WO2006130689A2

WO2006130689A2 - Detecting prostate cancer

Info

Publication number: WO2006130689A2
Application number: PCT/US2006/021118
Authority: WO
Inventors: Michael J. Wilson; Neil F. Wasserman; Akhouri Sinha
Original assignee: Regents Of The University Of Minnesota
Priority date: 2005-06-02
Filing date: 2006-06-01
Publication date: 2006-12-07
Also published as: WO2006130689A3

Abstract

This document provides methods and materials related to detecting prostate cancer. For example, methods and materials for assessing the likelihood of prostate cancer in a mammal by determining the level of a dipeptidylpeptidase IV polypeptide are provided. Methods and materials for assessing prostate cancer progression and prostate cancer regression in a mammal by determining the level of a dipeptidylpeptidase IV polypeptide also are provided.

Description

DETECTING PROSTATE CANCER

Statement as to Federally Sponsored Research Funding for the work described herein was provided by the federal government, which may have certain rights in the invention.

BACKGROUND

1. Technical Field

This document relates to methods and materials involved in assessing prostate cancer in a mammal.

2. Background Information

Other than skin cancer, prostate cancer is the most prevalent malignancy in adult males, and its incidence increases with age. In the United States, there are about 132,000 newly diagnosed cases of prostate cancer and more than 33,000 deaths from the disorder each year. Patients with early stage prostate cancer may exhibit no clinical symptoms or may exhibit symptoms that are similar to those of benign prostate conditions. The likelihood of being cured is higher if prostate cancer is diagnosed and treated early when still confined to the prostate. A primary diagnostic marker for prostate cancer is prostate specific antigen

(PSA). PSA is a tissue-specific serine protease produced by prostatic epithelial cells. The amount of PSA can correlate with the number of prostatic epithelial cells, and thus, the levels of PSA can be used as an indicator of abnormal prostate growth.

SUMMARY

This document provides methods and materials related to detecting prostate cancer. For example, this document provides methods for assessing the likelihood of prostate cancer in a mammal as well as kits that can be used to assess the likelihood of prostate cancer in a mammal. Detecting prostate cancer as described herein can allow clinicians to diagnose prostate cancer patients in an early and reliable manner, thereby increasing the patient's likelihood of being cured. As described herein, an elevated level of a dipeptidylpeptidase IV (DPIV) polypeptide can indicate that a mammal has prostate cancer with a volume of 4.0 cm³ or less. In addition, the level of DPIV polypeptide activity in prostate secretions from men with prostate cancer can be elevated as compared to those levels in secretions from men with no evidence of malignancy upon prostate biopsy. Further, men with cancer can exhibit elevated DPIV polypeptide activity in the peripheral zone (PZ), the zone most commonly involved with cancer in the prostate. Detecting prostate cancer as described herein also can allow clinicians to monitor prostate cancer progression in, for example, untreated patients, or to monitor prostate cancer regression in, for example, treated patients. Monitoring prostate cancer in untreated patients using the methods and materials provided herein can allow clinicians to not only determine when to start prostate cancer treatment but also which type of prostate cancer treatment to use. Monitoring prostate cancer in patients undergoing prostate cancer treatment can allow clinicians to determine the effectiveness of the treatment and make adjustments as needed.

In general, this document features a method for assessing the likelihood of prostate cancer in a mammal, wherein the volume of the prostate cancer is 4.0 cm³ or less. The method includes determining whether or not a prostate transition zone or prostate secretion sample from the mammal contains an elevated level of a DPIV polypeptide as compared to the level of the DPIV polypeptide in a control sample, wherein the presence of the elevated level indicates that the mammal likely has the prostate cancer. The volume of the prostate cancer can be between 0.5 and 4.0 cm³. The volume of the prostate cancer can be less than 0.5 cm³. The mammal can be a human. The sample can be a prostatic secretion sample. The sample can be a urine sample. The sample can be a urine sample containing the first one, two, three, four, or five mL of urine from the mammal following a prostate digital rectal examination. The sample can be a prostate transition zone sample. The sample can be a prostate transition zone sample obtained via endoscopy. The level of the DPIV polypeptide can be determined using an immunoassay. The level of the DPIV polypeptide can be determined by measuring DPIV polypeptide activity. The control sample can be from a prostate cancer-free mammal. The sample can be a prostatic secretion sample, and the elevated level can result in greater than 25 units of DPIV polypeptide activity per mL of the sample when the level of the DPIV polypeptide in the control sample results in between 15 and 22 units of DPIV polypeptide activity per mL of the control sample. The sample can be a prostatic secretion sample, and the elevated level can result in greater than 30 units of DPIV polypeptide activity per mL of the sample when the level of the DPIV polypeptide in the control sample results in between 15 and 22 units of DPIV polypeptide activity per mL of the control sample. The sample can be a prostate transition zone sample, and the elevated level can result in greater than 0.17 units of DPIV polypeptide activity per mg protein in the sample when the level of the DPIV polypeptide in the control sample results in less than 0.17 units of DPIV polypeptide activity per mg protein in the control sample. The sample can be a prostate transition zone sample, and the elevated level can result in greater than 0.21 units of DPIV polypeptide activity per mg protein in the sample when the level of the DPIV polypeptide in the control sample results in less than 0.17 units of DPIV polypeptide activity per mg protein in the control sample. The method can include determining whether or not the level of a neprilysin, tripeptidylpeptidase II, or aminopeptidase N polypeptide is reduced in the mammal as compared to the level of the neprilysin, tripeptidylpeptidase II, or aminopeptidase N polypeptide in a control mammal not having prostate cancer. The method can include determining whether or not the mammal contains an elevated level of prostate specific antigen. The control sample can be a sample from the mammal obtained earlier in life. The control sample can be a sample from the mammal obtained when the mammal did not have prostate cancer. In another embodiment, this document features a method for assessing the likelihood of prostate cancer in a mammal. The method includes determining whether or not a prostate peripheral zone sample from the mammal contains an elevated level of a DPIV polypeptide as compared to the level of the DPIV polypeptide in a control sample, wherein the presence of the elevated level indicates that the mammal likely has the prostate cancer. The volume of the prostate cancer can be between 0.5 and 4.0 cm³. The volume of the prostate cancer can be greater than 4.0 cm³. The mammal can be a human. The prostate peripheral zone sample can be a sample obtained via endoscopy. The level of the DPIV polypeptide can be determined using an immunoassay. The level of the DPTV polypeptide can be determined by measuring DPW polypeptide activity. The control sample can be from a prostate cancer-free mammal. The elevated level can result in greater than 0.09 units of DPIV polypeptide activity per mg protein in the sample when the level of the DPrV polypeptide in the control sample results in less than 0.09 units of DPrV polypeptide activity per mg protein in the control sample. The elevated level can result in greater than 0.12 units of DPIV polypeptide activity per mg protein in the sample when the level of the DPIV polypeptide in the control sample results in less than 0.09 units of DPIV polypeptide activity per mg protein in the control sample. The method can include determining whether or not the level of a neprilysin, tripeptidylpeptidase II, or aminopeptidase N polypeptide is reduced in the mammal as compared to the level of the neprilysin, tripeptidylpeptidase II, or aminopeptidase N polypeptide in a control mammal not having prostate cancer. The method can include determining whether or not the mammal contains an elevated level of prostate specific antigen. The control sample can be a sample from the mammal obtained earlier in life. The control sample can be a sample from the mammal obtained when the mammal did not have prostate cancer. hi another embodiment, this document features a method for monitoring prostate cancer progression in a mammal. The method includes determining whether or not a sample from the mammal contains an elevated level of a DPIV polypeptide as compared to the level of the DPIV polypeptide in a sample obtained from the mammal at an earlier point in time, wherein the elevated level indicates that the prostate cancer has progressed. The sample can be a prostatic secretion sample. The sample can be a urine sample. The sample can be a urine sample containing the first one, two, three, four, or five mL of urine from the mammal following a prostate digital rectal examination. The sample can be a prostate transition zone sample or a prostate peripheral zone sample. The sample can be obtained via endoscopy. hi another embodiment, this document features a method for detecting prostate cancer regression in a mammal. The method includes determining whether or not a prostate peripheral zone sample from the mammal contains an reduced level of a DPIV polypeptide as compared to the level of the DPIV polypeptide in a control sample obtained from the mammal at an earlier point in time, wherein the reduced level indicates that the prostate cancer has regressed. The sample can be obtained via endoscopy.

