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Description  |
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FIELD AND BACKGROUND OF THE INVENTION
As the primary organ of vision in vertebrate animals, the eye often compared to the usual photographic camera. In simplified terms, the eye has a lens system, a variable aperture system (the pupil) and a light-sensitive retina that corresponds
to the film. The focusing ability of the eye resides within the crystalline lens which comprises a strong elastic capsule filled with viscous, transparent fibers of crystalline and albuminoid proteins. Far more complex and fragile than that of a mere
camera lens, the crystalline lens of the eye is especially prone to damage due to disease, environmental factors and the aging process. For a review on the lens system, see, for example, Cotlier, E. in "Adler's Physiology of the Eye: Clinical
Application", 8th edition, R. A. Moses and W. M. Hart, editors; C. V. Mosby Co.; St. Louis, Mo., USA (1987), pages 268-272.
The unique ability of the lens to change its shape or curvature to automatically adjust the focus of the eye for objects at different distances is known as accommodation. Accommodation occurs by muscular contraction and relaxation causing the
elastic-like lens to change shape and increase its optical or refractive power. For a general discussion of the mechanism of accommodation, see Guyton, R. C. "Medical Physiology", 6th edition, W. B. Saunders Co.; Philadelphia, Pa., USA; 1981; page
724-735; Toats, F. M. Physiol. Rev. 52: 828, 1972; and Witkovsky, P., Annu. Rev. Physiol. 33: 257, 1971.
Accommodation is lost during the aging process primarily due to a hardening of the natural lens. Loss of accommodation due to a progressive denaturation of the lens proteins produces an abnormal condition commonly known as "presbyopia".
Presbyopia generally affects individuals in the early to mid forties and the resultant gradual loss of visual acuity is generally treated with bifocal spectacles. For general discussion of presbyopia, see "Merck Manual" 15th Edition, Merck, Sharpe and
Dohme publishers, 1987, page 2211 and Guyton, R. "Medical Physiology", 6th edition, W. B. Saunders Co.; Philadelphia, Pa., USA; 1981; page 724-735.
Lenticular cataracts are an especially common eye abnormality characterized by a progressive loss of vision beginning in middle age or later. In the early stage of cataract formation, the transparent protein fibers of the lens become denatured,
presumably by oxidative damage due to the normal aging process. The denatured proteins then coagulate, forming the characteristic opaque areas in place of the normal transparent protein fibers of the lens. In the advanced stage, calcium deposition
occurs on the coagulated proteins, thus further increasing the opacity. Cataract formation can also be accelerated by exposure to X-rays, heat from infrared rays, systemic disease (e.g. diabetes), uveitis, or systemic medications (e.g. corticosteroids). The degree of vision loss due to cataract formation can be ascertained by ophthalmoscopic slit-lamp examination which provides more details about the character, location, and extent the opacities. For a detailed discussion of cataract formation, see Van
Heynengen, R. Sci. Am. 233(6): 70, 1975 and "Merck Manual", 15th Edition, Merck, Sharp and Dohme publisher, 1987, page 2227.
Frequent refraction corrections help maintain useful vision during cataract development. When a cataract has obscured light transmission so greatly that useful vision is lost, surgical intervention is necessary to extract the lens. Lens
extraction can be accomplished by total removal of the lens, or by phacoemulsification of the lens followed by irrigation and aspiration, leaving the lens capsular sac intact. When this is done, however, the eye loses a large portion of its refractive
power. Post-operative refractive correction to alleviate the visual defect is accomplished by cataract spectacles, contact lens, or intraoperative implantation of an intraocular lens.
Traditionally, cataract spectacles have produced less than satisfactory results because of induced visual distortions such as aberrant depth perception. For example, cataract spectacles with thick lenses are known to induce a Galilean telescopic
effect which results in abnormally magnified images. Moreover, unilateral surgical removal of lenses makes correction of stereovision by such spectacles virtually impossible.
Contact lenses eliminates many of the aforementioned problems, however, the magnification problem remains. Furthermore, many patients are unable to tolerate contact lenses because of poor manual dexterity, insufficient tear production or lens
hygiene problems.
