Systemic coagulation activation in patients with glioblastoma

Systemic coagulation activation in patients with glioblastoma | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Systemic coagulation activation in patients with glioblastoma Justyna Joanna Rybus, Sandra Agnieszka Pilawska, Magdalena Dębicka, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6219685/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Purpose To assess if patients diagnosed with glioblastoma have higher markers of systemic coagulation activation in thrombin generation assay (TGA) and blood tests. Methods We collected preoperative blood samples from 42 glioblastoma patients and 35 healthy controls. We assessed fibrinogen concentration, fibrinogen degradation products (FDP) and D-dimer concentrations, plasminogen activator inhibitor concentration (PAI:Ag), antithrombin (AT) activity, thrombin-antithrombin complexes (TAT), the thrombin peak (P), endogenous thrombin potential (ETP), Lag Time and Time To Peak (TTP). Results Patients with glioblastoma showed higher values of following parameters preoperatively such as D-dimer concentration (1.8 ± 3.6 vs 0.35 ± 0.32 mg/l, p = 0.02), PAI:Ag (27.30 ± 8.90 vs 17.00 ± 9.34, p < 0.01) and TAT (15.20 ± 17.00 vs 4.56 ± 1.66, p < 0.01). Furthermore they showed longer Lag Time (3.50 ± 1.00 vs 3.02 ± 0.69, p = 0.01) and TTP (6.60 ± 1.50 vs 5.73 ± 1.04, p = 0.01) in TGA. Conclusion High values of d-dimer, PAI:AG and TAT concentrations are prothrombotic parameters but longer Lag Time and TTP did not indicate increased systemic coagulation activity. The above information sheds new light on coagulation disorders in glioblastoma patients. This may also indicate that TGA assessed by calibrated automated thrombogram (CAT) has limited ability to detect subtle abnormalities in the coagulation system. In both situations further studies are required. Thrombin Generation Assay Glioblastoma Systemic Coagulation Coagulation Disorders Figures Figure 1 Figure 2 Introduction The risk of venous thromboembolism (VTE) increases four- to sevenfold in patients with cancer in comparison to the general population [ 1 ]. Patients with brain tumors often suffer from motor deficits that can render them bedridden [ 2 ]. Brain tumors predispose to VTE more than cancer in other sites [ 3 ]. This fact is associated with many additive risk factors such as cancer itself, the thrombotic response to surgical trauma and limited postoperative mobility. Glioblastoma (GBM) causes chronic activation of the coagulation system. This phenomenon occurs because tumor cells contain transcripts for several coagulation factors and hemostasis regulating molecules. Furthermore, this molecular combination seems to differ between brain tumor subtypes [ 4 ]. This raises the question of whether the patient will benefit from pharmacological thromboprophylaxis or if it could be harmful. Results from conducted prospective studies are unclear, but there are trends indicating serious hemorrhagic complications after long-term administration of low-molecular-weight heparin (LMWH) compared to the placebo group [ 5 ]. Considering the high risk of VTE and complications associated with long-term pharmacological thromboprophylaxis, appropriate selection of patients for treatment should be made. Basic coagulation tests, such as prothrombin time (PT) and activated partial thromboplastin time (APTT), do not accurately represent the in vivo balance of coagulation. However, in patients with congenital deficiencies of natural anticoagulants (e.g., protein C, protein S, antithrombin), these parameters are often within reference ranges. This diagnostic gap in assessing the patient's hemostasis status is helped by the calibrated automated thrombogram (CAT), a global dynamic test that continuously and simultaneously examines both thrombin formation and its inhibition. Therefore, CAT is an invaluable tool for diagnosing, monitoring or implementing treatment for many hemorrhagic diseases. It is also highly relevant in the field of thrombosis for assessing the risk of VTE and determining a favorable time to administer antithrombotic drugs [ 6 ]. The aim of the study was to examine new biomarkers that could help profile patients, potentially allowing for the estimation of both pre- and postoperative VTE risk in patients with GBM. Materials and Methods Study population Forty-two patients with newly diagnosed glioblastoma who underwent surgical resection at the Department of Neurosurgery and Neurotraumatology of University Hospital in Kraków were enrolled in the study. The inclusion criteria were: age over 18 years, newly diagnosed brain tumor with confirmed histopathology: Glioblastoma, IDH-wildtype. Exclusion criteria were: continuous anticoagulant treatment with vitamin K antagonists, low-molecular-weight heparins or antiplatelet drugs, bacterial or viral infection, pregnancy. The control group, matched for age and gender, consisted of 35 healthy individuals (18 males and 17 females, median age of 50.29 ± 11.50 years) without a history of cancer, with no personal or family history of thrombosis or medications that could affect the clotting system. The control group was selected from among the patients of the hematology clinic where haematological diseases were excluded. Data evaluation The medical history of patients from the study group was evaluated in terms of sex, age, weight, height, BMI (body mass index), comorbidities, drug intake and complications during hospitalization. Thromboprophylaxis was assessed both in the preoperative and postoperative periods, including the timing of its administration. Other factors considered included preoperative steroid use, mean operative time, position during surgery and extent of resection. Laboratory tests, including complete blood count, sodium and potassium levels, glucose, creatinine and urea, C-reactive protein (CRP), albumin, fibrinogen, fibrinogen degradation products (FDP), antithrombin (AT) and D-dimers (DD) were established upon admission. Values of FDP were presented in 3 ranges − 0 when the FDP value was less than 5 mg/l, 1 when values were in the range of 5–15 mg/l and 2 when were above 20 mg/l. APTT and PT were assessed upon admission. Tumors were evaluated according to the World Health Organization (WHO) 2021 classification of Central Nervous System (CNS) tumors [ 7 ]. All patients were assessed using the Caprini Score for Venous Thromboembolism, the Karnofsky Scale and the ASA (American Society of Anaesthesiology) scale. Deep vein thrombosis was confirmed by compression ultrasound or computed tomography angiography (CT-angiography) in individual cases. Diagnosis was made by board certified vascular surgeon. The medical condition of patients in the study group was monitored for the period of 12 months following surgery. The study protocol was approved by the Bioethical Committee of the Jagiellonian