• D-Cycloserine destruction by alanine racemase and the limit of irreversible inhibition

      de Chiara C; Homsak M; Prosser GA, Douglas HL; Garza-Garcia A, Kelly G; Purkiss AG; Tate EW; de Carvalho LPS; Mycobacterial Metabolism and Antibiotic Research Laboratory, The Francis Crick Institute, London, UK. (2020)
    • Damage-induced apoptosis in intestinal epithelia from bcl-2-null and bax-null mice: investigations of the mechanistic determinants of epithelial apoptosis in vivo.

      Pritchard, D Mark; Potten, Christopher S; Korsmeyer, Stanley J; Roberts, Stephen A; Hickman, John A; CRC Molecular and Cellular Pharmacology Group, School of Biological Sciences, Stopford Building (G38), University of Manchester, Manchester M13 9PT, UK. (1999-12-02)
      The influence of bcl-2 and bax expression on apoptotic cell death in mouse intestinal epithelia was assessed using homozygously null mice. Apoptosis was induced in vivo by the enterotoxin 5-fluorouracil (5FU) or by gamma-irradiation and its cell positional incidence was assessed. 5FU and gamma-radiation treated bax-null mice surprisingly showed no reductions in apoptotic yield in the small intestine or midcolon at 4.5 h at cell positions in which both agents had previously been shown to strongly induce p53 protein expression. The colonic epithelia of 5FU treated bcl-2-null mice showed elevated levels of apoptosis at 4.5 h: from 48 apoptotic events in wild-type mice to 273 in the nulls, scoring 200 half crypts. The increase occurred specifically in the cell positions considered to harbour colonic stem cells, at the base of crypts, where there is selective expression of bcl-2. There was a modest but significant increase in apoptosis in the small intestine of the bcl-2-null mice although the epithelia of wild-type mice here are not immunohistochemically positive for bcl-2 protein. These findings show that bcl-2 plays a key role in determining the sensitivity of colonic stem cells to damage-induced death but that bax is not responsible for the p53-dependent induction of apoptosis in this context.
    • DCE-MRI biomarkers in the clinical evaluation of antiangiogenic and vascular disrupting agents.

      O'Connor, James P B; Jackson, Alan; Parker, Geoff J M; Jayson, Gordon C; Imaging Science and Biomedical Engineering, University of Manchester, Oxford Road, Manchester M13 9PT, UK.james.o'connor@manchester.ac.uk (2007-01-29)
      Dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) is now frequently used in early clinical trial assessment of antiangiogenic and vascular disrupting compounds. Evidence of drug efficacy and dose-dependent response has been demonstrated with some angiogenesis inhibitors. This review highlights the critical issues that influence T(1)-weighted DCE-MRI data acquisition and analysis, identifies important areas for future development and reviews the clinical trial findings to date.
    • Death of intestinal crypts and of their constituent cells after treatment by chemotherapeutic drugs.

      Moore, James V; Paterson Laboratories, CHristie Hospital and Holt Radium Institute, Manchester M20 9BX (1984-01)
      The number and spatial distribution of necrotic cells in the jejunal crypts of mice, has been measured after treatment by each of 6 cytotoxic drugs. At the LD10/8 day dose of each drug, the majority of necrotic cells were found below position 9 and numbers per crypt were similar for all drugs (approximately 8). These findings resemble those for radiation. However, major differences between agents were found in the calculated numbers of the microcolony-forming units (MFU) that determine overall crypt survival or ablation after high doses of cytotoxic agent. Numbers of MFU as assayed by radiation were approximately 80 per crypt, but only 2 when assayed by mechlorethamine hydrochloride, adriamycin and 5-fluorouracil, and 7 using BCNU. No crypts were destroyed by either cyclophosphamide or actinomycin D, despite the appearance of numerous necrotic cells in the lower part of the crypt. We conclude that in drug-treated intestine, necrotic cells may arise from a non-MFU compartment and the incidence and distributions of such cells are likely to be poor indicators of the response of the MFU.
    • The decay of autoradiographic grain number over crypt base columnar cells in murine ileum as a measure of their generation time.

