WO2009078876A1 - Procédé de dosage pour des réactions de transfert de groupes - Google Patents

Procédé de dosage pour des réactions de transfert de groupes Download PDF

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Publication number
WO2009078876A1
WO2009078876A1 PCT/US2007/088111 US2007088111W WO2009078876A1 WO 2009078876 A1 WO2009078876 A1 WO 2009078876A1 US 2007088111 W US2007088111 W US 2007088111W WO 2009078876 A1 WO2009078876 A1 WO 2009078876A1
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WO
WIPO (PCT)
Prior art keywords
antibody
sah
donor
tracer
reaction
Prior art date
Application number
PCT/US2007/088111
Other languages
English (en)
Inventor
Robert Lowery
Karen Kleman-Leyer
Matt Staeben
Thane Westermeyer
Original Assignee
Bellbrook Labs, Llc
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 Bellbrook Labs, Llc filed Critical Bellbrook Labs, Llc
Priority to EP07871707A priority Critical patent/EP2222708A1/fr
Publication of WO2009078876A1 publication Critical patent/WO2009078876A1/fr

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/44—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material not provided for elsewhere, e.g. haptens, metals, DNA, RNA, amino acids
    • 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
    • G01N33/536—Immunoassay; Biospecific binding assay; Materials therefor with immune complex formed in liquid phase
    • G01N33/542—Immunoassay; Biospecific binding assay; Materials therefor with immune complex formed in liquid phase with steric inhibition or signal modification, e.g. fluorescent quenching
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/90—Immunoglobulins specific features characterized by (pharmaco)kinetic aspects or by stability of the immunoglobulin
    • C07K2317/92—Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/90—Enzymes; Proenzymes
    • G01N2333/91—Transferases (2.)
    • G01N2333/91005—Transferases (2.) transferring one-carbon groups (2.1)
    • G01N2333/91011—Methyltransferases (general) (2.1.1.)

Definitions

  • linker positioned between the fluorophore and SAH, wherein the linker includes a chemical formula OfNH 2 -X-Z; wherein X is a saturated or unsaturated chain of 2 to 20 carbons and Z is a functional group capable of covalently binding to fluorophores or their activated derivatives; and wherein the group includes an NH 2 , SH, COOH or activated derivatives of COOH, such as succinimide or maleimide.
  • FIG. 3 illustrates use of FPIA to detect and quantify ADP formation, the donor product of the kinase reaction.
  • FIG. 14 shows examples of tracers used to quantify ADP formation.
  • FIG. 25 shows inhibition of SULTlEl by 2,6 Dichloro-4-nitrophenol (DCNP) measured with the FPIA-based assay.
  • FIG. 31 shows a standard curve for mock MT reaction with 1 O ⁇ M initial SAM showing stability of signal at room temperature over 20 hours.
  • SAH was added at the indicated concentrations and SAM, initially present at lO ⁇ M, was decreased proportionally to mimic the progress of a MT reaction.
  • Alexa 633 - ⁇ N- SAH tracer was present 2nM and Ab 3943 was used at its EC 8 O.
  • Acetyl transferase reaction acetyl CoenzymeA + acceptor -» acceptor-COCH 3 + CoenzymeA.
  • the screening is performed by a high- throughput screening technique, wherein the technique utilizes a multiwell plate or a microfluidic system.
  • bovine serum albumin BSA
  • KLH antigens generally elicit a stronger immune response in mammals, but also tend to be less soluble than BSA conjugates (Harlow, E. L., D., 1999).
  • Donor products with an adenine ring such as ADP and PAP may be attached to both carrier proteins via the C2, C8 and N6 amino group of adenine.
  • Donor products with a uridine ring can be conjugated via the C5 or C6 position of uridine.
  • Either type of nucleotide may be conjugated through the ribose 2' and 3 ' hydorxyls. Conjugation Methods
  • RTKs Growth factor receptor tyrosine kinases
  • RTKs are membrane- spanning proteins that transduce peptide growth factor signals from outside the cell to intracellular pathways that lead to activation of progrowth and anti-apoptotic genes.
  • the majority of the fifty-eight RTKs in humans are dominant oncogenes, meaning that aberrant activation or overexpression causes a malignant cell phenotype.
  • FP is a standard mode on several commercial HTS plate readers.
  • Figure 22 illustrates the effect of PAPS concentration on detection of enzymatically generated PAP.
  • the assay mixture included 200 ng of SULTlE l-6xHis (D) or assay buffer ( ⁇ ) was added to wells containing 30 mM phosphate (pH 7.4), 7 mM DTT, 8 mM MgCl 2 , 75 mM NaCl, 0.5 mg/mL BGG, 150 nM estradiol, 1 nM C8-PAP-F14 tracer, 12.5 uL Ab 3642, and varying concentrations of PAPS in a total assay volume of lOO ⁇ L.
  • D SULTlE l-6xHis
  • ⁇ assay buffer
  • DCNP was serially diluted two-fold into wells in 46 uL of phosphate assay buffer (30 mM KPO 4 (pH 6.5), 0.5 mg/mL BGG, 15 mM DTT, 1.6 mM MgCl 2 , 4 ⁇ M PAPS), followed by 50 ⁇ l of a 2X Antibody/tracer mix (5 uL 3642 Ab/ 2 nM Tracer C8- PAP-F 14), and 200 ng of SULTlE in a total volume of lOO ⁇ l. The plate was incubated at room temp for 30 min, and read on the Tecan Ultra. ⁇ mP values were calculated by subtracting the SULTlE reactions from the no SULTlE controls. All values represent the mean of replicates.
  • the reaction receptacle comprises a fluidic channel, and preferably, a microfluidic channel.
  • microfluidic refers to a channel or other conduit that has at least one cross-sectional dimension in the range of from about 1 micron to about 500 micron.
  • microfluidic devices useful for practicing the methods described herein include, e.g., those described in e.g., U.S. Pat. Nos. 5,942,443, 5,779,868, and International Patent Application No. WO 98/46438, the disclosures of which are incorporated herein by reference.
  • QBACTIVE ⁇ 6013956 1 that is attached to the body of the device and fluidly coupled to the reaction zone.
  • Such pipettor systems are described in, e.g., U.S. Pat. No. 5,779,868 (fully incorporated by reference).
  • the sampling Pipettor is serially dipped into different sources of test compounds which are separately and serially brought into the reaction zone to ascertain their affect, if any, on the reaction of interest.
  • kits designed to be used for detecting donor product or the catalytic activity generating the donor product through other means such as a homogenous assay, a homogeneous fluorescence intensity immunoassay, a homogeneous fluorescence lifetime immunoassay, a homogeneous fluorescence resonance energy transfer (FRET) immunoassay or a homogenous chemiluminescent immunoassay, or a non-homogenous assay such as enzyme- linked immunoassay (ELISA).
  • a homogenous assay such as a homogenous fluorescence intensity immunoassay, a homogeneous fluorescence lifetime immunoassay, a homogeneous fluorescence resonance energy transfer (FRET) immunoassay or a homogenous chemiluminescent immunoassay, or a non-homogenous assay such as enzyme- linked immunoassay (ELISA).
  • FRET fluorescence resonance energy transfer
  • ELISA enzyme- linked immunoassay

