Detecting targets using mass tags and mass spectrometry

Patent metadata

Field	Value
Publication number	US-12092643-B2
Application number	US-202017021659-A
Country	US
Kind code	B2
Filing date	Sep 15, 2020
Priority date	Jul 2, 2010
Publication date	Sep 17, 2024
Grant date	Sep 17, 2024

Abstract

Particular disclosed embodiments disclosed herein concern using a one or more various mass tags, which can be specifically deposited at targets through direct or indirect enzymatic-catalyzed transformation, to provide a method for identifying targets in tissue samples. The mass tags may be labeled with stable isotopes to produce mass tags having the same chemical structure but different masses. Mass codes produced by ionizing the mass tags are detected and/or quantified using mass spectrometry. The method can be used for multiplexed detection of multiple targets in a particular sample. In some embodiments, a map divided into sections representing sections of the tissue sample may be prepared, with the map sections including data corresponding to quantification data wherein the size of a mass peak is determined and correlated with the amount of a target for the corresponding tissue sample section.

First claim

The invention claimed is:

1. A conjugate comprising having a formula:

wherein
the ‘Mass Tag Precursor’ is a nitrophenyl moiety or a heteroaryl moiety;
R 16 is selected from ether, hydroxyl, —N(H)(R 11 ) or —NH 2 ;
R 11 is selected from an aliphatic group having between 1 and 10 carbon atoms, a heteroaliphatic group having between 1 and 10 carbon atoms, an aryl group, a heteroaryl group;
R 17 is selected from an aliphatic group having between 1 and 10 carbon atoms, a heteroaliphatic group having between 1 and 10 carbon atoms, an aryl group having 5 or 6 carbon atoms, or a heteroaryl group having 4 or 5 carbon atoms;
n is an integer ranging from 1 to 20;
Y is selected from oxygen, sulfur, or —N(R 11 )—, or —NH—;
the optional linker is selected from an aliphatic group, a heteroaliphatic group, or heterobifunctional linkers; and
the optional carrier is selected from a polymer, a biomolecule, a liposome, a micelle, or a nanoparticle;
wherein the nitrophenyl moiety has the formula:

where R 22 , R 23 , and R 24 are independently selected from an aliphatic group, a heteroaliphatic group, an aryl group, a heteroaryl group, a carbonyl group, a halogen, a hydrogen, a hydroxyl group, an isothiocyanate group, an isocyanate group, a nitrile group, a nitro group, a thiol, or any combination thereof, at least one of R 22 , R 23 , and R 24 is nitro;
Y is selected from oxygen, sulfur, or -(nitrogen —R 11 )—;
R 7 is selected from aliphatic or heteroaliphatic;
R 8 is selected from an aliphatic group, a heteroaliphatic group, a heterobifunctional linker, a charged moiety, a chromophore, or any combination thereof; and
R 11 is an aliphatic group, a heteroaliphatic group, an aryl group, or a heteroaryl group; and

wherein the heteroaryl moiety has a formula:

where R 25 , R 26 , and R 28 are independently selected from carbon or nitrogen, where at least one of R 25 , R 26 , R 27 , and R 28 is nitrogen;
each Y is independently selected from oxygen, sulfur, or -(nitrogen —R 11 )—;
R 8 comprises an aliphatic group, a heteroaliphatic group, a heterobifunctional linker, a charged moiety, a chromophore, or any combination thereof; and
R 11 is an aliphatic group, a heteroaliphatic group, an aryl group, or a heteroaryl group.

2. The conjugate of claim 1 , wherein the linker has the following general formula

where each X is independently selected from CH 2 , oxygen, sulfur, or -(nitrogen —R 11 )—;
R 18 is carbonyl or sulfoxyl; n is an integer ranging from 1 to 20; and p is 0 or 1.

3. The conjugate of claim 1 , wherein the nitrophenyl moiety has any of the formulas:

4. The conjugate of claim 1 , wherein the linker has the following general formula

where each X is independently selected from CH 2 , oxygen, sulfur, or -(nitrogen —R 11 )—; R 18 is carbonyl or sulfoxyl; n is an integer ranging from 1 to 20; and p is 0 or 1.

