MRI contrast agents and high-throughput screening by MRI

• US 8,568,690 B2
• Filed: 07/29/2008
• Issued: 10/29/2013
• Est. Priority Date: 07/31/2007
• Status: Active Grant

• Current Assignee: Board of Trustees of The University of Illinois
• Original Assignee: Board of Trustees of The University of Illinois
• Inventors: Lu, Yi, Yigit, Mehmet Veysel, Mazumdar, Debapriya
• Primary Examiner(s): Hartley, Michael G
• Assistant Examiner(s): PERREIRA, MELISSA JEAN
• Application Number:
  US12/182,018
• Publication Number:
  US 20090098550A1
• Time in Patent Office: 1,918 Days
• Field of Search: 424/9.32
• US Class Current: 424/9.1
• CPC Class Codes: A61K 49/1845 the small organic molecule being a carbohydrate (monosaccharides, discacharides)
• CPC Class Codes: C12N 15/111 General methods applicable to biologically active non-coding nucleic acids
• CPC Class Codes: C12N 15/115 Aptamers, i.e. nucleic acids binding a target molecule specifically and with high affinity without hybridising therewith ; Nucleic acids binding to non-nucleic acids, e.g. aptamers
• CPC Class Codes: C12N 2310/16 Aptamers
• CPC Class Codes: C12N 2310/3517 Marker; Tag
• CPC Class Codes: C12N 2320/10 in screening processes
• CPC Class Codes: C12Q 1/00 Measuring or testing processes involving enzymes, nucleic acids or microorganisms measuring or testing apparatus with condition measuring or sensing means, e.g. colony counters, C12M1/34; Compositions therefor; Processes of preparing such compositions

Abstract

The present invention provides an MRI contrast agent, comprising: MRI contrast agent particles, and oligonucleotides, attached to the particles.

Claims

1. An MRI contrast agent, comprising:

(i) MRI contrast agent particles that are biocompatible and biodegradable in vivo,

(ii) oligonucleotides, attached to the particles, wherein the oligonucleotides have one end attached to the particles and a second free end; and

(iii) an aptamer comprising an oligonucleotide sequence that is complementary to the free end of two oligonucleotides attached to different particles, wherein the aptamer is hybridized to the free end of the two oligonucleotides thereby forming a bridge between the particles, wherein the particles, the oligonucleotides, and the aptamer, together form aggregates in absence of the effector and wherein the aptamer, specific for an effector, undergoes a conformational change when bound to the effector, causing the particles to disaggregate.

2. The MRI contrast agent of claim 1, wherein the MRI contrast agent particles comprise gadolinium, manganese, iron, superparamagnetic iron oxide (SPIO), oral SPIO, ultrasmall superparamagnetic iron oxide (USPIO), paramagnetic fullerene pipes, fullerene nanoparticles, nanotubes, gadonanotubes, quantum dots coated or doped with a paramagnetic material, paramagnetic liposomes, a paramagnetic material or perfluorocarbon particles.

3. The MRI contrast agent of claim 1, wherein the effector is adenosine or a metal ion.

6. A method of forming an MRI image of a sample, comprising:

mixing the sample with the MRI contrast agent of claim 1; and

imaging the sample by MRI.

7. An MRI sensor system for screening a molecule against a test enzyme, comprising:

the MRI contrast agent of claim 1, and

a test enzyme,

wherein the test enzyme reacts with a corresponding substrate to form a product, and wherein the substrate binds to the aptamer but the product does not or wherein the substrate does not bind to the aptamer but the product does.

11. The MRI sensor system of claim 7, wherein the MRI contrast agent particles comprise a paramagnetic material.

12. The MRI sensor system of claim 7, wherein the MRI contrast agent particles comprise at least one member selected from the group consisting of gadolinium, manganese, iron, SPIO, oral SPIO, USPIO, paramagnetic fullerene pipes, fullerene nanoparticles, nanotubes, gadonanotubes, quantum dots coated or doped with a paramagnetic material, paramagnetic liposomes and perfluorocarbon particles.

14. A method of producing an MRI image in vivo, comprising:

administering the MRI contrast agent of claim 1 to a subject; and

imaging the subject by MRI.

18. The MRI contrast agent of claim 1, wherein the effector is a cancer antigen, growth factor, nucleic acid binding protein, anthrax, small pox, a pollutant, cocaine, or human immuno-deficiency virus (HIV).

21. The MRI contrast agent of claim 1, wherein the effector is prostate specific membrane antigen (PSMA), tenasin-C, basic fibroplastic growth factor, platelet derived growth factor (PDGF), nuclear factor kB, transcription factor E2F, or gonadotropin-releasing hormone.

26. The MRI contrast agent of claim 1, wherein the MRI contrast agent particles comprise a paramagnetic material.

29. The MRI contrast agent of claim 1, wherein the effector is thrombin.

4. An MRI contrast agent, comprising:

MRI contrast agent particles that are biocompatible and biodegradable in vivo;

oligonucleotides, attached to the particles, wherein the oligonucleotides have one end attached to the particles and a second free end; and

a nucleic acid enzyme specific for an effector, comprising

(a) an oligonucleotide substrate strand comprising an oligonucleotide sequence that is complementary to the free end of two oligonucleotides attached to different particles, wherein the oligonucleotide substrate strand is hybridized to the free end of the two oligonucleotides thereby forming a bridge between the particles, and

(b) an oligonucleotide enzyme strand which is hybridized to the oligonucleotide substrate strand,

wherein the particles, the oligonucleotides, the oligonucleotide substrate strand and the oligonucleotide enzyme strand, together form aggregates in absence of the effector; and

wherein the oligonucleotide enzyme strand cleaves the oligonucleotide substrate strand in the presence of the effector, causing the particles to disaggregate.

