Title:  Surfactants that mimic the glycocalyx

United States Patent:  6,759,388

Issued:  July 6, 2004

Inventors:  Marchant; Roger E. (Cleveland Heights, OH); Zhang; Tianhong (Rockport, MA); Qiu; Yongxing (Atlanta, GA); Ruegsegger; Mark A. (Cleveland Heights, OH)

Assignee:  Nanomimetics, Inc. (Cleveland Heights, OH)

Appl. No.:  302195

Filed:  April 29, 1999

Abstract

Comblike, surfactant polymers for changing the surface properties of biomaterials are provided. Such surfactant polymers comprise a polymeric backbone of repeating monomeric units having functional groups for coupling with side chains, a plurality of hydrophobic side chains linked to said backbone via the functional groups, and a plurality of hydrophilic side chains linked to said backbone via the functional groups. The hydrophobic side chains comprise an alkyl group comprising from 2 to 18 methylene groups. The alkyl groups are linked to the polymeric backbone through ester linkages, secondary amine linkages, or, preferably, amide linkages. The hydrophilic side chain is selected from the group consisting of: a neutral oligosaccharide, which, preferably, has weight average molecular weight of less than 7000; a charged oligosaccharide, preferably a negatively charged oligosaccharide having a weight average molecular weight of less than 10,000; an oligopeptide of from about 3 to about 30 amino acid residues, said oligopeptide having an amino acid sequence which interacts with protein receptors on the surface of cells; and combinations thereof. Methods of making the surfactant polymers and using the surfactant polymers to alter the surface properties of a biomaterial are also provided.

SUMMARY OF THE INVENTION

In accordance with the present invention, novel, comblike, surfactant polymers which are useful for changing the surface properties of biomaterials are provided. Such surfactant polymers comprise a polymeric backbone of repeating monomeric units having functional groups for coupling with side chains, a plurality of hydrophobic side chains linked to said backbone via the functional groups, and a plurality of hydrophilic side chains linked to said backbone via the functional groups. The hydrophobic side chains comprise an alkyl group (CH₃ (CH₂ --)_n) comprising from about 2 to 18 methylene groups. The alkyl groups are linked to the polymeric backbone through ester linkages, secondary amine linkages, or, preferably, amide linkages. The hydrophilic side chain is selected from the group consisting of: a neutral oligosaccharide, which, preferably, has weight average molecular weight of less than 7000; a charged oligosaccharide, preferably a negatively charged oligosaccharide having a weight average molecular weight of less than 10,000; an oligopeptide of from about 3 to about 30 amino acid residues, said oligopeptide having an amino acid sequence which interacts with protein receptors on the surface of cells; and combinations thereof. A preferred amino acid sequence which is known to interact with molecules on the surface of cells is arginine (R), glycine (G), and aspartic acid (D). The hydrophilic side chains are linked to the polymeric backbone through ester linkages, secondary amine linkages, or, preferably, amide linkages. It has been shown that effective surface modification of biomaterials can be accomplished using the surfactant polymers of the present invention without any requirements for surface coupling chemistries.

The present invention also provides an implantable, biomaterial having a comblike surfactant polymer of the present invention coated on a hydrophobic surface thereof.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides novel comblike surfactant polymers that mimic the glycocalyx. The glycocalyx is the oligosaccharide-rich region on the surface of cells. The glycocalyx serves to prevent undesirable biological adhesions, while proteins embedded in the cell membrane glycocalyx serve to promote desirable specific adhesions.

The comblike surfactant polymers of the present invention comprise a flexible polymeric backbone which is linked to a plurality of hydrophobic side chains and to a plurality of hydrophilic side chains. The polymeric backbone is conformationally flexible. Preferably, the polymeric backbone is formed from a homopolymer that contains a plurality of functional side groups such as for example OH groups, COOH groups, or NH₂ groups. Although less preferred, the polymeric backbone may be formed from a copolymer which has a combination of functional side groups. For example, the copolymer may have OH side groups and NH₂ side groups. Suitable homopolymers for forming the comblike surfactant polymer are, by way of example, polylysine, poly(vinyl alcohol) or poly(vinyl amine). Preferably, the polymeric backbone is formed from a poly(vinyl amine).

The hydrophopobic side chains comprise from about 2 to 18 methylene groups, preferably from about 4 to about 12 methylene groups, more preferably from about 4 to about 10 methylene groups, and are linked to the polymeric backbone via an ester linkage, a secondary amine linkage, or, preferably, an amide linkage. Preferably, the hydrophobic side chains are attached to the polymeric backbone by reacting an alkanoyl (CH₃ (--CH₂ --)_n CO--)or an alkanal (CH₃ (CH₂ --)_n CHO)with the homopolymer of the backbone using conventional procedures. For example, a plurality of hydrophobic side chains may be linked to poly(vinyl amine) by standard to procedures that use the corresponding N-alkanoyloxy succinimide and poly(vinyl amine) as reactants.

