WO2007122291A1 - Titanocènes pour le marquage de réactifs de liaison biospécifiques - Google Patents

Titanocènes pour le marquage de réactifs de liaison biospécifiques Download PDF

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Publication number
WO2007122291A1
WO2007122291A1 PCT/FI2007/050208 FI2007050208W WO2007122291A1 WO 2007122291 A1 WO2007122291 A1 WO 2007122291A1 FI 2007050208 W FI2007050208 W FI 2007050208W WO 2007122291 A1 WO2007122291 A1 WO 2007122291A1
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WO
WIPO (PCT)
Prior art keywords
labelling
conjugate
chelates
reactant
reactant according
Prior art date
Application number
PCT/FI2007/050208
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English (en)
Inventor
Jari Hovinen
Original Assignee
Wallac Oy
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 Wallac Oy filed Critical Wallac Oy
Publication of WO2007122291A1 publication Critical patent/WO2007122291A1/fr

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
    • C07F17/00—Metallocenes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K49/00—Preparations for testing in vivo
    • A61K49/0002—General or multifunctional contrast agents, e.g. chelated agents

Definitions

  • This invention relates to novel titanocene derivatives tethered to reporter groups which can be introduced into biospecific binding reactants.
  • Oligonucleotides, DNA and RNA can be transformed statistically by bisulfite-catalyzed transamination of cytosine residues [Hayatsu, H., 1976, Biochemistry, 15, 2677, Draper, D. E., Gold, L., 1980, Biochemistry, 19, 1774, Molander, J., Hurskainen, P., Hovinen, J., Lahti, M., L ⁇ nnberg, H., 1993, Bio- conjugate Chem., 4, 362, Adarichev, V.A., Kalachnikov, S. M., Kiseliova, A.V., Dymshits, G.
  • N7 of guanine residues react but also other bases such as N1 of adenine residues [Lippert, B. 2000, Coordination Chemistry Rev., Lippert B., 1989, Progr. Inorg. Chem., 37, 1 , Lippert B., Cisplatin, Chemistry and Biochemistry of a Leading Anticancer Drug/; Ed.
  • Titanocene dichloride and its analogues have been shown to have significant activity against a various cancer cell lines [K ⁇ pf-Maier, P., K ⁇ pf, H. Chem. Rev., 1987, 87, 1137, Clarke, M.J., Zhu, F., Frasca, D. R., 1999, Chem. Rev., 99, 2511 , Harding, M. M., Mokshdi, G., 2000, Curr. Med. Chem., 7, 1289, Yang, P., Guo, M., 1999, Coord. Chem. Rev., 185, 189, Melendez, E., 2002, Crit. Rev.
  • titanocene tethered to an acid chloride was allowed to react with various alcohols and amines giving rise to a multitude of titanocene conjugates.
  • One of the obstacles in the use of titanocens has been their very low aqueous solubility. Accordingly, several attempts to increase their hydro- philicity have been reported.
  • US 5,296,237 teaches to mix metal- locenes with polyols and various additives to give rise to water-soluble metal- locene-complexes. Although these metallocene-complex compositions can be used as cytostatics in cancer therapy, they are not suitable for analytical pur- poses.
  • the fluorescent labels used for biomolecule derivatization are most commonly organic dyes. Although organic chromophores can be utilized in several applications, these labels suffer from many commonly known drawbacks such as Raman and Raylegh scattering, low water solubility and concentration quenching. Instead, the unique properties of lanthanide(lll) chelates, such as high water solubility and strong long decay-time luminescence, make them ideal labels for numerous assays. Furthermore, large Stokes shift and very sharp emission bands enable the simultaneous use of four lanthanides (i.e. Eu, Tb, Sm, Dy) in the analysis. Time-resolved fluori- metric assays based on lanthanide chelates have found increasing applications in diagnostics, research and high throughput screening.
