EP0651650A1 - PROCEDE POUR TRAITER DES AFFECTIONS OPHTALMIQUES A L'AIDE DU FACTEUR TRANSFORMANT DE CROISSANCE-$g(b) - Google Patents

PROCEDE POUR TRAITER DES AFFECTIONS OPHTALMIQUES A L'AIDE DU FACTEUR TRANSFORMANT DE CROISSANCE-$g(b)

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Publication number
EP0651650A1
EP0651650A1 EP93917021A EP93917021A EP0651650A1 EP 0651650 A1 EP0651650 A1 EP 0651650A1 EP 93917021 A EP93917021 A EP 93917021A EP 93917021 A EP93917021 A EP 93917021A EP 0651650 A1 EP0651650 A1 EP 0651650A1
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EP
European Patent Office
Prior art keywords
tgf
retinal
neovascularization
ocular
eye
Prior art date
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EP93917021A
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German (de)
English (en)
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EP0651650A4 (fr
Inventor
Bert M. Glaser
Bruce B. Pharriss
Ann F. Hanham
George A. Ksander
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Celtrix Pharmaceuticals Inc
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Celtrix Pharmaceuticals Inc
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Publication of EP0651650A1 publication Critical patent/EP0651650A1/fr
Publication of EP0651650A4 publication Critical patent/EP0651650A4/fr
Ceased legal-status Critical Current

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/18—Growth factors; Growth regulators
    • A61K38/1841—Transforming growth factor [TGF]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A61P27/02—Ophthalmic agents

Definitions

  • This invention is a method for treating a variety of ophthalmic disorders in which wound healing is impaired or requires modulation, including macular holes, macular degeneration, retinal detachment and tears, retinal edema, retinal vascular disorders, retinal neovascularization, wound healing disorders, proliferative disorders, anti-degenerative disorders, anti-angiogenesis disorders, dry eye syndromes, uveitis, secondary cataracts, corneal epithelial wounds, corneal neovascularization, Sj ⁇ gren's syndrome, and surgical wounds.
  • the method entails application of Transforming Growth Factor-j8 (TGF-3) to the affected region.
  • TGF-32 is the preferred form of TGF-/3.
  • Other growth factors which have wound healing and neurotrophic effects may also be applied.
  • ophthalmic disorders affect the retina, lens and cornea.
  • retinal disorders are macular holes and degeneration, retinal tears, diabetic retinopathy, and miscellaneous disorders.
  • the most important disorders of the lens are cataracts and refractive errors.
  • the most important disorders of the cornea are those related to corneal defects, including corneal ulcers and wounds and the consequences of dry eye / Sj ⁇ gren's syndrome. These are discussed briefly below.
  • the retina is the light-sensitive portion of the eye. Supported by the choroid and retinal pigment cells and found at the posterior of the eye, the retina contains the cones and rods which detect colors. When the rods and cones are excited, they transmit signals which pass through successive neurons in the retina to the optic nerve and finally to the cerebral center, where a "visual picture" is integrated.
  • the macula lutea In the center of the retina is the macula lutea, which is about 1/3 to 1/2 cm in diameter.
  • the macula provides detailed vision, particularly in the center (the fovea) , because the cones are higher in density. Blood vessels, ganglion cells, inner nuclear layer and cells, and the plexiform layers are all displaced to one side (rather than resting above the cones) , thereby allowing light a more direct path to the cones.
  • the choroid Under the retina are the choroid, a collection of blood vessels embedded within a fibrous tissue, and the deeply pigmented epithelium, which overlays the choroid layer.
  • the choroidal blood vessels provide nutrition to the retina (particularly its visual cells) .
  • the retina may tear, form holes and separate from the underlying choroid.
  • macular holes which produce blurred central vision or metamorphopsia.
  • the cause of most macular holes is unknown.
  • trauma, cystic degeneration, and vitreoretinal traction have all been associated with hole formation.
  • Full thickness macular holes also appear following myopic degeneration, laser photocoagulation, lightning strike and pilocarpine administration. There also is a higher frequency of macular holes after cataract extraction. 24
  • the idiopathic senile macular hole is a disorder occurring generally in healthy women who are in their sixth decade of life or beyond.
  • the more severe holes involve the full thickness of the macula and are surrounded by a halo of retinal detachment.
  • In the early stage there may be a sudden decrease or distortion in vision. But early changes are difficult for physicians to spot. Patients may experience sudden vision changes or may not notice symptoms if the condition slowly evolves.
  • Partial holes that is, holes which are partial in depth or shape, having a new moon or horseshoe shape, are worth diagnosing early, particularly if an effective way to stop progression to macular holes were available.
  • retinal vascular and macular diseases which may exhibit voids, tears, or separations in the retina resulting from lack of fibrous or supporting tissue.
  • Age-related macular degeneration is the major cause of severe visual loss in United States citizens over the age of 55.
  • Most AMD patients have a build up of deposits within and under the retinal pigment epithelium in the macular region resulting in atrophy of the retina and the retinal pigment epithelium.
  • the retinal pigment cells are long-lived. They scavenge for photoreceptor discs from the rods and cones for years and accumulate intracellular wastes.
  • the incompletely digested residues reduce cytoplasmic space (Feeny-Burns, L. et al.. Invest. Qphthal. Mol. Vis. Sci. (1984) 25:195- 200) and interfere with metabolism.
  • Age-related macular degeneration is a sight-threatening disorder which occurs in either an atrophic or (less commonly) an exudative form.
  • AMD Age-related macular degeneration
  • blood vessels grow from the choriocapillaris through defects in Bruch's membrane, and in some cases the underlying retinal pigment epithelium (RPE) .
  • RPE retinal pigment epithelium
  • the retina may tear or separate from the choroid, and the choroid may rupture, for a wide variety of reasons.
  • Other situations in which tissue separation is observed include such widely disparate conditions as detachment of retina and pigment epithelium, degenerative myopia, as may be evidenced by visible breaks in Bruch's membrane (lacquer cracks) , acute retinal necrosis syndrome (ARN) , and traumatic chorioretinopathies or contusion (Purtscher's Retinopathy) .
  • retinal disorders include edema and ischemic conditions.
  • Macular and retinal edema are often associated with metabolic illnesses such as diabetes mellitus.
  • Retinal edema is found in a large percentage of individuals who have undergone cataract extraction and other surgical procedures upon the eye. Edema is also found with accelerated or malignant hypertension.
  • Macular edema is a common complication of prolonged inflammation via uveitis, Eales disease, or other diseases. Local edema is associated with multiple cytoid bodies ("cotton bodies") as a result of AIDS.
  • Retinal ischemia can occur from either choroidal or retinal vascular diseases, such as central or branch retinal vein occlusion, collagen vascular diseases and thrombocytopenic purpura. Retinal vasculitis and occlusion is seen with Eales disease and systemic lupus erythematosus.
  • Proliferative Diabetic Retinopathy PDR
  • the initiating event may be inadequate tissue oxygenation which causes vasodilation. Inadequate oxygenation may occur after the arterial basement membrane has thickened with diabetes-related deposits and because of endothelial cell proliferation, which is associated with pericyte degeneration. Basement membrane thickening and loss of pericytes are believed to result from low insulin and hyperglycemia, two important metabolic abnormalities of diabetes.
  • fibroblast growth factors FGF
  • TGF- ⁇ and ⁇ tumor necrosis factor
  • others have proposed that because retinal blood vessels appear to have a unique response to diabetic ischemia, there may be specific retina-derived growth factors.