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  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Immunology (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Molecular Biology (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Hematology (AREA)
  • Biochemistry (AREA)
  • Medicinal Chemistry (AREA)
  • Urology & Nephrology (AREA)
  • Biomedical Technology (AREA)
  • Food Science & Technology (AREA)
  • Biophysics (AREA)
  • Analytical Chemistry (AREA)
  • Microbiology (AREA)
  • Cell Biology (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Physics & Mathematics (AREA)
  • Genetics & Genomics (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Biotechnology (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
  • Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

La présente invention concerne des procédés pour détecter, quantifier et cribler à haut débit des produits donneurs et les activités catalytiques générant les produits donneurs dans des réactions de transfert de groupes. Les enzymes catalysant les réactions de transfert de groupes comprennent les méthyltransférases, les sulfotransférases, les kinases, les glycosyltransférases, l'uridine glucuronide transférase, les UDP-glucuronosyltransférases, les acétyl transférases, les glutathione transférases, et les ADP-ribosyltransférases. L'invention concerne en outre des dosages immunologiques, des anticorps et des kits qui peuvent être utilisés pour appliquer les procédés de l'invention.
PCT/US2007/088111 2007-12-18 2007-12-19 Procédé de dosage pour des réactions de transfert de groupes WO2009078876A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
EP07871707A EP2222708A1 (fr) 2007-12-18 2007-12-19 Procédé de dosage pour des réactions de transfert de groupes

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US95851507A 2007-12-18 2007-12-18
US11/958,515 2007-12-18

Publications (1)

Publication Number Publication Date
WO2009078876A1 true WO2009078876A1 (fr) 2009-06-25

Family

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Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2007/088111 WO2009078876A1 (fr) 2007-12-18 2007-12-19 Procédé de dosage pour des réactions de transfert de groupes

Country Status (2)

Country Link
EP (1) EP2222708A1 (fr)
WO (1) WO2009078876A1 (fr)

Cited By (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012016704A1 (fr) 2010-08-05 2012-02-09 Cellzome Ag Procédés pour l'identification de molécules interagissant avec la méthyltransférase et pour la purification de protéines de type méthyltransférase
WO2014204861A1 (fr) * 2013-06-17 2014-12-24 The Trustees Of Columbia University In The City Of New York Procédés universels de profilage de la méthylation
EP2968386A1 (fr) * 2013-03-15 2016-01-20 Hunan Skyworld Biotechnologies Co. Immunoessai de la s-adénosylméthionine faisant appel à des analogues de cette dernière et thérapeutique personnalisée
WO2016008671A1 (fr) 2014-07-15 2016-01-21 Valitacell Limited Procédé de mesure de concentration d'anticorps dans un échantillon
JP2018516858A (ja) * 2015-04-06 2018-06-28 アルツス バイオシステムズ リミテッド ライアビリティ カンパニー ヘテロ環式化合物のバイオコンジュゲート
US10337049B2 (en) 2013-06-17 2019-07-02 The Trustees Of Columbia University In The City Of New York Universal methylation profiling methods
US10591463B2 (en) 2014-04-04 2020-03-17 Valitacell Limited Method of predicting phenotypic instability in a cell
US10626436B2 (en) 2015-04-01 2020-04-21 Valitacell Limited Method of determining a compositional or functional characteristic of a cell culture media
US20200339970A1 (en) * 2017-12-20 2020-10-29 Sevivo Llc Methods and kits for quantifying adenosine-containing molecules

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6063581A (en) * 1992-01-22 2000-05-16 Axis-Shield Asa Immunoassay for homocysteine
WO2006127008A1 (fr) * 2005-05-26 2006-11-30 Bellbrook Labs, Llc Procede d’analyse destine aux reactions a transfert de groupe

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6063581A (en) * 1992-01-22 2000-05-16 Axis-Shield Asa Immunoassay for homocysteine
WO2006127008A1 (fr) * 2005-05-26 2006-11-30 Bellbrook Labs, Llc Procede d’analyse destine aux reactions a transfert de groupe

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
LOWERY ROBERT G ET AL: "Transcreener: screening enzymes involved in covalent regulation.", EXPERT OPINION ON THERAPEUTIC TARGETS FEB 2006, vol. 10, no. 1, February 2006 (2006-02-01), pages 179 - 190, XP009103618, ISSN: 1744-7631 *

Cited By (17)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012016704A1 (fr) 2010-08-05 2012-02-09 Cellzome Ag Procédés pour l'identification de molécules interagissant avec la méthyltransférase et pour la purification de protéines de type méthyltransférase
EP2968386A4 (fr) * 2013-03-15 2017-06-07 Hunan Skyworld Biotechnologies Co. Immunoessai de la s-adénosylméthionine faisant appel à des analogues de cette dernière et thérapeutique personnalisée
EP3915584A3 (fr) * 2013-03-15 2022-02-16 Xiujuan Hao Immunoessai de la s-adénosylméthionine faisant appel à des analogues de cette dernière et thérapeutique personnalisée
EP2968386A1 (fr) * 2013-03-15 2016-01-20 Hunan Skyworld Biotechnologies Co. Immunoessai de la s-adénosylméthionine faisant appel à des analogues de cette dernière et thérapeutique personnalisée
JP2016513806A (ja) * 2013-03-15 2016-05-16 フナン スカイワールド バイオテクノロジーズ コンパニー Ｓ−アデノシルメチオニンの類似体を用いたｓ−アデノシルメチオニンのイムノアッセイおよび個別化治療法
US10337049B2 (en) 2013-06-17 2019-07-02 The Trustees Of Columbia University In The City Of New York Universal methylation profiling methods
WO2014204861A1 (fr) * 2013-06-17 2014-12-24 The Trustees Of Columbia University In The City Of New York Procédés universels de profilage de la méthylation
US10591463B2 (en) 2014-04-04 2020-03-17 Valitacell Limited Method of predicting phenotypic instability in a cell
CN106662581A (zh) * 2014-07-15 2017-05-10 瓦里泰细胞有限公司 一种测定样品中抗体浓度的方法
WO2016008671A1 (fr) 2014-07-15 2016-01-21 Valitacell Limited Procédé de mesure de concentration d'anticorps dans un échantillon
EP3795999A1 (fr) 2014-07-15 2021-03-24 Valitacell Limited Procédé permettant de déterminer l'abondance d'une molécule cible dans un échantillon
US10626436B2 (en) 2015-04-01 2020-04-21 Valitacell Limited Method of determining a compositional or functional characteristic of a cell culture media
JP2018516858A (ja) * 2015-04-06 2018-06-28 アルツス バイオシステムズ リミテッド ライアビリティ カンパニー ヘテロ環式化合物のバイオコンジュゲート
JP2021152027A (ja) * 2015-04-06 2021-09-30 フーナン スカイワールド バイオテクノロジーズ コンパニー ヘテロ環式化合物のバイオコンジュゲート
JP7327898B2 (ja) 2015-04-06 2023-08-16 タイツォウ フイフェン ヘタイ バイオテクノロジー カンパニー リミテッド ヘテロ環式化合物のバイオコンジュゲート
JP7344928B2 (ja) 2015-04-06 2023-09-14 タイツォウ フイフェン ヘタイ バイオテクノロジー カンパニー リミテッド ヘテロ環式化合物のバイオコンジュゲート
US20200339970A1 (en) * 2017-12-20 2020-10-29 Sevivo Llc Methods and kits for quantifying adenosine-containing molecules

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Publication number Publication date
EP2222708A1 (fr) 2010-09-01

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