title: An in vitro method which assesses corneal epithelial toxicity due to antineoplastic, preservative and antimicrobial agents.
abstract: We developed an in vitro model for studying the cytotoxicity of pharmacologic agents on corneal epithelium employing 3H-thymidine incorporation. Primary rabbit corneal epithelial cell cultures were established, and the cells plated prior to each experiment. 3H-thymidine incorporation was measured after the addition of drug or vehicle to these confluent cells, and dose-response curves were generated. Marked inhibition of 3H-thymidine incorporation was reached at chemotherapeutic concentrations achieved clinically for cytosine arabinoside (10(-7) M), methotrexate (10(-3) M), and 5-fluorouracil (10(-6) M). A 10(-4) M concentration of 2-deoxycytidine, a naturally occurring competitive inhibitor of cytosine arabinoside, protected cells up to a concentration of 10(-5) M. We utilized these data to undertake an in vivo prophylaxis study in 13 leukemia patients receiving high-dose iv cytosine arabinoside. Topical deoxycytidine 10(-4) M and 1% prednisolone phosphate, given 12 hours prior to the start of antileukemic therapy, were effective in reducing symptoms and signs of keratitis; both were better than historical placebo-treated eyes. Ophthalmic preservatives were studied in vitro at concentrations used clinically: benzalkonium chloride (BAC) (0.004-0.02%) was the most toxic, thimerosal (TMS) (0.001-0.004%) intermediate, and chlorobutanol (CHB) (0.2-0.5%) the least toxic. Antiviral agents (final concentration) included: trifluridine (TFT) (1.0%), ethyldeoxuridine (EDU) (2.0%), and idoxuridine (IDU) (0.1%). Dose but not time-dependent concentrations of these 3 agents were noted to cause toxicity; however, (E)-5(2-bromovinyl)-2'-deoxyuridine (BVDU) (0.1%) was non-toxic. Similarly, tobramycin and amikacin were significantly less toxic than gentamicin and neomycin in this system. These in vitro cytotoxicity data correlate well with previous in vivo and pre-clinical corneal epithelial toxicity studies. Our model may be useful in the toxicologic study of future topical ophthalmic agents.
authors.affiliation: Department of Medicine, University Hospitals of Cleveland, Case Western Reserve University, Ohio 44106.
authors.forename: H M
authors.initials: HM
authors.lastname: Lazarus
authors.forename: P S
authors.initials: PS
authors.lastname: Imperia
authors.forename: R E
authors.initials: RE
authors.lastname: Botti
authors.forename: R J
authors.initials: RJ
authors.lastname: Mack
authors.forename: J H
authors.initials: JH
authors.lastname: Lass
mesh_terms: D000328:Adult; D000368:Aged; D000617:Aminoglycosides / Q000633:toxicity*; D000818:Animals; D000970:Antineoplastic Agents / Q000633:toxicity*; D000998:Antiviral Agents / Q000633:toxicity*; D002478:Cells, Cultured; D003315:Cornea / Q000187:drug effects*; D003561:Cytarabine / Q000009:adverse effects / Q000627:therapeutic use; D003841:Deoxycytidine / Q000627:therapeutic use; D004311:Double-Blind Method; D004848:Epithelium / Q000187:drug effects; D005260:Female; D006689:Hodgkin Disease / Q000150:complications / Q000188:drug therapy; D006801:Humans; D007634:Keratitis / Q000139:chemically induced / Q000517:prevention & control; D007938:Leukemia / Q000150:complications / Q000188:drug therapy; D008297:Male; D008875:Middle Aged; D011310:Preservatives, Pharmaceutical / Q000633:toxicity*; D011817:Rabbits; D013936:Thymidine / Q000378:metabolism; D014116:Toxicology / Q000379:methods*
chemical_list: D000617:Aminoglycosides; D000970:Antineoplastic Agents; D000998:Antiviral Agents; D011310:Preservatives, Pharmaceutical; D003561:Cytarabine; D003841:Deoxycytidine; D013936:Thymidine