• Deriving absolute values of alpha and beta for dose fractionation, using dose-incidence data.

      Hendry, Jolyon H; Moore, James V; Radiobiology Section, Paterson Laboratories, CHristie Hospital and Holt Radium Institute, Manchester M20 9BX (1985-09)
      A method is described for calculating absolute "operational" values of the parameters alpha and beta that characterise dose fractionation data for tissues, by using the steepness of dose-incidence curves measured around each dose per fraction investigated. The values are deduced from published mouse lethality data after irradiation of the bone marrow (alpha = 0.9 Gy-1; beta = 0.06 Gy-2), the lung (alpha = 0.14 Gy-1; beta = 0.02 Gy-2), and the oesophagus (alpha = 0.06 Gy-1; beta = 0.004 Gy-2). With bone marrow and other hierarchical renewal tissues, the operational values apply also to the inactivation of target cells which are the stem cells in the tissue. For other tissue types the interpretation of the values is unknown. The operational values are useful in characterising the steepness of dose-incidence curves for normal tissue injury after different fractionation schedules.
    • Deriving cell survival curves from the overall responses of irradiated tumours: analysis of published data for tumour spheroids.

      Moore, James V; West, Catharine M L; Hendry, Jolyon H; Department of Radiobiology, Paterson Institute for Cancer Research, Christie Hospital and Holt Radium Institute, Manchester, UK. (1987-09)
      Curves of growth delay (GD) or 'cure' after graded doses of radiation have been analysed for 16 lines of human and animal tumours grown as multicellular spheroids in vitro. Dose-survival curves were derived for those cellular units from which spheroids regrow after unsuccessful irradiation (spheroid-regenerating cellular units, SRU). For 10 sets of data from 6 spheroid lines, the Do's and extrapolation numbers of the SRU derived by GD could be compared with the response of the clonogenic cells of the spheroids. For Do, a good correlation (r = 0.910) was found between the two; this was true also for Do derived from curves of spheroid 'cure' (7 sets of data from 6 spheroid lines) and clonogenic cells (r = 0.986). Using GD, the correlation of extrapolation numbers was less good (r = 0.682), the values for SRU commonly being higher than those for clonogenic cells. This may reflect features of the growth curves of spheroids after the lower range of doses of radiation. For human and animal tumour spheroids of 250 microns or less, derived Do ranged from 0.5 to 2.5 Gy. For spheroids of 350 microns or more, derived Do for animal tumour lines ranged from 3.4 to 4.2 Gy, for human lines from 1.5 to 2.1 Gy.
    • Description and basic cell kinetics of the murine pericryptal fibroblast sheath.

      Neal, J Valerie; Potten, Christopher S; Paterson Laboratories, Christie Hospital, and Holt Radium Institute, Manchester (1981-01)
      The size of the pericryptal fibroblast sheath (PCFS) population was determined by scoring serial sections. There are 38 and 124 PCFS cells per murine small intestinal and colonic crypt, respectively. The cells of the PCFS are slightly weighted towards the lower two-thirds of the crypt in their distribution. The ratio of epithelial cells to PCFS cells is approximately 6.5:1. The in vivo cell kinetics were analysed under control and stressed (fasted-refed) conditions. The control labelling index increases from 8.9% in the small intestine and 7.0% in the colon to peaks 49% and 113% respectively above these values 24 hours after 3HTdR administration. Labelling is observed at all crypt levels equally, and no evidence of vertical migration of labelled PCFS cells was found. Colonic epithelial and PCFS cells show a similar pattern of response to feeding after a fast of 72 hours with respect to time, but a different distribution of response in terms of crypt position.
    • Design, implementation and operation of a reading center platform for clinical studies.

      Clin, L; Leitritz, M; Dietter, J; Dynowski, Marek; Burgert, O; Ueffing, M; Thies, C; School of Informatics, Reutlingen University, Germany (2017)
      Clinical reading centers provide expertise for consistent, centralized analysis of medical data gathered in a distributed context. Accordingly, appropriate software solutions are required for the involved communication and data management processes. In this work, an analysis of general requirements and essential architectural and software design considerations for reading center information systems is provided. The identified patterns have been applied to the implementation of the reading center platform which is currently operated at the Center of Ophthalmology of the University Hospital of Tübingen.
    • Desmosomal form, fate, and function in mammalian epidermis.