  • Uveitis refers to inflammation of the uveal tract. It includes ulceris, cyclitis and iridocyclitis and choroiditis and usually occurs with inflammation of additional structures of the eye. These disorder has a variety of causes but is typically treated with systemic steroids, topical steroids or cyclosporin. The disease frequently presents with a chronic inflammation occurring either in the anterior segment (70%) or in the posterior segment (30%) which is complicated by episodes of severe exacerbation that may not be controllable with conventional medications. Reports in the literature suggest that 30,000 individuals become legally blind each year in the United States from uveitis. In addition, an estimated 20,000 individuals suffer significant loss of visual acuity from this disorder. Additional means to control this condition, without suppressing infection fighting abilities with steroids, would be highly beneficial. Cataracts
  • Cataracts are opacities in what should be perfectly clear lenses. Cataracts interfere with the vision by causing blurred vision, glare, altered color perception and monocular diplopia. They are related to a variety of factors, including x-ray exposure and metabolic diseases such as diabetes, Wilson's disease (copper accumulation) and galactosemia. Cataracts are also a side effect of cortisone, methotrexate and nitrogen mustard therapy.
  • the cornea and conjunctiva are vulnerable to damage from pathogenic agents or direct trauma, drying associated with disorders of tearing, exposure to radiant energy (ultraviolet light, sun and welding guns) , allergens such as pollen and mold, and infectious agents. Keratoconjunctivitis can also occur in patients with Stevens-Johnson syndrome, Wegener's granulomatosis, rheumatoid arthritis, atopic dermatitis and cicatricial pemphigoid. Corneal ulcers may occur.
  • corneal surgery After corneal surgery, the cornea must heal.
  • Popular types of corneal surgery include cataract extraction, with or without lens replacement; corneal transplants, to treat viral infection or penetrating keratoplasty (PKP) ; glaucoma filtration surgery; and radial keratotomy and other types of surgery to correct refraction.
  • PGP penetrating keratoplasty
  • Cataract incisions are full thickness wounds in the cornea which are as large as 8 mm in length with conventional intraocular lenses (IOLs) and as small as 3 mm or less with foldable silicone IOLs. These wounds typically heal without difficulty, although they take several months to stabilize and are associated with warpage of the corneal tissues leading to permanent astigmatism. Treatment which could speed stabilization of vision and avoid astigmatism would be highly desirable.
  • Penetrating keratoplasty and corneal transplant are characterized by full-thickness wounds around the entire circumference of the cornea. These wounds tend to remain weak for one or more years. Patients experience drift in visual acuity and increasing risk of wound dehiscence and/or endophthalmitis. It would be highly desirable to stabilize visual acuity and accelerate wound maturation as early as possible, to avoid sight-threatening adverse effects.
  • Radial keratotomy is the most widespread technique for altering the shape of the cornea. The most commonly used form of RK is based on the placement of 4-8 surgical incisions in a radial pattern across the cornea. These incisions are typically 70-80% of the depth of the cornea, and are therefore non-penetrating wounds.
  • Sj ⁇ gren's syndrome is an immune system disorder which manifests itself in the eyes as conjunctival and corneal dryness (keratoconjunctivitis sicca syndrome) and a gritty sensation in the eyes. This is due to lack of tear resulting from destruction of the lacrimal (or tear) glands by progressive mononuclear cell infiltrate and scarring of the gland. If the cornea is too dry, corneal ulcerations can develop. "There is currently no effective treatment for the ongoing exocrine gland destruction. Treatment is geared toward symptomatic relief of mucosal dryness . . . and includes artificial tears [and] ophthalmologic lubricating ointments.” Harrison's Principles of Internal Medicine. 12th ed. , McGraw-Hill, pages 1449-50, 1991.
  • Neovascularization is a serious complication of a large variety of ocular disorders affecting the various tissues of the eye because it can lead to blindness. Corneal neovascularization occurs in many conditions and diseases, including trauma, chemical burns and corneal transplantation.
  • Corneal transplantation is successful in many patients because of the absence of blood vessels in the corneal tissue. Because there are no blood vessels in the cornea, the circulating components of the immune system are not exposed to the new cornea and there is normally no problem of host-graft rejection. Induction of neovascularization in the cornea would expose the cornea to the immune system and lead to graft rejection. In addition, a subsequent graft is less likely to be successful, too.
  • Treatments of these various causes of neovascularization may include the administration of immunosuppressives to modulate the inflammatory process, including neovascularization. However, immunosuppressives may inhibit appropriate wound healing in the cornea and interfere with the ability to fight infections. Delayed wound healing leaves the cornea vulnerable to infections for longer periods. Hence, vision-threatening infections can result from current treatments.
  • Neovascularization of the iris, and its attendant scarring can result in glaucoma and blindness. Neovascularization of this portion of the eye can arise as a consequence of diabetic retinopathy, venous occlusion, ocular tumors and retinal detachment. Most commonly, laser treatment to cauterize the blood vessels is tried; however, that has the attendant risk of causing additional scarring.
  • Retinal and intravitreal neovascularization occurs in a wide range of disorders including diabetic retinopathy, vein occlusions, sickle cell retinopathy, retinopathy of prematurity, retinal detachment, ocular ischemia and trauma.
  • Subretinal pigment epithelial (RPE) and sub- retinal neovascularization are common, yet very severe, disorders of the eye.
  • the growth of new blood vessels interferes with the normal anatomy of the visual and pigmentary cells in the eye, leading to severe visual loss.
  • the new blood vessels leak fluid and blood under the macula causing marked distortion and loss of vision.
  • these blood vessels develop in the avascular foveal region of the eye, the result is central visual loss and legal blindness.
  • Choroidal neovascularization is caused by such retinal disorders as age-related macular degeneration, presumed ocular histoplasmosis syndrome, myopic degeneration, angioid streaks and ocular trauma. Macular degeneration was discussed above. Choroidal neovascularization has proven recalcitrant to treatment in most cases. In only 10% of cases can laser photocoagulation be attempted. There is no other treatment available. Even with successful laser photocoagulation, neovascularization recurs in about 60- 70% of eyes.
  • TGF-3 The family of peptides known as TGF-3 can both regulate cell growth and differentiation. These polypeptides can both stimulate and inhibit cell proliferation depending largely on the cell type and environment. TGFs of some type have been found in almost all tissues from all species of animals which have been examined so far.
  • TGF-/32 is a well-characterized material. As noted above, it is a polypeptide and has a molecular weight of about 25,000 D and is a dimer composed of two 12,500 D subunits which are linked by a disulfide (Chiefetz et al.. Cell (1987) 4.8:408-415; Ikeda et al.. Biochemistry (1987) 26:2406-2410) and has been isolated from bovine demineralized bone (Seyedin et al., J. Biol. Chem. (1987) 262:1946-1949) .
  • porcine platelets (Cheifetz et al., Cell (1987) 8:409-415), human prostatic adenocarcinoma cell line, PC-3 (Ikeda et al., 1987, Biochemistry 26:2406-2410). etc.
  • Methods for separating and purifying TGF-02 are given in U.S. Patent 4,774,322 to Seyedin et al.
  • TGF-31 and TGF-32 are found in many of the same cells. However, their mature sequences have only about 75-80% homology (Derynck et al., EMBO J. (1987) 2: 3737- 3743) . It has been established that the several species of TGF-S are coded for by different genes. (Madisen et al., DNA (1988) 7: 1-8) It has been observed that TGF- ⁇ (only TGF-31 is mentioned in the article's materials section) appeared to seal the edge of surgical retinotomy in rabbits (See, Smiddy et al., "Transforming Growth Factor-j8—A Biologic Chorioretinal Glue", Arch. Qphthal. Mol. (1989) 107:577- 580) . Smiddy et al. showed the formation of fibrotic tissue around the retinotomy which sealed the retina to the choroid layer.
