Funding Sources

We are very thankful to all our funding sources (current and past), which include:
 

                      

                      

                      

 

ERC Consolidator Award: Background and key findings:


The proposal revolves around understanding and characterising hormone dependent
breast cancer. There are 55,000 new cases of breast cancer very year in the UK
alone, a full 75% of these are driven by the estrogen receptor (ER) pathway.
Drugs that block ER activity have had a dramatic impact on survival rates, but
a third (~12,000) of patients will relapse and die of metastatic breast cancer.
We and others have recently shown that the three key factors that make an ER
pathway function are ER, FoxA1 and GATA3. Importantly, these factors are
mutated in cancer, but we don’t know what these mutations do. In addition,
recent evidence has suggested that parallel hormonal pathways, particularly
progesterone receptor (PR) and androgen receptor (AR) can impact ER activity,
potentially by competing with ER for DNA access.

The key goals of the proposal were to understand how the hormonal pathways
influence ER activity and to exploit these findings therapeutically. In
addition, a key goal was to identify what the role of ER-associated proteins,
specifically GATA3 and FoxA1 were in disease progression. The overall
objectives were to delineate mechanisms of estrogen receptor activity in breast
cancer, with a focus on identifying and exploiting the changes in protein
fidelity and the potential for transcription factor cross-talk.

We have been exploring the role of progesterone (P4) in contexts where ER
(ESR1) is mutated and our data suggest that the ESR1 mutation associated with a
third of distant metastases, retains sensitivity to P4. In addition, we have
attempted to tackle a couple of the key controversies and complexities in the
field. We have recently explored whether MPA behaves the same as P4/progestins
and have some data to suggest that they are not the same. MPA appears to be
able to induce changes in ER binding, independent of PR, which is molecular
evidence that MPA is not the same as other progestins or P4.

We discovered an interaction between FoxA1 and MLL3, the methyltransferase that
deposits the histone marks at enhancer elements. We could show that MLL3 is
recruited to ER enhancers, via FoxA1, where it contributes to deposition of
methyl groups to form H3K4me1 and H3K4me2 marks. This suggests that FoxA1
binding precedes acquisition of the enhancer histone marks.

We have established a method called qPLEX-RIME. The qPLEX-RIME approach
combines the RIME method with chemical ways of tagging proteins, so that we can
mix multiple samples in one Mass Spec run and importantly we can get
quantitative information about changes in interacting proteins. We have applied
this method to discover novel FoxA1 and GATA3 interacting proteins which has
been informative. We have identified FoxA1 interacting proteins that only
associate with FoxA1 in drug resistant contexts, namely NFIB and ETV6. We have
validated these finding and are currently exploring the functional role of
these factors.

We have found an unexpected functional connection between GATA3 and TET2. TET2
is the enzyme that converts DNA methylation to 5hmC and there is literature
suggesting that 5hMC can modulate gene expression and we hypothesise that the
connection between GATA3 and TET2 may be intimately involved in this. This body
of work has been completed and has been written up for submission.

To complement the qPLEX-RIME data, we have established and utilised genome-wide
CRISPR screens and have applied these to identify genes involved in treatment
response and resistance. We discovered a role for ARID1A in the response to
three different anti-growth agents. ARID1A was the most essential gene for
tamoxifen and fulvestrant to work, but was the the number one 'hit' in JQ1 (BET
inhibitor) treated cells, but in the opposite direction, where loss of ARID1A
sensitised cells to JQ1. We identified the underlying mechanistic explanation
for this, whereby ARID1A recruits HDAC1 to maintain a deacetylated state. Loss
of ARID1A results in decreased HDAC1 binding and a gain in acetylation, which
is subsequently read by BRD (BET) proteins. This is important because ARID1A is
mutated and inactivated in 1 in 8 women with metastatic ER+ breast cancer.

Publications arising from the ERC funded work:


Mohammed, H, Taylor, C, Brown, G, Papachristou, E, Carroll, J.S and D’Santos,
C. Rapid Immunoprecipitation Mass spectrometry of Endogenous protein (RIME) for
analysis of chromatin complexes. Nature Protocols, 2016, 11: 316-326


Jozwik, K, Chernukhin, I, Serandour, A.A, Nagarajan, S and Carroll J.S. FOXA1
directs H3K4 monomethylation at enhancers via recruitment of the
methyltransferase MLL3. Cell Reports, 2016, 17: 2715-2723


Carroll, J.S, Hickey, T.E, Tarulli, G, Williams, M and Tilley, W.D. Deciphering
the divergent roles of progestogens in breast cancer, Nature Reviews Cancer,
2017, 17: 54-64

Papachristou, E.K , Kishore, K, Holding, A.N, Harvey, K, Roumeliotis, T.I,
Chilamakuri, C.S.R, Omarjee, S, Chia, K-M, Swarbrick, A, Lim, E, Markowetz, F,
Eldridge, M, Siersbaek, R, D’Santos, C.S and Carroll, J.S. Quantitative
Multiplexed Rapid Immunoprecipitation Mass spectrometry of Endogenous proteins
(qPLEX-RIME) for monitoring the Dynamics of Chromatin-Associated Complexes,
Nature Communications, 2018, 9:2311. doi: 10.1038


Serandour, A.A, Mohammed, H, Miremadi, A, Mulder, K.W and Carroll J.S. TRPS1
regulates estrogen-receptor binding and histone acetylation at enhancers.
Oncogene, 2018, doi: 10.1038/s41388-018-0312-2


Glont ,S. E, Chernukhin, I and Carroll J.S. Comprehensive Genomic Analysis
Reveals that the Pioneering Function of FOXA1 Is Independent of Hormonal
Signaling. Cell Rep. 2019, 26:2558-2565


Nagarajan, S, Rao, S.V, Chernukhin, I, Sutton, J, Cheeseman, D, Dunn, S,
Papachristou, E.K, Gonzalez Prada, J-E, Couturier, D-L, Kumar, S, Kishore, K,
Chilamakuri, CSR, Glont, S-E, Goode, E.A, Brodie, C, Guppy, N, Natrajan, R,
Bruna, A, Caldas, C, Russell, A.I, Siersbæk, R, Yusa, K and Carroll, J. S.
ARID1A dictates HDAC1/BRD4 activity, intrinsic proliferative capacity and
breast cancer treatment response. Nature Genetics, 2020, 52(2):187-197.



 

We would also like to thank other organisations that have contributed to our work including: The ROAN Charitable Trust and Deloitte.

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Jason.Carroll@cruk.cam.ac.uk


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