Investigating the functional importance of estrogen receptor alpha mutations in endocrine- resistant, metastatic breast cancer
File(s)
Author(s)
Goldsbrough, Isabella
Type
Thesis
Abstract
The growth of between 70% and 80% of breast cancers is driven by the expression
of estrogen receptor-α(ER). This understanding has led to the development of endocrine
therapies which inhibit ER activity. While these have vastly improved patient survival,
around 30% of patients will relapse, invariably developing metastatic, incurable disease.
Recurring mutations in the ER gene, ESR1, are detected in more than 20% of endocrinerefractory,
metastatic tumours, implicating these as an important mechanism of
resistance. However, the molecular consequences of many of these mutations are, as
yet, poorly-defined. Here, I have explored the transcriptomic actions of ESR1 mutations
using CRISPR-cas9 knock-in mutant models in the MCF-7 breast cancer cell line. I show
that MCF7 cells expressing the L536R, Y537C, Y537N, Y537S or the D538G ESR1
mutations express a novel isoform of the transcription factor GATA4. Elevated expression
of this isoform is seen in a broad range of cell line and patient-derived ER mutant models.
Furthermore, I have generated knock-in models of the ESR1 exon 5 mutation E380Q. I
show that, unlike wild-type (WT) parental cells, cells expressing the E380Q mutant are
estrogen-independent for their growth. Moreover, I have shown that functionally
homozygous, but not heterozygous E380Q mutant cells exhibit decreased sensitivity to
selective estrogen receptor modulators and degraders (SERM/SERDs). Surprisingly,
transcriptomic analysis by RNA-sequencing revealed that, unlike L536/Y537/D538
mutations, cells expressing E380Q have markedly reduced ER signalling relative to WT
cells. Mutant clones were found to express an E380Q-specific gene signature that was
associated with poor relapse-free survival in ER-positive breast cancer patients treated
with endocrine therapy. Taken together, the results presented in this thesis highlight a
role for the E380Q mutation in conferring resistance to estrogen-deprivation therapy. The
significance of this mutation in mediating resistance to SERMs and SERDs is less clear,
and may be influenced by continued expression of a WT allele. Crucially, work in this
thesis provides evidence that the E380Q mechanism of action is distinct from that of
mutations in the L536/Y537/D538 ER mutational hotspot.
of estrogen receptor-α(ER). This understanding has led to the development of endocrine
therapies which inhibit ER activity. While these have vastly improved patient survival,
around 30% of patients will relapse, invariably developing metastatic, incurable disease.
Recurring mutations in the ER gene, ESR1, are detected in more than 20% of endocrinerefractory,
metastatic tumours, implicating these as an important mechanism of
resistance. However, the molecular consequences of many of these mutations are, as
yet, poorly-defined. Here, I have explored the transcriptomic actions of ESR1 mutations
using CRISPR-cas9 knock-in mutant models in the MCF-7 breast cancer cell line. I show
that MCF7 cells expressing the L536R, Y537C, Y537N, Y537S or the D538G ESR1
mutations express a novel isoform of the transcription factor GATA4. Elevated expression
of this isoform is seen in a broad range of cell line and patient-derived ER mutant models.
Furthermore, I have generated knock-in models of the ESR1 exon 5 mutation E380Q. I
show that, unlike wild-type (WT) parental cells, cells expressing the E380Q mutant are
estrogen-independent for their growth. Moreover, I have shown that functionally
homozygous, but not heterozygous E380Q mutant cells exhibit decreased sensitivity to
selective estrogen receptor modulators and degraders (SERM/SERDs). Surprisingly,
transcriptomic analysis by RNA-sequencing revealed that, unlike L536/Y537/D538
mutations, cells expressing E380Q have markedly reduced ER signalling relative to WT
cells. Mutant clones were found to express an E380Q-specific gene signature that was
associated with poor relapse-free survival in ER-positive breast cancer patients treated
with endocrine therapy. Taken together, the results presented in this thesis highlight a
role for the E380Q mutation in conferring resistance to estrogen-deprivation therapy. The
significance of this mutation in mediating resistance to SERMs and SERDs is less clear,
and may be influenced by continued expression of a WT allele. Crucially, work in this
thesis provides evidence that the E380Q mechanism of action is distinct from that of
mutations in the L536/Y537/D538 ER mutational hotspot.
Version
Open Access
Date Issued
2021-11
Date Awarded
2022-04
Copyright Statement
Creative Commons Attribution NonCommercial NoDerivatives Licence
Advisor
Ali, Simak
Buluwela, Lakjaya
Sponsor
Medical Research Council (Great Britain)
AstraZeneca (Firm)
Grant Number
G01369
Publisher Department
Department of Surgery & Cancer
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)