Anticoagulant and antiprotease profiles of a novel natural heparinomimetic mannopentaose phosphate sulfate (PI-88).
Heparinomimetic mannopentaose phosphate sulfate (PI-88) (Progen Industries Ltd. Brisbane, Australia), currently developed as an anticoagulant and antiproliferative agent, mainly is composed of a pentomannan. However, tetrasaccharide and disaccharide components are also present. The molecular profile and the anticoagulant potency of PI-88 are investigated in this study. Gel permeation chromatography and polyacrylamide gel electrophoresis analyses were carried out to determine the molecular profile and separation of components of PI-88, respectively. Potentiation of antithrombin III (ATIII) and heparin cofactor-II (HC-II) activity were measured using chromogenic substrate assay. In order to determine anticoagulant and antiprotease effects of PI-88, various global anticoagulant tests, such as the prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT), Hep-test (Haemachem Inc., St. Louis), ecarin clotting time (ECT), activated clotting time (ACT), and thromboelastography (TEG) were used. Anti-Xa and anti-IIa activities also were measured. The effect of PI-88 on the release of tissue factor pathway inhibitor (TFPI) was performed in nonhuman primates who received PI-88 and in endothelial cell culture systems. The relative susceptibility of PI-88 to heparinase I, protamine sulfate (PS), and platelet factor 4 (PF4) also was evaluated. The high-performance liquid chromatography profiles of PI-88 showed that its average molecular weight is approximately 2300 Da. Separation and gradient electrophoretic patterns of PI-88 showed that it is composed of five different fractions. This agent activates HC-II through inhibiting the thrombin generation but not inhibiting ATIII. Although PI-88 produced a concentration-dependent prolongation of all of the clotting tests, ECT gave the best correlation in the dose-response curve (ECT, r2 = 0.94; TT, r2 = 0.84; APTT, r2 = 0.69). Heparinomimetic mannopentaose phosphate sulfate (PI-88) exhibited marked inhibition of FIIa, but not of FXa. Heparinase I failed to produce significant neutralization of PI-88 in all the assays used, whereas PS and PF4 partially neutralized the effects of this compound. Heparinomimetic mannopentaose phosphate sulfate (PI-88) produced fivefold increase in the TFPI levels at 15 minutes after intravenous (IV) injection to primates. The incubation of PI-88 in endothelial cell culture system also showed a strong effect on TFPI release. These results suggest that PI-88 exhibited strong antithrombotic and anticoagulant activity in addition to its known antiproliferative properties. Because of the molecular characteristics and the dual nature of the pharmacologic action of PI-88, this agent represents an attractive pharmacologic agent for the control of thrombotic and proliferative disorders.
Demir
M
M
Loyola University Medical Center, Maywood, IL 60153, USA.
Iqbal
O
O
Hoppensteadt
D A
DA
Piccolo
P
P
Ahmad
S
S
Schultz
C L
CL
Linhardt
R J
RJ
Fareed
J
J
D000925:Anticoagulants / Q000737:chemistry / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology*; D001780:Blood Coagulation Tests; D002240:Carbohydrate Sequence; D004305:Dose-Response Relationship, Drug; D019763:Heparin Lyase / Q000494:pharmacology; D006801:Humans; D008074:Lipoproteins / Q000187:drug effects; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009844:Oligosaccharides / Q000737:chemistry / Q000494:pharmacology*; D011480:Protease Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D013917:Thrombin / Q000037:antagonists & inhibitors
D000925:Anticoagulants; D000970:Antineoplastic Agents; D008074:Lipoproteins; D009844:Oligosaccharides; D011480:Protease Inhibitors; C051928:lipoprotein-associated coagulation inhibitor; C120158:phosphomannopentaose sulfate; D013917:Thrombin; D019763:Heparin Lyase