Abstract 4286: Proteolytic degradation of BIGH3 can be quantified non-invasively in serum with biomarker potential for patients with non-small cell lung cancer
Authors: Pedersen Rasmus S. , Karsdal Morten , Willumsen Nicholas 
Published: March 2024

Abstract
Introduction: Transforming growth factor beta induced protein ig-h3 (BIGH3/TGFBI) is expressed widely by multiple cell types, including macrophages and fibroblasts, and is participating in various biological processes including adhesion, migration, angiogenesis and wound healing. BIGH3 is known to bind multiple collagens and are often embedded in the matrix where it seems to function as a linker between ECM and cell surfaces. The increased proteolytic activity found in non-small cell lung cancer (NSCLC) may degrade BIGH3 that affect this interaction as well as generate small peptide fragments that can serve as novel non-invasive biomarker targets if released to circulation. The aim of this study is to develop a tool to quantify degraded BIGH3 non-invasively and explore its potential as a biomarker in NSCLC.
Methods: An ELISA, named nordicBIGH3M-N targeting a cleaved fragment of BIGH3 was developed to reflect BIGH3 degradation and enable serological quantification. We incubated recombinant BIGH3 with and without collagenase to confirm that nordicBIGH3M-N only measured degraded BIGH3. In addition, we generated matrixes from normal fibroblasts and cancer associated fibroblasts (CAFs) grown with or without tgf-b. These matrixes were cleaved with collagenase to confirm that NordicBIGH3M-N could be generated from degradation of matrix. nordicBIGH3M-N was also measured in serum from 39 patients with NSCLC (18 patients with adenocarcinoma and 21 patients with squamous cell carcinoma) and 35 healthy controls. BIGH3M-N levels in serum from patients with the two subtypes of NSCLC and healthy controls was compared using Dunn’s multiple comparison test and area under receiver operation characteristic curves (AUROC).
Results: BIGH3M-N was only detectable after incubating recombinant BIGH3 and fibroblasts matrices with collagenase. A 7-fold increase of BIGH3M-N levels was seen between the collagenase degraded matrix from tgf-b treated fibroblasts compared to the degraded matrix from untreated fibroblasts. No difference in BIGH3M-N levels was observed between degraded matrix from tgf-b treated or untreated CAFs. BIGH3M-N was significantly elevated in patients with squamous cell carcinoma compared to healthy controls (p = 0.006, AUROC = 0.73) and patients with adenocarcinoma (p = 0.022, AUROC = 0.78). However, there was no significant difference between BIGH3M-N levels in serum from patients with adenocarcinoma and healthy controls (p&gt;0.99, AUROC = 0.51).
Conclusion: Degradation of BIGH3 can be reflected by non-invasive quantification of the cleaved fragment of BIGH3, BIGH3M-N, in serum. BIGH3M-N is a promising biomarker in NSCLC with potential for discriminating between subtypes. Moreover, BIGH3M-N is connected to fibroblast matrix biology so the optimal use for this biomarker might be in combination with other ECM biomarkers.
Citation Format: Rasmus S. Pedersen, Morten Karsdal, Nicholas Willumsen. Proteolytic degradation of BIGH3 can be quantified non-invasively in serum with biomarker potential for patients with non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4286.