Preparation of biocompatible materials by immobilization of apyrase

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A prosthetic polymer material is made non-thrombogenic by immobilizing apyrase on its surface. Immobilization is preferably carried out by hydrolytically activating the surface of a polyamide polymer or a polyethylene terphthalate polymer, and treating the hydrolyzed polymer with a solution of cross-linking agent and a solution of apyrase. The apyrase converts adenosine diphosphate to adenosine monophosphate and adenosine whereby the formation of thrombi is inhibited.

The present application is a continuation of application Ser. No. 165,708 filed July 3, 1980, now abandoned.

This invention relates to a process for preparing biocompatible polymer and non-polymer materials by immobilising apyrase on their surface, apyrase being an enzyme able to catalyse the conversion of adenosine diphosphate (ADP) to adenosine monophosphate (AMP), followed by the conversion of this latter to adenosine. The invention also relates to the manufactured articles obtained by said process.

The possibility of imparting biocompatible properties to various types of material is of enormous practical importance at the present time. In this respect, materials exist which because of their good mechanical, machining and strength properties and the absence of toxicity would find immediate application in the construction of protheses of medium or long duration for implantation purposes, or in the construction of elements of auxiliary machines for use in extracorporal circulation, such as renal dialysis apparatus or heart-lung machines.

For this purpose, materials could be used lying within a very wide range from aliphatic or aromatic polyamide polymers, polyesters, polycarbonates, polyurethanes and PVC to special metal alloys. Unfortunately, the use of these materials is strongly limited by their generally poor biocompatibility testing. This is because as soon as a foreign material is inserted into the blood circulation, it immediately gives rise to the formation of thrombi by the initiation of an extremely complicated process.

It is believed that there is firstly an adhesion of the blood platelets to the material surface, with the consequent release from the platelets of ADP and serotonin, which then cause platelet aggregation. The platelet aggregation is itself a fundamental stage in the formation of thrombi. This is because it gives rise to the release of phospholipids which are essential in the blood coagulation process, by promoting the conversion of fibrinogen into fibrin.

The role of ADP as a platelet aggregation inducer, and thus as an initiator of the thrombi formation process, is widely known. See for example the fundamental work of A. Gaarder and A. Hellem in Nature 192, 531 (1961). It is therefore apparent that the immobilisation of an enzyme such as apyrase able to convert into AMP and adenosine the ADP produced by the adhesion of the platelets to the surface of the material which is placed in contact with the blood can inhibit subsequent platelet aggregation, so blocking the formation of thrombi. In this respect, it has been found, and forms the subject matter of the present invention, that the immobilisation of apyrase on the surface of thrombogenic materials gives these latter satisfactory biocompatible properties.

These properties do not depend on the system used for immobilising the enzyme. This can be done by adsorption and subsequent cross-linkage on the surface of polymer materials, or even on metal surfaces, for example on the surface of needles used for the arterio-venous connections in extracorporal circulation.

Alternatively, the apyrase can be bonded covalently to functional groups present on the surface of materials of which the biocompatibility testing is to be increased. It is indeed possible, if necessary, to previously activate the material so as to release reactive groups on to its surface which can be used for immobilising the apyrase.

For example, the enzyme can be attached covalently to the amino (or carboxyl) groups of aliphatic or aromatic polyamides subjected to mild surface hydrolysis. In the same manner, carboxyl groups of surface-hydrolysed polyesters can be used.

The invention is described in detail by the following examples, which however are not to be considered limiting.