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Anti-platelet treatments in cancer: Basic and clinical research
Thrombosis Research 164 (2018) S106–S111
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Full Length Article
Anti-platelet treatments in cancer: Basic and clinical research ⁎
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Paolo Gresele , Marco Malvestiti, Stefania Momi Center of Thrombosis and Haemostasis, Department of Medicine, University of Perugia, Perugia, Italy
A R T I C L E I N F O
A B S T R A C T
Keywords: Cyclooxygenase Metastasis P2Y12 Phosphodiesterases Platelets Platelet-therapy
Over the past few decades the central role that platelets play in cancer development and progression, and especially in metastasis, has been elucidated. The molecular mechanisms responsible for initiating and mediating tumor cell-induced platelet aggregation and secretion have been largely unravelled. Considerable mechanistic insight into how platelets contribute to tumor angiogenesis, immunoevasion and cancer cell invasion have been clarified and, consequently, platelets have been identified as potential new drug targets for cancer therapy. This article gives an overview of the platelet-targeted pharmacologic approaches that have been attempted in the prevention of cancer development, progression and metastasis, including the application of antiplatelet drugs currently used for cardiovascular disease and of new and novel strategies.
1. Introduction The involvement of platelets in cancer growth and metastasis is a longstanding concept. An inverse correlation between platelet count and disease-specific survival has been described for several cancers and numerous basic and clinical research observations show that platelets affect disease burden and treatment efficacy in cancer patients, and participate in cancer metastasis. Platelets physically and functionally interact with various tumor cells through surface receptors including integrins. β1 integrins and β3 integrins in particular participate in platelet–tumor cell interaction and in tumor metastasis [1]. Platelets display their pro-metastastic role through the secretion of pro-metastatic factors (e.g. autotaxin), chemoattraction of granulocytes to platelet/tumor aggregates, activation of epithelial-mesenchimal-like transition (EMT) via direct contact with tumor cells and/or secretion of TGFβ1, release of microvesicles containing microRNA-223 which targets the tumor suppressor EPB41L3, adhesion to tumor cells providing physical protection from direct contact with NK cells [2]. The interplay between platelets and tumor cells involves the formation of platelet–tumor cell aggregates in the circulation, with platelets forming a shield for tumor cells allowing the latter to escape natural killer (NK) cell and tumor necrosis factor (TNF)α cytotoxic activity. Platelets may also help tumor cell extravasation to the metastatic niche. On the other hand platelet activation releases plateletderived growth and proangiogenic factors that may, contribute to tumor growth and angiogenesis. Platelets flowing through the tumor vasculature get activated,
adhere to the neovascular endothelium, aggregate and become part of the tumor's microenvironment, thus potentially influencing the tumor parenchyma and stroma of most solid tumors, particularly carcinomas. Platelets contribute to tumor persistence in the circulation by shielding tumor cells from destruction by NK cell [3–5] and suppress IFN-γ secretion by NK through the interaction between glucocorticoid-induced TNF-related ligand (GITR), a member of the TNF receptor superfamily that acts as a NK-inhibitory ligand on the surface of NK and its ligand (GITRL) present on platelet's surface [10–14]. Platelets contribute to metastasis and distant tumor growth by aiding tumor cell attachment to the endothelium and by releasing angiogenic and growth factors, such as vascular endothelial growth factor (VEGF) [6,7] and transforming growth factor-β (TGF-β) [8,9]. Tumor cell invasion through the extracellular matrix (ECM) is a crucial step in tumor metastasis, and this process is mediated by proteolytic enzymes, such as matrix metalloproteinases (MMPs), that degrade the ECM surrounding blood vessels to allow cancer cells to penetrate in tissue. Indeed, the ECM surrounding blood vessels plays a critical role in the limitation of extravasation and intravasation of tumor cells. Platelet-derived TGF-β1 is crucial in inducing tumor growth and metastasis by up-regulating MMP-2 and MMP-9 [15] and by activating the TGFβ/SMAD and NF-kB pathways in cancer cells [16]. The importance of platelets in cancer growth is supported by the finding that platelet depletion results in a marked reduction of tumor growth in an orthotopic model of ovarian cancer, in vivo models of experimental pulmonary metastasis and in a murine model of spontaneous metastasis [17]. The interaction between cancer cells and/or the neovessels
⁎ Corresponding author at: Section of Internal and Cardiovascular Medicine, Department of Medicine, University of Perugia, Nuovo Polo Didattico, Strada Vicinale Via delle Corse, 06132 Perugia, Italy. E-mail address: [email protected] (P. Gresele).
https://doi.org/10.1016/j.thromres.2017.12.016 Received 24 November 2017; Accepted 27 December 2017 0049-3848/ © 2018 Elsevier Ltd. All rights reserved.
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prevention of CRC [35–38], cumulatively showed a significantly reduced risk of developing an adenoma on follow-up (0.83, 95%CI = 0.72 to 0.96, p = 0.012) [39]. In carriers of hereditary predisposition to non-polyposis colon cancer aspirin was shown to prevent the development of cancer [40] and to prolong survival when started after diagnosis [41]. Given that in most of these trials high doses of aspirin were used, the doubt that the effect of aspirin would be essentially due to an action on tumor cell COX-2, requiring higher doses, and that therefore long term platelet-selective low-dose, aspirin would not prevent CRC, was risen. Therefore, a follow-up cumulative observation study of four large randomized trials on aspirin (75–300 mg/daily) versus control in primary and secondary prevention of vascular events, involving over 14,000 patients followed-up for a median of 18.3 years, was performed showing that allocation to aspirin significantly reduced the 20-year risk of colon cancer (HR 0.76, 95%CI 0.6- to 0.96, p = 0.02) and cancer-related mortality (HR 0.65, 95%CI 0.48 to 0.88, p = 0.005), but not that of rectal cancer, and that the benefit increased with treatment duration such that at least 5 years of treatment were required to reduce colon cancer risk. Moreover, benefit was evident with aspirin doses as low as 75 mg/day, with no further increase of benefit with greater doses [42]. This is an important observation because it is known that the risk of major bleeding with aspirin is dosedependent [43,44]. Other subsequent meta-analyses of individual patient data from large aspirin studies of aspirin in cardiovascular prevention showed that aspirin reduced mortality also from non-gastrointestinal cancers [45], prevented metastasis from all cancers [46] and that the effect was only evident from 5 years of continued treatment onward [47]. These conclusions have been criticized because of the retrospective nature of the analysis on non-pre-specified end-points, because most of the recorded cancer events occurred well after the end of the randomized follow-up phase of the studies, and because some important trials, like the Women's Health Study, had been excluded from the analysis. Therefore, several ad hoc designed adjuvant trials of low-dose aspirin have been initiated in the last few years, in patients with various types of newly diagnosed cancer, and the protocols of some large, ongoing, primary cardiovascular prevention trials with aspirin (ASCEND, ACCEPT-D, ARRIVE and ASPREE) have been modified to collect prospective information about cancer incidence. The results of these ongoing studies will finally provide prospective evidence on the role of chronic aspirin in chemoprevention.
