• E-cadherin inhibits cell surface localization of the pro-migratory 5T4 oncofetal antigen in mouse embryonic stem cells.

      Spencer, Helen L; Eastham, Angela M; Merry, Catherine L R; Southgate, Thomas D; Perez-Campo, Flor-Maria; Soncin, Francesca; Ritson, Sarah; Kemler, Rolf; Stern, Peter L; Ward, Christopher M; et al. (2007-08)
      Epithelial-mesenchymal transition (EMT) events occur during embryonic development and are important for the metastatic spread of epithelial tumors. We show here that spontaneous differentiation of mouse embryonic stem (ES) cells is associated with an E- to N-cadherin switch, up-regulation of E-cadherin repressor molecules (Snail and Slug proteins), gelatinase activity (matrix metalloproteinase [MMP]-2 and -9), and increased cellular motility, all characteristic EMT events. The 5T4 oncofetal antigen, previously shown to be associated with very early ES cell differentiation and altered motility, is also a part of this coordinated process. E- and N-cadherin and 5T4 proteins are independently regulated during ES cell differentiation and are not required for induction of EMT-associated transcripts and proteins, as judged from the study of the respective knockout ES cells. Further, abrogation of E-cadherin-mediated cell-cell contact in undifferentiated ES cells using neutralizing antibody results in a reversible mesenchymal phenotype and actin cytoskeleton rearrangement that is concomitant with translocation of the 5T4 antigen from the cytoplasm to the cell surface in an energy-dependent manner. E-cadherin null ES cells are constitutively cell surface 5T4 positive, and although forced expression of E-cadherin cDNA in these cells is sufficient to restore cell-cell contact, cell surface expression of 5T4 antigen is unchanged. 5T4 and N-cadherin knockout ES cells exhibit significantly decreased motility during EMT, demonstrating a functional role for these proteins in this process. We conclude that E-cadherin protein stabilizes cortical actin cytoskeletal arrangement in ES cells, and this can prevent cell surface localization of the promigratory 5T4 antigen.
    • An E. coli ada transgenic clone of Nicotiana tabacum var. Xanthi has increased sensitivity to the mutagenic action of alkylating agents, maleic hydrazide and gamma-rays.

      Velemínský, J; Angelis, K; Babůrek, I; Gichner, T; Satava, J; Bríza, J; Margison, Geoffrey P; Institute of Experimental Botany, Academy of Sciences, Prague, Czech Republic. (1994-05-01)
      Two transgenic clones X3 and X15 of Nicotiana tabacum var. Xanthi, heterozygous in two genes (a1 and a2) for chloroplast differentiation and transformed with the E. coli DNA repair gene ada cloned downstream from the 1' direction of the dual mas promoter, differed in the expression of the ada gene, in the number of copies of integrated T-DNA and in the response to the mutagenic action of alkylating and non-alkylating agents. The X3 genome contained four copies and the X15 genome one copy of T-DNA, nevertheless the expression of the ada gene, measured by the activity of O6-alkylguanine DNA alkyltransferase (ATase), was about six times higher in X15 than in X3. ATase activity in both clones was highest in extracts from callus whereas very low (X15) or no (X3) activity was detected in leaf extracts. This may explain the lack of difference between X15 and non-transformed tobacco (NTX) in the frequency of N-methyl-N-nitrosourea (MNU)-induced somatic mutations in leaves. In contrast, the frequency of somatic mutations in X3 was about 2-5 times higher than in NTX and X15 after the same doses of MNU, methyl methanesulfonate, maleic hydrazide and gamma-rays. Alteration of plant gene(s) essential in mutation pathway(s) by insertion of T-DNA or by somaclonal variation may explain the higher sensitivity of the X3 clone.
    • The E. coli ogt gene.

      Margison, Geoffrey P; Cooper, Donald P; Potter, P M; Carcinogenesis Department, Paterson Institute for Cancer Research, Christie Hospital and Holt Radium Institute, Manchester, Great Britain. (2010-08-18)
    • The E1E4 protein of human papillomavirus type 16 associates with a putative RNA helicase through sequences in its C terminus.