  • TGF-/32 has been found to stimulate collagen glycoprotein synthesis as well as cellular proliferation and migration involved in the wound healing process.
  • TGF-3 has been found to inhibit [ 3 H]thymidine incorporation by retinal pigment epithelial cells which is stimulated by platelet-derived growth factor, a-FGF, b-FGF and EGF. According to Leschey, this could be due to TGF-/3 being linked to a strong inhibitory pathway capable of overriding stimulatory signals from other growth factors. Leschey et al., Invest. Qphthalmol. Vis. Sci. (1990) 3 ,:839-46.
  • TGF-3 positively modulates the bioactivity of FGF in corneal endothelial cells.
  • This invention is a method of significantly improving the ocular vision in retinal disorders of the mammalian eye, in which the retinal disorders are characterized by a loss or impending loss of fibrous tissue, and in which the method comprises administering to the mammal about 1 to 10 ⁇ g of TGF-/3.
  • the type of TGF-0 is TGF-02.
  • TGF-j8 is administered by intraocular, subretinal, subscleral, intrascleral, intrachoroidal and subconjunctival injection or by topical, oral or parenteral modes of administration.
  • the method comprises two additional preceding steps: removing the vitreous humor from the eye; and peeling the epiretinal membrane, if present, from the retina.
  • TGF-3 is administered in an effective amount as a concentrated solution by cannula to the portion of the retina requiring treatment.
  • the retinal disorder to be treated is a macular hole.
  • a method of maintaining or improving the ocular vision in macular degeneration The method calls for administering to the mammal an amount of TGF-/3 effective to stabilize or improve vision.
  • a method of maintaining or improving the ocular vision in cystoid macular edema The method calls for administering to the mammal an amount of TGF-3 effective to stabilize or improve vision.
  • the method of treating retinal disorders comprises the steps of removing the vitreous humor from the eye; removing the epiretinal membrane, if present, from the eye; and administering a concentrated solution of TGF-jS by cannula to place the TGF-/3 solution immediately over the portion of the retina requiring treatment.
  • a method for treating an individual with an ophthalmic disorder or poor healing in the eye which can benefit from the administration of TGF-3 orally, topically or systemically to an individual in need of such treatment.
  • TGF- ⁇ is administered in an amount sufficient to improve healing.
  • the ocular disorder is selected from the group consisting of retinal and corneal wounds, macular degeneration, secondary cataracts, corneal disease and dry eye / Sj ⁇ gren's syndrome.
  • TGF-3 is administered by intraocular injection or by application to the cornea.
  • TGF- ⁇ can be applied to the cornea by means of eyedrops or a timed release capsule placed in the cul de sac.
  • the method provides for administration of TGF-3 in an amount sufficient to promote healing and reduce symptoms associated with poor healing.
  • the amount of TGF-3 administered is at least about 0.5 to 50 ⁇ g of TGF-3 per treated eye.
  • a method for treating a mammal who has undergone or is about to undergo or is undergoing ophthalmic surgery to promote healing without excessive scarring comprising administering to said mammal TGF-/3 in an amount sufficient to promote healing without excessive scarring.
  • the ophthalmic surgery can be, but is not limited to, cataract extraction, with or without lens replacement; corneal transplants, to treat viral infection or penetrating keratoplasty (PKP) ; glaucoma filtration surgery; and radial keratotomy and other types of surgery to correct refraction.
  • a method for treating a mammal for ocular neovascularization comprising administering to a mammal an effective amount of TGF-/32.
  • a method for treating a mammal for uveitis in which the method comprises injecting an effective amount of TGF-3 intraocularly.
  • the Inventors propose that the administered TGF-/3 aids healing by modulating a fibrotic response in the ocular tissues.
  • TGF- ⁇ aids healing by modulating a fibrotic response in the ocular tissues.
  • application of an effective dose of TGF-3 to the retina appears to provide positive neural regenerative effects as evidenced by the fact that such TGF- ⁇ application significantly improves the eyesight of the individuals having the macular holes.
  • the method of this invention is suitable for the treatment of ophthalmic disorders, particularly retinal disorders involving macular degeneration, neovascularization, holes, separations, tears, and the like in the retina or between the retina and its underlying choroidal tissue, or involving choroidal tissue, as described above.
  • Optid disorder refers to physiologic abnormalities of the eye. They may involve the retina, the vitreous humor, lens, cornea, sclera or other portions of the eye, or physiologic abnormalities which adversely affect the eye, such as inadequate tear production.
  • Retinal wounds include, but are not limited to, tears and holes in the retina and separation from the underlying choroid. Retinal wounds appear after trauma, cystic degeneration, vitreoretinal traction, myopic degeneration, laser photocoagulation, lightning strike, pilocarpine administration and cataract extraction. To help the retina heal in a modulated process, TGF-/3 can be administered.
  • Macular degeneration is characterized by the excessive buildup of fibrous deposits in the macula and retina and the atrophy of the retinal pigment epithelium.
  • the administration of TGF-3 can help promote healing of the atrophied retinal pigment epithelium in a controlled fashion, which is designed to limit excessive fibroproliferation that may occur without such treatment.
  • Secondary cataracts are opacities in the ocular lens which interfere with vision. Secondary cataracts occur after x-ray exposure, in diabetes, Wilson's disease and galactosemia, and as side effects in drug therapy. TGF-3 can be used to promote healing of the lens after damage in a modulated fashion which is designed to limit hyperproliferation which can occur naturally.
  • diseased corneal tissue includes damage to the cornea by a variety of causes including, but not limited to, trauma, dry eyes (in which the conjunctiva on the inside of the eyelid may abrade the cornea) , excessive light, allergens and infectious agents.
  • TGF-3 can be used to promote gradual healing of diseased corneal tissues and avoid excessive scarring which can interfere with vision.
  • Sj ⁇ gren's syndrome is an autoimmune disorder which frequently is characterized by dry eyes, due to destruction of the tear glands by the autoimmune process.
  • TGF- ⁇ can be used to control at least the ocular manifestations of Sj ⁇ gren's syndrome. While not wishing to be bound by any particular theory, it appears that first TGF-3 can promote gradual healing without scarring of the tear gland and that second, TGF-3 also promotes healing of corneal epithelial wounds which arise from the dry eye syndrome caused by lack of tear glands.
  • “Ocular neovascularization” is herein defined as the unwanted new growth of blood vessels into the ocular tissues. Unchecked, such growth can result in blindness.
  • the ocular tissues which can be invaded by neovascularization include the cornea, iris, retina, vitreous, and choroid. Many diseases and conditions cause or contribute to ocular neovascularization.
  • causes of corneal neovascularization include but are not limited to trauma, chemical burns or corneal transplantation.
  • causes of neovascularization of the iris include but are not limited to diabetic retinopathy, vein occlusion, ocular tumor and retinal detachment.
  • causes of retinal and intravitreal neovascularization include but are not limited to diabetic retinopathy, vein occlusion, sickle cell retinopathy, retinopathy of prematurity, retinal detachment, ocular ischemia and trauma.
  • causes of choroidal neovascularization include but are not limited to retinal disorders of age-related macular degeneration, presumed ocular histoplasmosis syndrome, myopic degeneration, angioid streaks and ocular trauma.
  • Treating a mammal for ocular neovascularization is herein defined as treating ocular neovascularization which has already become detectable.