In another embodiment, this document features a method for monitoring a response to prostate cancer treatment in a mammal having prostate cancer with a volume of 4.0 cm³ or less. The method includes determining whether or not a sample obtained from the mammal after the treatment contains a reduced level of a DPrV polypeptide as compared to the level of the DPIV polypeptide in a control sample obtained from the mammal at an earlier point in time, wherein the reduced level indicates that the prostate cancer treatment reduced the volume of prostate cancer in the mammal. The sample can be a prostatic secretion sample. The sample can be a urine sample. The sample can be a urine sample containing the first one, two, three, four, or five mL of urine from the mammal following a prostate digital rectal examination. The sample can be a prostate transition zone sample or a prostate peripheral zone sample. The sample can be obtained via endoscopy. The prostate cancer treatment can be a radiation therapy. The prostate cancer treatment can be a hormone therapy.

In another embodiment, this document features a method for monitoring a response to prostate cancer treatment in a mammal (e.g., human) having prostate cancer. The method includes determining whether or not a prostate peripheral zone sample obtained from the mammal after the treatment contains a reduced level of a DPIV polypeptide as compared to the level of the DPIV polypeptide in a control sample obtained from the mammal at an earlier point in time, wherein the reduced level indicates that the prostate cancer treatment reduced the volume of prostate cancer in the mammal.

In another embodiment, this document features a kit containing an anti- DPrV polypeptide antibody and an antibody selected from the group consisting of an anti-prostate specific antigen antibody, an anti-neprilysin polypeptide antibody, an anti-tripeptidylpeptidase II polypeptide antibody, and an anti-aminopeptidase N polypeptide antibody.

Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Although methods and materials similar or equivalent to those described herein can be used to practice the invention, suitable methods and materials are described below. AU publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and not intended to be limiting.

The details of one or more embodiments of the invention are set forth in the accompanying drawings and the description below. Other features, objects, and advantages of the invention will be apparent from the description and drawings, and from the claims.

DESCRIPTION OF THE DRAWINGS

Figure 1 is a graph plotting the ROC curve from the logistic regression of DPrV polypeptide activities in prostatic secretion and serum PSA values. Area under the curve is 0.80 for DPIV polypeptide activities and is 0.57 for PSA. A cutoff at 26.4 U/mL DPIV polypeptide activity gives a sensitivity of 0.765 and a specificity of 0.867.

DETAILED DESCRIPTION This document provides methods and materials related to detecting prostate cancer. For example, this document provides methods for assessing the likelihood of prostate cancer in a mammal as well as kits that can be used to assess the likelihood of prostate cancer in a mammal. This document also provides methods and materials for assessing prostate cancer progression, prostate cancer regression, prostate cancer recurrence, and a patient's response to prostate cancer therapy. The methods and materials provided herein can be used to determine whether or not a mammal has prostate cancer or is likely to have prostate cancer. The mammal can be a human, non-human primate, goat, horse, cow, pig, dog, or cat. A mammal can be classified as having prostate cancer or as being likely to have prostate cancer by assessing the level of a DPIV polypeptide in a sample from the mammal. For example, a mammal found to have an elevated level of a DPIV polypeptide in the transition zone of the prostate or in a prostate secretion can be classified as having (or as being likely to have) prostate cancer with a volume of 4.0 cm³ or less. In some cases, a mammal found to have a reduced level of a DPIV polypeptide in the transition zone of the prostate or in a prostate secretion can be classified as having (or as being likely to have) prostate cancer with a volume of greater than 4.0 cm³. In some cases, a mammal found to have an elevated level of a DPIV polypeptide in the peripheral zone of the prostate can be classified as having (or as being likely to have) prostate cancer.

The term "elevated" as used herein with reference to a DPIV polypeptide level refers to any level of a DPIV polypeptide that is greater than either the level of a DPIV polypeptide found in a control sample or the average level of a DPIV polypeptide found in samples from a population of normal healthy mammals (e.g., a population of humans known to be free of prostate cancer). The term "reduced" as used herein with reference to a DPIV polypeptide level refers to any level of a DPIV polypeptide that is less than either the level of a DPIV polypeptide found in a control sample or the average level of a DPIV polypeptide found in samples from a population of normal healthy mammals (e.g., a population of humans known to be free of prostate cancer). A control sample can be a comparable sample obtained from a mammal without prostate cancer or from the same mammal but at a time when the mammal was known to be free of prostate cancer. For example, when assessing a particular mammal for the likelihood of having prostate cancer, the level of a DPIV polypeptide found in a prostate sample from that mammal can be compared to the level of DPIV polypeptide found in a sample obtained from that same mammal at a point in that mammal's life when it was known to be free of prostate cancer. Any population size can be used to determine the average level of a DPIV polypeptide found in samples from a population of normal healthy mammals. For example, a population of 2, 3, 4, 5, 10, 15, 20, 25, 30, 40, 50, or more humans free of prostate cancer can be used to determine the average level of a DPIV polypeptide in samples from a population of normal healthy mammals. An elevated level of a DPIV polypeptide can be 1 , 2, 3, 4, 5, 10, 20, 30, 50, or more percent higher than that level found in a control sample or the average level of a DPIV polypeptide found in samples from a population of normal healthy mammals. In some cases, an elevated level of a DPIV polypeptide can be 1, 2, 3, 4, 5, 10, 50, 100, or more fold higher than that level found in a control sample or the average level of a DPIV polypeptide found in samples from a population of normal healthy mammals. A reduced level of a DPIV polypeptide can be 1, 2, 3, 4, 5, 10, 20, 30, 50, or more percent lower than that level found in a control sample or the average level of a DPIV polypeptide found in samples from a population of normal healthy mammals. In some cases, a reduced level of a DPIV polypeptide can be 1, 2, 3, 4, 5, 10, 50, 100, or more fold lower than that level found in a control sample or the average level of a DPIV polypeptide found in samples from a population of normal healthy mammals.