      Doorbar, John; Elston, Robert C; Napthine, Sawsan; Raj, Kenneth; Medcalf, Elizabeth; Jackson, Deborah; Coleman, Nick; Griffin, Heather M; Masterson, Philip; Stacey, Simon N; et al. (2000-11)
      Human papillomavirus type 16 (HPV16) infects cervical epithelium and is associated with the majority of cervical cancers. The E1E4 protein of HPV16 but not those of HPV1 or HPV6 was found to associate with a novel member of the DEAD box protein family of RNA helicases through sequences in its C terminus. This protein, termed E4-DBP (E4-DEAD box protein), has a molecular weight of 66,000 (66K) and can shuttle between the nucleus and the cytoplasm. It binds to RNA in vitro, including the major HPV16 late transcript (E1E4. L1), and has an RNA-independent ATPase activity which can be partially inhibited by E1E4. E4-DBP was detectable in the cytoplasm of cells expressing HPV16 E1E4 (in vivo and in vitro) and could be immunoprecipitated as an E1E4 complex from cervical epithelial cell lines. In cell lines lacking cytoplasmic intermediate filaments, loss of the leucine cluster-cytoplasmic anchor region of HPV16 E1wedgeE4 resulted in both proteins colocalizing exclusively to the nucleoli. Two additional HPV16 E1E4-binding proteins, of 80K and 50K, were identified in pull-down experiments but were not recognized by antibodies to E4-DBP or the conserved DEAD box motif. Sequence analysis of E4-DBP revealed homology in its E4-binding region with three Escherichia coli DEAD box proteins involved in the regulation of mRNA stability and degradation (RhlB, SrmB, and DeaD) and with the Rrp3 protein of Saccharomyces cerevisiae, which is involved in ribosome biogenesis. The synthesis of HPV16 coat proteins occurs after E1E4 expression and genome amplification and is regulated at the level of mRNA stability and translation. Identification of E4-DBP as an HPV16 E1E4-associated protein indicates a possible role for E1E4 in virus synthesis.
    • E2 enzymes in genome stability: pulling the strings behind the scenes

      Osborne, Hugh C; Irving, E.; Forment, J. V.; Schmidt, Christine K; Manchester Cancer Research Centre, Division of Cancer Sciences, School of Medical Sciences, Faculty of Biology, Medicine, and Health, University of Manchester, 555 Wilmslow Road, Manchester M20 4GJ (2021)
      Ubiquitin and ubiquitin-like proteins (UBLs) function as critical post-translational modifiers in the maintenance of genome stability. Ubiquitin/UBL-conjugating enzymes (E2s) are responsible, as part of a wider enzymatic cascade, for transferring single moieties or polychains of ubiquitin/UBLs to one or multiple residues on substrate proteins. Recent advances in structural and mechanistic understanding of how ubiquitin/UBL substrate attachment is orchestrated indicate that E2s can exert control over chain topology, substrate-site specificity, and downstream physiological effects to help maintain genome stability. Drug discovery efforts have typically focussed on modulating other members of the ubiquitin/UBL cascades or the ubiquitin-proteasome system. Here, we review the current standing of E2s in genome stability and revisit their potential as pharmacological targets for developing novel anti-cancer therapies.
    • EACR-MRS conference on Seed and Soil: In Vivo Models of Metastasis.

      Teles Alves, I; Cohen, N; Ersan, P; Eyre, Rachel; Godet, I; Holovanchuk, D; Jackstadt, R; Kyjacova, L; Mahal, K; Noguera-Castells, A; et al. (2017-12)
      New experimental tools are urgently required to better understand the metastatic process. The importance of such tools is underscored by the fact that many anti-cancer therapies are generally ineffective against established metastases. This makes a major contribution to the fact that metastatic spread is responsible for over 90% of cancer patient deaths. It was therefore timely that the recent "Seed and Soil: In Vivo Models of Metastasis" conference held in Berlin, Germany (27-29 of November 2017) aimed to give an in-depth overview of the latest research models and tools for studying metastasis, and to showcase recent findings from world-leading metastasis researchers. This Meeting Report summarises the major themes of this ground-breaking conference.
    • EANO-EURACAN clinical practice guideline for diagnosis, treatment, and follow-up of post-pubertal and adult patients with medulloblastoma