  • “Mammals” are defined as humans and mammalian farm and sport animals and pets. Farm animals include, but are not limited to, cows, hogs, and sheep. Sport animals include, but are not limited to, dogs and horses.
  • the category pets includes, but is not limited to, cats, dogs, and hamsters.
  • the method can involve the placement of at least an effective amount of a growth factor such as TGF- ⁇ , preferably TGF-/S2, on the ophthalmic abnormality.
  • TGF-/3 or TGF-32 are placed on the macular hole itself and/or the edges of the macular hole.
  • Such treatments provide improvement of vision and healing by decreasing the thickness of the edge of the hole.
  • the edges of the hole appear to adhere to choroid or reconnect with the posterior hyaloid membrane.
  • use of the growth factor on other retinal abnormalities is effective.
  • TGF-32 is applied using known surgical techniques, such as those described in the example which follows. It is desirable that the TGF-/32 stay in place for a substantial period of time after application. For instance, a day is typically considered adequate for this purpose. To help retain
  • TGF-/S2 in place, known pharmaceutic combinations may be used.
  • Hyaluronic acid is typically used in the eye for this purpose; however, as indicated by the data below, hyaluronic acid does not appear to increase treatment effectiveness.
  • a TGF-3 dose of at least 1000 ng is preferred for at least partial alleviation of macular hole detachment. More preferred is a TGF-/3 dose of at least about 1300 ng. These doses appear to be the approximate dosage for improvement of vision (at least two lines on the Snellen Vision Chart) .
  • suitable ophthalmic carriers such as normal saline solution
  • This inventive treatment is applicable to retinal disorders, particularly to macular degeneration and holes, where it promotes healing and significantly improves vision.
  • the treatment also may be used on peripheral retinal holes and tears.
  • the formulation, method of administration and dosage will depend upon the disorder to be treated, the point at which the disorder is being treated, and perhaps other aspects of the medical history of the patient. These factors are readily determinable in the course of therapy. Suitable patients with an ophthalmic disorder can be identified by medical history, physical findings and laboratory tests. The medical history reveals such facts as time of onset of symptoms such as red sclera, pain, photophobia, dry or gritty eyes, and vision changes, such as blurred vision not correctable with eyeglasses and double vision in an eye. Patients sometimes complain of inability to engage in their usual activities, such as watching television or driving a car at night.
  • Patients with ophthalmic disorders associated with impaired healing may have physical findings such as injected sclera, cotton-wool spots on the retina, a macular hole, bleeding behind the retina.
  • Indicative laboratory results include low levels of TGF-3 in the serum or in eye tissues, such as the vitreous.
  • TGF-3 may be administered by any of a variety of routes known in the art, including but not limited to, intraocular, subretinal, subscleral, intrascleral, intrachoroidal, and subconjunctival injection, depending on the nature and location of the pathology being treated. Also contemplated in the present invention are administration by intravenous injection, subcutaneous injection, or oral administration, provided that sufficient TGF-3 reaches the condition being treated. In one preferred embodiment, a concentrated solution of TGF- ⁇ is injected into the eye and placed immediately over the lesion, for example, on the retina.
  • TGF- ⁇ may be administered in any pharmaceutically acceptable formulation, including, but not limited to, solutions, suspensions, and timed-release preparations, such as microcapsular particles and implantable articles.
  • the preferred dose is greater than about 1000 ng (measured at an absorbance of 210 or 280 nm wavelength) .
  • all weights of TGF- ⁇ are based on measurements performed at 210 or 280 nm wavelength. More preferably, the dose is about 1300 ng. For some conditions, the preferred dose is about 2500 ng.
  • TGF-/32 may be administered in a slow-release device embedded in the tissue stroma or in a compartment adjacent to the affected tissue.
  • TGF-3 in a pellet of ethylene vinyl copolymer 2 mm in diameter could be surgically implanted in the vitreous cavity or suprachoroidal space to release TGF-/3 over time.
  • This modality is believed to be particularly beneficial for neovascularization of the iris or choroid.
  • Patients at risk for ophthalmic healing problems include those who have undergone or about to undergo surgery. Examples of such surgery include, but are not limited to.
  • Cataract extraction with or without lens replacement
  • Corneal transplant for treating viral infections or penetrating keratoplasty (PKP) ;
  • TGF-/3 promotes prompt, gradual healing without excessive fibrous tissue formation.
  • growth factors which have both wound healing and neurotrophic effects can be applied in certain of these inventive treatments. These factors include, but are not limited to, acidic and basic fibroblast growth factor, insulin, insulin-like growth factor, platelet-derived growth factor, nerve growth factor, epidermal growth factor, transforming growth factor ⁇ , colony-stimulating factor, keratinocyte growth factor, and tissue plasminogen activator.
  • EXAMPLE 1 In this example, the effectiveness of TGF-32 in alleviating macular holes is shown. Clinical data are provided in a table which follows this example.
  • Each macular hole was graded as Stage 2, 3, or 4 according to the criteria described by Gass (Arch. Qphthalmol. (1988) 106:629-39) . Briefly, eyes with Stage 2 holes have a retinal dehiscence along the margin of the area of deep retinal cyst formation. In Stage 3, typically there is a full-thickness hole with overlying operculum. Macular holes are classified as Stage 4 when a posterior vitreous detachment is present. Treatment was scheduled within 2 weeks of the baseline examination. Under the criteria, patients were excluded if they had greater than 2+ nuclear sclerotic or posterior subcapsular lens changes. Patients were followed for 6-10 months, with mean follow-up of 8 months. Treat ent Groups
  • Doses of 70 ng, 330 ng, and 1330 ng of TGF-02 were administered.
  • the 70 ng dose was chosen to provide a negative control for the higher doses.
  • the 330 ng dose was believed to be at the low end of the effective range, and the 1330 ng dose was believed to be well within the effective range.
  • an epiretinal membrane was found; however, no definite edges of this membrane could be found. Where encountered, the epiretinal membrane was peeled from the surface of the retina and removed from the eye. In other cases, no definite epiretinal membrane could be found; however, there appeared to be some gelatinous condensation on the inner surface of the retina surrounding the macular hole for approximately 200-400 ⁇ , with a firm adhesion along the margin of the macular hole. This was carefully dissected where possible; however, great care was exercised in order to limit traction on the edges of the macular hole or damage to the nerves. After a short period of time to allow peripheral fluid to drain posteriorly, the fluid that had migrated posteriorly was aspirated. Typically, approximately 0.1-0.5 cc of fluid had reaccumulated on the retinal surface during this time. The center of the macular hole was gently aspirated to remove the last remaining amounts of fluid in the region of the macular hole.
  • TGF-/32 was thawed and mixed within 2 hours of use.
  • the TGF-32 was supplied by Celtrix Pharmaceuticals, Inc., Santa Clara, CA, and kept at -70°C until ready for use. It was always used within 2 hours of thawing and kept on ice until used.
  • the TGF-32 was highly purified (greater than 95% purity) , and derived from bovine bone.
  • the reconstituted formulation contained either 70, 330, or 1330 ng/0.1 cc of TGF-/32 after dilution with a diluent solution.
  • Eyes were randomly assigned a dose of TGF-02. About 0.1 cc of TGF- ⁇ 2 solution was gently infused into the macular hole. In about 50% of eyes, a comparable volume of hyaluronic acid was also introduced in order to determine if this might maintain the presence of the TGF-32 solution, thereby improving efficacy. After surgery, the patient was instructed to lie in a supine position for the first 24 hours following surgery; thereafter, the patient was instructed to remain in a face-down position as much as possible over the ensuing two weeks. After surgery, patients were examined at 1 day,
  • Fluorescein angiography was performed at 4 to 6 weeks, 3 months, and 6 months. Best corrected Snellen visual acuity, intraocular pressure, lens status, bubble size, status of macular hole, and occurrence of adverse effects were determined at each examination.
