• The Saccharomyces cerevisiae actin cytoskeletal component Bsp1p has an auxiliary role in actomyosin ring function and in the maintenance of bud-neck structure.

      Wright, Daniel J; Munro, Ewen; Corbett, Mark; Bentley, Adam J; Fullwood, Nigel J; Murray, Stephen M; Price, Clive; Biomedical Sciences Unit, Biological Sciences, Lancaster University, Lancaster LA1 4YQ, United Kingdom. (2008-04)
      Iqg1p is a component of the actomyosin contractile ring that is required for actin recruitment and septum deposition. Cells lacking Iqg1p function have an altered bud-neck structure and fail to form a functional actomyosin contractile ring resulting in a block to cytokinesis and septation. Here it is demonstrated that increased expression of the actin cytoskeleton associated protein Bsp1p bypasses the requirement for contractile ring function. This also correlates with reduced bud-neck width and remedial septum formation. Increased expression of this protein in a temperature-sensitive iqg1-1 background causes remedial septum formation at the bud neck that is reliant upon chitin synthase III activity and restores cell separation. The observed suppression correlates with a restoration of normal bud-neck structure. While Bsp1p is a component of the contractile ring, its recruitment to the bud neck is not required for the observed suppression. Loss of Bsp1p causes a brief delay in the redistribution of the actin cytoskeleton normally observed at the end of actin ring contraction. Compromise of Iqg1p function, in the absence of Bsp1p function, leads to a profound change in the distribution of actin and the pattern of cell growth accompanied by a failure to complete cytokinesis and cell separation.
    • The Saccharomyces cerevisiae spindle pole body (SPB) component Nbp1p is required for SPB membrane insertion and interacts with the integral membrane proteins Ndc1p and Mps2p.

      Araki, Yasuhiro; Lau, Corine K; Maekawa, Hiromi; Jaspersen, Sue L; Giddings, Thomas H; Schiebel, Elmar; Winey, Mark; The Paterson Institute for Cancer Research, Christie Hospital NHS Trust, Manchester M20 4BX, United Kingdom. (2006-04)
      The spindle pole body (SPB) in Saccharomyces cerevisiae functions to nucleate and organize spindle microtubules, and it is embedded in the nuclear envelope throughout the yeast life cycle. However, the mechanism of membrane insertion of the SPB has not been elucidated. Ndc1p is an integral membrane protein that localizes to SPBs, and it is required for insertion of the SPB into the nuclear envelope during SPB duplication. To better understand the function of Ndc1p, we performed a dosage suppressor screen using the ndc1-39 temperature-sensitive allele. We identified an essential SPB component, Nbp1p. NBP1 shows genetic interactions with several SPB genes in addition to NDC1, and two-hybrid analysis revealed that Nbp1p binds to Ndc1p. Furthermore, Nbp1p is in the Mps2p-Bbp1p complex in the SPB. Immunoelectron microscopy confirmed that Nbp1p localizes to the SPB, suggesting a function at this location. Consistent with this hypothesis, nbp1-td (a degron allele) cells fail in SPB duplication upon depletion of Nbp1p. Importantly, these cells exhibit a "dead" SPB phenotype, similar to cells mutant in MPS2, NDC1, or BBP1. These results demonstrate that Nbp1p is a SPB component that acts in SPB duplication at the point of SPB insertion into the nuclear envelope.
    • Sacral insufficiency fracture following pelvic radiotherapy in gynaecological malignancies: development of a predictive model

      Cachero, S.; Gkantia, M.; Bates, A. S.; Frechter, S.; Blackie, L.; McCarthy, Amy; Sutcliffe, B.; Strano, A.; Aso, Y.; Jefferis, G.; et al. (2020)
      Animal behavior is encoded in neuronal circuits in the brain. To elucidate the function of these circuits, it is necessary to identify, record from and manipulate networks of connected neurons. Here we present BAcTrace (Botulinum-Activated Tracer), a genetically encoded, retrograde, transsynaptic labeling system. BAcTrace is based on Clostridium botulinum neurotoxin A, Botox, which we engineered to travel retrogradely between neurons to activate an otherwise silent transcription factor. We validated BAcTrace at three neuronal connections in the Drosophila olfactory system. We show that BAcTrace-mediated labeling allows electrophysiological recording of connected neurons. Finally, in a challenging circuit with highly divergent connections, BAcTrace correctly identified 12 of 16 connections that were previously observed by electron microscopy.
    • Safeguarding the future of independent, academic clinical cancer research in Europe for the benefit of patients.