In some cases, a reference chart can be used to determine whether or not a particular level of a DPIV polypeptide in a sample is elevated, normal, or reduced. For example, a reference chart can contain the normal range of DPIV polypeptide levels found in prostate secretions, prostate transition zone tissues, and prostate peripheral zone tissues from humans free of prostate cancer. Using this reference chart, any level of a DPIV polypeptide measured in a sample can be classified as being an elevated level, a normal level, or a reduced level.

As described herein, the presence of an elevated level of a DPIV polypeptide in the transition zone of the prostate or in a prostate secretion can indicate that the mammal has or is likely to have prostate cancer with a volume of 4.0 cm³ or less. The presence of a reduced level of a DPIV polypeptide in the transition zone of the prostate or in a prostate secretion can indicate that the mammal has or is likely to have prostate cancer with a volume of greater than 4.0 cm³. The presence of an elevated level of a DPIV polypeptide in the peripheral zone of the prostate can indicate that the mammal has or is likely to have prostate cancer, m some cases, the prior presence of an elevated level of a DPIV polypeptide in the transition zone of the prostate or in a prostate secretion followed by a subsequent presence of a reduced level of a DPIV polypeptide in the transition zone of the prostate or in a prostate secretion can indicate that the mammal has or is likely to have prostate cancer with a volume of greater than 4.0 cm³.

Any method can be used to determine the volume of prostate cancer within a mammal. For example, biopsy samples or imaging techniques (e.g., ultrasound, computed tomography, or magnetic resonance imaging) can be used to determine the volume of prostate cancer. In some cases, prostate cancer volume can be a calculated volume of cancer (cVca) computed based on PSA levels. For example, the cVca can be equal to the cancer-specific PSA/(PSA in serum/cm³ cancer, where the volume of cancer is a function of the Gleason score; e.g., Gleason grade 1 = 20, grade 2 = 10, grade 3 = 4, and grade 5 = 1 ng/mL per cm³. In addition, a mammal's prostate cancer volume can be classified, based on estimated cVca values, as low (O.05 cm³), moderate (0.5-4.0 cm³), or high (>4.0 cm³). Cancer specific PSA can be calculated as the portion of PSA in serum that arises from malignant prostate epithelium and represents total PSA minus the PSA from benign prostatic hyperplaisic (BPH) tissue. The PSA from BPH tissue can be calculated as equal to the epithelial component fraction of prostate x PSA/cm³ produced by epithelial tissue x prostate volume. For this purpose, the amount of serum PSA attributed to normal epithelium can be 33 ng/mL per cm³ epithelium, and 20% of prostate volume can be estimated to be composed of epithelium. Thus, the PSA from BPH epithelium can be estimated by multiplying the total prostate volume determined by ultrasound x 0.066 ng/mL per cm³. In the case of humans, a DPIV polypeptide can be a human DPIV polypeptide such as a polypeptide having the amino acid sequence set forth in Genbank Accession No. P27487. In the case of non-human mammals, a DPIV polypeptide can be a corresponding DPIV polypeptide. For example, in the case of cats, a DPrV polypeptide can be a cat DPIV polypeptide such as a polypeptide having the amino acid sequence set forth in Genbank Accession No. Q9N217. In the case of cows, a DPIV polypeptide can be a bovine DPIV polypeptide such as a polypeptide having the amino acid sequence set forth in Genbank Accession No. P81425.

The level of a DPIV polypeptide within any sample can be determined. For example, the level of a DPIV polypeptide within a prostate secretion or a prostate tissue sample can be determined. Typically, samples containing prostate tissue, material produced by prostate tissue, or a mixture thereof are used as described herein to assess prostate cancer. Examples of such samples include, without limitation, prostate tissue biopsy samples, biopsy samples obtained from the transitional zone of the prostate gland, biopsy samples obtained from the peripheral zone of the prostate gland, biopsy samples obtained from a combination of the peripheral and transitional zones of the prostate gland, prostatic secretions, seminal plasma, blood samples, serum samples, and urine samples. In some cases, the level of a DPIV polypeptide in a sample obtained from non-prostate tissue (e.g., kidney, liver, and intestine tissue) can be used to assess prostate cancer as described herein provided that the level of DPIV polypeptide within that sample correlates with the level typically found in prostate tissue or prostate secretions under normal and prostate cancer conditions.

Any method can be used to obtain a sample from a mammal. For example, prostate biopsy tissue (e.g., transitional zone tissue, peripheral zone tissue, or a mixture thereof) can be collected using ultrasound guidance via a transrectal approach or via transperineal biopsy. In some cases, prostate massage can be used to obtain prostatic secretions. Blood samples can be obtained via venous puncture techniques. Serum samples can be prepared from whole blood using standard methods such as centrifuging blood samples that have been allowed to clot. Plasma samples can be obtained by centrifuging blood samples that were treated with an anti-coagulant such as heparin, hi some cases, a urine sample can be collected from a mammal and used to assess prostate cancer as provided herein.. For example, the first 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, or more mL of urine released following a digital rectal examination can be collected and used as provided herein.

