Circulating exosomes as diagnostic/prognostic indicators and therapeutic targets of melanoma and other cancers

1. A method of measuring total protein content per exosome in a subject having metastatic cancer comprising:

obtaining a sample from the subject having metastatic cancer;

isolating exosomes from the sample;

passing the isolated exosomes through a machine suitable for counting the number of isolated exosomes from the sample;

contacting the isolated exosomes with reagents suitable to measure total protein content in the isolated exosomes; and

measuring the total protein content per exosome based on said passing and said contacting.

2. A method of predicting metastasis in a subject having melanoma comprising:

obtaining isolated exosomes from the subject;

contacting the isolated exosomes with two or more antibodies selected from the group consisting of an anti-MET antibody, an anti-TYRP2 antibody, an anti-VLA-4 antibody, an anti-Hsp-90 antibody, and an anti-Hsp-70 antibody, wherein said antibodies bind to their respective target protein, if expressed by the isolated exosomes, to form two or more different antibody-target protein complexes;

detecting the two or more different antibody-target protein complexes formed as a result of said contacting;

quantifying amounts of the two or more different antibody-target protein complexes formed based on said detecting;

determining the difference in the quantified amounts of the two or more different antibody-target protein complexes formed in the isolated exosomes and amounts of the two or more different antibody-target protein complexes in a reference exosome sample; and

predicting metastasis in the subject having melanoma based on said determining, wherein an increased amount of the two or more different antibody-target protein complexes in the isolated exosomes relative to the reference exosome sample predicts metastasis in the subject.

3. A method of predicting the occurrence of metastatic melanoma in a subject, said method comprising:

obtaining a peripheral blood cell sample from a subject having melanoma;

contacting the peripheral blood cell sample with a collection of antibodies, said collection comprising an anti-MET antibody, an anti-CD45 antibody, and an anti-CD117 antibody, wherein the collection of antibodies bind to their respective target proteins, if present in the sample;

detecting cells in the sample that are positive for CD117 and MET expression (CD117 + /MET + ), but negative for CD45 expression (CD45 − ) based on said contacting;

quantifying the number of cells in the sample that are CD117 + /MET + /CD45 − based on said detecting;

determining the difference between the number of cells in the peripheral blood cell sample from the subject that are CD117 + /MET + /CD45 − and the number of cells that are CD117 + /MET + /CD45 − in a reference peripheral blood cell sample; and

predicting the occurrence of metastatic melanoma in said subject based on said determining, wherein an increase in the number of peripheral cells that are CD117 + /MET + /CD45 − in the sample from the subject relative to the reference cell sample predicts the occurrence of metastatic melanoma.

4. The method according to claim 3 , wherein the collection of antibodies further comprises an anti-TIE2 antibody and said predicting involves detecting and quantifying the number of cells in the peripheral blood sample that are positive for CD117, MET, and TIE2 but negative for CD45, wherein an increase in the number of cells positive for CD117, MET, and TIE2 but negative for CD45 in the peripheral blood sample relative to the reference sample predicts the occurrence of metastatic disease.

5. A method of monitoring metastatic disease treatment in a subject comprising:

obtaining a first and then a second sample of isolated exosomes, at different points in time, from the subject being treated for a metastatic disease;

contacting the isolated exosomes in the first and second samples with two or more antibodies selected from the group consisting of an anti-MET antibody, an anti-TYRP2 antibody, an anti-VLA-4 antibody, an anti-Hsp-90 antibody, and an anti-Hsp-70 antibody, wherein said antibodies bind to their respective target protein, if expressed by isolated exosomes of the first and second samples, to form two or more different antibody-target protein complexes;

detecting the two or more different antibody-target protein complexes formed as a result of said contacting in each of the first and second samples of isolated exosomes:

quantifying the amount of the two or more different antibody-target protein complexes formed based on said detecting in each of the first and second samples of isolated exosomes;

determining the difference between quantified amounts of the two or more different antibody-target protein complexes formed in the first sample of isolated exosomes and amounts of the corresponding two or more different antibody-target protein complexes formed in the second sample of isolated exosomes; and

identifying whether the subject is responding to the metastatic disease treatment based on said determining,

wherein a decrease in the quantified amounts of the two or more different antibody-target protein complexes formed in the second sample of isolated exosomes compared to the first sample of isolated exosomes indicates the subject is responding favorably to the metastatic disease treatment, and

wherein no change in the quantified amounts of the two or more different antibody-target protein complexes formed in second sample of isolated exosomes compared to the first sample of isolated exosomes indicates the subject is not responding favorably to the metastatic disease treatment, said method further comprising modifying the course of treatment for the subject upon a determination that the subject is not responding favorably to the metastatic disease treatment.

6. The method according to claim 2 , wherein the amounts of the two or more different antibody-target protein complexes in the reference exosome sample are the average amounts of the two or more different antibody-target protein complexes in two or more samples from healthy, cancer-free subjects.

7. The method according to claim 2 , wherein the reference exosome sample is a sample obtained from the subject at an earlier time point.

8. The method according to claim 2 , wherein said detecting is carried out by enzyme linked immunosorbent assay.

9. The method according to claim 3 , wherein the number of cells positive for CD117 and MET expression but negative for CD45 expression in the reference cell sample is the average number of cells positive for CD117 and MET expression but negative for CD45 expression in two or more cell samples from healthy, cancer-free subjects.

10. The method according to claim 3 , wherein the reference cell sample is a sample obtained from the subject at an earlier time point.

11. The method according to claim 3 , wherein said detecting is carried out by flow cytometry analysis.