Multisignal labeling reagents and processes and uses therefor

• US 9,156,986 B2
• Filed: 03/14/2011
• Issued: 10/13/2015
• Est. Priority Date: 04/03/2003
• Status: Active Grant

• Current Assignee: Enzo Life Sciences Incorporated (Enzo Biochem Incorporated)
• Original Assignee: Enzo Life Sciences Incorporated (Enzo Biochem Incorporated)
• Inventors: Coleman, Jack, Rabbani, Elazar, Stavrianopoulos, Jannis, Pande, Praveen
• Primary Examiner(s): Riley, Jezia
• Application Number:
  US13/065,101
• Publication Number:
  US 20110318788A1
• Time in Patent Office: 1,674 Days
• Field of Search: 546/26, 536/26.6, 435/91.1
• US Class Current: 1/1
• CPC Class Codes: C07D 219/08 Nitrogen atoms
• CPC Class Codes: C07D 405/14 containing three or more hetero rings
• CPC Class Codes: C07D 491/22 in which the condensed system contains four or more hetero rings
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C07H 21/04 with deoxyribosyl as saccharide radical
• CPC Class Codes: C09B 11/00 Diaryl- or thriarylmethane dyes
• CPC Class Codes: C09B 11/08 Phthaleins; Phenolphthaleins; Fluorescein
• CPC Class Codes: C09B 11/24 Phthaleins containing amino groups ; Phthalanes; Fluoranes; Phthalides; Rhodamine dyes; Phthaleins having heterocyclic aryl rings; Lactone or lactame forms of triarylmethane dyes
• CPC Class Codes: C09B 57/02 Coumarine dyes
• CPC Class Codes: C12Q 1/68 involving nucleic acids
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: G01N 33/533 with fluorescent label
• CPC Class Codes: G01N 33/582 with fluorescent label

Abstract

Provided are compounds comprising two DNA supramolecular binding molecules covalently joined by a linker group. Also provided are multisignal labeling reagents comprising (i) an oligomer of nucleotides or nucleotide analogs; (ii) a DNA supramolecular binding molecule noncovalently bound to the oligomer; and (iii) a first reactive group or a first partner of a first binding pair covalently bound to the oligomer. Additionally provided are methods of producing multisignal labeling reagents.

Claims

1. A compound comprising two DNA supramolecular binding molecules covalently joined by a linker group, the compound further comprising a detectable label bound thereto selected from a fluorescent compound, a phosphorescent compound, a chemiluminescent compound, a chelating compound, a magnetic compound and an energy transfer compound, wherein the detectable label is not either of the two DNA supramolecular binding molecules, and wherein said supramolecular binding molecules are each independently selected from a minor groove binder, and a major groove binder.

2. The compound of claim 1, wherein at least one of the DNA supramolecular binding molecules is a minor groove binder.

3. The compound of claim 2, wherein the minor groove binder is a diacrylamidine, or a bis-benzimidazole.

4. The compound of claim 2, wherein the minor groove binder is DAPI, berenil, pentamidine, distamycin A, or Hoechst 33258.

5. The compound of claim 1, wherein at least one of the DNA supramolecular binding molecules is a major groove binder.

6. The compound of claim 5, wherein the major groove binder is selected from a zinc finger, a leucine zipper and a helix-turn-helix motif.

10. The compound of claim 1 or claim 7, wherein the linker is flexible.

11. The compound of claim 1 or claim 7, wherein the linker is an unsubstituted C₁-C₂₀ straight-chain, branched or cyclic alkyl, alkenyl or alkynyl group, a substituted C₁-C₂₀ straight-chain, branched or cyclic alkyl, alkenyl or alkynyl group wherein one or more C, CH or CH₂ groups are substituted with an O atom, N atom, S atom, NH group, CO group or OCO group, or an unsubstituted or substituted aromatic group.

12. The compound of claim 1 or claim 7, wherein the linker is —

(CH₂)_1-10—

NH—

(CH₂)_1-10—

.

13. The compound of claim 1 or claim 7, wherein the linker is —

(CH₂)_1-5—

NH—

(CH₂)_1-5—

.

14. The compound of claim 1 or claim 7, wherein the linker is —

(CH₂)₃—

NH—

(CH₂)₄—

.

15. The compound of claim 1 or claim 7 wherein the detectable label is a fluorophore.

16. The compound of claim 15, wherein the fluorophore is selected from a symmetric or asymmetric cyanine, a merocyanine, a styryl moiety, an oxazine, a xanthene, a coumarin or an iminocoumarin.

17. The compound of claim 16, wherein the fluorophore is a xanthene.

18. The compound of claim 17, wherein the xanthene is a rhodamine or rhodamine derivative.

19. The compound of claim 17, wherein the xanthene is a rosamine or rosamine derivative.

20. The compound of claim 17, wherein the xanthene is a rhodol or rhodol derivative.

21. The compound of claim 16, wherein the fluorophore is a coumarin.

22. The compound of claim 21, wherein the coumarin is an iminocoumarin.

23. The compound of claim 1 or claim 7 wherein the detectable label is a chemiluminescent or phosphorescent moiety.

24. The compound of claim 1 or claim 7, wherein the detectable label is non-covalently bound to the compound via a binding pair.

25. The compound of claim 1 or claim 7, wherein the detectable label is covalently bound to the compound.

26. The compound of claim 25, wherein the detectable label is bound to the linker group.

27. The compound of claim 1, further comprising a third DNA supramolecular binding molecule that is an intercalator covalently joined by a linker group that is linked to the linker group that joins the two DNA supramolecular binding molecules.

28. The compound of claim 1:

7. A compound comprising two DNA supramolecular binding molecules covalently joined by a linker group, the compound further comprising a detectable label bound thereto selected from a fluorescent compound, a phosphorescent compound, a chemiluminescent compound, a chelating compound, a magnetic compound and an energy transfer compound, wherein the detectable label is not either of the two DNA supramolecular binding molecules, and wherein one of said DNA supramolecular binding molecules is selected from a minor groove binder, and a major groove binder, and the other DNA supramolecular binding molecule is an intercalator.

8. The compound of claim 7, wherein the intercalator is selected from an acridine intercalator, a coumarin intercalator, a psoralen intercalator, a phenanthridine intercalator, a quinoxaline intercalator, an anthracycline intercalator, and a metallo-intercalator.

9. The compound of claim 8, wherein the intercalator is selected from 9-aminoacridine, 7-hydroxy-4-methylcoumarin, 7-amino-4-methylcoumarin, 4-methyl-7-sulphato-methylcoumarin, 8[[[(diethylamino)methyl]propyl]oxy]psoralen, 5(N-piperadinyl)-8-methoxypsoralen, ethidium bromide, thiazole orange, 6-(-4′

-carboxyphenyl)-3,8-diamine-5-methyl phenanthridinium chloride, doxorubicin, daunomycin, [Pt(tpy)(SCH₂CH₂OH)]⁺, and [Rh(phi)(Me₂trien)]³⁺.