      Chwalinski, S; Potten, Christopher S; Department of Pathophysiology, Institute of Rheumatology, Warsaw, Poland. (1991-01)
      The decay in the number of grains over [3H]-thymidine labelled crypt base columnar cells (BCC) in autoradiographs of the ileum of BDF1 mice has been studied. The results revealed that using the conventional grain count halving (GCH) method it is possible to obtain an estimation of the generation time (Tc) of the proliferative BCC cells in the Paneth cell zone (PC-zone) of 18.8 +/- 0.74 h. This lies within the range obtained by the percent labelled mitoses (PLM) method, but is shorter than most values obtained by stathmokinetic methods. The present data show no evidence for a shortening of the cell cycle 3 days after irradiation (8 Gy) which is contrary to some earlier observations. Some reasons for this discrepancy are discussed. The comparatively high labelling index of the BCC allows a larger amount of data to be easily collected, compared with the PLM technique, and correction factors which take into account the complicated shape of the bottom of the crypt are not required.
    • Decoding the interdependence of multiparametric magnetic resonance imaging to reveal patient subgroups correlated with survivals

      Li, C; Wang, S; Liu, P; Torheim, Turid; Boonzaier, NR; van Dijken, BR; Schonlieb, CB; Markowetz, F; Price, SJ; Cambridge Brain Tumor Imaging Laboratory, Division of Neurosurgery, Department of Clinical Neurosciences, University of Cambridge, Addenbrooke's Hospital, Cambridge, UK (2019)
      Glioblastoma is highly heterogeneous in microstructure and vasculature, creating various tumor microenvironments among patients, which may lead to different phenotypes. The purpose was to interrogate the interdependence of microstructure and vasculature using perfusion and diffusion imaging and to investigate the utility of this approach in tumor invasiveness assessment. A total of 115 primary glioblastoma patients were prospectively recruited for preoperative magnetic resonance imaging (MRI) and surgery. Apparent diffusion coefficient (ADC) was calculated from diffusion imaging, and relative cerebral blood volume (rCBV) was calculated from perfusion imaging. The empirical copula transform was applied to ADC and rCBV voxels in the contrast-enhancing tumor region to obtain their joint distribution, which was discretized to extract second-order features for an unsupervised hierarchical clustering. The lactate levels of patient subgroups, measured by MR spectroscopy, were compared. Survivals were analyzed using Kaplan-Meier and multivariate Cox regression analyses. The results showed that three patient subgroups were identified by the unsupervised clustering. These subtypes showed no significant differences in clinical characteristics but were significantly different in lactate level and patient survivals. Specifically, the subtype demonstrating high interdependence of ADC and rCBV displayed a higher lactate level than the other two subtypes (P?=?.016 and P?=?.044, respectively). Both subtypes of low and high interdependence showed worse progression-free survival than the intermediate (P?=?.046 and P?=?.009 respectively). Our results suggest that the interdependence between perfusion and diffusion imaging may be useful in stratifying patients and evaluating tumor invasiveness, providing overall measure of tumor microenvironment using multiparametric MRI.
    • Decoding the regulatory network of early blood development from single-cell gene expression measurements.

      Moignard, V; Woodhouse, S; Haghverdi, L; Lilly, A J; Tanaka, Y; Wilkinson, A; Buettner, F; Macaulay, I; Jawaid, W; Diamanti, E; et al. (2015-02-09)
      Reconstruction of the molecular pathways controlling organ development has been hampered by a lack of methods to resolve embryonic progenitor cells. Here we describe a strategy to address this problem that combines gene expression profiling of large numbers of single cells with data analysis based on diffusion maps for dimensionality reduction and network synthesis from state transition graphs. Applying the approach to hematopoietic development in the mouse embryo, we map the progression of mesoderm toward blood using single-cell gene expression analysis of 3,934 cells with blood-forming potential captured at four time points between E7.0 and E8.5. Transitions between individual cellular states are then used as input to develop a single-cell network synthesis toolkit to generate a computationally executable transcriptional regulatory network model of blood development. Several model predictions concerning the roles of Sox and Hox factors are validated experimentally. Our results demonstrate that single-cell analysis of a developing organ coupled with computational approaches can reveal the transcriptional programs that underpin organogenesis.
    • Decrease of pro-angiogenic monocytes predicts clinical response to anti-angiogenic treatment in patients with metastatic renal cell carcinoma (mRCC)

      Oudard, S; Benhamouda, N; Escudier, B; Ravel, P; Kothari, D; Mehmud, F; Levionnois, E; Sevin, E; Negrier, S; Barthelemy, P; et al. (2015)
    • Decreased expression of Yes-associated protein is associated with outcome in the luminal A breast cancer subgroup and with an impaired tamoxifen response.

