Determination of complex subclonal structures of hematological malignancies by multiplexed genotyping of blood progenitor colonies.

Current next-generation sequencing (NGS) technologies allow unprecedented insights into the mutational profiles of tumors. Recent studies in myeloproliferative neoplasms have further demonstrated that, not only the mutational profile, but also the order in which these mutations are acquired is relevant for our understanding of the disease. Our ability to assign mutation order from NGS data alone is, however, limited. Here, we present a strategy of highly multiplexed genotyping of burst forming unit-erythroid colonies based on NGS results to assess subclonal tumor structure. This allowed for the generation of complex clonal hierarchies and determination of order of mutation acquisition far more accurately than was possible from NGS data alone.Work in ARG lab has been supported by the Leukemia and Lymphoma Society (grant 7001-12), the National Institute of Health Research (grant NF-SI-0512-10079) and core support grants by the MRC and Wellcome Trust to the Cambridge Institute for Medical Research (100140/Z/12/Z) and Wellcome Trust-MRC Cambridge Stem Cell Institute (097922/Z/11/Z). Work in ARG's laboratory has in addition been supported by Cancer Research UK (grants C1163/A12765 and C1163/A21762), Bloodwise (grant 13003) and the Wellcome Trust (grant 104710/Z/14/Z

Kinase joins the chaperone club: Androgen-regulated kinome reveals choline kinase alpha as a potential drug target in prostate cancer.

To identify clinically relevant downstream effectors of androgen signaling, the androgen-regulated kinome was defined in prostate cancer (PCa). Within this study, choline kinase α (CHKA) was identified as an androgen receptor chaperone that is both a biomarker of progression and a potential therapeutic target for PCa.This work was supported by a Cancer Research UK program grant (to DEN)

Pro-neural transcription factors as cancer markers.

BACKGROUND: The aberrant transcription in cancer of genes normally associated with embryonic tissue differentiation at various organ sites may be a hallmark of tumour progression. For example, neuroendocrine differentiation is found more commonly in cancers destined to progress, including prostate and lung. We sought to identify proteins which are involved in neuroendocrine differentiation and differentially expressed in aggressive/metastatic tumours. RESULTS: Expression arrays were used to identify up-regulated transcripts in a neuroendocrine (NE) transgenic mouse model of prostate cancer. Amongst these were several genes normally expressed in neural tissues, including the pro-neural transcription factors Ascl1 and Hes6. Using quantitative RT-PCR and immuno-histochemistry we showed that these same genes were highly expressed in castrate resistant, metastatic LNCaP cell-lines. Finally we performed a meta-analysis on expression array datasets from human clinical material. The expression of these pro-neural transcripts effectively segregates metastatic from localised prostate cancer and benign tissue as well as sub-clustering a variety of other human cancers. CONCLUSION: By focussing on transcription factors known to drive normal tissue development and comparing expression signatures for normal and malignant mouse tissues we have identified two transcription factors, Ascl1 and Hes6, which appear effective markers for an aggressive phenotype in all prostate models and tissues examined. We suggest that the aberrant initiation of differentiation programs may confer a selective advantage on cells in all contexts and this approach to identify biomarkers therefore has the potential to uncover proteins equally applicable to pre-clinical and clinical cancer biology.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

A glycolytic phenotype is associated with prostate cancer progression and aggressiveness: a role for monocarboxylate transporters as metabolic targets for therapy.

Metabolic adaptation is considered an emerging hallmark of cancer, whereby cancer cells exhibit high rates of glucose consumption with consequent lactate production. To ensure rapid efflux of lactate, most cancer cells express high levels of monocarboxylate transporters (MCTs), which therefore may constitute suitable therapeutic targets. The impact of MCT inhibition, along with the clinical impact of altered cellular metabolism during prostate cancer (PCa) initiation and progression, has not been described. Using a large cohort of human prostate tissues of different grades, in silico data, in vitro and ex vivo studies, we demonstrate the metabolic heterogeneity of PCa and its clinical relevance. We show an increased glycolytic phenotype in advanced stages of PCa and its correlation with poor prognosis. Finally, we present evidence supporting MCTs as suitable targets in PCa, affecting not only cancer cell proliferation and survival but also the expression of a number of hypoxia-inducible factor target genes associated with poor prognosis. Herein, we suggest that patients with highly glycolytic tumours have poorer outcome, supporting the notion of targeting glycolytic tumour cells in prostate cancer through the use of MCT inhibitors.Pertega-Gomes N. and Sousa S. received fellowships from the Portuguese Foundation for Science and Technology (FCT), refs. SFRH/BD/61027/2009, and PTDC/SAU-MET/113415/2009, respectively. Felisbino S. received a fellowship from the Sao Paulo Research Foundation (FAPESP) ref. 2013/08830-2 and 2013/06802-1. We thank the core facilities at the Cancer Research UK Cambridge Institute led by James Hadfield (Genomics), Matt Eldridge (Bioinformatics) and Allen Hazelhurst (BRU). We also thank the support and critical advice on the project given by Christian Frezza and Marco Sciacovelli from The MRC Cancer Cell Unit and Professor Rui Henrique from Portuguese Institute of Oncology for providing samples from patients with metastatic prostate cancer.This is the final version of the article. It first appeared from Wiley via http://dx.doi.org/10.1002/path.454