Implantation of endocapsular lens or (as more commonly known) intraocular lens (IOL), on the other hand, has been widely accepted as the treatment of choice for correcting visual impairments following removal of diseased lenses. The remarkable
success of IOL implantation is due to significant improvements in surgical instrumentation and technique as well as in the design and construction materials of IOLs.
Microsurgical procedures have been developed to remove cataract lenses through very small incisions in the capsular sac (see, for example, Arshinoff, S. A. Curr. Can. Ophthalmic Pract. 4(2):64, 1986 and Welsh, R. C. et al. Cataract Surg NOW
1(2): 21-22, 1983 for a discussion of capsulotomy surgical techniques). An IOL is then gently inserted into the intact capsular sac, positioned in place, and the wound is then closed with fine sutures. Conventional IOLs are generally fitted with
surgical loops to fix and/or maintain the IOL in position. Materials which are used to fabricate the IOLs are typically rigid or semi-rigid plastics such as polymethyl methacrylate. Newer and softer fabrication materials include biocompatible hydrogels
and silicones. For a general discussion of IOL development, see for example, Apple, D. J., Geiser, S. C. and Isenberg, R. A. "Evolution of Intraocular lenses," University of Utah Printing Service, 1985).
Conventional IOLs, however, have a number of deficiencies associated with their use. For a review of the complications of conventional IOLs, see for example, Apple, D. J. et al., (1984) Ophthalmology, Vol. 29, No. 1; Drews, R. C. (1982). Trans. Ophthal. Soc. U.K., Vol. 102, page 498; DeVore, D. P. (1991) J. Long-Term Effects of Medical Implants, Vol. 2, in press. For example, implantation of conventional IOLs are known to induce excessive accumulation of epithelial cells lining the lens
capsule. This interface of cells, in turn, results in opacification of the lens as well as a variety of pathological conditions which include pupillary occlusions, iris atrophy and secondary glaucoma. Moreover, Mechanical dislocation of the IOL
frequently results in damage to the corneal endothelium. Another notable drawback is that none of the IOLs of the prior art have accommodative capacity.
With the capsular bag intact, a safe, effective injectable material that could be used to refill the capsular bag and which simulates the natural lens would be desirable. This material should have an index of refraction similar to that of the
natural lens, but variable so that any refractive errors might be corrected. Such injectable lens would still be able to accommodate and therefore provide a dramatic advantage over current intraocular lens implants.
SUMMARY OF THE INVENTION
The present invention relates to a method of making an intraocular lens using injectable collagen-based compositions. Chemically modified collagen compositions with varying indices of refraction can be used in filling the intact lens capsular
bag following removal of the damaged lens. The new prosthetic lens, having predetermined index of refraction, are capable of accommodation. The intraocular lens prepared by the method of the present invention can be used to replace natural or diseased
lenses for treatment of cataracts, presbyopia, myopia and hyperopia.
The modified collagen compositions of the invention comprise viscous yet injectable solutions of purified, collagen modified with glutaric anhydride or other acylating agent or a sulfonating agent or a combination of the foregoing. The modified
collagen solutions can be prepared with a specific index of refraction to provide optimum visual acuity. The choice of chemical modifier(s) and the extent of modification will ultimately determine the indices of refraction as well as the biological
stability of-the resultant IOL.
The collagen-based preparations readily adhere to tissue, such as a lens capsule, and exhibit sufficient surface tension to prevent the solution from flowing out the capsular sac. Moreover, the injectable collagen solutions are transparent,
biologically stable, and appear to inhibit epithelial cell proliferation. No epithelial cell accumulation was noted after injection into the lens capsule in rabbit model studies.
Upon injection into the capsular sac, the collagen solution either remains in its original unpolymerized state or may undergo subsequent polymerization to form a gelatinous lens. Polymerization in the injection site may occur spontaneously or
can be initiated by chemical or enzymatic means, or by photoinitiation.
Accordingly, it is an object of the invention to provide a method for preparing IOLs using injectable biologically compatible, modified collagen compositions. The modified collagen compositions are viscous, but injectable, transparent and have a
predetermined refractive index. The range of the index of refraction is between about 1.2 and about 1.6. They are prepared by chemical modification of collagen derived from various animal sources with glutaric anhydride, other acylating agents,
sulfonating agents or combinations thereof. The refractive index may be modulated by choosing the appropriate modification agent.