      Lehn, S; Tobin, N; Sims, A; Stål, O; Jirström, K; Axelson, H; Landberg, Göran; Center for Molecular Pathology, Department of Laboratory Medicine, Lund University, Skåne University Hospital, 205 02 Malmö, Sweden (2014)
      Yes-associated protein (YAP1) is frequently reported to function as an oncogene in many types of cancer, but in breast cancer results remain controversial. We set out to clarify the role of YAP1 in breast cancer by examining gene and protein expression in subgroups of patient material and by downregulating YAP1 in vitro and studying its role in response to the widely used anti-estrogen tamoxifen.
    • Decreasing sensitivity to cytotoxic agents parallels increasing tumorigenicity in human fibroblasts.

      Kinsella, Anne R; Haran, M Sally; Paterson Institute for Cancer Research, Christie Hospital and Holt Radium Institute, Manchester, United Kingdom. (1991-04-01)
      Human embryo fibroblasts of common genetic origin but exhibiting a range of phenotypes from normal to aggressively tumorigenic have been used to study resistance to the cytotoxic drugs methotrexate and N-(phosphonacetyl)-L-aspartate. Measurement of the intrinsic sensitivities of these cells to the two drugs in standard survival assays, in normal fetal bovine serum, showed increasing resistance to parallel increasing tumor-igenicity. Tumor cells were totally resistant to 10 mM N-(phosphonacetyl)-L-aspartate whereas the 50% lethal dose for methotrexate for the tumor cells was 500 nM compared with 50 nM for the normal diploid parent cell line. The difference in resistance between the immortal and tumorigenic cell lines was eliminated for both methotrexate and N-(phosphonacetyl)-L-aspartate, when the experiments were repeated in the presence of dialyzed fetal bovine serum, but could be restored by the addition of either hypoxanthine (100 microM) or uridine (10 microM). This suggested an important role for the salvage pathways of purine and pyrimidine biosynthesis in the increased resistance of the more tumorigenic cell lines. The implications of these data in relation to cancer chemotherapy will be discussed.
    • Deduction of the clonogen content of intestinal crypts: a direct comparison of two-dose and multiple-dose methodologies.

      Roberts, Stephen A; Hendry, Jolyon H; Potten, Christopher S; Cancer Research Campaign, Biomathematics and Computing Unit, Paterson Institute for Cancer Research, Christie Hospital (NHS) Trust, Manchester, United Kingdom. (1995-03)
      A microcolony assay was used in conjunction with fractionated gamma irradiation to determine the number of clonogens in murine intestinal crypts with varying doses of irradiation used in the determination. The experimental design allows direct comparison between two-dose methodologies, employing one and two (or two or four) equal dose fractions, and multiple-dose methodologies involving determination of the crypt survival curves for a number of fractionation regimens using equal doses per fraction. The two-dose methodology yielded estimates of clonogen number of between 3 and 4 at low delivered dose (single and double fractions each of 6.5-7.5 Gy), rising to around 40 at high biological doses (two and four fractions each of 5.75 or 6.5 Gy). The multifraction methodology yielded estimates of clonogen number which increased from 13 after a single fraction to values of 26 and 22 after three and four fractions. However, the latter values were reduced to 11 and 9, and showed little evidence of any dependence on fraction number, when data pertaining to high biologically effective doses were excluded. Hence it is concluded that the high values for clonogen number typically deduced from such multiple-dose protocols, compared with the generally lower (but dose-dependent) values obtained from two-dose protocols, may be explained at least partially by the higher biological doses generally employed in the multiple-dose protocols.
    • A deep learning framework for predicting response to therapy in cancer

      Sakellaropoulos, T; Vougas, K; Narang, S; Koinis, F; Kotsinas, A; Polyzos, A; Moss, TJ; Piha-Paul, S; Zhou, H; Kardala, E; et al. (2019)
      A major challenge in cancer treatment is predicting clinical response to anti-cancer drugs on a personalized basis. Using a pharmacogenomics database of 1,001 cancer cell lines, we trained deep neural networks for prediction of drug response and assessed their performance on multiple clinical cohorts. We demonstrate that deep neural networks outperform the current state in machine learning frameworks. We provide a proof of concept for the use of deep neural network-based frameworks to aid precision oncology strategies.
    • Defective NOTCH signalling drives smooth muscle cell death and differentiation in bicuspid aortic valve aortopathy.