University in Kraków, Poland (approval number 1072.6120.92.2019), and written informed consent was obtained from all participants. Blood Sampling and Laboratory Assays Samples containing 20 ml of blood were collected into tubes with EDTA and 3.2% sodium citrate by sterile and atraumatic venipuncture on the day of the scheduled surgery. The procedure was repeated 24 and 72 hours postoperatively. Plasma samples were frozen at -80°C until analyzed. All tests, including a thrombin generation assay - calibrated automated thrombogram (TGA-CAT), of both the control and study groups, were conducted in the same laboratory using identical equipment and by the same practitioner. A thrombin generation assay (TGA) was assessed by a calibrated automated thrombogram (CAT). The plasma samples were measured in the 96-well plate fluorometer (Ascent Reader, Thermolabsystems OY, Helsinki, Finland), equipped with the 390/460 filter set and a dispenser. In brief, 80 µl of platelet-poor plasma was combined with 20 µl of tissue factor (TF)-based activator (Diagnostica Stago, Asnières, France) and 20 µl of FluCa solution (Diagnostica Stago, Asnières, France). A dedicated software program (Thrombinoscope, Synapse BV, Maastricht, The Netherlands) provided the evaluation of plasma thrombogenic potential [ 8 , 9 ]. Four parameters of TGA-CAT were assessed: thrombin peak (maximum concentration of thrombin formed during the recording time) - nmol (P), endogenous thrombin potential (the area under the curve) - nmol/min (ETP), lag time - min (LT), time to peak - min (TTP). Statistics Elements of descriptive statistics were used. Continuous variables are expressed as mean ± standard deviation, and proportions are presented as percentage value. Statistical analysis was performed using STATISTICA v. 13.0 (Statsoft, Poland). Pearson's Chi-square test was used to assess proportions, and the t-test was used to assess continuous variables. Pearson's correlation coefficient was used to evaluate correlations between variables. Differences were considered significant at p < 0.05. Results We analyzed prospectively collected data from 42 patients with newly diagnosed glioblastoma confirmed in histopathology examination. Mean age was 65.19 ± 7.02 years. Mean BMI score was 28.44 ± 5.73 kg/m2 and mean ASA scale score 2.26 ± 0.59. In the Caprini Score the average patient got 6.46 ± 1.05 points. Two patients (4.76%) suffered from venous thromboembolism during hospitalization and 3 (7.14%) after discharge of which one experienced VTE in both periods. Patient’s comorbidities are shown in Fig. 1 . Mean Karnofsky’s score was 76.9 ± 14.23. Anticoagulation therapy was withheld in all patients for a minimum of 24 hours before surgery, while antiplatelet therapy was withheld for 7 days preoperatively. Four patients (9.52%) received a perioperative antithrombotic drug (low-dose heparin). Twenty-seven patients (64.29%) received dexamethasone preoperatively at an average dose of 8 mg. In twenty-two patients (52.38%) gross-total resection (GTR) was achieved, 8 patients (19.05%) underwent subtotal resection (STR) and in 12 patients (28.57%) biopsy of the lesion was performed. Sixteen patients (38.10%) got prophylactic dosage of LWMH postoperatively on average on the second day after surgery (2.25 ± 1.29 days) due to a prolonged period of immobilization. Seven patients (16.67%) died before discharge. Five of them suffered from postoperative complications in the form of hematoma in the operation site (3 cases), epidural hematoma and intracerebral hematoma. The study group had significantly higher concentrations of D-dimer (1.8 ± 3.6 vs 0.35 ± 0.32, p = 0.02), PAI:Ag (27.30 ± 8.90 vs 17.00 ± 9.34, p < 0.01) and TAT (15.20 ± 17.00 vs 4.56 ± 1.66, p < 0.01). In TGA-CAT, the following parameters were also longer in the study group than in the control group: Lag Time (3.5 ± 1.00 vs 3.02 ± 0.69, p = 0.01) and TTP (6.60 ± 1.50 vs 5.73 ± 1.04, p = 0.01). There were no statistically significant differences between the groups in ETP (1513.10 ± 316.60 vs 1590.48 ± 266.84, p = 0.28) and Peak (292.20 ± 83.00 vs 307.74 ± 55.84, p = 0.37). A comparison of results between study and control groups is presented in Fig. 2 . Discussion In this prospective cohort study, we observed elevated levels of D-dimer, thrombin-antithrombin complexes, and PAI:Ag, alongside prolonged lag time and time to peak. This inconsistency can be explained in several ways. In certain diseases, such as cancer, microangiopathy can alter platelet function. Reduced platelet numbers or dysfunction can shorten the thrombin activation process, leading to prolonged lag time and TTP. At the same time, this can promote clot formation in the microcirculation, as evidenced by elevated D-dimer levels. In the context of glioblastoma patients, prolonged lag time and time to peak may suggest that a hypercoagulable state is not detected by standard tests, such as CAT, or that these tests lack the sensitivity to identify subtle clotting abnormalities. This indicates that thrombotic mechanisms in these patients may be more complex or less predictable. This is one of the very few studies demonstrating systemic coagulation processes in glioblastoma compared to healthy subjects. Most studies use patients with meningiomas as controls, which may show some coagulation abnormalities but not as pronounced as those seen in malignant brain neoplasms. A study by Yerrabothala et al. found that in patients with GBM and meningiomas, activation of the coagulation system was evidenced by increased peak TG compared to controls. In nine patients with GBM, peak TG was also reduced after surgery. [ 10 ] Thrombin generation assay (TGA) assessed by calibrated automated thrombogram (CAT) is undoubtedly a useful tool in the detection and management of thrombotic and bleeding disorders. However, there are limitations to its implementation. Firstly, TGA-CAT has limited ability to detect subtle abnormalities in the coagulation system, particularly in cases where dysfunction is complex and occurs in the early stages of clot activation. This is especially relevant in cancer patients, where thrombotic mechanisms may be difficult to capture. Moreover, TGA-CAT results may vary depending on the quality and handling of the sample, which can affect reproducibility. Factors such as sample storage time or blood collection techniques can influence the outcomes. The lack of standardization, with different laboratories using varying calibration methods and test settings, also leads to challenges in comparing results across centers. Finally, while TGA-CAT effectively assesses thrombin activation and clot formation, it has limited capability to evaluate processes related to microthrombosis and fibrinolysis. In advanced cancer states, where the primary mechanism is thrombus breakdown, TGA-CAT may not provide a complete picture of coagulation activity. Coagulation