It is another object of the invention to provide collagen-based IOLs to replace diseased or natural lenses following their removal. The IOL should be clear, transparent, biologically stable and be able to inhibit or stabilize lens capsule
epithelial cell accumulation. The IOL should exhibit refractive index similar to the natural lens (core-1.406 and cortex 1.386) or varied to correct various refractive errors following lens removal.
It is yet another object of the invention to provide collagen-based IOLs which can be used to replace an excised lens such that the IOL would not only correct any refractive errors of vision resulting from lens removal but would also allow the
lens system to continue to accommodate. The collagen-based IOL can be used in patients to treat visual defects resulting from cataracts, presbyopia, myopia, hyperopia and trauma.
These and other objects of the invention will be apparent in light of the detailed description below.
DESCRIPTION OF THE DRAWINGS
FIG. 1 illustrates that glutaric modified collagen solutions, prepared in accordance with Example 5, are psuedoplastic and thus exhibit shear thinning as shear rate increases. The rheometric analysis was performed on a Rheometrics System IV
rheometer (Rheometrics Piscataway, N.J.) using a 50 mm cone and plate at 0.04 radians. The temperature employed was 30.degree. C.
FIG. 2 illustrates that glutaric modified collagen solutions, prepared in accordance with Example 5, are viscoelastic. The G' component represents the elastic or storage modulus and the G" component represents the viscous or loss modulus. The
conditions employed are the same as described in FIG. 1.
FIG. 3 illustrates an absorbance spectrum of a purified, acid soluble collagen solution (collagen concentration of 30 mg/ml) undergoing limited fibrillogenesis. At T=0 the solution pH was adjusted to 7.2 using 2.5N NaOH and mixed well at
25.degree. C.
DETAILED DESCRIPTION OF THE INVENTION
All patents, patent applications, and literature references are hereby incorporated by reference in their entirety.
As defined herein, the term "injectable collagen composition" refers to an injectable, chemically modified, biologically compatible collagen composition which, when injected into an evacuated lens capsular sac, forms a prosthetic lens. The
injectable collagen composition fills the capsular sac and conforms to the dimensions of a natural crystalline lens which, in a human adult, measures approximately 4 mm in thickness and 9 mm in diameter. The term "biologically compatible" refers to
collagen modified in accordance with the present invention which when incorporated or implanted into or placed adjacent to the biological tissue of a subject, does not deteriorate appreciably over time or induce an immune response or deleterious tissue
reaction after such incorporation or implantation or placement.
The type of collagen useful in preparing the IOL of this invention is selected from the following groups: purified Type I collagen, Type IV collagen and Type III collagen, intact collagen-rich tissue or a combination of any of the foregoing.
Preferred as a collagen starting material is purified Type I collagen derived from animal tissue or predominantly Type I collagenous product prepared from human tissue. Type I collagen is ubiquitous and readily extracted from animal tissues such as
dermis and tendon. Common sources are bovine tendon and hide and rat tail tendon. Extraction from human tissues is difficult. U.S. Pat. No. 4,969,912, "Human Collagen Processing and Autoimplant Use", describes unique methods to disperse and
solubilize human tissue.
A variety of collagen solubilization procedures that are well known in the art can be used to prepare the modified collagen solutions useful for the instant invention. Native collagen is liberated from non-collagen connective tissue constituents
(lipids, sugars, proteins, etc.) and isolated after subjecting it to proteolytic enzymatic treatment by an enzyme other than collagenase. Suitable proteolytic enzymes include pronase and pepsin. The enzymatic treatment removes most of the immunogenic
non-helical portions of native collagen (telopeptide) and provides a monomeric collagen material which is soluble in dilute acidic aqueous media. A solution containing the crude solubilized collagen is then subjected to a series of treatments to purify
the soluble atelopeptide collagen by separating it from insoluble collagen, protease and noncollagen products resulting from the proteolytic enzymatic procedure. Conventional methods for preparing pure, acid soluble, monomeric collagen solutions by
dispersing and solubilizing native collagen are described, for example, in U.S. Pat. Nos. 3,934,852; 3,121,049; 3,131,130; 3,314,861; 3,530,037; 3,949,073; 4,233,360 and 4,488,911. A method for preparing solubilized collagen solution is provided in
the examples that follow.
