Cancer-Associated Thrombosis in Cirrhotic Patients with Hepatocellular Carcinoma.

It is common knowledge that cancer patients are more prone to develop venous thromboembolic complications (VTE). It is therefore not surprising that patients with hepatocellular carcinoma (HCC) present with a significant risk of VTE, with the portal vein being the most frequent site (PVT). However, patients with HCC are peculiar as both cancer and liver cirrhosis are conditions that can perturb the hemostatic balance towards a prothrombotic state. Because HCC-related hypercoagulability is not clarified at all, the aim of the present review is to summarize the currently available knowledge on epidemiology and pathogenesis of non-malignant thrombotic complications in patients with liver cirrhosis and HCC. They are at increased risk to develop both PVT and non-splanchnic VTE, indicating that both local and systemic factors can foster the development of site-specific thrombosis. Recent studies have suggested multiple and often interrelated mechanisms through which HCC can tip the hemostatic balance of liver cirrhosis towards hypercoagulability. Described mechanisms include increased fibrinogen concentration/polymerization, thrombocytosis, and release of tissue factor-expressing extracellular vesicles. Currently, there are no specific guidelines on the use of thromboprophylaxis in this unique population. There is the urgent need of prospective studies assessing which patients have the highest prothrombotic profile and would therefore benefit from early thromboprophylaxis

Neutrophil extracellular traps in central nervous system disorders: mechanisms, implications, and emerging perspective

Neutrophil Extracellular Traps (NETs), as a crucial defense mechanism of neutrophils, have garnered increasing attention in recent years for their roles in central nervous system (CNS) disorders. This review comprehensively summarizes the fundamental characteristics and formation mechanisms of NETs, while highlighting the latest research advances regarding their involvement in various CNS diseases. Specific mechanistic insights are discussed, including how NETs exacerbate ischemic stroke through immunothrombosis, promote blood-brain barrier disruption in multiple sclerosis, and contribute to neuroinflammation in Alzheimer’s disease. The paper systematically explores the potential mechanistic contributions of NETs to disease pathogenesis and progression, as well as their prospects as diagnostic biomarkers and therapeutic targets. Through an in-depth analysis of the multifaceted roles of NETs in CNS pathologies, this review aims to provide novel insights and references for advancing the understanding, clinical diagnosis, and therapeutic management of central nervous system disorders

Neutrophil extracellular traps in cancer and cancer-associated thrombosis

Cancer is associated with a hypercoagulable state, and venous thromboembolism (VTE) may be the first sign of occult cancer. Cancer screening of all patients presenting with VTE would, however, overload the healthcare system and burden patients with unnecessary investigations. Current data suggest that neutrophil extracellular traps (NETs), prothrombotic nuclear content released by neutrophils upon strong stimulation, are central in cancer biology. This thesis aimed at a clinical investigation of the role of coagulation in advanced cancer and the role of NETs in cancer-associated thrombosis. In Study I, we evaluated the recently developed RIETE risk score to identify patients presenting with VTE and a simultaneous high risk of occult cancer. The risk score failed to identify VTE patients with a high risk of occult cancer, illustrating the need for the development of risk score models in this population. In Study II, we developed an enzyme-linked immunosorbent assay for the quantification of nucleosomal citrullinated histone H3 (H3Cit-DNA), a protein-DNA complex generated during NET formation. The assay was rigorously validated revealing high accuracy. All assay components are furthermore commercially available, enabling rapid dissemination and implementation of the assay within the field of NETs research. Study III was an exploratory study investigating several biomarkers reflecting neutrophil activation, NET formation, coagulation, and fibrinolysis and their association with mortality in 106 terminal cancer patients. Markers of neutrophil activation and NETs were associated with mortality in univariate and multivariate Cox regression. Several prior studies have revealed that markers of coagulation and fibrinolysis are associated with prognosis in cancer patients. However, no studies have investigated terminal cancer patients, and to our surprise, we did not find an association between poor prognosis and markers of coagulation and fibrinolysis. Study IV was a prospective cohort study of 500 patients presenting with acute VTE. Venous blood was sampled at the time of VTE, and markers of NETs and neutrophil activation were analyzed. H3Cit-DNA and cell-free DNA were associated with cancer diagnosis during a one-year follow-up in univariate analyses, but only H3Cit-DNA remained significant after adjustments in multivariate analyses, which could indicate a role of NETs in the development of cancer-associated thrombosis. In summary, there are as of date no accurate risk scores identifying VTE patients with underlying cancer. Through the development of an assay quantifying the NET marker H3Cit- DNA in human plasma, we found that H3Cit-DNA is elevated in advanced cancer and in patients presenting with VTE and an underlying cancer, contributing to the growing evidence of the role of NETs in cancer and cancer-associated thrombosis. Further research will determine the diagnostic potential of NETs