  • Treatment effects were assessed using logistic regression.
  • the dependent variable was improvement in visual acuity of two or more lines on the Snellen Chart, and the independent variables were TGF-/32 dose and hyaluronic acid use.
  • the posterior hyaloid surface was completely separated from the retina in all eyes with Stage 2 and Stage 3 macular holes. However, in all eyes with Stage 2 macular holes, this separation of the posterior hyaloid surface from the retina extended marginal dehiscence and formed an operculum that was elevated from the posterior hyaloid surface. In one eye, the retina was torn in the inferotemporal quadrant between the ora and the equator and accompanied by an adjacent intraretinal hemorrhage. This retinal tear was treated without consequence with transscleral cryopexy.
  • the anterior chamber had only trace amounts of flare and cell in all eyes except six.
  • a fine, red-brown precipitate was found on the endothelial surface of the cornea along with minimal striae.
  • the precipitate and striae resolved within two weeks without sequelae. All eyes with intact lenses had mild to moderate posterior lens feathering which resolved within two weeks.
  • the macular region could be adequately examined using a biomicroscope with a contact lens or a 78 diopter lens.
  • Microscopic retinal detachment and retinal thickening surrounding the macular hole could be readily assessed at this time.
  • flattening of the detachment and thinning of the adjacent retina to a normal-appearing thickness occurred in 12/12 eyes treated with 330 ng of TGF-02 without hyaluronic acid and 11/11 eyes treated with 1330 ng of TGF-/32 without hyaluronic acid.
  • only 6/11 eyes treated with 70 ng TGF-32 without hyaluronic acid had the edges of the macular hole flatten after 4-6 weeks.
  • hyaluronic acid significantly suppressed the rate of flattening of the retina around the macular hole. Visual acuity did not improve in eyes with no improvement in retinal flattening. Final visual acuity improved two lines or more in 10/11 eyes treated with 1330 ng TGF-/32 without hyaluronic acid, 4/12 eyes treated with 330 ng TGF-j32 without hyaluronic acid, and 5/11 eyes treated with 70 ng TGF-02 without hyaluronic acid.
  • fluorescein angiography revealed a central hyperfluorescent window defect corresponding to the base of the macular hole in most eyes.
  • this method limits manipulation of the retina and induces chorioretinal adhesion using TGF-3.
  • the edges of the macular holes were flattened in 23/23 (100%) eyes treated with 330 or 1330 ng TGF-02 without hyaluronic acid.
  • a fine bead of fibrous tissue could be observed along the margin of the macular hole after it flattened. This fibrous tissue was accompanied with good visual recovery (vision improvement of two or more Snellen lines) .
  • TGF- ⁇ 2 might cause excessive fibrosis which can increase macular contraction and result in proliferative vitreoretinopathy (PVR) .
  • PVR proliferative vitreoretinopathy
  • TGF-32 is known to be present in significant concentration in eyes with PVR, and has been implicated in its formation (See, Connor, J. Clin. Invest. (1989) 83.H661-66). However, the concentration of TGF-32 used in these cases was significantly less than that seen in eyes with PVR. Notably, neither of these potential complications was seen in any of the 60 eyes treated; and all eyes were observed for at least six months after treatment. Visual acuity improved in eyes with flattened edges of the macular hole, resorption of subretinal fluid, and thinning of the adjacent retina in response to
  • TGF- / 32 also enhanced recovery of the photoreceptor outer segment function, possibly by neural regeneration or stimulation of accessory tissues which in turn help stabilize and align neural retinal cells.
  • EXAMPLE 2 In this example, treatment of AMD is described.
  • the therapeutic efficacy and safety of locally administered bovine TGF-02 (bTGF-B2) was comparison to a placebo or no treatment in patients with exudative age-related macular degeneration and visual acuity of 20/160 or better.
  • the primary efficacy variable was change in visual acuity from baseline in the respective treatment groups.
  • the secondary efficacy variable was stabilization (within 2 lines on ETDRS) or improvement
  • At least one high risk characteristic a. subretinal serous exudate b. subretinal hemorrhagic exudate c. presence of lipid 4. Best corrected vision 20/160 or better as measured by ETDRS (Early Treatment Diabetic
  • Each patient was randomly assigned to one of three groups. Stratification of patients at baseline was performed to ensure that all groups were represented with minimum bias. The stratification parameters included visual acuity (20/100 mid point) , age (70 years) , and visual loss duration (3 months) .
  • the patients in Group I received 50 ⁇ l (665 ng) of bTGF-B2 subretinally in the area of the neovascular net, with another 150 ⁇ l (1995 ng) of bTGF-B2 applied directly onto the fovea at its interface with the vitreous cavity.
  • Subjects assigned to Group II were treated in the same manner as those in Group I except that they received a placebo (vehicle) solution identical to the drug product but without bTGF-B2, while those in Group III were untreated.
  • the amount of serous and hemorrhagic exudate, size of the neurosensory detachment, presence of epiretinal membrane, presence and size of hyperfluorescence from classic/ occult neovascularization, total lesion size and foveal involvement were also measured on fluorescein angiography and ICG (indocyanine green) angiography.
  • bTGF-B2 extracted from bovine bone in a highly purified form (>95% purity) and supplied as a concentrated acidic solution (Vial 1, MS 2004) which was mixed with diluent containing human serum albumin (Vial 2, MS 2005) prior to subretinal or intravitreal injection.
  • the final concentration of bTGF-B2 following mixing with the diluent was 2660 ng per 200 ⁇ l volume of solution.
  • the placebo preparation was an acidic solution without bTGF-B2 (Vial 1, MS 2009) which was mixed with diluent (Vial 2, MS 2005) prior to subretinal or intravitreal injection.
  • the total dose administered was 200 ⁇ l (2660 ng) of bTGF-/32 (Group I) , or 200 ⁇ l of placebo (Group II) .
  • 50 ⁇ l was injected into the subretinal space through a site at least one disc diameter from the center of the fovea, and 150 ⁇ l applied directly onto the retinal surface with underlying neovascularization.
  • One treatment was given following vitrectomy, except for Group III patients who received neither a vitrectomy nor the experimental or placebo solutions (i.e. will remain untreated).
  • Baseline findings not consistent with the requirements of the study were cause for discontinuation of the patient from the study.
  • Baseline studies included the following. A complete medical and ophthalmologic history were obtained.
  • Stereo angiographic views were taken during the transit phase, and at 30, 40, 60, 90 seconds, 2, 3, 5, and 10 minutes centered on field II. At 10 minutes there were also a stereo view of field I of the eye. Additional views of the opposite eye and of other fields were obtained at the discretion of the treating ophthalmologist.
  • the ICG angiogram were taken using a similar protocol, but ICG views were taken at 40 minutes as well. All study photographs and angiograms were labeled with the patient's code at the clinic. Photographs and angiograms were read by an observer masked with respect to patient information and randomization code.
  • Patients eligible for the study were randomly assigned to treatment groups, using a computer program to stratify on the basis of baseline visual acuity (20/100) , age (70 years) and visual loss duration (3 months) .
  • the treatment of Group I and II patients was scheduled within one week (seven days) of the date the baseline tests are completed. All surgeries were performed under either local anesthesia with sedation or general anesthesia. After the eye was prepped and draped, a lid speculum was positioned. Two conjunctival flaps were made laterally and medially. A 3 mm infusion cannula was to be placed 4 mm posterior to the limbus, and held in place with a pre-placed 4-0 white silk mattress suture.