Any method that can be used to determine the level of a DPIV polypeptide in a sample. For example, the level of a DPIV polypeptide can be determined by measuring the level of mRNA that encodes a DPIV polypeptide, by measuring the level of the DPIV polypeptide itself, or by measuring the level of DPIV polypeptide activity. Any method can be used to measure the level of mRNA that encodes a DPIV polypeptide including, without limitation, northern blotting techniques, slot blotting techniques, quantitative reverse transcriptase polymerase chain reaction (RT-PCR) techniques, or chip hybridization techniques. In addition, any method can be used to measure the level of the DPIV polypeptide itself including, without limitation, immunoassays such as ELISA or immunoblotting techniques. Any method can be used to measure the level of DPIV polypeptide activity including, without limitation, enzyme assays and ligand binding assays. In some cases, the level of DPIV polypeptide activity can be determined by measuring the amount of hydrolysis products generated by a DPIV polypeptide from naturally-occurring or synthetic substrates such as L-glycylprolyl-p-nitroanalide or ala-pro- chlormethylcoumarin(CMAC). The amount of hydrolysis products can be measured using any standard photometric or fluorometric method. In some cases, the level of DPIV polypeptide activity can be determined using binding or signal transduction assays that monitor the interaction of a DPIV polypeptide with a specific ligand such as plasminogen type II or angiostatin 2epsilon. Prior to determining the level of a DPIV polypeptide in a sample, the sample can be subjected one or more standard preparatory procedures. For example, to measure the level of DPIV polypeptide activity, a tissue biopsy sample can be fixed (e.g., fixed in formalin), rinsed, and exposed to detergent (e.g., triton-x-100) to yield an extract. The detergent can be in a buffer that preserves a DPIV polypeptide activity (e.g., a phosphate buffered saline at neutral pH). The total protein concentration in the extract can be determined using any standard method such as Bradford reagent or bicinchoninic acid techniques.

When the level of DPIV polypeptide activity is determined measuring enzymatic hydrolysis of, for example, a synthetic substrate such as L glycylprolyl-p- nitroanalide, 1 unit of enzyme activity can be the amount of DPIV polypeptide catalyzing the formation of 1 mmol p-nitroanalide/minute. For example, a human having a level of DPIV polypeptide activity such that a prostatic secretion sample contains more than 23, 24, 25, 26, 27, 28, 29, 30, or more units/mL compared to a control sample (e.g., a prostate secretion sample from a healthy human with no evidence of prostate cancer) having 15 to 22 units/mL can be classified as having or as likely to have prostate cancer with a volume of 4.0 cm³ or less. In another example, a human having a level of DPIV polypeptide activity in a prostate transitional zone sample that is greater than 0.17, 0.18, 0.19, 0.2, or more units/mg protein compared to a control sample (e.g., a prostate transitional zone sample from a healthy human with no evidence of prostate cancer) having between 0.11 and 0.17 units/mg protein can be classified as having or as likely to have prostate cancer with a volume of 4.0 cm³ or less, hi an additional example, a human having a level of DPIV polypeptide activity in a prostate peripheral zone sample that is greater than 0.09, 0.1, 0.12. 0.13, 0.14, 0.15, or more units/mg protein compared to a control sample (e.g., a prostate peripheral zone sample from a healthy human with no evidence of prostate cancer) having between 0.05 and 0.09 units/mg protein can be classified as having or as likely to have prostate cancer.

In some cases, the level of a DPIV polypeptide can be determined as a ratio with the level of another polypeptide such as a polypeptide that is expressed at low levels in prostate cancer (e.g., a neprilysin polypeptide, a tripeptidylpeptidase II (TPII) polypeptide, or a aminopeptidase N polypeptide). For example, the ratio of the level of a DPIV polypeptide to the level of a neprilysin polypeptide within a prostate secretion sample (e.g., a urine sample or blood sample) can be compared to a control ratio to assess prostate cancer as described herein.

Once identified as having or as likely to have prostate cancer, the mammal can begin prostate cancer treatment or can undergo additional testing to confirm or evaluate the prostate cancer. For example, a mammal classified as likely having prostate cancer as described herein can initiate one or more prostate cancer treatments such as surgery, radiation therapy, or chemotherapy. Surgical treatments can be radical prostatectomy, cryosurgery, or orchiectomy. Radiation therapy can be external beam radiation therapy or interstitial implantation of radioisotopes (e.g., brachytherapy). Chemotherapy can include treatments with leutenizing hormone releasing hormone (LHRK-) agonists, estrogen, or steroidal or non-steroidal anti- androgens. In some cases, a mammal classified as likely having prostate cancer as described herein can undergo diagnostic tests such as biopsy procedures, ultrasound procedures, or magnetic resonance imaging to confirm the existence or severity of prostate cancer.

The methods and materials provided herein can be used to assess prostate cancer progression and regression as well as to assess a mammal's response to a prostate cancer treatment. For example, the presence of an elevated level of a DPIV polypeptide in a sample (e.g., a prostate transitional zone sample, a prostate peripheral zone sample, or a prostate secretion sample) that is higher than an earlier measured level that was also elevated can indicate that that mammal's prostate cancer is progressing. In some cases, the presence of a reduced level of a DPIV polypeptide in a sample (e.g., a prostate transitional zone sample, a prostate peripheral zone sample, or a prostate secretion sample) that is lower than an earlier measured level that was elevated can indicate that that mammal's prostate cancer is regressing.

In one embodiment, the methods and materials provided herein can be applied to a treatment option described in the art as "watchful waiting" or "expectant management." A watchful waiting approach to prostate cancer management can include closely monitoring the prostate cancer without surgery or radiation therapy. For example, a baseline level of a DPIV polypeptide can be established in a human at the time of initial prostate cancer diagnosis. Additional samples (e.g., blood samples, urine samples, or prostate tissue samples) can be obtained at intervals (e.g., every month, every other month, every six months, or every twelve months) after diagnosis in order to assess whether or not the level of DPIV polypeptide changed as compared to the patient's baseline level or a level determined from any earlier collected sample. An increase in the level of a DPIV polypeptide relative to the baseline level can indicate that the volume of the prostate cancer has increased and that more active treatment should be started. In some cases, a decrease in the level of a DPIV polypeptide in a prostate transitional zone sample or prostate secretion sample relative to the baseline level can indicate that the volume of the prostate cancer has increased above 4.0 cm³ and that more active treatment should be started. Such treatment can include surgery, radiation therapy, chemotherapy, or a combination thereof.

The methods and materials provided herein also can be used to monitor the response of patients undergoing prostate cancer treatment (e.g., surgery, radiation therapy, or chemotherapy). For example, a baseline level of a DPIV polypeptide can be determined prior to initiating the prostate cancer therapy. Additional samples (e.g., blood samples, urine samples, or prostate tissue samples) can be obtained at intervals (e.g., every week, every other week, every month, every other month, or every six months) during or after the therapy in order to assess whether or not the level of DPIV polypeptide changed as compared to the patient's baseline level or a level determined from any earlier collected sample. A decrease in the level of a DPIV polypeptide relative to the baseline level can indicate that the volume of the prostate cancer has decreased and that the prostate cancer treatment is effective. An increase in the level of a DPIV polypeptide relative to the baseline level can indicate that the volume of the prostate cancer has increased, that the prostate cancer treatment is not effective, and that an additional or alternative prostate cancer treatment should be initiated.