Review of Trousseau phenomenon - pathomechanism, diagnosis, treatment and risk of cancer

Introduction: Trousseau's phenomenon, also referred to as malignancy-associated thrombosis, represents a hypercoagulable state commonly encountered in oncology patients, contributing substantially to morbidity and mortality. This review examines the underlying mechanisms of Trousseau’s phenomenon, including the release of tumor-derived procoagulants and immune-inflammatory interactions, alongside contemporary diagnostic methodologies and emerging biomarker candidates. Additionally, therapeutic strategies, with a focus on anticoagulation management, are discussed, highlighting the clinical and prognostic significance of Trousseau’s phenomenon in evaluating cancer progression and risk stratification. The evolving understanding of this condition underscores the necessity of interdisciplinary collaboration in its clinical management and ongoing research efforts. Aim of these study: The aim of this study was to explore the issue of hypercoagulability in the cancer patient population and to investigate the underlying mechanisms contributing to its development. State of knowledge: It is well-established in scientific literature that oncology patients are at a significantly increased risk for thromboembolic events. Neoplasms promote a hypercoagulable state and its associated complications through diverse and complex pathophysiological mechanisms. Conclusions: Cancer-associated thrombosis is a significant clinical challenge in oncology patients. Understanding the underlying mechanisms, identifying specific risk factors, and ensuring early diagnosis are essential for improving prognosis and optimizing therapeutic outcomes