5. The conjugate of claim 1 , wherein the heteroaryl moiety has a formula:

6. The conjugate of claim 1 , wherein

 is tyramide or a tyramide derivative.

7. The conjugate of claim 1 , wherein the ‘Mass Tag Precursor’ includes a nitrophenyl moiety and wherein the charged moiety is a charged amino acid moiety or a tetra-alkyl ammonium species.

8. The conjugate of claim 7 , wherein the charged amino acid moiety is one of arginine, lysine, or histidine.

9. The conjugate of claim 1 , wherein the ‘Mass Tag Precursor’ includes a nitrophenyl moiety and wherein R 23 is nitro.

10. The conjugate of claim 9 , wherein R 23 is located ortho to the group R 7 .

11. The conjugate of claim 1 , wherein the ‘Mass Tag Precursor’ includes a nitrophenyl moiety and wherein R 7 is —[CH(Me)]—.

12. The conjugate of claim 1 , wherein the ‘Mass Tag Precursor’ includes a nitrophenyl moiety and wherein R 8 is substituent having a m/z value ranging from about 100 to about 5000.

13. The conjugate of claim 12 , wherein the m/z value ranges from about 500 to about 2000.

14. The conjugate of claim 1 , the ‘Mass Tag Precursor’ includes a heteroaryl moiety and wherein Y is sulfur.

15. The conjugate of claim 1 , the ‘Mass Tag Precursor’ includes a heteroaryl moiety and wherein each of R 25 , R 26 , R 27 , and R 28 are nitrogen.

16. The conjugate of claim 1 , the ‘Mass Tag Precursor’ includes a heteroaryl moiety.

17. The conjugate of claim 1 , wherein the ‘Mass Tag Precursor’ includes a nitrophenyl moiety.

18. A conjugate comprising having a formula:

wherein
the ‘Mass Tag Precursor’ is a nitrophenyl moiety or a heteroaryl moiety;
R 16 is hydroxyl;
R 11 is selected from an aliphatic group, a heteroaliphatic group, an aryl group, a heteroaryl group;
R 17 is selected from an aliphatic group, a heteroaliphatic group; n is an integer ranging from 1 to 20;
Y is selected from oxygen, sulfur, or —N(R 11 )—, or —NH—;
the optional linker is selected from an aliphatic group, a heteroaliphatic group, or heterobifunctional linkers; and
the optional carrier is selected from a polymer, a biomolecule, a liposome, a micelle, or a nanoparticle;
wherein the nitrophenyl moiety has the formula:

where R 22 , R 23 , and R 24 are independently selected from an aliphatic group, a heteroaliphatic group, an aryl group, a heteroaryl group, a carbonyl group, a halogen, a hydrogen, a hydroxyl group, an isothiocyanate group, an isocyanate group, a nitrile group, a nitro group, a thiol, or any combination thereof, at least one of R 22 , R 23 , and R 24 is nitro;
Y is selected from oxygen, sulfur, or -(nitrogen —R 11 )—;
R 7 is selected from aliphatic or heteroaliphatic;
R 8 is selected from an aliphatic group, a heteroaliphatic group, a heterobifunctional linker, a charged moiety, a chromophore, or any combination thereof; and
R 11 is an aliphatic group, a heteroaliphatic group, an aryl group, or a heteroaryl group; and

wherein the heteroaryl moiety has a formula:

where R 25 , R 26 , R 27 , and R 28 are independently selected from carbon or nitrogen, where at least one of R 25 , R 26 , R 27 , and R 28 is nitrogen;
each Y is independently selected from oxygen, sulfur, or -(nitrogen —R 11 )—;
R 8 comprises an aliphatic group, a

Assignees

• Ventana Med Syst Inc
• Ventana Medic Systems Inc

Inventors

• Bieniarz Christopher
• Froehlich Jan
• Kelly Brian Daniel
• Hong Rui
• Lefever Mark
• Miller Phillip
• Wang Hong

Classifications

• G01N33/6848 Primary

  Methods of protein analysis involving mass spectrometry · CPC title