Abstract 2579: Integrated sequencing analysis reveals prognostic thyroid cancer-associated fibroblast subpopulations

Abstract Thyroid cancers exist on a continuum of differentiation. Our group recently defined dynamic changes in the tumor microenvironment co-occurring with tumor histotypes and de-differentiation. Here, we integrate 423, 733 cells from seven distinct single-cell RNA-sequencing cohorts to define unique thyroid cancer- specific fibroblast subtypes occurring across the histologic and mutational spectrum of thyroid cancer. We identify distinct inflammatory, perivascular-like, and myofibroblastic cancer-associated (myCAF) populations. The myCAF population, specifically, is only found in malignant samples, and mapping myCAFs to three thyroid cancer bulk RNA-sequencing cohorts reveals an association with disease progression, with hazard ratio of 3.0 (95% CI, 1.7-5.3). Within this myCAF population, we identify subpopulations with immune-regulating phenotypes that are specifically associated with de-differentiated tumors. Additionally, we identify an invasive tumor cell phenotype in both differentiated and de-differentiated tumors marked by TNC and LAMB3. Ligand-receptor interaction analysis indicates robust interaction of myCAFs and invasive tumor cells through TGF-beta, tenascin, and collagen signaling pathways. Multiplex immunofluorescence, immunohistochemistry, and spatial transcriptomics analysis of 28 tumors (13 differentiated, 8 de-differentiated, 7 with areas of differentiated and de-differentiated histopathology) confirms robust upregulation of myCAFs in de-differentiated tumors. Further, myCAFs spatially localize next to the leading edge of de-differentiated tumors and near invasive cells from differentiated tumors, supporting a tumor-promoting role for myCAFs in aggressive thyroid carcinomas. In summary, our study provides an integrated atlas of thyroid cancer fibroblast subtypes, spatial analysis on patient samples, and an association of fibroblasts with disease progression. Citation Format: Matthew A. Loberg, George J. Xu, Claudia C. Wahoski, Kailey P. Caroland, Megan L. Tigue, Sheau-Chiann Chen, Hua-Chang Chen, Quanhu Sheng, Eric C. Huang, Fei Ye, Ryan H. Belcher, Ethan Lee, Vivian L. Weiss. Integrated sequencing analysis reveals prognostic thyroid cancer-associated fibroblast subpopulations [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 2579.