The Early Effects of Rapid Androgen Deprivation on Human Prostate Cancer.

The androgen receptor (AR) is the dominant growth factor in prostate cancer (PCa). Therefore, understanding how ARs regulate the human transcriptome is of paramount importance. The early effects of castration on human PCa have not previously been studied 27 patients medically castrated with degarelix 7 d before radical prostatectomy. We used mass spectrometry, immunohistochemistry, and gene expression array (validated by reverse transcription-polymerase chain reaction) to compare resected tumour with matched, controlled, untreated PCa tissue. All patients had levels of serum androgen, with reduced levels of intraprostatic androgen at prostatectomy. We observed differential expression of known androgen-regulated genes (TMPRSS2, KLK3, CAMKK2, FKBP5). We identified 749 genes downregulated and 908 genes upregulated following castration. AR regulation of α-methylacyl-CoA racemase expression and three other genes (FAM129A, RAB27A, and KIAA0101) was confirmed. Upregulation of oestrogen receptor 1 (ESR1) expression was observed in malignant epithelia and was associated with differential expression of ESR1-regulated genes and correlated with proliferation (Ki-67 expression).We thank CRUK, The NIHR, The Academy of Medical Sciences(RG:63397) and the National Cancer Research Prostate Cancer: Mechanisms of Progression and Treatment (ProMPT) collaborative (G0500966/75466), Hutchison Whampoa Limited, the Human Research Tissue Bank (Addenbrooke’s Hospital, supported by the NIHR Cambridge BRC), and Cancer Research UK

Ultra-sensitive detection of circulating tumour DNA enriches for patients with greater risk of recurrence in clinically localised prostate cancer

Funding: C.E.M. and H.D. were supported by the Cancer Research UK Cambridge Centre, John Black Charitable Foundation and Prostate Cancer Foundation. H.D. and V.J.G. acknowledge infrastructure support from the National Institute for Health Research (NIHR) Cambridge Biomedical Research Centre (BRC-1215- 20014).Peer reviewe

Dynamic partitioning of branched-chain amino acids-derived nitrogen supports renal cancer progression

Publisher Copyright: © 2022, The Author(s).Metabolic reprogramming is critical for tumor initiation and progression. However, the exact impact of specific metabolic changes on cancer progression is poorly understood. Here, we integrate multimodal analyses of primary and metastatic clonally-related clear cell renal cancer cells (ccRCC) grown in physiological media to identify key stage-specific metabolic vulnerabilities. We show that a VHL loss-dependent reprogramming of branched-chain amino acid catabolism sustains the de novo biosynthesis of aspartate and arginine enabling tumor cells with the flexibility of partitioning the nitrogen of the amino acids depending on their needs. Importantly, we identify the epigenetic reactivation of argininosuccinate synthase (ASS1), a urea cycle enzyme suppressed in primary ccRCC, as a crucial event for metastatic renal cancer cells to acquire the capability to generate arginine, invade in vitro and metastasize in vivo. Overall, our study uncovers a mechanism of metabolic flexibility occurring during ccRCC progression, paving the way for the development of novel stage-specific therapies.Peer reviewe

The architecture of clonal expansions in morphologically normal tissue from cancerous and non-cancerous prostates