irrorating tumors and platelets leads to the triggering of adhesion and activation of the latter, a process also called tumor-cell induced platelet aggregation (TCIPA). TCIPA is mediated by multiple agents: tissue factor (TF), thrombin, adenosine diphosphate (ADP), TxA2, and MMP [18]. Therefore the use of antiplatelet agents as adjuvant cancer therapy or the exploitation of platelets as carriers of antitumor agents are attractive approaches. 2. Cyclooxygenase inhibitors Cyclooxygenase (COX) is the enzyme transforming arachidonic acid into the precursor (PGG2-PGH2) of different prostaglandins (PGs) or of thromboxanes (Tx), depending on the cell type. Arachidonic acid is cleaved from membrane phospholipids by phospholipase A2 and is then transformed by cyclooxygenases (COXs) into PGH2, which is rapidly converted into prostanoids by different PGor Tx-synthases [19]. The COX-1 isoenzyme preferentially couples with TXA2 synthase (in platelets) and PGF2 synthase (PGFS), whereas COX-2 is associated mainly with PGI2 synthase (PGI-S) (in endothelial cells) and PGE synthase (PGEs) in several cell types, including mucosal cells of the gastrointestinal tract [20]. Evidence exists for an antitumor activity of aspirin, which acts as an active-site acetylating agent inducing an irreversible inactivation of COX-1 (through acetylation of Ser 529) and COX-2 (through acetylation of Ser 516), with the consequent suppression of prostaglandin and thromboxane production [21,22]. The inhibition of platelet TxA2 production by aspirin is the basis of the most widely diffused antiplatelet therapy in clinical use. 2.1. Preclinical studies Early observations showed that tumor metastases were reduced in rats treated with aspirin [23] and that prostaglandin concentration was raised in rat colorectal tumor tissue [24,25], opening the way to the studies on a potential benefit of aspirin in cancer. Supporting observations showed that oral administration of aspirin significantly inhibited the incidence and number of invasive, azoxymethane-induced adenocarcinomas of the colon in rats [26] as well as the onset of lung tumors induced by a tobacco-specific nitrosamine in mice [27]. Aspirin, administered to mice at a dose equivalent to low-dose in humans (around 100 mg/day), induced apoptosis of colorectal cancer (CRC) cells by a mechanism involving the downregulation of the IL-6-STAT3 signalling pathway [28]. The antitumor activity of aspirin is considered to be driven by the inhibition of COX-2, which is overexpressed in cancer cells [29,30], but also by the inhibition of platelet adhesion to tumor cells, and of TCIPA, essential steps in the protective effects by platelets from immune surveillance against cancer cells [31]. CRC cells (HT29) previously exposed to platelets in vitro transform into mesenchymal-like cancer cells which, injected into the tail vein of immunodeficient NOD-SCID mice, produce a higher number of lung metastases compared to HT29 cells not exposed to platelets, an effect associated with enhanced systemic biosynthesis of TXA2 and PGE2 in vivo. Aspirin administration prevented the increased rate of metastasis as well as the enhanced production of TXA2 and PGE2 induced by platelet-primed HT29 cells [32].
3. PDE inhibitors Phosphodiesterases (PDEs) catalyze the hydrolysis of cyclic (c) AMP and cGMP, two powerful intracellular inhibitory second messengers, to inactive 5′AMP and 5′GMP. Of the over 60 different PDEs known, platelets express PDE2, PDE3, and PDE5 and their inhibition rises the intraplatelet levels of the two cyclic nucleotides thus inhibiting platelet activation [48]. Among the isoenzyme-selective PDE inhibitors developed as antiplatelet agents, cilostazol and dipyridamole have been explored as possible adjuvant therapy in cancer. 3.1. Cilostazol Cilostazol (6-(4-(1-cyclohexyl-1H-tetrazol-5-yl)butoxy)-3,4-dihydro-2(1H)-quinolinone) is a specific and strong inhibitor of PDE3 in platelets and smooth muscle cells, it is in clinical use since 1999 for the treatment of intermittent claudication [49] and has shown protective effects on ischemic cardiovascular events [48]. Its effect on cancer has been only evaluated in preclinical studies.
2.2. Clinical studies From a case-control study, chronic aspirin use among CRC-patients was significantly lower than among age/sex-matched non-CRC-controls [33]. Later, a meta-analysis cumulating data from several case-control studies in CRC patients, involving > 30,000 subjects, concluded that regular aspirin use was associated with a reduced risk of CRC (pooled odds ratio (OR) 0.62, 95%CI 0.58–0.67, p < 0.0001) [34]. A few small randomized, double-blind, placebo-controlled trials, involving around 3000 participants, evaluating chronic aspirin for the secondary
3.1.1. Preclinical studies Cilostazol suppressed the migration of human colon cancer cells, DLD-1, induced by soluble fibronectin or fetal bovine serum in a S107
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recommended further studies [66]. Despite these apparently positive results, it must be considered that they came from small and mostly uncontrolled studies, and that a number of other small trials examining the potential usefulness of dipyridamole to enhance chemotherapeutic efficacy in sarcoma, colorectal, breast, renal, and prostate cancers failed to show meaningful improvement in response [67].
phagokinetic assay and it suppressed cancer cell invasion induced by fetal bovine serum in a trans-cellular migration assay, showing that cilostazol inhibits colon cancer cell motility [50]. The adjuvant antitumor effect of cilostazol in association with cisplatin was assessed in a xenograft tumor model in nude mice injected with cells derived from a human oral carcinoma or with heLa cells or with the respective cisplatin-resistant cell lines. A synergic antitumor effect of the combination of cilostazol and cisplatin was observed in cisplatin-resistant cell lines, with a significant increase in the number of apoptotic cancer cells [51]. In vitro, the addition of B16 tumor cells to washed platelets induced aggregation, an effect inhibited by cilostazol [52]. Moreover, cilostazol administered 2 h before the injection of human adenocarcinoma cells in nude mice strongly inhibited lung metastasis, while it was ineffective when administered 72 h after tumor cell injection, suggesting that platelet aggregation induced by tumor cells plays an important role in metastasis development [53].
