Original article
Semen AMACR protein as a novel method for detecting prostate cancer

https://doi.org/10.1016/j.urolonc.2018.09.010Get rights and content

Abstract

Background

Alpha methylacyl A coenzyme racemase (AMACR) has shown to be an excellent immunohistological biomarker for prostate cancer (CaP). Given the connection between prostate and urethra, we hypothesized that semen ejaculate would be an ideal specimen for detection of CaP specific biomarkers, such as AMACR. This study explores the detection of semen AMACR protein in men with and without CaP.

Methods

Semen ejaculates from 28 biopsy proven CaP patients prior to radical prostatectomy and 15 age-comparable controls were analyzed. An indirect sandwich ELISA chemiluminescence assay was used to detect semen AMACR, PSA, and Matriptase proteins. Tissue AMACR protein was quantified in 12 corresponding prostatectomy specimens using automated quantitative analysis (AQUA).

Results

Semen AMACR protein was detected in 23 of 28 (82%) CaP patients and 23 of 24 (96%) CaP patients with significant tumor volume (>0.5 cc or 0.3 g). Among the 5 cancer patients with undetectable semen AMACR, 4 patients had small tumor volumes (<1% or 0.3 g). Semen AMACR protein was also detected in 7 of 15 (47%) control noncancer patients. Using 76 ng/ml as a cutoff value, 20 of 28 (71%) patients and 20 of 24 (83%) patients with significant tumor volume were positive for semen AMACR protein, whereas only 5 of 15 (33%) age-comparable controls were positive. AMACR levels degrade with time.

Conclusions

This is the first study to demonstrate that AMACR protein is detectable in semen ejaculate. The higher AMACR levels detected in cancer patients suggests that semen AMACR protein may be useful as a noninvasive test for prostate cancer. Further validation is warranted.

Introduction

CaP is the most common age-related cancer for men in the United States [1]. Early CaP usually has no symptoms and is most commonly detected through screening tests such as the prostate specific antigen (PSA) blood test and digital rectal exam. Because PSA elevation can be due to benign processes such as hypertrophy and inflammation, it has only an approximately 30% specificity for detection of CaP [2]. While the current cutoff value for selecting patients for further work-up is a PSA >4.0 ng/ml, 25% to 30% of CaP are detected with PSAs between 2.5 and 4.0 ng/ml, and some of these cancers are clinically significant [3]. Men with persistently elevated PSAs are subject to prostate biopsy, a test with the potential for significant side effects and inaccuracies given 60% of annual biopsies are repeat biopsies after initial negative findings [4], [5], [6]. A noninvasive, sensitive and specific test to better identify patients at high risk for CaP, particularly clinically significant cancer (cancer with significant volume) is in urgent need.

Alpha methylacyl A coenzyme racemase (AMACR) is a peroxisomal and mitochondrial enzyme involved in the ß-oxidation of branched fatty acids. AMACR was identified as a CaP biomarker by molecular profiling [7]. It is shown to be an excellent immunohistological biomarker for CaP and high-grade prostate intraepithelial neoplasia with a sensitivity of 97% and specificity of 92%, whereas hyperplasia or normal prostate have low or undetectable AMACR [8], [9], [10]. AMACR immunohistochemical staining is commonly used in diagnosing small, morphologically ambiguous CaP [8], [10], [11], [12]. AMACR has also found to be moderately overexpressed in some other malignancies, such as colon cancer, papillary renal cell carcinoma, pancreatic carcinoma, nephrogenic adenoma, and dysplastic esophageal epithelium [9], [13], [14], [15], [16], [17]. Although blood AMACR has been reported undetectable, studies have shown that AMACR protein and mRNA can be detected from urine cellular component collected postprostate biopsy or massage [18], [19]. A humoral immune response against AMACR in CaP patients has also been reported recently [20].

Seminal ejaculate consists of sperm and secretions with 30% of the fluid originating from the prostate. Given CaP originates from ductal epithelial cells, we explored whether seminal AMACR is detectable in men with CaP and hypothesized that semen AMACR protein levels correlate with CaP volume.

Section snippets

Subjects

After institutional review board approval, prospectively 28 patients with biopsy proven CaP (53–71 yrs.) and 15 age-comparable controls (60–77 yrs., PSA < 2 ng/ml) were recruited at UW Hospital and Clinics for this study. Subsequently, 26 of the 28 cancer patients underwent a radical prostatectomy. The information obtained from all subjects included demographics, PSA test result, histological diagnosis of prostate biopsy or prostatectomy and stage of disease if applicable. Total tumor volume

Results

Separated seminal serum from 28 CaP patients and 15 age comparable controls with very low serum PSA levels (<2 ng/ml) were tested (Table 1). Gleason score of 7 (79%) and tumor stage T2 (82%) were most common. Semen samples with PSA <200 µg/ml or undetectable matriptase by ELISA were considered degraded and another sample from the patient was obtained.

Analysis of mean ± SD seminal fluid AMACR protein was 220.4 ± 196.4 ng/ml in patients with cancer and 109.5 ± 149.4 ng/ml in patients without

Discussion

New biomarkers for the detection of CaP are desperately needed. To date, little work has been performed on analyzing seminal fluid. The putative CaP biomarkers (beta HCG and GSTP1) can be detected in semen ejaculate [31], [32]. More recently, PCA3, a noncoding mRNA in prostate was found to be overexpressed in CaP cells and can be detected from shed CaP cells in prostatic fluid and urine. The reported sensitivity and specificity are 65% and 82%, for seminal fluid PCA3 test, 43% and 61% for urine

Conclusions

This is the first study to demonstrate that AMACR protein is detectable in semen ejaculate. The higher semen AMACR level detected in cancer patients with significant cancers suggests that semen AMACR protein may be useful as a noninvasive test. These findings need further validation in a larger prospective screening trial.

Declaration of conflict of interest

Funding source: University of Wisconsin Robert Draper Technology Fund (WH).

Conflict of Interest: DFJ and WH have a patent pending on AMACR filed by the Wisconsin Alumni Research Foundation (WARF). This technology has been licensed by Gregor Diagnostics. DFJ serves as a scientific advisor to Gregor Diagnostics.

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