has been proposed as both a biomarker and a prognostic factor in multiple neoplasms [ 11 ]. In the context of glioblastoma monocyte-to-lymphocyte ratio, systemic immune-inflammatory index (SII), platelet-to-lymphocyte ratio, and platelet-to-fibrinogen ratio have been suggested as prognostic factors. A high preoperative SII level is an independent risk factor for glioblastoma prognosis [ 12 ]. A Study by Navone et al. showed that PT and aPTT were significantly reduced in glioblastoma patients compared to meningioma patients. On the other hand, d-dimer, von Willebrand Factor levels, and leukocyte count were significantly higher. [ 13 ] Moreover, these findings correlated with overall survival [ 13 ]. Mafia et al. confirmed these results and showed that concentration of von Willebrand factor can be a prognostic factor in glioblastoma patients [ 14 ]. A model based on five genes related to coagulation has been proposed in glioblastoma prognosis [ 15 ]. This study demonstrated that glioblastoma has a distinct coagulation signature and explored this as a potential future target for treatment. Thromboembolic events have been implicated in glioblastoma patients, with an estimated frequency of up to 30% [ 16 ]. Average time to the development of thromboembolic events in glioblastoma patients was found to be 6.5 months [ 17 ]. Factors associated with the development of thromboembolic events in these patients include BMI ≥ 35, Karnofsky scale grade ≤ 80, history of venous thromboembolism, and steroid therapy. In our group of patients mean BMI score was 28.44 so it was elevated but still significantly lower than in the mentioned study. On the other hand the mean Karnofsky’s score was 76.9. Two patients (4.76%) suffered from venous thromboembolism during hospitalization and 3 (7.14%) after discharge of which one experienced VTE in both periods. A study by Kapetein et al. involving nearly 1,000 patients with glioblastoma demonstrated that these patients are at significant risk of both venous and arterial thrombosis, as well as major bleeding [ 18 ]. In this study, thromboembolic events were more frequent in patients that underwent surgery compared to those who had a biopsy [ 19 ]. Comparatively, very few studies have focused on biomarkers. Despite the fact that primary brain neoplasms are almost exclusively limited to the brain convexity, malignant gliomas show significant impact on systemic coagulation. A study by Cui et al. demonstrated that in cancer patients (excluding brain neoplasms) systemic coagulation is activated only in the presence of metastatic disease [ 19 ]. In this study, the only common coagulation factor that was elevated regardless of metastatic diseases was D-dimer. Other factors such as TAT, PIC, tPAI, FDP were elevated only in the metastatic group [ 19 ]. Similarly, a study by Lundebech et al. concluded that hypercoagulable state occurs in patients with lung and head and neck cancers, as evidenced by elevated TAT and F1 + F2 [ 20 ]. Kirwan et al. suggested that circulating tumor cells are primarily responsible for the activation of systemic coagulation in cancer patients [ 21 ]. This phenomenon warrants further exploration in brain tumor patients. However, the presence of systemic circulating cells in brain cancer is limited due to a blood-brain barrier. Unfortunately, we were unable to find studies comparing the coagulation state of patients with glioblastoma and those with other cancers. A limitation of our study is the small cohort of patients with glioblastoma. In addition, TGA-CAT is a test with no standards, and only a comparison with healthy individuals can give perspective on its results. More research needs to be done to properly assess the potential of TGA-CAT. Moreover, TGA-CAT has limited ability to detect subtle abnormalities in the coagulation system. Other tools should also be tested in assessing the coagulation system in patients with GBM. Conclusions Primary brain neoplasms are rarely considered tumors with systemic effects, as the disease is typically confined to the brain. This study emphasizes that glioblastoma should be regarded as a systemic disease. Patients with glioblastoma showed higher values of following parameters preoperatively such as d-dimer, PAI:Ag and TAT. They also showed longer Lag Time and TTP than the control group. While high values of d-dimer, PAI:AG and TAT concentrations are prothrombotic parameters, longer Lag Time and TTP did not indicate increased systemic coagulation activity. The above information sheds new light on coagulation disorders in glioblastoma patients. This may also indicate that TGA-CAT has limited ability to detect subtle abnormalities in the coagulation system. In both situations further studies are required. Primary VTE prophylaxis and its biomarkers should be further investigated until more effective strategies and therapies for glioma are identified. Declarations Funding: Statutory founds from Jagiellonian University Medical College. FOunding number N41/DBS/000992 Competing Interests: Authors are required to disclose financial or non-financial interests that are directly or indirectly related to the work submitted for publication. Interests within the last 3 years of beginning the work (conducting the research and preparing the work for submission) should be reported. Interests outside the 3-year time frame must be disclosed if they could reasonably be perceived as influencing the submitted work. Author Contributions: All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by Justyna Rybus, Magdalena Dębicka, Teresa Iwaniec, Sandra Pilawska and Roger Krzyżewski. The first draft of the manuscript was written by Sandra Pilawska, Justyna Rybus and Roger Krzyżewski and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript. Research Data Policy and Data Availability: The datasets generated and analysed during the current study are available from the corresponding author on reasonable request. Ethics approval statement: Study was approved by Jagiellonian University Bioethical Comiettie Consent to participate: Informed consent was obtained from all individual participants included in the study. References Timp JF, Braekkan SK, Versteeg HH, Cannegieter SC. Epidemiology of cancer-associated venous thrombosis. Blood. 2013 Sep 5;122(10):1712-23. doi: 10.1182/blood-2013-04-460121. Epub 2013 Aug 1. PMID: 23908465. Rolston JD, Han SJ, Bloch O, Parsa AT. What clinical factors predict the incidence of deep venous thrombosis and pulmonary embolism in neurosurgical patients? J Neurosurg. 2014 Oct;121(4):908-18. doi: 10.3171/2014.6.JNS131419. Epub 2014 Aug 1. PMID: 25084467. Jeraq M, Cote DJ, Smith TR. Venous Thromboembolism in Brain Tumor Patients. Adv Exp Med Biol. 2017;906:215-228. doi: 10.1007/5584_2016_117. 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Incidence and determinants of thrombotic and bleeding complications in patients with glioblastoma. J Thromb Haemost. 