This document also provides kits that can be used to assess prostate cancer. For example, reagents used to determine the level of a DPIV polypeptide in a sample can be combined as an article of manufacture such as a kit. In one embodiment, a kit can contain reagents for the immunodetection of a DPIV polypeptide. For example, a kit can contain an anti-DPIV polypeptide antibody. In some cases, such a kit can include suitable reagents for detecting the binding of the anti-DPIV polypeptide antibody to a DPIV polypeptide with or without additional antibodies such as antibodies having the ability to bind a neprilysin polypeptide, a tripeptidylpeptidase II polypeptide, an aminopeptidase N polypeptide, PSA, a chromogranin A polypeptide, a prostate stem cell antigen, or a prostate-specific membrane antigen. In some cases, a kit can contain a synthetic or naturally-occurring substrate for a DPIV polypeptide (e.g., L-glycylprolyl-p-nitroanalide or ala-pro-CMAC). Such kits can include buffers for lysing cells, assay reaction buffers, a positive control sample (e.g., a solution containing active DPIV polypeptide), or combinations thereof.

The reagents within a kit can be housed together in various combinations or can be packaged in separate vials or containers. The kits provided herein also can include labels or packaging inserts setting out instructions for preparation and use.

The invention will be further described in the following examples, which do not limit the scope of the invention described in the claims.

EXAMPLES

Example 1 - DPIV activities and prostate cancer Patient group

Seventy-one men at high risk for prostate cancer and about to undergo an ultrasound-guided biopsy of the prostate were recruited into this study. These patients generally were referred for prostate biopsy due to an elevation of serum PSA (PSA >4 ng/mL) or an abnormal digital rectal examination. Informed consent was obtained to collect (a) expressed prostatic secretion (EPS) before undergoing the ultrasound guided biopsy procedure, (b) a biopsy core from the peripheral zone (PZ) and a biopsy core from the transition zone (TZ) during the biopsy procedure, and (c) clinical information (e.g., serum PSA levels, ultrasound imaging and prostate zone volumes, and pathological reports on the biopsies taken) relevant to the analysis of DPIV activity data. The EPS samples were collected by prostate massage from 40 of the 71 men. Secretions were centrifuged at 960 X g for 10 minutes at 4°C to remove cells. The supernatant was frozen in powdered dry ice, and then stored at -20° C until assayed for DPIV activity.

Ultrasound imaging and prostate biopsy

Ultrasound imaging of patients for biopsy was done with a biplane transrectal ultrasound probe utilizing high frequency transducers (greater than 5.5 MHz). Gray-scale images were recorded in the axial and longitudinal planes at multiple sites through the prostate and recorded on film for hard copy interpretation. The same person performed the ultrasound imaging, biopsy procedure, and prostate zone measurements. The prostate volume measurements were obtained by measuring the maximum anteroposterior (AP) diameter either in the axial or longitudinal sagittal plane, the maximum transverse (Trans) diameter in the axial plane, and the length in the sagital plane. The volume was calculated using the following formula for a prolate ellipsoid:

Vcc = cm AP diameter X cm Trans diameter X cm Length Total volume and volume of the inner glandular tissues containing the TZ were obtained. The volume of the PZ was approximated by subtracting the TZ volume from the total volume. Central zone volume, which is not separately discernable on ultrasound and is negligible in size in older men due to compression from surrounding zonal growth, was included in the PZ measurement.

Multiple biopsies were obtained from each prostate for clinical evaluation using ultrasound guidance via a transrectal approach and a Bard Biopty spring- loaded device (C. R. Bard, Inc., Covington, GA) with a 19-gauge needle producing a 1.25 cm length core. Systematic biopsy specimens were taken from the right and left sides of the prostate, including anterolateral tissues (3-6 each side depending on prostate size). Three cores were obtained from hypoechoic PZ foci suspicious for carcinoma. Biopsy specimens were segregated into separate formalin-containing bottles specifically labeled according to site of specimen origin.

Single additional biopsy samples for this study were removed from normal appearing tissue areas in the TZ and PZ that were not grossly suspicious for cancer based upon ultrasound imaging, were placed in 0.1% Triton X-100, and were frozen at -70⁰C for DPIV assay.

DPIV activity assay

Prostate biopsies were homogenized in 0.1% Triton X-100 using 10 strokes with a teflon-glass tissue grinder at 4° C followed by centrifugation at 4° C for 10 minutes at 960 x g. The supernatant was removed for protein and DPIV activity measurements. Protein concentration was determined by the bicinchoninic acid method (Smith et al, Anal. Biochem., 150:76-85 (1985). DPIV activity was measured at room temperature in a final volume of 100 μL containing 70 mmol/L glycine NaOH buffer (pH 8.7), 1.4 mmol/L glycylprolyl-p-nitoanalide (Sigma Chemical Co., St. Louis, MO), and appropriately diluted prostate biopsy extract or prostatic secretion in a 96-well, flat-bottom microtiter plate. Substrate was added to start the reaction. The microtiter plate was read immediately and then at timed intervals at 405 nm in a 3550 plate reader (BioRad, Richmond, CA). The amount of product formed was determined from a standard curve of p-nitroanalide. One unit of enzyme activity was defined as the amount of enzyme catalyzing the formation of 1 μmol p-nitroanalide/minute.

Calculating cancer-specific PSA and volume of cancer (cVca)

Cancer specific PSA was computed as the portion of PSA in serum that arose from malignant prostate epithelium and represents total PSA-PSA from BPH tissue. The PSA from BPH tissue was calculated as equal to the epithelial component fraction of prostate x PSA/cm³ produced by epithelial tissue x prostate volume. For this purpose, the amount of serum PSA attributed to normal epithelium was 33 ng/mL per cm³ epithelium, and 20% of prostate volume was estimated to composed of epithelium. Thus, the PSA from BPH epithelium was estimated by multiplying the total prostate volume determined by ultrasound x 0.066 ng/mL per cm³. The cVca was equal to the cancer-specific PSA/(PSA in serum/ cm³ cancer, where the volume of cancer is a function of the Gleason score; i.e. Gleason grade 1 = 20, grade 2 = 10, grade 3 = 4, and grade 5 = 1 ng/mL per cm³. Patient data were grouped as to estimated volume of cancer as cVca <0.05 (low), 0.5-4.0 (moderate), and >4.0 cm³ (high).

