Whole-genome single-cell copy number profiling from formalin-fixed paraffin-embedded samples

• Published in: Nature medicine Vol. 23; no. 3; pp. 376 - 385
• Main Authors: Martelotto, Luciano G, Baslan, Timour, Kendall, Jude, Geyer, Felipe C, Burke, Kathleen A, Spraggon, Lee, Piscuoglio, Salvatore, Chadalavada, Kalyani, Nanjangud, Gouri, Ng, Charlotte K Y, Moody, Pamela, D'Italia, Sean, Rodgers, Linda, Cox, Hilary, da Cruz Paula, Arnaud, Stepansky, Asya, Schizas, Michail, Wen, Hannah Y, King, Tari A, Norton, Larry, Weigelt, Britta, Hicks, James B, Reis-Filho, Jorge S
• Format: Journal Article
• Language: English
• Published: New York Nature Publishing Group US 01.03.2017; Nature Publishing Group
• Subjects: 38/23; 45/22; 692/699/67/1347; 692/699/67/2329; 692/699/67/69; Biomedicine; Breast cancer; Breast Neoplasms - genetics; Breast Neoplasms - pathology; Cancer cells; Cancer Research; Carcinoma, Ductal, Breast - genetics; Carcinoma, Ductal, Breast - pathology; Carcinoma, Intraductal, Noninfiltrating - genetics; Carcinoma, Intraductal, Noninfiltrating - pathology; Cell Nucleus; Disease Progression; DNA Copy Number Variations - genetics; DNA sequencing; Female; Flow Cytometry; Formaldehyde; Genetic aspects; Genetic diversity; Genomes; Genomics; Health aspects; Heterogeneity; Humans; In Situ Hybridization, Fluorescence; Infectious Diseases; MCF-7 Cells; Metabolic Diseases; Methods; Microscopy, Confocal; Molecular Medicine; Multiplex Polymerase Chain Reaction; Neurosciences; Paraffin Embedding; Sequence Analysis, DNA; Single-Cell Analysis; Subpopulations; technical-report; Tissue Fixation; Tumors; New York; United States > US
• ISSN: 1078-8956; 1546-170X
• DOI: 10.1038/nm.4279

A method enabling copy-number analysis of single cells from formalin-fixed paraffin-embedded samples is described, validated and applied to analyze samples of synchronous ductal carcinoma in situ and invasive breast carcinoma. A substantial proportion of tumors consist of genotypically distinct subpopulations of cancer cells. This intratumor genetic heterogeneity poses a substantial challenge for the implementation of precision medicine. Single-cell genomics constitutes a powerful approach to resolve complex mixtures of cancer cells by tracing cell lineages and discovering cryptic genetic variations that would otherwise be obscured in tumor bulk analyses. Because of the chemical alterations that result from formalin fixation, single-cell genomic approaches have largely remained limited to fresh or rapidly frozen specimens. Here we describe the development and validation of a robust and accurate methodology to perform whole-genome copy-number profiling of single nuclei obtained from formalin-fixed paraffin-embedded clinical tumor samples. We applied the single-cell sequencing approach described here to study the progression from in situ to invasive breast cancer, which revealed that ductal carcinomas in situ show intratumor genetic heterogeneity at diagnosis and that these lesions may progress to invasive breast cancer through a variety of evolutionary processes.