Page 19 - Read Online

Page 274 Cancer Drug Resist 2018;1:266-302 I http://dx.doi.org/10.20517/cdr.2018.18

Disruption of the DNA damage response, frequently seen as a consequence of genetic mutation, can lead
to more rapid onset of tumorigenesis. However, inhibiting those pathways which are still intact provides a
potential therapeutic strategy for targeting tumours that have become dependent on their activity. For this
reason, inhibitors of ATR and Chk1 represent a potential new class of anti-cancer therapies, currently in
clinical trials. Here, we describe a mechanism, involving the Chk1 targeted phosphosite Thr505 on the NF-
κB subunit, RelA, and how mutation of this site results in earlier onset of MYC-driven lymphomagenesis in
vivo. We describe a positive feedback loop in which Chk1 phosphorylation of RelA at T505, together with
the NF-κB subunit c-Rel, drives the expression of the ATR checkpoint kinase regulator Claspin in response
to DNA replication stress in cancer cells. This in turn is required for maintenance of Chk1 activity. Loss of
a single allele of the Clspn gene in mice is sufficient to drive earlier tumorigenesis and low levels of CLSPN
mRNA expression are associated with worse survival in some forms of human cancer. Importantly, disrup-
tion of this pathway leads to resistance of cells to treatment with Chk1 inhibitors, a result with implications
for the potential use of these drugs as cancer therapies. Taken together, these data suggest that Claspin
mRNA levels may be a potential biomarker for the response to novel Chk1 inhibitors in the clinic, and also
a prognostic factor in some cancers.

18. Novel strategies to target RNA splicing to overcome androgen receptor splice variant-7
signalling in castration resistant prostate cancer

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A. Sharp , J. C. Welti , W. Yuan , A. Paschalis , I. Figueiredo , C. Bertan , V. S. Gil , D. N. Ro-
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drigues , E. Knight , J. Ning , J. Francis , D. Dolling , A. Neeb , G. Boysen , M. Crespo , B. Al-
Lazikani , S. Carreira1, J. Luo , A. Swain , S. R. Plymate , J. S. de Bono 1
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1 Division of Clinical Studies, The Institute of Cancer Research, UK
2 Tumour Profiling Unit, The Institute of Cancer Research, UK
3 Cancer Research UK Cancer Therapeutics Unit, The Institute of Cancer Research, UK
4 Johns Hopkins University School of Medicine, USA
5 University of Washington, USA
# The two authors contributed equally to this Abstract
The constitutively active androgen receptor (AR) splice variant-7 (AR-V7) drives persistent AR signalling
in castration resistant prostate cancer (CRPC). We demonstrate that AR-V7 protein increases as castration
resistance emerges and associates with resistance to endocrine therapies. In addition, AR-V7 does not cor-
relate with full length AR protein or AR copy number, suggesting that aberrant RNA splicing is critical for
AR splice variant (AR-SV) generation. Targeting RNA splicing is an attractive strategy to prevent AR-V7
generation and we demonstrate a novel role of bromodomain and extra-terminal (BET) family protein in-
hibitors in supressing RNA splicing. Our data show that BRD2, BRD3 and BRD4 RNA expression correlate
with AR driven transcription in CRPC biopsies. Consistent with this, chemical BET inhibition (BETi), and
BET family protein knockdown, reduced AR-V7 expression and AR signalling in prostate cancer models.
In addition, we show BETi regulate RNA processing reducing alternative splicing and AR-V7 expression.
Furthermore, BETi reduce growth of prostate cancer cells and patient derived organoids with known AR
mutations, AR amplification and AR-V7 expression. Consistent with this, BETi, unlike enzalutamide, de-
creases persistent AR signalling and growth of a patient derived xenograft with AR amplification and AR-
V7 expression. Despite these promising data, the pleotropic effects of BETi on cellular pathways and treat-
ment related toxicities remain a concern. In light of this, we determined critical factors for BETi mediated
AR-V7 regulation in CRPC. Interrogation of BETi treated RNA expression data and a focused splicing fac-
tor siRNA screen identified splicing factor B (anonymised; Sf-B) crucial for RNA splicing, AR-V7 genera-
tion and PC cell growth. In addition; Sf-B expression correlates with AR signalling and is associated with
reduced clinical benefit from current endocrine therapies. Based on our results we propose that inhibition
of Sf-B provides a novel approach to target AR-V7 and other AR-SVs in CRPC.