Alistair G Rust

Wellcome Trust Sanger Institute, Cambridge, England, United Kingdom

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Publications (99)982.36 Total impact

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    ABSTRACT: Familial adenomatous polyposis (FAP) and MUTYH-associated polyposis (MAP) are inherited disorders associated with multiple colorectal adenomas that lead to a very high risk of colorectal cancer. The somatic mutations that drive adenoma development in these conditions have not been investigated comprehensively. Here we perform analysis of paired colorectal adenoma and normal tissue DNA from individuals with FAP or MAP, sequencing 14 adenoma whole exomes (eight MAP; six FAP), 55 adenoma targeted exomes (33 MAP, 22 FAP) and germline DNA from each patient, and a further 63 adenomas by capillary sequencing (41 FAP, 22 MAP). With these data we examine the profile of mutated genes, the mutational signatures and the somatic mutation rates, observing significant diversity in the constellations of mutated driver genes in different adenomas, and find loss-of-function mutations in WTX (9%; P < 9.99e-06); a gene implicated in regulation of the WNT pathway and p53 acetylation. These data extend our understanding of the early events in colorectal tumorigenesis in the polyposis syndromes.
    No preview · Article · Sep 2015 · The Journal of Pathology
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    ABSTRACT: The introduction of highly selective ABL-tyrosine kinase inhibitors (TKIs) has revolutionized therapy for chronic myeloid leukemia (CML). However, TKIs are only efficacious in the chronic phase of the disease and effective therapies for TKI-refractory CML, or after progression to blast crisis (BC), are lacking. Whereas the chronic phase of CML is dependent on BCR-ABL, additional mutations are required for progression to BC. However, the identity of these mutations and the pathways they affect are poorly understood, hampering our ability to identify therapeutic targets and improve outcomes. Here, we describe a novel mouse model that allows identification of mechanisms of BC progression in an unbiased and tractable manner, using transposon-based insertional mutagenesis on the background of chronic phase CML. Our BC model is the first to faithfully recapitulate the phenotype, cellular and molecular biology of human CML progression. We report a heterogeneous and unique pattern of insertions identifying known and novel candidate genes and demonstrate that these pathways drive disease progression and provide potential targets for novel therapeutic strategies. Our model greatly informs the biology of CML progression and provides a potent resource for the development of candidate therapies to improve the dismal outcomes in this highly aggressive disease. © 2015 Giotopoulos et al.
    No preview · Article · Aug 2015 · Journal of Experimental Medicine
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    ABSTRACT: B-cell precursor acute lymphoblastic leukemia (B-ALL) is amongst the leading causes of childhood cancer-related mortality. Its most common chromosomal aberration is the ETV6-RUNX1 fusion gene, with ~25 % of ETV6-RUNX1 patients also carrying PAX5 alterations. We have recreated this mutation background by inter-crossing Etv6-RUNX1 (Etv6 (RUNX1-SB) ) and Pax5 (+/-) mice and performed an in vivo analysis to find driver genes using Sleeping Beauty transposon-mediated mutagenesis and also exome sequencing. Combination of Etv6-RUNX1 and Pax5 (+/-) alleles generated a transplantable B220 + CD19+ B-ALL with a significant disease incidence. RNA-seq analysis showed a gene expression pattern consistent with arrest at the pre-B stage. Analysis of the transposon common insertion sites identified genes involved in B-cell development (Zfp423) and the JAK/STAT signaling pathway (Jak1, Stat5 and Il2rb), while exome sequencing revealed somatic hotspot mutations in Jak1 and Jak3 at residues analogous to those mutated in human leukemias, and also mutation of Trp53. Powerful synergies exists in our model suggesting STAT pathway activation and mutation of Trp53 are potent drivers of B-ALL in the context of Etv6-RUNX1;Pax5 (+/-) .