4. ADP receptor antagonists Purinergic receptors (P2 receptors) are present on the platelet surface and bind ADP triggering platelet aggregation [68]. P2Y12, the Gαicoupled ADP receptor on platelets [69], is the target of all currently used antiplatelet ADP antagonists [70]. On the other hand, P2Y12 has also been shown to be present in some tumor cells (intestinal epithelial carcinoma cells, glioma C6 cells, ovarian cancer cells, etc.) in which it regulates cell growth [71]. In a model of lung metastasis induced by Lewis Lung Carcinoma cells (LLC), P2Y12-deficient mice showed a lower tumor burden, confirming a role of platelet P2Y12 in tumor metastasis [72]. P2Y12 deficiency diminished the ability of LLC cells to induce platelet release of active TGFβ1, resulting in a reduced platelet-induced EMT-like transformation of LLC cells, a prerequisite for LLC cell metastasis. In accordance, platelet P2Y12 deficiency resulted in significantly less lung metastasis in a B16 melanoma experimental murine metastasis model [72].
3.2. Dipyridamole Dipyridamole (2,6–bis (diethanolamino)–4,8-dipiperidino-pyrimido 5,4-d pyrimidine), synthesized more than fifty years ago and also used as a coronary vasodilator, is currently in clinical use as an antithrombotic drug for secondary prevention of stroke in association with aspirin. Dipyridamole inhibits platelet function not only by acting as an inhibitor of PDE5 and PDE3 but also by inhibiting the reuptake of adenosine by red blood cells. In this way dipyridamole enhances the plasma levels of this platelet-inhibitor and vasodilatory nucleoside, and contribute to the scavenging of free radicals that inactivate vessel wall COX, thus enhancing PGI2 biosynthesis [48, 54–57].
4.1. Clopidogrel Clopidogrel, a thienopyridine, is an oral P2Y12 inhibitor acting as a pro-drug that requires metabolic conversion to its active form by the liver. Indeed, about 85% of the absorbed clopidogrel is metabolized into an inactive metabolite (SR26334) by plasmatic carboxylases and the remaining 15% is metabolized, by a two-step biotransformation process dependent on cytochrome P450 isoenzymes in the liver [73,74], in R130964, the active circulating metabolite that irreversibly blocks P2Y12 [75–77].
3.2.1. Preclinical studies Dipyridamole inhibited TCIPA and reduced liver metastases in athymic nude mice injected with a human pancreatic adenocarcinoma cell line (RWP-2) [58]. Moreover, dipyridamole has shown potential benefit against cancer multidrug resistance, a phenomenon that decreases the clinical benefit of several anticancer agents, including doxorubicin. Treatment with dipyridamole of mice injected with doxorubicin-resistant melanoma cells significantly delayed the growth of tumors compared to treatment with doxorubicin alone due to a significant increase in the intratumoral accumulation of doxorubicin [59]. Indeed, dipyridamole was shown to increase the concentration of several anticancer drugs (5-fluorouracil, methotrexate, piperidine, vincristine, cisplatin) in cancer cells [60,61]. In a xenograft mouse model of breast cancer cell-induced tumor, administration of dipyridamole reduced metastasis formation by decreasing the activation of the Wnt, ERK1/2-MAPK and NF-kB signalling pathways and the infiltration of macrophages and of myeloid-derived suppressor cells in primary tumors, suggesting that dipyridamole could be a promising agent for breast-cancer treatment [62].
4.1.1. Preclinical studies P2Y12-mediated platelet activation contributed to the release of proangiogenic factors, such as VEGF that help the growth of the tumor. P2Y12 antagonists, beyond their antiplatelet activity, may reduce angiogenic proteins release [78]. SR 25989, an enantiomer of clopidogrel lacking anti-aggregatory activity but provided with anti-angiogenic properties [79], inhibited pulmonary metastases in a model of subcutaneously-injected highly metastatic melanoma B16 cells in mice [80]. 4.1.2. Clinical studies A randomized phase II study in patients with metastatic breast cancer explored whether inhibition of platelet function would decrease circulating tumor cells thus potentially reducing metastasis. Treatment with clopidogrel or aspirin for 1 month did not reduce the proportion of patients with detectable circulating tumor cells (CTC), however baseline CTC number was lower than expected and study size rather small, making the study inconclusive [81]. A recent retrospective, observational study of clopidogrel in a cohort of 41,403 newly diagnosed patients with colorectal, breast and prostate cancer reported (after adjusting for relevant confounders) no significant differences in cancer-related mortality for either colorectal, breast or prostate cancer [82].
