Ivacaftor (VX-770)

Ivacaftor (VX-770) (10 μM) significantly increases the forskolin-stimulated Cl^- secretion (I_T) by ~4-fold with an EC50 of 100 nM in the recombinant Fisher rat thyroid (FRT) cells expressing G551D gating mutation of CFTR, and by ~6-fold with an EC50 of 25 nM in the recombinant cells expressing temperature-corrected F508del processing mutation of CFTR. Consistent with the increases in the forskolin-stimulated I_T, this compound increases the open probability (P_o) of G551D-, F508del-, and wild-type CFTR by ~6-fold, ~5-fold and ~2-fold, respectively, indicating that it acts directly on CFTR to increase its gating activity. In primary cultured human CF bronchial epithelia (HBE) carrying the G551D and F508del CFTR mutations, Ivacaftor potently increases the forskolin-stimulated I_T by ~10-fold from 5% to a maximum level of 48% of that measured in non-CF HBE, with an EC50 of 236 nM displaying ~70-fold more potency compared with the commonly used CFTR potentiator genistein, which has an EC50 of 16 μM. In HBE with F508del homozygous CFTR, it causes a significant increase in the forskolin-stimulated I_T with an EC50 of 22 nM, to a less extent from 4% to 16% of non-CF HBE compared with the effect in G551D/F508del HBE. Due to CFTR potentiation, this compound inhibits excessive ENaC-mediated Na⁺ and fluid absorption with an IC50 of 43 nM, and decreases the response, resulting in an increase in the surface fluid and cilia beat frequency (CBF) in G551D/F508del HBE. [1]

• Registros de cámara de Ussing

  El efecto de Ivacaftor (VX-770) sobre la secreción de Cl^- mediada por CFTR se caracteriza midiendo la I_T mediada por CFTR en cámaras utilizando células tiroideas de rata Fisher (FRT) recombinantes que expresan G551D o F508del CFTR. Las células se cultivan en insertos de cultivo celular Costar Snapwell mantenidos a 37 °C antes del registro. Los insertos de cultivo celular se montan en una cámara de Ussing para registrar la I_T en el modo de fijación de voltaje (V_hold = 0 mV). Para las células FRT, la membrana basolateral es un gradiente de Cl^- basolateral a apical. La solución de baño basolateral contiene 135 mM de NaCl, 1,2 mM de CaCl₂, 1,2 mM de MgCl₂, 2,4 mM de K₂HPO₄, 0,6 mM de KHPO₄, 10 mM de N-2-hidroxietilpiperazina-N

• Líneas celulares

  FRT and NIH 3T3 cells

• Concentraciones

  100/25 nM

• Tiempo de incubación

  24 h

• Método

  Cells were treated with various concentrations of Ivacaftor (VX-770).

• Modelos animales

  Male Sprague-Dawley rats

• Dosificaciones

  3 mg/kg

• Administración

  i.v.