      Allen, Terence D; Potten, Christopher S; Paterson Laboratories, Christie Hospital and Holt Radium Institute, Manchester (1975-04)
    • Destabilization of CHK2 by a missense mutation associated with Li-Fraumeni Syndrome.

      Lee, S B; Kim, S H; Bell, D W; Wahrer, D C; Schiripo, T A; Jorczak, M M; Sgroi, D C; Garber, J E; Li, F P; Nichols, K E; et al. (2001-11-15)
      Li Fraumeni Syndrome (LFS) is a multicancer phenotype, most commonly associated with germ-line mutations in TP53. In a kindred with LFS without an inherited TP53 mutation, we have previously reported a truncating mutation (1100delC) in CHK2, encoding a kinase that phosphorylates p53 on Ser(20). Here, we describe a CHK2 missense mutation (R145W) in another LFS family. This mutation destabilizes the encoded protein, reducing its half-life from >120 min to 30 min. This effect is abrogated by treatment of cells with a proteosome inhibitor, suggesting that CHK2(R145W) is targeted through this degradation pathway. Both 1100delC and R145W germ-line mutations in CHK2 are associated with loss of the wild-type allele in the corresponding tumor specimens, and neither tumor harbors a somatic TP53 mutation. Our observations support the functional significance of CHK2 mutations in rare cases of LFS and suggest that such mutations may substitute for inactivation of TP53.
    • Destructive and topical treatments of skin lesions in organ transplant recipients and relation to skin cancer

      Green, Adèle C; Way, M.; Oster, M.; Plasmeijer, E. I.; Jiyad, Z.; O'Rourke, P.; Miura, K.; Campbell, S.; Isbel, N.; Chambers, D. C.; et al. (2020)
      Various treatments of keratotic skin lesions and early skin cancers are performed in organ transplant recipients (OTRs) at high risk of skin malignancies but the frequency of their use is unknown. We prospectively assessed the frequency of use of cryotherapy, diathermy, and topical therapies and also investigated their associations with background incidence of histologically-confirmed squamous-cell carcinoma (SCC) and basal cell carcinoma (BCC) in a cohort of OTRs in Queensland, Australia. Median follow-up ranged from 1.7 to 3.2 years across organ transplant groups. Among 285 kidney, 125 lung and 203 liver transplant recipients [382 (62%) male, 380 (62%) immunosuppressed > 5 years, 394 (64%) previously diagnosed with skin cancer], 306 (50%) reported treatment of skin lesions with major types of non-excision therapies during follow-up: 278 (45%) cryotherapy or diathermy; 121 (20%) topical treatments. Of these 306, 150 (49%) developed SCC at double the incidence of those who did not receive these treatments, as assessed by incidence rate ratio (IRR) adjusted for age, sex, type of organ transplant, skin color and history of skin cancer at baseline, calculated by multivariable Poisson regression (IRRadj = 2.1, 95% confidence interval (CI) 1.4-3.1). BCC incidence was not associated with these therapies. Skin lesions in OTRs that are treated with cryotherapy, diathermy, or topical treatment warrant judicious selection and careful follow-up.
    • A detailed mammosphere assay protocol for the quantification of breast stem cell activity.

      Shaw, Frances L; Harrison, Hannah; Spence, Katherine; Ablett, Matthew P; Simões, Bruno M; Farnie, Gillian; Clarke, Robert B; Breast Biology Group, School of Cancer and Enabling Sciences, Paterson Institute for Cancer Research, The University of Manchester, Wilmslow Road, Manchester, UK. (2012-06)
      Since the discovery that neural tissue contains a population of stem cells that form neurospheres in vitro, sphere-forming assays have been adapted for use with a number of different tissue types for the quantification of stem cell activity and self-renewal. One tissue type widely used for stem cell investigations is mammary tissue, and the mammosphere assay has been used in both normal tissue and cancer. Although it is a relatively simple assay to learn, it can be difficult to master. There are methodological and analytical aspects to the assay which require careful consideration when interpreting the results. We describe here a detailed mammosphere assay protocol for the assessment of stem cell activity and self-renewal, and discuss how data generated by the assay can be analysed and interpreted.
    • Detailed mapping and loss of heterozygosity analysis suggests a suppressor locus involved in sporadic breast cancer within a distal region of chromosome band 17p13.3.