The biological stability of the collagen based IOL of the invention appears to be affected by the solubility characteristics of the starting collagen as well as the extent of chemical modification. Completely solubilized modified collagen
generally does not produce an IOL that is resistant to high concentrations of neutral proteolytic enzymes under laboratory testing conditions. Hence, in practicing this invention, it is preferred that prior to chemical modification, a solubilized
collagen solution is converted to partially fibrillar collagen. Chemical modification of a partially fibrillar collagen solution results in a modified collagen composition which is clear, transparent and injectable. The use of partially fibrillized
collagen as the preferred starting material for the modification process results in an IOL with improved resistance to degradation by neutral proteolytic enzymes, such as trypsin.
To prepare a partially fibrillized collagen solution, a solubilized collagen solution is adjusted to pH between about 7.0 and 7.6, preferably about 7.4, and allowed to undergo limited fibrillogenesis at a temperature between about 25.degree. C.
and 40.degree. C., preferably about 37.degree. C. , for a period of between about 10 and 30 minutes, preferably about 20 minutes.
The extent of fibrillogenesis can be ascertained by measuring the increase in turbidity of the solubilized collagen solution by absorption spectroscopy (FIG. 3). In general, fibrillogenesis is permitted to continue until the turbidity of the
solution becomes about 20% to 60% greater, preferably about 25% greater, than the absorbance of the initial solution.
Without being bound to any mechanism or theory of collagen fibrillogenesis in this invention, it is believed that a portion of the solubilized collagen molecules undergoes self-assembly to form microfibrils which can be observed microscopically.
For a discussion of the fibrillogenesis process, see Nimni, M. E. "Collagen: Vol. I Biochemistry," CRC Press, Inc.; Boca Raton, Fla., USA (1988), pages 7-16 and Silver, F. H. in "Biological Materials: Structure, Mechanical Properties, and Modeling of
Soft Tissues", New York University Press, New York; N.Y. (1987), pages 137-163.
After chemical modification, the partially fibrillar collagen composition loses its turbidity and turns clear and transparent. Collagen microfibrils in the chemically modified partially fibrillar collagen solutions can no longer be observed
microscopically. The modified partially fibrillar collagen solution is believed to include modified collagen molecules and modified collagen aggregates containing collagen molecules.
In general, useful chemical modifier agents are those which react covalently with solubilized or partially fibrillar collagen to produce a modified collagen composition which is injectable, optically transparent, biologically stable and
compatible, and which resists epithelialization and degradation by proteolytic enzymes.
Depending on the choice of modifier and the extent of modification, chemical modification of collagen imparts an appropriate index of refraction to the composition. Other desirable features which may be imparted to the modified collagen
composition include UV blockage ability, and polymerizability.
Suitable acylating agents for use in the instant invention include aliphatic, alicyclic and aromatic anhydrides or acid halides. Non-limiting examples of acylating agents include glutaric anhydride, succinic anhydride, lauric anhydride,
diglycolic anhydride, methylsuccinic anhydride, methyl glutaric anhydride, dimethyl glutaric anhydride, succinyl chloride, glutaryl chloride, lauryl chloride, phthalic anhydride, methacrylic anhydride, trifluoroacetic anhydride, styrene/maleic anhydride
co-polymer, and ethylene/maleic anhydride copolymer. These chemicals are available from Aldrich Chemical Company (Milwaukee, Wis.). Preferred acylating agent for use in the present invention are glutaryl anhydride, methacrylic anhydride,
trifluoroacetic anhydride, ethylene/maleic anhydride copolymer, and phthalic anhydride. An effective amount of an acylating agent is broadly between about 0.5 and 20% wt total collagen, preferably between about 3 and 10% total collagen in solution.
