Molecular Analysis of Tumor Samples
This invention relates to methods for diagnosing cancer, e.g., cancer of epithelial origin, by detecting the presence of tumor cells in a sample, based (at least in some embodiments) on the quantification of levels of four biomarkers, MUC1, EGFR, EpCAM, and HER2. In some embodiments, the methods are performed using diagnostic magnetic resonance (DMR), e.g., with a portable relaxometer or MR imager.
1 . A method for diagnosing a tumor in a subject, the method comprising:
obtaining a sample from the subject;
detecting levels of MUC-1, HER2, EGFR, and EpCAM in the sample; and
comparing the levels of MUC-1, HER2, EGFR, and EpCAM in the sample to reference levels; and
diagnosing cancer in a subject who has levels of MUC-1, HER2, EGFR, and EpCAM above the reference levels.
2 . A method for detecting the presence of tumor cells in a sample, the method comprising:
detecting levels of MUC-1, HER2, EGFR, and EpCAM in the sample; and
comparing the levels of MUC-1, HER2, EGFR, and EpCAM in the sample to reference levels;
wherein the presence in a sample of levels of MUC-1, HER2, EGFR, and EpCAM above the reference levels indicates the presence of tumor cells in the sample.
3 . The method of claim 1 , wherein detecting levels of MUC-1, HER2, EGFR, and EpCAM in the sample comprises contacting the sample with antibodies or antigen-binding fragments thereof that bind to MUC-1, HER2, EGFR, and EpCAM.
4 . The method of claim 3 , wherein the antibodies are labeled.
5 . The method of claim 4 , wherein the antibodies are labeled with magnetic nanoparticles.
6 . The method of claim 5 , wherein the magnetic nanoparticles are monocrystalline iron oxide nanoparticles (MIONs).
7 . The method of claim 3 , wherein a single undivided sample is contacted with a mixture of antibodies, or antigen-binding fragments thereof, that bind to MUC-1, HER2, EGFR, and EpCAM, substantially simultaneously.
8 . The method of claim 3 , wherein the sample is subdivided into at least four subparts, and each antibody, or antigen-binding fragment thereof, that binds to MUC-1, HER2, EGFR, or EpCAM is contacted with a single subpart.
9 . The method of claim 1 , wherein the levels of MUC-1, HER2, EGFR, and EpCAM are detected using diagnostic magnetic resonance (DMR); direct magnetic detection; optical detection methods, fluorescence detection; or electric measurements.
10 . The method of claim 1 , wherein the sample comprises blood or a subfraction thereof.
11 . The method of claim 1 , wherein the sample comprises a biopsy sample.
12 . The method of claim 11 wherein the biopsy sample is a fine needle aspirate (FNA), endoscopic biopsy, or core needle biopsy.
13 . The method of claim 1 , wherein the sample comprises cells from the pancreas, lung, breast, prostate, kidney, stomach, esophagus, bladder, endometrial, cervix, biliary, thyroid ovary or colon of the subject.
14 . The method of claim 1 , wherein the tumor is a pancreas, lung, breast, prostate, kidney, stomach, esophagus, bladder, endometrial, cervix, biliary, thyroid ovary or colon tumor.
15 . A kit comprising reagents for detection of tumor cells, wherein the reagents comprise a panel of antigen-binding reagents consisting of:
antibodies or antigen-binding fragments thereof that bind to MUC1,
antibodies or antigen-binding fragments thereof that bind to EGFR,
antibodies or antigen-binding fragments thereof that bind to HER2, and
antibodies or antigen-binding fragments thereof that bind to EpCAM.
16 . The kit of claim 15 , wherein the antibodies or antigen-binding fragments thereof are linked to superparamagnetic nanoparticles.
17 . The kit of claim 15 , wherein the antibodies or antigen-binding fragments thereof are linked to superparamagnetic nanoparticles via trans-cyclooctene (TCO)/tetrazine (Tz) chemistry.
18 . A method for isolating tumor cells from a sample, the method comprising:
providing a sample comprising or suspected of comprising tumor cells;
contacting the sample with:
antibodies or antigen-binding fragments thereof that bind to MUC1,
antibodies or antigen-binding fragments thereof that bind to EGFR,
antibodies or antigen-binding fragments thereof that bind to HER2, and
antibodies or antigen-binding fragments thereof that bind to EpCAM;
under conditions sufficient for the antibodies or antigen-binding fragments thereof to bind to tumor cells in the sample; and
removing the antibodies or antigen-binding fragments thereof that are bound to tumor cells from the sample,
thereby isolating tumor cells from the sample.
19 . The method of claim 18 , wherein the antibodies or antigen-binding fragments thereof are linked to superparamagnetic nanoparticles, and the antibodies or antigen-binding fragments thereof that are bound to tumor cells are removed from the sample by application of a magnetic field to the sample.
20 . The method of claim 18 , wherein the sample comprises blood from a subject.
21 . The method of claim 20 , wherein the method further comprises returning the blood to the subject after removal of the tumor cells.