  • the heterogeneous DELFIA ® (Dissociation-Enhanced Lanthanide Fluorescence Immuno- assay) technique is applied in assays requiring exceptional sensitivity, robustness and a multi-label approach.
  • Development of highly luminescent stable chelates extends the use of time resolution to homogeneous assays, based on fluorescence resonance energy transfer (TR-FRET), fluorescence quenching (TR-FQA) or changes in luminescence properties of a chelate during a binding reaction.
  • TR-FRET fluorescence resonance energy transfer
  • TR-FQA fluorescence quenching
  • the main object of the present invention is to provide titanocene derivatives which allow labelling of biospecific binding reactants. These derivatives can be used in diagnostic and therapeutic applications. Accordingly, they are suitable for in vitro diagnostics, molecular imaging, and they can be used as radiopharmaceuticals.
  • the major advantages of the present invention are: (i) The present titanocenes are derivatives of metal chelates with high water solubility. Accordingly, they do not suffer from the hydrophobi- city of titanocene dichloride. (ii) The present labelling reactants do not suffer from concentration quenching, and thus they are highly suitable for multilabelling. (iii) Since they bind to the biomolecule statistically, the labelling degree can be controlled simply by the ratio of the reactants. (iv) The present reporter groups can be used in the labelling of oligonuc- leotides, LNA, DNA, RNA, phosphopeptides and phosphoproteins.
  • the present invention concerns a labelling reactant of a formula (I) suitable for labelling of biospecific binding reactants
  • L 1 and L 2 are a linkers, same or different or not present; G 1 and G 2 are conjugate groups, same or different, or not present; X is Cl or
  • the conjugate groups can be, for example, a luminescent or non- luminescent lanthanide(lll) chelates, labels suitable for SPECT or PET applications, biotin or a spin label.
  • the invention concerns a biospecific binding reactant conjugated with the labelling reactant according to this invention.
  • the target can be, for example, a synthetic oligonucleotide, DNA or RNA as well as a phosphopeptide or a phos- phoprotein.
  • the molecule of formula (I) reacts statistically, and the degree of labelling can be adjusted by controlling the ratio of the reaction components.
  • the molecule of formula (I) reacts with phosphoproteins and phosphopeptides in physiological pH with no need of phosphoelimination at highly basic conditions.
  • Ar is cyclopenta- 1 ,3-diene.
  • the conjugate groups G 1 -G 3 are reporter groups, such as metal chelates.
  • reporter groups such as metal chelates.
  • luminescent or non-luminescent lanthanide(lll) chelates particularly chelates of europium (Eu), terbium (Tb) or gadolinium (Gd).
  • Particularly preferable lanthanide (III) chelates for this purpose are luminescent chelates based on triazacycloalkanes, 2-(3-(pyridin-2-yl)-1 H-pyrazol-1 -yl)pyridine and terpyridine and nonluminescent chelates based on pyridine-2,6-diyl-bis(methylenenitrilo) tetrakis(acetic acid) and diethylenetriaminepentaacetic.
  • other metal chelates can be used.
  • One of the conjugate groups G 1 -G 3 can also be a group useful for immobilizing the target biomolecule to as solid support.
  • the conjugate groups G 1 -G 3 are metal chelates comprising terpyridine, 2-(3-(pyridin-2-yl)-1 H- pyrazol-1-yl)pyridine or diethylene triamine bound to a group of formula (II)
  • R 1 is methylene or 1-butynyl and is bound further to L 1 , L 2 or A of formula (I) and R 2 is a single bond or alkyl, where the alkyl is methylene or ethylene, and is bound further to the terpyridine, the 2-(3-(pyridin-2-yl)-1 H- pyrazol-1-yl)pyridine or the diethylene triamine.