2022 Jul;20(7):1665-1673. doi: 10.1111/jth.15739. Epub 2022 May 9. PMID: 35460331; PMCID: PMC9320838. Cui C, Gao J, Li J, Yu M, Zhang H, Cui W. Value of TAT and PIC with D-dimer for cancer patients with metastasis. Int J Lab Hematol. 2020 Aug;42(4):387-393. doi: 10.1111/ijlh.13194. Epub 2020 Apr 6. Erratum in: Int J Lab Hematol. 2023 Dec;45(6):1020. doi: 10.1111/ijlh.14169. PMID: 32250048. Lundbech M, Krag AE, Christensen TD, Hvas AM. Thrombin generation, thrombin-antithrombin complex, and prothrombin fragment F1+2 as biomarkers for hypercoagulability in cancer patients. Thromb Res. 2020 Feb;186:80-85. doi: 10.1016/j.thromres.2019.12.018. Epub 2019 Dec 28. Erratum in: Thromb Res. 2022 Aug;216:130-132. doi: 10.1016/j.thromres.2021.11.017. PMID: 31918352. Kirwan CC, Descamps T, Castle J. Circulating tumour cells and hypercoagulability: a lethal relationship in metastatic breast cancer. Clin Transl Oncol. 2020 Jun;22(6):870-877. doi: 10.1007/s12094-019-02197-6. Epub 2019 Aug 31. PMID: 31473984; PMCID: PMC7188731. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6219685","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":433393850,"identity":"d899edbf-e69c-43db-bb83-d4131d738095","order_by":0,"name":"Justyna Joanna Rybus","email":"","orcid":"","institution":"Szpital Uniwersytecki w Krakowie","correspondingAuthor":false,"prefix":"","firstName":"Justyna","middleName":"Joanna","lastName":"Rybus","suffix":""},{"id":433393852,"identity":"0aec3279-97a9-4f61-91be-d12bf4944e6f","order_by":1,"name":"Sandra Agnieszka Pilawska","email":"","orcid":"","institution":"Szpital Uniwersytecki w Krakowie","correspondingAuthor":false,"prefix":"","firstName":"Sandra","middleName":"Agnieszka","lastName":"Pilawska","suffix":""},{"id":433393853,"identity":"cdb4522b-907b-4db1-afad-8d38cdc64389","order_by":2,"name":"Magdalena Dębicka","email":"","orcid":"","institution":"Szpital Uniwersytecki w Krakowie","correspondingAuthor":false,"prefix":"","firstName":"Magdalena","middleName":"","lastName":"Dębicka","suffix":""},{"id":433393854,"identity":"eda0d5a5-f990-4611-89d8-c31a4c8e26af","order_by":3,"name":"Roger Marek Krzyżewski","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABD0lEQVRIie3PsWrDMBCA4TMCe7ngjgoJzSuoGGJCh76KgsFaVDoUgkaVgLX0Afw4LoZMJtAto02gc7O11IE6SespNhkL1Y9Ap4MPbACb7e/G2wH948ttZu9SMtS/hFxIgGWHu4eExrxuFcQPk1RUW6zjcbDhJbwvcgg7yLgoHm8KkLOnlEfBIJE43XDupOscZsvzhFIZN1+vGKF8NbrX6kjIIMmB5Z1EfB6IS+fmS9YKg7Qh+14iVo4GyZBGLpGuREYb4vQRLMhQs5hRfCOjfRIjLUr+8rwW2Pkvnql2WkVsYkS1S+vozjdyXn4sbq9DT583gAygOXDF2w3PfvYdeeXp9rN2c5q6ic1ms/2zvgGsG1ezY7qnygAAAABJRU5ErkJggg==","orcid":"","institution":"Jagiellonian University","correspondingAuthor":true,"prefix":"","firstName":"Roger","middleName":"Marek","lastName":"Krzyżewski","suffix":""},{"id":433393855,"identity":"42608526-f1a8-4c0a-9f49-2979f3974f26","order_by":4,"name":"Joanna Zdziarska","email":"","orcid":"","institution":"Szpital Uniwersytecki w Krakowie","correspondingAuthor":false,"prefix":"","firstName":"Joanna","middleName":"","lastName":"Zdziarska","suffix":""},{"id":433393856,"identity":"cbad1d3a-d36f-49e2-857d-b1b8daf60c08","order_by":5,"name":"Teresa Iwaniec","email":"","orcid":"","institution":"Szpital Uniwersytecki w Krakowie","correspondingAuthor":false,"prefix":"","firstName":"Teresa","middleName":"","lastName":"Iwaniec","suffix":""},{"id":433393857,"identity":"2602e743-afd7-4671-8523-d64b4478e8c4","order_by":6,"name":"Borys Maria Kwinta","email":"","orcid":"","institution":"Jagiellonian University","correspondingAuthor":false,"prefix":"","firstName":"Borys","middleName":"Maria","lastName":"Kwinta","suffix":""},{"id":433393858,"identity":"f9558f2a-c0ab-48b9-87c2-4221512ae1c9","order_by":7,"name":"Krzysztof Stachura","email":"","orcid":"","institution":"Jagiellonian University","correspondingAuthor":false,"prefix":"","firstName":"Krzysztof","middleName":"","lastName":"Stachura","suffix":""}],"badges":[],"createdAt":"2025-03-13 11:38:21","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6219685/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6219685/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":79562233,"identity":"616e8f47-9923-43c4-9b98-4a59c6bf74ba","added_by":"auto","created_at":"2025-03-31 08:55:32","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":141389,"visible":true,"origin":"","legend":"\u003cp\u003eComorbidities in patient’s population\u003c/p\u003e","description":"","filename":"Fiugre1.png","url":"https://assets-eu.researchsquare.com/files/rs-6219685/v1/931adcc30b3330d91658a7f5.png"},{"id":79564225,"identity":"44648d6b-4541-41da-9f44-6e42570f37da","added_by":"auto","created_at":"2025-03-31 09:11:32","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":135423,"visible":true,"origin":"","legend":"\u003cp\u003eA comparison of test results between study and control group. A - TTP in GBM and control group, B - TAT in GBM and control group, C - Peak in GBM and control group, D - ETP in GBM and control group, E - PAI:Ag in GBM and control group, F - DD in GBM and control group.\u003c/p\u003e","description":"","filename":"Fiugre2.png","url":"https://assets-eu.researchsquare.com/files/rs-6219685/v1/b4a0c52213028e9d57ed4f30.png"},{"id":80446562,"identity":"d72c47de-30cd-4fa0-97af-b14bffe3de18","added_by":"auto","created_at":"2025-04-12 11:31:32","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":704508,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6219685/v1/0e5c62bd-e490-482b-82af-3d7400ced531.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Systemic coagulation activation in patients with glioblastoma","fulltext":[{"header":"Introduction","content":"\u003cp\u003eThe risk of venous thromboembolism (VTE) increases four- to sevenfold in patients with cancer in comparison to the general population [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. Patients with brain tumors often suffer from motor deficits that can render them bedridden [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. Brain tumors predispose to VTE more than cancer in other sites [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. This fact is associated with many additive risk factors such as cancer itself, the thrombotic response to surgical trauma and limited postoperative mobility. Glioblastoma (GBM) causes chronic activation of the coagulation system. This phenomenon occurs because tumor cells contain transcripts for several coagulation factors and hemostasis regulating molecules. Furthermore, this molecular combination seems to differ between brain tumor subtypes [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. This raises the question of whether the patient will benefit from pharmacological thromboprophylaxis or if it could be harmful. Results from conducted prospective studies are unclear, but there are trends indicating serious hemorrhagic complications after long-term administration of low-molecular-weight heparin (LMWH) compared to the placebo group [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Considering the high risk of VTE and complications associated with long-term pharmacological thromboprophylaxis, appropriate selection of patients for treatment should be made.