Nuevos biomarcadores trombóticos en tumores gastrointestinales

El tromboembolismo venoso es una comorbilidad relativamente frecuente en los pacientes oncológicos, así como la segunda causa de muerte en esta población. Se han empleado diferentes tratamientos anticoagulantes para prevenir esta complicación, pero su baja eficiencia en población no seleccionada y el aumento de sangrado asociado, han llevado a que la tromprofilaxis generalizada sea descartada. Este escenario destaca en los tumores gastrointestinales, ya que, aunque son altamente trombogénicos, también se asocian a un alto riesgo de sangrado. Así, es necesario identificar nuevos marcadores de riesgo trombótico que permitan seleccionar a aquellos pacientes que podrían beneficiarse de la tromboprofilaxis primaria a pesar del riesgo de sangrado. Por ello, el objetivo de esta tesis es identificar nuevos biomarcadores trombóticos en pacientes con cáncer colorrectal y cáncer gástrico avanzado. En cáncer colorrectal, nos centramos en el estudio de la hepsina, una serín-proteasa implicada en invasión tumoral y en la activación de la vía extrínseca de la coagulación. Retrospectivamente, reclutamos casi 300 pacientes con cáncer colorrectal localizado o metastásico. Mediante inmunohistoquímica, medimos la intensidad de tinción de hepsina en la biopsia primaria al diagnóstico y la correlacionamos con variables clínicas como la incidencia acumulada de trombosis, la recaída metastásica, o la supervivencia global, entre otras. Posteriormente, realizamos estudios in vitro e in vivo basados en células de cáncer colorrectal transfectadas establemente con el gen de la hepsina, y determinamos el efecto de su sobreexpresión en ensayos de migración, invasión, proliferación y generación de trombina. Finalmente, de cara a una aplicación como diana terapéutica, utilizamos las técnicas de "virtual screening" y "docking molecular" para identificar fármacos aprobados por la Administración de Alimentos y Medicamentos (FDA) que pudiesen inhibir la actividad de hepsina, así como suprimir los efectos de su sobreexpresión en las células tumorales. Como resultados principales, describimos la hepsina como un biomarcador de trombosis y recaída metastásica en pacientes con cáncer colorrectal localizado. En concordancia, la sobreexpresión de hepsina en células de cáncer colorrectal aumentaba significativamente su fenotipo invasivo y su capacidad de generación de trombina. Finalmente, descubrimos Venetoclax y Suramin como dos nuevos inhibidores de hepsina que, además, suprimían sus efectos protumorales y trombogénicos. En conclusión, la hepsina expresada en el tumor primario es un nuevo parámetro potencialmente útil para mejorar la predicción del riesgo trombótico en pacientes con cáncer colorrectal localizado, así como una nueva diana cuya inhibición podría prevenir la metástasis y trombosis en dichos pacientes. En cáncer gástrico avanzado, mediante un "array" de expresión no dirigido, comparamos retrospectivamente la expresión génica en el tumor primario de 48 pacientes que sufrieron tromboembolismo venoso durante el seguimiento y 49 controles sin dicha complicación, apareados por "propensity score matching". Una vez identificados los genes asociados significativamente a la ocurrencia de tromboembolismo venoso, reclutamos una nueva cohorte de 44 pacientes con dicha comorbilidad y 39 controles. En el tumor primario, medimos por PCR digital la expresión absoluta de los genes previamente identificados, y comprobamos si esta seguía la misma tendencia respecto al tromboembolismo venoso que en la primera cohorte, así como su potencial para estratificar a aquellos pacientes de alto riesgo. Como resultados principales, en la primera cohorte, identificamos 15 genes cuya expresión diferencial se asociaba con la ocurrencia de tromboembolismo venoso, siendo además aplicables tanto al subtipo de cáncer gástrico intestinal como difuso. En la cohorte de validación, 3 de estos genes, EPS8, PRKD3 y SAA1, siguieron la misma tendencia de expresión respecto a la trombosis, y además, su expresión absoluta distinguió dos grupos de pacientes