Single-Cell Multi-Omics Sequencing to Understand the Nature, Extent and Biology of Cellular Heterogeneity in Six Special Breast Cancer Cases

Abstract

Breast tumors consist of different subpopulations of cells with potentially distinctive properties such as treatment-resistance or metastatic potential. Bulk sequencing methodologies have limited capacity to disclose the full extent, nature and biology of cellular heterogeneity in cancer, precluding the development of better anti-cancer modalities.
Single-cell sequencing techniques allow the study of the subclonal architecture of tumors and reveal the co-occurrence of (driver) mutations as well as their order of acquisition over molecular pseudo-time. Recently novel single-cell multi-omics methods have been developed. Importantly, such technologies now enable us to study the diversity of cancer cell states (determined by the interplay of their genome, epigenome and transcriptome) that arises within a tumor, at its most fundamental level, the cell. Genome-plus-transcriptome sequencing (G&T-seq) enables characterizing the DNA and RNA of the same single cell in parallel.

In this project, six special breast tumors were profiled for the first time using 10X RNA sequencing and G&T-seq: one polymorphous carcinoma, one secretory carcinoma and one bone-, one chondroid-, and two myxoid-matrix producing metaplastic carcinomas. We were able to computationally identify the normal and cancer cell states based on their genomic and transcriptomic profiles. The DNA copy number landscape of the polymorphous and the matrix producing carcinomas are highly rearranged and disclosed clear genetic alterations present in subclonal populations of cells. None of the rearrangements observed in the matrix producing carcinomas, were shared between tumors. No aberrations were observed in the secretory carcinoma, in accordance with literature.

At the transcriptomic level we found the presence of fibroblasts, endothelial and immune cells, in addition to the cancerous cells. The identification of specific marker genes and pathways, as well as the integration with pathology information is ongoing.
Lastly, we were able to unambiguously study for the first time the effect of DNA copy number state on the transcriptome in breast tumors using G&T-seq.