      Franceschi, E; Hofer, S; Brandes, AA; Frappaz, D; Kortmann, RD; Bromberg, J; Dangouloff-Ros, V; Boddaert, N; Hattingen, E; Wiestler, B; et al. (2019)
      The European Association of Neuro-Oncology (EANO) and EUropean RAre CANcer (EURACAN) guideline provides recommendations for the diagnosis, treatment, and follow-up of post-pubertal and adult patients with medulloblastoma. The guideline is based on the 2016 WHO classification of tumours of the CNS and on scientific developments published since 1980. It aims to provide direction for diagnostic and management decisions, and for limiting unnecessary treatments and cost. In view of the scarcity of data in adults with medulloblastoma, we base our recommendations on adult data when possible, but also include recommendations derived from paediatric data if justified. Our recommendations are a resource for professionals involved in the management of post-pubertal and adult patients with medulloblastoma, for patients and caregivers, and for health-care providers in Europe. The implementation of this guideline requires multidisciplinary structures of care, and defined processes of diagnosis and treatment.
    • EARL: a multicentre phase III randomised trial to evaluate the efficacy of endobronchial electrocautery with autofluorescence bronchoscopy (AFB) surveillance versus AFB surveillance alone in high-grade bronchial dysplasia

      Kalinke, L.; Thakrar, R.; Daniels, H.; Rintoul, R.; Booton, R.; Hackshaw, Allan; Janes, S.; Lungs for Living Research Unit, London, United Kingdom (2020)
      Introduction: Bronchial pre-invasive lesions are precursors of squamous cell cancer. However, not every pre-invasive lesion is destined to this; some lesions may remain stable or even regress to normal epithelium. Performing surveillance with autofluorescence bronchoscopy (AFB), we have shown that 50% of high-grade lesions (HGLs), i.e. severe dysplasia/carcinoma in-situ, progress to cancer (unpublished). Their treatment remains controversial. Guidelines advocate surgical management. This carries inherent risk and its benefit has not been proven in a randomised trial; new lesions also develop, so a tissue-sparing strategy is needed. Methods: We are conducting a Cancer Research UK-funded, international multi-centre randomised trial to examine whether thermal ablation using bronchoscopic electrocautery of HGLs prevents their progression to cancer. Participants will be randomised to electrocautery and AFB surveillance or AFB surveillance alone. The trial will run across four centres (University College London Hospital, Wythenshawe, Papworth and VUmc, Amsterdam)and opens in December 2019. We aim to recruit 106 participants within 3 years. Referral criteria are: 1. Individuals with HGLs found incidentally on bronchial biopsy or at resection margins post-operatively (even if found up to one year previously) 2. Individuals with abnormal sputum cytology, but normal CT and/or bronchoscopy All participants will have a chest CT, pulmonary function tests and AFB at baseline, and then AFB at 6, 12, 24 and 36 months. Participants can receive two electrocautery treatments per year over the 3-year trial period. Results: The primary endpoint is the time to progression of any HGL in a patient to invasive lung cancer. Secondary outcomes are (i) cancer-free and overall survival, (ii) health-related quality of life and (iii) cost-effectiveness. Conclusion: The EARL trial is the first Phase III randomised trial of an endobronchial therapy for the prevention of lung cancer. If successful, it will address the unmet clinical need for a proven tissue-sparing therapy for this high-risk population.
    • Early chromatin unfolding by RUNX1: a molecular explanation for differential requirements during specification versus maintenance of the hematopoietic gene expression program.

