Protease-activated Receptor-2 (PAR-2)-mediated Nf-κB Activation Suppresses Inflammation-associated Tumor Suppressor MicroRNAs in Oral Squamous Cell Carcinoma

Abstract

Oral cancer is the sixth most common cause of death from cancer with an estimated 400,000 deaths worldwide and a low (50%) 5-year survival rate. The most common form of oral cancer is oral squamous cell carcinoma (OSCC). OSCC is highly inflammatory and invasive, and the degree of inflammation correlates with tumor aggressiveness. The G protein-coupled receptor protease-activated receptor-2 (PAR-2) plays a key role in inflammation. PAR-2 is activated via proteolytic cleavage by trypsin-like serine proteases, including kallikrein-5 (KLK5), or by treatment with activating peptides. PAR-2 activation induces G protein-␣-mediated signaling, mobilizing intracellular calcium and Nf-B signaling, leading to the increased expression of pro-inflammatory mRNAs. Little is known, however, about PAR-2 regulation of inflammation-related microRNAs. Here, we assess PAR-2 expression and function in OSCC cell lines and tissues. Stimulation of PAR-2 activates Nf-B signaling, resulting in RelA nuclear translocation and enhanced expression of pro-inflammatory mRNAs. Concomitantly, suppression of the anti-inflammatory tumor suppressor microRNAs let-7d, miR-23b, and miR-200c was observed following PAR-2 stimulation. Analysis of orthotopic oral tumors generated by cells with reduced KLK5 expression showed smaller, less aggressive lesions with reduced inflammatory infiltrate relative to tumors generated by KLK5-expressing control cells. Together, these data support a model wherein KLK5-mediated PAR-2 activation regulates the expression of inflammation-associated mRNAs and microRNAs, thereby modulating progression of oral tumors. Squamous cell carcinoma of the oral cavity (OSCC)2 is a highly inflammatory disease that ranks as the 6th most frequently diagnosed cancer worldwide, with a poor 5-year survival rate of about 50% (1). Early diagnosis of OSCC is challenging, predominantly because early oral cancers and premalignant lesions are often subtle and asymptomatic. Although many patients present for diagnosis with stage III or IV disease, premalignant lesions in the oral mucosa often precede invasive OSCC (2). Furthermore, unlike most solid tumors that are monoclonal in origin, multiple distinct foci of dysplastic cells may be present in the oral mucosa. These epithelial dysplasia are categorized as mild, moderate, severe, or carcinoma in situ based on the histologic abnormalities present in the oral epithelium (3). Studies show that ϳ12-36% of epithelial dysplasia progress to carcinoma (4, 5); however, current approaches do not enable accurate identification of premalignant lesions likely to undergo malignant transformation.The link between inflammation and cancer is now well established, and inflammatory mediators are present in the microenvironment of virtually all solid tumors (6 -10). Many studies have associated proteinase-activated receptor-2 (PAR-2) with both inflammation and tumor progression (11-15); however, the expression of PAR-2 in OSCC has not been evaluated. PAR-2 is a G protein-coupled receptor that is activated by trypsin-...

miR‐200c‐3p spreads invasive capacity in human oral squamous cell carcinoma microenvironment

“…Many previous reports have indicated that miR-200c attenuates cell invasion and metastasis by downregulating the master regulators of the epithelial-to-mesenchymal transition (EMT)-such as ZEB1 and ZEB2-in several cancers, 17,[41][42][43] including head and neck cancer. 44 In contrast, other studies propose that increased miR-200c expression is an indicator of malignancy, as it is often found to be overexpressed in the sera of cancer patients as compared with levels in healthy controls.…”

“…The majority of miRNA expression profiling studies have used either cancer cell lines or tissue samples; however, the resulting data were always controversial when discussing the role of individual miRNAs . Several factors are likely responsible for these inconsistencies, including methodological and statistical variation, as well as the presence of heterogeneous expression patterns in OSCC cell lines and tumor tissue samples …”

“…7 Consequentially, aberrant miRNA expression is common in various cancers, and miRNA signatures have been helpful as diagnostic, prognostic, and therapeutic biomarkers for cancers, including OSCC. 8 The majority of miRNA expression profiling studies have used either cancer cell lines 9,10 or tissue samples 4,[11][12][13][14][15][16][17][18] ; however, the resulting data were always controversial when discussing the role of individual miRNAs. 4 Several factors are likely responsible for these inconsistencies, including methodological and statistical variation, as well as the presence of heterogeneous expression patterns in OSCC cell lines and tumor tissue samples.…”

Proteases: Pivot Points in Functional Proteomics

“…Epigenetic silencing of the HTRA1 gene in cancer cells may be caused by histone deacetylase targeting of the promoter, or transcription repression of the methylated promoter by binding of methyl-CpG-binding domain protein 2 (MBD2) [138]. Silencing of tumor suppressor microRNAs via protease-activated PAR and Nf-κB signaling, and of caspase-8 expression by DNA methylation are yet a few other epigenetic mechanisms associated with the development of certain cancers [139,131]. …”

“…Many other kallikreins may also be suitable cancer biomarkers. hK5 proteolytically activates PAR-2, leading to Nf-κB activation and downregulation of tumor suppressor microRNAs in oral squamous cell carcinoma [139]. Plasma kallikrein is capable of activating the complement system, linking inflammatory responses to many cancer-related processes [151].…”