Abstract 1606: Chromosomal instability drives spatial and temporal phenotypic diversity in Schwann cancer cells
Authors: Yan Haixi , Demeulemeester Jonas , Verfaillie Annelien , Martin Cristina Cotobal , Kaufmann Tom , Schwarz Roland , Voet Thierry , Zaccaria Simone , Flanagan Adrienne , Tarabichi Maxime , Van Loo Peter 
Published: June 2022

Abstract
Background Malignant peripheral nerve sheath tumors (MPNSTs) are rare, highly aggressive soft tissue sarcomas derived from Schwann cells with poor prognosis. Previous studies, although limited in scale and depth, have shown that these tumors are typically genomically complex, with a moderate burden of single nucleotide variants but extensive copy number aberrations (CNAs).
Methods We applied a multi-omics approach to scrutinize the evolution and heterogeneity of a primary MPNST and five recurrence regions from one patient with an NF1 phenotype. Techniques utilized included bulk whole genome sequencing (WGS), single-cell DNA sequencing (scDNA-seq), single-cell RNA sequencing (scRNA-seq), simultaneous genome and transcriptome sequencing (G&T-seq), spatial transcriptomics, and laser capture microdissection (LCM)-based spatial genomics.
Results We find significant heterogeneity in copy number profiles, suggestive of ongoing chromosomal instability and evolution. We perform in-depth tumor phylogenetic reconstruction from bulk WGS data. scDNA-seq revealed further CNA heterogeneity across and within regions, allowing us to refine the tree down to single-cell resolution. De novo somatic variant calling in scDNA-seq data confirms the reconstructed tree and identifies additional subclonal mutations.
We also profile different populations of tumor and non-tumor cells and confirmed this with a genotyping approach. CNA profiles inferred from scRNA-seq reflect the within-region heterogeneity seen in scDNA-seq. We find tumor cells with similar CNA profiles typically cluster together by their gene expression profiles as well, implying that the majority of gene expression heterogeneity in this tumor is underpinned by copy number changes. By using CNA profiles as a native barcoding system, we link genetic subclones to clusters of cells and their transcriptomes to characterize the effects of gene dosage.
Finally, we explore the spatial relationships between tumor subclones using LCM. We detect heterogeneity within tissue sections evidencing local clonal expansions and CNA events that follow spatial distributions. Using spatial transcriptomics, we show tumor cells are homogeneously admixed with tumor microenvironment populations.
Conclusions Our work demonstrates the power of combining spatial multi-omics at the single-cell and bulk levels to study cancer evolution. This enables a spatio-temporal representation of a tumor’s development with annotation of genetic and transcriptomic events.
Citation Format: Haixi Yan, Jonas Demeulemeester, Annelien Verfaillie, Cristina Cotobal Martin, Tom Kaufmann, Roland Schwarz, Thierry Voet, Simone Zaccaria, Adrienne Flanagan, Maxime Tarabichi, Peter Van Loo. Chromosomal instability drives spatial and temporal phenotypic diversity in Schwann cancer cells [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 1606.