title: Preformulation Studies of a Liposomal Formulation Containing Sirolimus for the Treatment of Dry Eye Disease.
abstract: OBJECTIVE
  The aim of this study was to develop and characterize a liposomal product containing sirolimus to be administered subconjunctivally for the treatment of nonresponsive keratoconjunctivitis sicca (KCS) or dry eye.
  
  
  METHODS
  Formulations were prepared using an ethanol injection method and an adaptation of the heating method in pursuance of the most suitable methodology for future industrial production. Liposomes were loaded with either a high dose of 1 mg/mL of sirolimus or a less toxic dose of 0.4 mg/mL. The effects of critical process and formulation parameters were investigated. Liposomes were characterized in terms of size, zeta potential, polydispersity, differential scanning calorimetry, morphology, entrapment efficiency, phospholipid content, thermal stability, and sterility. The formulation was evaluated clinically in dogs with spontaneous KCS.
  
  
  RESULTS
  Sterile liposomal dispersions with sizes ranging from 140 to 211 nm, were successfully obtained. High entrapment efficiency of 93%-98% was achieved. The heating method allowed an easier production of liposomes with high entrapment efficiency, to significantly shorten production time and the elimination of the use of alcohol. The poor stability of the obtained liposomes in aqueous dispersion made the inclusion of a lyophilization step necessary to the manufacturing process. In vivo testing of the liposomal sirolimus formulations in the spontaneous KCS dog model have produced promising results, particularly with a sirolimus dose of 1 mg/mL, indicating the need for further development and study of proposed formulations in the treatment of canine KCS. Clinical improvement in tear production in dogs with spontaneous KCS treated with the 1 mg/mL dose product was observed.
  
  
  CONCLUSIONS
  The heating method allowed easier production of high entrapment efficiency liposomes to significantly shorten production time and the elimination of the use of alcohol. Tear production was increased in dogs administered with the formulation.
authors.affiliation: 1 Faculty of Chemistry, National Autonomous University of Mexico , Mexico City, Mexico .
authors.forename: Mónica Anayántzin
authors.initials: MA
authors.lastname: Linares-Alba
authors.affiliation: 2 Veterinary Specialty Hospital "Oftalvet," Mexico City, Mexico .
authors.forename: Magda Berenice
authors.initials: MB
authors.lastname: Gómez-Guajardo
authors.affiliation: 3 Faculty of Veterinary Medicine, Sao Paulo State University , Botucatu, SP, Brazil .
authors.forename: Joice Furtado
authors.initials: JF
authors.lastname: Fonzar
authors.affiliation: 4 College of Veterinary Medicine, University of Florida , Gainesville, Florida.
authors.forename: Dennis E
authors.initials: DE
authors.lastname: Brooks
authors.affiliation: 2 Veterinary Specialty Hospital "Oftalvet," Mexico City, Mexico .
authors.forename: Gustavo Adolfo
authors.initials: GA
authors.lastname: García-Sánchez
authors.affiliation: 1 Faculty of Chemistry, National Autonomous University of Mexico , Mexico City, Mexico .
authors.forename: Maria Josefa
authors.initials: MJ
authors.lastname: Bernad-Bernad
mesh_terms: D000818:Animals; D002626:Chemistry, Pharmaceutical; D004195:Disease Models, Animal; D004285:Dogs; D000431:Ethanol / Q000737:chemistry; D056965:Injections, Intraocular; D007638:Keratoconjunctivitis Sicca / Q000175:diagnosis / Q000188:drug therapy*; D008081:Liposomes; D010316:Particle Size; D020123:Sirolimus / Q000008:administration & dosage* / Q000009:adverse effects / Q000627:therapeutic use*
chemical_list: D008081:Liposomes; D000431:Ethanol; D020123:Sirolimus