Methods for diagnosing glaucoma and for screening therapeutic agents for their usefulness in treating glaucoma are disclosed.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to the field of glaucoma diagnosis and treatment. More specifically, the present invention provides methods for diagnosis of glaucoma by measuring the amount of GR.beta. present in the trabecular meshwork of a patient's eye.

2. Description of the Related Art

Glaucoma is usually diagnosed by monitoring a patient's visual field loss, changes in the appearance of their optic disc, and their intraocular pressure. Glaucoma is currently treated using one or more of three strategies to lower the elevated intraocular pressure associated with the disease: with pharmaceuticals (such as beta-blockers, carbonic anhydrase inhibitors, miotics or prostaglandins), with laser trabeculoplasty, and/or with glaucoma filtration surgery. All of these therapies indirectly lower intraocular pressure but do not address the underlying disease process occurring in the trabecular meshwork. It would be advantageous to be able to diagnose glaucoma before a patient begins experiencing a loss in their visual field and deterioration of their optic disc.

There is a large body of evidence suggesting that glucocorticoids are involved in the generation of ocular hypertension and glaucoma (Clark 1995). The human glucocorticoid receptor (hGR) and its isoforms, hGR.alpha. (SEQ ID NO:3) and hGR.beta. (SEQ ID NO:1), are described in Encio and Detera-Wadleigh (1991) (See also FIG. 1). Several investigators have shown that the human trabecular meshwork (TM) contains the classical glucocorticoid receptor (hGR.alpha.) (Weinreb et al. 1981; Hernandez et al. 1983). Recently, the expression of an alternatively spliced form of the human glucocorticoid receptor (hGR.beta.) was discovered in non-ocular tissues and cells (Bamberger et al. 1995; Oakley et al. 1996). This alternatively spliced form of hGR is expressed as a protein (SEQ ID NO:2) which no longer binds glucocorticoids, but is able to interfere with the activated form of the normal glucocorticoid receptor and block or alter physiological functions of the glucocorticoid receptor.

SUMMARY OF THE INVENTION

The present invention is directed to methods for diagnosing glaucoma by testing a person for aberrant hGR.beta. expression. In preferred embodiments, a decrease in hGR.beta. expression in the trabecular meshwork of glaucomatous eyes as compared to hGR.beta. expression in non-glaucomatous eyes. Also set forth are methods for screening for therapeutic agents useful for treating glaucoma.

DESCRIPTION OF PREFERRED EMBODIMENTS

Surprisingly, it has been found that cultured human trabecular meshwork cell lines express the proteins from both an alternate splice form of the human glucocorticoid receptor (GR.beta.; SEQ ID NO:1), as well as the normal glucocorticoid receptor (GR.alpha.; SEQ ID NO:3). Glaucomatous TM cells have less GR.beta. protein and therefore are more susceptible to endogenous and exogenous glucocorticoids. It is believed that the elevated intraocular pressure associated with primary open-angle glaucoma may be due to the aberrant expression of GR.beta. in the trabecular meshwork. Therefore, determining that an individual abnormally expresses GR.beta. in their trabecular meshwork or other tissues can lead to a diagnosis of glaucoma.

The present invention further provides a method for determining whether a candidate substance has therapeutic value in treating glaucoma by determining whether the candidate substance interacts with the GR.beta. protein (SEQ ID NO:2) or alters the expression of GR.beta. (SEQ ID NO:1). This can be done using ligand binding assays or GR.beta. functional assays.

Diagnosing aberrant GR.beta. expression or defects in the GR gene which encodes GR.beta. can be done by using procedures well known to those skilled in the art (Caskey 1993). For example, subjects could be screened for the presence of a genetic defect in GR.beta. by analyzing the DNA derived from peripheral blood leukocytes. Types of DNA analyses could include, but would not be limited to: restriction fragment length polymorphisms (RFLP), single-stranded conformation polymorphisms (SSCP), polymerase chain reaction (PCR), denaturing gradient gel electrophoresis, allele specific oligonucleotide ligation assay, and allele specific hybridization assay. In addition, trabecular meshwork, or other relevant cells from subjects could be analyzed for GR.beta. expression by a number of techniques such as reverse-transcription polymerase chain reaction (RT-PCR), immunoassays, GR functional assays, etc.
 

Claim 1 of 1 Claim

1. A method for diagnosing glaucoma in a patient, said method comprising the steps: (a) obtaining a biological sample from the trabecular meshwork of said patient; and (b) analyzing said sample for expression of GR.beta. (SEQ ID NO:1); wherein a decrease in expression of GR.beta. as compared to expression of GR.beta. in a patient not suffering from glaucoma indicates a diagnosis of glaucoma.

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