Trousseau’s Syndrome Revisited: Tissue Factor-Bearing Microparticles in Pancreatic Cancer.

Abstract

Thromboembolic disease is a major complication of malignant disease, and pancreatic cancer ranks among the malignancies associated with the highest rates of thrombosis. Using laser injury in a mouse model of thrombosis, we previously demonstrated that tissue factor-bearing microparticles circulating in normal plasma accumulate in the developing thrombus through an interaction mediated by P-selectin and PSGL-1. Microparticle tissue factor contributes significantly to fibrin propagation within the thrombus. Based upon this observation, we evaluated the hypothesis that elevated numbers of tissue factor-bearing microparticles might be one of the causes of cancer-associated thrombosis. Microparticles are cell-derived vesicular structures under 1000 nm in diameter. Since the light scattering methodology used in commercial flow cytometers cannot determine particle size when the diameter of the particle (200–1000 nm) is the same order of magnitude as the wavelength of the incident light employed (488 nm), we applied a novel instrumentation to determine the size, size distribution, and concentration of tissue factor-bearing microparticles. An NPE Systems Quanta flow cytometer, using impedance to measure particle size, was extensively modified for this microparticle application. Application of 780 nm fluorescent calibration beads yielded a major population from 740 to 820 nm. Application of 520 nm fluorescent beads yielded a major population from 500 to 540 nm. Using a high affinity antibody to tissue factor, tissue factor-bearing microparticles were measured in platelet-poor plasma derived from normal subjects and subjects with surgically unresectable or metastatic pancreatic cancer. Tissue factor bearing microparticles were detected in significantly more subjects with pancreatic cancer (12 of 17, 70%) than healthy individuals in whom tissue factor-bearing microparticles remained below the level of detection in all but a single individual (1 of 11, P=0.005). By comparison, tissue factor-bearing microparticles were not detected in subjects with advanced stage (III or IV) non-small cell lung cancer (N=9). In subjects with advanced pancreatic cancer, the mean concentration of tissue factor bearing microparticles was 444,000 particles/microliter with a mean diameter ranging between 371 nm to 727 nm. The two patients with pancreatic cancer in this study who suffered a recent lower extremity venous thrombosis had high measurable tissue factor-bearing microparticles levels with a mean concentration of 975,000 particles/microliter. These results indicate that tissue factor-bearing microparticles are present in the plasma of many patients with advanced pancreatic cancer and suggest that tissue factor-bearing microparticles may be central to the pathogenesis of cancer-associated thrombosis.

Platelets and Metastasis: New Implications of an Old Interplay

“…MPs are found in the blood of cancer patients. Patients with pancreatic, colorectal, brain, prostate, and breast cancer have higher levels of plasma TF/PS-expressing MPs and higher MP-associated pro-thrombotic activity than healthy subjects, especially during advanced stages of disease and after chemotherapy or radiotherapy (104)(105)(106)(107)(108)(109)(110). Metastatic cancer patients had particularly high plasma levels of TF + MPs across a range of cancer types (109).…”

Tissue Factor–Bearing Microparticles and Cancer

“…35 Notably, the reported number of TF-bearing microparticles observed by light scatter flow cytometry was 10,000-fold less than what we have observed using an impedance-based flow cytometer. 36 The in vitro measurement of microparticles expressing TF is usually performed either indirectly by monitoring the TF-dependent generation of thrombin or factor Xa using a chromogenic substrate or directly by flow cytometric analysis. Whereas standard light scatter flow cytometry is widely available and allows for accurate cell identification through surface antigen immunophenotyping and sizing, its use in sizing microparticles is subject to limitations.…”

“…2 illustrates the detection of TF-bearing microparticles in cell-free media conditioned with ASPC-1 pancreatic cancer cells. Using the impedance-based flow cytometer, we can also detect TF-bearing microparticles in the plasma of patients with cancer 36 and are currently conducting a case-control study in cancer patients who present with an acute venous thromboembolic event. We hypothesize that these circulating microparticles play a pathogenic role in the development of thrombosis in cancer.…”