con un riesgo de tromboembolismo venoso significativamente diferente. En conclusión, identificamos 3 genes cuya expresión en cáncer gástrico avanzado podría mejorar la predicción del tromboembolismo venoso.Venous thromboembolism is a relatively common comorbidity in cancer patients and the second leading cause of death in this population. Different anticoagulant treatments have been used to prevent this complication, but their low efficacy in unselected populations and the increased risk of bleeding associated with this tretaments have led to the abandonment of widespread thromboprophylaxis. This scenario stands out in gastrointestinal tumours, as although they are highly thrombogenic, they are also associated with a high risk of bleeding. Therefore, it is necessary to identify new thrombotic risk markers that can help select those patients who could benefit from primary thromboprophylaxis, despite the risk of bleeding. Thus, the objective of this thesis is to identify new thrombotic biomarkers in patients with colorectal cancer and advanced gastric cancer. In colorectal cancer, we focused on the study of hepsin, a serine protease involved in tumour invasion and activation of the extrinsic coagulation pathway. Retrospectively, we recruited nearly 300 patients with localized or metastatic colorectal cancer. Using immunohistochemistry, we measured hepsin staining intensity in the primary biopsy at diagnosis and correlated it with clinical variables such as cumulative incidence of thrombosis, metastatic relapse, overall survival, among others. Subsequently, we conducted in vitro and in vivo studies using colorectal cancer cells stably transfected with the hepsin gene, and determined the effect of its overexpression in migration, invasion, proliferation and thrombin generation assays. Finally, with a view toward therapeutic targeting, we used virtual screening and molecular docking techniques to identify FDA-approved drugs that could inhibit hepsin activity and suppress the effects of its overexpression in tumour cells. As main results, we described hepsin as a biomarker of thrombosis and metastatic relapse in patients with localized colorectal cancer. In line with this, hepsin overexpression in colorectal cancer cells significantly increased their invasive phenotype and thrombin generation capacity. Finally, we identified Venetoclax and Suramin as two new hepsin inhibitors that also suppressed its pro-tumoural and pro-thrombogenic effects. In conclusion, hepsin expressed in the primary tumour is a new potentially useful parameter for improving thrombotic risk prediction in patients with localized colorectal cancer, as well as a new target whose inhibition could prevent metastasis and thrombosis in these patients. In advanced gastric cancer, using a non-targeted expression array, we retrospectively compared gene expression in the primary tumour of 48 patients who developed venous thromboembolism during follow-up and 49 controls without this complication, matched by propensity score matching. Once genes significantly associated with the occurrence of venous thromboembolism were identified, we recruited a new cohort of 44 patients with this comorbidity and 39 controls. In the primary tumour, we measured the absolute expression of the previously identified genes by digital PCR and checked whether they followed the same trend concerning venous thromboembolism as in the first cohort, as well as their potential for stratifying patients at high risk. As main results, in the first cohort, we identified 15 genes whose differential expression was associated with the occurrence of venous thromboembolism, applicable to both intestinal and diffuse gastric cancer subtypes. In the validation cohort, 3 of these genes, EPS8, PRKD3 and SAA1, followed the same expression trend with respect to thrombosis, and their absolute expression distinguished two groups of patients with a significantly different risk of venous thromboembolism. In conclusion, we identified 3 genes whose expression in advanced gastric cancer could improve the prediction of venous thromboembolism