      Negrouk, A; Lacombe, D; Cardoso, F; Morin, F; Carrasco, E; Maurel, J; Maibach, R; Aranda, E; Marais, Richard; Stahel, R; et al. (2017)
      Cancer is a complex disease that is constantly evolving. It is now the most common cause of death in Europe after cardiovascular diseases. There are inequalities among European countries, potentially unsustainable healthcare systems impacting quality of cancer care and increasing number of patients with cancer with rare conditions. Clinical and translational research are the backbone in establishing scientific advances as novel treatments and advancing progress to the benefit of patients. Commercially sponsored clinical trials are responsible for developing new medicines that can treat various disease areas, including cancer. It is important to note, however, that these clinical trials only assess the viability of compounds that are chosen by a commercial entity that funds the entire process. By their design and focus, these trials need to fulfil commercial interests and market expectations, which do not always coincide with patients' needs. As soon or even before novel treatments and compounds obtain formal market authorisation, academia will take these existing and new medicines to further conduct research in order to optimise their use, develop new combinations and with a strong focus on the patients and their needs. Established standard of care most commonly relies on clinical cancer research stemming from non-commercial entities, cooperative groups or academic clinical research. This article provides a consensus on the definition of academic research, illustrates its added value and suggests and calls to European Union institutions to support this type of research for the benefit of patients.
    • Safety and immunogenicity of TA-HPV, a recombinant vaccinia virus expressing modified human papillomavirus (HPV)-16 and HPV-18 E6 and E7 genes, in women with progressive cervical cancer.

      Kaufmann, Andreas M; Stern, Peter L; Rankin, Elaine M; Sommer, Harald; Nuessler, Volkmar; Schneider, Achim; Adams, Malcolm; Onon, Toli S; Bauknecht, Thomas; Wagner, Uwe; et al. (2002-12)
      PURPOSE: Cervical cancer, the second most common malignancy in women worldwide, is almost invariably associated with infection by human papillomavirus (HPV). HPV-16 or -18 is commonly present in 70% of cervical cancers. HPV-positive tumor cells present antigens of the viral protein in the context of human leukocyte antigen (HLA) class I that can be recognized by CTLs. We have conducted a study in patients with early-stage cervical cancer to assess the safety and immunological effects of vaccination with TA-HPV, a live recombinant vaccinia virus expressing modified forms of the HPV-16 and -18 E6 and E7 proteins. EXPERIMENTAL DESIGN: Twenty-nine patients with clinical International Federation of Gynecologists and Obstetricians (FIGO) stage Ib or IIa cervical cancer were given two vaccinations with TA-HPV at least 4 weeks apart, starting 2 weeks before radical hysterectomy. Patients were monitored closely for side effects of the vaccination. Serial blood samples were examined for HPV-specific CTLs or changes in levels of antibodies to HPV-16 or -18 E6 and E7 proteins and to vaccinia virus. RESULTS: Vaccination with recombinant vaccinia was well tolerated in all patients with only mild to moderate local toxicity, and no serious adverse events were attributable to the vaccine. After a single vaccination, HPV-specific CTLs were found in four patients (HLA A1, A3, three patients; HLA A1, A24, one patient). Eight patients developed HPV-specific serological responses. CONCLUSIONS: This study confirmed the safety and immunogenicity of the vaccine in a proportion of those patients vaccinated. Additional clinical studies using TA-HPV in combination with an additional experimental vaccine for HPV-16 are currently under way.
    • Safety and tolerability of PCK3145, a synthetic peptide derived from prostate secretory protein 94 (PSP94) in metastatic hormone-refractory prostate cancer.