3.2.2. Clinical studies Dipyridamole was administered to thirty melanoma patients for a period of 11 years. Twenty six of them, with relatively advanced disease, had a five-year survival of 74% compared with an expected 32% [63]. Combination therapy using 5-fluorouracil (5-FU), cisplatin (CDDP) and dipyridamole (4 mg/kg/d) was explored in 28 patients with advanced gastric cancer and appeared to be safe and well tolerated and to potentiate the cytotoxicity of 5-FU [64]. Dipyridamole was used in combination with 5FU/LV and mitomycin for unresectable pancreatic cancer and a 39% response rate and 70% one-year survival rate in 38 patients were reported, resulting in a prolongation of survival compared to chemotherapeutic agents alone [65]. A phase II trial by the same team reported potential improvement in survival and resectability of localized unresectable pancreatic cancer with dipyridamole and
4.2. Ticagrelor Ticagrelor belongs to the cyclopentyl-triazolo-pyrimidine family, does not require metabolic activation to exert its action [83] and is a S108
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The first approach was investigated with the commonly used chemotherapeutic agent doxorubicin. Doxorubicin was loaded into platelets, by natural uptake through the open canalicular system, and these were then infused in a mouse lymphoma model showing an enhanced therapeutic efficacy and reduced adverse effects, compared to doxorubicin injected as such [95]. The second approach consists in coating nanoparticles containing anticancer drugs with membranes isolated from platelets. Nanoparticles as carriers are widely investigated for anticancer therapy, and some of them are currently used clinically (e.g. liposome containing doxorubicin). Indeed, the coating of nanoparticles using human platelets may potentially provide some benefits, such as the prolongation of drug circulation time, reduced immunogenicity, and selective targeting [96]. Coated nanoparticles are composed of a nanogel-based inner core loaded with anticancer drug, e.g. doxorubicin, and a platelet membrane–based outer shell. MDA-MB-231 (breast cancer) tumor-bearing nude mice were used as model, and the antitumor effect of the engineered nanoparticles loaded with doxorubicin was monitored by in vivo fluorescence imaging and histology. The i.v. infusion of plateletcoated nanoparticles loaded with doxorubicin effectively eliminated circulating tumor cells in vivo and inhibited the development of tumor metastases and the effect is significantly increased compared to nanoparticles loaded with doxorubicin but not coated with platelet membranes [97]. Recent evidence is accumulating that platelets may deliver nucleic acids to target cells in vivo. In a recent article, it was shown that platelets through their microparticles can infiltrate tumor mass and they can deliver their microRNAs content. The transfer of platelet miRNAs to tumor cells results in downregulation of tumor cell genes and inhibition of solid tumor growth, pointing out the role of miR-24 as a major PMPderived regulator of tumor growth in two cancer cell lines [98]. Platelet conjugated with an innovative therapeutic antibody, used in cancer immunotherapy, can interact with primary melanoma tumors partially removed by surgery and with circulating melanoma tumor cells improving survival. Platelets were conjugated with antibodies against programmed-cell-death protein 1 ligand (PDL-1). The antibodyconjugated platelets were injected via the tail vein in mice, got activated in the surgical bed due to the resection of tumor and shed platelet-derived microparticles that bound the cancer cells remained after surgical resection of tumor, thus completing tumor cell removal [99]. The antibody used was an immune checkpoint blocker, preventing PD1 expressed on T lymphocytes (T cells) from binding its PDL1 on cancer and on antigen-presenting cells (APCs), and thus from inactivating lymphocytes, in this way allowing cancer cells to evade attack. The mechanism of microparticle-mediated antitumor action was evaluated with in vitro (in a transwell system) and in vivo assays (using directly antibody-conjugated microparticles), showing that activated platelets in situ release factors that can recruit more immune cells to infiltrate the tumor microenvironment [99].
reversible oral P2Y12 receptor antagonist [84]. 4.2.1. Preclinical studies The potential of ticagrelor to inhibit cancer cell adhesion and metastasis was explored in intravenous and intrasplenic B16-F10 melanoma metastasis mouse models. Treatment with ticagrelor (10 mg/kg) markedly reduced lung and liver metastases and improved survival compared to saline-treated animals. A similar effect was observed in mice receiving intravenous 4T1 breast cancer cells, with significant reductions in lung and bone marrow metastases by intraperitoneal ticagrelor treatment, by inhibiting GPIIbIIIa activity [85]. 4.2.2. Clinical studies There are no clinical studies directly assessing the effects of treatment with ticagrelor on cancer and metastasis. In the ticagrelor arm of the PLATO trial, a study comparing the combination of ticagrelor and aspirin with aspirin and clopidogrel in patients with acute coronary syndromes (ACS), cancer incidence showed a trend towards a reduction with ticagrelor [86]. In contrast, there were significantly more cancer deaths in the ticagrelor arms of the PEGASUS trial [87] beyond one year therapy [88]. 4.3. Potential procancerogenic effects of P2Y12 antagonists Prasugrel (2-acetoxy-5-[α-cyclopropylcarbonyl-2-fluorobenzyl]4,5,6,7-tetrahydrothieno (3,2-c) pyridine) is a thienopyridine pro-drug metabolized in vivo to an active metabolite that irreversibly binds the platelet P2Y12 receptor. Recently, a potential association between prasugrel use and enhanced cancer incidence has been raised based on studies with prasugrel in rats and mice suggesting a weak dose-related increase of cancers of the intestine and lung [89,90]. A post hoc analysis of the TRITON-TIMI 38 trial, a study comparing the combination of prasugrel and aspirin with clopidogrel and aspirin in patients undergoing percutaneous coronary intervention (PCI), showed that the frequency of new, non-benign neoplasms and cancer deaths was higher among prasugrel-treated patients (1.6 vs. 1.2%, relative risk 1.29, 95%CI 0.96 to 1.75), with colon and lung cancer contributing to most of the difference [91]. Consequently, a comprehensive neoplasm ascertainment process was implemented for the TRILOGY ACS trial, a subsequent trial comparing prasugrel with clopidogrel, both in association with aspirin, in medically-treated ACS patients, but a similar risk of developing cancer in the two treatment arms was shown, albeit the statistical power was limited given the low event rates [92]. A recent systematic review and meta-analysis involving a total of nine studies, six randomized controlled trials, including the TRITONTIMI 38, and three retrospective cohort studies, for a total number of 282,084 participants, with follow-up ranging from a minimum of 6 to a maximum of 33 months, revealed that cancer event rate did not differ between prasugrel- and clopidogrel-exposed patients, and between thienopyridine-treated and aspirin or placebo-treated patients, thus not supporting concerns for a class effect of thienopyridines in increasing cancer [93]. Moreover, a very large population-based, historical cohort Israelian study, including 183,912 subjects, showed that aspirin users had a lower risk of cancer as compared with antiplatelet non-users (HR 0.46, 95%CI 0.44–0.49), and that aspirin plus clopidogrel users had a lower cancer risk than aspirin users (HR 0.92, 0.86–0.97), data confirming a protective effect of antiplatelet therapy on cancer incidence [94].
6. Conclusions Platelets definitely play a pathogenic role in cancer development and metastasis, by participating in primary tumor growth and in all the steps of the metastatic process. Therefore, pharmacological approaches specifically targeting platelet interactions with tumor cells represent an attractive, adjuvant anticancer therapy. Despite the great advances in the knowledge on the role of platelets in cancer, however, the identification of specific pharmacologic targets has lagged behind so far. New developments in biotechnological methods implying platelet use, may represent the way out to the current impasse, leading to targeted approaches to cancer cell-specific marks, and/or to the development of new drugs not interfering with the hemostatic platelet function but with the pro-cancer activities of platelets. These developments may provide an exciting advancement in the
5. Innovative therapeutic landscapes: platelets as drug carriers Two approaches have been adopted in using platelets as drug carriers in cancer therapy: encapsulating an anticancer drug within intact platelets or covering nanoparticle-containing anticancer drugs with platelet membranes. S109
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treatment of cancer patients and especially in the prevention of metastasis.
