WO1992012122A1 - Aminoalkylaldehydes a protection n - Google Patents

Aminoalkylaldehydes a protection n Download PDF

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Publication number
WO1992012122A1
WO1992012122A1 PCT/US1991/009791 US9109791W WO9212122A1 WO 1992012122 A1 WO1992012122 A1 WO 1992012122A1 US 9109791 W US9109791 W US 9109791W WO 9212122 A1 WO9212122 A1 WO 9212122A1
Authority
WO
WIPO (PCT)
Prior art keywords
protected
aminoalkylaldehyde
reagents
crosslinking
aminoalkylaldehydes
Prior art date
Application number
PCT/US1991/009791
Other languages
English (en)
Inventor
Maciej Adamczyk
Yon-Yi Chen
Original Assignee
Abbott Laboratories
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Abbott Laboratories filed Critical Abbott Laboratories
Publication of WO1992012122A1 publication Critical patent/WO1992012122A1/fr

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Classifications

    • 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
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54353—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals with ligand attached to the carrier via a chemical coupling agent
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C271/00—Derivatives of carbamic acids, i.e. compounds containing any of the groups, the nitrogen atom not being part of nitro or nitroso groups
    • C07C271/06—Esters of carbamic acids
    • C07C271/08—Esters of carbamic acids having oxygen atoms of carbamate groups bound to acyclic carbon atoms
    • C07C271/10—Esters of carbamic acids having oxygen atoms of carbamate groups bound to acyclic carbon atoms with the nitrogen atoms of the carbamate groups bound to hydrogen atoms or to acyclic carbon atoms
    • C07C271/18—Esters of carbamic acids having oxygen atoms of carbamate groups bound to acyclic carbon atoms with the nitrogen atoms of the carbamate groups bound to hydrogen atoms or to acyclic carbon atoms to carbon atoms of hydrocarbon radicals substituted by doubly-bound oxygen atoms

