A mouse model to test the in vivo efficacy of chemical chaperones

In vitro studies in transfected cells have indicated that chemical chaperon es including glycerol (0.5-1.2 M) and trimethylamine oxide (TMAO, 50-100 mM ) can correct defective trafficking of some proteins, including Delta F508 CFTR in cystic fibrosis and AQP2 mutants in nephrogenic diabetes insipidus. To develop a mouse model to test the efficacy of chemical chaperones in vi vo, glycerol and TMAO were administered by intraperitoneal (i.p.), subcutan eous (s.c.), and oral routes. Glycerol and TMAO assays that utilized 1-5 mu L of tail vein blood were developed. Administration by the i.p. and s.c. r outes gave maximum serum glycerol concentrations of similar to 100 mM, leve ls that were well below the effective in vitro concentrations. Single i.p. or s.c. doses of TMAO (7 g/kg, 8% solution in water) resulted in serum [TMA O] greater than 50 mM, with a long half-life (t(1/2) similar to 18-21 h). S ustained high serum and tissue [TMAO] > 52 mM for 3 days was achieved by s. c. administration of TMAO (7 g/kg) in water every 8 h. Although similar to 50% of the mice died with this multiple-dose regimen, the remaining mice ha d nearly normal liver, renal, and pancreatic function. A lower dose of TMAO (5 g/kg) given by the s.c. route every 8 h resulted in serum [TMAO] concen tration of 22 mM. a level that was well tolerated by all mice for 72 h. The se mice also had high [TMAO] in urine, 400 mM. These results demonstrate th at potentially therapeutic concentrations of TMAO can be sustained in mice in vivo, permitting the testing of chemical chaperones in transgenic mouse models of diseases caused by defective protein trafficking. (C) 1999 Elsevi er Science Inc.