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Statement of conflict of interest [32]
The authors declare that there are no conflicts of interest associated with this manuscript.
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Full Length Article
Anti-platelet treatments in cancer: Basic and clinical research ⁎
T
Paolo Gresele , Marco Malvestiti, Stefania Momi Center of Thrombosis and Haemostasis, Department of Medicine, University of Perugia, Perugia, Italy
A R T I C L E I N F O
A B S T R A C T
Keywords: Cyclooxygenase Metastasis P2Y12 Phosphodiesterases Platelets Platelet-therapy
Over the past few decades the central role that platelets play in cancer development and progression, and especially in metastasis, has been elucidated. The molecular mechanisms responsible for initiating and mediating tumor cell-induced platelet aggregation and secretion have been largely unravelled. Considerable mechanistic insight into how platelets contribute to tumor angiogenesis, immunoevasion and cancer cell invasion have been clarified and, consequently, platelets have been identified as potential new drug targets for cancer therapy. This article gives an overview of the platelet-targeted pharmacologic approaches that have been attempted in the prevention of cancer development, progression and metastasis, including the application of antiplatelet drugs currently used for cardiovascular disease and of new and novel strategies.
1. Introduction The involvement of platelets in cancer growth and metastasis is a longstanding concept. An inverse correlation between platelet count and disease-specific survival has been described for several cancers and numerous basic and clinical research observations show that platelets affect disease burden and treatment efficacy in cancer patients, and participate in cancer metastasis. Platelets physically and functionally interact with various tumor cells through surface receptors including integrins. β1 integrins and β3 integrins in particular participate in platelet–tumor cell interaction and in tumor metastasis [1]. Platelets display their pro-metastastic role through the secretion of pro-metastatic factors (e.g. autotaxin), chemoattraction of granulocytes to platelet/tumor aggregates, activation of epithelial-mesenchimal-like transition (EMT) via direct contact with tumor cells and/or secretion of TGFβ1, release of microvesicles containing microRNA-223 which targets the tumor suppressor EPB41L3, adhesion to tumor cells providing physical protection from direct contact with NK cells [2]. The interplay between platelets and tumor cells involves the formation of platelet–tumor cell aggregates in the circulation, with platelets forming a shield for tumor cells allowing the latter to escape natural killer (NK) cell and tumor necrosis factor (TNF)α cytotoxic activity. Platelets may also help tumor cell extravasation to the metastatic niche. On the other hand platelet activation releases plateletderived growth and proangiogenic factors that may, contribute to tumor growth and angiogenesis. Platelets flowing through the tumor vasculature get activated,
adhere to the neovascular endothelium, aggregate and become part of the tumor's microenvironment, thus potentially influencing the tumor parenchyma and stroma of most solid tumors, particularly carcinomas. Platelets contribute to tumor persistence in the circulation by shielding tumor cells from destruction by NK cell [3–5] and suppress IFN-γ secretion by NK through the interaction between glucocorticoid-induced TNF-related ligand (GITR), a member of the TNF receptor superfamily that acts as a NK-inhibitory ligand on the surface of NK and its ligand (GITRL) present on platelet's surface [10–14]. Platelets contribute to metastasis and distant tumor growth by aiding tumor cell attachment to the endothelium and by releasing angiogenic and growth factors, such as vascular endothelial growth factor (VEGF) [6,7] and transforming growth factor-β (TGF-β) [8,9]. Tumor cell invasion through the extracellular matrix (ECM) is a crucial step in tumor metastasis, and this process is mediated by proteolytic enzymes, such as matrix metalloproteinases (MMPs), that degrade the ECM surrounding blood vessels to allow cancer cells to penetrate in tissue. Indeed, the ECM surrounding blood vessels plays a critical role in the limitation of extravasation and intravasation of tumor cells. Platelet-derived TGF-β1 is crucial in inducing tumor growth and metastasis by up-regulating MMP-2 and MMP-9 [15] and by activating the TGFβ/SMAD and NF-kB pathways in cancer cells [16]. The importance of platelets in cancer growth is supported by the finding that platelet depletion results in a marked reduction of tumor growth in an orthotopic model of ovarian cancer, in vivo models of experimental pulmonary metastasis and in a murine model of spontaneous metastasis [17]. The interaction between cancer cells and/or the neovessels
⁎ Corresponding author at: Section of Internal and Cardiovascular Medicine, Department of Medicine, University of Perugia, Nuovo Polo Didattico, Strada Vicinale Via delle Corse, 06132 Perugia, Italy. E-mail address: [email protected] (P. Gresele).
https://doi.org/10.1016/j.thromres.2017.12.016 Received 24 November 2017; Accepted 27 December 2017 0049-3848/ © 2018 Elsevier Ltd. All rights reserved.
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prevention of CRC [35–38], cumulatively showed a significantly reduced risk of developing an adenoma on follow-up (0.83, 95%CI = 0.72 to 0.96, p = 0.012) [39]. In carriers of hereditary predisposition to non-polyposis colon cancer aspirin was shown to prevent the development of cancer [40] and to prolong survival when started after diagnosis [41]. Given that in most of these trials high doses of aspirin were used, the doubt that the effect of aspirin would be essentially due to an action on tumor cell COX-2, requiring higher doses, and that therefore long term platelet-selective low-dose, aspirin would not prevent CRC, was risen. Therefore, a follow-up cumulative observation study of four large randomized trials on aspirin (75–300 mg/daily) versus control in primary and secondary prevention of vascular events, involving over 14,000 patients followed-up for a median of 18.3 years, was performed showing that allocation to aspirin significantly reduced the 20-year risk of colon cancer (HR 0.76, 95%CI 0.6- to 0.96, p = 0.02) and cancer-related mortality (HR 0.65, 95%CI 0.48 to 0.88, p = 0.005), but not that of rectal cancer, and that the benefit increased with treatment duration such that at least 5 years of treatment were required to reduce colon cancer risk. Moreover, benefit was evident with aspirin doses as low as 75 mg/day, with no further increase of benefit with greater doses [42]. This is an important observation because it is known that the risk of major bleeding with aspirin is dosedependent [43,44]. Other subsequent meta-analyses of individual patient data from large aspirin studies of aspirin in cardiovascular prevention showed that aspirin reduced mortality also from non-gastrointestinal cancers [45], prevented metastasis from all cancers [46] and that the effect was only evident from 5 years of continued treatment onward [47]. These conclusions have been criticized because of the retrospective nature of the analysis on non-pre-specified end-points, because most of the recorded cancer events occurred well after the end of the randomized follow-up phase of the studies, and because some important trials, like the Women's Health Study, had been excluded from the analysis. Therefore, several ad hoc designed adjuvant trials of low-dose aspirin have been initiated in the last few years, in patients with various types of newly diagnosed cancer, and the protocols of some large, ongoing, primary cardiovascular prevention trials with aspirin (ASCEND, ACCEPT-D, ARRIVE and ASPREE) have been modified to collect prospective information about cancer incidence. The results of these ongoing studies will finally provide prospective evidence on the role of chronic aspirin in chemoprevention.