Definitions

  • the present invention relates to the preparation and use of heterobifunctional crosslinking reagents.
  • the present invention is related to N-protected aminoalkylaldehydes such as N-t-butylcarbonylaminoalkyl- aldehydes and N-benzyloxycarbonylaminoaldehydes, which are particularly useful as blocked crosslinking reagents.
  • bifunctional reagents i.e., reagents which have two reactive groups capable of reacting with, and forming bridges between, e.g., the side chains of amino acids and proteins
  • Homobifunctional reagents i.e., reagents carrying two identical reactive groups
  • Homobifunctional reagents are commonly used but their use is limited due to several inherent problems including random collisional crosslinking, crossreaction time, difficulty in controlling the reaction, and nonselective crosslinking [Kanaoka, et al., Chem. Phar . Bull., 32 (10) 3926 (1984)].
  • heterobifunctional reagents where the two reactive groups are sufficiently different to permit well-controlled seguential reactions of each group, are more desirable.
  • Even more desirable heterobifunctional reagents are those in which one of the reactive functional groups can be initially masked and subsequently can be readily unmasked and then used for coupling in a well-defined sequence of reaction steps, wherein uncontrolled crosslinking, often leading to undesired polymeric products, can be avoided [Weltman, et al., BioTechni ⁇ ues, 148-152 (Sept/Oct 1983); Jou, et al., Methods in Enzymology, Academic Press, Inc., vol.
  • heterobifunctional crosslinking reagents A number of different types have been described. For example, Kanaoka, et al., supra, describe multifunctional crosslinking reagents which are photoactivatable, thiol—directed fluorescent reagents.
  • Such heterobifunctional crosslinking reagents include m- maleimidobenzoyl-N-hydroxysuccinimide (MBS) for preparing antibody-beta-galactosidase conjugates, Fab'-beta- galactosidase conjugates, and the like; succinimidyl 4-(N- maleimidoethyl)cyclohexane-l-carboxylate (SMCC) for conjugating rabbit Fab' to horseradish peroxidase, conjugation of alkaline phosphatase and human IgG, and the like; sulfo-MBS for a comparison of maleimide containing heterobifunctional crosslinking reagents in the conjugation of Fab' fragments to horseradish peroxidase, and the like; and N-succinimidyl 3-2(2-pyridyldithio)propionate (SPDP) for introducing thiol groups into proteins and methods for
  • SUBSTITUTESHEET forming protein conjugates, preparation of antibody-toxin conjugates, and the like.
  • crosslinking reagents are very important in the rapidly growing field of biotechnology and immunology.
  • immunochemistry crosslinking applications wherein conjugates used in drug carrier systems, antibody production and enzyme immunoassays employing SPDP have been described.
  • Intramolecular crosslinking has been used to introduce additional tertiary structure into proteins, e.g., enzymes, in order to attempt to increase their conformational stability and to measure interresidue distances in proteins.
  • intermolecular crosslinking can be used to bind proteins of the same or different kinds to each other (protein-protein conjugation) and to modify cell membranes or other macromolecular assemblies.
  • BOC t-butoxycarbonyl
  • t-BOC N-t-butoxycarbonyl
  • protection of an amino group employing t-butyl carbamate has been described [Green, T.W., "Protective Groups in Organic synthesis", John Wiley & Sons, New York, page 232 [1981].
  • the lability and lipophilicity of the BOC group renders it unsuitable.
  • Cbz benzyloxycarbonyl
  • the present invention provides a method for preparing crosslinking reagents comprising novel N-protected aminoalkylaldehydes having aldehyde and amino termini which are useful for preparing conjugates with biologically active compounds having free amino and aldehyde groups.
  • the present invention relates to a simple and efficient method for preparing N-t-butyloxycarbonyl- aminoalkylaldehydes and N-benzyloxycarbonylaminoalkyl- aldehydes which are capable of being conjugated to such biologically active compounds by reductive amination with the free amino groups thereof.
  • N- benzyloxycarbonylaminoalkylalcoholsandN-butyloxycarbonyl- aminoalkylalcohols are oxidized with a Dess-Martin periodinane reagent to provide such novel N-protected aminoaldehydes for use as heterobifunctional crosslinking reagents.
  • the resulting crosslinking reagents are particularly useful for preparing labeled reagents useful in immunoassays, radioimmunoassays, and the like, which require the conjugation of a biologically active compound with a detectable moiety for use as a labeled reagent therein.
  • FIG. 1 illustrates the schematic pathway for preparing
  • N-t-butyloxycarbonylaminoalkylaldehyde according to the present invention.
  • Fig. 2 illustrates the schematic pathway for preparing
  • N-benzyloxycarbonylaminoalkylaldehyde according to the present invention.
  • N-Boc-aminoalkylalcohols (2.), as well as N-Cbz-aminoalkylalcohols (5 . ), can be readily oxidized with Dess-Martin periodinane reagents [Dess, et al., J. Org. Chem. , 48:4156-4158 (1983)] in methylene chloride to give desired N-protected aminoalkylaldehydes (3 . and 6.) .
  • the use of hypervalent iodine in organic synthesis is described in Moriarty, et al., Ace. Chem. Res. , 19:244 (1986).
  • the Dess-Martin reagent originally described by Dess, et al., supra, page 4155, is a mild oxidative procedure which does not involve acid and does not generate any undesired by-products. After mild alkaline treatment, crude products exhibited only 1 spot by thin layer chromatography (silica gel, ethyl acetate/hexanes [1:1] as an eluent) and provided satisfactory 1H NMR spectra. The products can be used as crosslinking reagents without further purification.
  • crosslinking reagents prepared according to the present invention are particularly useful for preparing labeled antibody conjugates for use in homogeneous and heterogeneous immunoassay systems known in the art, such as competitive immunoassays, sandwich immunoassays, immunometric assays, and the like, to determine the amount
  • SUBSTITUTESHEET of analyte present in a test sample depend upon the ability of an immunoglobulin, i.e., a whole antibody or fragment thereof, to bind to a specific analyte wherein a conjugate comprising an antibody to such analyte conjugated with a label or detectable moiety known in the art is employed to determine the extent of such binding.
  • detectable moieties or labels include, but are not intended to be limited to, enzymes, chromogens, luminescent compounds, phosphorescent compounds, chemiluminescent compounds, fluorescent compounds, and the like.
  • the extent of binding is determined by the amount of the detectable moiety present in the conjugate which either has or has not participated in a binding reaction with the analyte, wherein the amount of the detectable moiety detected and measured can be correlated to the amount of analyte present in the test sample.
  • the free aldehyde of the N-protected aminoalkylaldehyde can be reacted by reductive amination with any free amino group.
  • the protecting group can be removed from the heterobifunctional reagent by mild reduction for the removal of Cbz and by mild acid for the removal of BOC. Once the protecting group has been removed, a free amino group is available to which can be linked any molecule having a free aldehyde group.
  • the free aldehyde of the N-protected amino alkylaldehyde can be linked to the amino group of a protein rather than, for example, a column support.
  • the protecting group can be removed from the heterobifunctional linker to expose a free amino group which can be reacted with any free aldehyde groups present, such as on a column support.
  • crosslinking reagents of the present invention include, but are not intended to be limited to, antibody mediated delivery systems by providing a therapeutic antibody conjugate that selectively localizes to tumor cell sites and requires that an adequate number of the drug molecules reach their site of action within the tumor cells where they can then exert a cytotoxic or cytostatic effect; covalent modification of antibodies and the design and synthesis of specialized crosslinking reagents in order to retain the homogeneous binding properties of the antibody conjugate; in vivo imaging applications which require discrete functional elements
  • SUBSTITUTESHEET and, in particular, must have chelator groups capable of strong association to any one of the family of radioactive metal cations; and the like.

