This proposal centers on the mechanisms of cancer-associated thrombosis, also known as Trousseau's syndrome. In many cancers, platelet-mediated thrombosis is the leading cause of morbidity. Platelets are present in blood at levels higher than any other cell and their activation and aggregation leads to thrombosis. Due to the presence of numerous storage granules, platelets are able to uptake a number of important mediators. Several lines of experimental evidence from our and other groups suggest that the presence of tumor changes platelet transcriptome, secretome (storage granules content) as well as platelet activation status resulting in so-called ?tumor-educated platelets?. We have shown that platelets from tumor-bearing mice and platelets from cancer patients carry not only proteins or tumor origin but also tumor-specific RNAs. To understand how tumor RNAs find their way to platelets, we focus on vesicles called exosomes, which are produced by aggressive tumors and contain tumor-specific RNAs and proteins. We demonstrated that tumor exosomes are efficiently taken in by platelets in vitro and in vivo, resulting in changes of platelet transcriptome and platelet activation. Capitalizing on these preliminary results we hypothesized that tumor exosomes, containing tumor signature, are efficiently taken in by platelets via CD63 and platelet secretory granules machinery (VAMPs). This results in changes in platelet transcriptome and, eventually, in platelet hyperactivation leading to thrombosis.
AIM1. To determine the mechanisms of exosome uptake by platelets. Using exo from prostate cancer cells and from patients, we will assess the role of CD63 and its glycosylation in exo uptake by platelets in vitro and in vivo. The role of platelet endocytosis/exocytosis machinery in exosomes uptake will be tested using VAMP8, VAMP3, Arf6 KO platelets.
AIM2. To define the mechanisms of platelet activation by exo in vivo and identify potential targets for intervention. Platelet activation by tumors and tumor-derived exo will be monitored in the presence or absence of CD63 blocking reagents in vivo.
AIM3. To define tumor-specific signature in both, exo and circulating platelets isolated from blood of cancer patients. We will focus on prostate cancer patients and will measure selected tumor-specific markers in exosomes and platelets by qPCR. Platelets from patients before and 3 and 6 months after prostatectomy will be assayed for tumor-specific reporters and activation status. These studies are designed to define the specific mechanisms and consequences of platelets activation by tumor exosomes and develop new therapeutic strategies to interfere with cancer-associated thrombosis.
It remains rather unknown that the major and, in many cancers, the leading cause of morbidity and mortality in cancer patients is cancer-associated thrombosis (also known as Trousseau's syndrome). While our anti-cancer therapeutic strategies have improved, resulting in a longer life span of cancer patients, the problem of thromboembolism becomes even more important. While numerous clinical studies emphasize an importance of this problem, very little is known about its specific molecular mechanisms. One of the main reasons is that such studies require a rare combination of expertise in tumor biology and relevant experimental models, as well as in biology of thrombosis, first of all in platelet biology. In this project, we will capitalize on our extensive preliminary data and many years of expertise in cancer and platelet research. Several lines of experimental evidence suggest that the presence of tumor alters platelets and resulting ?tumor-educated platelets? carry proteins of tumor origin and tumor-specific RNAs. To understand how tumor RNAs find their way to platelets, we will focus on vesicles called exosomes, which are produced by aggressive tumors and contain tumor- specific RNAs and proteins. We demonstrated that tumor exosomes are efficiently taken in by platelets in vitro and in vivo, resulting in changes of platelet transcriptome. This results in platelet hyperactivation leading to thrombosis. Our studies are designed to define the specific mechanisms and consequences of platelets activation by tumor exosomes. Most importantly, we will focus on the identification and validation of new therapeutic strategies to prevent cancer-induced thrombosis.