    Full-text · Article · Aug 2015 · BMC Cancer
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    ABSTRACT: Although nearly half of human melanomas harbor oncogenic BRAF(V600E) mutations, the genetic events that cooperate with these mutations to drive melanogenesis are still largely unknown. Here we show that Sleeping Beauty (SB) transposon-mediated mutagenesis drives melanoma progression in Braf(V600E) mutant mice and identify 1,232 recurrently mutated candidate cancer genes (CCGs) from 70 SB-driven melanomas. CCGs are enriched in Wnt, PI3K, MAPK and netrin signaling pathway components and are more highly connected to one another than predicted by chance, indicating that SB targets cooperative genetic networks in melanoma. Human orthologs of >500 CCGs are enriched for mutations in human melanoma or showed statistically significant clinical associations between RNA abundance and survival of patients with metastatic melanoma. We also functionally validate CEP350 as a new tumor-suppressor gene in human melanoma. SB mutagenesis has thus helped to catalog the cooperative molecular mechanisms driving BRAF(V600E) melanoma and discover new genes with potential clinical importance in human melanoma.
    No preview · Article · Apr 2015 · Nature Genetics
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    Full-text · Dataset · Apr 2015
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    ABSTRACT: Alterations of genes encoding transcriptional regulators of lymphoid development are a hallmark of B-progenitor acute lymphoblastic leukemia (B-ALL), and most commonly involve PAX5, encoding the DNA-binding transcription factor paired-box 5. The majority of PAX5 alterations in ALL are heterozygous, and key PAX5 target genes are expressed in leukemic cells, suggesting that PAX5 may be a haploinsufficient tumor suppressor. To examine the role of PAX5 alterations in leukemogenesis, we performed mutagenesis screens of mice heterozygous for a loss-of-function Pax5 allele. Both chemical and retroviral mutagenesis resulted in a significantly increased penetrance and reduced latency of leukemia, with a shift to B-lymphoid lineage. Genomic profiling identified a high frequency of secondary genomic mutations, deletions and retroviral insertions targeting B-lymphoid development, including Pax5, and additional genes and pathways mutated in ALL, including tumor suppressors, Ras and JAK-STAT signaling. These results show that in contrast to simple Pax5 haploinsufficiency, multiple sequential alterations targeting lymphoid development are central to leukemogenesis and contribute to the arrest in lymphoid maturation characteristic of ALL. This cross-species analysis also validates the importance of concomitant alterations of multiple cellular growth, signaling and tumor suppression pathways in the pathogenesis of B-ALL. Copyright © 2015 American Society of Hematology.
    Full-text · Article · Apr 2015 · Blood
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    ABSTRACT: BRAF (v-raf murine sarcoma viral oncogene homolog B) inhibitors elicit a transient anti-tumor response in ∼80% of BRAF V600-mutant melanoma patients that almost uniformly precedes the emergence of resistance. Here we used a mouse model of melanoma in which melanocyte-specific expression of Braf V618E (analogous to the human BRAF V600E mutation) led to the development of skin hyper-pigmentation and nevi, as well as melanoma formation with incomplete penetrance. Sleeping Beauty insertional mutagenesis in this model led to accelerated and fully penetrant melanomagenesis and synchronous tumor formation. Treatment of Braf V618E trans-poson mice with the BRAF inhibitor PLX4720 resulted in tumor regression followed by relapse. Analysis of transposon insertions identified eight genes including Braf, Mitf, and ERas (ES-cell expressed Ras) as candidate resistance genes. Expression of ERAS in human melanoma cell lines conferred resistance to PLX4720 and induced hyperphosphorylation of AKT (v-akt murine thy-moma viral oncogene homolog 1), a phenotype reverted by com-binatorial treatment with PLX4720 and the AKT inhibitor MK2206. We show that ERAS expression elicits a prosurvival signal associated with phosphorylation/inactivation of BAD, and that the resistance of hepatocyte growth factor-treated human melanoma cells to PLX4720 can be reverted by treatment with the BAD-like BH3 mimetic ABT-737. Thus, we define a role for the AKT/BAD pathway in resistance to BRAF inhibition and illustrate an in vivo approach for finding drug resistance genes. melanoma | drug resistance | BRAF inhibitors | mouse models
    Full-text · Article · Jan 2015 · Proceedings of the National Academy of Sciences
