| Popis: |
The tumor microenvironment is a complex milieu relevant to the progression of cancer. A detailed analysis needs to be performed for each cancer type to identify relevant cell types and secreted factors promoting metastasis. Fibrolamellar carcinoma (FLC) is an aggressive liver cancer that predominantly afflicts adolescents and young adults who lack predisposing conditions, and patients with FLC are characterized by a heterozygous deletion on chromosome 19 that creates an oncogenic gene fusion, DNAJB1-PRKACA. The extracellular environment of FLC tumors is poorly characterized and may contribute to cancer progression, metastasis, and/or drug resistance. To bridge this knowledge gap, we performed experiments to assess pathways relevant to proteoglycans, a major component of the extracellular matrix. We first analyzed gene expression data from FLC and non-malignant liver tissue (n=27) to identify changes in pathways pertinent to glycosaminoglycan (GAG) biosynthesis. We then developed and implemented a novel chemical analytics method, utilizing custom molecular standards, to quantify the abundance of different types of GAGs in patient tumor samples by liquid chromatography with tandem mass spectrometry. We then measured the mRNA and protein levels of different GAG-associated proteins, followed by immunohistochemical confirmation. Finally, we performed the first single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) on FLC tumors to define the heterogeneous cellular landscape and to identify cell types likely responsible for producing the most dominant GAG-associated protein in FLC. We found that chondroitin sulfate (CS) but not heparan sulfate (HS) or hyaluronic acid (HA) biosynthesis genes are dramatically increased in FLC, especially the rate-limiting enzyme CS GalNAc transferase 1 (CSGALNACT1). We also determined that CS chains are significantly elevated in FLC. Further analysis showed that versican (VCAN) is the primary CS-associated protein in FLC, that CSGALNACT1 and VCAN are correlated with DNAJB1-PRKACA levels, and that VCAN is more highly expressed in FLC compared to almost all other cancer types. Finally, the single cell ATAC-seq analyses demonstrated for the first time the cellular heterogeneity of FLC and revealed that the VCAN locus is active in tumor epithelial cells but is most active in proliferating stellate cells. This study merges chemical, molecular, and single-cell analyses to uncover the marked elevation of chondroitin sulfate proteoglycans in FLC tumors. Our results suggest that VCAN may play a critical role in FLC tumor growth and/or metastasis. We also identified the presence of activated stellate cells in FLC tumors and suggest a role in propagating fibrosis. Future studies are required to identify whether and how attenuation of CS and/or VCAN mitigates proliferative and/or metastatic potential in FLC. Citation Format: Adam Francisco, Jine Li, Alaa Farghili, Matt Kanke, Bo Shui, Wencheng Zhang, Paul Soloway, Zhangjie Wang, Lola Reid, Praveen Sethupathy. Chemical, molecular, and single cell analysis reveal chondroitin sulfate proteoglycan aberrancy in fibrolamellar carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3830. |
