Biocompatible fluorescent metal oxide nanoparticles

• US 20080226562A1
• Filed: 12/22/2006
• Published: 09/18/2008
• Est. Priority Date: 12/22/2005
• Status: Active Grant

• Current Assignee: VisEn Medical, Inc. (Revvity, Inc.)
• Original Assignee: VisEn Medical, Inc. (Revvity, Inc.)
• Inventors: Groves, Kevin, Rajopadhye, Milind
• Granted Patent: US 9,913,917 B2
• Time in Patent Office: Days
• US Class Current: 424/9.6
• CPC Class Codes: A61K 49/0002 General or multifunctional contrast agents, e.g. chelated agents
• CPC Class Codes: A61K 49/0032 Methine dyes, e.g. cyanine dyes
• CPC Class Codes: A61K 49/0056 Peptides, proteins, polyamino acids
• CPC Class Codes: A61K 49/0093 Nanoparticle, nanocapsule, nanobubble, nanosphere, nanobead, i.e. having a size or diameter smaller than 1 micrometer, e.g. polymeric nanoparticle
• CPC Class Codes: A61K 49/1833 having a (super)(para)magnetic core coated or functionalised with a small organic molecule oligomeric, polymeric, dendrimeric A61K49/1851
• CPC Class Codes: A61K 49/186 the organic macromolecular compound being polyethyleneglycol [PEG]
• CPC Class Codes: A61K 49/1866 the nanoparticle having a (super)(para)magnetic core coated or functionalised with a peptide, e.g. protein, polyamino acid
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: B82Y 5/00 Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery

Abstract

The invention relates to highly fluorescent metal oxide nanoparticles to which biomolecules and other compounds can be chemically linked to form biocompatible, stable optical imaging agents for in vitro and in vivo applications. The fluorescent metal oxide nanoparticles may also be used for magnetic resonance imaging (MRI), thus providing a multi modality imaging agent.

Claims

1. A fluorescent metal oxide nanoparticle comprising:

(a) a core comprising metal oxide;

(b) a polymer coating chemically linked to the core; and

(c) a plurality of fluorochromes chemically linked to the coating, wherein the nanoparticle has a molar extinction coefficient of at least 1,000,000 M<sup>−

1</sup>cm<sup>−

1</sup>, and emits fluorescent light upon illumination with light absorbable by the plurality of fluorochromes.

2. The nanoparticle of claim 1, wherein the molar extinction coefficient is from about 1,000,000 M⁻

1cm<sup>−

1</sup> to about 30,000,000 M<sup>−

1</sup>cm<sup>−

1</sup>, is from about 1,250,000 M<sup>−

1</sup>cm<sup>−

1</sup> to about 20,000,000 M<sup>−

1</sup>cm<sup>−

1</sup>, is from about 1,500,000 M<sup>−

1</sup>cm<sup>−

1</sup> to about 10,000,000 M<sup>−

1</sup>cm<sup>−

1</sup>, or is from about 1,750,000 M<sup>−

1</sup>cm<sup>−

1</sup> to about 7,500,000 M<sup>−

1</sup>cm<sup>−

1</sup>.

3. The nanoparticle of claim 1, wherein the molar extinction coefficient is from about 2,000,000 M⁻

1cm<sup>−

1</sup> to about 5,000,000 M<sup>−

1</sup>cm<sup>−

1</sup>.

4. The nanoparticle of claim 3, wherein the nanoparticle comprises from about 4 to about 100 fluorochromes chemically linked to the coating.

5. The nanoparticle of claim 4, wherein the nanoparticle comprises from about 20 to about 60 fluorochromes chemically linked to the coating.

