A new long acting ophthalmic formulation of carteolol containing alginic acid.

Alginic acid was evaluated as a potential vehicle in ophthalmic solutions for prolonging the therapeutic effect of carteolol. This anionic vehicle was expected to slow down drug elimination by the lacrimal flow, both by undergoing in-situ gel formation and by interacting with the mucus. In vitro studies indicated that carteolol is released slowly from alginic acid formulations, suggesting an ionic interaction. The adhesive behavior of alginic acid solution was better than that of another polymer, hydroxyethylcellulose (HEC). Intraocular pressure (IOP) measurements of rabbit eyes treated with a 1% carteolol formulation with or without alginic acid showed that this polymer significantly extended the duration of the pressure-reducing effect of carteolol to 8 h. The increased ocular bioavailability of 1% carteolol in the presence of alginic acid led to an equivalent concentration in the target tissue although administration was only once a day compared with twice a day for 1% carteolol alone. The overall results of this study indicate that the alginic-acid vehicle is an excellent drug carrier, well tolerated, and could be used for the development of a long-acting ophthalmic formulation of carteolol.

Interactions of human lymphoblasts with targeted vesicles containing Sendai virus envelope proteins.

We have studied the internalization of targeted fusogenic liposome content to leukemic T cells (CEM) in vitro. We describe a method for the covalent coupling of T101 antibody to the surface of liposomes and the incorporation of fusogenic viral protein into the liposome membrane. Hygromycin B, an impermeant inhibitor of protein synthesis, was encapsulated in the targeted fusogenic liposomes and delivered directly to the cytoplasm of leukemic T cells by fusion between the two membranes. The cytotoxic effect was measured by [3H]thymidine incorporation. We show that CEM are rapidly and specifically killed by the drug encapsulated in the targeted fusogenic liposomes. This effect is due to the binding of the liposome by means of the antibody and then to the fusion of the liposome with the targeted cell membrane, mediated by F protein.

F protein-F protein interaction within the Sendai virus identified by native bonding or chemical cross-linking.

The spatial arrangement of the F protein spike in the Sendai virus was studied after purifying the protein and reconstituting it in lipid vesicles (Sechoy, O., Philippot, J. R., and Bienvenue, A. (1986) Biochim. Biophys. Acta 857, 1-12). The different components of the F protein spikes were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under various conditions of treatment, i.e. at different temperatures and sodium dodecyl sulfate concentrations, using different detergents for F protein solubilization (Triton X-100 and octyl glucoside), by fast protein liquid chromatography analysis, and by chemical cross-linking between subunits with bifunctional agents such as dimethyl adipimidate and dithiobis(succinimidyl propionate). The F protein spike appeared to be a structurally stable complex, composed of a noncovalent association of four homooligomers, each consisting of two peptides, F1 and F2, linked by a disulfide bond. Octyl glucoside and Triton X-100 solubilized the F protein, preserving the tetramer, which is probably the native form. Using chemical cross-linking, a covalent bond was formed between two monomers. We hypothesize that the tetrameric form of the F protein in its native form (spike) consists of two identical dimers that can be chemically cross-linked in a stable complex.

Preparation and characterization of F-protein vesicles isolated from Sendai virus by means of octyl glucoside.

We have demonstrated that Triton X-100 is always present in F-protein vesicles at concentrations that can provoke cell lysis. In order to avoid any misinterpretation of the fusogenic capacity of this protein, we solubilized the Sendai virus using octyl glucoside, which can be totally removed from the F protein preparation in less than 16 h by dialysis in the presence of absorbent beads. F-glycoprotein preparations preserved their ability to lyse erythrocytes in the presence of lectins and to induce cell-vesicle fusion as demonstrated by ESR studies. These vesicles were characterized by electron microscopy and SDS-polyacrylamide gel electrophoresis. Lipid analysis of these preparations by thin-layer chromatography indicated that they had the same proportion of lipids as virus envelopes, with slight variations in the sphingomyelin content and the cholesterol/phospholipid molar ratio. F-protein vesicles of different sizes can be obtained by adding exogenous lipids before detergent removal. The hemolytic activity of the vesicles was retained over a large range of lipid concentrations. We conclude that F-protein vesicles prepared with octyl glucoside are convenient tools for studying the fusogenic mechanism of this protein and improving the fusion process between liposomes and cells.