Ged Byrne

Plasma levels of soluble CD105 correlate with metastasis in patients with breast cancer

CD105 (endoglin), a receptor for transforming growth factor (TGF) β1 and β3 in vascular endothelial cells, is highly up‐regulated in blood vessels of tissues where neovascularisation occurs. It modulates endothelial‐mesenchymal signalling and is essential for angiogenesis. Indeed, CD105 knock‐out mice die from malvascularisation by 11.5 day p.c. In the present study CD105, TGFβ1 and CD105/TGFβ1 complexes were quantified in plasma samples from 77 healthy individuals and 92 patients with early stage breast cancer prior to any treatment. When compared with normal controls, both CD105 and CD105/TGFβ1 complex levels were significantly elevated in breast cancer patients, whereas TGFβ1 levels were lower in cancer patients. The most important finding to emerge was that CD105 levels were significantly increased in patients who developed distant metastasis compared with disease‐free patients. While there was no significant difference between CD105 levels in controls compared to disease‐free patients, it was significantly higher in patients with metastatic disease. Thus patients who had died following local relapse or distant metastases possessed the highest levels of CD105. Neither CD105/TGFβ1 complex nor TGFβ1 levels correlated with tumour progression. Our data indicate that CD105 might be a valuable novel angiogenic marker for identifying breast cancer patients who are at high risk of developing metastasis. Int. J. Cancer (Pred. Oncol.) 89:122–126, 2000.

Early changes in the haemostatic and procoagulant systems after chemotherapy for breast cancer.

Venous thromboembolism (VTE) following breast cancer chemotherapy is common. Chemotherapy-induced alterations in markers of haemostasis occur during chemotherapy. It is unclear how rapidly this occurs, whether this is upregulated in patients developing VTE and whether changes predict for VTE. Markers of haemostasis, functional clotting assays and vascular endothelial growth factor were measured before chemotherapy and at 24 h, 4 days, 8 days and 3 months following commencement of chemotherapy in early and advanced breast cancer patients and in age- and sex-matched controls. Duplex ultrasound imaging was performed after 1 month or if symptomatic. Of 123 patients, 9.8% developed VTE within 3 months. Activated partial thromboplastin time (APTT), prothrombin time (PT), D-dimer, fibrinogen, platelet count, VEGF and fibrinogen were increased in cancer. Fibrinogen, D-dimer, VEGF and tissue factor were increased, at baseline, in patients subsequently developing VTE. D-dimer of less than 500 ng ml−1 has a negative predictive value of 97%. Activated partial thromboplastin time, PT and thrombin–antithrombin showed significantly different trends, as early as within 24 h, in response to chemotherapy in patients subsequently developing VTE. Markers of coagulation and procoagulants are increased, before chemotherapy, in patients who subsequently develop VTE. A group of patients at minimal risk of VTE can be identified, allowing targeted thrombopropylaxis to the higher risk group.

Surrogate markers of tumoral angiogenesis.

Background Angiogenesis is a prerequisite for tumor growth and metastasis. Vascular cell adhesion molecule-1 (VCAM-1) is expressed on endothelial cells as a result of vascular endothelial growth factor (VEGF) stimulation. Purpose To determine if measurement in serum of VEGF or VCAM-1 provides an accurate measure of tumor angiogenesis. Methods VCAM-1 and VEGF were measured in the serum of women with early and advanced breast cancer by ELISA. Levels were compared to levels of VCAM-1 and VEGF in women with normal breasts and levels of the endothelial glycoprotein von Willebrand factor. Levels of VEGF and VCAM-1 in women with early breast cancer were correlated with established clinicopathological prognostic markers and intratumoral microvessel density (IMD). Results In early breast cancer serum VCAM-1 correlated closely with the microvessel density in tumors (r=0.61, p<0.001). Women with lymph node-positive and high-grade tumors had higher levels of serum VCAM-1 than women with lymph node-negative and low-grade tumors. Serum VEGF demonstrated no correlation with established prognostic features or IMD. Levels of VCAM-1 and VEGF were raised in women with advanced breast cancer. Conclusion Serum VCAM-1 is a surrogate marker of angiogenesis in breast cancer and its measurement may help in the assessment of antiangiogenic drugs currently in phase II trials.

Prevention of venous thromboembolism in surgical patients with breast cancer

No randomized trial has yet studied venous thromboembolism (VTE) prophylaxis in patients undergoing surgery for breast cancer.

Radioimmunoassay for the measurement of thrombospondin in plasma and breast cyst fluid: Validation and clinical application

Thrombospondin is an adhesive protein that has been implicated in malignancy, specifically in tumour progression and angiogenesis. We developed a radioimmunoassay for the measurement of thrombospondin in plasma and breast cyst fluid. The assay exhibited high accuracy, with recoveries of 102-136% and acceptable imprecision, with an intra-assay coefficient of variation (CV) of < 7·5% across the analytical range 30-1000 ng/mL and inter-assay CV of 4·4% and 7·7% at 152 and 224 ng/mL, respectively. Thrombospondin measured in the breast cyst fluid of patients with gross cystic disease of the breast showed that patients with type II (Na+) cysts had significantly higher concentrations than type I (K+) cysts. The plasma thrombospondin reference range was determined as 131-274 ng/mL. Patients with breast cancer had significantly higher plasma thrombospondin concentrations than normal individuals or patients with benign breast disease. Plasma thrombospondin was higher in breast cancer patients with lymph node involvement.