  • the tip of the cannula was seen within the vitreous cavity prior to the onset of infusion.
  • Two additional scleroto ies were made at 10 o'clock and 2 o'clock meridians, 4 mm posterior to the limbus.
  • a light pipe and vitreous cutter were then introduced.
  • a core vitrectomy was performed in the involved eye.
  • the vitreous cutter was removed and replaced with a cannula having a flexible silicone tip.
  • the cannula was then connected to the aspiration system of the vitrectomy machine and aspiration set at 150 mm Hg.
  • the infusion bottle was positioned approximately 50 cm above the level of the patient's head.
  • the tip of the cannula was positioned approximately 1 mm above the surface of the retina just below the superotemporal arcade. Full aspiration was applied, and the cannula gently elevated. After the posterior hyaloid surface was elevated in the area just inferior to the superotemporal arcade, the cannula was used to extend this posterior hyaloid detachment as far as possible out to the equator. In some cases, additional manipulation was needed at the disc in order to complete the detachment of the posterior hyaloid surface. Once this was accomplished, the vitreous cutter was introduced and total pars plana vitrectomy performed, removing the vitreous as far out to the periphery as possible.
  • the flexible-tipped cannula was then reintroduced and again positioned approximately 1 mm above the retinal surface, and gently moved side to side. If the entire posterior hyaloid was removed, there was no bending of the cannula tip.
  • the instruments were then removed from the eye, and replaced with scleral plugs.
  • the peripheral retina were examined with indirect ophthalmoscopy and scleral depression in order to ensure that no retinal tears occurred.
  • the scleral plugs were then removed, and the light pipe and flexible-tipped cannula were reintroduced.
  • a fluid-air exchange was performed aspirating all fluid over the optic disc.
  • the instruments were then removed from the eye and replaced with scleral plugs. Fifteen minutes were allowed for peripheral fluid to drain posteriorly.
  • the scleral plugs were once again removed, and the light pipe and flexible-tipped cannula were reintroduced. Additional fluid that migrated posteriorly was aspirated.
  • the patient received acetazolamide, 500 mg intravenously (IV) and continued on acetazolamide 250 mg by mouth or IV every six hours, for the next 24 hours.
  • the patient was instructed to lie in a supine position for the first 24 hours following surgery; thereafter to remain in a facedown position as much as possible over the ensuing five days.
  • followup on day 1 and week 1 included intraocular pressure, the size of the remaining gas bubble, the appearance of the lens, adverse events. concomitant medications.
  • Post-Treatment included best corrected visual acuity for both distance and near vision using the EDTRS eye chart and Bailey-Lovie word chart, respectively; refraction; intraocular pressure; presence of epiretinal membrane; results of fluorescein angiography (3, 6 and 12 months only); results of ICG angiography in selected patients; lens status; occurrence of adverse events; and assessment of concomitant medications.
  • EXAMPLE 3 In this example, a process for the treatment of subretinal pigment epithelial (sub-RPE) or subretinal neovascularization using TGF-/32 is described.
  • sub-RPE subretinal pigment epithelial
  • the study involves 50 patients who satisfy the study criteria for sub-RPE or sub-retinal neovascularization. Patients are chosen according to the following criteria:
  • Baseline studies include visual acuity measurement (standardized Snellen and ETDRS eye charts) and biomicroscopy, as well as both fluorescein and ICG angiography to document the presence of the sub-RPE or sub-retinal neovascularization. After treatment, patients are followed for one year.
  • Safety and efficacy assessments include visual acuity measurements, biomicroscopic visualization of the fovea, and fluorescein and ICG angiography.
  • Surgical Procedures All surgery is performed under either local anesthesia with sedation or general anesthesia. Two conjunctival flaps are made laterally and medially. Two additional sclerotomies are made at 10 o'clock and 2 o'clock meridians, 4 mm posterior to the limbus. A light pipe and vitreous cutter are then introduced. At this point, a core vitrectomy is performed in the involved eye. After the core vitrectomy, the vitreous cutter is removed and replaced with a cannula having a flexible silicone tip. Then the cannula is connected to an aspiration system. The tip of the cannula is inserted and positioned approximately 1 mm above the retinal surface but below the superotemporal arcade.
  • the posterior hyaloid surface is elevated in the area just inferior to the superotemporal arcade, the posterior hyaloid is detached as far as possible out to the equator. In some cases, additional manipulation is needed at the disc in order to complete the detachment of the posterior hyaloid surface. Once this is accomplished, a total pars plana vitrectomy is performed by removing the vitreous as far out to the periphery as possible. The retina is examined to assure that no retinal tears have occurred.
  • TGF-32 Freshly thawed TGF-32 is suspended in buffer containing 2% human serum albumin for a concentration of 1330 ng/100 ⁇ l TGF-/32 solution.
  • TGF-02 solution (either 50 ⁇ l or 200 ⁇ l) is applied to the area of neovascularization.
  • TGF-02 solution (either 50 ⁇ l or 200 ⁇ l) is applied to the area of neovascularization.
  • TGF-/32 a bent, tapered 33-gauge cannula is used to enter the subretinal space at a site at least one disc diameter from the center of the fovea.
  • gentle injection of 50 ⁇ l containing 1330 ng/ 100 ⁇ l TGF-32 is performed.
  • the additional 150 ⁇ l of TGF-02 is injected within the vitreous cavity just over the area of neovascularization.
  • the conjunctival flaps and sclerotomies are closed.
  • the intraocular pressure is checked and the intravitreal bubble adjusted to achieve normal pressure.
  • the patient receives acetazolamide, 500 mg IV, and is continued on acetazolamide, 250 mg PO or IV every six hours, for the next 24 hours.
  • the patient is instructed to lie in a supine position for the first 24 hours following surgery; thereafter, the patient is instructed to remain in a face-down position as much as possible over the next five days.
  • Patients are examined at one day, two weeks, four to six weeks, and at three, six and twelve months after surgery. The patients are examined for best corrected visual acuity for both distance and near vision, refraction, intraocular pressure, size of neovascular net, presence of epiretinal membrane, presence of hyperfluorescence on fluorescein angiography, lens status, and results of ICG angiography.
  • a treatment is considered successful if: corrected visual acuity improves (about two lines) or stabilizes in patients whose vision is 20/200 or better; or corrected visual acuity improves to 20/200 in patients whose vision is worse than 20/200, or - there is a decrease in the size of the neovascular net.
  • inventive treatment is also considered to be beneficial in other ocular disorders such as retinal edema, retinal vascular disorders, wound healing disorders, proliferative ocular disorders, anti- degenerative disorders, anti-angiogenesis disorders, dry eye syndromes, uveitis, and various retinal detachments.
  • ocular disorders such as retinal edema, retinal vascular disorders, wound healing disorders, proliferative ocular disorders, anti- degenerative disorders, anti-angiogenesis disorders, dry eye syndromes, uveitis, and various retinal detachments.
  • EXAMPLE 4 To investigate the effects of TGF-j8l and TGF-32 alone on neovascularization in vivo, various doses of TGF- / 8 were implanted into the clear cornea of rabbits, and the neovascular response was measured over time. Five to seven pound male and female New Zealand White rabbits were used. The animals were anesthetized with subcutaneous injections of 20 mg of xylazine and 80 mg of keta ine every other day for a total of 4 anesthesias. While under anesthesia on day 6, the animals were euthanized with an intracardiac injection of 325 mg of sodium pentobarbital.
