Method of diagnosing and monitoring bladder cancer

Abstract

The present invention provides a method for the diagnosing and monitoring of bladder cancer.

Claims

1. A method for diagnosing the presence of bladder cancer, the method comprising: analyzing human blood, plasma, serum, tears, sweat, vaginal discharge, saliva or urine from the subject at risk for developing bladder cancer with an assay that specifically detects any of the uroplakins in the sample.

2. (canceled)

3. The method of claim 1 in which the uroplakins are: Uroplakin Ia, Uroplakin Ib, Uroplakin II, or Uroplakin III.

4. The method of claim 1 wherein the uroplakin is detected by an analytical method such as, but not limited to, ELISA, radioimmunoassay, competitive binding assay, Western blot, or dot blot.

5. The method of claim 1 wherein the level of the uroplakin above a cutoff is indicative of the presence of bladder cancer in an individual suspected of having bladder cancer.

Description

BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWING

[0032] Example 1 Demonstrating a positive correlation between bladder cancer and the blood level of UPK II.

[0033] FIG. 1, is a dot blot comparing the plasma level of UPK II of a woman with Stage IV bladder versus that of three dilutions of plasma from a healthy sixty-five year old man.

[0034] The normal plasma was from a healthy adult White male. The patient plasma was from a 59 year old White female who was initially diagnosed with localized bladder cancer. The patient had a complete cystectomy twenty months prior to the collection of the plasma sample. The patient was later diagnosed as Stage IV when lung metastases were detected, which was prior to the collection of the sample. Based upon the plasma half-life of human serum albumin, which is twenty-one days, the blood level of uroplakin II had undergone 28.5 half lives and would be reduced to less than two hundred millionth its pre-operative level. See Anderson et al., Mol. Cell. Proteomics 1.11(2002):845-867. Even if the half-life of UPK II in plasma were four times as long as that of albumin, the UPK II blood level would have been reduced to less than one per cent of its pre-operative level.

[0035] A small volume of diluted patient plasma and 1, 3 and 10 volumes of the normal plasma, relative to the bladder cancer patient plasma, were spotted on two strips of nitrocellulose. The other binding sites on the nitrocellulose were covered by incubating the nitrocellulose in a solution of bovine serum albumin.

[0036] A mouse monoclonal antibody generated against a peptide of human Uroplakin II was obtained from a commercial vendor. This antibody has been shown to react exclusively with normal urothelium or urothelium-derived cancer tissue. See Hoang et al., Arch. Pathol. Lab. Med. 138.7(2014):943-949. The antibody was biotinylated and reacted with streptavidin Alkaline-Phosphatase to produce an antibody with the reporter Alkaline-Phosphatase attached. The experimental and control strips were put into incubation media with or without the anti-UPK II Alkaline Phosphatase complex, respectively. Both were washed, then put into buffer containing the substrate for alkaline phosphatase to visualize the UPK II.

[0037] Both strips of nitrocellulose show positive reactions in nearly all spots including the controls. Reaction product in the no antibody controls is due to a component of human plasma that reacts with the substrate for alkaline phosphatase. The plasma from the healthy male had much more of this component than the patient plasma, which was nearly zero.

[0038] The normal plasma shows a non-linear increase in UPK II with the amount of plasma analyzed.

[0039] The relative quantity of UPK II in the patient plasma cannot be determined exactly, but it is greater than the 1 volume and less than the 10 volume of Normal Plasma. Regardless of the amount of UPK II in the patient plasma relative to the healthy individual, the amount is much greater than the nearly zero quantity that would be expected twenty months after the UPK II producing tissues had been surgically removed from the patient.

[0040] Since UPK II and the other uroplakins are synthesized only by urothelial cells, the UPK II in the patient plasma must have been synthesized by the metastases of the bladder cancer.

[0041] These results indicate that the level of and of the four uroplakin proteins in blood and other human bodily fluid could be used in diagnostic assays for the diagnosing and monitoring of bladder cancer.

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