Abstract 2431: Single cell gene expression profiling in breast cancer cells with the Her2/neu gene knockout by CRISPR-Cas9

Abstract

Abstract The Her2/neu gene is amplified and overexpressed in 15%-30% of breast cancers. The overexpression of this oncogene is strongly correlated with decreased survival, increased cancer relapse, and poor prognosis. Although therapy has been successfully developed to target this oncogene, a better understanding of this oncogene will provide further insight for breast cancer biology and future drug development. In recent years, the CRISPR-Cas9 system has emerged as an efficient method for genetic engineering, enabling targeted gene knockout with minimal off-target effects. In this study, we employed a plasmid-based CRISPR-Cas9 system to knock out the Her2/neu gene in breast cancer cells. Puromycin selected, GFP-positive (two indicators of the incorporation of the CRISPR-Cas9 system), Her2-negative individual cells were index-sorted using a BD FACSMelody™ system into individual wells of a 96-well BD™ Precise plate with sample barcoding and molecular indexing. A whole transcriptome amplification (WTA) assay was performed to obtain a gene expression profile for each of the Her2/neu knockout cells. Her2 gene knockout was performed on two cell lines: T47D, a ductal carcinoma-derived breast cancer cell line with no Her2 amplification that shows low to intermediate Her2/neu expression; and SKBR3, an adenocarcinoma-derived breast cancer cell line known for Her2/neu gene amplification and overexpression. For comparison, WTA assays were also carried out on parental cells without gene editing. Comparison of the gene expression profiles reveals downstream changes in gene expression correlated to the Her2 knockout. The profiling of gene expression in Her2/neu CRISPR-Cas9 knockout cells on a single cell level promises to provide insight into the mechanisms in these aggressive cancers that could help future drug discovery. For Research Use Only. Not for use in diagnostic or therapeutic procedures. 23-19341-00 Citation Format: Xiaoyang Wang, Chip Lomas, Michael A. Tycon, Craig Betts, Suzanne Weaver. Single cell gene expression profiling in breast cancer cells with the Her2/neu gene knockout by CRISPR-Cas9 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2431. doi:10.1158/1538-7445.AM2017-2431