      Stack, Maria; Jones, David; White, Gavin R M; Liscia, D S; Venesio, T; Casey, G; Crichton, D; Varley, Jennifer; Mitchell, Erika L D; Heighway, Jim; et al. (1995-11)
      The chromosome region 17p13.3 is thought to encode a tumour suppressor gene involved in sporadic breast cancer and other malignancies. Physical ordering of markers has been carried out by a series of multicolour fluorescent in situ hybridisation (FISH) experiments, using isolated yeast artificial chromosomes (YACs) and cosmids. Eight polymorphic markers ordered within this new physical map and one external marker were used to investigate the pattern of loss of heterozygosity in a panel of 40 sporadic breast tumour patients. The data revealed a region of high loss (60%) within distal 17p13.3, defined by markers D17S926, D17S695 and D17S849 which mapped close together. A contig of YACs was constructed physically linking these three markers.
    • A detailed study of loss of heterozygosity on chromosome 17 in tumours from Li-Fraumeni patients carrying a mutation to the TP53 gene.

      Varley, Jennifer; Thorncroft, Mary R; McGown, Gail; Appleby, J; Kelsey, Anna M; Tricker, K J; Evans, D Gareth R; Birch, Jillian M; CRC Department of Cancer Genetics, Paterson Institute for Cancer Research, Manchester, UK. (1997-02-20)
      We have studied a total of 36 tumours from 28 patients with germline mutations to the TP53 gene for loss of heterozygosity at TP53 using techniques of both direct sequencing and restriction fragment length polymorphism analysis. All patients were from families conforming to the definition of classical Li-Fraumeni syndrome (LFS) or were Li-Fraumeni-like (LFL). The data we have obtained show that loss of the wild-type TP53 gene is observed in under half (44%) of all tumours, and that the pattern of LOH at TP53 may be mutation specific. LOH has been observed in premalignant as well as invasive tumours. Two tumours (6%) show loss of the mutant allele and retention of the wild-type. To confirm that TP53 is indeed the target for LOH events on chromosome 17, we have used additional microsatellite repeats to examine patterns of allelic imbalance along the length of chromosome 17. Data from this analysis indicate that TP53 is the target of loss, but reveal some other interesting patterns of allelic imbalance at other loci on chromosome 17.
    • Detecting gas-induced vasomotor changes via blood oxygenation level-dependent contrast in healthy breast parenchyma and breast carcinoma.

      Wallace, T; Patterson, A; Abeyakoon, O; Bedair, R; Manavaki, R; McLean, M; O'Connor, James P B; Graves, M; Gilbert, F; Department of Radiology, University of Cambridge, Cambridge Biomedical Campus, Cambridge, UK (2016-02-21)
      To evaluate blood oxygenation level-dependent (BOLD) contrast changes in healthy breast parenchyma and breast carcinoma during administration of vasoactive gas stimuli.
    • Detection by DNA polymerase I of breaks produced in rat liver chromatin in vivo by alklating agents

      Saffhill, Roy; Cooper, Helen K; Itzhaki, Ruth F; Paterson Laboratories, Christie Hospital and Holt Radium Institute, Manchester (1974)
    • Detection of acrolein and crotonaldehyde DNA adducts in cultured human cells and canine peripheral blood lymphocytes by 32 P-postlabeling and nucleotide chromatography

      Wilson, Vincent L; Foiles, Peter G; Chung, Fung-Lung; Povey, Andrew C; Frank, Anthony A; Harris, Curtis C; Molecular Genetics/Oncology Laboratory, Department of Pathology and The Children's Hospital kempe Research Center, The Children's Hospital and the Department of Pathology, University of Colorado Health Sciences Center, Denver, CO 80218 (1991)
    • The detection of alkylation damage in the DNA of human gastrointestinal tissues.