      Hawkins, Robert E; Daigneault, Luc; Cowan, Richard A; Griffiths, Richard W; Panchal, Chandra; Armstrong, Anne C; Fenemore, Jackie; Irvine, Alan; Sereda, Kasia; Dulude, Hélène; et al. (2005-09)
      BACKGROUND: The safety, tolerability, and pharmacokinetic and preliminary efficacy of PCK3145 were determined in patients with metastatic hormone-refractory prostate cancer. PATIENTS AND METHODS: PCK3145 was administered in ascending doses of 5, 20, 40, and 80 mg/m2 3 times per week for 4 weeks to cohorts of 4 patients. Dose escalation was based on dose-limiting toxicity (DLT). Pharmacokinetic profiles, tumor burden, and tumor markers (including prostate-specific antigen [PSA] and matrix metalloproteinase-9 [MMP-9] levels) were assessed. Sixteen patients received PCK3145. The median age was 66 years, and the median PSA level was 232.5 microg/L. A total of 32 cycles of therapy were administered. RESULTS: The most common adverse events reported were pain and nausea. The only DLT was a grade 4 cardiac arrhythmia in a patient treated at the 80-mg/m2 dose level. Pharmacokinetic analysis using a 2-compartment model indicated that the mean area under the curve values increased as the dose range increased, and the mean elimination half-life ranged from 0.35 hours to 1.45 hours. The best tumor response was stable disease in 10 patients and progressive disease in 5 patients. No PSA responses were observed, but 1 patient showed a marked reduction in PSA of 41% at cycle 2. A substantial reduction in MMP-9 levels was observed in patients with baseline levels of MMP-9 > 100 microg/L. CONCLUSION: PCK3145 was safe and well tolerated at all doses. Efficacy observations were encouraging, and the biologic activity of PCK3145 in reducing MMP-9 level may suggest a potential role of this peptide in the regulation of metastatic tumor growth.
    • Safety, tolerability, pharmacokinetics and pharmacodynamics of the oral cyclin-dependent kinase inhibitor AZD5438 when administered at intermittent and continuous dosing schedules in patients with advanced solid tumours.

      Boss, D S; Schwartz, G K; Middleton, M R; Amakye, D D; Swaisland, H; Midgley, R S; Ranson, Malcolm R; Danson, Sarah; Calvert, H; Plummer, R; et al. (2010-04)
      BACKGROUND: AZD5438 is an orally bioavailable inhibitor of cyclin E-cdk2, cyclin A-cdk2 and cyclin B-cdk1 complexes. Three phase I studies assessed the clinical safety, tolerability, pharmacokinetics and pharmacodynamics of AZD5438 when administered in different dosing schedules. PATIENTS AND METHODS: AZD5438 was administered four times daily, once every 7 days (study 1), for 14 consecutive days followed by 7 days of rest (study 2), or continuously (study 3), to patients with advanced solid tumours. Dose escalation proceeded until the emergence of dose-limiting toxic effects. RESULTS: Sixty-four patients were included across the three studies (19, 17 and 28, respectively). Nausea and vomiting were the most common adverse events. When dosed continuously, 40 mg four times daily was considered intolerable, and due to safety issues, all studies were terminated prematurely. Consequently, no intolerable dose was identified during the weekly schedule. Pharmacokinetics demonstrated dose-proportional exposure, high interpatient variability and accumulation after multiple doses. Skin biopsies indicated reduced retinoblastoma protein phosphorylation at cdk2 phospho-sites; other pharmacodynamic assessments did not reveal consistent trends. CONCLUSIONS: AZD5438 was generally well tolerated in a weekly dosing schedule, but not in continuous schedules. The clinical development programme for AZD5438 was discontinued owing to tolerability and exposure data from these studies.
    • Salvage stereotactic body radiotherapy (SBRT) for intraprostatic relapse after prostate cancer radiotherapy: An ESTRO ACROP Delphi consensus

