CA2553769A1 - Npc1l1 (npc3) and methods of identifying ligands thereof - Google Patents

Npc1l1 (npc3) and methods of identifying ligands thereof Download PDF

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Publication number
CA2553769A1
CA2553769A1 CA002553769A CA2553769A CA2553769A1 CA 2553769 A1 CA2553769 A1 CA 2553769A1 CA 002553769 A CA002553769 A CA 002553769A CA 2553769 A CA2553769 A CA 2553769A CA 2553769 A1 CA2553769 A1 CA 2553769A1
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Prior art keywords
azetidinone
glucuronide
substituted
value
npc1l1
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CA002553769A
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French (fr)
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CA2553769C (en
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Margarita Garcia-Calvo
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Merck Sharp and Dohme LLC
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/60—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving cholesterol
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/5005—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
    • G01N33/5008—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
    • G01N33/5076—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics involving cell organelles, e.g. Golgi complex, endoplasmic reticulum
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2500/00—Screening for compounds of potential therapeutic value

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Immunology (AREA)
  • Molecular Biology (AREA)
  • General Health & Medical Sciences (AREA)
  • Zoology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Biomedical Technology (AREA)
  • Biochemistry (AREA)
  • Analytical Chemistry (AREA)
  • Wood Science & Technology (AREA)
  • Urology & Nephrology (AREA)
  • Biophysics (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Medicinal Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Genetics & Genomics (AREA)
  • Microbiology (AREA)
  • Toxicology (AREA)
  • Biotechnology (AREA)
  • Cell Biology (AREA)
  • Hematology (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Food Science & Technology (AREA)
  • General Engineering & Computer Science (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Peptides Or Proteins (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

The present invention provides human, rat and mouse NPCIL1 polypeptides and polynucleotides encoding the polypeptides. Methods for detecting ligands which bind to NPClL1 and block intestinal cholesterol absorption are provided. Also included is a method of identifying ligands which bind to NPCILI using membranes derived from brush border membrane preparations. Compounds that bind to NPCILI can be used for inhibiting intestinal cholesterol absorption in a subject.

Claims (20)

1. A method for identifying a ligand of NPC 1 L1 comprising:
contacting human NPC1L1 with a detectably labeled substituted 2-azetidinone glucuronide and a candidate compound; and determining whether said candidate compound binds to human NPC1L1;
wherein binding of said candidate compound to human NPC1L1 modulates binding of said detectably labeled substituted 2-azetidinone glucuronide to human NPC1L1, wherein the detectably labeled substituted 2-azetidinone glucuronide has a binding affinity KD value for human NPC1L1 that is 200nM or lower, and wherein said modulation indicates that the candidate compound is a ligand that binds to human NPC1L1.
2. The method of claim 1, wherein the KD value is 100nM or lower.
3. The method of claim 1, wherein the KD value is 50nM or lower.
4. The method of claim 1, wherein the KD value is 20nM or lower.
5. The method of claim 1, wherein the KD value is 10nM or lower.
6. The method of claim 1, wherein the substituted 2-azetidinone-glucuronide is selected from the group consisting of a compound of Formula I and a compound of Formula II.
7. The method of claim 6, wherein the substituted 2-azetidinone-glucuronide comprises a detectable label from the group consisting of 35S and 125I.
S. The method of claim 7, wherein the detectable label is 35S.
9. The method of claim 6, wherein the substituted 2-azetidinone-glucuronide is a compound of Formula II, wherein R9 comprises an -SO2- group.
10. The method of claim 9, wherein the substituted 2-azetidinone-glucuronide of Formula II is labeled with 35S.
11. A method for identifying a ligand of NPC1L1 comprising:
contacting human NPC1L1 with a detestably labeled substituted 2-azetidinone glucuronide of Formula II and a candidate compound; and determining whether said candidate compound binds to human NPC1L1;
wherein binding of said candidate compound to human NPC1L1 modulates binding of said detestably labeled substituted 2-azetidinone glucuronide of Formula II
to human NPC1L1, and wherein said modulation indicates that the candidate compound is a ligand that binds to human NPC1L1.
12. The method of claim 11, wherein R9 of the detestably labeled substituted 2-azetidinone glucuronide of Formula II comprises an -SO2- group.
13. The method of claim 11, wherein the detestably labeled substituted 2-azetidinone glucuronide of Formula II is labeled with 35S.
14. The method of claim 11, wherein the detestably labeled substituted 2-azetidinone glucuronide of Formula II has a binding affinity K D value for human NPC1L1 that is 200nM or lower.
15. The method of claim 14, wherein the K D value is 100nM or lower.
16. The method of claim 14, wherein the K D value is 50nM or lower.
17. The method of claim 14, wherein the K D value is 20nM or lower.
18. The method of claim 14, wherein the K D value is 10nM or lower.
19. The method of claim 1 wherein the detestably labeled substituted 2-azetidinone glucuronide is labeled with 35S.
20. The method of claim 1 wherein the detestably labeled substituted 2-azetidinone glucuronide is 35S-labeled compound 2.
CA2553769A 2004-01-16 2005-01-14 Npc1l1 (npc3) and methods of identifying ligands thereof Expired - Fee Related CA2553769C (en)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
US53734104P 2004-01-16 2004-01-16
US60/537,341 2004-01-16
PCT/US2005/001469 WO2005069900A2 (en) 2004-01-16 2005-01-14 Npc1l1 (npc3) and methods of identifying ligands thereof