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    ABSTRACT: To provide a more comprehensive understanding of the genes and evolutionary forces driving colorectal cancer (CRC) progression, we performed Sleeping Beauty (SB) transposon mutagenesis screens in mice carrying sensitizing mutations in genes that act at different stages of tumor progression. This approach allowed us to identify a set of genes that appear to be highly relevant for CRC and to provide a better understanding of the evolutionary forces and systems properties of CRC. We also identified six genes driving malignant tumor progression and a new human CRC tumor-suppressor gene, ZNF292, that might also function in other types of cancer. Our comprehensive CRC data set provides a resource with which to develop new therapies for treating CRC.URL: http://www.nature.com/ng/journal/vaop/ncurrent/full/ng.3175.html
    No preview · Article · Jan 2015 · Nature Genetics
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    ABSTRACT: Next-generation sequencing of human tumours has refined our understanding of the mutational processes operative in cancer initiation and progression, yet major questions remain regarding factors that induce driver mutations, and the processes that shape their selection during tumourigenesis. We performed whole-exome sequencing (WES) on adenomas from three mouse models of non-small cell lung cancer (NSCLC), induced by exposure to carcinogens (Methyl-nitrosourea (MNU) and Urethane), or by genetic activation of Kras (KrasLA2). Although the MNU-induced tumours carried exactly the same initiating mutation in Kras as seen in the KrasLA2 model (G12D), MNU tumours had an average of 192 non-synonymous, somatic single nucleotide variants (SNVs), compared to only 6 in tumours from the KrasLA2 model. In contrast, the KrasLA2 tumours exhibited a significantly higher level of aneuploidy and copy number alterations (CNAs) compared to the carcinogen-induced tumours, suggesting that carcinogen and genetically-engineered models adopt different routes to tumour development. The wild type (WT) allele of Kras has been shown to act as a tumour suppressor in mouse models of NSCLC. We demonstrate that urethane-induced tumours from WT mice carry mostly (94%) Q61R Kras mutations, while those from Kras heterozygous animals carry mostly (92%) Q61L mutations, indicating a major role of germline Kras status in mutation selection during initiation. The exome-wide mutation spectra in carcinogen-induced tumours overwhelmingly display signatures of the initiating carcinogen, while adenocarcinomas acquire additional C>T mutations at CpG sites. These data provide a basis for understanding the conclusions from human tumour genome sequencing that identified two broad categories based on relative frequency of SNVs and CNAs1, and underline the importance of carcinogen models for understanding the complex mutation spectra seen in human cancers.
    Full-text · Article · Nov 2014 · Nature
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    ABSTRACT: The ability of retroviruses and transposons to insert their genetic material into host DNA makes them widely used tools in molecular biology, cancer research and gene therapy. However, these systems have biases that may strongly affect research outcomes. To address this issue, we generated very large datasets consisting of [Formula: see text] to [Formula: see text] unselected integrations in the mouse genome for the Sleeping Beauty (SB) and piggyBac (PB) transposons, and the Mouse Mammary Tumor Virus (MMTV). We analyzed [Formula: see text] (epi)genomic features to generate bias maps at both local and genome-wide scales. MMTV showed a remarkably uniform distribution of integrations across the genome. More distinct preferences were observed for the two transposons, with PB showing remarkable resemblance to bias profiles of the Murine Leukemia Virus. Furthermore, we present a model where target site selection is directed at multiple scales. At a large scale, target site selection is similar across systems, and defined by domain-oriented features, namely expression of proximal genes, proximity to CpG islands and to genic features, chromatin compaction and replication timing. Notable differences between the systems are mainly observed at smaller scales, and are directed by a diverse range of features. To study the effect of these biases on integration sites occupied under selective pressure, we turned to insertional mutagenesis (IM) screens. In IM screens, putative cancer genes are identified by finding frequently targeted genomic regions, or Common Integration Sites (CISs). Within three recently completed IM screens, we identified 7%-33% putative false positive CISs, which are likely not the result of the oncogenic selection process. Moreover, results indicate that PB, compared to SB, is more suited to tag oncogenes.