Cancer-associated thrombosis: An overview of mechanisms, risk factors, and treatment

Cancer-associated thrombosis is a major cause of mortality in cancer patients, the most common type being venous thromboembolism (VTE). Several risk factors for developing VTE also coexist with cancer patients, such as chemotherapy and immobilisation, contributing to the increased risk cancer patients have of developing VTE compared with non-cancer patients. Cancer cells are capable of activating the coagulation cascade and other prothrombotic properties of host cells, and many anticancer treatments themselves are being described as additional mechanisms for promoting VTE. This review will give an overview of the main thrombotic complications in cancer patients and outline the risk factors for cancer patients developing cancer-associated thrombosis, focusing on VTE as it is the most common complication observed in cancer patients. The multiple mechanisms involved in cancer-associated thrombosis, including the role of anticancer drugs, and a brief outline of the current treatment for cancer-associated thrombosis will also be discussed

Study of the thrombogenicity induced by the cytotoxic treatment of malignant disease

Cancer and its treatment are frequently complicated by the development of venous thromboembolism (VTE). Interestingly, VTE incidence rates vary according to tumour type and the chemotherapy regimen administered. Yet, the precise mechanisms responsible for the increase in VTE in cancer patients remain unclear. Tissue factor (TF), the primary cellular initiator of the coagulation cascade, is over-expressed in many solid malignancies, particularly pancreatic cancer, and the number of circulating TF+ microparticles (MP) are increased in cancer patients. It is hypothesised that the increased risk of VTE in cancer patients generated by cytotoxic treatment may be partly attributed to the apoptotic process, involving the exposure of procoagulant phosphatidylserine (PS) on tumour cells or other chemotherapy damaged cells and increased release of TF+ MP into the circulation. The aim of this study was to explore how coagulation can be initiated in cancer cells and be potentiated by chemotherapy, with respects to the specific expression of TF and PS, and generation of MP.Flow cytometry was used to evaluate expression of cell surface TF and level of apoptosis in untreated/doxorubicin (Dox)-treated cancer cell lines, and number of MP in platelet-free plasma (PFP) from untreated pancreatic cancer patients, myeloproliferative disorder (MPD) patients, and multiple myeloma (MM) patients before, during, and after chemotherapy. The cell proliferation of untreated/Dox-treated cancer cells was assessed by an MTS assay. Procoagulant activity (PCA) of untreated/Dox-treated cancer cells and their isolated cell-free supernatants containing MP, and also MM and MPD patient PFP was measured using a prothrombin time assay. Enzyme-linked immunosorbent assay quantified levels of TF in unfiltered/0.1 μm filtered cell-free supernatants isolated from cancer cell lines, and serum levels of soluble cell adhesion molecules CD106 and CD54 from MM patients.Pancreatic (AsPC-1, CFPAC-1, MIA-PaCa-2), ovarian (A270, ES2, SKOV-3), colorectal (CaCo-2, LoVo), breast (MCF-7, MDA-MB-231, T47D), and haematological (JJN3, U937) cancer cells were found to support coagulation in a cell number-dependent manner, defined by a logarithmic relationship that was consistent across all cell lines. Furthermore, single cell clotting time (CT) was determined for each cancer cell line from the slope and y axis intercept of a logarithmically transformed data plot. A near linear relationship was observed between TF expression and single cell CT where a higher expression of TF results in a proportionally faster CT (P = 0.01). In addition, tumour cell-derived MP were shown to be procoagulant and the majority of procoagulant potential could be removed by passing isolated cell-free supernatants through a 0.1 μm filter. A dose-dependent CT was observed with AsPC-1, CFPAC-1, ES2, SKOV-3, LoVo, and MDA-MB-231 cell-free supernatants.The cytotoxic chemotherapeutic agent Dox was found to decrease the number of viable ovarian (ES2), breast (MDA-MB-231, T47D), and haematological (MM.1S, U937) cancer cells in a time- and dose-dependent manner, and cell death was shown to be induced by apoptosis and subsequently necrosis at higher drug concentrations. Cell surface expression of PS was found to increase following Dox treatment, while TF was not upregulated. Furthermore, Dox was shown to dose-dependently increase PCA in all cancer cells examined, although the effect of cell-free supernatants on PCA was less consistent; ES2, MDA-MB-231, and U937 cell-free supernatants isolated from Dox-treated cells demonstrated increased PCA at 0.01 and 0.1 μM concentrations. The Dox-induced increase in PCA of cancer cells and cell-free supernatants were found to correlate with tumour cell viability (r = 0.79 to 0.97, P < 0.01).Numbers of TF+ MP were significantly higher in 35 untreated pancreatic cancer patients in comparison with 15 MM patients prior to chemotherapy (P < 0.0005). Furthermore, numbers of endothelial cell-derived MP (EMP), monocyte-derived MP, plasma cell-derived MP, and PS+ MP, were significantly higher in 6 MPD patients in comparison with 15 MM patients prior to chemotherapy (P < 0.005), but not levels of platelet-derived MP (PMP) or TF+ MP. Markers of endothelial dysfunction, including EMP and soluble cell adhesion molecules CD106 and CD54, were elevated after thalidomide (Thal)- or lenalidomide (Len)-based therapies in MM patients. Furthermore, PCA was significantly increased in MM patients after treatment (P = 0.007), and also levels of PMP, plasma cell-derived MP, and PS+ MP (P < 0.05), but not monocyte-derived MP (P = 0.33) and TF+ MP (P = 0.41).In summary, this study shows that across a range of tumour sites a consistent relationship is seen between procoagulant potential and both cell number and TF cell surface expression. Dox can increase PCA of cancer cells through reduced cell viability that leads to PS exposure. Importantly, this Dox-induced procoagulant mechanism was not found to involve an upregulation of surface TF antigen on ovarian, breast, or haematological cancer cells. In MM patients treated with Thal- or Len-based therapies increased PCA was observed, which may be mediated by endothelial dysfunction and increased generation of MP

