Binding between heparin and the integrin VLA-4

Abstract

SummaryHeparin possesses antimetastatic effects that were related to various molecular mechanisms beyond anticoagulant activities. The ability of heparin to interfere with the function of adhesion receptors in the metastatic course appears as a promising therapeutic approach. This refers to numerous findings that heparin attenuates metastasis in a selectin-dependent manner. We recently demonstrated that heparin interferes with the integrin VLA-4 on murine melanoma cells binding to VCAM-1. To confirm this activity and to obtain further insight into molecular recognition of heparin by VLA-4, we investigated the inhibition of VLA-4 mediated binding of human melanoma MV3 cells to immobilised VCAM-1 by different heparins. The size of heparin has an important impact on inhibition. Unfractionated heparin (UFH) and tinzaparin, a low-molecular-weight heparin (LMWH) representing a mean of about 18-20 monomers, displayed high Keywords Heparin, melanoma cell adhesion, VLA-4, antimetastatic activity inhibitory activity. Fractionating tinzaparin to 14-18 monomers reduced inhibition slightly, while the pentasaccharide fondaparinux was without effects. To confirm molecular recognition of tinzaparin by VLA-4, a surface acoustic wave-biosensor was applied. A VLA-4 containing membrane preparation of MV3 cells was immobilised at the sensors to allow for detection of kinetic binding constants of tinzaparin compared to VCAM-1. Tinzaparin binds to VLA-4 with affinity in the low micromolar range (4.61×10 -6 M), which clearly indicates specific molecular recognition. Furthermore, tinzaparin displays a nearly identical k off compared to VCAM-1 (5.13×10 -3 s -1 versus 3.44×10 -3 s -1) which is evident for interference with the ligand binding. The data provide evidence for a direct confirmation of heparin binding to VLA-4 and thus, contribute to understand the antimetastatic activity of heparin.

Heparin plays a key regulatory role via a p53/FAK‐dependent signaling in melanoma cell adhesion and migration

“…In fact, previous studies, performed with radioactive or fluorescent heparin derivatives showed that these compounds were bound and internalized by SMC (20), human endothelial cells (21) and rat neurons (33). Other reports show that heparin binds to specific cell surface adhesion molecules, for example, integrins to modulate the adhesive capabilities of different cell lines (34,35). The present study clearly demonstrates that M5 melanoma cells uptake heparin which results in its cytoplasmic and nuclear localization.…”

Heparin and its derivatives - Present and future

“…Such an activity is reported by Schlesinger et al (7) who demonstrate that the low-molecular-weight heparin (LMWH) tinzaparin interferes with integrin α4β1/VLA-4 binding to its counter-receptor vascular cell adhesion molecule-1 (VCAM-1). They used an acoustic wave biosensor to analyse these interactions.…”

The role of VLA-4 binding for experimental melanoma metastasis and its inhibition by heparin

“…Although we described numerous structure-activity relationships of heparin-VLA-4 binding [36] the relevance of this interaction has never been addressed in vivo. Here we show that VLA-4 on murine melanoma cell line B16F10 contributes dominantly to its experimental metastasis.…”

“…The interaction of B16F10 and B16F10-VLA-4kd cells with the ligand VCAM-1 was investigated under physiological flow conditions in a parallel plate flow chamber as described before [35,36]. Briefly, VCAM-1 Fc chimera (0.2 μg) was coated on glass slides which were incorporated into a parallel plate flow chamber and mounted on an inverted microscope.…”