title: siRNA-mediated knockdown of P450 oxidoreductase in rats: a tool to reduce metabolism by CYPs and increase exposure of high clearance compounds.
abstract: OBJECTIVE
  To develop a tool based on siRNA-mediated knockdown of hepatic P450 oxidoreductase (POR) to decrease the CYP-mediated metabolism of small molecule drugs that suffer from rapid metabolism in vivo, with the aim of improving plasma exposure of these drugs.
  
  
  METHODS
  siRNA against the POR gene was delivered using lipid nanoparticles (LNPs) into rats. The time course of POR mRNA knockdown, POR protein knockdown, and loss of POR enzyme activity was monitored. The rat livers were harvested to produce microsomes to determine the impact of POR knockdown on the metabolism of several probe substrates. Midazolam (a CYP3A substrate with high intrinsic clearance) was administered into LNP-treated rats to determine the impact of POR knockdown on midazolam pharmacokinetics.
  
  
  RESULTS
  Hepatic POR mRNA and protein levels were significantly reduced by administering siRNA and the maximum POR enzyme activity reduction (~85%) occurred 2 weeks post-dose. In vitro analysis showed significant reductions in metabolism of probe substrates due to POR knockdown in liver, and in vivo POR knockdown resulted in greater than 10-fold increases in midazolam plasma concentrations following oral dosing.
  
  
  CONCLUSIONS
  Anti-POR siRNA can be used to significantly reduce hepatic metabolism by various CYPs as well as greatly increase the bioavailability of high clearance compounds following an oral dose, thus enabling it to be used as a tool to increase drug exposure in vivo.
authors: Department of Pharmacokinetics, Pharmacodynamics, and Drug Metabolism,, Merck Research Laboratories, Merck & Co., Inc.,, West Point,, Pennsylvania, 19486, USA, rob@aviditynano.com.
authors: Rob S
authors: RS
authors: Burke
authors: Inthirai
authors: I
authors: Somasuntharam
authors: Paul
authors: P
authors: Rearden
authors: Duncan
authors: D
authors: Brown
authors: Sujal V
authors: SV
authors: Deshmukh
authors: Martha A
authors: MA
authors: DiPietro
authors: Jillian
authors: J
authors: DiMuzio
authors: Roy
authors: R
authors: Eisenhandler
authors: Scott E
authors: SE
authors: Fauty
authors: Christopher
authors: C
authors: Gibson
authors: Marian E
authors: ME
authors: Gindy
authors: Kelly A
authors: KA
authors: Hamilton
authors: Ian
authors: I
authors: Knemeyer
authors: Kenneth A
authors: KA
authors: Koeplinger
authors: Hae Won
authors: HW
authors: Kwon
authors: Traci Q
authors: TQ
authors: Lifsted
authors: Karsten
authors: K
authors: Menzel
authors: Mihir
authors: M
authors: Patel
authors: Nicole
authors: N
authors: Pudvah
authors: Deanne Jackson
authors: DJ
authors: Rudd
authors: Jessica
authors: J
authors: Seitzer
authors: Walter R
authors: WR
authors: Strapps
authors: Thomayant
authors: T
authors: Prueksaritanont
authors: Charles D
authors: CD
authors: Thompson
authors: Jerome H
authors: JH
authors: Hochman
authors: Brian A
authors: BA
authors: Carr
mesh_terms: D000818:Animals; D002626:Chemistry, Pharmaceutical; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D004008:Diclofenac / Q000378:metabolism; D055785:Gene Knockdown Techniques / Q000379:methods*; D066298:In Vitro Techniques; D008297:Male; D008861:Microsomes / Q000187:drug effects / Q000201:enzymology; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology; D008874:Midazolam / Q000378:metabolism; D053758:Nanoparticles; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000494:pharmacology*; D051381:Rats
chemical_list: D034741:RNA, Small Interfering; D004008:Diclofenac; D003577:Cytochrome P-450 Enzyme System; D008874:Midazolam