The hydrophilic side chain is selected from the group consisting of a neutral oligosaccharide which preferably has a weight average molecular weight of less than 7000; a charged oligosaccharide, preferably a negatively charged oligosaccharide having a weight average molecular weight of less than 10,000; an oligopeptide of from about 3 to about 30 amino acid residues said oligopeptide having an amino acid sequence which interacts with protein receptors on the surface of cells; and combinations thereof. The hydrophilic side chains are linked to the polymeric backbone through an ester linkage, a secondary amine linkage, or preferably an amide linkage. In a preferred embodiment a plurality of oligopeptides and a plurality of oligosaccharides are attached to the polymeric backbone.

To form a coating which blocks adhesion of non-specific plasma proteins on the surface of the substrate, the surfactant polymer, preferably, comprises a plurality of hydrophilic side chains formed from oligosaccharides. Such surfactant polymers may be non-ionic or ionic. Suitable oligosaccharides include, but are not limited to, neutral oligosaccharides having a weight average molecular weight of less than about 7000. Examples of such neutral oligosaccharides include dextran, which is composed of .alpha.(1.fwdarw.4) linked glucose residues, and an oligomaltose, which is composed of .alpha.(1.fwdarw.4) linked glucose residues. Such oligosaccharide side chains are attached to the reactive amines of the poly(vinyl amine) by standard procedures that employ the polymer and the corresponding lactone form of the oligosaccharide as reactants.

The hydrophilic side chains may also be formed from charged oligosaccharides such as, for example, the oligosaceharides that are obtained from heparin. The heparin oligosaccharides are hydrated and negatively charged which provides an additional electrostatic repulsive force that further repels plasma proteins and cellular elements such as platelets. The heparin oligosaccharide contains the unique pentasaccharide sequence that is essential for heparin's anticoagulant activity. The heparin product of deaminative cleavage of heparin possesses a terminal 2,5 anhydromanose unit. In a preferred embodiment, the terminal aldehyde of the 2,5 anhydromannose is reacted with the amines on the polymeric backbone via reductive amination to form a secondary amine.

Other suitable charged oligosaccharides for forming coatings which are non-adhesive for plasma proteins include dermatan sulfate, and dextran sulfate, which are hydrated and negatively charged and serve to repulse proteins and platelets. Preferably, these charged oligosaccharides are linked to the backbone by an amide linkage which is formed by converting the reducing end of the oligosaccharide to a lactone and then selectively reacting the lactone with the functional amine groups on the homopolymer. Alternatively, the charged oligosaccharides are linked to the backbone by reductive amination, which involves a reaction between an amine group on the polymer and a terminal aldehyde on the oligosaccharide. The resulting linkage is a secondary amine.

The ratio of hydrophobic side chains to hydrophilic oligosaccharide chains on the polymer backbone is designed to achieve a hydrophilic to hydrophobic balance that allows the surfactant to adsorb onto the hydrophobic surface of the biomaterial and, preferably, that allows the hydrophilic side chains to extend from the surface of the biomaterial into a surrounding aqueous medium. The hydrophilic to hydrophobic balance depends on the density of the hydrophobic and hydrophilic side chains and the length of the hydrophobic side chains and hydrophilic side chains. Adhesion of the adsorbing polymer onto the hydrophobic surface of the biomaterial is enhanced by increasing the length, i.e., the number of methylene groups of the hydrophobic side chain, by increasing the density of the hydrophobic side chains relative to the hydrophilic side chains, and/or by reducing the length of the hydrophilic side chains. Thus, for a surfactant polymer in which the hydrophilic side chains are formed from dextran side chains composed of 9 glucose residues and from hydrophobic alkanoyl side chains comprising 4 methylene groups, the preferred ratio of hydrophilic side chains to a hydrophobic side chains is from about 2:1 to about 1:6, more preferably from about 1:1 to about 1:5 If the hydrophobic side chains comprise 10 methylene groups and the hydrophilic side chains are dextran side chains composed of 9 glucose residues, the preferred ratio of hydrophilic side chains to hydrophobic side chains from 3:1 to about 1:5, more preferably from about 2:1 to about 1.3.

In those cases where the ratio of oligosaccharide side chains to hydrophobic side chains on the resulting surfactant polymer is low, i.e., about 1:1, there may be some unreacted functional groups, such as for example residual amine groups, which are available to bind to plasma proteins. Preferably, such unreacted functional groups are blocked or capped by further reacting the resulting surfactant polymer with organic molecules that are small relative to the hydrophilic side chains prior to application of the surfactant to the biomaterial. Suitable small organic molecules are, by way of example, glucose, maltose, and acetaldehyde.