irrorating tumors and platelets leads to the triggering of adhesion and activation of the latter, a process also called tumor-cell induced platelet aggregation (TCIPA). TCIPA is mediated by multiple agents: tissue factor (TF), thrombin, adenosine diphosphate (ADP), TxA2, and MMP [18]. Therefore the use of antiplatelet agents as adjuvant cancer therapy or the exploitation of platelets as carriers of antitumor agents are attractive approaches. 2. Cyclooxygenase inhibitors Cyclooxygenase (COX) is the enzyme transforming arachidonic acid into the precursor (PGG2-PGH2) of different prostaglandins (PGs) or of thromboxanes (Tx), depending on the cell type. Arachidonic acid is cleaved from membrane phospholipids by phospholipase A2 and is then transformed by cyclooxygenases (COXs) into PGH2, which is rapidly converted into prostanoids by different PGor Tx-synthases [19]. The COX-1 isoenzyme preferentially couples with TXA2 synthase (in platelets) and PGF2 synthase (PGFS), whereas COX-2 is associated mainly with PGI2 synthase (PGI-S) (in endothelial cells) and PGE synthase (PGEs) in several cell types, including mucosal cells of the gastrointestinal tract [20]. Evidence exists for an antitumor activity of aspirin, which acts as an active-site acetylating agent inducing an irreversible inactivation of COX-1 (through acetylation of Ser 529) and COX-2 (through acetylation of Ser 516), with the consequent suppression of prostaglandin and thromboxane production [21,22]. The inhibition of platelet TxA2 production by aspirin is the basis of the most widely diffused antiplatelet therapy in clinical use. 2.1. Preclinical studies Early observations showed that tumor metastases were reduced in rats treated with aspirin [23] and that prostaglandin concentration was raised in rat colorectal tumor tissue [24,25], opening the way to the studies on a potential benefit of aspirin in cancer. Supporting observations showed that oral administration of aspirin significantly inhibited the incidence and number of invasive, azoxymethane-induced adenocarcinomas of the colon in rats [26] as well as the onset of lung tumors induced by a tobacco-specific nitrosamine in mice [27]. Aspirin, administered to mice at a dose equivalent to low-dose in humans (around 100 mg/day), induced apoptosis of colorectal cancer (CRC) cells by a mechanism involving the downregulation of the IL-6-STAT3 signalling pathway [28]. The antitumor activity of aspirin is considered to be driven by the inhibition of COX-2, which is overexpressed in cancer cells [29,30], but also by the inhibition of platelet adhesion to tumor cells, and of TCIPA, essential steps in the protective effects by platelets from immune surveillance against cancer cells [31]. CRC cells (HT29) previously exposed to platelets in vitro transform into mesenchymal-like cancer cells which, injected into the tail vein of immunodeficient NOD-SCID mice, produce a higher number of lung metastases compared to HT29 cells not exposed to platelets, an effect associated with enhanced systemic biosynthesis of TXA2 and PGE2 in vivo. Aspirin administration prevented the increased rate of metastasis as well as the enhanced production of TXA2 and PGE2 induced by platelet-primed HT29 cells [32].
3. PDE inhibitors Phosphodiesterases (PDEs) catalyze the hydrolysis of cyclic (c) AMP and cGMP, two powerful intracellular inhibitory second messengers, to inactive 5′AMP and 5′GMP. Of the over 60 different PDEs known, platelets express PDE2, PDE3, and PDE5 and their inhibition rises the intraplatelet levels of the two cyclic nucleotides thus inhibiting platelet activation [48]. Among the isoenzyme-selective PDE inhibitors developed as antiplatelet agents, cilostazol and dipyridamole have been explored as possible adjuvant therapy in cancer. 3.1. Cilostazol Cilostazol (6-(4-(1-cyclohexyl-1H-tetrazol-5-yl)butoxy)-3,4-dihydro-2(1H)-quinolinone) is a specific and strong inhibitor of PDE3 in platelets and smooth muscle cells, it is in clinical use since 1999 for the treatment of intermittent claudication [49] and has shown protective effects on ischemic cardiovascular events [48]. Its effect on cancer has been only evaluated in preclinical studies.
2.2. Clinical studies From a case-control study, chronic aspirin use among CRC-patients was significantly lower than among age/sex-matched non-CRC-controls [33]. Later, a meta-analysis cumulating data from several case-control studies in CRC patients, involving > 30,000 subjects, concluded that regular aspirin use was associated with a reduced risk of CRC (pooled odds ratio (OR) 0.62, 95%CI 0.58–0.67, p < 0.0001) [34]. A few small randomized, double-blind, placebo-controlled trials, involving around 3000 participants, evaluating chronic aspirin for the secondary
3.1.1. Preclinical studies Cilostazol suppressed the migration of human colon cancer cells, DLD-1, induced by soluble fibronectin or fetal bovine serum in a S107
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recommended further studies [66]. Despite these apparently positive results, it must be considered that they came from small and mostly uncontrolled studies, and that a number of other small trials examining the potential usefulness of dipyridamole to enhance chemotherapeutic efficacy in sarcoma, colorectal, breast, renal, and prostate cancers failed to show meaningful improvement in response [67].
phagokinetic assay and it suppressed cancer cell invasion induced by fetal bovine serum in a trans-cellular migration assay, showing that cilostazol inhibits colon cancer cell motility [50]. The adjuvant antitumor effect of cilostazol in association with cisplatin was assessed in a xenograft tumor model in nude mice injected with cells derived from a human oral carcinoma or with heLa cells or with the respective cisplatin-resistant cell lines. A synergic antitumor effect of the combination of cilostazol and cisplatin was observed in cisplatin-resistant cell lines, with a significant increase in the number of apoptotic cancer cells [51]. In vitro, the addition of B16 tumor cells to washed platelets induced aggregation, an effect inhibited by cilostazol [52]. Moreover, cilostazol administered 2 h before the injection of human adenocarcinoma cells in nude mice strongly inhibited lung metastasis, while it was ineffective when administered 72 h after tumor cell injection, suggesting that platelet aggregation induced by tumor cells plays an important role in metastasis development [53].