\u003c/p\u003e \u003cp\u003eBasic coagulation tests, such as prothrombin time (PT) and activated partial thromboplastin time (APTT), do not accurately represent the in vivo balance of coagulation. However, in patients with congenital deficiencies of natural anticoagulants (e.g., protein C, protein S, antithrombin), these parameters are often within reference ranges. This diagnostic gap in assessing the patient's hemostasis status is helped by the calibrated automated thrombogram (CAT), a global dynamic test that continuously and simultaneously examines both thrombin formation and its inhibition. Therefore, CAT is an invaluable tool for diagnosing, monitoring or implementing treatment for many hemorrhagic diseases. It is also highly relevant in the field of thrombosis for assessing the risk of VTE and determining a favorable time to administer antithrombotic drugs [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe aim of the study was to examine new biomarkers that could help profile patients, potentially allowing for the estimation of both pre- and postoperative VTE risk in patients with GBM.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eStudy population\u003c/h2\u003e \u003cp\u003eForty-two patients with newly diagnosed glioblastoma who underwent surgical resection at the Department of Neurosurgery and Neurotraumatology of University Hospital in Krak\u0026oacute;w were enrolled in the study. The inclusion criteria were: age over 18 years, newly diagnosed brain tumor with confirmed histopathology: Glioblastoma, IDH-wildtype. Exclusion criteria were: continuous anticoagulant treatment with vitamin K antagonists, low-molecular-weight heparins or antiplatelet drugs, bacterial or viral infection, pregnancy.\u003c/p\u003e \u003cp\u003eThe control group, matched for age and gender, consisted of 35 healthy individuals (18 males and 17 females, median age of 50.29\u0026thinsp;\u0026plusmn;\u0026thinsp;11.50 years) without a history of cancer, with no personal or family history of thrombosis or medications that could affect the clotting system. The control group was selected from among the patients of the hematology clinic where haematological diseases were excluded.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eData evaluation\u003c/h3\u003e\n\u003cp\u003eThe medical history of patients from the study group was evaluated in terms of sex, age, weight, height, BMI (body mass index), comorbidities, drug intake and complications during hospitalization. Thromboprophylaxis was assessed both in the preoperative and postoperative periods, including the timing of its administration. Other factors considered included preoperative steroid use, mean operative time, position during surgery and extent of resection. Laboratory tests, including complete blood count, sodium and potassium levels, glucose, creatinine and urea, C-reactive protein (CRP), albumin, fibrinogen, fibrinogen degradation products (FDP), antithrombin (AT) and D-dimers (DD) were established upon admission. Values of FDP were presented in 3 ranges \u0026minus;\u0026thinsp;0 when the FDP value was less than 5 mg/l, 1 when values were in the range of 5\u0026ndash;15 mg/l and 2 when were above 20 mg/l. APTT and PT were assessed upon admission.\u003c/p\u003e \u003cp\u003eTumors were evaluated according to the World Health Organization (WHO) 2021 classification of Central Nervous System (CNS) tumors [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. All patients were assessed using the Caprini Score for Venous Thromboembolism, the Karnofsky Scale and the ASA (American Society of Anaesthesiology) scale. Deep vein thrombosis was confirmed by compression ultrasound or computed tomography angiography (CT-angiography) in individual cases. Diagnosis was made by board certified vascular surgeon.\u003c/p\u003e \u003cp\u003eThe medical condition of patients in the study group was monitored for the period of 12 months following surgery. The study protocol was approved by the Bioethical Committee of the Jagiellonian University in Krak\u0026oacute;w, Poland (approval number 1072.6120.92.2019), and written informed consent was obtained from all participants.\u003c/p\u003e\n\u003ch3\u003eBlood Sampling and Laboratory Assays\u003c/h3\u003e\n\u003cp\u003eSamples containing 20 ml of blood were collected into tubes with EDTA and 3.2% sodium citrate by sterile and atraumatic venipuncture on the day of the scheduled surgery. The procedure was repeated 24 and 72 hours postoperatively. Plasma samples were frozen at -80\u0026deg;C until analyzed. All tests, including a thrombin generation assay - calibrated automated thrombogram (TGA-CAT), of both the control and study groups, were conducted in the same laboratory using identical equipment and by the same practitioner.\u003c/p\u003e \u003cp\u003eA thrombin generation assay (TGA) was assessed by a calibrated automated thrombogram (CAT). The plasma samples were measured in the 96-well plate fluorometer (Ascent Reader, Thermolabsystems OY, Helsinki, Finland), equipped with the 390/460 filter set and a dispenser. In brief, 80 \u0026micro;l of platelet-poor plasma was combined with 20 \u0026micro;l of tissue factor (TF)-based activator (Diagnostica Stago, Asni\u0026egrave;res, France) and 20 \u0026micro;l of FluCa solution (Diagnostica Stago, Asni\u0026egrave;res, France). A dedicated software program (Thrombinoscope, Synapse BV, Maastricht, The Netherlands) provided the evaluation of plasma thrombogenic potential [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eFour parameters of TGA-CAT were assessed: thrombin peak (maximum concentration of thrombin formed during the recording time) - nmol (P), endogenous thrombin potential (the area under the curve) - nmol/min (ETP), lag time - min (LT), time to peak - min (TTP).\u003c/p\u003e\n\u003ch3\u003eStatistics\u003c/h3\u003e\n\u003cp\u003eElements of descriptive statistics were used. Continuous variables are expressed as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation, and proportions are presented as percentage value. Statistical analysis was performed using STATISTICA v. 13.0 (Statsoft, Poland). Pearson's Chi-square test was used to assess proportions, and the t-test was used to assess continuous variables. Pearson's correlation coefficient was used to evaluate correlations between variables. Differences were considered significant at p\u0026thinsp;\u0026lt;\u0026thinsp;0.05.