Billie M. York

Glutathione depletion in the lens of galactosemic and diabetic rats

Depletion of lens glutathione (GSH) occurs quickly and drastically following induction of diabetes or galactosemia in rats as well as in lens culture. The explanation for this dramatic loss of GSH has been investigated by many laboratories but the solution has been elusive. There are several possible causes for the change in the reducing power of the lens under hyperglycemia. (a) The enzyme glutathione reductase which reduces oxidized glutathione to GSH is inhibited. (b) The cofactor NADPH which both the aldose reductase of polyol pathway and glutathione reductase require becomes depleted under hyperglycemia to the point that there is an insufficient amount for glutathione reduction. (c) Membrane permeability is increased, due to osmotic-induced lens hydration. We explored all the above possibilities in the mechanism of GSH depletion and studied the effect of aldose reductase inhibitor (ARI) on osmotic change. We found that under hyperglycemic condition, there was no change in the enzyme glutathione reductase activity. There was an initial drop in NADPH level but there was sufficient remaining for glutathione reductase use. Both NADPH and glutathione depletion could be prevented completely by ARI. In addition, ARI could also prevent any hyperglycemic-induced abnormal transport and leakage of amino acids. We have therefore concluded that only the decreased membrane transport of amino acids which are needed for glutathione biosynthesis and the simultaneous loss of GSH through leaky membrane as initiated by the polyol pathway can be responsible for the drastic GSH depletion.

Reformulation and Drop Size of Apraclonidine Hydrochloride

We performed a prospective, double-masked, placebo-controlled, six-period, cross-over study in which normal subjects were randomly assigned to treatment and compared three different formulations of apraclonidine hydrochloride (the present commercially available formulation, and formulations with hydroxypropylmethylcellulose or lysolecithin). We also evaluated the efficacy of a 16-microliters and 30-microliters drop size. The magnitude and duration of decrease in intraocular pressure was comparable for all formulations. Most subjects tolerated all formulations well with only a few reporting any side effects. The best-tolerated formulation was 0.5% apraclonidine hydrochloride delivered with a 16-microliters drop size. Dry mouth developed frequently with the commercially available 1% apraclonidine solution. Blurred vision complicated the use of the formulation containing hydroxypropylmethylcellulose. Both dry mouth (P less than .05) and blurred vision (P = .004) were statistically significant side effects.

The effect of an aldose reductase inhibitor on lens phosphorylcholine under hyperglycemic conditions: biochemical and NMR studies.

Phosphorylcholine (P-choline) is a precursor of the phospholipids in the lens membrane. A human lens normally contains approx. 1 mM P-choline but this is significantly lowered in some cataractous lenses. A normal rat lens contains a very high concentration (11 mM). We found that rat lens P-choline was depleted drastically when the lenses were exposed to hyperglycemic conditions either in culture, with galactose or xylose, or in vivo by streptozotocin-induced diabetes. The lens P-choline level was measured by fractionating the organic phosphates in the lens homogenate using an ion exchange column, or by quantitating the P-choline 31P NMR intensity in intact lenses. The results of both the chemical method and the noninvasive method agreed remarkably well. Besides the change in P-choline, the choline influx was also drastically reduced both in lenses from diabetic rats and in lenses incubated with 30 mM xylose. In addition, the ATP concentration was greatly diminished under similar conditions. The changes in P-choline, choline, and ATP could all be prevented in the presence of an aldose reductase inhibitor (ARI). It is thus concluded that these changes in phospholipid precursors may result from lenticular membrane defects caused by hyperglycemic stress. The effect of the lowered precursors on lipid biosynthesis was observed, and surprisingly showed a more rapid phospholipid-biosynthesis in the 2-week diabetic rat lens than in the 3-day diabetic rat lens.

Method for treating hypertension with 2,3-diamino-1,4-butanedithiol; 4,5-diamino-1,2-dithiane; and N-acyl and N-alkyl derivatives thereof

Disclosed are 2,3-diamino-1,4-butanedithiol; 4,5-diamino-1,2-dithiane; and their N-acyl and N-alkyl derivatives. Also disclosed are processes for preparing the disclosed compounds; pharmaceutical compositions comprising such compounds; and a method of treatment comprising administering such compounds and compositions when an antihypertensive effect or radioprotective effect is indicated.

