18F labeling of proteins using sortases

Patent metadata

Field	Value
Publication number	US-11850216-B2
Application number	US-202016737620-A
Country	US
Kind code	B2
Filing date	Jan 8, 2020
Priority date	Nov 13, 2013
Publication date	Dec 26, 2023
Grant date	Dec 26, 2023

Abstract

The present invention, in some aspects, provides methods, reagents, compositions, and kits for the radiolabeling of proteins, for example, of proteins useful for positron emission tomography (PET) or single-photon emission computed tomography (SPECT) (e.g., for diagnostic and therapeutic applications), using sortase-mediated transpeptidation reactions. Some aspects of this invention provide methods for the conjugation of an agent, for example, a radioactive agent or molecule to diagnostic or therapeutic peptides or proteins. Compositions comprising sortagged, radiolabeled proteins as well as reagents for generating radiolabeled proteins are also provided. Kits comprising reagents useful for the generation of radiolabeled proteins are provided, as are precursor proteins that comprise a sortase recognition motif.

First claim

What is claimed is:

1. A radioactive protein of Formula (II):

wherein
represents a protein;
L 1 is a peptide linker comprising -LPXTGGGK-, -LPXTGGG-, -NPXTGGGK-, -NPXTGGG-, -LPXTAAA-, -NPXTAAA-, -LPXTGGGGG-, or -LPGAG-, wherein each instance of X is independently any amino acid; and
L 2 is a linker comprising optionally substituted heterocyclylene.

2. A pharmaceutical composition comprising the radioactive protein of claim 1 and a pharmaceutically acceptable carrier.

3. The radioactive protein of claim 1 , wherein is an antibody, a nuclear factor, a neuropeptide, a receptor protein, an enzyme, a structural protein, or a fragment thereof.

4. The radioactive protein of claim 1 , wherein L 1 is -LPXTGGG-.

5. The radioactive protein of claim 1 , wherein L 2 is of Formula (ii):

wherein
R t is hydrogen, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted aryl, optionally substituted heteroaryl, or optionally substituted heterocyclyl;
R s is a bond, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted arylene, optionally substituted heteroarylene, or optionally substituted heterocyclylene;
each instance of R c1 is hydrogen, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted aryl, optionally substituted heteroaryl, or optionally substituted heterocyclyl; or two R c1 are joined to form an optionally substituted carbocyclic or optionally substituted heterocyclic ring;
R c2 is a bond, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted arylene, optionally substituted heteroarylene, or optionally substituted heterocyclylene;
m is 0, 1, 2, 3, 4, 5, 6, 7, or 8, as valency permits;
a is a point of attachment to L 1 ; and
b is a point of attachment to 18 F.

6. The radioactive protein of claim 5 , wherein L 2 is of Formula (ii-a):

wherein n is an integer between 1 and 8, inclusive.

7. The radioactive protein of claim 6 , wherein R t is optionally substituted methyl, optionally substituted pyridine, or optionally substituted phenyl.

8. The radioactive protein of claim 1 , wherein L 2 is of Formula (iii):

wherein
R t is hydrogen, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted aryl, optionally substituted heteroaryl, or optionally substituted heterocyclyl;
R s is a bond, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted arylene, optionally substituted heteroarylene, or optionally substituted heterocyclylene;
each instance of R c1 is hydrogen, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted aryl, optionally substituted heteroaryl, or optionally substituted heterocyclyl;
R c2 is a bond, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted arylene, optionally substituted heteroarylene, or optionally substituted heterocyclylene;
m is 0, 1, 2, 3, 4, 5, 6, 7, or 8, as valency permits;
a is a point of attachment to L 1 ; and
b is a point of attachment to 18 F.

9. The radioactive protein of claim 8 , wherein L 2 is of Formula (iii-a):

10. The radioactive protein of claim 9 , wherein -L 2 - 18 F is of Formula (iii-b):

wherein
each instance of R p is independently hydrogen, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclyl, hydroxyl, or optionally substituted amino; provided at least one RP comprises 18 F; and
p is 1, 2, 3, 4, or 5.

11. The radioactive protein of claim 10 , wherein -L 2 - 18 F is of Formula (iii-b1):

wherein
L 3 is a bond, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted arylene, optionally substituted heteroarylene, or optionally substituted heterocyclylene;
R N1 is independently hydrogen, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted aryl, optionally substituted heteroaryl, or a nitrogen protecting group; and
each of R s1 , R s2 , R s3 , and R s4 is independently hydrogen, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted aryl, optionally substituted heteroaryl, or an oxygen protecting group.

12. The radioactive protein of claim 10 , wherein -L 2 - 18 F is of the formula:

13. The radioactive protein of claim 10 , wherein -L 2 - 18 F is of Formula (iii-b2):

wherein
L 3 is a bond, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted arylene, optionally substituted heteroarylene, or optionally substituted heterocyclylene;
R N1 is independently hydrogen, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclyl, or a nitrogen protecting group;
each instance of R q1 is independently hydrogen, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclyl, hydroxyl, or optionally substituted amino; and
q1 is 0, 1, 2, 3, or 4.

14. The radioactive protein of claim 10 , wherein -L 2 - 18 F is of the formula:

15. The radioactive protein of claim 10 , wherein -L 2 - 18 F is of Formula (iii-b3):

wherein
R N1 is independently hydrogen, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted aryl, optionally substituted heteroaryl, or a nitrogen protecting group; and
each instance of R q2 is independently hydrogen, optionally substituted aliphatic, optionally substituted heteroaliphatic, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclyl, hydroxyl, or optionally substituted amino; and
q2 is 0, 1, 2, 3, or 4.

16. The radioactive protein of claim 15 , wherein -L 2 - 18 F is of the formula:

17. The radioactive protein of claim 1 , wherein -L 2 - 18 F is of Formula (iii-c):

Assignees

• Whitehead Inst Biomedical Res
• Massachusetts Gen Hospital

Inventors

• Rashidian Mohammad
• Ploegh Hidde L
• Weissleder Ralph
• Keliher Edmund J

Classifications

• A61K51/1096 Primary

  radioimmunotoxins, i.e. conjugates being structurally as defined in A61K51/1093, and including a radioactive nucleus for use in radiotherapeutic applications · CPC title

• A61K51/08

  Peptides, e.g. proteins {, carriers being peptides, polyamino acids, proteins} · CPC title

• C07H5/02

  to halogen · CPC title

• C07K16/2845

  against integrin beta2-subunit-containing molecules, e.g. CD11, CD18 · CPC title

• C12N9/52 Primary

  derived from bacteria {or Archaea} · CPC title