Background: Up to 80% of cases of prostate cancer present with multifocal independent tumour lesions leading to the concept of a field effect present in the normal prostate predisposing to cancer development. In the present study we applied Whole Genome DNA Sequencing (WGS) to a group of morphologically normal tissue (n = 51), including benign prostatic hyperplasia (BPH) and non-BPH samples, from men with and men without prostate cancer. We assess whether the observed genetic changes in morphologically normal tissue are linked to the development of cancer in the prostate. Results: Single nucleotide variants (P = 7.0 × 10–03, Wilcoxon rank sum test) and small insertions and deletions (indels, P = 8.7 × 10–06) were significantly higher in morphologically normal samples, including BPH, from men with prostate cancer compared to those without. The presence of subclonal expansions under selective pressure, supported by a high level of mutations, were significantly associated with samples from men with prostate cancer (P = 0.035, Fisher exact test). The clonal cell fraction of normal clones was always higher than the proportion of the prostate estimated as epithelial (P = 5.94 × 10–05, paired Wilcoxon signed rank test) which, along with analysis of primary fibroblasts prepared from BPH specimens, suggests a stromal origin. Constructed phylogenies revealed lineages associated with benign tissue that were completely distinct from adjacent tumour clones, but a common lineage between BPH and non-BPH morphologically normal tissues was often observed. Compared to tumours, normal samples have significantly less single nucleotide variants (P = 3.72 × 10–09, paired Wilcoxon signed rank test), have very few rearrangements and a complete lack of copy number alterations. Conclusions: Cells within regions of morphologically normal tissue (both BPH and non-BPH) can expand under selective pressure by mechanisms that are distinct from those occurring in adjacent cancer, but that are allied to the presence of cancer. Expansions, which are probably stromal in origin, are characterised by lack of recurrent driver mutations, by almost complete absence of structural variants/copy number alterations, and mutational processes similar to malignant tissue. Our findings have implications for treatment (focal therapy) and early detection approaches

Discovery of PTN as a serum-based biomarker of pro-metastatic prostate cancer.

Distinguishing clinically significant from indolent prostate cancer (PC) is a major clinical challenge. We utilised targeted protein biomarker discovery approach to identify biomarkers specific for pro-metastatic PC. Serum samples from the cancer-free group; Cambridge Prognostic Group 1 (CPG1, low risk); CPG5 (high risk) and metastatic disease were analysed using Olink Proteomics panels. Tissue validation was performed by immunohistochemistry in a radical prostatectomy cohort (n = 234). We discovered that nine proteins (pleiotrophin (PTN), MK, PVRL4, EPHA2, TFPI-2, hK11, SYND1, ANGPT2, and hK14) were elevated in metastatic PC patients when compared to other groups. PTN levels were increased in serum from men with CPG5 compared to benign and CPG1. High tissue PTN level was an independent predictor of biochemical recurrence and metastatic progression in low- and intermediate-grade disease. These findings suggest that PTN may represent a novel biomarker for the presence of poor prognosis local disease with the potential to metastasise warranting further investigation

Independence of HIF1a and androgen signaling pathways in prostate cancer

Funder: Cancer Research UK; doi: http://dx.doi.org/10.13039/501100000289Abstract: Background: Therapeutic targeting of the androgen signaling pathway is a mainstay treatment for prostate cancer. Although initially effective, resistance to androgen targeted therapies develops followed by disease progression to castrate-resistant prostate cancer (CRPC). Hypoxia and HIF1a have been implicated in the development of resistance to androgen targeted therapies and progression to CRCP. The interplay between the androgen and hypoxia/HIF1a signaling axes was investigated. Methods: In vitro stable expression of HIF1a was established in the LNCaP cell line by physiological induction or retroviral transduction. Tumor xenografts with stable expression of HIF1a were established in castrated and non-castrated mouse models. Gene expression analysis identified transcriptional changes in response to androgen treatment, hypoxia and HIF1a. The binding sites of the AR and HIF transcription factors were identified using ChIP-seq. Results: Androgen and HIF1a signaling promoted proliferation in vitro and enhanced tumor growth in vivo. The stable expression of HIF1a in vivo restored tumor growth in the absence of endogenous androgens. Hypoxia reduced AR binding sites whereas HIF binding sites were increased with androgen treatment under hypoxia. Gene expression analysis identified seven genes that were upregulated both by AR and HIF1a, of which six were prognostic. Conclusions: The oncogenic AR, hypoxia and HIF1a pathways support prostate cancer development through independent signaling pathways and transcriptomic profiles. AR and hypoxia/HIF1a signaling pathways independently promote prostate cancer progression and therapeutic targeting of both pathways simultaneously is warranted