The following is a brief summary of the U.S. PATENT # 5278189 that was issued on January 11, 1994 to Pauling and Rath. This is some what edited for brevity and readability, but is otherwise a true copy of the patent. For complete text go to U.S. Patent site or refer here.

U.S. PATENT # 5278189

Prevention and treatment of occlusive cardiovascular disease with ascorbate and substances that inhibit the binding of lipoprotein (A)

WKU Patent Number: 05278189
TTL Title of Invention: Prevention and treatment of occlusive cardiovascular disease with ascorbate and substances that inhibit the binding of lipoprotein (A)
ISD Issue Date: 19940111 [1994 Jan 11]
NAM Inventor Name: Rath; Matthias W.
NAM Inventor Name: Pauling; Linus C.

Rath, M. & L. Pauling, “Solution of the puzzle of human cardiovascular disease: Its primary cause is ascorbate deficiency leading to the deposition of lipoprotein(a) and fibrinogen/fibrin in the vascular wall,” J. Orthomolecular Med. (In Press 1991).

ABST Abstract A method is provided for prevention and treatment of cardiovascular disease, such as atherosclerosis, by administering therapeutically effective dosages of a drug comprised of ascorbate, lipoprotein(a) binding inhibitors, and antioxidants.

TECHNICAL FIELD

The present invention relates generally to the prevention and treatment of cardiovascular disease and more particularly to methods and compounds that inhibit the binding of lipoprotein (a) to components of the arterial wall.

BACKGROUND OF THE INVENTION

Lipoprotein(a) (”Lp(a)”) was first identified by Blumberg, B. S., et al. (1962) J. Clin. Invest. 41: 1936-1944 , and Berg, K. (1963) Acta Pathol. 59: 369-382. The structure of Lp(a) resembles that of low-density lipoprotein (”LDL”) in that both share a lipid apoprotein composition, mainly apolipoprotein B-100 (”apo B”), the ligand by which LDL binds to the LDL receptors present on the interior surfaces of arterial walls.

The unique feature of Lp(a) is an additional glycoprotein, designated apoprotein(a), apo(a), which is linked to apo B by disulfide groups. The cDNA sequence of apo(a) shows a striking homology to plasminogen, with multiple repeats of kringle 4, one kringle 5, and a protease domain. The isoforms of apo(a) vary in the range of 300 to 800 kDa and differ mainly in their genetically determined number of kringle 4 structures. McLean, J. W., et al. (1987) Nature 300: 132-137. Apo(a) has no plasmin-like protease activity. Eaton, D. L., et al., (1987) Proc. Natl Acad. Sci. USA, 84: 3224-3228.

Serine protease activity, however, has been demonstrated. Salonen, E., et al. (1989) EMBO J. 8: 4035-4040. Like plasminogen, Lp(a) has been shown to bind to lysine-sepharose, immobilized fibrin and fibrinogen, and the plasminogen receptor on endothelial cells. Harpel, P.C., et al. (1989) Proc. Natl. Acad. Sci. USA 86:3847-3851; Gonzalez-Gronow, M., et al. (1989) Biochemistry 28: 2374-2377; Miles, L. et al. (1989) Nature 339: 301-302; Hajjar, K. A., et al. (1989) Nature 339: 303-305. Furthermore, Lp(a) has been demonstrated to bind to other components of the arterial wall like fibronectin and glycosaminoglycans. The nature of these bindings, however, is poorly understood.

Essentially all human blood contains lipoprotein(a); however, there can a thousand-fold range in its plasma concentration between individuals. High levels of Lp(a) are associated with a high incidence of cardiovascular disease. Armstrong, V. W., et al. (1986) Atherosclerosis 62: 249-257; Dahlen, G., et al. (1986) Circulation 74: 758-765; Miles, L. A., et al. (1989) Nature 339: 301-302; Zenker, G., et al. (1986). Stroke 17: 942-945 (The term occlusive cardiovascular disease will be used hereafter as including all pathological states leading to a narrowing and/or occlusion of blood vessels throughout the body, but particularly atherosclerosis, thrombosis and other related pathological states, especially as occurs in the arteries of the heart muscle and the brain.)

