntegrilin, a commercially
available platelet-specific aIIbb3 integrin antagonist, was
administered to mice after establishment of bony metastases
in a study by Boucharaba and colleagues, evaluating the role
of platelet-derived lysophosphatidic acid. This resulted in
thrombocytopenia, decreased circulating Lpa plasma levels
and a significant reduction in the number of osteolytic bony
metastases [51].
Wenzel et al. have recently reported successful in vivo
reduction of pulmonary metastases in a murine model of
breast cancer using the platelet aggregation inhibitor cilostazol.
By administrating liposomal cilostazol intravenously, they
observed decreased ex vivo platelet aggregability and decreased
platelet-tumor complex formation [161]. Similar results were
obtained using liposomal dipyramidole [162]. Few studies
evaluating the common ADP receptor inhibitors, clopidogrel
and ticlopidine, h ave been reported a nd they demonstrated
limited success [163].
Conclusion
Platelets play a multifaceted and important role in cancer
biology (Table 3). The existing research suggests a compelling
biological rationale for attempting to disrupt tumor-platelet
cross-talk, with the goal of down-regulating tumor invasion,

angiogenesis and spread. In the laboratory, platelet receptors,
both constitutive and activation dependent, such a s G P1b/IX/
V, P-selectin and alphaIIb-beta3 integrin, c an promote the
progression and metastases of various tumor types and are
obvious targets for further clinical study [164]. Additionally,
control of the platelet reservoir of angiogenic proteins, which
are both secreted and sequestered in a selective manner,
represents an approach to angiogenic control within t he tumor
microenvironment.
The study of platelet inhibitors in the clinical setting will
require a careful consideration of not only cancer type but stage
of disease targeted. Importantly, appropriate trial endpoints
must be chosen that are not by des ign predicated on dir ect and
toxic tumor effects and secondary rapid cell kill and tumor
shrinkage. A potential barrier that surrounds chronic admin-
istration of antiplatelet agents in the setting of active malig-
nancy is directly related to the paramount role that platelets
play in maintaining hemostasis. Currently available oral
antiplatelet agents irreversibly inhibit their target, making the
risk of bleeding more difficult to mitigate. Future work in the
development of novel agents would ideally yield a molecule
able to inhibit platelet-tumor interaction while maintaining
sufficient platelet function to prevent bleeding. Potential new
classes of a gents include antibodies against P-selectin, platelet-
specific oral integrin i nhibitors, PAR-1 antagon ists and block -
ade of platelet-derived LPA.
How should we combine antiplatelet therapy with con-
ventional cancer cell-directed therapy? Will other host factors
that influence platelet activation, such as diabetes, be
important in patient selection [135]? Existing antiplatelet

drugs, such as aspirin and clopidogrel, remain understudied
as ad juvants t o conventional chemotherapeutic and hor-
monal therapies, particularly in animal models and the
clinical setting. Increasing our translational database on the
anticancer biology of a ntiplatelet strategies to i nclude com-
bination therapy and studies directed at prevention vs. l ow
burden vs. h igh burden disease are imperative f or the
successful clinical translation of results. Importantly, we have
learned much from the use of antiplatelet therapy in the
treatment of cardiovascular d isease, such as t he concept of
drug resistance. These considerations might be applied
prospectively in oncologic s tudies. F uture c linical trials
formally addressing the role of antiplatelet therapy will need
rigorous attention to patient selection, combination therapy
with existing agents and trial e ndpoints but offer the
hematologic community a significant opportunity to poten-
tially improve cancer outcomes.
Table 3 Overview of important platelet-cancer cell interactions and their po ten tial influence on cancer p rogression. A full discussion of t hese interactions is
found in the text. These observa tions reflect in vitro and murine model data
Platelet-related mechanism Effect
Platelet activation
Increased in patients with cancer
Facilitated by contact with tumor cells
and tumor release/production of platelet
agonists such as ADP and thrombin
Platelet activation enhances tumor cell-induced platelet aggregation, releases
chemotactic cytokines, proteolytic enzymes and platelet microparticles that can support
cancer growth and extravasation as well as angiogenesis
Platelet activation provides a procoagulant surface to facilitate cancer-related
coagulation

Inhibition of key platelet activation and aggregation receptors decreases metastases
Tumor-cell-induced platelet aggregation (TCIPA) Platelet aggregation correlates with metastatic potential in in vivo and in vitro models
Protection of tumor cells from environment Platelets provide mechanical shielding from physical stressors
Platelet-derived proteins down-regulate immune cells, thereby impairing their antitumor
activity
Production of platelet microparticles (PMPs) Transfer of receptors to tumor cell membranes, which may increase invasiveness. May
regulate MMP production and influence invasion
Release of angiogenic proteins Platelets contain pro- and anti-angiogenic proteins packaged into distinct alpha
granules, which can be differentially released to support angiogenesis
Prevention of tumor hemorrhage Platelets maintain tumor vascular integrity and reduce tumor hemorrhage
Platelet-enhanced metastases Platelets facilitate tumor cell migration and extravasation
244 N. M. Bambace and C. E. Holmes
Ó 2011 International Society on Thrombosis and Haemostasis
Disclosure of Conflict of Interests
The authors state that they have no conflict of interest.
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