MK-927

Ampholyte carbonic anhydrase inhibitors (including MK-927) have previously been shown to elicit greater intraocular pressure reduction when applied in ionized form at moderately acidic or alkaline pH compared to application of un-ionized drug at neutral pH. Since ionized solutions should be far more membrane-impermeant than un-ionized solutions, we attempted to solve this apparent paradox. We studied the distribution at 0.5-18 hr of MK-927 in eye tissues and fluid after topical instillation of one drop of a 0.5% solution at pH 4.9, 7.0 and 9.1. Measured drug concentrations at 30 min in corneal epithelium and stroma are approximately 4.5-fold greater after acidic or alkaline application than after neutral application, with the highest levels being found in corneal epithelium. The same pattern is seen in ciliary process, the site of aqueous humor production. Here drug concentration is approximately 60% of that in cornea at 30 min. Free drug concentrations in ciliary process were used to compute the time course of maximal carbonic anhydrase inhibition for the three modes of application. Drug concentration in sclera, uvea and aqueous humor at all times are all low by comparison, suggesting drug movement is from cornea to ciliary process via the corneo-scleral junction. The k(in) for proton movement across the corneal epithelium was measured (k(in) = 12.4 hr-1) from which a permeability coefficient (P = 2.7 x 10(-2) cm sec-1) was computed. Separate analysis was made of the pH status of the cornea 30 min and 1 hr following instillation of 1 drop 2% acidic (pH 4.9) and alkaline (pH 9.1) MK-927, with sodium sulfadiazine and pilocarpine.HCl as pH controls. Stromal bicarbonate at 30 min was approximately halved after acidic drops and doubled after alkaline drops consistent with pH decrease or increase of nearly 0.3 U. The results are in accord with classical schemes for amine permeation of the cornea at acidic pH when consideration is given to movement of acid equivalents and corneal pH. Thus drug inside the eye is in relatively (> 95%) lipophilic form except that trapped in the cornea shortly after acidic or alkaline applications.

Many functions of polymorphonuclear leukocytes (PMNs) appear to alter and be affected by changes in the intracellular and/or extracellular acid-base milieu, suggesting that carbonic anhydrase (CA) may be important. Although small amounts of CA activity in PMNs have been reported, it has not been characterized fully. We therefore studied isolated mature circulating human PMNs and cultured HL-60 cells, an undifferentiated myelopoietic cell line, and compared these to human red cells (RBCs) for CA activity. Activity and sulfonamide inhibition were measured by a modified micromethod assay. Isoenzyme and total CA concentrations were determined by radioimmunoassay for human isozyme CA I, differential inhibition by MK-927, inhibition by 0.2% sodium dodecyl sulfate (SDS), and quantitative sulfonamide binding. Total CA activity (units/106 cells) was 0.04 in PMNs, 0.06 in HL-60 cells, and 0.62 in RBCs. Human PMNs have a total CA concentration of 1.3 μM, of which 0.9 μM is CA I and the remainder is CA II. Total loss of CA activity with 100 μM ethoxzolamide and 0.2% SDS ruled out significant CA III or CA IV activity. Subcellular fractionation of PMNs revealed that all CA activity was cytosolic. The absence of CA activity in mitochondrial and microsomal membrane fractions argues against any mitochondrial CA V or membrane-bound CA IV contribution to total CA activity. Neutrophils contain both CA I and II isozymes in roughly the same proportion as RBCs but at much lower concentrations, suggesting that in the course of maturation the CA content of neutrophils is regulated differently from that in erythrocytes. J. Leukoc. Biol. 55: 343–348; 1994.