Comparison of the Pharmacokinetics and Pharmacodynamics of the Aldose Reductase Inhibitors, AL03152 (RS), AL03802 (R), and AL03803 (S)

The pharmacokinetics of AL03152 (RS) and its enantiomers, AL03802 (R) and AL03803 (S), were studied in the Sprague–Dawley rat following intravenous bolus administration. The enantiomers had differing pharmacokinetic profiles, while the racemic compound exhibited pharmacokinetic parameters approximating the mean values of the individual enantiomers. The total clearance (CLT) values of the two enantiomers were similar, but the intrinsic clearance (Clint) was much greater for the S-enantiomer than for the R-enantiomer. The volume of distribution (Vss) for AL03802 (R) was threefold greater than that for AL03803 (S). The stereoselectivity in Vss could not be totally accounted for by the slight difference in serum protein binding of the isomers and resulted in a difference in the half-lives of the enantiomers. Only the R-isomer exhibited a persistent terminal elimination phase, consistent with more extensive tissue binding than the S-isomer. AL03152 enantiomers were equivalent in potency assessed from in vitro IC50 values toward rat lens aldose reductase and rat kidney L-hexonate dehydrogenase and lens EC50 values in diabetic rats.

Pharmacokinetics and efficacy of structurally related spirohydantoin and spirosuccinimide aldose reductase inhibitors

1. Six potent aldose reductase inhibitors (ARI), three spirohydantoin (I to III) and three spirosuccinimide (IV to VI) compounds, showed similar IC50 activities in vitro for the inhibition of rat lens aldose reductase, but their ED50 values in diabetic rats varied as much as 20-fold in the lens and 50-fold in the sciatic nerve tissue. Pharmacokinetic studies were undertaken to investigate these findings. Structure-pharmacokinetic relationships were studied following i.v. administration to cynomolgus monkeys. 2. The clearance (CL) of each spirosuccinimide ARI was faster (greater than 5 times) than that of the corresponding spirohydantoin compound. In both series the CL values of the C(4) methyl and methoxy analogues were 4-fold greater than those for the unsubstituted compounds, although the CL values of the methoxy and methyl derivatives in the same series were not significantly different. 3. The volumes of distribution (Vss) of the spirohydantoins were about one-half those of the corresponding spirosuccinimides, and the Vss values of the parent compounds of both ARI series did not differ dramatically from those of their methyl and methoxy analogues. 4. All six compounds were eliminated from plasma in a biexponential fashion. The half-lives (lambda 1 and lambda 2) of the spirohydantoin compounds were much longer than those of the corresponding spirosuccinimide compounds, and the unsubstituted compounds had longer half-lives than their methyl and methoxy derivatives. The longest lambda 1 and lambda 2 half-lives were observed for imirestat, while two of the spirosuccinimides had the shortest half-lives. 5. These results indicate that the relationships observed between the in vitro and in vivo activities of the six ARI can be attributed to structurally dependent differences in metabolic clearance.

Role of Aldose Reductase and Effects of Aldose Reductase Inhibitors in Ocular Tissue Aging Phenomena in the Diabetic Rat

Studies of the role of aldose reductase in sugar cataractogenesis have been aided by the synthesis and characterization of inhibitors of this enzyme which can prevent lens opacification in galactosemic or diabetic rats1,2. The anti-cataract activity of aldose reductase inhibitors correlates with inhibition of polyol accumulation in the lenses of treated galactosemic or diabetic animals; in untreated diabetic rats, lens sorbitol levels may be elevated 100-fold over the normal value3. Hyperglycemia produces an increase in glucose content of tissues not responsive to insulin. Due to increased substrate availability, the rate of sorbitol formation by action of aldose reductase is greatly accelerated relative to conversion of this metabolite to fructose via sorbitol dehydrogenase1:

Brinzolamide and brimonidine for treating ocular conditions

Glucose\quad + \quad NADPH\quad + \quad {H^{ + }}\;\xrightarrow{{Aldose\,Reductase}}\;Sorbitol\quad + \quad NAD{P^{ + }}