The therapeutic activity of nystatin (NYS) incorporated in multilamellar liposomes (L-NYS) was studied in vivo. Hale-Stoner mice injected intravenously with various doses of L-NYS and free NYS showed a significant reduction in toxicity of NYS after the NYS was incorporated into liposomes (maximal tolerated doses, 16 and 4 mg/kg of body weight, respectively). The maximal tolerated dose of free NYS had no effect in the treatment of mice infected with Candida albicans, whereas L-NYS at an equivalent dose improved the survival of mice. A marked increase in survival was observed when L-NYS was administered in higher and multiple doses (total doses up to 80 mg/kg). Liposome encapsulation thus provided a means for intravenous administration of NYS, reducing its toxicity and making it an active systemic antifungal agent.The treatment of fungal infections remains a major problem in spite of the availability of effective antifungal drugs such as polyenes (3). Most of the available polyene antibiotics have toxic side effects that limit their clinical application (5, 6). In addition, the hydrophobic nature of nystatin (NYS) has precluded its systemic administration. It has been used as suspensions prepared in various ways and administered to patients orally (1,2,7,14,17). However, most of these studies failed to document a beneficial effect of NYS administration against systemic fungal infections (2, 14, 17).We have recently demonstrated that liposome encapsulation improves the therapeutic index of amphotericin B both against experimental murine candidiasis (10, 11) and in the treatment of fungal infections in patients with leukemia and lymphoma (9). NYS in liposomal form (L-NYS) was a good additional drug prototype to study antifungal activity because of the structural similarities NYS shares with amphotericin B. The present communication reports on the in vivo toxicity and antifungal efficacy of L-NYS as compared with those of the free drug.MATERIALS AND METHODS Drug, lipids, and reagents. NYS (bulk powder) was obtained from Lederle Laboratories, Pearl River, N.Y. Chromatographically pure dimyristoyl phosphatidylcholine (DMPC) and dimyristoyl phosphatidylglycerol (DMPG) were purchased from Avanti Polar Lipids, Birmingham, Ala.Liposome preparation. Multilamellar vesicles were prepared as described previously (11). NYS was solubilized in methanol (1 mg/ml) and stored at 4°C, protected from light. Phospholipids DMPC and DMPG (7:3) were mixed with a methanol solution of NYS, and the organic solvents were evaporated under vacuum by using a rotary evaporator. The dried drug-lipid film was suspended in 0.9% pyrogen-free saline and hand shaken, allowing liposomes to form. The suspension was then centrifuged at 100,000 x g for 1 h, and the pellet was resuspended in the saline. Animals and model of experimental candidiasis. HaleStoner mice, 6 to 8 weeks old (body weight, 20 to 25 g), were purchased from the University of Texas Science Park, Bastrop. The mice (eight per group) were injected via the tail vein with 0.2 ml of Candida a...