      Hoogenkamp, Maarten; Lichtinger, Monika; Krysinska, Hanna; Lancrin, Christophe; Clarke, Deborah; Williamson, Andrew J K; Mazzarella, Luca; Ingram, Richard; Jorgensen, Helle; Fisher, Amanda; et al. (2009-07-09)
      At the cellular level, development progresses through successive regulatory states, each characterized by their specific gene expression profile. However, the molecular mechanisms regulating first the priming and then maintenance of gene expression within one developmental pathway are essentially unknown. The hematopoietic system represents a powerful experimental model to address these questions and here we have focused on a regulatory circuit playing a central role in myelopoiesis: the transcription factor PU.1, its target gene colony-stimulating-factor 1 receptor (Csf1r), and key upstream regulators such as RUNX1. We find that during ontogeny, chromatin unfolding precedes the establishment of active histone marks and the formation of stable transcription factor complexes at the Pu.1 locus and we show that chromatin remodeling is mediated by the transient binding of RUNX1 to Pu.1 cis-elements. By contrast, chromatin reorganization of Csf1r requires prior expression of PU.1 together with RUNX1 binding. Once the full hematopoietic program is established, stable transcription factor complexes and active chromatin can be maintained without RUNX1. Our experiments therefore demonstrate how individual transcription factors function in a differentiation stage-specific manner to differentially affect the initiation versus maintenance of a developmental program.
    • Early detection of melanoma in specialised primary care practice in Australia

      Green, Adèle C; Pandeya, N.; Morton, S.; Simonidis, J.; Whiteman, D. C.; Population Health Department, QIMR Berghofer Medical Research Institute, 300 Herston Road, Herston, Queensland, 4006, Australia; CRUK Manchester and Faculty of Biology, Medicine and Health, University of Manchester, Manchester, (2020)
      Background: Primary care skin cancer clinics facilitate early treatment of melanoma in Australia. We investigated the clinical and histopathological features of melanomas diagnosed and treated in an established clinic in Brisbane. Methods: Retrospective audit of medical records of patients diagnosed with in situ or invasive primary cutaneous melanoma in a primary care clinic specializing in skin cancer, 2000-2017. Demographic and clinical data were standardly extracted by a medically-trained investigator. We used descriptive analyses to assess characteristics of patients and melanomas, and examine surgical management according to tumour thickness. Results: Of 380 patients (median age 57 years; 57 % male) newly diagnosed with 497 histologically-confirmed primary cutaneous melanomas, 369 were in situ and 128 invasive. Of the 369 in situ melanomas, 143 (39 %) were on the trunk and 87 (24 %) on the head and neck; 247 (67 %) were diagnosed by shave biopsy; and 141 (38 %) referred for wide local excision (WLE). Of the 128 invasive melanomas, only 21 (16 %) had thickness ≥ 0.8 mm and these occurred more often on head and neck than thin invasive melanomas (p = 0.02). The majority of invasive melanomas were diagnosed by excision biopsy, and WLE was carried out in a median of 3 days (melanomas ≥ 0.8 mm) and 2 days (<0.8 mm). The doctor detected the majority of in situ (83 %) and thin invasive (73 %) melanomas during surveillance, compared with 48 % of thicker invasive melanomas ≥ 0.8 mm (p < 0.001). Conclusion: In Australia, specialised primary care practice plays a major role in detection and treatment of early primary melanoma.
    • Early detection of melanoma: a consensus report from the Australian Skin and Skin Cancer Research Centre Melanoma Screening Summit

      Janda, M; Cust, AE; Neale, RE; Aitken, JF; Baade, PD; Green, Adèle C; Khosrotehrani, K; Mar, V; Soyer, HP; Whiteman, DC; et al. (2020)
    • Early gene signalling-dependent and -independent induction of apoptosis in Ramos human B cells can be inhibited by over-expression of Bcl-2.

      Ning, Z Q; Norton, John D; Johnson, Diane; Murphy, J J; Division of Life Sciences, King's College London, UK. (1995-10-04)
      We have previously shown that calcium ionophore-induced apoptosis of Ramos human B cells is preceded by the induced expression of early response genes, implying a requirement for new gene expression in this mode of programmed cell death. We have found in the present studies that inhibitors of macromolecular synthesis, cycloheximide and actinomycin D, are also potent inducers of apoptosis in the same Ramos cell model. These drugs trigger apoptosis through apparently early gene signalling-independent pathways. Although different mechanisms for induction of apoptosis exist in Ramos cells, enforced over-expression of Bcl-2 protects cells from apoptosis induced in response to different agents, demonstrating that Bcl-2 blocks a final common pathway for programmed cell death in the Ramos cell model.
    • Early human hemogenic endothelium generates primitive and definitive hematopoiesis in vitro.