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  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Immunology (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Urology & Nephrology (AREA)
  • Biomedical Technology (AREA)
  • Hematology (AREA)
  • Molecular Biology (AREA)
  • Biotechnology (AREA)
  • Microbiology (AREA)
  • Food Science & Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Cell Biology (AREA)
  • Peptides Or Proteins (AREA)
  • Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)

Abstract

Procédé de synthétisation de nouveaux aminoalkylaldéhydes à protection N ayant des terminaisons aldéhyde et amino et qui sont utiles en temps que réactifs réticulants dans la préparation de conjugués avec des composés biologiquement actifs. Des alcools de N-benzyloxy-carbonylaminoalkyles et des alcools de N-butyloxy-carbonylaminoalkyles sont oxydés avec de la périodinane Dess-Martin pour produire des aminoaldéhydes à protection N destinés à être utilisés comme réactifs hétérobifonctionnels de réticulation.
PCT/US1991/009791 1990-12-27 1991-12-27 Aminoalkylaldehydes a protection n WO1992012122A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US63468090A 1990-12-27 1990-12-27
US634,680 1990-12-27

Publications (1)

Publication Number Publication Date
WO1992012122A1 true WO1992012122A1 (fr) 1992-07-23

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WO (1) WO1992012122A1 (fr)

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7459469B2 (en) 2004-11-10 2008-12-02 Targacept, Inc. Hydroxybenzoate salts of metanicotine compounds
US8017785B2 (en) 2006-05-09 2011-09-13 Astrazeneca Ab Salt forms of (2S)-(4E)-N-methyl-5-[3-(5-isopropoxypyridin)y1]-4-penten 2-amine
EP2371818A1 (fr) 2004-11-10 2011-10-05 Targacept, Inc. Sels hydroxybenzoates de composés de métanicotine
US8461344B2 (en) 2006-05-09 2013-06-11 Targacept, Inc. Polymorph forms of (2S)-(4E)-N-methyl-5-[3-(5-isopropdxypyridin)yl]-4-penten-2-amine
US8703802B2 (en) 2010-05-20 2014-04-22 Targacept, Inc. Process for the preparation of aryl substituted olefinic amines

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US1927858A (en) * 1930-12-27 1933-09-26 Ig Farbenindustrie Ag Urethane derivatives and alpha process for their production

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US1927858A (en) * 1930-12-27 1933-09-26 Ig Farbenindustrie Ag Urethane derivatives and alpha process for their production

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
J. MED. CHEM., Volume 28, No. 3, issued March 1985, B.H. LEE et al., "Artificial Siderophores", see pages 317-323. *
J. URY. CHEM., Volume 48, No. 22, issued November 1983, D.B. DESS, "Readily Accesible 12-I-51 Oxidant for the Conversion of Primary and Secondary Alcohols to Aldphydes and Ketones", see pages 4155 to 4156. *
T. GREENE, "Protective Groups In Organic Synthesis", published 1981 by JOHN WILEY & SONS, see pages 218 to 224, 232, 233, 239 to 247. *
TETRAHEDRON, Volume 31, No. 23, issued 1975, L.J. GRAY, "The Synthesis of an oc-Azaornithine Derivative and its Reaction with Trypisia", see pages 2940-2943. *

Cited By (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7459469B2 (en) 2004-11-10 2008-12-02 Targacept, Inc. Hydroxybenzoate salts of metanicotine compounds
EP2371818A1 (fr) 2004-11-10 2011-10-05 Targacept, Inc. Sels hydroxybenzoates de composés de métanicotine
US8053451B2 (en) 2004-11-10 2011-11-08 Targacept, Inc. Hydroxybenzoate salts of metanicotine compounds
US8580826B2 (en) 2004-11-10 2013-11-12 Targacept, Inc. Hydroxybenzoate salts of metanicotine compounds
US8778978B2 (en) 2004-11-10 2014-07-15 Targacept, Inc. Hydroxybenzoate salts of metanicotine compounds
US9107915B2 (en) 2004-11-10 2015-08-18 Targacept, Inc. Hydroxybenzoate salts of metanicotine compounds
US8017785B2 (en) 2006-05-09 2011-09-13 Astrazeneca Ab Salt forms of (2S)-(4E)-N-methyl-5-[3-(5-isopropoxypyridin)y1]-4-penten 2-amine
US8461344B2 (en) 2006-05-09 2013-06-11 Targacept, Inc. Polymorph forms of (2S)-(4E)-N-methyl-5-[3-(5-isopropdxypyridin)yl]-4-penten-2-amine
US8703802B2 (en) 2010-05-20 2014-04-22 Targacept, Inc. Process for the preparation of aryl substituted olefinic amines

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