Core 2 β-1, 6-N-acetylglucosaminyltransferase-1 expression in prostate biopsy specimen is an indicator of prostate cancer aggressiveness

https://doi.org/10.1016/j.bbrc.2016.01.011Get rights and content

Highlights

  • GCNT1 expression in biopsy specimen could predict prostate cancer recurrence.

  • GCNT1 expressing tumor forms large tumor with in vivo experiment.

  • GCNT1 expression is closely rerated with prostate cancer aggressiveness.

Abstract

Introduction

To avoid over-treatment of early stage prostate cancer (PCa), predictive biomarkers for PCa aggressiveness which can be obtained during pre-treatment evaluation are essential. Core 2 β-1, 6-N-acetylglucosaminyl-transferase-1 (GCNT1) is a key enzyme that forms core 2 branched O-glycans, the expression of which is associated with aggressive potential of prostate cancer. We examined whether GCNT1 expression in prostate biopsy specimen can predict cancer recurrence after radical prostatectomy for the patients with with PCa. We then investigated molecular background for aggressive malignant potential mediated by GCNT1 expression.

Methods

Paraffin-embedded PCa biopsy specimens were immunohisto-chemically tested for GCNT1 expression using an anti-GCNT1 monoclonal antibody. We also examined the role of GCNT1 in PCa progression using cell lines which express high or low levels of GCNT1.

Results

GCNT1 expression correlated with D′ Amico's recurrence risk classification. The GCNT1-positive rate in organ confined PCa was significantly lower than that in PCa with extra-prostatic extension. GCNT1-negative tumors were associated with significantly better prostate-specific antigen (PSA)-free survival compared with GCNT1-positive tumors. Multivariate analysis revealed that GCNT1 expression status was an independent risk factor for PSA recurrence after radical prostatectomy. Subsequent basic study revealed that GCNT1-over-expressing cells produced a significantly larger amount of growth factors when co-cultured with prostate stromal cells compared with GCNT1-knocked down cells and formed larger tumors.

Conclusions

GCNT1 expression in prostate biopsy specimen is a significant and independent predictor of recurrence after radical prostatectomy, which can be used in pre-treatment decision making for the patient. Further validation study is necessary to establish clinical implication of GCNT1 in management of PCa.

Introduction

In the western countries, prostate cancer (PCa) is the most common malignancy in men and the second-leading cause of cancer-related death [1], [2]. Its incidence is rapidly increasing in the Asia–Pacific region [3]. One of the most critical issues related to PCa in clinical practice is over-diagnosis and over-treatment [4]. Over-treatment of indolent PCa with a low malignant potential is a major issue because aggressive treatment of PCa is sometimes associated with adverse events. A promising alternate modality to prevent overtreatment may be active surveillance [5]; however, the identification of suitable patients for aggressive treatment is associated with difficulties. An appropriate tool for patient selection for active surveillance is still lacking. Therefore, development of novel biomarkers of PCa aggressiveness is of vital importance for the prevention of PCa over-treatment.

Preoperative serum PSA levels and biopsy Gleason score are conventional and powerful predictors of biological outcomes after radical prostatectomy [6], [7]. To improve the risk stratification for PCa recurrence after primary treatment in patients with localized PCa, many investigators have sought biomarkers that reflect the aggressive potential of PCa [8]. However, the majority of reported biomarkers have not been validated for providing information that is more useful than that provided by conventional clinicopathological parameters. With a novel biomarker representing the malignant potential of PCa, more accurate prediction of PSA recurrence and appropriate treatment selection may be possible.

Core 2 β-1, 6-N-acetylglucosaminyltransferase-1 (GCNT1) [9], [10] is a key enzyme that synthesizes core 2 branched O-glycans by catalyzing the transfer of N-acetylglucosamine (GlcNAc) from uridine diphosphate (UDP)-GlcNAc with a β1, 6-linkage to α-GalNAc of a core 1 O-glycan (Fig. 1A). Previously, it was shown that the expression of core 2 branched O-glycans is closely correlated with the malignant potential of colorectal cancer [11] and pulmonary adenocarcinoma [12]. Using a polyclonal antibody [10], we already have demonstrated that immunohistochemical testing of GCNT1 was closely related with the aggressive potential of PCa [13], testicular cancer [14], and bladder cancer [15]. However, polyclonal antibody has a weakness in specificity and reproducibility as a clinically useful biomarker. We then established a monoclonal antibody against GCNT1 by peptide immunization [16]. The monoclonal anti-GCNT1 successfully detected GCNT1 expression in paraffin-embedded specimen obtained by radical prostatectomy and post-digital rectal examination urine samples. Although the GCNT1 status were closely related with the aggressive potential of PCa in clinical samples [16], the mechanism of tumor progression still remained unclear. Furthermore, to develop a novel indicator of cancer aggressiveness it is essential to determine the malignant potential before the initiation of treatment for PCa. Therefore, it is very important to test whether GCNT-1 expression in prostate biopsy specimen can predict recurrence after radical prostatectomy.

