Abstract 314: Fixative eXchange (FX)-seq reveals single nucleus transcriptional landscapes of archived clinical cancer FFPE specimens

Abstract

Abstract The wide availability of FFPE tissues stored in hospitals and biobanks could potentially be an invaluable resource to gain insights for improved medical treatment towards precision medicine by combining genomic and transcriptomic data with pathological annotations and medical records. Nevertheless, the majority of current single-cell sequencing methods depend on fresh or fresh-frozen samples to generate high-quality transcriptome data due to the limited reverse transcription (RT) yields in FFPE samples. Consequently, single-cell level transcriptomic studies of FFPE samples have been a longstanding challenge. Here, we present Fixative-eXchange (FX)-seq, a highly scalable snRNA-seq method for heavily paraformaldehyde (PFA)-fixed and/or FFPE samples. We successfully analyzed a total of nearly 320k nuclei from various samples, including PFA-fixed tissue, FFPE blocks, FFPE and hematoxylin and eosin (H&E)-stained sections from mouse brain and human clinical cancer specimens. In a human gastrointestinal stromal tumor (GIST) FFPE sample, we observed rare transcriptomic transitions toward increased epigenetic modifications affected by acquired resistance in response to prolonged imatinib treatment. Furthermore, in an archived human colorectal cancer (CRC) FFPE sample, spatially-contextualized FX-seq with pathologist annotation distinguishes tumor cells from epithelial progenitors along with identification of the tumor microenvironment (TME) such as cancer-associated fibroblasts (CAFs). We then proceeded to investigate transcriptional programs associated with drug resistance and lack of response in immunotherapy observed in CRC patients. With a hypothesis-driven curation, we compared transcription profiles at the single-nucleus level observed in MSS and MSI-H CRC patients from archived FFPE blocks and found distinctive transcriptional profiles from each group. Overall, FX-seq holds promise for clinical research, precision medicine, biomarker discovery, and molecular diagnosis using archived clinical FFPE specimens. Citation Format: Han-Eol Park, Yu Tak Lee, Jaewon Lee, Junsuk Lee, Hyelin Ji, Young-Lan Song, Yoon Dae Han, Hyunki Kim, Chang Ho Sohn. Fixative eXchange (FX)-seq reveals single nucleus transcriptional landscapes of archived clinical cancer FFPE specimens [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 314.