Cancer-Associated Ischemic Stroke: The Bergen NORSTROKE Study

Background: Stroke and cancer are both leading causes of death and disability in the Western world. Cancer can lead to a hypercoagulable state that can cause ischemic stroke. Mechanisms include disturbance of the coagulation cascade, tumor substance secretion, infections and non-bacterial endocarditis. Many types of cancer also share a similar risk factor profile to that of ischemic stroke. The cancer-stroke connection has not previously been studied in Norway. Further knowledge on which subgroups of patients are at highest risk for both stroke and cancer, as well as on how to recognize a cancer-associated stroke is needed. Methods and materials: The studies 1-3 are the foundation of the present thesis. All papers have used data from The Norwegian Stroke Research Registry (NORSTROKE). NORSTROKE is a comprehensive, prospective registry that since February 2006 has included all stroke patients admitted to the Stroke Unit at Haukeland University Hospital. For the present studies, the medical charts of all stroke patients were reviewed for collection of cancer diagnoses. Cancer data was quality assured through linking NORSTROKE to The Cancer Registry of Norway. Results: From August 2006 through August 2011, 1511 patients with ischemic stroke were registered in NORSTROKE. In study I, patients with cancer post stroke were excluded. In study I, a total of 1456 patients were included, of whom 229 (15.7 %) had cancer prior to index stroke. The prevalence of stroke was higher in stroke patients compared to the general population below the age of 70. In study II, patients with cancer pre stroke were excluded. In study II, a total of 1282 patients were included, of whom 55 (4.3 %) were diagnosed with cancer post index stroke. In study III, the inclusion period was extended and patients with inactive cancer were excluded. In study III, 1646 patients were included in the study, of whom 82 (5.0 %) had active cancer. The most common cancer types overall were cancer of the; colon, prostate, breast, lung, bladder, uterus and ovaries as well as of unknown primary site. Patients with active cancer had similar risk factors to patients without a history of cancer. Active cancer in stroke patients was associated with smoking, age, increased D-dimer and decreased hemoglobin. Active cancer was also associated with stroke of unknown etiology. Conclusions: The prevalence of cancer was higher in stroke patients compared to the general population below the age of 70. Findings in patients with cancer-associated ischemic stroke were elevated D-dimer, signalling hypercoagulation, lower hemoglobin and a history of smoking. These factors may also be used to predict active cancer in stroke patients and thus indicate which stroke patients could be screened for underlying cancer

Promotion Mechanisms of Stromal Cell-Mediated Lung Cancer Development Within Tumor Microenvironment

Siyu Wu,1,&ast; Yumeng Hu,1,&ast; Bowen Sui2 1Heilongjiang University of Chinese Medicine, Harbin, People’s Republic of China; 2First Affiliated Hospital, Heilongjiang University of Chinese Medicine, Harbin, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Bowen Sui, Email [email protected]: Lung cancer, with its high incidence and mortality rates, has garnered significant attention in the medical community. The tumor microenvironment (TME), composed of tumor cells, stromal cells, extracellular matrix, surrounding blood vessels, and other signaling molecules, plays a pivotal role in the development of lung cancer. Stromal cells within the TME hold potential as therapeutic targets for lung cancer treatment. However, the precise and comprehensive mechanisms by which stromal cells contribute to lung cancer progression have not been fully elucidated. This review aims to explore the mechanisms through which stromal cells in the tumor microenvironment promote lung cancer development, with a particular focus on how immune cells, tumor-associated fibroblasts, and endothelial cells contribute to immune suppression, inflammation, and angiogenesis. The goal is to provide new insights and potential strategies for the diagnosis and treatment of lung cancer.Keywords: tumor microenvironment, lung cancer, stromal cells, immune cells, cancer-associated fibroblasts, endothelial cell