In this study, we attempted to demonstrate that GCNT1 expression in prostate biopsy specimens was a significant and independent predictor for PSA recurrence after radical prostatectomy. Furthermore, we investigated possible roles of GCNT1 in PCa progression using cell lines which express high or low levels of GCNT1 with in vitro and in vivo experimental systems.

Section snippets

Cells

The human androgen-dependent PCa cell line LNCaP and human androgen-independent cell lines DU145 and PC-3 were purchased from the American Type Culture Collection (Rockville, MD, USA). Human prostate stromal cells (PrSCs) were purchased from the Cell Scientific Laboratory (Tokyo, Japan).

Stable transfection

LNCaP cells and DU145 cells were maintained in RPMI1640 medium supplemented with 10% FBS. The cells were plated in a 35-mm cell culture dish 24 h before transfection. The cells were transfected using the X-treme

GCNT1 expression in biopsy specimen positively correlated with extra-prostatic capsule extension and PSA recurrence

To evaluate the role of GCNT1 in PCa agressivenes, PCa biopsy specimens were immunohistochemically analyzed using the anti-human GCNT1 mAb. The results demonstrated that GCNT1 was barely expressed in normal prostate gland, whereas PCa cells often expressed significant levels of GCNT1 (Fig. 2A). The GCNT1-positive rate significantly elevated according to the D′ Amico's recurrence risk classification [19]. In the pathological parameters, extra-prostatic extension (pT3/4) and lymph node metastasis

Discussion

Aberrant glycosylation of cell surface glycoproteins plays an important role in cancer initiation, proliferation, invasion, and metastasis [23], [24], [25]. Biosynthesis of oligosaccharides on glycoproteins is performed in concert by several glycosyltransferases; GCNT1 is one of the glycosyltransferases that forms the core 2 O-glycans on the surface of lymphocytes and various cancer cells [13], [14], [26], [27]. It is noteworthy that the present study clearly demonstrated that

Conflict of interest

The authors have declared that no conflict of interest exists.

Acknowledgments

The authors thank Drs. Minoru Fukuda and Shigeru Tsuboi for useful discussions. The authors also thank Ms. Sayaka Yamada and Ms. Yukie Nishizawa for technical supports. This work was supported by JSPS KAKENHI (Grant Numbers 24791631 and 15K10569 to YT and 15H02563 to CO), and the Suzuki Urological Research Foundation (to YT).

References (33)

  • R. Siegel et al.

    Cancer statistics, 2013

    CA Cancer J. Clin.

    (2013)
  • L. Klotz

    Prostate cancer overdiagnosis and overtreatment

    Curr. Opin. Endocrinol. Diabetes Obes.

    (2013)
  • J.I. Epstein et al.

    Prediction of progression following radical prostatectomy. A multivariate analysis of 721 men with long-term follow-up

    Am. J. Surg. Pathol.

    (1996)
  • M.W. Kattan et al.

    A preoperative nomogram for disease recurrence following radical prostatectomy for prostate cancer

    J. Natl. Cancer Inst.

    (1998)
  • M.F. Bierhuizen et al.

    Expression cloning of a cDNA encoding UDP-GlcNAc:Gal beta 1-3-GalNAc-R (GlcNAc to GalNAc) beta 1-6GlcNAc transferase by gene transfer into CHO cells expressing polyoma large tumor antigen

    Proc. Natl. Acad Sci. U. S. A.

    (1992)
  • K. Shimodaira et al.

    Carcinoma-associated expression of core 2 beta-1,6-N-acetylglucosaminyltransferase gene in human colorectal cancer: role of O-glycans in tumor progression

    Cancer Res.

    (1997)
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