Aberrant expression of KLK6 is a common feature for many human malignancies and numerous studies evaluated KLK6 as a prognostic biomarker (reviewed in [
3]). Consistent with most reports, we found high KLK6 expression in primary tumor samples of 42.6 % HNSCC patients. However, while induced KLK6 expression alone or in combination with other KLK family members was associated with poor progression-free and overall survival in colorectal cancer [
31,
32], gastric cancer [
33,
34], pancreatic ductal adenocarcinoma [
35], ovarian cancer [
36,
37], lung cancer [
38], and intracranial tumors [
39,
40], a high KLK6 expression pattern served as favorable prognostic biomarker in our OPSCC and LSCC patient cohorts. These data suggest a context-specific role of KLK6 in regulating the malignant progression and response to therapy, with both tumor promoting and tumor protective functions depending on the cellular origin of the tumor tissue. However, it is worth noting that the tumor promoting function of KLK6 in most human cancers has been deduced from the association of its expression and clinical or pathological features. But only few studies address the underlying molecular mechanisms
in vitro, and confirmations in preclinical model systems are missing.
So far, only two studies investigated the prognostic value of other KLKs in laryngeal cancer, while to the best of our knowledge no study was published on oropharyngeal SCCs. Both studies reported a remarkable down-regulation of either KLK4 or KLK11 transcript levels in laryngeal cancer as compared to their non-malignant counterparts [
41,
42]. Similar to our data on KLK6, patients with KLK11-positive tumors had a favorable prognosis [
41], and low KLK4 expression predicted short-term relapse and poor disease-free survival [
42]. Finally, down-regulation of KLK13 was reported in oral SCC cell lines and low KLK13 expression in primary oral cancer significantly correlated with regional lymph node metastasis [
43,
44].
A tumor protective role of KLK6 was demonstrated in breast cancer, where it was identified originally as a putative tumor suppressor due to its down-regulation during metastasis [
22]. Tumor-specific loss of KLK6 expression in breast cancer cells is mediated by epigenetic silencing initiated by hyper-methylation at the proximal promoter [
14]. It will be interesting to address whether a similar mode of regulation also occurs in primary HNSCC with low KLK6 expression, and in local recurrence or metastasis that develop in HNSCC patients with treatment failure.
Pampalakis and colleagues demonstrated that restoration of physiological KLK6 levels in breast cancer cell lines reverted the malignant phenotype
in vitro and
in vivo [
14]. They provided compelling evidence that KLK6 may act as suppressor for tumor progression by promoting a mesenchymal-to-epithelial transition [
14]. In line with our findings in HeLa cells, reactivation of KLK6 in breast cancer cells was associated with prominent down-regulation of Vimentin in the absence of a parallel rise in E-cadherin (unpublished data). These data suggest a common inhibitory function of KLK6 on Vimentin expression in breast and mucosal epithelial cells; however, the molecular mechanism remains to be fully elucidated. Moreover, silencing of KLK6 in a HNSCC cell line with prominent KLK6 expression resulted in an EMT-like phenotype with strong up-regulation of Vimentin, complete loss of E-cadherin and deregulation of other well-established molecular markers of EMT. On the cellular level loss of KLK6 was associated with accelerated migration and invasion as well as impaired response to irradiation, well-known characteristics of EMT and most likely responsible for the poor clinical outcome of HNSCC patients with low KLK6 expression. Our data are in contrast to recent findings in other tumor cells lines derived from skin and colon cancer in which KLK6 promotes tumor cell migration and invasion by either E-cadherin ectodomain shedding [
45], activation of protease-activated receptors [
8] or so far unknown molecular mechanisms [
46]. Although the principle nature for these cell type specific differences remains elusive, Pampalakis and colleagues claimed that KLK6 regulates EMT in breast cancer cells via the TGF-β pathway [
14,
47]. However, neither FaDu cells with silenced KLK6 expression nor HeLa cells with ectopic overexpression exhibited obvious changes in SMAD2/3 phosphorylation, questioning a major impact of KLK6 on canonical TGF-β signaling, at least in mucosal tumor cells. Intriguingly, our study demonstrated nuclear accumulation of β-catenin and activation of TCF-dependent gene expression upon KLK6 silencing in FaDu cells. Nuclear accumulation of β-catenin was also evident in KLK6
low tumor samples of HNSCC patients. Intracellular expression of β-catenin has been reported in the context of invasive growth and metastasis of oral carcinoma cells as well as poor prognosis [
48]. β-catenin is a component of the cell-cell adhesion complex and anchored to cadherin-related proteins, which sequester β-catenin at the cell periphery. Consequently, nuclear accumulation of β-catenin might result from down-regulation of E-cadherin during EMT after silencing of KLK6
in vitro or loss of E-cadherin expression in KLK6
low tumor cells during malignant progression of HNSCCs. In addition, β-catenin is also a key regulator of the Wnt signaling. Wnt/β-catenin is one of the crucial pathways in the maintenance of the self-renewal capacity of stem cells, and its inhibition is a promising approach to target cancer stem cells from HNSCC [
49,
50]. In contrast to our data, KLK6-induced nuclear translocation of β-catenin was demonstrated in mouse keratinocytes and human lung cancer cell lines
in vitro [
45,
51], further supporting a diverse and context dependent function of KLK6 in malignant progression.