Statistical Analysis

Statistical analysis was used to explore associations between DPIV activities in EPS, the total volume of the prostate, and the volumes of the TZ and PZ, as well as the associations between DPIV activities in secretions, TZ, and PZ and prostate cancer. Data were summarized based on the biopsy diagnosis. Since DPIV activities did not appear to be normally distributed, the Wilcoxon Rank Sum Test was used to compare the median of the variables. Pearson's correlation analysis was used to investigate the linear relationship between DPIV activities in expressed prostatic secretions and DPIV activities in both the TZ and PZ. Using DPIV in expressed prostatic secretions as a response variable, simple linear regression was used to evaluate whether age has an effect on the DPIV activity values. The paired T-test was adopted to compare the mean differences of DPIV activities and volumes of TZ and PZ for each subject among the NEM patients.

Patient characteristics There was no difference in the mean age or serum PSA levels of patients when the patients were grouped according to one of three biopsy diagnoses: (1) no evidence of malignancy (NEM), (2) prostatic high-grade intraepithelial neoplasia or atypical small acinar proliferation (PIN/atypia), or (3) cancer (Table 1). DPIV activities were studied in 71 patients, but prostatic secretions were not collected from all men from whom biopsies were obtained. Secretions were collected from 15 of 28 men with NEM, 7 of 12 patients with PIN/atypia, and 18 of 31 men with a diagnosis of cancer. The majority of patients with biopsy diagnosed prostate cancer had Gleason scores of 6 and 7.

Table 1. Patient characteristics and diagnostic groups

Diagnostic Biopsy EPS group Number Age PSA Number Age PSA

NEM 28 70.1±l.l^a 7.8±1.5 15 68.9±1.6 8.5±2.5

PTN/Atypia 12 70.1±2.0 6.9±2.1 7 68.9±3.4 7.4±4.9

Cancer 31 70.4±1.4 8.4±1.6 18 70.8±2.0 7.4±1.9

Gleason 3 2 2

Gleason 6 10 7

Gleason 7 17 7

Gleason 8 2 2 a = S.E. Prostatic tissue sources of secreted DPIV

For men with NEM, the specific activity of DPIV per unit of protein in TZ biopsies was significantly greater than that in the PZ (p=0.008; Table 2). In addition, in men with NEM, the TZ was significantly larger than the PZ. In contrast, DPIV activity in secretion was inversely correlated with TZ volume, that is the correlation of DPIV activity in secretion with TZ volume is negative (p=-0.287, p=0.076), whereas the correlation of DPIV activity in secretion with PZ volume was positive (p=0.358, p=0.151). Thus, association between TZ volume and DPIV activity in secretion was statistically highly significant (p=0.006), and the association between PZ volume and DPIV activity in secretion was also significant (p=0.022). Since DPIV is a component of membrane vesicles (prostasomes) secreted by the prostate gland, the negative association of secreted DPIV activities with TZ volume and positive association of secreted DPIV activities with PZ volume indicates that the PZ releases more prostasomes into the secretion than does the TZ.

(U/mg P)= Units Activity/mg Protein (U/mL)= Units Activity/mL prostatic secretion SE = standard error of mean a= NEM TZ DPrV activity vs. NEM PZ DPIV activity, ρ<0.008 (Paired T-test) b= NEM TZ volume vs. NEM PZ volume, ρθ.0001 (Paired T-test) c = Cancer vs. NEM p<0.0001 (ANOVA), pO.OOOl (Wilcoxon) d = Cancer vs. NEM p<0.0353 (ANOVA), pθ.0601 (Wilcoxon) e = Cancer vs. NEM ρ<0.0363 (ANOVA), ρ<0.0258 (Wilcoxon)

Although the higher DPIV specific activity in the TZ biopsies would suggest that the TZ might secrete more DPIV than the PZ, there was no correlation of DPIV activities in EPS with specific activities in either the PZ or TZ biopsies (Table 3). In addition, among all patients as a collective group, secreted DPIV activity was also not associated with total volume of the prostate, individual volumes of the TZ or PZ, level of DPIV specific activity in either the PZ or TZ samples, nor age of the patients (Tables 2 and 3). Taken together, these results indicate that with respect to prostate function, DPIV in secretion arises from both the PZ and TZ, but that more DPIV may remain cell associated in the TZ.

Table 3. Correlation between variables.

*statistically different

DPIV activities in relation to prostate cancer

The mean and median activities of DPIV in EPS from men with prostate cancer were significantly higher than in men with NEM upon prostate biopsy (Table 2). When the mean and median values were both examined statistically using ANOVA and the Wilcoxon Rank Sum tests, respectively, cancer associated increases in DPIV activities were detected in the individual prostate zones. Patients with cancer had higher mean DPW activity values in biopsies of both the PZ and TZ as compared with the non-cancer group (Table 2). The difference in PZ DPIV activities was highly significant (pO.OOOl) using either ANOVA or Wilcoxon tests, and the DPIV activities in the TZ were also statistically different for the cancer versus NEM groups using ANOVA (p=0.035) and marginally significant upon Wilcoxon test analysis (p=0.060). These results indicate that DPIV activity is elevated in both the TZ and PZ in prostate cancer, with a greater magnitude of response in the PZ.

Correlations between the variables listed in Table 2, including DPIV levels in prostate tissues, prostatic secretions, and cancer volume, are presented in Table 3. There was a significant correlation between the volume of cancer (cVca) and DPIV activity in the PZ (p= 0.035).

DPIV activities in men with cancer of low, moderate, and high volume

The activities of DPIV were also evaluated with respect to the calculated volume of cancer (cVca) using the approach of D'Amico and Propert (Int. J. Radiat. Oncol. Biol. Phys., 35:273-279 (1996)). The cancer patients were grouped according to Olumi et al. (Urol., 56:273-277 (2000)) as to low, moderate, and high cVca (Table 4).

The correlation of cVca with total prostate and TZ volumes, and serum PSA levels, reflect the role of these components in calculating cVca. The DPIV activities in the PZ were 43% and TZ 38% greater in the moderate versus low cVca group, however, these differences in grouped data were not statistically different, perhaps due to the small number of patients in the higher cVca group. There was no significant correlation of secretory DP IV activity with cVca, but there was a significant correlation between DP IV activity in the PZ (p=0.3793, p=0.035) but not the TZ with cVca (Table 3). There was also a significant correlation of serum

PSA levels with DP TV activities in the PZ (p=0.38, ρ=0.05) but not the TZ. These data indicate that PZ DP IV activities are more selectively elevated in cancer.

Table 4. DPIV activities in secretions, PZ extracts, and TZ extracts from men with cancer of low, moderate, and high calculated volume of cancer cVca .

