Abstract 5116: Alpha-2-macroglobulin in the tumor microenvironment of pancreatic ductal adenocarcinoma to regulate versican proteolysis

Abstract Background: Versican (VCAN), a large chondroitin sulfate matrix proteoglycan is upregulated in the pancreatic ductal adenocarcinoma (PDAC) stroma and plays an immunosuppressive role. In contrast, the VCAN V1 isoform can be cleaved by ADAMTS proteases 1, 4, 5, 9, 15 and 20 into the fragment versikine which is known to be immunostimulatory. PDAC displays poor VCAN proteolysis and CD8+ T-cell infiltration, but interestingly overexpresses ADAMTS 1, 4, 5 and 9. Alpha-2-macroglobulin (A2M) has been shown to inhibit ADAMTS 4 and 5 and may be overexpressed in the PDAC tumor microenvironment (TME). This complex interplay is currently not well understood and is investigated here. Methods: The PDAC TCGA PanCancer Atlas dataset was used to compare relative levels of ADAMTS protease expression at the bulk RNA seq level. A TNM plot was used to compare levels of ADAMTS proteases between normal adjacent (NA) and tumor tissues. Patient-derived tissue microarrays (TMAs) (n=66) were used to correlate VCAN proteolysis, ADAMTS 4 and 5 as well as A2M expression. To explore the causative link between A2M and VCAN proteolysis, a major drawback of the well-studied Pan02 injectable model of PDAC is the lack of intense desmoplasia seen in the TME of human PDAC. To overcome this limitation, KPC tumor organoids and cancer-associated fibroblasts (1:10) were co-allografted into the flanks of B6 mice, followed by validation of cytokeratin-17/19 and alpha-smooth muscle actin expression using immunohistochemistry. Results: ADAMTS 1, 4 and 9 have similar relative levels of expression in PDAC; ADAMTS 5 and 15 slightly lower [median RSEM log2(value+1) = 10.2, 9.91, 9.54, 7.25, 5.33] and ADAMTS 20 is almost absent. ADAMTS 4, 5, 9 and 15 have significantly higher expression in PDAC tumor tissue versus (vs) normal adjacent (NA) pancreas tissue (log2FC= 2.16, 1.9, 1.57, 1.4, p-values <0.05). In patient-derived TMAs, PDAC tumors show higher levels of ADAMTS 4 expression vs. NA pancreas tissue (58.42% vs 44.12% of samples are ADAMTS 4hi). ADAMTS 5 expression is high in both tumor and NA tissue (90.5% vs 90%) which may be attributed to residual tumor tissue in the NA samples. Both ADAMTS 4 and 5 correlate with poor VCAN proteolysis. A2M RNA (log2FC= 2.09, p-value <0.05) and protein levels are significantly higher in tumor tissue vs. NA pancreas (67.41% vs 33.33% of samples are A2Mhi) and correlate with poor VCAN proteolysis. The murine co-allograft model resembles the human PDAC TME closely as evidenced by the pattern of cytokeratin-17/19 staining on epithelial cells and alpha-smooth muscle actin on cancer-associated fibroblasts. Conclusions: High A2M expression correlates with high ADAMTS 4/5 expression and poor VCAN proteolysis, indicating a potential for A2M to aid in VCAN accumulation in the TME. The co-allograft model will be used to explore the role of A2M further in the PDAC TME. Citation Format: Somak Chaudhuri, Yousef Gadalla, Katherine A. Johnson, Philip B. Emmerich, Sean G. Kraus, Cheri A. Pasch, Dustin A. Deming. Alpha-2-macroglobulin in the tumor microenvironment of pancreatic ductal adenocarcinoma to regulate versican proteolysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5116.