Recent Progress 2015
Prostate cancer (PCa) is the third leading cause of cancer-related mortality. Initial response to hormone therapy is almost universally good but progression to castration-resistance PCa (CRPC) is inevitable and lethal. Failure to develop effective therapies is a consequence of genetic heterogeneity and a lack of pathologically defined subtypes that predict patient outcome. Furthermore, our inadequate knowledge of the genetic lesions that drive tumour progression has hampered efforts for effective treatment strategies, resulting in overtreatment or treatment failure. Thus, the main focus of the Prostate Oncobiology group is to advance the understanding of the molecular mechanisms and cellular heterogeneity in prostate tumourigenesis to predict PCa patients’ outcomes and pursue the development of better and personalised therapeutics.
Molecular mechanisms driving prostate tumourigenesis
Understanding the molecular mechanisms underlying tumour initiation and progression is critical for development of more effective prostate cancer therapy. The recent technologies to combine pathological and morphological changes with cancer genome sequencing are providing the means to highlight multiple somatic alterations that occur in cancer, such as mutations in metabolic pathways. One of key PCa pathways alterations identified are the ETS transcription factor gene fusions in ~50% of human prostate cancer cases, making them the most frequent gene fusions associated with human malignancy. The recently developed genetically engineered mouse models (GEMM) based on these key ETS fusions, that more closely replicate specific disease phenotypes, suggest a different role for ERG and ETV1 in prostate tumourigenesis. Specifically, ETV1 directs androgen synthesis, providing new insights into how this oncogene acts to promote hormone unresponsive lethal disease. Indeed, findings from our genetically engineered mice and genomic analysis led us to predict a worse outcome in patients by ETV1 over-expression and the ETV1-associated network.
A major reprogramming of cellular energy metabolism is observed in cancer cells to support continuous cell growth and proliferation, replacing the metabolic program that operates in most normal tissues. This elevation in the rates of glucose uptake, but reduced rates of oxidative phosphorylation by tumours in the presence of oxygen known as aerobic glycolysis, was first noted by Otto Warburg. Since then, glycolytic fuelling has been shown to be associated with activated oncogenes (e.g., RAS, MYC) and mutant tumour suppressors (e.g., TP53). Moreover, gain-of-function mutations in metabolic enzymes such as the isocitrate dehydrogenase 1/2 (IDH) have been reported as driver mutations in glioma and other human tumours. Such findings suggest that targeting metabolic reactions could be a promising therapeutic strategy.
Figure 1. Metabolic profiling of human prostate cell lines. RWPE-1 cells, human immortalised normal prostate cells; control (CTL) and ETV1-expressing (ETV1) conditions. LNCaP, human androgen-dependent prostate cancer cells growing in normal (FBS) and androgen-deprived (AD) conditions; shRNA silencing ETV1 (ETV1 k/d) and non-targeting shRNA control (CTL).
As an initial step in identifying novel or understudied roles of genomic aberrations such as the ETV1 fusion in cancer metabolism, we will focus on its targets involved in steroid biosynthesis, and energy metabolism. We have performed mass spectrometry-based metabolome analysis on human prostate cancer cells after ETV1 overexpression and silencing (Figure 1). The initial metabolome data analysis, in comparison with gene expression profiling obtained in the same cell lines support our transcriptome findings, and indicates that ETV1 promotes glucose intake and lactate production, and increases lipid metabolism. Thus, we will focus on enzymes that are (i) ETV1 direct targets showing altered expression in ETV1-expressing non-tumorigenic and cancer prostate cells, (ii) highlighted in metabolome and gene expression studies as involved in the reprogrammed de novo lipid synthesis, and (iii) scored in lipid metabolism drug screening. Then, we will test the requirements for selected ETV1 metabolism-associated targets for tumour growth in vivo using our GEMM and preclinical xenograft mouse models. We anticipate that ETV1 metabolic targets and their combinatorial effect with current treatments will improve current therapeutics. Thus, studying and elucidating the relationship of metabolic disorders and cancer should provide new avenue for molecular intervention and may also help to promote a new field for ETV1-expression tumours (e.g. prostate cancer, melanoma, and gastro-intestinal stromal tumours) that will provide metabolism-based targets for cancer patients.
Identification of prostate tumour-initiating cells as novel biomarkers and targets for therapeutics
A central challenge in cancer is whether oncogenic transformation of different cells of origin within an adult tissue gives rise to distinct tumour subtypes that differ in their prognosis and/or heterogeneous treatment response. Thus, a key problem remains the identification of the cell type capable of initiating and sustaining growth of the tumour. Being able to target neoplastic cell populations based on unique surface-marker expression patterns has provided a new avenue in cancer research to study directly the cells thought to be at the root of the disease. Our group would like to identify cells of origin of prostate cancer, and the pathways responsible for the transformation of normal target cells into self-renewing cancer cells.
As a first step to identify tumour-initiating cells, further characterisation of the prostate compartment is needed. Thus, we will initially focus on the characterisation of the basal and luminal epithelial cell compartments in normal and regenerating androgen-deprived mouse prostate. Given the limitation of cell surface markers and histological features defining prostate epithelial cells, gene expression analyses of the cell populations based on current markers may not adequately reflect the difference between the basal and luminal compartments. Single cell analysis overcomes this limitation, and allows identification of novel surface markers to classify the prostatic epithelial compartments. The establishment of self-organising organoids in ex vivo culture has become an emerging paradigm for the study of tissue stem cells and tumorigenic potential. In addition, organoids can reconstitute either normal or transformed prostate in vivo. These findings indicate that prostate organoids represent an excellent system for investigating prostate biology. Thus, to test their differentiation potential and lineage specifications ex vivo, we are establishing organoids cultures from primary mouse prostate cells (Figure 2). Next, we will investigate whether the novel basal and/or luminal subpopulations defined in our single-cell screening might have differential tumorigenic potential by shRNA and CRISPR/Cas9 targeting for genetic engineering of PCa tumour phenotypes in vitro, as shown for intestinal organoids. Furthermore, we will perform mechanistic studies of therapeutic response and resistance comparing organoids from different cells of origin. Thus, the individuality of the single-cell, and the information it contains, is likely to be the key to therapeutically targeting every cell in a tumour.
Figure 2. Generation of mouse prostate epithelial organoids. Bright field images of a Day-8 prostate organoid.
The focus of the Prostate Oncobiology group research is to facilitate the development of patient-specific therapies, and serve as a valuable resource in understanding the roles of cancer metabolism and cancer stem cells regulatory networks. In addition, the data generated from this research will be a valuable resource for novel biomarkers and therapeutic targets for epithelial tumours.