Comparison of DPIV activity and PSA levels as a biomarker for prostate cancer

In order to evaluate DPIV as a marker for distinguishing men with prostate cancer from those with no evidence of malignancy, an ROC (receiver operating characteristic) curve was calculated (Figure 1). The logistic regression data suggest that the optimal cutoff point is at 26.4 U/mL. This cutoff value gives a sensitivity of 0.765 and a specificity of 0.867 in detecting prostate cancer. The area under the ROC curve is 0.80, which contrasts markedly from 0.57 for serum PSA discrimination for the same patients. The positive correlation coefficient for the EPS DPIV level (β=0.0967, ρ=0.0118) yielded from the logistic model indicates that a higher DPIV value is more accurate in distinguishing the malignancy of the disease.

The results provided herein demonstrate that DPIV activities in prostatic secretions arise from both the PZ and TZ, but that a greater proportion of DPIV activities in secretion may come from the PZ since there was a negative correlation of TZ volume and DPIV activities in secretion. With the onset of cancer, DPIV activities increase in the PZ, the intra-prostatic site of origin of the majority of prostate cancers, and to a limited extent in the TZ (Table 2). The increases in PZ DPIV activities were reflected in increased activities in EPS. Increased DPIV activities in the PZ tissues of patients with biopsy evidence of cancer may reflect increased DPIV activities in the cancer tissue itself, or in normal or benign glands near the cancers in which DPIV activities are elevated. The increase in TZ DPIV activities in men with cancer may also reflect a field effect on expression of DPIV in benign glands from adjacent cancer in the PZ itself, or cancer invading from the PZ into the TZ. hi some cases, elevated TZ DPIV activities may reflect the presence of TZ cancers, which occur in about 30% of patients. Although serum PSA measurements and ultrasound imaging of the prostate have revolutionized detection of prostate cancer, overlapping PSA values between men with early, organ-confined adenocarcinoma and with BPH diminish the effectiveness of detecting early stage prostate cancer (Gretzer and Partin, Urol. Clin. N. Amer., 30:677 (2003) and Han et al, Med. Clin. N. Amer., 88:245 (2004)). hi addition to detection, treatment decision options for men who have serum PSA values between 4 to 20 ng/niL and a biopsy-determined Gleason score of between 5 and 7 need to be better defined since the aggressive nature of the cancer cannot be accurately predicted. Attempts at using PSA density (PSA/mL prostate) or PSA velocity (change in PSA over time) to refine prostate disease evaluation have had limited success.

As demonstrated herein, there was a statistically significant relationship of the ratio of PZ:TZ volume with cancer (also reflected in an increased PSA density). The increase in DPTV activities in EPS distinguishing men with cancer is reflected by increased activities in the PZ, but also by reduced activities proportional to increased TZ volume in NEM. However, the area of 0.80 under the ROC curve gives a strong indication for utilization of DPIV measurements in evaluating prostate disease, hi perspective, the area under the ROC curve for serum PSA in these patients was 0.57. Since DPIV is not specific to the prostate, increases in serum DPIV above a threshold level can be used to identify patients with prostate cancer.

A threshold level of serum DPIV can reflect levels of DPIV released in different tissues into the circulatory system of an individual. Threshold levels of DPIV can be established by analysis of collective data of men with no evidence of prostate malignancy. For an individual patient, a serum DPIV level determined before need to biopsy because of suspicion of prostate cancer can be used as a threshold.

A clinically practical approach to using DPIV measurements in evaluating prostate disease can involve using a post-prostate digital rectal examination urine sample containing EPS deposited into the urethra from the pressure placed on the gland during the examination. The measurement of DPIV together with markers that decrease in prostate cancer, e.g., neprilysin, tripeptidylpeptidase II, or aminopeptidase N, can provide ratios related to the presence of prostate cancer and its aggressive nature.

Ratio of DPIV and TPII as a marker for prostate cancer The activity levels of DPIV and TPII polypeptides were measured in BPH tissue and prostate cancer tissue. The homogenized tissues were examined by histology of frozen sections. Prostate cancer tissues were 70 percent or more cancer, and no detectable cancer was in the BPH tissues.

The level of TPII prostate cancer tissue was reduced as compared to the levels measured in BPH tissue (Table 5). The ability of the level of a DPIV polypeptide to detect early stage prostate cancer can be enhanced when expressed in a ratio with the level of a polypeptide that is down regulated in prostate cancer (e.g., a TPII polypeptide or a neprilysin polypeptide). For example, the ratio of DPΓV:TPII in BPH tissue was 5.8, but 67 in prostate cancer tissue.

Table 5. DPIV and TPII activities in extracts of BPH and prostate cancer tissues.

Prostate N DPIV (mU/mg protein) TPII (mU/mg protein)

BPH 8 167 ±23 (± SEM) 29.1 ±4.6 Cancer 7 355 ±62 (p<0.05) 5.3 ±3.1 (pO.Ol) OTHER EMBODIMENTS

It is to be understood that while the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

Claims

WHAT IS CLAIMED IS:

1. A method for assessing the likelihood of prostate cancer in a mammal, wherein the volume of said prostate cancer is 4.0 cm³ or less, said method comprising determining whether or not a prostate transition zone or prostate secretion sample from said mammal comprises an elevated level of a DPIV polypeptide as compared to the level of said DPIV polypeptide in a control sample, wherein the presence of said elevated level indicates that said mammal likely has said prostate cancer.

2. The method of claim 1 , wherein the volume of said prostate cancer is between 0.5 and 4.0 cm³.

3. The method of claim 1, wherein the volume of said prostate cancer is less than 0.5 cm³.

4. The method of claim 1, wherein said mammal is a human.

5. The method of claim 1, wherein said sample is a prostatic secretion sample.

6. The method of claim 1 , wherein said sample is a urine sample.

7. The method of claim 1, wherein said sample is a urine sample comprising the first 3 mL of urine from said mammal following a prostate digital rectal examination.

8. The method of claim 1, wherein said sample is a prostate transition zone sample.

9. The method of claim 1, wherein said sample is a prostate transition zone sample obtained via endoscopy.

10. The method of claim 1 , wherein the level of said DPIV polypeptide is determined using an immunoassay.

11. The method of claim 1 , wherein the level of said DPIV polypeptide is determined by measuring DPIV polypeptide activity.

12. The method of claim 1, wherein said control sample is from a prostate cancer-free mammal.

13. The method of claim 1 , wherein said sample is a prostatic secretion sample, and wherein said elevated level results in greater than 25 units of DPIV polypeptide activity per mL of said sample when the level of said DPIV polypeptide in said control sample results in between 15 and 22 units of DPIV polypeptide activity per mL of said control sample.