4. ADP receptor antagonists Purinergic receptors (P2 receptors) are present on the platelet surface and bind ADP triggering platelet aggregation [68]. P2Y12, the Gαicoupled ADP receptor on platelets [69], is the target of all currently used antiplatelet ADP antagonists [70]. On the other hand, P2Y12 has also been shown to be present in some tumor cells (intestinal epithelial carcinoma cells, glioma C6 cells, ovarian cancer cells, etc.) in which it regulates cell growth [71]. In a model of lung metastasis induced by Lewis Lung Carcinoma cells (LLC), P2Y12-deficient mice showed a lower tumor burden, confirming a role of platelet P2Y12 in tumor metastasis [72]. P2Y12 deficiency diminished the ability of LLC cells to induce platelet release of active TGFβ1, resulting in a reduced platelet-induced EMT-like transformation of LLC cells, a prerequisite for LLC cell metastasis. In accordance, platelet P2Y12 deficiency resulted in significantly less lung metastasis in a B16 melanoma experimental murine metastasis model [72].
3.2. Dipyridamole Dipyridamole (2,6–bis (diethanolamino)–4,8-dipiperidino-pyrimido 5,4-d pyrimidine), synthesized more than fifty years ago and also used as a coronary vasodilator, is currently in clinical use as an antithrombotic drug for secondary prevention of stroke in association with aspirin. Dipyridamole inhibits platelet function not only by acting as an inhibitor of PDE5 and PDE3 but also by inhibiting the reuptake of adenosine by red blood cells. In this way dipyridamole enhances the plasma levels of this platelet-inhibitor and vasodilatory nucleoside, and contribute to the scavenging of free radicals that inactivate vessel wall COX, thus enhancing PGI2 biosynthesis [48, 54–57].
4.1. Clopidogrel Clopidogrel, a thienopyridine, is an oral P2Y12 inhibitor acting as a pro-drug that requires metabolic conversion to its active form by the liver. Indeed, about 85% of the absorbed clopidogrel is metabolized into an inactive metabolite (SR26334) by plasmatic carboxylases and the remaining 15% is metabolized, by a two-step biotransformation process dependent on cytochrome P450 isoenzymes in the liver [73,74], in R130964, the active circulating metabolite that irreversibly blocks P2Y12 [75–77].
3.2.1. Preclinical studies Dipyridamole inhibited TCIPA and reduced liver metastases in athymic nude mice injected with a human pancreatic adenocarcinoma cell line (RWP-2) [58]. Moreover, dipyridamole has shown potential benefit against cancer multidrug resistance, a phenomenon that decreases the clinical benefit of several anticancer agents, including doxorubicin. Treatment with dipyridamole of mice injected with doxorubicin-resistant melanoma cells significantly delayed the growth of tumors compared to treatment with doxorubicin alone due to a significant increase in the intratumoral accumulation of doxorubicin [59]. Indeed, dipyridamole was shown to increase the concentration of several anticancer drugs (5-fluorouracil, methotrexate, piperidine, vincristine, cisplatin) in cancer cells [60,61]. In a xenograft mouse model of breast cancer cell-induced tumor, administration of dipyridamole reduced metastasis formation by decreasing the activation of the Wnt, ERK1/2-MAPK and NF-kB signalling pathways and the infiltration of macrophages and of myeloid-derived suppressor cells in primary tumors, suggesting that dipyridamole could be a promising agent for breast-cancer treatment [62].
4.1.1. Preclinical studies P2Y12-mediated platelet activation contributed to the release of proangiogenic factors, such as VEGF that help the growth of the tumor. P2Y12 antagonists, beyond their antiplatelet activity, may reduce angiogenic proteins release [78]. SR 25989, an enantiomer of clopidogrel lacking anti-aggregatory activity but provided with anti-angiogenic properties [79], inhibited pulmonary metastases in a model of subcutaneously-injected highly metastatic melanoma B16 cells in mice [80]. 4.1.2. Clinical studies A randomized phase II study in patients with metastatic breast cancer explored whether inhibition of platelet function would decrease circulating tumor cells thus potentially reducing metastasis. Treatment with clopidogrel or aspirin for 1 month did not reduce the proportion of patients with detectable circulating tumor cells (CTC), however baseline CTC number was lower than expected and study size rather small, making the study inconclusive [81]. A recent retrospective, observational study of clopidogrel in a cohort of 41,403 newly diagnosed patients with colorectal, breast and prostate cancer reported (after adjusting for relevant confounders) no significant differences in cancer-related mortality for either colorectal, breast or prostate cancer [82].