Investigation of Proposed Mechanisms of Chemotherapy-Induced Venous Thromboembolism Endothelial Cell Activation and Procoagulant Release Due to Apoptosis

Venous thromboembolism (VTE) during chemotherapy is common, with 7% mortality in metastatic breast cancer (MBC). In a prospective cohort study of patients with breast cancer, we investigated whether vascular endothelial cell activation (VECA), and whether apoptosis, is the cause of chemotherapy-induced VTE. Methods: Serum markers of VECA, E-selectin (E-sel), vascular cell adhesion molecule 1 (VCAM-1) and d-dimer (fibrin degradation and hypercoagulability marker) were measured prechemotherapy and at 1, 4, and 8 days following chemotherapy. Clinical deep vein thrombosis (DVT) or pulmonary embolism and occult DVT detected by duplex ultrasound imaging were recorded as VTE-positive (VTE+). In patients with MBC, hypercoagulable response to chemotherapy was compared between patients with and without cancer progression. Development of VTE and cancer progression was assessed 3 months following starting chemotherapy. Results: Of the 134 patients, 10 (7.5%) developed VTE (6 [17%] of 36 MBC receiving palliation, 0 of 11 receiving neoadjuvant to downsize tumor, and 4 [5%] of 87 early breast cancer receiving adjuvant chemotherapy, P = .06). Levels of E-sel and VCAM-1 decreased in response to chemotherapy (P < .001) in both VTE+ and patients not developing VTE (VTE−). However, decrease in VECA markers was similar in VTE+ and VTE− patients, implying this is not the cause of VTE. In patients with MBC following chemotherapy, d-dimer (geometric mean) increased by 36% in the 21 patients with MBC responding to chemotherapy but steadily decreased by 11% in the 15 who progressed (day 4, P < .01), implying patients with tumor response (apoptosis) had an early hypercoagulable response. Conclusions: During chemotherapy for breast cancer, VECA is induced; however, this is not the primary mechanism for VTE. Chemotherapy-induced apoptosis may enhance hypercoagulability and initiate VTE.

Current UK practice of thromboprophylaxis for breast surgery

Perioperative thromboprophylaxis is routine in most surgical specialties. A meta-analysis of 31 trials of lowdose heparin prophylaxis in patients undergoing general surgery in 1988 reported an overall incidence of deep vein thrombosis (DVT) diagnosed by 125I-radiolabelled fibrinogen uptake of 27·0 per cent. A clinical diagnosis of DVT was made in only 3·0 per cent1. A more recent meta-analysis of eight randomized trials in general surgery comparing low molecular weight heparin or unfractionated heparin with placebo or no treatment confirmed a 72 per cent reduction in asymptomatic DVT and a 75 per cent reduction in clinical pulmonary embolism2. There are no published clinical trials on thromboprophylaxis in breast surgery. Theoretically, patients who have surgery lasting over 1 h, who require general anaesthesia and who may have cancer-induced hypercoagulability are at risk of venous thromboembolism (VTE). Many surgeons avoid anticoagulants because of the perceived risk of wound haematoma, a complication that occurs in up to 6 per cent of women undergoing breast-conserving surgery even in the absence of anticoagulants3. This study explored current policies for thromboprophylaxis among breast surgeons in the UK.

A Scheme for the Development and Validation of Enzyme Linked Immunosorbent Assays (ELISA) for Measurement of Angiogenic Biomarkers in Human Blood

In this chapter the authors describe a protocol that can be applied to design and validate an ELISA technique using commercially available reagents. This often proves an economical alternative to purchasing off-the-shelf-kits. In addition, this protocol allows custom validation of the ELISA to a specific purpose. We have illustrated our protocol using VCAM as an example, however the principles and techniques described can equally be applied to all ELISA based techniques.

Serum soluble VCAM: A surrogate marker of angiogenesis

Purpose: Angiogenesis is essential for tumour growth and metastasis. Vascular cell adhesion molecule-l (VCAM) and endothelial-selectin (ESEL) are expressed on activated endothelial cells and we hypothesized that their measurement in serum would be an accurate measurement of tumour angiogenssis Methods: Preoperative serurn levels of VCAM, ESEL and VWF (another EC marker) were measured by enzyme-linked immunosorbent assay (ELISA) in 93 women with early breast cancer and levels were correlated with histological prognostic features and the microvessel density in each tumour (assessed by CD31 imrnunostaining). Sequential serum samples were taken from 55 women with advanced breast cancer, immediately prior to a change in hormonal therapy and 3 months later. Changes in serum VCAM, ESEL and VWF were c:ompared to the response of the disease assessed by UICC criteria at 6 months. Results: In early breast cancer serum VCAM, not ESEL or VWF, correlated with the microvessel density (r = 0.66, p c 0.001) in each tumour whilst in advanced disease serum VCAM levels, not ESEL or VWF, rose in those women whose disease progressed (p < 0.001) but levels remained unchanged or fell in those women whose disease remained stable or showed a partial response to therapy. Conclusion: Serum VCAM is a surrogate marker of angiogenesis in breast cancer and its measurement may help in the assessment of antiangiogenic drugs currently in phase II trials.