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eWe analyzed prospectively collected data from 42 patients with newly diagnosed glioblastoma confirmed in histopathology examination. Mean age was 65.19\u0026thinsp;\u0026plusmn;\u0026thinsp;7.02 years. Mean BMI score was 28.44\u0026thinsp;\u0026plusmn;\u0026thinsp;5.73 kg/m2 and mean ASA scale score 2.26\u0026thinsp;\u0026plusmn;\u0026thinsp;0.59. In the Caprini Score the average patient got 6.46\u0026thinsp;\u0026plusmn;\u0026thinsp;1.05 points. Two patients (4.76%) suffered from venous thromboembolism during hospitalization and 3 (7.14%) after discharge of which one experienced VTE in both periods. Patient\u0026rsquo;s comorbidities are shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e. Mean Karnofsky\u0026rsquo;s score was 76.9\u0026thinsp;\u0026plusmn;\u0026thinsp;14.23.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eAnticoagulation therapy was withheld in all patients for a minimum of 24 hours before surgery, while antiplatelet therapy was withheld for 7 days preoperatively. Four patients (9.52%) received a perioperative antithrombotic drug (low-dose heparin). Twenty-seven patients (64.29%) received dexamethasone preoperatively at an average dose of 8 mg.\u003c/p\u003e \u003cp\u003eIn twenty-two patients (52.38%) gross-total resection (GTR) was achieved, 8 patients (19.05%) underwent subtotal resection (STR) and in 12 patients (28.57%) biopsy of the lesion was performed. Sixteen patients (38.10%) got prophylactic dosage of LWMH postoperatively on average on the second day after surgery (2.25\u0026thinsp;\u0026plusmn;\u0026thinsp;1.29 days) due to a prolonged period of immobilization. Seven patients (16.67%) died before discharge. Five of them suffered from postoperative complications in the form of hematoma in the operation site (3 cases), epidural hematoma and intracerebral hematoma.\u003c/p\u003e \u003cp\u003eThe study group had significantly higher concentrations of D-dimer (1.8\u0026thinsp;\u0026plusmn;\u0026thinsp;3.6 vs 0.35\u0026thinsp;\u0026plusmn;\u0026thinsp;0.32, p\u0026thinsp;=\u0026thinsp;0.02), PAI:Ag (27.30\u0026thinsp;\u0026plusmn;\u0026thinsp;8.90 vs 17.00\u0026thinsp;\u0026plusmn;\u0026thinsp;9.34, p\u0026thinsp;\u0026lt;\u0026thinsp;0.01) and TAT (15.20\u0026thinsp;\u0026plusmn;\u0026thinsp;17.00 vs 4.56\u0026thinsp;\u0026plusmn;\u0026thinsp;1.66, p\u0026thinsp;\u0026lt;\u0026thinsp;0.01). In TGA-CAT, the following parameters were also longer in the study group than in the control group: Lag Time (3.5\u0026thinsp;\u0026plusmn;\u0026thinsp;1.00 vs 3.02\u0026thinsp;\u0026plusmn;\u0026thinsp;0.69, p\u0026thinsp;=\u0026thinsp;0.01) and TTP (6.60\u0026thinsp;\u0026plusmn;\u0026thinsp;1.50 vs 5.73\u0026thinsp;\u0026plusmn;\u0026thinsp;1.04, p\u0026thinsp;=\u0026thinsp;0.01). There were no statistically significant differences between the groups in ETP (1513.10\u0026thinsp;\u0026plusmn;\u0026thinsp;316.60 vs 1590.48\u0026thinsp;\u0026plusmn;\u0026thinsp;266.84, p\u0026thinsp;=\u0026thinsp;0.28) and Peak (292.20\u0026thinsp;\u0026plusmn;\u0026thinsp;83.00 vs 307.74\u0026thinsp;\u0026plusmn;\u0026thinsp;55.84, p\u0026thinsp;=\u0026thinsp;0.37). A comparison of results between study and control groups is presented in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eIn this prospective cohort study, we observed elevated levels of D-dimer, thrombin-antithrombin complexes, and PAI:Ag, alongside prolonged lag time and time to peak. This inconsistency can be explained in several ways. In certain diseases, such as cancer, microangiopathy can alter platelet function. Reduced platelet numbers or dysfunction can shorten the thrombin activation process, leading to prolonged lag time and TTP. At the same time, this can promote clot formation in the microcirculation, as evidenced by elevated D-dimer levels.\u003c/p\u003e \u003cp\u003eIn the context of glioblastoma patients, prolonged lag time and time to peak may suggest that a hypercoagulable state is not detected by standard tests, such as CAT, or that these tests lack the sensitivity to identify subtle clotting abnormalities. This indicates that thrombotic mechanisms in these patients may be more complex or less predictable.\u003c/p\u003e \u003cp\u003eThis is one of the very few studies demonstrating systemic coagulation processes in glioblastoma compared to healthy subjects. Most studies use patients with meningiomas as controls, which may show some coagulation abnormalities but not as pronounced as those seen in malignant brain neoplasms. A study by Yerrabothala et al. found that in patients with GBM and meningiomas, activation of the coagulation system was evidenced by increased peak TG compared to controls. In nine patients with GBM, peak TG was also reduced after surgery. [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]\u003c/p\u003e \u003cp\u003eThrombin generation assay (TGA) assessed by calibrated automated thrombogram (CAT) is undoubtedly a useful tool in the detection and management of thrombotic and bleeding disorders. However, there are limitations to its implementation. Firstly, TGA-CAT has limited ability to detect subtle abnormalities in the coagulation system, particularly in cases where dysfunction is complex and occurs in the early stages of clot activation. This is especially relevant in cancer patients, where thrombotic mechanisms may be difficult to capture. Moreover, TGA-CAT results may vary depending on the quality and handling of the sample, which can affect reproducibility. Factors such as sample storage time or blood collection techniques can influence the outcomes. The lack of standardization, with different laboratories using varying calibration methods and test settings, also leads to challenges in comparing results across centers. Finally, while TGA-CAT effectively assesses thrombin activation and clot formation, it has limited capability to evaluate processes related to microthrombosis and fibrinolysis. In advanced cancer states, where the primary mechanism is thrombus breakdown, TGA-CAT may not provide a complete picture of coagulation activity.\u003c/p\u003e \u003cp\u003eCoagulation has been proposed as both a biomarker and a prognostic factor in multiple neoplasms [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. In the context of glioblastoma monocyte-to-lymphocyte ratio, systemic immune-inflammatory index (SII), platelet-to-lymphocyte ratio, and platelet-to-fibrinogen ratio have been suggested as prognostic factors. A high preoperative SII level is an independent risk factor for glioblastoma prognosis [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. A Study by Navone et al. showed that PT and aPTT were significantly reduced in glioblastoma patients compared to meningioma patients. On the other hand, d-dimer, von Willebrand Factor levels, and leukocyte count were significantly higher. [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e] Moreover, these findings correlated with overall survival [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. Mafia et al. confirmed these results and showed that concentration of von Willebrand factor can be a prognostic factor in glioblastoma patients [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. A model based on five genes related to coagulation has been proposed in glioblastoma prognosis [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. This study demonstrated that glioblastoma has a distinct coagulation signature and explored this as a potential future target for treatment.