For some time, general medical practice has focused on the role of LDL, the so called “bad cholesterol,” in occlusive cardiovascular disease. A great many studies have been published ostensibly linking occlusive cardiovascular disease with elevated levels of LDL. As a result, most therapies for the treatment and prevention of arteriosclerosis rely on drugs and methods for the reduction of serum levels of LDL’s. Such therapies have had mixed results. The efficacy of such approaches to the problem of occlusive cardiovascular disease continues to be major source of debate.
There exists therefore a need for a drug therapy for reducing the binding of Lp(a) to vessel walls, for reducing the overall level of Lp(a) in the circulatory system and for promoting the release of existing deposits of Lp(a) on vessel walls.

SUMMARY OF THE INVENTION

The foregoing needs in the treatment and prevention of cardiovascular disease are met by the methods and compositions of the present invention.

A method is provided for the treatment of occlusive cardiovascular disease, comprising the step of administering to a subject an effective amount of ascorbate and one or more binding inhibitors, as a mixture or as a compound comprising ascorbate covalently linked with binding inhibitors, which inhibit the binding of Lp(a) to blood vessel walls, such as arterial walls. This effect may also be obtained by administering an effective amount of one or more inhibitors, without ascorbate.

The term binding inhibitor throughout the specification and claims is intended to include all substances that have an affinity for the lysine binding site present on the interior walls of blood vessels, particularly arteries, the site of Lp(a) binding. Most of these substances compete with lasmin for the lysine binding site and some of these compounds, in high doses, are in clinical use for the treatment of hyperfibrinolytic states.

A method is further provided for the prevention of atherosclerosis comprising the step of administering to a subject an effective amount of ascorbate and one or more binding inhibitors as previously discussed but further comprising one or more antioxidants. The term antioxidant throughout the specification and the claims is intended to exclude ascorbate which has as one of its chemical properties a potent antioxidant effect.

It is thus an object of the invention to provide a method for treatment of occlusive cardiovascular disease by administering to a subject an effective amount of ascorbate and one or more binding inhibitors, or an effective amount of one or a mixture of binding inhibitors.

It is another object of the invention to provide a method for preventing of occlusive cardiovascular disease, by administering to a subject an amount of ascorbate effective to lower the amount of Lp(a) in the plasma of the subject.

Yet another object of the present invention is to provide a method for prevention of cardiovascular disease by administering to a subject an effective amount of ascorbate and one or more binding inhibitors, or an effective amount of one or more binding inhibitors.

A further object of the present invention is to provide a pharmaceutically acceptable agent for the treatment of occlusive cardiovascular disease.

Still another object of the present invention is to provide a pharmaceutically acceptable agent for the prevention of cardiovascular disease.

These and other objects will be more readily understood upon consideration of the following detailed descriptions of embodiments of the invention and the drawings.

DETAILED DESCRIPTION OF THE INVENTION

Our invention is based in part on our discovery that animals which have lost the ability to produce ascorbate, such as higher primates and guinea pigs, uniformly produce Lp(a). Most animals which possess the ability to synthesize ascorbate generally do not produce Lp(a). Further, we have found that ascorbate deficiency in humans and guinea pigs tends to raise Lp(a) levels and causes atherosclerosis by the deposition of Lp(a) in the arterial wall, from which we conclude that ascorbate administration lowers plasma Lp(a) levels.

We have also discovered that substances that inhibit the binding of Lp(a) to components of the arterial wall, particularly to fibrinogen, fibrin and fibrin degradation products herein identified as binding inhibitors, such as lysine or .epsilon.-aminocaproic acid used alone or in combination with ascorbate, cause release of Lp(a) from the arterial wall. Thus, ascorbate and such binding inhibitors are not only useful for the prevention of occlusive cardiovascular disease, but also for the treatment of such disease. The present invention, then, provides methods and pharmaceutical agents for the both the treatment and prevention of occlusive cardiovascular disease in vivo.

GENERAL APPLICATIONS

The present invention provides a method and pharmaceutical agent for the treatment and prevention of occlusive cardiovascular disease generally, by administering to a subject an effective amount of ascorbate and one or more binding inhibitors. Throughout the specification and claims, the term binding inhibitor is intended to cover any substance which has as at least one of its chemical properties the ability to inhibit the binding of Lp(a) to blood vessel wall components, particularly to fibrin or fibrinogen.