      Garcia-Alegria, E; Menegatti, S; Fadlullah, Muhammad Z H; Menendez, P; Lacaud, Georges; Kouskoff, Valerie; Developmental Haematopoiesis Group, Faculty of Biology, Medicine and Health, The University of Manchester, Manchester M13 9PT, UK (2018)
      The differentiation of human embryonic stem cells (hESCs) to hematopoietic lineages initiates with the specification of hemogenic endothelium, a transient specialized endothelial precursor of all blood cells. This in vitro system provides an invaluable model to dissect the emergence of hematopoiesis in humans. However, the study of hematopoiesis specification is hampered by a lack of consensus in the timing of hemogenic endothelium analysis and the full hematopoietic potential of this population. Here, our data reveal a sharp decline in the hemogenic potential of endothelium populations isolated over the course of hESC differentiation. Furthermore, by tracking the dynamic expression of CD31 and CD235a at the onset of hematopoiesis, we identified three populations of hematopoietic progenitors, representing primitive and definitive subsets that all emerge from the earliest specified hemogenic endothelium. Our data establish that hemogenic endothelium populations endowed with primitive and definitive hematopoietic potential are specified simultaneously from the mesoderm in differentiating hESCs.
    • Early response gene expression in Ras oncoprotein signalling.

      Travers, H; Atherton, Graham T; Deed, R W; Norton, John D; CRC Department of Gene Regulation, Paterson Institute for Cancer Research, Christie Hospital NHS Trust, Manchester. (1996-02)
    • Early response gene signalling cascades activated by ionising radiation in primary human B cells.

      Wilson, R E; Taylor, S L; Atherton, Graham T; Johnston, D; Waters, C M; Norton, John D; CRC Department of Carcinogenesis, Paterson Institute for Cancer Research, Christie Hospital NHS Trust, Manchester, UK. (1993-12)
      We have used a panel of 13 protein kinase C-responsive immediate early gene probes to dissect the cellular signalling pathways activated by ionising gamma radiation in primary human B cells. Of these 13 genes, a delayed transient induction was observed for only 8: c-fos, c-jun, jun-B, jun-D, c-myc, ergI/krox 24 and two 'anonymous' genes, 3L3 and 19A. Expression of c-myc and c-fos mRNAs was paralleled by the appearance of their encoded proteins suggesting that these oncoproteins may couple radiation signalling to cellular responses. Of three protein kinase C-coupled transcription factors examined by gel retardation assay, (AP1, NF kappa B, EgrK/Krox24) only NF kappa B and, to a lesser extent, AP1 was stimulated in response to irradiation. These observations are not obviously compatible with a simple model invoking protein kinase C in radiation signalling in primary B cells and suggest that the pleiotropic effects of ionising radiation on this cell type are mediated through a distinct cellular signalling cascade.
    • Early response gene signalling in bryostatin-stimulated primary B chronic lymphocytic leukaemia cells in vitro.

      Ning, Z Q; Hirose, Tohru; Deed, Richard W; Newton, J; Murphy, J J; Norton, John D; Division of Life Sciences, King's College London, U.K. (1996-10-01)
      The protein kinase C activator bryostatin induces differentiation and antagonizes the effects of tumour-promoting phorbol esters in a number of different cell types. We show here that bryostatin preferentially inhibits phorbol 12-myristate 13-acetate (PMA)-induced proliferation compared with differentiation in a number of different B chronic lymphocytic leukaemia (BCLL) cell populations examined. By using a panel of 11 early-response gene probes in Northern hybridization analysis, we found that the profile of genes induced in response to bryostatin and PMA was qualitatively similar and displayed comparable sensitivities to inhibition with the serine-threonine kinase inhibitor 1-(5-isoquinolinylsulphonyl)-2-methylpiperazine hydrochloride (H7), consistent with common signalling through protein kinase C. However, the nuclear oncogene. c-myc, which was induced strongly in response to PMA treatment, was only marginally up-regulated by bryostatin. In addition, bryostatin selectively inhibited the magnitude of PMA-responsive induction of c-myc, to a degree commensurate with its antagonistic effects seen at the biological level. Finally, an anti-sense oligonucleotide blockade of c-myc inhibited PMA-induced proliferation but not the differentiation of BCLL cells, implicating this nuclear oncogene as an important determinant distinguishing PMA from bryostatin-coupled biological responses and also as a candidate third-messenger effector target for the anti-tumour effects of bryostatin.
    • Early stage NSCLC - challenges to implementing ctDNA-based screening and MRD detection.

