The target of ezetimibe is Niemann-Pick C1-Like 1 (NPC1L1)

doi:10.1073/pnas.0500269102

The target of ezetimibe is Niemann-Pick C1-Like 1 (NPC1L1)

Departments of ^aMetabolic Disorders, ^fDrug Metabolism Labeled Compound Synthesis, ^gImmunology, ^hPharmacology, ⁱMedicinal Chemistry, and ^kTarget Validation, Merck Research Laboratories, Rahway, NJ 07065; and Departments of ^dCardiovascular/Metabolic Diseases, ^eChemistry, and ^jDiscovery Technologies, Schering-Plough Research Institute, Kenilworth, NJ 07033

Edited by Joseph L. Goldstein, University of Texas Southwestern Medical Center, Dallas, TX (received for review January 12, 2005)

Abstract

Ezetimibe is a potent inhibitor of cholesterol absorption that has been approved for the treatment of hypercholesterolemia, but its molecular target has been elusive. Using a genetic approach, we recently identified Niemann-Pick C1-Like 1 (NPC1L1) as a critical mediator of cholesterol absorption and an essential component of the ezetimibe-sensitive pathway. To determine whether NPC1L1 is the direct molecular target of ezetimibe, we have developed a binding assay and shown that labeled ezetimibe glucuronide binds specifically to a single site in brush border membranes and to human embryonic kidney 293 cells expressing NPC1L1. Moreover, the binding affinities of ezetimibe and several key analogs to recombinant NPC1L1 are virtually identical to those observed for native enterocyte membranes. K _D values of ezetimibe glucuronide for mouse, rat, rhesus monkey, and human NPC1L1 are 12,000, 540, 40, and 220 nM, respectively. Last, ezetimibe no longer binds to membranes from NPC1L1 knockout mice. These results unequivocally establish NPC1L1 as the direct target of ezetimibe and should facilitate efforts to identify the molecular mechanism of cholesterol transport.

Footnotes

↵ b To whom correspondence should be addressed at: Department of Metabolic Disorders, Merck Research Laboratories, Mail Stop RY50G241, P.O. Box 2000, Rahway, NJ 07065. E-mail: margarita_garcia-calvo{at}merck.com.
↵ c M.G.-C., J.L., H.G.B., B.E.H., D.A.B., M.P.B., J.H.C., H.R.D., D.C.D., P.A.D., M.P.G., M.H., D.E.M., A.O., K.A.O., S.P.N.I., D.E.S., M.M.S., Y.S.T., A.M.M., F.U., S.W.A., K.T.C., and N.A.T. are employees of Merck and/or Schering-Plough, which comarket ezetimibe (Zetia and Vytorin) as a cholesterol-absorption inhibitor.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations: NPC1L1, Niemann-Pick C1-Like 1; HEK, human embryonic kidney; SSD, sterol-sensing domain; EZE-glue, ezetimibe glucuronide; BBM, brush border membranes.