      Jereczek-Fossa, B. A.; Marvaso, G.; Zaffaroni, M.; Gugliandolo, S. G.; Zerini, D.; Corso, F.; Gandini, S.; Alongi, F.; Bossi, A.; Cornford, P.; et al. (2021)
      Background and purpose: Between 30% and 47% of patients treated with definitive radiotherapy (RT) for prostate cancer are at risk of intraprostatic recurrence during follow-up. Re-irradiation with stereotactic body RT (SBRT) is emerging as a feasible and safe therapeutic option. However, no consensus or guidelines exist on this topic. The purpose of this ESTRO ACROP project is to investigate expert opinion on salvage SBRT for intraprostatic relapse after RT. Materials and methods: A 40-item questionnaire on salvage SBRT was prepared by an internal committee and reviewed by a panel of leading radiation oncologists plus a urologist expert in prostate cancer. Following the procedure of a Delphi consensus, 3 rounds of questionnaires were sent to selected experts on prostate re-irradiation. Results: Among the 33 contacted experts, 18 (54.5%) agreed to participate. At the end of the final round, participants were able to find consensus on 14 out of 40 questions (35% overall) and major agreement on 13 questions (32.5% overall). Specifically, the consensus was reached regarding some selection criteria (no age limit, ECOG 0-1, satisfactory urinary flow), diagnostic procedures (exclusion of metastatic disease, SBRT target defined on the MRI) and therapeutic approach (no need for concomitant ADT, consideration of the first RT dose, validity of Phoenix criteria for salvage SBRT failure). Conclusion: While awaiting the results of ongoing studies, our ESTRO ACROP Delphi consensus may serve as a practical guidance for salvage SBRT. Future research should address the existing disagreements on this promising approach.
    • SAMHD1 sheds moonlight on DNA double-strand break repair.

      Cabello-Lobato, Maria J; Wang, Siyue; Schmidt, Christine K; Division of Cancer Sciences, School of Medical Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester Cancer Research Centre Building, 555 Wilmslow Road, Manchester (2017-09-29)
      SAMHD1 (sterile α motif and histidine (H) aspartate (D) domain-containing protein 1) is known for its antiviral activity of hydrolysing deoxynucleotides required for virus replication. Daddacha et al. identify a hydrolase-independent, moonlighting function of SAMHD1 that facilitates homologous recombination of DNA double-strand breaks (DSBs) by promoting recruitment of C-terminal binding protein interacting protein (CTIP), a DNA-end resection factor, to damaged DNA. These findings could benefit anticancer treatment.
    • Sample pooling strategies for SARS-CoV-2 detection

      Lagopati, N.; Tsioli, P.; Mourkioti, I.; Polyzou, A.; Papaspyropoulos, A.; Zafiropoulos, A.; Evangelou, K.; Sourvinos, G.; Gorgoulis, Vassilis G; Molecular Carcinogenesis Group, Department of Histology and Embryology, School of Medicine, National Kapodistrian University of Athens (NKUA), Greece. (2020)
      The worldwide COVID-19 pandemic outburst has caused a serious public health issue with increasing needs of accurate and rapid diagnostic and screening testing. This situation requires an optimized management of the chemical reagents, the consumables, and the human resources, in order to respond accurately and effectively, controlling the spread of the disease. Testing on pooled samples maximizes the number of tested samples, by minimizing the time and the lab supplies needed. The general conceptualization of the pooling method is based on mixing samples together in a batch. Individual testing is needed only if a specific pool exhibits a positive result. The development of alternative hybrid methods, based on "in house" protocols, utilizing commercially available consumables, in combination with a reliable pooling method would provide a solution, focusing on the better exploitation of the personnel and the lab supplies, allowing for rapid screening of a population in a reasonably short time.
    • Sarcospan, the 25-kDa transmembrane component of the dystrophin-glycoprotein complex.

      Crosbie, R H; Heighway, Jim; Venzke, D P; Lee, J C; Campbell, K P; Howard Hughes Medical Institute, Department of Physiology and Biophysics, University of Iowa College of Medicine, Iowa City, Iowa 52242, USA. (1997-12-12)
      The dystrophin-glycoprotein complex is a multisubunit protein complex that spans the sarcolemma and forms a link between the subsarcolemmal cytoskeleton and the extracellular matrix. Primary mutations in the genes encoding the proteins of this complex are associated with several forms of muscular dystrophy. Here we report the cloning and characterization of sarcospan, a unique 25-kDa member of this complex. Topology algorithms predict that sarcospan contains four transmembrane spanning helices with both N- and C-terminal domains located intracellularly. Phylogenetic analysis reveals that sarcospan's arrangement in the membrane as well as its primary sequence are similar to that of the tetraspan superfamily of proteins. Sarcospan co-localizes and co-purifies with the dystrophin-glycoprotein complex, demonstrating that it is an integral component of the complex. We also show that sarcospan expression is dramatically reduced in muscle from patients with Duchenne muscular dystrophy. This suggests that localization of sarcospan to the membrane is dependent on proper dystrophin expression. The gene encoding sarcospan maps to human chromosome 12p11.2, which falls within the genetic locus for congenital fibrosis of the extraocular muscle, an autosomal dominant muscular dystrophy.
    • Scanning electron microscopy of human metaphase chromosomes.