      Abbosh, Christopher; Birkbak, Nicolai J; Swanton, Charles; Cancer Research UK Lung Cancer Centre of Excellence London and Manchester, University College London Cancer Institute, London, UK (2018-07-03)
      Circulating tumour DNA (ctDNA) refers to the fraction of cell-free DNA in a patient's blood that originates from a tumour. Advances in DNA sequencing technologies and our understanding of the molecular biology of tumours have resulted in increased interest in exploiting ctDNA as a tool to facilitate earlier detection of cancer and thereby improve therapeutic outcomes by enabling early intervention. ctDNA analysis might also have utility in the adjuvant therapeutic setting by enabling the identification of patients at a high risk of disease recurrence on the basis of the detection of post-surgical minimal (or molecular) residual disease (MRD). This approach could provide the capability to adapt clinical trials in the adjuvant setting in order to optimize risk stratification, and we argue that this objective is achievable with current technologies. Herein, we evaluate contemporary next-generation sequencing (NGS) approaches to ctDNA detection with a focus on non-small-cell lung cancer. We explain the technical and analytical challenges to low-frequency mutation detection using NGS-based ctDNA profiling and evaluate the feasibility of ctDNA profiling in both screening and MRD assessment contexts.
    • Early steps in the free radical polymerisation of 3,4-dihydroxyphenylalanine (dopa) into malanin

      Chedekel, M R; Land, Edward J; Thompson, A; Truscott, T G; Division of Environmental Chemistry, Department of Environmental Health Sciences, Johns Hopkins University School of Hygiene and Public Health, MD 21205, U.S.A. (1984)
    • Early studies on human chromosomes.

      Harnden, David G; Paterson Institute for Cancer Research, Christie Hospital, Manchester, UK. (1996-02)
      The author describes his introduction to the field of cytogenetics, with his first viewing of himself, cytogenetically, down the microscope, and the progression of human cytogenetics as an area of study up to its modern integration with molecular genetics and computer technology.
    • Early-response gene signalling is induced by angiogenic oligosaccharides of hyaluronan in endothelial cells. Inhibition by non-angiogenic, high-molecular-weight hyaluronan.

      Deed, Richard W; Rooney, P; Kumar, Patricia; Norton, John D; Smith, J; Freemont, A J; Kumar, Shant; Paterson Institute for Cancer Research, Manchester, UK. (1997-04-10)
      The degradation products of hyaluronan are known to stimulate endothelial-cell proliferation and to promote neovascularization associated with angiogenesis, whilst native high-molecular-weight hyaluronan is inhibitory to these processes. To investigate the cellular signalling pathways coupled to hyaluronan-induced responses in angiogenesis, we have analyzed early-response gene expression in vitro, in cultured bovine aortic endothelial cells. Angiogenic oligosaccharides of hyaluronan induced rapid transient up-regulation of the immediate early genes c-fos, c-jun, jun-B, Krox-20 and Krox-24. In contrast, native hyaluronan when used alone failed to elicit a significant change in expression of any of the genes tested, and when used in combination with angiogenic oligosaccharides of hyaluronan, gave a dose-dependent inhibition of induced gene expression. However, prior addition of angiogenic hyaluronan, as little as one minute before addition of high-molecular-weight hyaluronan, abrogated this inhibition, suggesting that positive or negative responses associated with hyaluronan signalling are integrated at a very early stage following receptor binding. Conversely, prior addition of high-molecular-weight hyaluronan led to an irreversible block in gene expression and proliferative response. These data are consistent with native hyaluronan antagonizing the angiogenic response in part by blocking a signalling cascade at or immediately following ligand-receptor interaction. Finally, we demonstrated that chronic exposure to oligosaccharides of hyaluronan is essential for cell proliferation, indicating that short-term immediate early-gene signalling is insufficient to elicit the proliferation of endothelial cells.