      Hall, C N; Badawi, A F; O'Connor, Peter J; Saffhill, Roy; Department of Surgery, Wythenshawe Hospital, Manchester. (1991-07)
      Damage arising from putative environmental sources has been found in the DNA of the gastric and colorectal mucosae of patients presenting with gastrointestinal disorders from the South Manchester area. O6-Methylguanine (O6-MeG) in the range 0.010- greater than 0.300 mu moles mole-1 adenine was heterogeneously distributed both between and within individuals. The pattern of alkylation of tissue DNA appears to differ when comparison is made between gastric and colorectal samples. Most of the gastric tumour DNA samples were alkylated (5/6; 0.087 +/- 0.097), whereas the DNA of the associated mucosa was alkylated less frequently (2/7) and to a lesser extent; (0.017 +/- 0.030; P = 0.07). Conversely, colorectal tumour DNA was alkylated infrequently (1/7) and to a lower extent (0.003 +/- 0.007) than the DNA of the adjacent mucosa (8/10 samples alkylated with a mean of 0.083 +/- 0.106; P = less than 0.01), or indeed of any other tissue. Although increased levels of DNA damage in tissue associated with malignant disease have been indicated by independent studies of DNA damage at other cancer sites, significant differences were not observed in the present report, neither was there any suggestion of a relationship with smoking or alcohol consumption. The data provided by this report indicate that exposure to putative environmental alkylating agents occurs in the UK at levels comparable to those previously detected in areas of higher cancer risk. Although we cannot determine the extent to which this DNA damage is attributable to normal background exposures, it is evident that the alkylation of tissue DNA occurs and is not uniform. In conjunction with other reports, therefore these differences may begin to provide indications of mechanisms that could be of relevance in the aetiology of gastrointestinal cancers.
    • Detection of BRAF mutations in the tumour and serum of patients enrolled in the AZD6244 (ARRY-142886) advanced melanoma phase II study.

      Board, Ruth E; Ellison, G; Orr, M C M; Kemsley, K R; McWalter, G; Blockley, L Y; Dearden, S P; Morris, C; Ranson, Malcolm R; Cantarini, M V; et al. (2009-11-17)
      BACKGROUND: This study investigated the potential clinical utility of circulating free DNA (cfDNA) as a source of BRAF mutation detection in patients enrolled into a phase II study of AZD6244, a specific MEK1/2 inhibitor, in patients with advanced melanoma. METHODS: BRAF mutations were detected using Amplification Refractory Mutation System allele-specific PCR. BRAF mutation status was assessed in serum-derived cfDNA from 126 patients enrolled into the study and from 94 matched tumour samples. RESULTS: Of 94 tumour samples, 45 (47.9%) were found to be BRAF mutation positive (BRAF+). Serum-derived cfDNA was BRAF+ in 33 of 126 (26.2%) samples, including in five samples for which tumour data were unavailable. Of BRAF+ tumours, 25 of 45 (55.6%) were BRAF+ in cfDNA. In three cases in which the tumour was negative, cfDNA was BRAF+. Progression-free survival (PFS) of patients with BRAF+ tumour and cfDNA was not significantly different compared with tumour BRAF+ but cfDNA BRAF-negative patients, indicating that cfDNA BRAF detection is not associated with poorer prognosis on PFS in stage III/IV advanced melanoma. CONCLUSIONS: These data demonstrate the feasibility of BRAF mutation detection in cfDNA of patients with advanced melanoma. Future studies should aim to incorporate BRAF mutation testing in cfDNA to further validate this biomarker for patient selection.
    • Detection of circulating anticomplementary factors in chronic lung diseases.

      Hilton, A M; Moore, Michael; Howat, J M T; Paterson Laboratories, Christie Hospital and Holt Radium Institute, Manchester. (1978-02)
    • Detection of concomitant formation of O6-carboxymethyl- and O6-methyl-2'-deoxyguanosine in DNA exposed to nitrosated glycine derivatives using a combined immunoaffinity/HPLC method.

      Harrison, Kathryn L; Jukes, Rebekah; Cooper, Donald P; Shuker, David E; MRC Toxicology Unit, Hodgkin Building, University of Leicester, P.O. Box 138, Lancaster Road, Leicester LE1 9HN, U.K. (1999-01)
      A previous observation that an N-nitroso-N-carboxymethyl derivative reacts with DNA to give both O6-carboxymethyl-2'-deoxyguanosine (O6-CMdGuo) and O6-methyl-2'-deoxyguanosine (O6-MedGuo) [Shuker, D. E. G., and Margison, G. P. (1997) Cancer Res. 57, 366-369] has been confirmed using a range of nitrosated glycine derivatives [N-acetyl-N'-nitroso-N'-prolylglycine (APNG), azaserine (AS), and potassium diazoacetate (KDA)]. In addition, mesyloxyacetic acid (MAA) was also found to give both O6-adducts in DNA. O6-CMdGuo and O6-MedGuo were assessed in enzymatic hydrolysates of treated calf thymus DNA using a combined immunoaffinity/HPLC/fluorescence procedure. The ratio of O6-CMdGuo to O6-MedGuo varied somewhat between the different compounds with APNG giving the most methylation (O6-CM:O6-Me ratio of 10) and AS the least (39), with KDA and MAA giving intermediate amounts (16 and 18, respectively). The formation of O6-MedGuo by the four compounds probably arises through decarboxylation at various stages in the decomposition pathways, but the exact mechanisms remain to be clarified. The formation of O6-MedGuo from reactions of nitrosated glycine derivatives with DNA in vitro may explain the frequent detection of this adduct in human gastrointestinal DNA, as nitrosation of dietary glycine may occur. O6-CMdGuo is likely to be a useful biomarker of this pathway in vivo and has been detected in human tissues.
    • The detection of cyclobutane thymine dimers, (6-4) photolesions and the Dewar photoisomers in sections of UV-irradiated human skin using specific antibodies, and the demonstration of depth penetration effects.

