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Clinical relevance of the deregulated kallikrein-related peptidase 8 mRNA expression in breast cancer: a novel independent indicator of disease-free survival

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Abstract

Examining for new BC biomarkers has proven that kallikrein-related peptidase (KLK) family members represent promising serum and/or tissue molecular tools for early diagnosis, effective prognosis, and treatment monitoring of patients. The aim of this study was to investigate, the previously unexplored, prognostic significance of KLK8 in BC. KLK8 mRNA expression was quantitatively analyzed in 150 cancerous and 100 corresponding normal breast tissue specimens via a SYBR Green-based Real-Time PCR methodology. Expression data and patients’ clinicopathological parameters were used for extensive biostatistical analyses, including internal validation. KLK8 mRNA expression was significantly downregulated in the cancerous tissue part relative to the non-cancerous counterpart (P < 0.001), in the majority of the paired breast tissue samples. KLK8 expression was associated with advanced TNM stage (P = 0.019) and positive nodal status involvement (P = 0.044). Triple negative (TNBC) and HER2 overexpressing tumors exhibited higher KLK8 expression levels (P < 0.001), compared to Luminal A and B molecular subtypes. Kaplan–Meier survival curve analysis revealed that BC patients with high KLK8 expression had significantly shorter disease-free survival (DFS) intervals (P < 0.001) compared to those belonging in the KLK8-low expression group. Cox univariate analysis confirmed the association between KLK8 expression, analyzed as a continuous variable, and poor patients’ outcome (Hazard ratio [HR] = 3.28, P < 0.001). Most importantly, multivariate analysis showed that KLK8 expression is a strong and independent predictor of adverse DFS in BC ([HR] = 2.74; P = 0.002). Our results show that KLK8 mRNA expression is associated with aggressive tumor characteristics and it can serve as a novel independent biomarker of unfavorable prognosis for BC patients.

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Abbreviations

AUC:

Area under the curve

BC:

Breast cancer

CI:

Confidence interval

Ct:

Threshold cycle

DFS:

Disease-free survival

ECM:

Extracellular matrix

EMT:

Epithelial to mesenchymal transition

ER:

Estrogen receptor

HER2:

Human epidermal growth factor receptor 2

HPRT1 :

Hypoxanthine phosphoribosyltransferase 1

IHC:

Immunohistochemistry

KLK:

Kallikrein-related peptidase

MMPs:

Matrix metalloproteinases

PR:

Progesterone receptor

ROC:

Receiver operating characteristic

RQ units:

Relative quantification units

RT-qPCR:

Quantitative real-time PCR

TNBC:

Triple negative breast cancer

TNM:

Tumor-node-metastasis

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Acknowledgments

This research has been co-financed by the European Union (European Social Fund—ESF) and Greek national funds through the Operational Program “Education and Lifelong Learning” of the National Strategic Reference Framework (NSRF)—Research Funding Program: THALES–UoA–BIOPROMO, MIS 377046.

Competing interests

The authors declare that they have no competing interests.

Ethical standards

All research procedures that took place during the completion of our study comply with the ethical standards of the 1975 Declaration of Helsinki, as revised in 2008, and were approved by the institutional review board of ‘Saint Savvas’ Anticancer Hospital. Additionally, written informed consent was obtained from all BC patients participating in the study.

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, University of Athens, Panepistimiopolis, 15701, Athens, Greece

    Kleita Michaelidou & Andreas Scorilas

  2. First Department of Oncology, St. Savvas Anticancer Hospital, 171, Alexandras Avenue, 11522, Athens, Greece

    Alexandros Ardavanis

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  1. Kleita Michaelidou
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  2. Alexandros Ardavanis
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  3. Andreas Scorilas
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Correspondence to Andreas Scorilas.

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10549_2015_3470_MOESM1_ESM.tif

Supplementary Fig. 1 Relative quantification of KLK8 mRNA expression via RT-qPCR. Dissociation curves of HPRT1 (A) and KLK8 (B) amplicon, along with corresponding 3.0 % agarose gel electrophoresis of the RT-qPCR products of randomly selected breast tissue samples (C). Standard curves for HPRT1 and KLK8, constructed using serial dilutions of calibrator cDNA, covering several orders of magnitude (D). M: molecular weight marker; PC: positive control, NC: negative control

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Michaelidou, K., Ardavanis, A. & Scorilas, A. Clinical relevance of the deregulated kallikrein-related peptidase 8 mRNA expression in breast cancer: a novel independent indicator of disease-free survival. Breast Cancer Res Treat 152, 323–336 (2015). https://doi.org/10.1007/s10549-015-3470-8

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