Publications (2)

Publication Number Publication Date
CA2553769A1 true CA2553769A1 (en) 2005-08-04
CA2553769C CA2553769C (en) 2011-01-04

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Family Applications (1)

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CA2553769A Expired - Fee Related CA2553769C (en) 2004-01-16 2005-01-14 Npc1l1 (npc3) and methods of identifying ligands thereof

Country Status (6)

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US (1) US7901893B2 (en)
EP (1) EP1723414A4 (en)
JP (1) JP4590417B2 (en)
CA (1) CA2553769C (en)
MX (1) MXPA06008124A (en)
WO (1) WO2005069900A2 (en)

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US20040132058A1 (en) * 2002-07-19 2004-07-08 Schering Corporation NPC1L1 (NPC3) and methods of use thereof
ATE485267T1 (en) 2003-12-23 2010-11-15 Astrazeneca Ab DIPHENYLAZETIDINONE DERIVATIVES WITH CHOLESTERINE ABSORPTION INHIBITING EFFECT
AU2004308332B2 (en) 2003-12-23 2008-04-10 Merck Sharp & Dohme Corp. Anti-hypercholesterolemic compounds
MXPA06008124A (en) 2004-01-16 2007-01-26 Merck & Co Inc Npc1l1 (npc3) and methods of identifying ligands thereof.
JP2008543837A (en) * 2005-06-15 2008-12-04 メルク エンド カムパニー インコーポレーテッド Anti-hypercholesterolemic compound
UY29607A1 (en) 2005-06-20 2007-01-31 Astrazeneca Ab CHEMICAL COMPOUNDS
MY148538A (en) 2005-06-22 2013-04-30 Astrazeneca Ab Novel 2-azetidinone derivatives as cholesterol absorption inhibitors for the treatment of hyperlipidaemic conditions
SA06270191B1 (en) 2005-06-22 2010-03-29 استرازينيكا ايه بي Novel 2-Azetidinone Derivatives as Cholesterol Absorption Inhibitors for the Treatment of Hyperlipidaemic Conditions
DE102005055726A1 (en) * 2005-11-23 2007-08-30 Sanofi-Aventis Deutschland Gmbh Hydroxy-substituted diphenylazetidinones, processes for their preparation, medicaments containing these compounds and their use
US7910698B2 (en) 2006-02-24 2011-03-22 Schering Corporation NPC1L1 orthologues
AR060623A1 (en) 2006-04-27 2008-07-02 Astrazeneca Ab COMPOUNDS DERIVED FROM 2-AZETIDINONE AND A PREPARATION METHOD
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WO2009138035A1 (en) * 2008-05-13 2009-11-19 中国科学院上海生命科学研究院 Screening method for medicine of blocking cholesterol absorption based on analyzing subcellular localization of npc1l1 protein
CN102869382A (en) * 2010-06-10 2013-01-09 (株)Ad生物技 Composition for inhibiting hyperlipidemia and obesity by inhibiting intestinal absorption of cholesterol
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Also Published As

Publication number Publication date
JP4590417B2 (en) 2010-12-01
JP2007526998A (en) 2007-09-20
CA2553769C (en) 2011-01-04
MXPA06008124A (en) 2007-01-26
EP1723414A2 (en) 2006-11-22
WO2005069900A2 (en) 2005-08-04
US7901893B2 (en) 2011-03-08
EP1723414A4 (en) 2008-03-26
US20100009461A1 (en) 2010-01-14
WO2005069900A3 (en) 2006-06-08
WO2005069900B1 (en) 2006-08-03

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