      Chadwick, Caroline A; Potten, Christopher S; Nikaido, O; Matsunaga, T; Proby, C; Young, A R; CRC Department of Epithelial Biology, Paterson Institute for Cancer Research, Christie Hospital NHS Trust, Manchester, UK. (1995-05)
      Ultraviolet irradiation of skin induces various DNA photolesions. Here we demonstrate that irradiation of human buttock skin with 300 nm UVR in situ induces thymine dimers and 6-4 photoproducts. Irradiation with 260 nm immediately followed by UVA (320 nm) induces the Dewar photoisomers of the 6-4 lesions. All three lesions can be detected in methanol-fixed paraffin sections using specific monoclonal antibodies. The sections have been analysed in an automated image analysis system (Discovery) and the level of immuno-DAB-peroxidase measured in individual epidermal cell nuclei as absorption at 460 nm (integrated optical density). The staining patterns with the antibodies showed no detectable change with epidermal depth by eye after 300 nm irradiation, however, the machine detected a fall off with depth of about 2.5% per cell layer. Following irradiation with a shorter wavelength (260 nm) there was a rapid fall off in staining with depth easily detectable by eye and machine (39% per cell layer).
    • Detection of EBV DNA in post-nasal space biopsy tissue from asymptomatic EBV-seropositive individuals.

      Lees, Janice F; Goodeve, A C; Arrand, J E; Ghosh, Anna K; Jones, P H; Arrand, John R; Paterson Institute for Cancer Research, Christie Hospital, Manchester, United Kingdom. (1992-05)
      The association between EBV and nasopharyngeal carcinoma (NPC) has been well documented although the precise role of the virus in the genesis of the tumour is not understood. We undertook this study to examine the prevalence of EBV infection in nasopharyngeal tissue obtained from 33 healthy individuals not considered to be at risk of developing NPC. Using polymerase chain amplification (PCR) and in situ hybridization we have identified EBV DNA in 70% (23/33) of the tissues examined. Our data demonstrate that EBV is present at the site of tumour development in the low-risk population and by inference that the virus is also present before the onset of disease in the high-risk group. This survey supports the concept of NPC pathogenesis as a multifactorial process.
    • Detection of hTERT protein by flow cytometry.

      Ali, A; Chopra, Rajesh; Robertson, J; Testa, Nydia G; Department of Experimental Haematology, Paterson Institute for Cancer Research, Manchester, UK. (2000-12)
      Telomerase is a telomere-specific DNA polymerase consisting of protein and RNA components, which is activated in germline cells and the majority of cancers and serves to counter the consequences of telomere shortening. The protein component, hTERT, is believed to be the catalytic subunit of human telomerase and its expression at the mRNA level correlates well with telomerase activity in vitro. Current techniques for assaying telomerase activity detect only the mean activity in a sample and are unable to isolate specific cell sub-populations. This report describes the development and validation of a cellular, immunofluorescence-based flow cytometry assay that allows detection of intranuclear hTERT while maintaining identifiable cell population characteristics. The assay was shown to be both sensitive to changes in telomerase expression and was semi-quantitative. In both cell line differentiation experiments and in primary cells, a good correlation existed between hTERT expression measured by flow cytometry and telomerase activity detected by the telomeric repeat amplification protocol (TRAP). The method developed offers a quick, simple and reproducible cellular-based assay for hTERT expression. This assay will provide a useful, new tool for future investigations, facilitating the analysis of hTERT expression in mixed cell populations.