25. The nanoparticle of claim 1, wherein a fluorochrome is linked to the polymer via a peptide linker.

26. The nanoparticle of claims 25, wherein the peptide comprises a protease cleavage site.

27. The nanoparticle of claim 1, wherein the metal oxide core is magnetic.

28. The nanoparticle of claim 1, wherein the metal oxide core is paramagnetic.

29. The nanoparticle of claim 28, wherein the metal oxide core is superparamagnetic.

30. The nanoparticle of claim 1, further comprising a biological modifier.

31. The nanoparticle of claim 30, wherein the biological modifier is polyethylene glycol or a derivative thereof.

32. The nanoparticle of claim 1, further comprising a biomolecule chemically linked to the polymer.

33. The nanoparticle of claim 32, wherein the biomolecule is covalently linked to the polymer through an amine moiety.

34. The nanoparticle of claim 32, wherein the biomolecule is a peptide, an antibody or an antigen binding fragment thereof, or a small molecule.

35. The nanoparticle of claim 32, wherein the biomolecule is a bivalent or multivalent binding molecule.

36. The nanoparticle of claim 32, wherein the biomolecule is an N,N-dipicolylamine moiety.

37. The nanoparticle of claim 32, wherein the biomolecule is protamine.

38. The nanoparticle of claim 32, wherein, when the biomolecule is a peptide, fluorescence of the nanoparticle increases once the peptide has been cleaved with an enzyme.

39. The nanoparticle of claim 32, comprising from about 1 to about 100 biomolecules chemically linked to the nanoparticle.

40. The nanoparticle of claim 32, further comprising a metal chelator chemically linked to the polymer.

41. The nanoparticle of claim 32, further comprising a radiolabel chemically linked to the polymer.

42. The nanoparticle of claim 32, further comprising a photosensitizer or phototherapy reagent chemically linked to the polymer.

43. The nanoparticle of claim 42, wherein the photosensitizer is Photofrin.

44. The nanoparticle of claim 43, wherein the nanoparticle has an absorption maxima and an emission maxima in the range from about 400 nm to about 1,200 nm.

45. The nanoparticle of claims 44, wherein the nanoparticle has an absorption maxima and an emission maxima in the range from about 600 nm to about 900 nm.

46. The nanoparticle of claim 44, wherein the fluorochrome is a red or near infrared fluorochrome.

47. The nanoparticle of claim 46, wherein the fluorochrome is a carbocyanine fluorochrome.

48. The nanoparticle of claim 46, wherein the fluorochrome is Cy5, Cy5.5, Cy7, VivoTag680, VivoTag-S680 or VivoTag-S750.

49. The nanoparticle of claim 48, wherein the nanoparticle has a diameter from about 10 nm to about 100 nm.

50. The nanoparticle of claim 48, wherein the nanoparticle is biocompatible.

51. A method of in vivo optical imaging, the method comprising:

a) administering to a subject nanoparticles of any one of claims 1-50;

b) allowing the nanoparticles to distribute in the subject;

c) exposing the subject to (i) light of a wavelength absorbable by the fluorochrome of the nanoparticle and/or (ii) magnetic radiation; and

d) detecting an optical and/or magnetic signal emitted by the nanoparticle.

52. The method of claim 51, wherein the signal emitted by the nanoparticles is used to construct an image.

53. The method of claim 52, wherein the image is a tomographic image.

54. The met hod of claim 53, wherein the tomographic image is co-registered with an image obtained by magnetic resonance.

55. The method of claim 51, wherein steps (a)-(d) are repeated at predetermined intervals thereby to permit evaluation of the emitted signals of the nanoparticles in the subject over time.

56. The method of claim 51, further comprising the step of positioning a detection device to detect optical and/or magnetic signals emitted from the subject.

57. The method of claim 51, wherein the subject is an animal or a human.

58. The method of claim 51, wherein in step (a) two or more imaging probes whose signal properties are distinguishable from one another are administered to a subject, wherein at least one of the imaging probes is a nanoparticle.

59. The method of claim 51, wherein a nuclear imaging agent is administered to the subject.

60. The method of claim 59, wherein the nuclear imaging agent chemically linked to the nanoparticle.

61. The method of claim 59, wherein the nuclear imaging agent can be detected by CT, PET, SPECT or MR imaging.

62. The method of claim 51, wherein the illuminating and detecting steps are performed using an endoscope, catheter, tomographic system, hand-held optical imaging system, or an intraoperative microscope.

