Investigating the tumour suppressor function of RUNX1 in breast cancer

Sweeney, Kerri Louise (2023) Investigating the tumour suppressor function of RUNX1 in breast cancer. PhD thesis, University of Glasgow.

Abstract

Despite the remarkable advances in the understanding of breast cancer since the 1940s, and therefore in the treatment of the disease, there are still patients whose cancer either cannot be addressed using current targeted therapies or whose cancer does not respond to therapy in the desired manner. These issues highlight the need to identify novel prognostic/therapeutic targets in breast cancer in order that these unmet clinical needs can be addressed.

Various sequencing studies have facilitated the identification of driver mutations in breast cancer and mapping of the breast cancer genomic landscape. Such studies resulted in the identification of RUNX1 as a gene of interest in breast cancer as tumours frequently harboured loss-of-function mutations in RUNX1, particularly those classed as oestrogen receptor(ER)-positive. Previously, RUNX1 has been shown to be an important player in leukaemias where it elicits both a tumour suppressor and oncogenic role depending on context. Likewise, studies focusing on its role in breast cancer have revealed that while ER-positive breast cancers often have RUNX1 mutations, indicating a tumour suppressive role, high levels of RUNX1 expression in the triple-negative (ER-/PR-/HER2-) subtype has been correlated with poor outcome in patients.

Using genetically engineered mouse models (GEMM) for mammary tumorigenesis (with or without the additional mammary-specific deletion of Runx1), our laboratory provided the first definitive in vivo confirmation of RUNX1 acting to restrict tumour development in preclinical models of breast cancer. Accelerated emergence and increased numbers of multifocal and multicentric lesions were observed upon depletion of Runx1, indicating a possible role for RUNX1 in regulating the stem-like potential of the mammary epithelium. Evidence from one of these models suggests the early expansion of a tumour-initiating sub-population of cells with an elevated stem/progenitor-like potential, which may at least partly explain this phenotype.

In this thesis, the mechanisms behind the tumour suppressive functions of RUNX1 in the mammary epithelium were explored, with a particular focus on the transcriptional alterations that are initiated upon deleting this transcription factor. RNA-Sequencing highlighted a myriad of biological functions and pathways that this protein is essential for and offers insights into the various potential mechanisms by which it exerts its tumour-suppressing functions. Notably, the data indicates altered expression of stem-related genes in RUNX1-deleted cells, and thereby offers potential mechanisms that may underpin RUNX1-dependent control over the stem cell compartment (including the regulation of stem-associated genes such as Aldh1a1, Aldh1a7, and Bcl11b), thereby offering some explanation as to why tumours, in which RUNX1 is deleted, exhibit accelerated onset.

Using an immortalised mammary cell line, these findings were validated using mammosphere and 3D colony formation assays for the analysis of stem/progenitorlike potential. While CRISPR/Cas9-mediated deletion of RUNX1 in cells elevates the stem-like potential of mammary epithelial cells, ectopic expression of RUNX1 conversely decreased mammosphere and colony formation capabilities. Additionally, loss of RUNX1 function appears to enhance the stemness-promoting effects of WNT3A-treatment, while its overexpression dampens the stemnesspromoting abilities of WNT3A. Furthermore, stem-like genes such as Aldh1a1 and Aldh1a7 were enriched in Runx1-deleted 3D colonies. Together, this work indicates that RUNX1 is an essential factor for the regulation of the stem-like compartment of mammary epithelial cells, and its loss may result in the expansion of a cellular subpopulation that is more vulnerable to transformation following oncogenic insult.

Collectively, this research offers some valuable insights into the potential functional and molecular mechanisms behind the tumour suppressive function of RUNX1 in the mammary gland, and provides promising avenues to explore in order to extend our understanding of this transcription factor in breast cancer.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Subjects: R Medicine > RC Internal medicine > RC0254 Neoplasms. Tumors. Oncology (including Cancer)
• Colleges/Schools: College of Medical Veterinary and Life Sciences > School of Cancer Sciences
• Supervisor's Name: Blyth, Prof. Karen and Cameron, Prof. Ewan
• Date of Award: 2023
• Depositing User: Theses Team
• Unique ID: glathesis:2023-83472
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 08 Mar 2023 14:04
• Last Modified: 08 Mar 2023 14:04
• Thesis DOI: 10.5525/gla.thesis.83472
• URI: https://theses.gla.ac.uk/id/eprint/83472