Three-dimensional modelling identifies novel genetic dependencies associated with breast cancer progression in the isogenic MCF10 model.

Sarah L Maguire,Aditi Gulati,Christopher J Lord,Simon Vyse,Paul Huang,Holly Barker,Gillian Farnie,Barrie Peck,Rachael Natrajan,Keith Brennan,Patty T Wai,James Campbell,Frances Daley

doi:10.1002/path.4778

Abstract

The initiation and progression of breast cancer from the transformation of the normal epithelium to ductal carcinoma in situ (DCIS) and invasive disease is a complex process involving the acquisition of genetic alterations and changes in gene expression, alongside microenvironmental and recognized histological alterations. Here, we sought to comprehensively characterise the genomic and transcriptomic features of the MCF10 isogenic model of breast cancer progression, and to functionally validate potential driver alterations in three‐dimensional (3D) spheroids that may provide insights into breast cancer progression, and identify targetable alterations in conditions more similar to those encountered in vivo. We performed whole genome, exome and RNA sequencing of the MCF10 progression series to catalogue the copy number and mutational and transcriptomic landscapes associated with progression. We identified a number of predicted driver mutations (including PIK3CA and TP53) that were acquired during transformation of non‐malignant MCF10A cells to their malignant counterparts that are also present in analysed primary breast cancers from The Cancer Genome Atlas (TCGA). Acquisition of genomic alterations identified MYC amplification and previously undescribed RAB3GAP1–HRAS and UBA2–PDCD2L expressed in‐frame fusion genes in malignant cells. Comparison of pathway aberrations associated with progression showed that, when cells are grown as 3D spheroids, they show perturbations of cancer‐relevant pathways. Functional interrogation of the dependency on predicted driver events identified alterations in HRAS, PIK3CA and TP53 that selectively decreased cell growth and were associated with progression from preinvasive to invasive disease only when cells were grown as spheroids. Our results have identified changes in the genomic repertoire in cell lines representative of the stages of breast cancer progression, and demonstrate that genetic dependencies can be uncovered when cells are grown in conditions more like those in vivo. The MCF10 progression series therefore represents a good model with which to dissect potential biomarkers and to evaluate therapeutic targets involved in the progression of breast cancer. © 2016 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Highlights

The initiation and progression of breast cancer from the transformation of the normal epithelium to carcinoma in situ and invasive disease is a multifaceted process that results in the acquisition of multiple genomic alterations, including changes in genomic copy number, structural rearrangements, acquisition of mutations, altered gene expression, and pathway dysregulation [1,2,3,4]
We have performed a comprehensive analysis of both the genomes and transcriptomes of the MCF10 cell line series that represent different stages of breast cancer progression when grown in vivo, and have demonstrated that these cell lines harbour relevant driver alterations seen in primary breast cancers, and represent a good model for studying breast cancer progression with in vitro spheroid models
The patterns of genomic copy number alterations are similar between the cell lines; there are key differences, suggestive of subclonal selection from the parental MCF10A cells

Summary

Introduction

The initiation and progression of breast cancer from the transformation of the normal epithelium to carcinoma in situ and invasive disease is a multifaceted process that results in the acquisition of multiple genomic alterations, including changes in genomic copy number, structural rearrangements, acquisition of mutations, altered gene expression, and pathway dysregulation [1,2,3,4]. The transition through these states, i.e. non-invasive to invasive disease, is a well-defined and staged process, through which breast cancers progress to acquire the capacity to grow, persist, and eventually spread to secondary sites. This series of cell lines represents an isogenic model of disease progression, and provides a useful tool for the investigation of molecular changes during the progression of human breast neoplasia and the generation of tumour heterogeneity on a common genetic background [19]

Methods

Results

Conclusion