Abstract 7307: Potential role of alpha-2-macroglobulin in versican proteolysis regulation in colorectal cancer

Abstract Background: Versican is a large extracellular glycoprotein that acts as a barrier for immune cell infiltration in colorectal cancer (CRC). The V1 isoform of versican can be cleaved by ADAMTS 4 and 5 into the bioactive fragment versikine with immune stimulatory properties. Despite overexpression of ADAMTS 4 and 5 in CRC, versican may be poorly proteolyzed. ADAMTS 4 and 5 may be inhibited by the presence of an extracellular endopeptidase inhibitor, alpha 2 macroglobulin (A2M). Here we investigate A2M expression in CRC, its correlation with versican proteolysis status, and immune cell infiltration. Methods: The Cancer Genome Atlas (TCGA) Colorectal Adenocarcinoma PanCancer Atlas dataset was used to stratify CRC patients (n=594) into two cohorts: A2Mhi (n=57) (mRNA expression Z-score >1) and A2Mlo (n=341) (Z-score <0). Clinical characteristics and differences in gene expression were analyzed between the two groups. CRC patient-derived tumor samples (n=306) were analyzed for protein-level expression of versican (VCAN), versikine (Vkine), A2M, CD8A and CD4 via immunohistochemistry (IHC). A binning scale of 0 - 3 was used to assess staining intensity with 0/1 being low and 2/3 being high expression. The number of CD8A and CD4-positive lymphocytes in direct contact with tumors (tumor-infiltrating lymphocytes; TILs) were counted per high power field (HPF). Th1 (T helper 1 cell) and Treg (T regulatory cell) subsets of CD4+ TILs were further evaluated per 20X field of view (FOV) via dual immunofluorescence. Results: A2Mhiand A2Mlogroups have no differences in disease staging (q=.109), sex (q =0.992) or age distribution (q= 0.368). The A2Mhigroup has higher mRNA expression of VCAN (log2fold change = 2.40, q <.01), ADAMTS1 (2.3, q<.01), ADAMTS4 (1.74, q <.01), ADAMTS5 (1.47, q<.01), CD8A (1.48, q<.01), CD4 (1.91, q<.01), CXCL9 (1.88, q<.01), and CXCL10 (1.32, q<.01). Out of 306 patient-derived tumor samples, 82.9% of versican proteolytic prominent (VPP) (VCANLoVkineHi ) were A2MHi, 76.8% of versican proteolytic weak (VPW) (VCANHiand VkineLoor VkineHi) were A2MHi, and 85.1% of versican versikine low (VVL) were A2MHi. There is a trend towards significant interaction between A2M, versican and Th1 cells (p=0.0531), but none exist between A2M, versican and CD8+ T-cells/ Tregs. VPP and A2MHisamples have significantly higher median Th1s/FOV than VPW and A2MHisamples (VPP and A2MHimedian: 24.5 Th1s/FOV, VPW and A2MHi median: 9, p = .009). Conclusion: The expression of A2M can regulate ADAMTS protease function but does not appear to prevent VCAN proteolysis or impact T cell infiltration in CRCs. Citation Format: Yousef H. Gadalla, Somak Chaudhuri, Sean G. Kraus, Anna M. Field, Katherine A. Johnson, Cheri A. Pasch, Dustin A. Deming. Potential role of alpha-2-macroglobulin in versican proteolysis regulation in colorectal cancer [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 7307.