The inhibition of pancreatic cancer invasion-metastasis cascade in both cellular signal and blood coagulation cascade of tissue factor by its neutralisation antibody
Introduction
In cancer invasion and metastasis, the cancer cells degrade the basement membrane and intravasate into lymphatic or blood microvessels. The cells are then transported to a new location and become clogged within the microvessels, proceeding to grow following extravasation.1 These steps include cancer cell invasion, degradation of the basement membrane and stromal extracellular matrix (ECM), and formation of microthrombi. The matrix metalloproteinase (MMP) family represents important enzymes that degrade ECM and facilitate tumour invasion.2 Amongst them, MMP-9 is well-known as one of the most important factors in facilitating invasion and metastasis in pancreatic cancer.3
Tissue factor (TF), the initiating cell surface receptor for the coagulation cascade, activates factor VIIa. The TF-VIIa complex activates factor X, and consequently this protease cascade forms fibrin clots.4, 5 The relationship between cancer and blood coagulation was initially described by the French surgeon Trousseau.6 Cancer patients, especially those with pancreatic, stomach, and glioma cancer, often suffer from a state of hypercoagulation and venous thrombosis, leading to patient morbidity and mortality.7, 8, 9 In another study using a fibrinogen-deficient transgenic mouse model, fibrinogen appeared to be an important element of the metastatic potential of circulating tumour cells.10 Meanwhile, TF plays an important role in not only blood coagulation but also cell signalling in which the TF-VIIa complex phosphorylates extracellular-regulated kinase 1/2 (ERK1/2) via protease-activated receptor-2 (PAR-2).11 Moreover, its complex promotes the expression of interleukin-8 (IL-8) and invasion in breast cancer cell lines.12 However, the concrete involvement of TF in tumour invasion-metastasis has not yet been fully evaluated.
Human pancreatic cancer has one of the worst prognoses amongst cancers.13 Invasion and metastasis advancing beyond the pancreas are typical. Direct invasion to nearby organs, such as the stomach, duodenum, colon, spleen and kidney frequently occurs. Distant metastasis to the liver and peritoneal dissemination are also commonly seen.14, 15 In terms of the relationship between TF and pancreatic cancer, TF expression is an important early event in malignant transformation of the pancreas.16 TF expression may contribute to the aggressiveness of pancreatic cancer that would stimulate tumour invasiveness, and evaluation of the primary tumour for TF expression may identify patients with a poor prognosis.17, 18
Therefore, elucidation of the relationship between TF and pancreatic cancer invasion-metastasis may lead to the development of new therapeutic strategies as well as a better understanding of pancreatic cancer biology.
Section snippets
Cell lines
Human pancreatic cancer cell lines BxPC3, Panc1, Capan1, and MIA PaCa-2 were purchased from the American Type Culture Collection (Rockville, MD, USA). The cell lines were maintained in Dulbecco’s Modified Eagle’s Medium supplemented with 10% faetal bovine serum (FBS) (Cell Culture Technologies, Gaggenau-Hoerden, Germany), 100 units/mL streptomycin, and 2 mmol/L L-glutamine (Sigma, St. Louis, MO, USA) in an atmosphere of 5% CO2 at 37 °C.
Immunocytochemistry
Cells (1 × 105) were seeded on a 4-well culture slide (BD
TF-positive human pancreatic cancer cells, BxPC3, enhance invasion potential in vitro and in vivo
We used the BxPC3 cell line in a series of our experiments because real-time PCR analysis and immunostaining of TF showed that BxPC3 strongly expressed TF amongst the four pancreatic cancer cell lines (Fig. 1A). Then, we examined the status of TF expression for BxPC3 in vitro and in vivo. Immunostaining revealed TF expression specifically in cancer cells contacting with stromal tissues, namely the invasive front in vivo, although TF expression was uniformly observed in all cells in vitro (Fig. 1
Discussion
Pancreatic cancer is the most refractory neoplasm and possesses several clinicopathological characteristics. First, pancreatic cancer exerts extensive invasion and metastasis to other organs.14, 15 Second, thrombosis occurs most frequently in patients with pancreatic cancer.7 Third, expression of TF may contribute to the aggressiveness of pancreatic cancer that stimulates tumour invasiveness, and evaluation of the primary tumour for TF expression may identify patients with a poor prognosis.17,
Grant support
This work was supported partly by a Grant-in-Aid from the Third Term Comprehensive Control Research for Cancer, the Ministry of Health, Labor and Welfare (Matsumura, H19-025), Scientific Research on Priority Areas from the Ministry of Education, Culture, Sports, Science and Technology (Matsumura, 17016087), and the Japanese Foundation for Multidisciplinary Treatment of Cancer (Matsumura), Funding Program for World-Leading Innovative R&D on Science and Technology (FIRST Program) and the Princess
Conflict of interest statement
None declared.
Acknowledgements
We thank Dr. Miyawaki (RIKEN) for Venus. We thank N. Mie and M. Ohtsu for their technical assistance, and K. Shiina for her secretarial assistance.
References (28)
- et al.
Matrix metalloproteinases: molecular aspects of their roles in tumour invasion and metastasis
Eur J Cancer
(2000) Trousseau’s syndrome: multiple definitions and multiple mechanisms
Blood
(2007)- et al.
Incidence of venous thromboembolism in patients hospitalized with cancer
Am J Med
(2006) - et al.
Fibrinogen is an important determinant of the metastatic potential of circulating tumor cells
Blood
(2000) - et al.
Tissue factor-factor VIIa-specific up-regulation of IL-8 expression in MDA-MB-231 cells is mediated by PAR-2 and results in increased cell migration
Blood
(2004) - et al.
Antitumour activity of NK012, SN-38-incorporating polymeric micelles, in hypovascular orthotopic pancreatic tumour
Eur J Cancer
(2010) - et al.
Proteinase-activated receptor-2-mediated matrix metalloproteinase-9 release from airway epithelial cells
J Allergy Clin Immunol
(2000) - et al.
Platelets and fibrin(ogen) increase metastatic potential by impeding natural killer cell-mediated elimination of tumor cells
Blood
(2005) The pathogenesis of cancer metastasis: the ‘seed and soil’ hypothesis revisited
Nat Rev Cancer
(2003)- et al.
Histologic features of venous invasion, expression of vascular endothelial growth factor and matrix metalloproteinase-2 and matrix metalloproteinase-9, and the relation with liver metastasis in pancreatic cancer
Pancreas
(2002)