Surfactant polymers useful for changing the surface properties of a biomaterial may also be comprised of a polymeric backbone, a plurality of hydrophobic side chains, and a plurality of hydrophilic side chains formed from polyethylene oxide. Mono-aldehyde terminated polyethylene oxide side chains are linked to the polymeric backbone via reductive amination to form a secondary amine.

In another embodiment, the suactant polymer comprises a plurality of hydrophilic oligopeptide sides chains capable of interacting with specific protein receptors on the surface of animal cells such as, for example, endothelial cells. The oligopeptide side chains act as ligands for binding the cells to the surface of the biomaterial. The oligopeptide side chains comprise from about 3 to about 30 amino acid residues. Preferably, the oligopeptide comprises the amino acid sequence RGD, more preferably RGDS, most preferably RGDSP. Alternatively, the oligopeptide comprises one of the following amino acid sequences: (i) RRAR, (ii) RRRKRR, (iii) PPRRARVT, or (iv) PPREVVPRPRP. In a preferred embodiment the oligopeptide comprises the sequence GSSSGRGDSPX, wherein X is alanine or another hydrophobic amino acid residue. The oligopeptide ligands are linked to the homopolymer backbone by an ester linkage, a secondary amine linkage, or an amide linkage.

Comblike surfactant polymers that comprise hydrophobic side chains, oligopeptide side chains, and oligosaccharide side chains are useful for providing a glycocalyx-like coating on the surface of a biomaterial. Such glycocalyx-like coatings prevent adhesion of plasma proteins to the coated surface of the biomaterial as compared to an uncoated hydrophobic surface of the biomaterial. Such coatings promote adhesion of selected cells, particularly endothelial cells, to the coated surface of the biomaterial.

The surfactants are used to coat one or more surfaces of a hydrophobic biomaterial, including a flexible, hydrophobic material. Such biomaterials are used to make implantable medical devices such as for example heart valves, stents, vascular grafts and catheters. Preferably, the surfactant is used to coat a surface which will come into contact with the blood or other body fluid of the patient following implantation of the device in the patient. The substrate is any material demonstrating biocompatibility and sufficient hydrophobicity to bind the surfactant, such as, for example, graphite and polyethylene. Other suitable biomaterials include, for example: polystyrene, polyesters, for example: Dacron.RTM., carbon in pyrolytic carbon; polycarbonate; polymer membranes, for example, cellulose acetate, polyacrylonitrile; fluorocarbon polymers, for example Teflon.RTM., Impra.RTM. and Gortex.RTM.; polysulfones; polyvinyl chloride; silicone rubber for example, Silastic.RTM.; silicone polymers; polypropylene; and polyurethanes. Suitable biomaterials also include nonpolymeric materials, such as for example, titanium, stainless steel, silicon, glass; and mixtures or composites thereof, that have been treated in a manner which renders their surfaces hydrophobic. The selection of the biomaterial depends upon the mechanical and functional properties required for forming an implantable biomedical device

The comblike surfactant polymers of the present invention, preferably, are soluble in water and are easily applied to the surface of the biomaterial. Application is achieved by immersing the biomaterial in a solution, preferably an aqueous solution, comprising the surfactant polymer. The surfactant spontaneously attaches to the hydrophobic surface of the polymeric biomaterial to provide a monolayer which alters the surface properties of the hydrophobic surface. The monolayer is bonded to the biomaterial via hydrophobic interactions and is able to withstand a shear stress of 75 dynes/cm² as determined using a rotating disc system followed by infrared spectroscopic surface analysis. Following adsorption of the surfactant to the hydrophobic surface, the surfactant may be air dried and stored in the dry state.

Claim 1 of 11 Claims

What is claimed is:

1. A comblike surfactant polymer comprising:

a) a polymeric backbone of repeating monomeric units;

b) a plurality of hydrophobic side chains comprising from about 2 to about 18 methylene groups, said plurality of hydrophobic side chains being linked to said polymeric backbone by ester linkages, secondary amine linkages, amide linkages, or combinations thereof; and

c) a plurality of hydrophilic side chains linked to said polymeric backbone by ester linkages, secondary amine linkages, amide linkages, or combinations thereof, said hydrophilic side chains selected from the group consisting of: neutral oligosaccharide side chains having a weight average molecular weight of less than 7000; charged oligosaccharide side chains having a weight average molecular weight of less than 10,000; an oligopeptide of from about 3 to 30 amino acid residues, said oligopeptide having an amino acid sequence which interacts with protein receptors on the surface of cells; and combinations thereof.

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