5. The MRI contrast agent of claim 4, wherein the oligonucleotide substrate strand comprises:

first and second portions, hybridized to the free end of the two oligonucleotides attached to the different particles, and

wherein the first and second portions are cleaved by the oligonucleotide enzyme strand in the presence of the effector.

8. An MRI sensor system for screening a molecule against a test enzyme, comprising:

the MRI contrast agent of claim 4, and

a test enzyme that catalyzes conversion to the effector,

wherein the bridge is cleaved in the presence of the effector.

9. An MRI sensor system for screening a molecule against a test enzyme, comprising:

the MRI contrast agent of claim 4, and

a test enzyme that catalyzes conversion of a corresponding substrate to a product, wherein the corresponding substrate is the effector,

wherein the bridge is cleaved in the presence of the corresponding substrate.

10. An MRI sensor system for screening a molecule against a test enzyme, comprising:

the MRI contrast agent of claim 5

an assay enzyme, and

a test enzyme,

wherein the first and second portions are ligated by the assay enzyme in the presence of an effector,

the test enzyme reacts with a corresponding substrate to form a product, and the corresponding substrate or the product is the effector.

13. The MRI sensor system of claim 8, wherein the MRI contrast agent particles comprise a paramagnetic material.

15. A method of producing an MRI image in vivo, comprising:

administering the MRI contrast agent of claim 4 to a subject; and

imaging the subject by MRI.

19. The MRI contrast agent of claim 4, wherein the effector is a cancer antigen, growth factor, nucleic acid binding protein, anthrax, small pox, a pollutant, cocaine, adenosine, a metal ion, or human immuno-deficiency virus (HIV).

22. The MRI contrast agent of claim 4, wherein the effector is prostate specific membrane antigen (PSMA), tenasin-C, basic fibroplastic growth factor, platelet derived growth factor (PDGF), nuclear factor kB, transcription factor E2F, or gonadotropin-releasing hormone.

24. The MRI contrast agent of claim 4, wherein the MRI contrast agent particles comprise gadolinium, manganese, iron, superparamagnetic iron oxide (SPIO), oral SPIO, ultrasmall superparamagnetic iron oxide (USPIO), paramagnetic fullerene pipes, fullerene nanoparticles, nanotubes, gadonanotubes, quantum dots coated or doped with a paramagnetic material, paramagnetic liposomes, a paramagnetic material or perfluorocarbon particles.

27. The MRI contrast agent of claim 4, wherein the MRI contrast agent particles comprise a paramagnetic material.

16. An MRI contrast agent, comprising:

MRI contrast agent particles that are biocompatible and biodegradable in vivo;

oligonucleotides, attached to the particles, wherein the oligonucleotides have one end attached to the particles and a second free end; and

an aptazyme specific for an effector and a co-factor, comprising

(a) an oligonucleotide substrate strand comprising an oligonucleotide sequence that is complementary to the free end of two oligonucleotides attached to different particles wherein the oligonucleotide substrate strand is hybridized to the free end of the two oligonucleotides thereby forming a bridge between the particles, and

(b) an oligonucleotide enzyme strand hybridized to the oligonucleotide substrate strand, wherein the oligonucleotide enzyme strand comprises

(i) an aptamer portion comprising a binding site for the effector, and

(ii) an enzyme portion comprising a binding site for the co-factor when hybridized to the oligonucleotide substrate strand,

wherein the particles, the oligonucleotides, the oligonucleotide substrate strand and the oligonucleotide enzyme strand, together form aggregates in absence of the effector and the co-factor; and

wherein the oligonucleotide enzyme strand cleaves the oligonucleotide substrate strand in the presence of the effector and the co-factor, causing the particles to disaggregate.

17. A method of producing an MRI image in vivo, comprising:

administering the MRI contrast agent of claim 16 to a subject; and

imaging the subject by MRI.

20. The MRI contrast agent of claim 16, wherein the effector is a cancer antigen, growth factor, nucleic acid binding protein, anthrax, small pox, a pollutant, cocaine, adenosine, or human immuno-deficiency virus (HIV).

23. The MRI contrast agent of claim 16, wherein the effector is prostate specific membrane antigen (PSMA), tenasin-C, basic fibroplastic growth factor, platelet derived growth factor (PDGF), nuclear factor kB, transcription factor E2F, or gonadotropin-releasing hormone.

25. The MRI contrast agent of claim 16, wherein the MRI contrast agent particles comprise gadolinium, manganese, iron, superparamagnetic iron oxide (SPIO), oral SPIO, ultrasmall superparamagnetic iron oxide (USPIO), paramagnetic fullerene pipes, fullerene nanoparticles, nanotubes, gadonanotubes, quantum dots coated or doped with a paramagnetic material, paramagnetic liposomes, a paramagnetic material or perfluorocarbon particles.

28. The MRI contrast agent of claim 16, wherein the MRI contrast agent particles comprise a paramagnetic material.