      Harrison, OJ; Torrens, C; Salhiyyah, K; Modi, A; Moorjani, N; Townsend, Paul A; Ohri, SK; Cagampang, F; Institute of Developmental Sciences, Faculty of Medicine, University of Southampton, Southampton, UK (2019)
      OBJECTIVES: Bicuspid aortic valve disease is common and is associated with ascending aortic aneurysms. Vascular smooth muscle cell (VSMC) apoptosis is characteristic of the ascending aorta of bicuspid patients, and NOTCH1 gene mutations have also been linked to the disease. NOTCH signalling is a fundamental cell signalling pathway, which dictates cell fate decisions including apoptosis. Our objective was to elucidate the role of NOTCH signalling in VSMC apoptosis and differentiation in bicuspid aortopathy. METHODS: Ascending aortic biopsies were obtained from 19 bicuspid and 12 tricuspid aortic valve patients and were sub-classified into 4 groups according to the maximum ascending aortic diameter (aneurysmal ??45?mm). Apoptotic VSMCs were counted by light microscopy using a TUNEL assay. Gene expression of key regulators of NOTCH signalling (NOTCH1 and HES1), apoptosis (BAX and BCL-2) and VSMC differentiation (MYH11, CNN1 and MYH10) were quantified using quantitative real-time PCR. Primary VSMCs were cultured from 2 tricuspid aortic valve and 2 bicuspid aortic valve patients, NOTCH signalling was inhibited with N-[N-(3,5-Difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester, and the gene expression was again quantified. RESULTS: The apoptotic cell count was significantly higher in bicuspid aortic valve patients (3.2 cells/50 000??m2 vs 1.1 cells/50 000??m2; P?=?0.033). There was a trend towards lower apoptotic cell count in the aneurysmal versus non-aneurysmal tricuspid and bicuspid groups and an increased ratio of proapoptotic gene expression, which was not statistically significant. This was associated with a 2.8-fold increase in contractile gene expression (P?=?0.026) and a 2.0-fold increase in NOTCH signalling gene expression in bicuspid versus tricuspid aortic valve patients (P?=?0.022). NOTCH inhibition in cultured VSMCs induced a similar pattern of increased proapoptotic and procontractile gene expressions. CONCLUSIONS: This preliminary study suggests that NOTCH activation in the non-aneurysmal bicuspid aortas may underlie aortopathy by influencing VSMC apoptosis and differentiation. NOTCH signalling manipulation may provide a therapeutic target for preventing aneurysms in bicuspid patients. Further studies with larger sample sizes are needed to substantiate the present findings.
    • Defective responses of transformed keratinocytes to terminal differentiation stimuli. Their role in epidermal tumour promotion by phorbol esters and by deep skin wounding.

      Parkinson, Eric K; Department of Epithelial Kinetics, Paterson Laboratories, Christie Hospital and Holt Radium Institute, Wilslow Road, Manchester M20 9BX, UK. (1985-10)
      Epidermal tumourigenesis can be achieved in rodents by the application of a single subthreshold dose of a carcinogen (initiation) followed by repeated applications of a tumour promoter such as 12-0-tetradecanoyl phorbol, 13-acetate (TPA). TPA induces terminal differentiation in the majority of epidermal keratinocytes in vitro. However, transformed keratinocytes respond weakly to this terminal differentiation signal, and it is suggested that this property allows initiated cells and their progeny to obtain a selective advantage over their normal counterparts during promotion of papilloma formation by TPA. New data are reviewed which suggest that a putative wound hormone TGF-beta has similar differential effects on normal and transformed epithelial cells to those of TPA. It is proposed that the release of TGF-beta from platelets following deep skin wounding may be an explanation as to why wounding is a promoting stimulus but milder forms of epidermal injury are not. Weakly promoting hyperplasiogenic agents are also discussed within the context of a selection theory of tumour promotion.
    • Defining the myofibroblast: normal tissues, with special reference to the stromal cells of Wharton's jelly in human umbilical cord.

      Eyden, Brian P; Ponting, J; Davies, H; Bartley, C; Torgersen, E; Department of Histopathology, Christie Hospital National Health Service Trust, Manchester, UK. (1994-07)
      Cells differing widely in tissue distribution, immunophenotype and ultrastructure have been described as myofibroblasts. The definition of the myofibroblast was analysed as applied to normal tissues, with original observations on Wharton's jelly stromal cells as an example. Stromal cells in Wharton's jelly were studied by conventional histology, immunohistochemistry, and electron microscopy. The normal architecture of the cord was confirmed by light microscopy. Stromal cells and the smooth-muscle cells of the umbilical vessels were positive for vimentin, desmin and alpha-smooth muscle actin, while only the stromal cells were positive for prolyl 4-hydroxylase. Electron microscopy revealed variable but sometimes only moderate amounts of rough endoplasmic reticulum, bundles of smooth-muscle type filaments with focal densities, a large Golgi apparatus with collagen secretion granules, lipid and glycogen. There was no convincing evidence for either lamina or fibronexus junctions. The nature of the stromal cell was discussed in the light of these findings. It was concluded that a myofibroblastic designation was inappropriate and that these cells had phenotypic similarities to vascular smooth muscle cells. The possibility is proposed that most examples of spindle cells cited in the literature as being myofibroblasts and seen in normal tissues not subjected to trauma or showing pathology may be pericytic or smooth-muscle in nature.
    • The degradation of 5-iododeoxyuridine and 5-bromodeoxyuridine by serum from different sources and its consequences for the use of the compounds for incorporation into DNA.