      Allen, Terence D; Jack, Elspeth M; Harrison, Christine J; Claugher, D (1986)
      Preparative methods for scanning electron microscopy of chromosomes are dependent on the original source of material. Chromosomes extracted from unfixed metaphase cells via isolation buffers tend to show topography and surface morphology which may have been induced by the choice of isolation buffer itself. Furthermore, this type of preparation often precludes any chromosome identification, as many metaphases have been pooled, and also the chromosomes from these preparations are not suitable for the banding techniques regularly used in clinical cytogenetics. Our own approach has been to use the standard cytogenetic approach, starting with methanol-acetic acid fixed, air dried metaphase spreads, allowing both identification of individual chromosomes, and also the facility for various banding procedures such as G and C banding to be performed. Chromosomes are subsequently "reprepared" for SEM, using rehydration, glutaraldehyde fixation, and osmium impregnation using Thiocarbohydrazide (TCH). This method produces chromosomes which can be examined at high resolution, without metallic coating, for their topography, surface morphology and chromatin organisation, and the changes produced by banding techniques which give rise to a structural alterations resulting in differential staining in the light microscope.
    • Scanning electron microscopy of nuclear structure.

      Allen, Terence D; Rutherford, Sandra A; Murray, Stephen M; Drummond, Sheona P; Goldberg, Martin W; Kiseleva, Elena; Department of Structural Cell Biology, Paterson Institute for Cancer Research, University of Manchester, Manchester M20 4BX, United Kingdom. (2008)
      Accessing internal structure and retaining relative three dimensional (3D) organization within the nucleus has always proved difficult in the electron microscope. This is due to the overall size and largely fibrous nature of the contents, making large scale 3D reconstructions difficult from thin sections using transmission electron microscopy. This chapter brings together a number of methods developed for visualization of nuclear structure by scanning electron microscopy (SEM). These methods utilize the easily accessed high resolution available in field emission instruments. Surface imaging has proved particularly useful to date in studies of the nuclear envelope and pore complexes, and has also shown promise for internal nuclear organization, including the dynamic and radical reorganization of structure during cell division. Consequently, surface imaging in the SEM has the potential to make a significant contribution to our understanding of nuclear structure.
    • Scanning electron microscopy of the G-banded human karyotype.

      Harrison, C J; Britch, Martin; Allen, Terence D; Harris, R; Department of Medical Gentics, S Mary's Hospital, Manchester M13 0JH (1981-07)
    • Scanning electron microscopy of variations in human metaphase chromosome structure revealed by Giemsa banding.

      Harrison, Christine J; Allen, Terence D; Harris, R; Paterson Laboratories, Christie Hospital Manchester UK (1983)
    • Scanning the structure and antigenicity of HPV-16 E6 and E7 oncoproteins using antipeptide antibodies.

      Stacey, Simon N; Eklund, C; Jordan, Deborah; Smith, Nigel K; Stern, Peter L; Dillner, J; Arrand, John R; Cancer Research Campaign Department of Molecular Biology, Paterson Institute for Cancer Research, Christie Hospital, Manchester, UK. (1994-02)
      The structure and antigenicity of the HPV-16 E6 and E7 oncoproteins was studied using a set of antisera against overlapping synthetic peptides. We report that antigenic, mobile regions of the native proteins, as defined by reactivity with antipeptide antisera, occur at the N-termini of both E6 and E7 proteins, corresponding to regions of known or suspected protein-protein interactions. The putative zinc finger domains were consistently non-reactive, despite computer predictions of relatively high antigenicity, suggesting that the proposed zinc finger regions are held in stable secondary structures that the peptides were not able to mimic. In E6, the linker region between the two zinc fingers was antigenic, indicating that the two zinc finger structures might be able to articulate relative to one another by a flexible linker region. The highly antigenic N-terminal region of HPV-16 E7 was also found to be antigenic in E7 of both HPV-11 and HPV-18, indicating that the E7 proteins of different HPV types have similar antigenic structures. The identification of antigenic regions of the E6 and E7 proteins should be therefore be useful in the design of site-directed antibodies against E6 and E7 for numerous HPV types.
    • Scatter factor influences the formation of prostate epithelial cell colonies on bone marrow stroma in vitro.