3.2.2. Clinical studies Dipyridamole was administered to thirty melanoma patients for a period of 11 years. Twenty six of them, with relatively advanced disease, had a five-year survival of 74% compared with an expected 32% [63]. Combination therapy using 5-fluorouracil (5-FU), cisplatin (CDDP) and dipyridamole (4 mg/kg/d) was explored in 28 patients with advanced gastric cancer and appeared to be safe and well tolerated and to potentiate the cytotoxicity of 5-FU [64]. Dipyridamole was used in combination with 5FU/LV and mitomycin for unresectable pancreatic cancer and a 39% response rate and 70% one-year survival rate in 38 patients were reported, resulting in a prolongation of survival compared to chemotherapeutic agents alone [65]. A phase II trial by the same team reported potential improvement in survival and resectability of localized unresectable pancreatic cancer with dipyridamole and
4.2. Ticagrelor Ticagrelor belongs to the cyclopentyl-triazolo-pyrimidine family, does not require metabolic activation to exert its action [83] and is a S108
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The first approach was investigated with the commonly used chemotherapeutic agent doxorubicin. Doxorubicin was loaded into platelets, by natural uptake through the open canalicular system, and these were then infused in a mouse lymphoma model showing an enhanced therapeutic efficacy and reduced adverse effects, compared to doxorubicin injected as such [95]. The second approach consists in coating nanoparticles containing anticancer drugs with membranes isolated from platelets. Nanoparticles as carriers are widely investigated for anticancer therapy, and some of them are currently used clinically (e.g. liposome containing doxorubicin). Indeed, the coating of nanoparticles using human platelets may potentially provide some benefits, such as the prolongation of drug circulation time, reduced immunogenicity, and selective targeting [96]. Coated nanoparticles are composed of a nanogel-based inner core loaded with anticancer drug, e.g. doxorubicin, and a platelet membrane–based outer shell. MDA-MB-231 (breast cancer) tumor-bearing nude mice were used as model, and the antitumor effect of the engineered nanoparticles loaded with doxorubicin was monitored by in vivo fluorescence imaging and histology. The i.v. infusion of plateletcoated nanoparticles loaded with doxorubicin effectively eliminated circulating tumor cells in vivo and inhibited the development of tumor metastases and the effect is significantly increased compared to nanoparticles loaded with doxorubicin but not coated with platelet membranes [97]. Recent evidence is accumulating that platelets may deliver nucleic acids to target cells in vivo. In a recent article, it was shown that platelets through their microparticles can infiltrate tumor mass and they can deliver their microRNAs content. The transfer of platelet miRNAs to tumor cells results in downregulation of tumor cell genes and inhibition of solid tumor growth, pointing out the role of miR-24 as a major PMPderived regulator of tumor growth in two cancer cell lines [98]. Platelet conjugated with an innovative therapeutic antibody, used in cancer immunotherapy, can interact with primary melanoma tumors partially removed by surgery and with circulating melanoma tumor cells improving survival. Platelets were conjugated with antibodies against programmed-cell-death protein 1 ligand (PDL-1). The antibodyconjugated platelets were injected via the tail vein in mice, got activated in the surgical bed due to the resection of tumor and shed platelet-derived microparticles that bound the cancer cells remained after surgical resection of tumor, thus completing tumor cell removal [99]. The antibody used was an immune checkpoint blocker, preventing PD1 expressed on T lymphocytes (T cells) from binding its PDL1 on cancer and on antigen-presenting cells (APCs), and thus from inactivating lymphocytes, in this way allowing cancer cells to evade attack. The mechanism of microparticle-mediated antitumor action was evaluated with in vitro (in a transwell system) and in vivo assays (using directly antibody-conjugated microparticles), showing that activated platelets in situ release factors that can recruit more immune cells to infiltrate the tumor microenvironment [99].
reversible oral P2Y12 receptor antagonist [84]. 4.2.1. Preclinical studies The potential of ticagrelor to inhibit cancer cell adhesion and metastasis was explored in intravenous and intrasplenic B16-F10 melanoma metastasis mouse models. Treatment with ticagrelor (10 mg/kg) markedly reduced lung and liver metastases and improved survival compared to saline-treated animals. A similar effect was observed in mice receiving intravenous 4T1 breast cancer cells, with significant reductions in lung and bone marrow metastases by intraperitoneal ticagrelor treatment, by inhibiting GPIIbIIIa activity [85]. 4.2.2. Clinical studies There are no clinical studies directly assessing the effects of treatment with ticagrelor on cancer and metastasis. In the ticagrelor arm of the PLATO trial, a study comparing the combination of ticagrelor and aspirin with aspirin and clopidogrel in patients with acute coronary syndromes (ACS), cancer incidence showed a trend towards a reduction with ticagrelor [86]. In contrast, there were significantly more cancer deaths in the ticagrelor arms of the PEGASUS trial [87] beyond one year therapy [88]. 4.3. Potential procancerogenic effects of P2Y12 antagonists Prasugrel (2-acetoxy-5-[α-cyclopropylcarbonyl-2-fluorobenzyl]4,5,6,7-tetrahydrothieno (3,2-c) pyridine) is a thienopyridine pro-drug metabolized in vivo to an active metabolite that irreversibly binds the platelet P2Y12 receptor. Recently, a potential association between prasugrel use and enhanced cancer incidence has been raised based on studies with prasugrel in rats and mice suggesting a weak dose-related increase of cancers of the intestine and lung [89,90]. A post hoc analysis of the TRITON-TIMI 38 trial, a study comparing the combination of prasugrel and aspirin with clopidogrel and aspirin in patients undergoing percutaneous coronary intervention (PCI), showed that the frequency of new, non-benign neoplasms and cancer deaths was higher among prasugrel-treated patients (1.6 vs. 1.2%, relative risk 1.29, 95%CI 0.96 to 1.75), with colon and lung cancer contributing to most of the difference [91]. Consequently, a comprehensive neoplasm ascertainment process was implemented for the TRILOGY ACS trial, a subsequent trial comparing prasugrel with clopidogrel, both in association with aspirin, in medically-treated ACS patients, but a similar risk of developing cancer in the two treatment arms was shown, albeit the statistical power was limited given the low event rates [92]. A recent systematic review and meta-analysis involving a total of nine studies, six randomized controlled trials, including the TRITONTIMI 38, and three retrospective cohort studies, for a total number of 282,084 participants, with follow-up ranging from a minimum of 6 to a maximum of 33 months, revealed that cancer event rate did not differ between prasugrel- and clopidogrel-exposed patients, and between thienopyridine-treated and aspirin or placebo-treated patients, thus not supporting concerns for a class effect of thienopyridines in increasing cancer [93]. Moreover, a very large population-based, historical cohort Israelian study, including 183,912 subjects, showed that aspirin users had a lower risk of cancer as compared with antiplatelet non-users (HR 0.46, 95%CI 0.44–0.49), and that aspirin plus clopidogrel users had a lower cancer risk than aspirin users (HR 0.92, 0.86–0.97), data confirming a protective effect of antiplatelet therapy on cancer incidence [94].
6. Conclusions Platelets definitely play a pathogenic role in cancer development and metastasis, by participating in primary tumor growth and in all the steps of the metastatic process. Therefore, pharmacological approaches specifically targeting platelet interactions with tumor cells represent an attractive, adjuvant anticancer therapy. Despite the great advances in the knowledge on the role of platelets in cancer, however, the identification of specific pharmacologic targets has lagged behind so far. New developments in biotechnological methods implying platelet use, may represent the way out to the current impasse, leading to targeted approaches to cancer cell-specific marks, and/or to the development of new drugs not interfering with the hemostatic platelet function but with the pro-cancer activities of platelets. These developments may provide an exciting advancement in the
5. Innovative therapeutic landscapes: platelets as drug carriers Two approaches have been adopted in using platelets as drug carriers in cancer therapy: encapsulating an anticancer drug within intact platelets or covering nanoparticle-containing anticancer drugs with platelet membranes. S109
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treatment of cancer patients and especially in the prevention of metastasis.
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Statement of conflict of interest [32]
The authors declare that there are no conflicts of interest associated with this manuscript.
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