    Full-text · Article · Apr 2014 · PLoS Genetics
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    ABSTRACT: Medulloblastoma (MB) is the most common pediatric brain tumor, and in ~25% of cases it is driven by aberrant activation of the Sonic Hedgehog (SHH) pathway in granule neuron precursor cells (GNPs). In this study, we identified novel MB driver genes through a transposon mutagenesis screen in the developing brain of wild-type and Trp53 mutant mice. Twenty-six candidates were identified along with established driver genes such as Gli1 and Crebbp. The transcription factor FoxR2, the most frequent gene identified in the screen, is overexpressed in a small subset of human MB of the SHH subtype. Tgif2 and Alx4, two new putative oncogenes identified in the screen, are strongly expressed in the SHH subtype of human MB. Mutations in these two genes were mutually exclusive with mutations in Gli1 and tended to co-occur, consistent with involvement in the SHH pathway. Notably, Foxr2, Tgif2 and Alx4 activated Gli-binding sites in cooperation with Gli1, strengthening evidence they function in SHH signaling. In support of an oncogenic function, Foxr2 overexpression transformed NIH3T3 cells and promoted proliferation of GNPs, the latter of which was also observed for Tgif2 and Alx4. These findings offer forward genetic and functional evidence associating Foxr2, Tgif2 and Alx4 with SHH subtype MB.
    Preview · Article · Mar 2014 · Cancer Research
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    ABSTRACT: Retroviral insertional mutagenesis (RIM) is a powerful tool for cancer genomics that was combined in this study with deep sequencing (RIM/DS) to facilitate a comprehensive analysis of lymphoma progression. Transgenic mice expressing two potent collaborating oncogenes in the germ line (CD2-MYC, -Runx2) develop rapid onset tumours that can be accelerated and rendered polyclonal by neonatal Moloney murine leukaemia virus (MoMLV) infection. RIM/DS analysis of 28 polyclonal lymphomas identified 771 common insertion sites (CISs) defining a 'progression network' that encompassed a remarkably large fraction of known MoMLV target genes, with further strong indications of oncogenic selection above the background of MoMLV integration preference. Progression driven by RIM was characterised as a Darwinian process of clonal competition engaging proliferation control networks downstream of cytokine and T-cell receptor signalling. Enhancer mode activation accounted for the most efficiently selected CIS target genes, including Ccr7 as the most prominent of a set of chemokine receptors driving paracrine growth stimulation and lymphoma dissemination. Another large target gene subset including candidate tumour suppressors was disrupted by intragenic insertions. A second RIM/DS screen comparing lymphomas of wild-type and parental transgenics showed that CD2-MYC tumours are virtually dependent on activation of Runx family genes in strong preference to other potent Myc collaborating genes (Gfi1, Notch1). Ikzf1 was identified as a novel collaborating gene for Runx2 and illustrated the interface between integration preference and oncogenic selection. Lymphoma target genes for MoMLV can be classified into (a) a small set of master regulators that confer self-renewal; overcoming p53 and other failsafe pathways and (b) a large group of progression genes that control autonomous proliferation in transformed cells. These findings provide insights into retroviral biology, human cancer genetics and the safety of vector-mediated gene therapy.
