Heparin Inhibition of Endothelial Cell Proliferation and Organization Is Dependent on Molecular Weight

Abstract

Objective-Studies have shown improved survival in cancer patients treated with low molecular weight heparins (LMWHs). Tumors depend on an expanding vasculature, and heparins may affect vessel growth and function. We investigated the effect of heparins differing in M r on selected endothelial cell properties. Methods and Results-Human umbilical vein endothelial cells were cultured with fibroblast growth factor-2 and heparins differing in M r . Cell proliferation was assessed by [ 3 H]thymidine incorporation, and vascular organization was assessed by in vitro assays. Maximum inhibition of 94Ϯ2% was observed with 6-kDa LMWH, greater than the inhibition seen with unfractionated heparin (58Ϯ8%) or 3-kDa LMWH (60Ϯ9%, Pϭ0.02 for both). No inhibition of proliferation was observed with heparin tetrasaccharide, octasaccharide, or pentasaccharide (fondaparinux). Three-and 6-kDa fractions decreased endothelial tube formation in Matrigel to 58Ϯ15% and 67Ϯ9% (PϽ0.05), respectively, of that with fibroblast growth factor-2, whereas no inhibition was observed with unfractionated heparin, tetrasaccharide, pentasaccharide, or octasaccharide. LMWH (6 kDa) also inhibited vessel formation in a placental explant. Conclusions-Heparin inhibition of endothelial cell proliferation and organization requires a chain length of Ͼ8 saccharide units, with maximal inhibition at M r of 6 kDa. This M r dependence differs from that required for anticoagulant activity. Key Words: low molecular weight heparins Ⅲ cancer Ⅲ angiogenesis Ⅲ endothelium Ⅲ thrombosis U nfractionated heparin is a heterogeneous mixture of polysaccharide molecules with a mean M r between 12 and 15 kDa containing glycosaminoglycan chains from 200 to 300 saccharide units. 1 Low molecular weight heparins (LMWHs) are fragments of unfractionated heparin produced by controlled enzymatic or chemical depolymerization with lower mean M r between 3 and 6 kDa and chain lengths of 12 to 18 saccharide units. 2 These structural differences result in important functional and pharmacokinetic differences, inasmuch as LMWHs have an increased anti-Xa/anti-IIa activity ratio, reduced plasma protein binding, decreased interaction with platelets and platelet factor 4, a prolonged half-life, and increased bioavailability after subcutaneous administration. 2,3 These changes give LMWHs an improved therapeutic profile compared with unfractionated heparin and have led to their increasing use in the treatment of venous and arterial thrombosis. See page 1954Individual studies 4,5 and meta-analyses 6,7 have found significant improvements in 3-and 6-month mortality in patients with cancer who were treated with LMWHs compared with unfractionated heparin. This survival advantage was found in subgroups differing in age, sex, and primary site of malignancy and could not be attributed to differences in thrombosis or bleeding complications. 8 The mechanism of this effect remains unknown but appears to be independent of the anticoagulant action of LMWH and may be related to effects on the vasculature. Prior studie...

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