As used herein, the term “ascorbate” includes any pharmaceutically acceptable salt of ascorbate, including sodium ascorbate, as well as ascorbic acid itself. Binding inhibitors include, but are not limited to .epsilon.-aminocaproic acid, lysine, tranexamic acid (4-aminomethylcyclohexane carboxylic acid), p-aminomethylbenzoic acid, p-benzylamine sulfuric acid, o-N-acetyl-lysine-methyl ester, PROBUCOL (a compound comprised of 2 butyl hydroxy tocopherol groups linked together by a disulphide group), Aprotinin, trans-4-aminomethylcyclohexanecarboxylic acid (AMCA), and benzamidine derivatives such as amidinophenylpyruvic acid (APPA) and 1-naphthyl-(1)-3-(6-amidinonaphthyl-(2))-propanone-1 HCl (NANP).

An effective amount of a binding inhibitor or a mixture of binding inhibitors may also be used, without ascorbate. Other substances used in the treatment of occlusive cardiovascular disease may be co-administered, including antioxidants, such as tocopherol, carotene and related substances; vitamins; provitamins; trace elements; lipid-lowering drugs, such as hydroxy-methyl-glutaryl coenzyme A reductase inhibitors, nicotinic acid, fibrates, bile acid sequestrants; and mixtures of any two or more of these substances.

Although ascorbate can be used alone or in varying combinations with one or more representative constituents of the above classes of compounds, we prefer when treating a pre-existing cardiovascular condition to combine ascorbate with at least one each of the binding inhibitors, antioxidants and lipid lowering drugs elements in the dosages (per kilogram of body weight per day (Kg BW/d)) provided in Table 1. It should be noted that Table 1 provides differing concentration ranges of each constituent, depending upon whether the agent is to be administered orally or parenterally.

The variance in dosages is reflective of variation in disease severity. It will be realized therefore that if the subject has been diagnosed for advanced stages of atherosclerosis,dosages at the higher end of this range can be utilized. However, if only prevention of an atherosclerosis condition is the object, dosages at the lower end of this range can be utilized.

As an alternative, a pharmaceutical agent identical to the one just described, but omitting ascorbate, may be employed.

Where ascorbate and binding inhibitors are utilized in the same agent, they may simply be mixed or may be chemically combined using synthesis methods well known in the art, such as compounds in which ascorbate and the inhibitor are covalently linked, or form ionically bound salts. For example, ascorbate may be bound covalently to lysine, other amino acids, or .epsilon.-aminocaproic acid by ester linkages. Ascorbyl .epsilon.-aminocaproate is such an example. In this form the ascorbate moiety may be particularly effective in preventing undesirable lipid peroxidation.

In the case of oral administration, a pharmaceutically acceptable and otherwise inert carrier may be employed. Thus, when administered orally, the active ingredients may be administered in tablet form. The tablet may contain a binder such as tragacanth, corn starch or gelatin; a disintegrating agent, such as alginic acid, and/or a lubricant such as magnesium stearate. If administration in liquid form is desired, sweetening and/or flavoring agents may be used. If administration is by parenteral injection, in isotonic saline, a phosphate buffered solution or the like, may be used as a pharmaceutically acceptable carrier.

The advisability of using binding inhibitors in treating occlusive cardiovascular disease will depend to some extent on the subject’s general health, particularly with regard to hyperfibrinolytic conditions. Most binding inhibitors (except lysine) are used clinically to treat such conditions. As a result, monitoring of the subject’s coagulation and fibrinolytic system is recommended before and during treatment for occlusive cardiovascular disease. Long-term administration of binding inhibitors will require formulations in which the dosages of binding inhibitors are in the lower ranges of the dosages given in Table 1.

Prevention, as contrasted with treatment, of cardiovascular disease may be accomplished by oral or parenteral administration of ascorbate alone. Table 1 gives a range of ascorbate concentrations sufficient to lower the serum Lp(a) concentration.

Preferably the prevention of the occlusive cardiovascular disease according to the invention is accomplished by use of a physical mixture of ascorbate and one or more binding inhibitors, or by use of a compound comprising covalently linked ascorbate with one or more of the binding inhibitors, which inhibit binding of Lp(a) to the arterial wall. A binding inhibitor or mixture of binding inhibitors may also be administered without ascorbate to prevent Lp(a)-associated occlusive cardiovascular disease.

To optimize the therapeutic effect of the release of Lp(a) from the blood vessel walls, the ascorbate and the binding inhibitors described above may be separately administered. Further optimization of therapeutic effect can be gained by using a time release composition to achieve relatively constant serum concentrations of the agent through time.