Abstract A084: Cathepsin S Activity as a Biomarker for Early Detection of Pancreatic Cancer
Utsav Joshi, Heather Farris, Helene Gross, Nivedha Srikanth. Iyer, Manish Sampat, Timothy Pham, Jiayu Chen, Majid Ghassemian, Ida Deichaite, Pablo Tamayo, Karen Messer, Michael Heller, Geert Schmid-. Schonbein, Rebekah WhiteAbstract
Background:
The pancreas produces digestive proteases, which are often inappropriately activated in diseased states. High levels of digestive protease activity as well as secondary proteases, including cathepsins and metalloproteinases (MMPs), have been implicated in the pathophysiology of pancreatic ductal adenocarcinoma (PDAC). We hypothesize that elevated plasma protease activity can serve as a potential biomarker for early detection of PDAC.
Methods:
The Rapid Assay for Protease Detection (RAPD) assay is a novel assay developed at our institution that utilizes fluorescent charge-changing peptide substrates to produce a positively charged fluorescent product fragment upon cleavage by the target protease. The RAPD assay was used to analyze trypsin, chymotrypsin, cathepsin S (CTSS), and MMP-2 activity levels in treatment-naïve PDAC patients (N=35) and healthy (N=20) plasma samples. To further investigate specific proteases involved in PDAC, we compared human PDAC to control plasma using label-free quantitative peptidomics analysis of cleavage patterns. To explore mechanistic pathways upstream and downstream of these proteases, we examined de-identified bulk RNA-sequencing data, obtained through a commercial CLIA-certified assay, from 209 patients with PDAC from our institution.
Results:
Using the RAPD, we found that the activities of chymotrypsin, MMP2, and CTSS are significantly elevated in the plasma of PDAC patients compared to healthy subjects. A logistic regression model combining all 3 allowed perfect discrimination of PDAC from healthy controls in the training data, with cross-validated AUC of 0.96 (95% CI, 0.90 - 1.00). CTSS was identified as a lead biomarker because we were able to demonstrate higher activity in the small, combined group of stage 0 (carcinoma in situ) or stage I patients than in healthy subjects. CTSS-compatible peptides were more abundant than were trypsin- and chymotrypsin-compatible peptides overall, and we observed significantly elevated levels of CTSS compatible peptides in the plasma of PDAC patients compared to healthy subjects. Gene correlation assay displays many statistically significant correlations (p<0.05) between the CTSS gene and other genes involved in lipid metabolism, lysosomal activity, and autophagy. To determine the source of the plasma CTSS activity, we analyzed a publicly available dataset (www.proteinatlas.org) of single-cell RNA sequencing data from human PDAC patients. This data demonstrates that myeloid cells are not only one of the largest cell clusters but also the highest expressers of CTSS.
Conclusions:
In conclusion, this study provides compelling evidence for the importance of protease activity, particularly CTSS, in PDAC pathogenesis. We have developed a novel assay, the RAPD, that can rapidly and sensitively detect protease activity with extremely small (5-10 ml) sample requirement and potential for multiplex analysis of proteases as part of an early detection strategy for PDAC.
Citation Format:
Utsav Joshi, Heather Farris, Helene Gross, Nivedha Srikanth. Iyer, Manish Sampat, Timothy Pham, Jiayu Chen, Majid Ghassemian, Ida Deichaite, Pablo Tamayo, Karen Messer, Michael Heller, Geert Schmid-. Schonbein, Rebekah White. Cathepsin S Activity as a Biomarker for Early Detection of Pancreatic Cancer [abstract]. In: Proceedings of the AACR Conference on Pancreatic Cancer: New Frontiers in Biology and Therapeutic Development; 2026 Sep 25-28; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(18_Suppl_2):Abstract nr A084.