Abstract

L'invention concerne un nouveau réactif de marquage de formule (I) permettant de marquer des réactifs de liaison biospécifiques. Dans la formule (I), L1 et L2 sont un liant, identiques ou différents, ou absents, G1 et G2 sont des groupes conjugués, identiques ou différents, ou absents, X est Cl ou -A-G3, où A est soufre ou oxygène et G3 un groupe conjugué, et Ar est un noyau aromatique, à condition que les groupes conjugués (G1, G2 et G3) ne soient pas tous simultanément absents.
PCT/FI2007/050208 2006-04-21 2007-04-20 Titanocènes pour le marquage de réactifs de liaison biospécifiques WO2007122291A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US79372206P 2006-04-21 2006-04-21
US60/793,722 2006-04-21

Publications (1)

Publication Number Publication Date
WO2007122291A1 true WO2007122291A1 (fr) 2007-11-01

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Family Applications (1)

Application Number Title Priority Date Filing Date
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WO (1) WO2007122291A1 (fr)

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1997036931A1 (fr) * 1996-04-01 1997-10-09 British Nuclear Fuels Plc Systeme de dosage et nouveaux composes marques utiles pour ce dosage
WO1998007760A1 (fr) * 1996-08-16 1998-02-26 Targor Gmbh Systeme de catalyseur

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1997036931A1 (fr) * 1996-04-01 1997-10-09 British Nuclear Fuels Plc Systeme de dosage et nouveaux composes marques utiles pour ce dosage
WO1998007760A1 (fr) * 1996-08-16 1998-02-26 Targor Gmbh Systeme de catalyseur

Non-Patent Citations (8)

* Cited by examiner, † Cited by third party
Title
ANONYMOUS: "Novel preparation of transition metal dichloride complexes", RESEARCH DISCLOSURE, vol. 445, no. 105, May 2001 (2001-05-01), XP007128223, ISSN: 0374-4353 *
CAUSEY P W ET AL: "Synthesis, characterization, and assessment of cytotoxic properties of a series of titanocene dichloride derivatives", ORGANOMETALLICS, vol. 23, no. 19, 13 September 2004 (2004-09-13), pages 4486 - 4494, XP009090001, ISSN: 0276-7333 *
HARDING M M ET AL: "Antitumour metallocenes: structure-activity studies and interactions with biomolecules", CURRENT MEDICINAL CHEMISTRY, vol. 7, no. 12, December 2000 (2000-12-01), pages 1289 - 1303, XP009090003, ISSN: 0929-8673 *
HOGAN ET AL: "Synthesis and Cytotoxicity Studies of New Dimethylamino-Functionalized and Azole-Substituted Titanocene Anticancer Drugs", ORGANOMETALLICS, vol. 26, 2007, pages 2501 - 2506, XP009090033, ISSN: 0276-7333 *
JUTZI P ET AL: "Titanium and zirconium bent-sandwich complexes with the new (2-(diisopropylamino)ethyl)cyclopentadienyl ligand: catalysts for the polymerization of ethylene and the dehydrocoupling of phenylsilane", ORGANOMETALLICS, vol. 15, no. 15, 1996, pages 4153 - 4161, XP000905650, ISSN: 0276-7333 *
PAMPILLON C ET AL: "Synthesis and cytotoxicity studies of new dimethylamino-functionalised and heteroaryl-substituted titanocene anti-cancer drugs", JOURNAL OF ORGANOMETALLIC CHEMISTRY, vol. 692, no. 11, 5 April 2007 (2007-04-05), pages 2153 - 2159, XP022022455, ISSN: 0022-328X *
PINKAS J ET AL: "Synthesis and structure of bis{eta5-1,2,3,4-tetramethyl- 5-(dimethyl silylsulfido-kappaS) cyclopentadienyl}ti tanium(IV)", INORGANIC CHEMISTRY COMMUNICATIONS, vol. 7, no. 10, October 2004 (2004-10-01), pages 1135 - 1138, XP004587720, ISSN: 1387-7003 *
QIAN Y L ET AL: "Synthesis of terminal amino substituted cyclopentadienyl titanium complexes", CHINESE CHEMICAL LETTERS, vol. 7, no. 12, 1996, pages 1139 - 1142, XP009090041, ISSN: 1001-8417 *

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