Useful sulfonating agents for the preparation of modified collagen monomers of the present invention include aliphatic, alicyclic and aromatic sulfonic acids or sulfonyl halides. Non-limiting examples of sulfonating agents for use in the present
invention include anthraquinone-1, 5-disulfonic acid, 2-(chlorosulfonyl)-anthraquinone, 8-hydroxyquinoline sulfonic acid, 2-naphthalene- sulfonyl chloride, beta-styrene sulfonyl chloride, 2-acrylamido-2-methyl-1-propane sulfonic acid, aniline-2-sulfonic
acid, fluorosulfonylbenzene sulfonyl chloride, and poly (vinyl) sulfonic acid. These chemicals are also available from Aldrich Chemical Company (Milwaukee, Wis.). Preferred sulfonating agents for preparing the adhesive collagen materials are
beta-styrene sulfonyl chloride, and aniline-2-sulfonic acid. Such compounds, in non-toxic effective amounts, can be safely employed in collagen-based intraocular lens. An effective amount of sulfonating agent is broadly between about 0.5 and 20 wt % of
the total collagen, preferably between about 1 and 10 wt % of the total collagen in solution.
Non-limiting combinations of acylating agents and/or sulfonating agents include glutaric anhydride/beta-styrene sulfonyl chloride/methacrylic anhydride; glutaric anhydride/ethylene/maleic anhydride copolymer/methacrylic anhydride; glutaric
anhydride/polyvinyl sulfonic acid/methacrylic anhydride; and glutaric anhydride/ethylene/maleic anhydride copolymer/styrene/maleic anhydride copolymer. Preferred combinations for use in the present invention are glutaric anhydride/beta-styrene sulfonyl
chloride; glutaric anhydride/phthalic anhydride; and glutaric anhydride/aniline-2-sulfonic acid.
When combinations of two or more acylating agents, sulfonating agents, or mixtures of both agents are used for preparation of modified collagen composition, the total amount of chemical modifiers is preferably between about 3 and 10% wt of
collagen in solution. Excess quantities of chemical modifiers beyond the preferred range may result in a collagen composition that is biologically unstable and sensitive to tissue proteases.
Modification of collagen is carried out at alkaline pH, in a range between about 7.5 and 10.0, preferably between about 8.5 and 9.5, and most preferably at about pH 9.0. The acylation reaction can be monitored by the decrease in pH. The
reaction is terminated when the pH value remains stable at between about 5 and 8, preferably about 6.5 and 7.5. The reaction can also be monitored by removing aliquots and measuring the free amine concentration of the modified collagen solution as
compared to the starting solution of collagen.
The modification reaction should be complete in between about 5 and 90 minutes, preferably between about 20 and 40 minutes. The reactions should be carried out at temperatures between about 0.degree. C. and 37.degree. C., preferably between
about 4.degree. C. and 25.degree. C.
The reaction can be stopped by adjusting the pH to about 12.0 for about 2 minutes. This destroys residual, unreacted chemical modifiers. The modified collagen is then precipitated by reducing the pH using hydrochloric acid, acetic acid, nitric
acid, sulfuric acid, or other acid.
The amount of acid must be sufficient to precipitate out the chemically modified collagen. Generally precipitation occurs at a pH between about 3.5 and 6.0, preferably between about 4.0 and 5.0.
The precipitate of reacted collagen which now contains substituent groups reacted with amine groups (primarily epsilon-amino groups), is recovered from the mixture using conventional techniques such as centrifugation or filtration.
Centrifugation at between about 3,000 and 15,000 rpm for between about 20 and 60 minutes, preferably between about 4,000 and 12,000, for between about 20 and 30 minutes provides efficient recovery of the precipitate.
After recovery, the precipitate is washed with deionized water and subsequently dissolved in a physiological solution, e.g., phosphate buffer (0.1M) at about pH 7.2. It may be necessary to adjust the pH between about 7.0 and 7.5. This can be
done, for example, by the addition of sodium hydroxide solution.
Following dissolution of the precipitate, the solution is generally filtered by conventional filtering means, i.e. a 5 micron filter, and then centrifuged to remove air bubbles. At this point, the resulting solution containing chemically
modified collagen molecules and aggregates exhibits a viscous consistency, varying degrees of transparency and clarity, and a characteristic refractive index depending on the choice of chemical modifiers, the extent of acylation and on the state of
solubility of the starting collagen material.
The viscosity of the injectable modified collagen solution, determined at a temperature of about 25.degree. C., is broadly between about 30,000 centipoise and 300,000 centipoise, preferably between about 75,00 | | |