      Saffhill, Roy; Hume, W J; Paterson Laboratories, Christie Hospital, Manchester M20 9BX UK (1986-03)
      Enzymes are present in sera that convert 5-iododeoxyuridine (IdU) to deoxyuridine (dU) and 5-iodouracil (IU). Although in the presence of serum 5-bromodeoxyuridine (BrdU) is not subject to extensive debromination it is converted to 5-bromouracil (BrU) at approx. 50% of the rate for IdU. These conversions are likely brought about by the enzymes thymidylate synthetase and thymidine phosphorylase. In vivo and in culture the dU enters DNA as thymidine 5'-monophosphate (dTMP) via the de novo pathway. Deoxyuridine is often found as a contaminant of [3H]IdU and [3H]BrdU. For these reasons, complications can arise in the interpretation of experimental work using these radioactive compounds. The problems may be overcome by purifying the compounds by high performance liquid chromatography (HPLC) before use together with identification of the DNA components with which the 3H is associated by chromatographic analysis.
    • Degradation of adriamycin in aqueous sodium hydroxide: formation of a ring-A oxabicyclononenone

      Abdeen, Z; Bruce, J Malcolm; Guyan, Patricia M; Land, Edward J; Mukherjee, Tulsi; Univ., Dep. Chemistry, Manchester M13 9PL, Royaume-Uni (1985)
    • Deja Vu: EGF Receptors Drive Resistance to BRAF Inhibitors.

      Girotti, Maria Romina; Marais, Richard; Molecular Oncology Group, The Paterson Institute for Cancer Research, The University of Manchester, Manchester, United Kingdom. (2013-05)
      Summary: The promise of personalized medicine is upon us, and in some cancers, targeted therapies are rapidly becoming the mainstay of treatment for selected patients based on their molecular profile. The protein kinase BRAF is a driver oncogene in both thyroid cancer and melanoma, but while drugs that target BRAF and its downstream signaling pathway are effective in melanoma, they are ineffective in thyroid cancer. In this issue of Cancer Discovery, Montero-Conde and colleagues investigate why thyroid cancer is resistant to BRAF inhibitors despite the presence of BRAF mutation. Cancer Discov; 3(5); 487-90. ©2013 AACR.
    • The delay before onset of accelerated tumour cell repopulation during radiotherapy: a direct maximum-likelihood analysis of a collection of worldwide tumour-control data.

      Roberts, Stephen A; Hendry, Jolyon H; Department of Biomathematics and Computing, Paterson Institute for Cancer Research, Christie Hospital (NHS) Trust Withington, Manchester, UK. (1993-10)
      The worldwide collection of control data for head and neck tumours presented by Withers et al. (Withers, H.R., Taylor, J.M.G. and Maciejewski, B. Acta Oncol. 27: 131-146, 1988) was reanalysed using a model which includes an explicit lag phase before the onset of tumour clonogen repopulation. A direct maximum-likelihood approach was used and the methodology extended to include the computation of profile-likelihood confidence limits. A statistically significant (p = 0.02) lag of 29 days was obtained with 95% confidence limits covering the range 17-31 days. However, the confidence interval was disconnected, and excluded the period 21-23 days. The analysis gave a time factor of 0.66 Gy/day. The mean values confirm the conclusions drawn by the original authors using a two-stage (indirect) method, and the values are similar to those calculated here for another data set comprising 496 patients (lag period = 26 (19-33) days). However, the data set itself is retrospective, and potentially subject to a number of biases. Therefore any clinical conclusions can only be tentative. A new feature of the methodology is the computation of profile-likelihood confidence limits and this will be useful in future direct analyses of clinical data of this type. The more usually computed normal approximation to the confidence limits have been shown to be inadequate in this analysis, and either profile-likelihood limits or likelihood ratio tests must be employed to determine the significance of the model parameters.