    Full-text · Article · Feb 2014 · PLoS Genetics
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    ABSTRACT: A major challenge in cancer genetics is to determine which low-frequency somatic mutations are drivers of tumorigenesis. Here we interrogate the genomes of 7,651 diverse human cancers and find inactivating mutations in the homeodomain transcription factor gene CUX1 (cut-like homeobox 1) in ~1–5% of various tumors. Meta-analysis of CUX1 mutational status in 2,519 cases of myeloid malignancies reveals disruptive mutations associated with poor survival, highlighting the clinical significance of CUX1 loss. In parallel, we validate CUX1 as a bona fide tumor suppressor using mouse transposon-mediated insertional mutagenesis and Drosophila cancer models. We demonstrate that CUX1 deficiency activates phosphoinositide 3-kinase (PI3K) signaling through direct transcriptional downregulation of the PI3K inhibitor PIK3IP1 (phosphoinositide-3-kinase interacting protein 1), leading to increased tumor growth and susceptibility to PI3K-AKT inhibition. Thus, our complementary approaches identify CUX1 as a pan-driver of tumorigenesis and uncover a potential strategy for treating CUX1-mutant tumors.
    Full-text · Article · Dec 2013 · Nature Genetics
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    ABSTRACT: The most common risk factor for developing hepatocellular carcinoma (HCC) is chronic infection with hepatitis B virus (HBV). To better understand the evolutionary forces driving HCC, we performed a near-saturating transposon mutagenesis screen in a mouse HBV model of HCC. This screen identified 21 candidate early stage drivers and a very large number (2,860) of candidate later stage drivers that were enriched for genes that are mutated, deregulated or functioning in signaling pathways important for human HCC, with a striking 1,199 genes being linked to cellular metabolic processes. Our study provides a comprehensive overview of the genetic landscape of HCC.
    Full-text · Article · Dec 2013 · Nature Genetics
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    ABSTRACT: The Sleeping Beauty (SB) transposon mutagenesis screen is a powerful tool to facilitate the discovery of cancer genes that drive tumorigenesis in mouse models. In this study, we sought to identify genes that functionally cooperate with sonic hedgehog signaling to initiate medulloblastoma (MB), a tumor of the cerebellum. By combining SB mutagenesis with Patched1 heterozygous mice (Ptch1(lacZ/+)), we observed an increased frequency of MB and decreased tumor-free survival compared with Ptch1(lacZ/+) controls. From an analysis of 85 tumors, we identified 77 common insertion sites that map to 56 genes potentially driving increased tumorigenesis. The common insertion site genes identified in the mutagenesis screen were mapped to human orthologs, which were used to select probes and corresponding expression data from an independent set of previously described human MB samples, and surprisingly were capable of accurately clustering known molecular subgroups of MB, thereby defining common regulatory networks underlying all forms of MB irrespective of subgroup. We performed a network analysis to discover the likely mechanisms of action of subnetworks and used an in vivo model to confirm a role for a highly ranked candidate gene, Nfia, in promoting MB formation. Our analysis implicates candidate cancer genes in the deregulation of apoptosis and translational elongation, and reveals a strong signature of transcriptional regulation that will have broad impact on expression programs in MB. These networks provide functional insights into the complex biology of human MB and identify potential avenues for intervention common to all clinical subgroups.
