Reductive release probes containing a chemoselectively cleavable α-azidoether linker and methods of use thereof

• US 9,422,232 B2
• Filed: 07/07/2010
• Issued: 08/23/2016
• Est. Priority Date: 07/09/2009
• Status: Expired due to Fees

• Current Assignee: Board of Trustees of the Leland Stanford Junior University (Stanford University)
• Original Assignee: Board of Trustees of the Leland Stanford Junior University (Stanford University)
• Inventors: Franzini, Raphael M., Kool, Eric Todd
• Primary Examiner(s): Hartley, Michael G
• Assistant Examiner(s): SCHLIENTZ, LEAH H
• Application Number:
  US13/383,135
• Publication Number:
  US 20120178086A1
• Time in Patent Office: 2,239 Days
• US Class Current: 1/1
• CPC Class Codes: A61K 49/0017 Fluorescence in vivo
• CPC Class Codes: C07C 247/04 being saturated
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: C12Q 2563/107 fluorescence
• CPC Class Codes: G01N 33/532 Production of labelled immunochemicals

Abstract

Probes comprising one or more selectively cleavable α-azidoether moieties are provided; and linkers comprising the one or more selectively cleavable α-azidoether moieties. The α-azidoether moiety will undergo a Staudinger reaction with a suitable reducing agent, resulting in cleavage. The probes find use in a variety of detection assays, e.g. specific polynucleotide binding assays, polypeptide binding assays, etc. The cleavable linkers are suitable for synthetic reactions, e.g. to prepare probes of the invention; in the synthesis of cleavable peptide conjugates; and the like.

Claims

1. A selectively cleavable probe of structure (I):

2. The selectively cleavable probe of claim 1, wherein the compound is of structure (III):

6. The selectively cleavable probe of claim 1, wherein the detection moiety is selected from a fluorophore and a quencher.

7. The selectively cleavable probe of claim 1, wherein the specific binding moiety is selected from an antibody or fragment thereof, a polypeptide and a polynucleotide.

8. The selectively cleavable probe of claim 7, wherein the specific binding moiety is a polynucleotide of at least 6 bases in length and not more than about 100 bases in length.

9. The selectively cleavable probe of claim 8, wherein the detection moiety is a quencher, and said polynucleotide further comprises at least one fluorophore quenched by the detection moiety.

10. The selectively cleavable probe of claim 9, wherein the quencher is a dabsyl quencher.

3. A selectively cleavable probe of one of structures (VIIIa) and (VIIIb):

4. The selectively cleavable probe of claim 3, wherein the probe is of structure (IX):

5. The selectively cleavable probe of claim 4, wherein:

M³ comprises a fluorophore that is quenched by Q⁷.

11. The selectively cleavable probe of claim 3, wherein T¹⁴ and T¹⁵ are independently selected from one of the following structures, or a substituted version thereof:

12. The selectively cleavable probe of claim 3, wherein Q⁶ is selected from a fluorophore and a quencher.

13. The selectively cleavable probe of claim 3, wherein M² is selected from an antibody or fragment thereof, a polypeptide and a polynucleotide.

14. The selectively cleavable probe of claim 13, wherein M¹ is a polynucleotide of at least 6 bases in length and not more than about 100 bases in length.

15. The selectively cleavable probe of claim 14, wherein Q⁶ is a quencher, and said polynucleotide further comprises at least one fluorophore quenched by Q⁶.

16. The selectively cleavable probe of claim 15, wherein Q⁶ is a dabsyl quencher.