Purine based fluorescent dyes

• US 7,737,281 B2
• Filed: 07/07/2005
• Issued: 06/15/2010
• Est. Priority Date: 05/24/2005
• Status: Active Grant

• Current Assignee: Enzo Life Sciences Incorporated (Enzo Biochem Incorporated)
• Original Assignee: Enzo Life Sciences Incorporated (Enzo Biochem Incorporated)
• Inventors: Szczepanik, Maciej
• Primary Examiner(s): SEAMAN, D MARGARET M
• Application Number:
  US11/177,923
• Publication Number:
  US 20060269931A1
• Time in Patent Office: 1,804 Days
• Field of Search: 546/16, 546/270.1, 546/271.1
• US Class Current: 546/270.1
• CPC Class Codes: C07D 401/14 containing three or more hetero rings
• CPC Class Codes: C07D 405/14 containing three or more hetero rings
• CPC Class Codes: C07D 417/06 linked by a carbon chain containing only aliphatic carbon atoms
• CPC Class Codes: C07D 417/14 containing three or more hetero rings
• CPC Class Codes: C07D 473/00 Heterocyclic compounds containing purine ring systems
• CPC Class Codes: C07F 9/65616 containing the ring system having three or more than three double bonds between ring members or between ring members and non-ring members, e.g. purine or analogs
• CPC Class Codes: C09B 23/0091 having only one heterocyclic ring at one end of the methine chain, e.g. hemicyamines, hemioxonol styryl dyes see C09B23/14
• CPC Class Codes: C09B 23/04 one >CH- group, e.g. cyanines, isocyanines, pseudocyanines
• CPC Class Codes: C09B 23/06 three >CH- groups, e.g. carbocyanines
• CPC Class Codes: C09B 23/083 five >CH- groups
• CPC Class Codes: C09B 23/105 two >CH- groups
• CPC Class Codes: C09B 23/145 the ethylene chain carrying an heterocyclic residue, e.g. heterocycle-CH=CH-C6H5
• CPC Class Codes: C09B 23/146 (Benzo)thiazolstyrylamino dyes
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 2563/107 fluorescence

Abstract

The present invention provides novel purine-based fluorescent dyes that may be used for staining, localizing and otherwise labeling target molecules, such as nucleic acids, for detection, amplification and quantification.

Claims

1. A dye having the formula:

2. The dye of claim 1, wherein at least one of R¹, R², R³, R⁵, R⁶, R⁷, R⁸, R¹⁰, R¹¹, R¹³, R¹⁴, R¹⁵, R¹⁶, R¹⁷ or R¹⁸ comprises a reactive group.

3. The dye of claim 2, wherein said reactive group comprises a nucleophilic reactive group, an electrophilic reactive group, a terminal alkene, a terminal alkyne, a coordinate group or an alkylating agent.

4. The dye of claim 3, wherein said nucleophilic reactive group comprises a thiol, amine, hydrazine or hydroxyl group.

5. The dye of claim 3, wherein said electrophilic reactive group comprises an isocyanate, isothiocyanate, monochlorotriazine, dichlorotriazine, 4,6-dichloro-1,3,5-triazines, mono- or di-halogen substituted pyridine, mono- or di-halogen substituted diazine, maleimide, haloacetamide, aziridine, sulfonyl halide, acid halide, hydroxysuccinimide ester, hydroxysulfosuccinimide ester, imido ester, azidonitrophenol, azide, 3-(2-pyridyl dithio)proprionamide, glyoxal or aldehyde group.

6. The dye of claim 1, wherein said dye is linked to a target molecule.

7. The dye of claim 6, wherein said target molecule comprises a nucleoside, nucleotide, oligonucleotide, polynucleotide, peptide nucleic acid, protein, peptide, enzyme, antigen, antibody, hormone, hormone receptor, cellular receptor, lymphokine, cytokine, hapten, lectin, avidin, strepavidin, digoxygenin, carbohydrate, oligosaccharide, polysaccharide, lipid, glycolipid, viral particle, viral component, bacterial cell, bacterial component, eucaryotic cell, eukaryotic cell component, natural drug, synthetic drug, glass particle, glass surface, plastic particle, plastic surface, siliceous particle, siliceous surface, organic molecule, dye or a derivative thereof.

8. The dye of claim 7, wherein said nucleoside, nucleotide, oligonucleotide, or polynucleotide comprises one or more ribonucleoside moieties, ribonucleotide moieties, deoxyribonucleoside moieties, deoxyribonucleotide moieties, modified ribonucleosides, modified ribonucleotides, modified deoxyribonucleosides, modified deoxyribonucleotides, ribonucleotide analogues, deoxyribonucleotide analogues or any combination thereof.

9. The dye of claim 6, wherein said dye is linked to said target molecule through a linker arm.

10. The dye of claim 9, wherein said linker arm is attached to said target molecule through a bond which comprises a covalent bond, a non-covalent bond, a polar bond or a coordinate bond.

11. The oligonucleotide or polynucleotide of claim 8, wherein said oligonucleotide or polynucleotide comprises two or more dye molecules attached to separate nucleotides of said oligonucleotide or polynucleotide.

12. The oligonucleotide or polynucleotide of claim 11, wherein said two or more dye molecules comprise the same dye molecules.

13. The oligonucleotide or polynucleotide of claim 11, wherein said two or more dye molecules comprise different dye molecules.

14. A composite dye comprising the dye of claim 7, said dye being joined or attached to a second dye molecule.

15. The dye of claim 1, wherein said dye is attached to a target specific moiety.

16. The dye of claim 15, wherein said target specific moiety comprises a protein or a nucleic acid.

17. The dye of claim 16, wherein said protein comprises an antibody or a fragment thereof.

18. The dye of claim 16, wherein said nucleic acid comprises unmodified nucleotides, modified nucleotides, nucleotide analogues or any combination thereof.

19. The dye of claim 1, wherein said alkyl or alkoxy groups independently comprise from 1-18 carbons.

20. The dye of claim 19, wherein said alkyl or alkoxy groups independently comprise from 1-6 carbons.

21. The dye of claim 3 wherein said electrophilic reactive group comprises an isocyanate, isothiocyanate, monochlorotriazine, dichlorotriazine, 4,6-dichloro-1,3,5-triazines, mono- or di-halogen substituted pyridine, mono- or di-halogen substituted diazine, maleimide, haloacetamide, aziridine, sulfonyl halide, acid halide, hydroxysuccinimide ester, hydroxysulfosuccinimide ester, imido ester, azidonitrophenol, azide, 3-(2-pyridyl dithio)proprionamide, glyoxal or aldehyde group.

22. A dye having the formula

23. A dye having the formula