Clinical and Experimental Medicine

, Volume 18, Issue 2, pp 203–213 | Cite as

Kallikrein-related peptidase 6 (KLK6) expression differentiates tumor subtypes and predicts clinical outcome in breast cancer patients

  • Christoforos Haritos
  • Kleita Michaelidou
  • Konstantinos Mavridis
  • Ioannis Missitzis
  • Alexandros Ardavanis
  • John Griniatsos
  • Andreas Scorilas
Original Article


Novel molecular markers that address the heterogeneity of breast cancer (BC) and provide meaningful prognostic information for BC patients are needed. Kallikrein-related peptidase 6 (KLK6) is aberrantly expressed and functionally implicated in BC and, like other members of the KLK family, may prove a useful molecular tool for clinical management. Our objective was to assess, for the first time, the clinical relevance of KLK6 mRNA expression in BC. Total RNA was isolated from 165 breast tumors, as well as 100 adjacent non-cancerous tumor specimens. After cDNA synthesis, and following quality control, quantitative real-time PCR for KLK6 expression analysis took place. Receiver operating characteristic curves were constructed in order to assess the ability of KLK6 mRNA expression levels to differentiate between molecular BC subtypes. Survival analyses, using DFS as endpoint, were performed at the univariate and multivariate levels. Publicly available BC databases and online survival analysis tools were used to validate our findings. A significant downregulation of KLK6 mRNA expression was observed in BC tissue sections compared to the non-cancerous component (P < 0.001). The expression of KLK6 is positively associated with tumor grade (P = 0.038) and is overexpressed in TNBC and HER2-positive tumors (P < 0.001). Aberrant KLK6 expression predicts the clinical outcome of BC patients in terms of DFS, independently of currently used prognostic markers (HR = 7.11, 95% CI = 1.19–42.45). The differential expression of KLK6 and its association with unfavorable outcome in BC patients was validated via in silico analyses. Although an independent external cohort is necessary to confirm our findings, we proved for the first time that KLK6 can provide independent prognostic information for BC patients.


KLK6 Kallikreins KLKs Serine proteases Biological tumor marker Prognostic biomarker 



Area under the curve


Breast cancer


Confidence interval


Threshold cycle


Disease-free survival


Distant metastasis-free survival


Extracellular matrix


Epidermal growth factor receptor


Epithelial to mesenchymal transition


Estrogen receptor


Human epidermal growth factor receptor 2


Hypoxanthine phosphoribosyltransferase 1




Kallikrein-related peptidase


Protease-activated receptors


Progesterone receptor


Receiver operating characteristic

RQ units

Relative quantification units


Spearman correlation coefficient


Robust single sample predictor classification


Quantitative real-time PCR


Single sample predictors


Triple-negative breast cancer




Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards.

Informed consent

Informed consent was obtained from all individual participants included in the study.

Supplementary material

10238_2018_487_MOESM1_ESM.tif (9.5 mb)
Quality control of the developed qPCR assay for quantification of KLK6 expression. Dissociation curves of (A) HPRT1 and (B) KLK6 amplicons. (C) Corresponding 3.0% w/v agarose gel electrophoresis of the RT-qPCR products of randomly selected breast tissue samples. (D) Standard curves for HPRT1 and KLK6, constructed using serial dilutions of calibrator cDNA, covering several orders of magnitude. M: molecular weight marker; PC: positive control, NC: negative control. (TIFF 9682 kb)
10238_2018_487_MOESM2_ESM.doc (54 kb)
Supplementary material 2 (DOC 53 kb)


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Copyright information

© Springer International Publishing AG, part of Springer Nature 2018

Authors and Affiliations

  1. 1.The Breast ClinicSaint Savvas Anticancer HospitalAthensGreece
  2. 2.Department of Biochemistry and Molecular BiologyNational and Kapodistrian University of AthensAthensGreece
  3. 3.First Department of OncologySaint Savvas Anticancer HospitalAthensGreece
  4. 4.First Department of Surgery, Medical SchoolNational and Kapodistrian University of Athens, Laiko HospitalAthensGreece

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