Down-regulation of Monocyte Tissue Factor Mediated by Tissue Factor Pathway Inhibitor and the Low Density Lipoprotein Receptor-related Protein

Abstract

Inflammatory mediators like bacterial lipopolysaccharide induce monocytes to express tissue factor (TF), the cell-surface protein that triggers the blood clotting cascade in hemostasis and thrombotic disease. The physiologic ligand for TF is the serine protease, factor VIIa (FVIIa), and the resulting bimolecular enzyme, TF/ FVIIa, can be reversibly inhibited by tissue factor pathway inhibitor (TFPI). Culturing monocytic cells in the presence of both FVIIa and TFPI caused down-regulation of TF expression via reducing its half-life. To exert this effect, FVIIa had to be competent to bind both TF and TFPI, and TFPI had to contain the C-terminal domain required for binding to other cell-surface receptors, including the low density lipoprotein receptor-related protein (LRP). TF down-regulation by FVIIa plus TFPI was abrogated by the 39-kDa receptor-associated protein, which blocks binding of all known ligands to LRP. Furthermore, treatment with FVIIa plus TFPI caused monocyte TF to colocalize with ␣-adaptin, a component of clathrin-coated pits. Thus, in addition to reversibly inhibiting TF/FVIIa catalytic activity, TFPI also mediates the permanent down-regulation of cell-surface TF in monocytic cells via LRP-dependent internalization and degradation. This represents an unusual mechanism for receptor internalization, requiring ligand-dependent bridging of one cell-surface receptor (TF) to a second cell-surface receptor (LRP), the latter being capable of clathrin-mediated internalization.

High Turnover of Tissue Factor Enables Efficient Intracellular Delivery of Antibody–Drug Conjugates

“…Previous publications also indicate that TF has a higher turnover rate compared with EGFR and HER2. Hamik and colleagues demonstrated that the halflife of TF on monocytes was 3.7 hours, which could be reduced to 1.3 hours when TF was bound by TF protein inhibitor and FVII (38). Unstimulated EGFR and HER2 on the other hand have a half-life of 6 to 24 hours depending on the cell line used (39).…”

Tissue factor and tissue factor pathway inhibitor

“…TFPI does not cleave readily, and prevents the complex from engaging other molecules [5]. TFPI also causes monocytes to internalise and degrade TF-FVIIa complexes on the cell surface [43]. Circulating TFPIFxa-TF-FVIIa complexes are metabolised by the liver [35].…”

Factor VIIa Interaction With Tissue Factor and Endothelial Cell Protein C Receptor on Cell Surfaces

“…FVIIa bound to TF is internalized in many cell types (e.g. fibroblasts, monocytes, and baby hamster kidney [BHK] cells) (14)(15)(16). The majority of internalized FVIIa enters the endocytotic pathway and gets degraded but a small portion of it is recycled back to the cell surface as intact protein (14).…”

“…Although both FVIIa and active siteinhibited FVIIa (ASIS) bind to TF, there seems to be subtle differences between them in their internalization and recycling patterns (17). Interestingly, despite the continuous internalization of FVIIa via TF, TF levels on the cell surface remain relatively constant (14)(15)(16). It may be important to note that the rate of TF-mediated FVIIa endocytosis is not as rapid as one would expect for classical receptor-mediated endocytosis.…”

“…It may be important to note that the rate of TF-mediated FVIIa endocytosis is not as rapid as one would expect for classical receptor-mediated endocytosis. However, formation of the ternary complex with tissue factor pathway inhibitor (TFPI)-FXa significantly increases the rate of FVIIa internalization in fibroblasts (14) and down-regulates TF levels at the cell surface in both fibroblasts (14) and activated monocytes (15). Antibodies against low density lipoprotein receptor-related protein (LRP) or the receptor-associated protein, which block LRP-dependent internalization, reduce TFPI-mediated FVIIa internalization but not the FVIIa internalization observed in the absence of TFPI (14).…”