      Lang, Shona H; Clarke, Noel W; George, Nicholas J; Testa, Nydia G; CRC Section of Haemopoietic Cell and Gene Therapeutics, Patterson Institute for Cancer Research, Christie Hospital NHS Trust, Manchester, UK. shl2@imap.york.ac.uk (1999-06)
      Prostate cancer metastases form selectively in the bone marrow. Previously we demonstrated motility was important for the formation of primary prostatic epithelial cell colonies in bone marrow stroma (BMS) co-culture. In this study we looked at the influence of motility factors on the colony formation of epithelial cells derived from benign (bPEC) or malignant (mPEC) prostate tissue. After 7 days co-culture we found that anti-scatter factor consistently inhibited prostate epithelial cell colony formation on BMS (7/7 mPEC and 4/7 bPEC samples showed significant inhibition). Antibodies against bFGF and 5T4 did not significantly affect colony formation. Addition of fibroblast conditioned media (derived from benign prostates) to co-cultures stimulated the colony formation of bPEC (170%) and mPEC (252%). This stimulation was eliminated by depletion of SF from the conditioned media. Immunohistochemical staining found c-Met expression in 5/6 bPEC cultures and 7/9 mPEC cultures. When grown in BMS co-culture expression of c-Met was positive in 3/6 bPEC and 2/7 mPEC samples. In conclusion, scatter factor influences the in vitro formation of prostate epithelial cell colonies on BMS co-culture.
    • Scavenger receptor class B type I regulates cellular cholesterol metabolism and cell signaling associated with breast cancer development.

      Danilo, C; Gutierrez-Pajares, J; Mainieri, M; Mercier, I; Lisanti, Michael P; Frank, P (2013)
      Previous studies have identified cholesterol as an important regulator of breast cancer development. High-density lipoprotein (HDL) and its cellular receptor, the scavenger receptor class B type I (SR-BI) have both been implicated in the regulation of cellular cholesterol homeostasis, but their functions in cancer remain to be established.
    • The scavenging of oxidants by sulphasalazine and its metabolites. A possible contribution to their anti-inflammatory effects?

      Aruoma, O I; Wasil, M; Halliwell, B; Hoey, Brigid M; Butler, John; Department of Biochemistry, King's College, London, U.K. (1987-11-01)
      Sulphasalazine (Salazopyrin) and its metabolites sulphapyridine and 5-aminosalicylate are powerful scavengers of the hydroxyl radical, determined by pulse radiolysis and confirmed by assays based on deoxyribose degradation by hydroxyl radicals. 5-Aminosalicylate can also protect alpha 1-antiprotease against attack by the myeloperoxidase-derived oxidant hypochlorous acid. The ability to scavenge oxidants produced at sites of inflammation may contribute to the anti-inflammatory action of sulphasalazine and its metabolites.
    • SCE induction and harlequin staining in mycoplasma-contaminated Chinese hamster cells.

      White, Gavin R M; Ockey, Charles H; Paterson Laboratories, Christie Hospital and Holt Radium Institute, Withington, Manchester, M20 9BX, Great Britain (1985)
      Chinese hamster V79 and CHO cells infected with Mycoplasma hyorhinis show elevated sister-chromatid exchange (SCE) levels but normal cell proliferation and levels of chromosomal aberrations when compared with uninfected cells. Harlequin staining patterns differ from those seen with uninfected cells at similar levels of bromodeoxyuridine (BrdUrd), indicating that BrdUrd is rapidly depleted from the medium by the mycoplasmal uridine phosphorylase and therefore becomes unavailable over the two cell cycles necessary for harlequin staining. Continuous treatment with the antibiotic minocycline restores the SCE level and harlequin staining to that seen in uncontaminated cells. The results suggest that mycoplasma infection should be suspected if harlequin staining patterns indicate a sudden decrease in incorporation of BrdUrd in cells grown in normal levels of BrdUrd.