  • TGF-/31 and TGF-j82 and vehicle controls were placed in 2.5 isogel agarose (FMC Corp., Rockland, ME).
  • Porcine platelet-derived TGF-/31 and TGF-02 lyophilized without bovine serum albumin (BSA) were obtained from Drs. Anita Robert and Michael Sporn (NIH, Bethesda, MD) .
  • Porcine platelet-derived TGF-j ⁇ l lyophilized with BSA also was obtained from R&D systems, Inc. (Minneapolis, MN) .
  • the duplication of sources was used to help control for the method of procurement, handling and shipment variables.
  • TGF- ⁇ l and TGF-/S2 Prior to adding to the agarose, TGF- ⁇ l and TGF-/S2 were solubilized in 4 mM HC1. The agarose was heated to 60° C, added to the solubilized peptides and then allowed to gel at room temperature. The gelled agarose was then divided into 2
  • TGF- ⁇ l When TGF- ⁇ l was implanted into a nonvascular rabbit cornea, there was a dose-dependent stimulation of blood vessel growth in 82% of corneas implanted with 1, 5, 25 and 100 ng. The majority of corneas implanted with 1 ng of TGF-/S showed no neovascular ingrowth. The remaining 1-ng-treated corneas had sparse, short blood vessels. As the dose increase from 5 to 100 ng, the neovascularization became more dense, the blood vessels were longer, and the corneas became more edematous. At two days post-implantation, an intrastromal neovascular response was evident and became more prominent at days 4 and 6.
  • administering TGF-/31 or TGF-/S2 without any preexisting neovascularization may cause neovascularization to develop.
  • the triple pocket corneal assay includes first administration of an agent to produce neovascularization in one compartment, followed by implantation of TGF-31, TGF-02 or a control on both sides of the neovascularization.
  • neovascularization was induced by implanting a pellet containing either PGE1 (Upjohn Co. , Kalamazoo, MI) or TGF-alpha (Chemicon International, Inc. , El Segundo, CA) .
  • PGE1 Upjohn Co. , Kalamazoo, MI
  • TGF-alpha Chemicon International, Inc. , El Segundo, CA
  • the PGE1 was solubilized in absolute alcohol and then added to a casting solution of 10% ethylene vinyl coacetate polymer in methylene chloride to form pellets.
  • TGF- ⁇ was solubilized in 1 mM HC1 and then added to agarose and divided into implants.
  • TGF-31 and TGF-/S2 were used at doses of 1, 3, 5, 10, 25, 100 and 200 ng (6 corneas for each dose, except for 8 for the 100 ng dose) .
  • TGF-31 was implanted at the 50 ng dose.
  • Control agarose implants contained an equivalent volume of vehicle (4 mM HC1) or 100 ng of platelet-derived growth factor (PDGF from R&D Systems, Inc.) solubilized in 4 mM HC1.
  • the pockets for the secondary implants were formed by one-half-thickness incisions which were 1.5 mm long and perpendicular to and 1.5 mm from the limbus and 3-4 mm from the primary implant. Two pockets were formed on either side of the primary implant by gently inserting a cyclodialysis spatula into the incised edge of the cornea and advancing the spatula in a plane parallel to the curvature of the cornea to within 1.0 mm of the primary implant such that the pockets lay 1.5 mm from and parallel to the limbus.
  • Blood vessel lengths were measured adjacent to the TGF- ⁇ -containing implant (E) and the control implant (C) 2 mm from the center of the primary implant ( Figure 1) .
  • the relative lengths of the blood vessels in these areas were then expressed as a ratio: E/C (the length of the blood vessels in the area of the TGF-/3 implant divided by the length of the blood vessels in the area of the control implant) .
  • Percent stimulation or inhibition was calculated by subtracting 1.0 from E/C and multiplying by 100.
  • Measurements of blood vessel length adjacent to both the TGF-/3 and control implants were made from projected slides taken at days 2, 4 and 6. Serial 5-micron frozen sections were taken from a cornea implanted with 100 ng of TGF- ⁇ in a triple pocket assay, stained with hematoxylin and examined by light microscopy.
  • TGF-3 enhanced neovascularization in 89% of corneas at doses of 1, 3 and 5 ng. At 1 ng, neovascularization was enhanced by 47%; at 3 ng, 118%; and at 5 ng, 67% relative to control on day 4. TGF-01 stimulated neovascularization much more than TGF-32 at the 1 ng dose (about 55% and 40%, respectively) and 3 ng dose (180% and 56%, respectively).
  • the 50 ng dose of TGF-01 inhibited neovascularization by 59% on day 2, 49% on day 4 and 29% on day 6.
  • the dose of 10 ng appeared to be a transitional dose at which two of six corneas showed stimulation of neovascularization in the area of the TGF- ⁇ implant and four of six had neovascularization relative to the control.
  • both TGF- ⁇ l and TGF-02 can inhibit neovascularization caused by PGE1 or TGF- ⁇ .
  • the inhibition was found to be dose dependent, with doses greater than 10 ng inhibiting neovascularization.
  • the optimal dose in this experiment appeared to be about 100 ng.
  • TGF-/32 is superior in having less stimulatory effect and greater safety while offering equivalent neovascularization inhibition.
  • the inventors propose that the different effects above and below 10 ng may be due to the interplay of TGF- ⁇ on multiple functions, including causing chemotaxis in blood monocytes at about 0.1 to 1.0 pg/ml, inducing gene expression for interleukin-1 (at least IL-l3-specific mRNA has been observed in cultured monocytes) at 1.0 to 25 ng/ml, and inhibiting vascular endothelial cell proliferation at 0.1 to 10.0 ng/ml.
  • interleukin-1 at least IL-l3-specific mRNA has been observed in cultured monocytes
  • vascular endothelial cell proliferation appears to predominate; whereas, at higher doses, effects on cellular function which could inhibit neovascularization.
  • EXAMPLE 6 in this experiment 100 ng of PDGF and 100 ng of TGF-/31 and TGF-J2 (four corneas for each peptide) were the secondary implants after the initial 1.5 ug PGE1 neovascular stimulus.
  • TGF- ⁇ inhibited neovascularization relative to PDGF in 100% of corneas.
  • TGF- ⁇ l and TGF-32 showed comparable degrees of inhibition.
  • the average blood vessel length in the area of the TGF-/3 implant was 19%, 40% and 36% of the average blood vessel length in the area of the PDGF implant on days 2, 4 and 6, respectively (combined data for ⁇ l and ⁇ 2) .
  • TGF- ⁇ 100 ng of PDGF and 100 ng of TGF-01 and TGF-02 (six corneas for each peptide) were the secondary implants after the initial 300 ng TGF- ⁇ neovascular stimulus.
  • TGF-/3 also inhibited neovascularization stimulated by TGF- ⁇ at a dose of 300 ng.
  • TGF-/S inhibited neovascularization in 100% of corneas.
  • TGF-S1 and TGF-/32 had comparable degrees of inhibition.
  • the combined average blood vessel length in the area of the TGF- ⁇ implant was 47%, 51% and 47% of the blood vessel lengths around the control implant on days 2, 4 and 6, respectively.
  • the effect of TGF-/S2 on healing after corneal surgery for correction of myopia and hyperopia is determined by measuring the magnitude of effect that TGF-02 has on altering the corneal topography in three different types of corneal incisions.
  • Radial non-penetrating radial incisions are made.
  • One group of animals receives two radial incisions and the other group receives four radial incisions using a knife with micrometer adjusted to cut up to 90% of the central corneal thickness. Both eyes are operated. With the aid of an operating microscope, the 3.5 mm central optical zone centered over the pupil is demarcated with a marker.
