Cellular labeling and quantification for nuclear magnetic resonance techniques

• US 8,263,043 B2
• Filed: 04/16/2007
• Issued: 09/11/2012
• Est. Priority Date: 04/14/2006
• Status: Active Grant

• Current Assignee: Carnegie Mellon University
• Original Assignee: Carnegie Mellon University
• Inventors: Ahrens, Eric T., Srinivas, Mangala
• Primary Examiner(s): Hartley, Michael G
• Assistant Examiner(s): SAMALA, JAGADISHWAR RAO
• Application Number:
  US11/787,521
• Publication Number:
  US 20070253910A1
• Time in Patent Office: 1,975 Days
• Field of Search: 424/9.3, 424 96- 98, 424/489, 514/450, 514/832
• US Class Current: 424/9.34
• CPC Class Codes: A61B 5/055 involving electronic [EMR] or nuclear [NMR] magnetic resonance, e.g. magnetic resonance imaging
• CPC Class Codes: A61B 5/411 Detecting or monitoring allergy or intolerance reactions to an allergenic agent or substance
• CPC Class Codes: A61B 5/413 Monitoring transplanted tissue or organ, e.g. for possible rejection reactions after a transplant
• CPC Class Codes: A61B 5/415 the glands, e.g. tonsils, adenoids or thymus
• CPC Class Codes: A61B 5/416 the spleen
• CPC Class Codes: A61B 5/417 the bone marrow
• CPC Class Codes: A61B 5/418 lymph vessels, ducts or nodes
• CPC Class Codes: A61B 5/4519 Muscles A61B5/389, A61B5/224 take precedence
• CPC Class Codes: A61B 5/7285 for synchronising or triggering a physiological measurement or image acquisition with a physiological event or waveform, e.g. an ECG signal A61B6/5205, A61B6/541, A61B8/5207, A61B8/543 take precedence
• CPC Class Codes: A61K 49/1896 not provided for elsewhere, e.g. cells, viruses, ghosts, red blood cells, virus capsides
• CPC Class Codes: B82Y 5/00 Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery

Abstract

The disclosure provides, in part, a method for quantifying cell numbers in vivo. The disclosure comprises a method of quantifying labeled cells by nuclear magnetic resonance techniques and a computer method for the same.

Claims

1. A method for quantifying cell number in vivo, the method comprising:

a. administering to a subject cells that are labeled with a fluorocarbon imaging reagent, wherein at least a portion of the fluorocarbon imaging reagent was internalized into the cell;

b. examining at least a portion of the subject by ¹⁹F magnetic resonance imaging (MRI), thereby detecting the cells that are labeled with the fluorocarbon imaging reagent in the subject; and

c. quantifying the number of labeled cells in a region of interest (ROI).

2. The method of claim 1, wherein the fluorocarbon imaging reagent is a perfluoropolyether.

3. The method of claim 1, wherein the fluorocarbon imaging reagent is a perfluoro-crown ether.

4. The method of claim 1, wherein the imaging reagent is a perfluoro-15-crown-5-ether.

5. The method of claim 1, wherein the fluorocarbon is a perfluorinated polyether having an average formula:

XO(Y—

O)_nZ

wherein Y is selected from the group comprising;

6. The method of claim 1, wherein the fluorocarbon imaging reagent is selected from the group comprising:

a linear perfluoropolyether, a cyclic perfluoropolyether and a mixture thereof.

7. The method of claim 1, wherein prior to administration to the subject, the cells are contacted with a composition comprising the fluorocarbon imaging reagent, whereby the cells are labeled with the fluorocarbon imaging reagent.

8. The method of claim 7, wherein the composition comprising the fluorocarbon imaging reagent further comprises an uptake enhancing reagent.

9. The method of claim 8, wherein the uptake enhancing reagent comprises a compound selected from the group consisting of:

a. cationic lipid; and

b. cationic polypeptide.

10. The method of claim 9, wherein the cationic peptide is a protamine.

11. The method of claim 7, wherein the composition comprising the fluorocarbon imaging reagent further comprises:

a. a surfactant; and

b. a cationic lipid.

12. The method of claim 7, wherein the composition comprising the fluorocarbon imaging reagent further comprises Pluronic L-35.

13. The method of claim 7, wherein the fluorocarbon imaging reagent is formulated as an emulsion.

14. The method of claim 7, wherein the emulsion comprises particles having a mean diameter of between 30 and 500 nm.

15. The method of claim 1, wherein prior to administration to the subject, the cells are labeled with perfluorocarbon emulsion particles using electroporation.

16. The method of claim 1, wherein the cell is a mammalian cell.

17. The method of claim 1, wherein the cell is a cell of the immune system.

18. The method of claim 1, wherein the cell is a T cell.

19. The method of claim 1, wherein the cell is a dendritic cell.

20. The method of claim 1, wherein the cell is a stem cell.

21. The method of claim 1, further comprising collecting a ¹H data set.

22. The method of claim 1, further comprising generating and comparing a ¹⁹F image and a ¹H image.

23. The method of claim 1, wherein the cell is administered to the subject as part of a cellular therapeutic regimen.

24. The method of claim 1, wherein quantifying comprises using a calibrated ¹⁹F signal in the ROI.

25. The method of claim 24, wherein the cellular dose of fluorocarbon imaging reagent is calculated prior to administration of cells to the subject.

26. The method of claim 1, wherein examining comprises detecting a pre-calibrated ¹⁹F signal from which one can deduce a relationship between the ¹⁹F signal in the ROI and the representative number of ¹⁹F molecules or cell quantity.

27. The method of claim 1, wherein examining comprises detecting a contemporaneous signal in the ROI from which one can deduce a relationship between the signal and the representative number of ¹⁹F molecules or cell quantity.

28. The method of claim 1, wherein examining comprises detecting a post-calibrated ¹⁹F signal in the ROI from which one can deduce a relationship between the signal and the representative number of ¹⁹F molecules or cell quantity.

29. The method of claim 1, wherein quantifying is done by comparison to a calibrated external ¹⁹F reference during the ¹⁹F MRI scan of the subject.

30. The method of claim 29, wherein the reference is a cell free reference.

31. The method of claim 1, wherein quantifying comprises calculating ratios of the intensity of ¹⁹F signal and the volume of labeled cells in a ROI compared to a reference.

32. The method of claim 1, wherein quantifying the number of labeled cells comprises using at least one of the parameters from the group consisting of:

(i) the cellular dose of labeling agent (i.e., F_c) measured in vitro;

(ii) in vivo ¹⁹F MRI data set taken in the subject at one or more time points following labeled cell administration;

(iii) the voxel volume;

(iv) the in-plane voxel area (i.e., area of the image pixel);

(v) the MRI data set from the ¹⁹F reference standard;

(vi) the measured Johnson noise of the ¹⁹F MRI data in the subject material;

(vii) the measured signal-to-noise ratio (SNR) of one or more voxels of the ¹⁹F MRI data set in the subject material;

(viii) the measured SNR of one or more voxels of the ¹⁹F MRI data set from the reference standard;

(ix) the ¹⁹F NMR relaxation times (T1, T2, and T2*) of the subject material; and

(x) the ¹⁹F NMR relaxation times (T1, T2, and T2*) of the reference standard.

33. The method of claim 1, wherein quantifying the number of labeled cells comprises using the formula: