Chemical modifications of heparin that diminish its anticoagulant but preserve its heparanase-inhibitory, angiostatic, anti-tumor and anti-metastatic properties

Structural features of heparin potentially important for heparanase-inhibitoryactivity were examined by measuring the ability of heparin derivativesto affect the degradation of [³H]acetylated heparan sulphateby tumor cell heparanases. IC₅₀ values were determined usingan assay which distinguished degraded from undegraded substrateby precipitation of the latter with cetylpyridinium chloride(CPC). Removal of heparin's 2-O-sulphate and 3-O-sul-phate groupsenhanced heparanase-inhibitory activity (50%). Removal of itscarboxyl groups slightly lowered the activity (18%), while combiningthe treatments abolished the activity. At least one negativecharge on the iduronic acid/idose moiety, therefore, is necessaryfor heparanase-inhibitory activity. Replacing heparin's N-sulphategroups with N-acetyl groups reduced its activity (37%). Comparingthis heparin derivative with 2,3-O-de-sulphated heparin, theplacement of sulphate groups appears important for activitysince the two structures have similar nominal linear chargedensity. In addition, unsubstituted uronic acids are nonessentialfor inhibition since their modification (periodate-oxidation/borohydride-reduction)enhanced rather than reduced heparanase-inhibitory activity.The most effective heparanase inhibitors (2,3-O-desulphatedheparin, and [periodate-oxidized, borohydride-reduced] heparin)were tested in the chick chorioallantoic membrane (CAM) bioassayfor anti-angiogenic activity and found to be at least as efficaciousas heparin. 2,3-O-desulphated heparin also significantly decreasedthe tumor growth of a subcutaneous human pancreatic (Ca-Pan-2)adenocarcinoma in nude mice and prolonged the survival timesof C57BL/6N mice in a B16-F10 melanoma experimental lung metastasisassay. angiogenesis chemically-modified heparins endoglycosidase hepara sulphate cancer