  • Radial incisions start at the central optical zone and extend peripherally to within about 2-3 mm of the limbus.
  • Circular incisions are made with corneal trephines of different diameters and penetrate about 90% of the corneal depth. At the end of surgery, 2.0 ⁇ g or 5.0 ⁇ g TGF-,32 or control solution is applied to each eye.
  • TGF- ⁇ 2 is applied to one incision (after which a cup is placed over the incision to keep the medication from dispersing) and the other incision is not treated. This helps assess the effect of TGF- ⁇ 2 on the change in topography.
  • antibiotic ointment is topically applied to the eyes. The eyes are observed under the slit lamp and corneal topographic measurements are made, both before and at regular intervals after surgery. The slit lamp is used to evaluate corneal vascularization, epithelial healing, depth of incisions (to assess healing) and the amount of scar tissue formed. Corneal topographic measurements will help assess how symmetrically and quickly the eyes stabilize.
  • the cats are euthanized, and sections of the eyes are mounted on slide, stained and compared.
  • the eyes receiving TGF- ⁇ 2 treatment have rapid, strong healing and early stabilization of corneal topography.
  • bTGF-B2 was applied onto the macula of patients with CME following vitrectomy with the intent to reduce the magnitude of retinal edema and to improve visual acuity.
  • Study participants are examined at baseline and selected intervals post-treatment. Parameters to be assessed include visual acuity (EDTRS eye chart) , macular status (biomicroscopy and fluorescein angiography) , adverse events (lens status, intraocular pressure, and retinal detachment) , and concomitant medications.
  • CME cystoid macular edema
  • ocular disease e.g. macular degeneration, diabetic retinopathy, macular hole, retinal detachment, advanced glaucoma, etc.
  • macular degeneration e.g. macular degeneration, diabetic retinopathy, macular hole, retinal detachment, advanced glaucoma, etc.
  • Stereo red-free photographs are also taken of the macula at the beginning of the angiogram.
  • the transit frames of the angiogram are centered on the macular (field II) of the study eye, with frames taken at 30 seconds, 1 minute, 2 minutes, 5 minutes, and 10 minutes.
  • a stereo photograph of the disc (field I) is also taken at 10 minutes. All study photographs are labeled with the patient's study code and the date of the photograph, and subsequently graded by a certified grader who is masked with regard to either the patient's study code or the time the photographs were taken relative to treatment.
  • Treatment is scheduled within one week (seven days) of the date on which the baseline tests are completed. If for any reason treatment is postponed, all baseline studies must be repeated.
  • a vitreous cutter and fiberoptic light pipe are then introduced into the vitreous cavity.
  • the anterior and central vitreous are removed. Any visible vitreous adhesions to the anterior segment, iris, or lens capsule are severed.
  • the posterior vitreous is then removed with the vitreous cutter.
  • Each eye is examined for a posterior vitreous detachment using a flexible silicone tip cannula attached to the suction line of the vitreous cutter console.
  • a suction of 150 mm Hg is applied approximately 2-3 mm superior to the fovea, about 0.2 to 0.5 mm anterior to the retina, to determine if there is residual posterior cortical vitreous with an attached posterior hyaloid.
  • the silicone tip cannula is used to create a posterior vitreous detachment.
  • the posterior hyaloid is removed with the vitreous cutter to at least the equator.
  • the silicone tip cannula is used to verify that the posterior hyaloid has been completely removed posterior to the equator.
  • the vitreous cutter and fiberoptic light pipe is removed from the eye, and the sclerotomies closed with scleral plugs.
  • the peripheral retina is examined with indirect ophthalmoscopy and scleral depression to look for any peripheral retinal breaks. Peripheral breaks, if present, are treated with cryopexy.
  • the scleral plugs are then removed, and the light pipe and silicone tip cannula are reintroduced into the vitreous cavity.
  • a fluid-air exchange is performed, aspirating the intravitreal fluid over the optic nerve.
  • the instruments are removed, and the scleral plugs replaced. Fifteen minutes is allowed for accumulation of fluid from the peripheral retina and vitreous base around the optic nerve.
  • the scleral plugs are again removed, and the remaining fluid anterior to the optic nerve aspirated using the silicone tipped cannula.
  • the viscodissection cannula and tubing are then connected to a 1 cc syringe containing TGF-b2 freshly diluted in neutral buffer supplemented with 2% human serum albumin.
  • the tip of the viscodissection cannula is positioned directly over the fovea, taking care not to touch the fovea with the cannula.
  • a 0.1 ml aliquot (1330 ng) of TGF-32 is injected over the macula.
  • the instruments are removed from the eye, and the two superior sclerotomies closed with 7-0 vicryl suture.
  • the infusion cannula is removed, and the remaining sclerotomy closed with 7-0 vicryl suture.
  • the intraocular pressure is normalized with sterile air to achieve an intraocular pressure of about 10 mm Hg.
  • the conjunctiva is then closed with interrupted 6-0 collagen suture.
  • the patient is instructed to lie in a supine position for the first 24 hours following surgery; thereafter, the patient is instructed to remain in a facedown position as much as possible over the ensuing five days.
  • Intraocular pressure if abnormally high, IOP is treated first with topical aqueous suppressants such as 0-blockers or ⁇ -blockers; oral carbonic anhydrase inhibitors are only used if topical therapy has been deemed inadequate to control the intraocular pressure
  • topical aqueous suppressants such as 0-blockers or ⁇ -blockers
  • oral carbonic anhydrase inhibitors are only used if topical therapy has been deemed inadequate to control the intraocular pressure
  • the size of the gas bubble remaining the anterior segment, including the cornea, anterior chamber, and lens, with slit lamp biomicroscopy; adverse events; and concomitant medication.
  • the following parameters are assessed: best corrected vision using standard refraction and the ETDRS chart; intraocular pressure; lens status including the magnitude of nuclear sclerosis and posterior subcapsular cataract formation, both of which are graded on a scale of 0-4; the severity of cystoid macular edema is assessed by slit lamp biomicroscopy on a 0-4 scale; the severity of cystoid macular edema from fluorescein angiographic photographs by a certified grader who is masked with regard to patient identity and visitation date; the presence or absence of an epiretinal membrane; adverse events; and changes in concomitant medications.

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Abstract

Ce procédé convient pour le traitement d'une grande variété d'affections ophtalmiques, et notamment les perforations maculaires, la dégénérescence maculaire, ainsi que le décollement et les déchirures de la rétine, les cataractes, et les lésions cornéennes et sclérales. Ce procédé consiste à appliquer à la région affectée une quantité efficace de facteur transformant de croissance-β.
EP93917021A 1992-07-08 1993-07-08 PROCEDE POUR TRAITER DES AFFECTIONS OPHTALMIQUES A L'AIDE DU FACTEUR TRANSFORMANT DE CROISSANCE--g(b). Ceased EP0651650A4 (fr)

Applications Claiming Priority (5)

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US91083492A 1992-07-08 1992-07-08
US910834 1992-07-08
US877893A 1993-01-22 1993-01-22
US8778 1993-01-22
PCT/US1993/006420 WO1994001124A1 (fr) 1992-07-08 1993-07-08 PROCEDE POUR TRAITER DES AFFECTIONS OPHTALMIQUES A L'AIDE DU FACTEUR TRANSFORMANT DE CROISSANCE-$g(b)

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WO1994001124A1 (fr) 1994-01-20
EP0651650A4 (fr) 1996-04-17
JPH08502033A (ja) 1996-03-05
CA2138644A1 (fr) 1994-01-20

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