Next-generation sequencing of advanced prostate cancer treated with androgen-deprivation therapy.
BACKGROUND
Androgen-deprivation therapy (ADT) is standard treatment for locally advanced or metastatic prostate cancer (PCa). Many patients develop castration resistance (castration-resistant PCa [CRPC]) after approximately 2-3 yr, with a poor prognosis. The molecular mechanisms underlying CRPC progression are unclear.


OBJECTIVE
To undertake quantitative tumour transcriptome profiling prior to and following ADT to identify functionally important androgen-regulated pathways or genes that may be reactivated in CRPC.


METHODS
RNA sequencing (RNA-seq) was performed on tumour-rich, targeted prostatic biopsies from seven patients with locally advanced or metastatic PCa before and approximately 22 wk after ADT initiation. Differentially regulated genes were identified in treatment pairs and further investigated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) on cell lines and immunohistochemistry on a separate CRPC patient cohort. Functional assays were used to determine the effect of pathway modulation on cell phenotypes.


METHODS
We searched for gene expression changes affecting key cell signalling pathways that may be targeted as proof of principle in a CRPC in vitro cell line model.


CONCLUSIONS
We identified ADT-regulated signalling pathways, including the Wnt/β-catenin signalling pathway, and observed overexpression of β-catenin in a subset of CRPC by immunohistochemistry. We validated 6 of 12 (50%) pathway members by qRT-PCR on LNCaP/LNCaP-AI cell RNAs, of which 4 (67%) demonstrated expression changes consistent with RNA-seq data. We show that the tankyrase inhibitor XAV939 (which promotes β-catenin degradation) reduced androgen-independent LNCaP-AI cell line growth compared with androgen-responsive LNCaP cells via an accumulation of cell proportions in the G0/G1 phase and reduction in the S and G2/M phases. Our biopsy protocol did not account for tumour heterogeneity, and pathway inhibition was limited to pharmacologic approaches.


CONCLUSIONS
RNA-seq of paired PCa samples revealed ADT-regulated signalling pathways. Proof-of-principle inhibition of the Wnt/β-catenin signalling pathway specifically delays androgen-independent PCa cell cycle progression and proliferation and warrants further investigation as a potential target for therapy for CRPC.
Institute of Cancer Sciences, College of Medical, Veterinary and Life Sciences, University of Glasgow, Cancer Research UK Beatson Institute, Glasgow, UK; Cancer Research UK Beatson Institute, The Beatson Institute for Cancer Research, Glasgow, UK. Electronic address: p.rajan@beatson.gla.ac.uk.
Prabhakar
P
Rajan
Computational Genomics Analysis and Training Programme, Medical Research Council Functional Genomics Unit, Department of Physiology Anatomy and Genetics, University of Oxford, Oxford, UK.
Ian M
IM
Sudbery
Institute of Cancer Sciences, College of Medical, Veterinary and Life Sciences, University of Glasgow, Cancer Research UK Beatson Institute, Glasgow, UK; Cancer Research UK Beatson Institute, The Beatson Institute for Cancer Research, Glasgow, UK.
M Eugenia M
ME
Villasevil
Cancer Research UK Beatson Institute, The Beatson Institute for Cancer Research, Glasgow, UK.
Ernest
E
Mui
Cancer Research UK Beatson Institute, The Beatson Institute for Cancer Research, Glasgow, UK.
Janis
J
Fleming
Department of Urology, Newcastle upon Tyne Hospitals NHS Foundation Trust, Freeman Hospital, Newcastle upon Tyne, UK.
Mark
M
Davis
Institute of Cancer Sciences, College of Medical, Veterinary and Life Sciences, University of Glasgow, Cancer Research UK Beatson Institute, Glasgow, UK; Cancer Research UK Beatson Institute, The Beatson Institute for Cancer Research, Glasgow, UK.
Imran
I
Ahmad
Institute of Cancer Sciences, College of Medical, Veterinary and Life Sciences, University of Glasgow, Cancer Research UK Beatson Institute, Glasgow, UK.
Joanne
J
Edwards
Cancer Research UK Beatson Institute, The Beatson Institute for Cancer Research, Glasgow, UK.
Owen J
OJ
Sansom
Computational Genomics Analysis and Training Programme, Medical Research Council Functional Genomics Unit, Department of Physiology Anatomy and Genetics, University of Oxford, Oxford, UK.
David
D
Sims
Computational Genomics Analysis and Training Programme, Medical Research Council Functional Genomics Unit, Department of Physiology Anatomy and Genetics, University of Oxford, Oxford, UK.
Chris P
CP
Ponting
Computational Genomics Analysis and Training Programme, Medical Research Council Functional Genomics Unit, Department of Physiology Anatomy and Genetics, University of Oxford, Oxford, UK.
Andreas
A
Heger
Northern Centre for Cancer Care, Newcastle upon Tyne Hospitals NHS Foundation Trust, Freeman Hospital, Newcastle upon Tyne, UK.
Rhona M
RM
McMenemin
Northern Centre for Cancer Care, Newcastle upon Tyne Hospitals NHS Foundation Trust, Freeman Hospital, Newcastle upon Tyne, UK.
Ian D
ID
Pedley
Institute of Cancer Sciences, College of Medical, Veterinary and Life Sciences, University of Glasgow, Cancer Research UK Beatson Institute, Glasgow, UK; Cancer Research UK Beatson Institute, The Beatson Institute for Cancer Research, Glasgow, UK. Electronic address: h.leung@beatson.gla.ac.uk.
Hing Y
HY
Leung
D000368:Aged; D059447:Cell Cycle Checkpoints / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015870:Gene Expression; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D006575:Heterocyclic Compounds, 3-Ring / Q000494:pharmacology; D006801:Humans; D008297:Male; D008875:Middle Aged; D064129:Prostatic Neoplasms, Castration-Resistant / Q000737:chemistry / Q000188:drug therapy* / Q000235:genetics*; D012334:RNA, Neoplasm / Q000032:analysis*; D017423:Sequence Analysis, RNA / Q000379:methods; D059467:Transcriptome; D060449:Wnt Signaling Pathway / Q000235:genetics*; D051176:beta Catenin / Q000032:analysis / Q000235:genetics*
D004791:Enzyme Inhibitors; D006575:Heterocyclic Compounds, 3-Ring; D012334:RNA, Neoplasm; C544261:XAV939; D051176:beta Catenin
Androgen-deprivation therapy; Castration resistant; Prostate cancer; Wnt; β-catenin