\u003c/p\u003e \u003cp\u003eThromboembolic events have been implicated in glioblastoma patients, with an estimated frequency of up to 30% [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Average time to the development of thromboembolic events in glioblastoma patients was found to be 6.5 months [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. Factors associated with the development of thromboembolic events in these patients include BMI\u0026thinsp;\u0026ge;\u0026thinsp;35, Karnofsky scale grade\u0026thinsp;\u0026le;\u0026thinsp;80, history of venous thromboembolism, and steroid therapy. In our group of patients mean BMI score was 28.44 so it was elevated but still significantly lower than in the mentioned study. On the other hand the mean Karnofsky\u0026rsquo;s score was 76.9. Two patients (4.76%) suffered from venous thromboembolism during hospitalization and 3 (7.14%) after discharge of which one experienced VTE in both periods.\u003c/p\u003e \u003cp\u003eA study by Kapetein et al. involving nearly 1,000 patients with glioblastoma demonstrated that these patients are at significant risk of both venous and arterial thrombosis, as well as major bleeding [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e]. In this study, thromboembolic events were more frequent in patients that underwent surgery compared to those who had a biopsy [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Comparatively, very few studies have focused on biomarkers.\u003c/p\u003e \u003cp\u003eDespite the fact that primary brain neoplasms are almost exclusively limited to the brain convexity, malignant gliomas show significant impact on systemic coagulation. A study by Cui et al. demonstrated that in cancer patients (excluding brain neoplasms) systemic coagulation is activated only in the presence of metastatic disease [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. In this study, the only common coagulation factor that was elevated regardless of metastatic diseases was D-dimer. Other factors such as TAT, PIC, tPAI, FDP were elevated only in the metastatic group [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Similarly, a study by Lundebech et al. concluded that hypercoagulable state occurs in patients with lung and head and neck cancers, as evidenced by elevated TAT and F1\u0026thinsp;+\u0026thinsp;F2 [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. Kirwan et al. suggested that circulating tumor cells are primarily responsible for the activation of systemic coagulation in cancer patients [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. This phenomenon warrants further exploration in brain tumor patients. However, the presence of systemic circulating cells in brain cancer is limited due to a blood-brain barrier. Unfortunately, we were unable to find studies comparing the coagulation state of patients with glioblastoma and those with other cancers.\u003c/p\u003e \u003cp\u003eA limitation of our study is the small cohort of patients with glioblastoma. In addition, TGA-CAT is a test with no standards, and only a comparison with healthy individuals can give perspective on its results. More research needs to be done to properly assess the potential of TGA-CAT. Moreover, TGA-CAT has limited ability to detect subtle abnormalities in the coagulation system. Other tools should also be tested in assessing the coagulation system in patients with GBM.\u003c/p\u003e"},{"header":"Conclusions","content":"\u003cp\u003ePrimary brain neoplasms are rarely considered tumors with systemic effects, as the disease is typically confined to the brain. This study emphasizes that glioblastoma should be regarded as a systemic disease. Patients with glioblastoma showed higher values of following parameters preoperatively such as d-dimer, PAI:Ag and TAT. They also showed longer Lag Time and TTP than the control group. While high values of d-dimer, PAI:AG and TAT concentrations are prothrombotic parameters, longer Lag Time and TTP did not indicate increased systemic coagulation activity. The above information sheds new light on coagulation disorders in glioblastoma patients. This may also indicate that TGA-CAT has limited ability to detect subtle abnormalities in the coagulation system. In both situations further studies are required. Primary VTE prophylaxis and its biomarkers should be further investigated until more effective strategies and therapies for glioma are identified.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eFunding:\u003c/strong\u003e Statutory founds from Jagiellonian University Medical College. FOunding number N41/DBS/000992\u003c/p\u003e\n\u003cp\u003eCompeting Interests: Authors are required to disclose financial or non-financial interests that are directly or indirectly related to the work submitted for publication. Interests within the last 3 years of beginning the work (conducting the research and preparing the work for submission) should be reported. Interests outside the 3-year time frame must be disclosed if they could reasonably be perceived as influencing the submitted work.\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003cstrong\u003eAuthor Contributions:\u003c/strong\u003e All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by Justyna Rybus, Magdalena Dębicka, Teresa Iwaniec, Sandra Pilawska and Roger Krzyżewski. The first draft of the manuscript was written by Sandra Pilawska, Justyna Rybus and Roger Krzyżewski and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResearch Data Policy and Data Availability:\u003c/strong\u003e The datasets generated and analysed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval statement: Study was approved by Jagiellonian University Bioethical Comiettie\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent to participate:\u0026nbsp;\u003c/strong\u003eInformed consent was obtained from all individual participants included in the study.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eTimp JF, Braekkan SK, Versteeg HH, Cannegieter SC. Epidemiology of cancer-associated venous thrombosis. Blood. 2013 Sep 5;122(10):1712-23. doi: 10.1182/blood-2013-04-460121. Epub 2013 Aug 1. PMID: 23908465.\u003c/li\u003e\n\u003cli\u003eRolston JD, Han SJ, Bloch O, Parsa AT. 