Methods for detecting 5T4-positive circulating tumor cells and methods of diagnosis of 5T4-positive cancer in a mammalian subject

1. A method for detecting 5T4-positive circulating tumor cells in a mammalian subject suspected of having 5T4-positive cancer and treating the mammalian subject comprising:

obtaining a sample of blood from the mammalian subject;

testing the sample of blood from the mammalian subject, wherein the sample of blood comprises a cell population;

mounting the sample of blood on a substrate;

detecting the presence of a nucleated cell using a first marker in the sample of blood, wherein the first marker is DAPI;

detecting the presence of expression of a second marker on a circulating tumor cell in the sample of blood, wherein the second marker is cytokeratin;

detecting the absence of expression of a third marker on a circulating tumor cell in the sample of blood, wherein the third marker is CD45;

detecting the presence of expression of a fourth marker on a circulating tumor cell in the sample of blood, wherein the fourth marker is human 5T4 antigen;

analyzing the first, second, third, and fourth markers in the cell population to identify and characterize the circulating tumor cells;

quantifying the expression of human 5T4 antigen on the circulating tumor cells, assigning an H-score category, and summing the percentage of cells in each H-score category multiplied by the value of the H-score category to generate an H-score, wherein the H-score is used to select a 5T4-positive cancer patient population; and

administering an effective amount of an anti-5T4 antibody-drug conjugate to the mammalian subject in the 5T4-positive cancer patient population.

2. The method of claim 1 , wherein detecting the presence of 5T4-positive circulating tumor cells in the sample of blood indicates the presence of early stage 5T4-positive cancer in the mammalian subject.

3. The method of claim 1 , wherein detecting the absence of 5T4-positive circulating tumor cells in the sample of blood indicates a disease free state or a non-measurable disease state in the mammalian subject, wherein the disease is 5T4-positive cancer.

4. The method of claim 1 , wherein the presence or absence of the circulating tumor cells in the sample of blood indicates therapy management during 5T4-positive cancer therapy or cancer recovery.

5. The method of claim 1 , wherein the cell population is a mixed cell population.

6. The method of claim 1 , wherein the substrate is a planar substrate, a micro fluidic device, or a cartridge that holds an enriched population of cells.

7. The method of claim 1 , wherein mounting the sample on the substrate forms a biological monolayer.

8. The method of claim 1 , wherein the first marker, the second marker, the third marker, or the fourth marker is detected using fluorescence.

9. The method of claim 1 , wherein the first marker is used in analyzing the cell population by nuclear detail, nuclear contour, presence or absence of nucleoli, quality of cytoplasm, or quantity of cytoplasm.

10. The method of claim 9 , further comprising analyzing the cell population by measuring intact cells with a high nuclear to cytoplasmic ratio, intact cells with a low nuclear to cytoplasmic ratio, early apoptotic cells, or late apoptotic cells, and identifying the circulating tumor cells and circulating tumor cell clusters.

11. The method of claim 1 , further comprising a cytologic stain to identify the circulating tumor cell by morphology, size, or nuclear to cytoplasmic ratio.

12. The method of claim 11 , wherein the cytologic stain is Wright-Giemsa stain.

13. The method of claim 1 , further comprising analyzing the cell population by cell attachment to the substrate, scanning the cell population on the substrate, and imaging the cells by digital microscopy using relocation.

14. The method of claim 1 , wherein detecting the presence of 5T4-positive circulating tumor cells in the sample of blood indicates the presence of a 5T4-positive cancer in a mammalian subject.

15. The method of claim 14 , wherein the 5T4-positive cancer is selected from the group consisting of carcinomas of the bladder, breast, cervix, colon, endometrium, kidney, lung, esophagus, ovary, prostate, pancreas, skin, stomach, and testes.

16. The method of claim 1 , wherein the analyzing of the first, second, third, and fourth markers in the cell population is utilized to determine the inclusion or exclusion of mammalian subjects for the treatment of 5T4-positive cancer.