63. The method of claim 51, wherein the presence, absence, or level of emitted signal is indicative of a disease state.

64. The method of claim 51, wherein the method is used to detect and/or monitor a disease.

65. The method of claim 64, wherein the disease is selected from the group, consisting of bone disease, cancer, cardiovascular disease, environmental disease, dermatological disease, immunologic disease, inherited disease, infectious disease, inflammatory disease, metabolic disease, neurodegenerative disease, ophthalmic disease, and respiratory disease.

66. The method of claim 51, wherein, in step (a), cells labeled with the nanoparticle are administered to the subject.

67. The method of claim 66, wherein the signal emitted by the nanoparticle is used to monitor trafficking and localization of the cells.

68. An in vitro optical imaging method, the method comprising:

a) contacting a sample with the nanoparticles of any one of the claims 1-50;

b) allowing the nanoparticles to become activated by or to bind to a biological target;

c) optionally, removing unbound nanoparticles;

d) illuminating the sample with a light of a wavelength absorbable by the nanoparticles; and

e) detecting signal emitted from the nanoparticles thereby to determine whether the nanoparticles have been activated by or bound to the biological target.

69. The method of claim 68, wherein the sample is a biological sample.

6. A fluorescent metal oxide nanoparticle comprising:

(a) a core comprising metal oxide;

(b) a non-crosslinked polymer coating chemically linked to the core; and

(c) from about 10 to about 300 fluorochromes chemically linked to the coating.

7. The nanoparticle of claim 6, wherein the nanoparticle comprises from about 15 to about 100 fluorochromes per nanoparticle.

8. The nanoparticle of claim 7, wherein the nanoparticle comprises from about 20 to about 60 fluorochromes per particle.

9. A fluorescent metal oxide nanoparticle comprising:

(a) a core comprising metal oxide;

(b) a polymer coating chemically linked to the core; and

(c) a plurality of fluorochromes chemically linked to the polymer, wherein the nanoparticle, when illuminated with light of a wavelength absorbable by the plurality of fluorochromes chemically linked to the core retains at least 50% of the fluorescence obtained from substantially the same number of free fluorochromes when measured under the same conditions.

10. The nanoparticle of claim 9, wherein the nanoparticle retains from about 50% to about 100% of the fluorescence obtained from substantially the same number of free fluorochromes when measured under the same conditions.

11. The nanoparticle of claim 10, wherein the nanoparticle retains from about 75% to about 95% of the fluorescence obtained from substantially the same number of free fluorochromes when measured under the same conditions.

12. The nanoparticle of claim 10, wherein the nanoparticle comprises from about 100 to about 300 fluorochromes per particle.

13. The nanoparticle of claim 12, wherein the nanoparticle comprises from about 15 to about 100 fluorochromes per nanoparticle.

14. The nanoparticle of claim 13, wherein the nanoparticle comprises from about 20 to about 60 fluorochromes per particle.

15. The nanoparticle of claim 10, wherein the polymer is selected from the group consisting of a natural polymer, a synthetic polymer, and co-polymers thereof.

16. The nanoparticle of claim 10, wherein the polymer is polyacrylic acid, polyglutamic acid, polylactic acid, or polyethylene maleic acid.

17. The nanoparticle of claim 16, wherein the polymer is carboxymethyl-polyvinylalcohol.

18. The nanoparticle of claim 15, wherein the polymer comprises a plurality of carboxyl moieties.

19. The nanoparticle of claim 18, wherein the coating is chemically linked to the core through a subset of the plurality of the carboxyl moieties.

20. The nanoparticle of claim 15, wherein the polymer comprises a plurality of reactive amines.

21. The nanoparticle of claim 20, wherein the nanoparticle comprises from about 100 to about 300 reactive amines per particle.

22. The nanoparticle of claim 21, wherein the nanoparticle comprises from about 150 to about 250 reactive amines per particle.

23. The nanoparticle of claim 20, wherein a fluorochrome is linked to the polymer via an amine moiety.

24. The nanoparticle of claim 17, wherein the carboxymethyl-PVA is linked to the fluorochrome through a diamine moiety represented by:

NH₂—

(CH₂)_n—

NH₂, wherein n is an integer from 1 to 12.