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C., Ahmed, S. R., Guerin, S. J., Porter, M., Wishart, H. A., Ernstoff, M. S., Fadul, C. E., \u0026amp; Ornstein, D. L. (2021). Systemic coagulation is activated in patients with meningioma and glioblastoma. Journal of Neuro-Oncology, 155(2), 173\u0026ndash;180. https://doi.org/10.1007/s11060-021-03865-w\u003c/li\u003e\n\u003cli\u003eWahab R, Hasan MM, Azam Z, Grippo PJ, Al-Hilal TA. The role of coagulome in the tumor immune microenvironment. Adv Drug Deliv Rev. 2023 Sep;200:115027. doi: 10.1016/j.addr.2023.115027. Epub 2023 Jul 28. PMID: 37517779; PMCID: PMC11099942.\u003c/li\u003e\n\u003cli\u003eDuan X, Yang B, Zhao C, Tie B, Cao L, Gao Y. Prognostic value of preoperative hematological markers in patients with glioblastoma multiforme and construction of random survival forest model. BMC Cancer. 2023 May 12;23(1):432. doi: 10.1186/s12885-023-10889-0. PMID: 37173662; PMCID: PMC10176909. \u003c/li\u003e\n\u003cli\u003eNavone SE, Guarnaccia L, Locatelli M, Rampini P, Caroli M, La Verde N, Gaudino C, Bettinardi N, Riboni L, Marfia G, Campanella R. Significance and Prognostic Value of The Coagulation Profile in Patients with Glioblastoma: Implications for Personalized Therapy. World Neurosurg. 2019 Jan;121:e621-e629. doi: 10.1016/j.wneu.2018.09.177. Epub 2018 Oct 3. PMID: 30292037.\u003c/li\u003e\n\u003cli\u003eMarfia G, Navone SE, Fanizzi C, Tabano S, Pesenti C, Abdel Hadi L, Franzini A, Caroli M, Miozzo M, Riboni L, Rampini P, Campanella R. Prognostic value of preoperative von Willebrand factor plasma levels in patients with Glioblastoma. Cancer Med. 2016 Aug;5(8):1783-90. doi: 10.1002/cam4.747. Epub 2016 May 28. PMID: 27236861; PMCID: PMC4887291.\u003c/li\u003e\n\u003cli\u003eZhou M, Deng Y, Fu Y, Liang R, Liu Y, Liao Q. A new prognostic model for glioblastoma multiforme based on coagulation-related genes. Transl Cancer Res. 2023 Oct 31;12(10):2898-2910. doi: 10.21037/tcr-23-322. Epub 2023 Oct 10. PMID: 37969372; PMCID: PMC10643966.\u003c/li\u003e\n\u003cli\u003eKapteijn MY, Bakker N, Koekkoek JAF, Versteeg HH, Buijs JT. Venous Thromboembolism in Patients with Glioblastoma: Molecular Mechanisms and Clinical Implications. Thromb Haemost. 2024 Aug 21. doi: 10.1055/s-0044-1789592. Epub ahead of print. PMID: 39168144.\u003c/li\u003e\n\u003cli\u003eYust-Katz S, Mandel JJ, Wu J, Yuan Y, Webre C, Pawar TA, Lhadha HS, Gilbert MR, Armstrong TS. Venous thromboembolism (VTE) and glioblastoma. J Neurooncol. 2015 Aug;124(1):87-94. doi: 10.1007/s11060-015-1805-2. Epub 2015 May 19. PMID: 25985958.\u003c/li\u003e\n\u003cli\u003eKaptein FHJ, Stals MAM, Kapteijn MY, Cannegieter SC, Dirven L, van Duinen SG, van Eijk R, Huisman MV, Klaase EE, Taphoorn MJB, Versteeg HH, Buijs JT, Koekkoek JAF, Klok FA. Incidence and determinants of thrombotic and bleeding complications in patients with glioblastoma. J Thromb Haemost. 2022 Jul;20(7):1665-1673. doi: 10.1111/jth.15739. Epub 2022 May 9. PMID: 35460331; PMCID: PMC9320838.\u003c/li\u003e\n\u003cli\u003eCui C, Gao J, Li J, Yu M, Zhang H, Cui W. Value of TAT and PIC with D-dimer for cancer patients with metastasis. Int J Lab Hematol. 2020 Aug;42(4):387-393. doi: 10.1111/ijlh.13194. Epub 2020 Apr 6. Erratum in: Int J Lab Hematol. 2023 Dec;45(6):1020. doi: 10.1111/ijlh.14169. PMID: 32250048.\u003c/li\u003e\n\u003cli\u003eLundbech M, Krag AE, Christensen TD, Hvas AM. Thrombin generation, thrombin-antithrombin complex, and prothrombin fragment F1+2 as biomarkers for hypercoagulability in cancer patients. Thromb Res. 2020 Feb;186:80-85. doi: 10.1016/j.thromres.2019.12.018. Epub 2019 Dec 28. Erratum in: Thromb Res. 2022 Aug;216:130-132. doi: 10.1016/j.thromres.2021.11.017. PMID: 31918352.\u003c/li\u003e\n\u003cli\u003eKirwan CC, Descamps T, Castle J. Circulating tumour cells and hypercoagulability: a lethal relationship in metastatic breast cancer. Clin Transl Oncol. 2020 Jun;22(6):870-877. doi: 10.1007/s12094-019-02197-6. Epub 2019 Aug 31. PMID: 31473984; PMCID: PMC7188731.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Thrombin Generation Assay, Glioblastoma, Systemic Coagulation, Coagulation Disorders","lastPublishedDoi":"10.21203/rs.3.rs-6219685/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6219685/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003ePurpose\u003c/h2\u003e \u003cp\u003eTo assess if patients diagnosed with glioblastoma have higher markers of systemic coagulation activation in thrombin generation assay (TGA) and blood tests.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eWe collected preoperative blood samples from 42 glioblastoma patients and 35 healthy controls. We assessed fibrinogen concentration, fibrinogen degradation products (FDP) and D-dimer concentrations, plasminogen activator inhibitor concentration (PAI:Ag), antithrombin (AT) activity, thrombin-antithrombin complexes (TAT), the thrombin peak (P), endogenous thrombin potential (ETP), Lag Time and Time To Peak (TTP).\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003ePatients with glioblastoma showed higher values of following parameters preoperatively such as D-dimer concentration (1.8\u0026thinsp;\u0026plusmn;\u0026thinsp;3.6 vs 0.35\u0026thinsp;\u0026plusmn;\u0026thinsp;0.32 mg/l, p\u0026thinsp;=\u0026thinsp;0.02), PAI:Ag (27.30\u0026thinsp;\u0026plusmn;\u0026thinsp;8.90 vs 17.00\u0026thinsp;\u0026plusmn;\u0026thinsp;9.34, p\u0026thinsp;\u0026lt;\u0026thinsp;0.01) and TAT (15.20\u0026thinsp;\u0026plusmn;\u0026thinsp;17.00 vs 4.56\u0026thinsp;\u0026plusmn;\u0026thinsp;1.66, p\u0026thinsp;\u0026lt;\u0026thinsp;0.01). Furthermore they showed longer Lag Time (3.50\u0026thinsp;\u0026plusmn;\u0026thinsp;1.00 vs 3.02\u0026thinsp;\u0026plusmn;\u0026thinsp;0.69, p\u0026thinsp;=\u0026thinsp;0.01) and TTP (6.60\u0026thinsp;\u0026plusmn;\u0026thinsp;1.50 vs 5.73\u0026thinsp;\u0026plusmn;\u0026thinsp;1.04, p\u0026thinsp;=\u0026thinsp;0.01) in TGA.\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003eHigh values of d-dimer, PAI:AG and TAT concentrations are prothrombotic parameters but longer Lag Time and TTP did not indicate increased systemic coagulation activity. The above information sheds new light on coagulation disorders in glioblastoma patients. This may also indicate that TGA assessed by calibrated automated thrombogram (CAT) has limited ability to detect subtle abnormalities in the coagulation system. In both situations further studies are required.\u003c/p\u003e","manuscriptTitle":"Systemic coagulation activation in patients with glioblastoma","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-03-31 08:55:27","doi":"10.21203/rs.3.rs-6219685/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"cafdcd8c-6ef3-46e9-9cad-e41ba043d420","owner":[],"postedDate":"March 31st, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-04-12T11:23:25+00:00","versionOfRecord":[],"versionCreatedAt":"2025-03-31 08:55:27","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6219685","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6219685","identity":"rs-6219685","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}