14. The method of claim 1, wherein said sample is a prostatic secretion sample, and wherein said elevated level results in greater than 30 units of DPIV polypeptide activity per mL of said sample when the level of said DPIV polypeptide in said control sample results in between 15 and 22 units of DPIV polypeptide activity per mL of said control sample.

15. The method of claim 1 , wherein said sample is a prostate transition zone sample, and wherein said elevated level results in greater than 0.17 units of DPIV polypeptide activity per mg protein in said sample when the level of said DPIV polypeptide in said control sample results in less than 0.17 units of DPIV polypeptide activity per mg protein in said control sample.

16. The method of claim 1, wherein said sample is a prostate transition zone sample, and wherein said elevated level results in greater than 0.21 units of DPIV polypeptide activity per mg protein in said sample when the level of said DPIV polypeptide in said control sample results in less than 0.17 units of DPIV polypeptide activity per mg protein in said control sample.

17. The method of claim 1 , wherein said method comprising determining whether or not the level of a neprilysin, tripeptidylpeptidase II, or aminopeptidase N polypeptide is reduced in said mammal as compared to the level of said neprilysin, tripeptidylpeptidase II, or aminopeptidase N polypeptide in a control mammal not having prostate cancer.

18. The method of claim 1 , wherein said method comprising determining whether or not said mammal comprises an elevated level of prostate specific antigen.

19. The method of claim 1, wherein said control sample is a sample from said mammal obtained earlier in life.

20. The method of claim 1, wherein said control sample is a sample from said mammal obtained when said mammal did not have prostate cancer.

21. A method for assessing the likelihood of prostate cancer in a mammal, said method comprising determining whether or not a prostate peripheral zone sample from said mammal comprises an elevated level of a DPIV polypeptide as compared to the level of said DPIV polypeptide in a control sample, wherein the presence of said elevated level indicates that said mammal likely has said prostate cancer.

22. The method of claim 21 , wherein the volume of said prostate cancer is between 0.5 and 4.0 cm³.

23. The method of claim 21 , wherein the volume of said prostate cancer is greater than 4.0 cm³.

24. The method of claim 21, wherein said mammal is a human.

25. The method of claim 21 , wherein said prostate peripheral zone sample was obtained via endoscopy.

26. The method of claim 21 , wherein the level of said DPIV polypeptide is determined using an immunoassay.

27. The method of claim 21, wherein the level of said DPIV polypeptide is determined by measuring DPIV polypeptide activity.

28. The method of claim 21, wherein said control sample is from a prostate cancer-free mammal.

29. The method of claim 21, wherein said elevated level results in greater than 0.09 units of DPIV polypeptide activity per mg protein in said sample when the level of said DPIV polypeptide in said control sample results in less than 0.09 units of DPrV polypeptide activity per mg protein in said control sample.

30. The method of claim 21 , wherein said elevated level results in greater than 0.12 units of DPIV polypeptide activity per mg protein in said sample when the level of said DPIV polypeptide in said control sample results in less than 0.09 units of DPrV polypeptide activity per mg protein in said control sample.

31. The method of claim 21 , wherein said method comprising determining whether or not the level of a neprilysin, tripeptidylpeptidase II, or aminopeptidase N polypeptide is reduced in said mammal as compared to the level of said neprilysin, tripeptidylpeptidase II, or aminopeptidase N polypeptide in a control mammal not having prostate cancer.

32. The method of claim 21, wherein said method comprising determining whether or not said mammal comprises an elevated level of prostate specific antigen.

33. The method of claim 21, wherein said control sample is a sample from said mammal obtained earlier in life.

34. The method of claim 21, wherein said control sample is a sample from said mammal obtained when said mammal did not have prostate cancer.

35. A method for monitoring prostate cancer progression in a mammal, said method comprising determining whether or not a sample from said mammal comprises an elevated level of a DPIV polypeptide as compared to the level of said DPIV polypeptide in a sample obtained from said mammal at an earlier point in time, wherein said elevated level indicates that said prostate cancer has progressed.

36. The method of claim 35, wherein said sample is a prostatic secretion sample.

37. The method of claim 35, wherein said sample is a urine sample.

38. The method of claim 35, wherein said sample is a urine sample comprising the first 3 mL of urine from said mammal following a prostate digital rectal examination.

39. The method of claim 35, wherein said sample is a prostate transition zone sample or a prostate peripheral zone sample.

40. The method of claim 35, wherein said sample is obtained via endoscopy.

41. A method for detecting prostate cancer regression in a mammal, said method comprising determining whether or not a prostate peripheral zone sample from said mammal comprises an reduced level of a DPIV polypeptide as compared to the level of said DPIV polypeptide in a control sample obtained from said mammal at an earlier point in time, wherein said reduced level indicates that said prostate cancer has regressed.

42. The method of claim 41 , wherein said sample is obtained via endoscopy.

43. A method for monitoring a response to prostate cancer treatment in a mammal having prostate cancer with a volume of 4.0 cm³ or less, said method comprising determining whether or not a sample obtained from said mammal after said treatment comprises a reduced level of a DPIV polypeptide as compared to the level of said DPIV polypeptide in a control sample obtained from said mammal at an earlier point in time, wherein said reduced level indicates that said prostate cancer treatment reduced the volume of prostate cancer in said mammal.

44. The method of claim 43, wherein said sample is a prostatic secretion sample.

45. The method of claim 43, wherein said sample is a urine sample.

46. The method of claim 43, wherein said sample is a urine sample comprising the first 3 mL of urine from said mammal following a prostate digital rectal examination.

47. The method of claim 43, wherein said sample is a prostate transition zone sample or a prostate peripheral zone sample.

48. The method of claim 43, wherein said sample was obtained via endoscopy.

49. The method of claim 43, wherein said prostate cancer treatment is a radiation therapy.

50. The method of claim 43, wherein said prostate cancer treatment is a hormone therapy.

51. A method for monitoring a response to prostate cancer treatment in a mammal having prostate cancer, said method comprising determining whether or not a prostate peripheral zone sample obtained from said mammal after said treatment comprises a reduced level of a DPIV polypeptide as compared to the level of said DPIV polypeptide in a control sample obtained from said mammal at an earlier point in time, wherein said reduced level indicates that said prostate cancer treatment reduced the volume of prostate cancer in said mammal.

52. A kit comprising an anti-DPIV polypeptide antibody and an antibody selected from the group consisting of an anti-prostate specific antigen antibody, an anti-neprilysin polypeptide antibody, an anti-tripeptidylpeptidase II polypeptide antibody, and an anti-aminopeptidase N polypeptide antibody.