Abstract
Simple SummaryOral cancer is often deadly and severely painful. Because this form of cancer pain cannot be adequately treated with current medications including opiates, new treatment approaches are needed. Cathepsin S, a lysosomal cysteine protease may play a role in oral cancer pain through a protease-activated receptor-2 (PAR2)-dependent mechanism. We undertook a series of experiments to define the role of cathepsin S in oral cancer pain. We compared cathepsin S activity in human oral cancer tumors versus patient-matched normal tissue; a human oral cancer cell line versus a benign dysplastic oral keratinocyte cell line; and in an orthotopic xenograft tongue cancer mouse model versus normal controls in mice. We localized cathepsin S in macrophages and carcinoma cells in human oral cancers. The injection of cathepsin S caused nociception in a mouse model while the injection of oral cancer cells in which the gene for cathepsin S is deleted generated less nociception. Our findings will lay the foundations for clinical trials of cathepsin S inhibitors for treating oral cancer pain.Oral squamous cell carcinoma (SCC) pain is more prevalent and severe than pain generated by any other form of cancer. We previously showed that protease-activated receptor-2 (PAR2) contributes to oral SCC pain. Cathepsin S is a lysosomal cysteine protease released during injury and disease that can activate PAR2. We report here a role for cathepsin S in PAR2-dependent cancer pain. We report that cathepsin S was more active in human oral SCC than matched normal tissue, and in an orthotopic xenograft tongue cancer model than normal tongue. The multiplex immunolocalization of cathepsin S in human oral cancers suggests that carcinoma and macrophages generate cathepsin S in the oral cancer microenvironment. After cheek or paw injection, cathepsin S evoked nociception in wild-type mice but not in mice lacking PAR2 in Nav1.8-positive neurons (Par2Nav1.8), nor in mice treated with LY3000328 or an endogenous cathepsin S inhibitor (cystatin C). The human oral SCC cell line (HSC-3) with homozygous deletion of the gene for cathepsin S (CTSS) with CRISPR/Cas9 provoked significantly less mechanical allodynia and thermal hyperalgesia, as did those treated with LY3000328, compared to the control cancer mice. Our results indicate that cathepsin S is activated in oral SCC, and that cathepsin S contributes to cancer pain through PAR2 on neurons.
Highlights
Cathepsin S is a lysosomal cysteine protease that degrades proteins along the endocytic pathway, including the invariant chain necessary for MHC class II antigen processing and presentation [1]
To determine whether cathepsin S is activated in oral squamous cell carcinoma (SCC), we collected oral SCC specimens and matched normal oral mucosa from seven patients (Table 1)
A BMV157-labeled species of 25 kDa was activated in oral SCC versus normal tissue (Figure 1a)
Summary
Cathepsin S is a lysosomal cysteine protease that degrades proteins along the endocytic pathway, including the invariant chain necessary for MHC class II antigen processing and presentation [1]. Of the 11 human cathepsins, most require an acidic environment, such as the interior of lysosomes, to be active. Because of the capacity of cathepsin S to function at a neutral pH, it is active in the extracellular environment, where it degrades extracellular matrix proteins. Inhibition as a therapy is limited by the wide tissue expression of proteases and resultant side effects; for this reason, treatment with matrix metalloproteinase inhibitors is highly limited [3]. Cathepsin S, on the other hand, has limited tissue expression, primarily in antigen presenting cells in the lymph and spleen [4,5]. Cathepsin S is an attractive target, and cathepsin S inhibitors have been tested in clinical trials (NCT 00425321, 01515358); the inhibitors exhibit good safety profiles [6]
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