    Full-text · Article · Oct 2013 · Proceedings of the National Academy of Sciences
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    ABSTRACT: Background: Medulloblastomas, the most frequent malignant brain tumours affecting children, comprise at least 4 distinct clinicogenetic subgroups. Aberrant sonic hedgehog (SHH) signalling is observed in approximately 25% of tumours and defines one subgroup. Although alterations in SHH pathway genes (e.g. PTCH1, SUFU) are observed in many of these tumours, high throughput genomic analyses have identified few other recurring mutations. Here, we have mutagenised the Ptch+/- murine tumour model using the Sleeping Beauty transposon system to identify additional genes and pathways involved in SHH subgroup medulloblastoma development. Results: Mutagenesis significantly increased medulloblastoma frequency and identified 17 candidate cancer genes, including orthologs of genes somatically mutated (PTEN, CREBBP) or associated with poor outcome (PTEN, MYT1L) in the human disease. Strikingly, these candidate genes were enriched for transcription factors (p=2x10-5), the majority of which (6/7; Crebbp, Myt1L, Nfia, Nfib, Tead1 and Tgif2) were linked within a single regulatory network enriched for genes associated with a differentiated neuronal phenotype. Furthermore, activity of this network varied significantly between the human subgroups, was associated with metastatic disease, and predicted poor survival specifically within the SHH subgroup of tumours. Igf2, previously implicated in medulloblastoma, was the most differentially expressed gene in murine tumours with network perturbation, and network activity in both mouse and human tumours was characterised by enrichment for multiple gene-sets indicating increased cell proliferation, IGF signalling, MYC target upregulation, and decreased neuronal differentiation. Conclusions: Collectively, our data support a model of medulloblastoma development in SB-mutagenised Ptch+/- mice which involves disruption of a novel transcription factor network leading to Igf2 upregulation, proliferation of GNPs, and tumour formation. Moreover, our results identify rational therapeutic targets for SHH subgroup tumours, alongside prognostic biomarkers for the identification of poor-risk SHH patients.
    Full-text · Article · Jul 2013
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    ABSTRACT: We have developed Cake, a bioinformatics software pipeline that integrates four publicly available somatic variant-calling algorithms to identify single nucleotide variants with higher sensitivity and accuracy than any one algorithm alone. Cake can be run on a high-performance computer cluster or used as a standalone application. Cake is open-source and is available from http://cakesomatic.sourceforge.net/ CONTACT: da1@sanger.ac.uk SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
    Full-text · Article · Jun 2013 · Bioinformatics
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    ABSTRACT: We propose mutational genomics as an approach for identifying putative cancer pathways. This approach relies on expression profiling tumors that are induced by retroviral insertional mutagenesis. Akin to genetical genomics, this provides the opportunity to search for associations between tumor-initiating events (the viral insertion sites) and the consequent transcription changes, thus revealing putative regulatory interactions. An important advantage is that in mutational genomics the selective pressure exerted by the tumor growth is exploited to yield a relatively small number of loci that are likely to be causal for tumor formation. This is unlike genetical genomics which relies on the natural occurring genetic variation between samples to reveal the effects of a locus on gene expression. We performed mutational genomics using a set of 97 lymphoma from mice presenting with splenomegaly. This identified several known as well as novel interactions, including many known targets of Notch1 and Gfi1. In addition to direct one-to-one associations, many multilocus networks of association were found. This is indicative of the fact that a cell has many parallel possibilities in which it can reach a state of uncontrolled proliferation. One of the identified networks suggests that Zmiz1 functions upstream of Notch1. Taken together, our results illustrate the potential of mutational genomics as a powerful approach to dissect the regulatory pathways of cancer.
    No preview · Conference Paper · Jun 2013
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    Full-text · Dataset · Feb 2013
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    Full-text · Dataset · Feb 2013

Publication Stats

9k Citations
982.36 Total Impact Points

Institutions

  • 2009-2015
    • Wellcome Trust Sanger Institute
      • Experimental Cancer Genetics
      Cambridge, England, United Kingdom
  • 2004-2009
    • Institute for Systems Biology
      Seattle, Washington, United States
  • 2006
    • The Ohio State University
      • Department of Molecular and Cellular Biochemistry
      Columbus, Ohio, United States
  • 1998-2006
    • University of Hertfordshire
      • • Science and Technology Research Institute (STRI)
      • • School of Computer Science
      Hatfield, ENG, United Kingdom
  • 2002
    • The University of Western Ontario
      • Department of Computer Science
      London, Ontario, Canada