• The radial distribution of fibroblastic colony-forming units in mouse femoral marrow.

      Xu, Cheng Xiong; Hendry, Jolyon H (1981-09)
      The distribution of fibroblastic colony forming units (CFU-F) in mouse femoral marrow was investigated using a method which separates bone marrow cells into two regional fractions of varying size. It is shown that the concentration of CFU-F decreases almost linearly from the femoral axis (800-1,000 CFU-F per 10(7) bone marrow cells) to the bone surface (300 CFU-F per 10(7) cells). When irradiated axial or marginal cells were added into cultures as feeder cells, the colony numbers produced by both axial or marginal CFU-F were increased by about 50% and the higher concentration in the axial regions remained. The survival curve for CFU-F in both regions was characterised by a Do value of 1.54 +/- 0.11 Gy and an extrapolation number of 2.60 +/- 0.38. The results are compared to the distributions of haemopoietic stem cells and committed progenitor cells.
    • The radiation chemistry of some platinum-containing radiosensitizers and related compounds.

      Butler, John; Hoey, Brigid M; Swallow, A John (1985-04)
      Oxidation and reduction of cis- and trans-dichlorodiammine platinum II (cis- and trans-PDD), cis-dichlorobis(1-(2-hydroxyethyl)-2-methyl-5-nitroimidazole-N3)-p latinum II (cis-Flap), and cis-dichlorobis(isopropylamine)-trans-dihydroxyplatinum IV (Chip) have been studied using pulse radiolysis. Spectra corresponding to platinum in various oxidation states have been observed and several rate constants have been obtained. Reduction of all the compounds, except cis-Flap, produces species of a lower oxidation state of platinum which subsequently have both chloride ligands replaced. Ultimately, these products disproportionate. In the case of cis-Flap, reduction occurred on the nitroimidazole ligand. This was verified by the absence of platinum metal after disproportionation. Oxidation of all four compounds consists of production of a higher oxidation state of platinum followed by replacement of chloride ligands and finally disproportionation of the products. Only cis-Flap and Chip could be reduced by oxidized DNA bases. The one-electron reduction potential of cis-Flap was found to be -370 +/- 10 mV. trans-Flap had almost the same value. It was not possible to measure the potentials of the other compounds since their ligands were replaced rapidly but it is estimated that the one-electron reduction potentials decrease in the order cis- or trans-Flap greater than Chip greater than cis-PDD greater than trans-PDD.
    • Radiation damage and repair phenomena.

      Fox, Brian W; Lajtha, L G; Paterson Laboratories, Christie Hospital and Holt Radium Institute, Manchester (1973-01)
    • A radiation hybrid panel for human chromosome 6q.

      Orphanos, Vassilis; Greaves, Martin J; Santibanez-Koref, Mauro F; Fox, M; Edwards, Y H; Boyle, John M; Cancer Research Campaign Department of Cancer Genetics, Christie CRC Research Centre, Manchester, UK. (1995-04)
      A panel of 63 radiation-reduced hybrids has been derived from a mouse cell line containing a neo-marked human Chromosome (Chr) 6, primarily to provide a resource for higher resolution localization of new markers. Hybrids were generated with radiation doses of 40-400 Gy, selected in G418, and were shown by PCR to contain the neo gene. PCR was also used to score the retention of 15 loci that map from 6q13 to q25.2 of the current consensus map, plus six other loci assigned to 6q26-q27. An average retention frequency of 27.8% was observed, with the highest frequencies at D6S313 and D6S280 (63.5%) located near the centromere at 6q13, and at D6S283 (68.5%) at 6q16.3-q21, presumably close to the neo integration site. Lowest frequencies (4.8%) were observed for telomeric markers. All markers segregated independently except D6S297 and D6S193. Agreement and some improvement to the current consensus map of 6q was made by mapping 12 loci by the non-parametric statistical method of Falk. In addition, deletion mapping with informative hybrids allowed the ordering of six loci from 6q26 to q27 and permitted some integration of maps of this region.
    • Radiation induced hemopoiesis in adult mouse liver.

      Testa, Nydia G; Hendry, Jolyon H (1977-03)
      Repeated whole-body irradiation of adult mice induced hemopoiesis in liver, as shown by the presence of stem cells (CFUS), progenitor cells of granulocytes and macrophages (CFUC) and foci of granulocytic cells. The largest numbers of CFUS (up to 700) were found 24 to 47 days after four doses of 450 rad x-rays given at 24 day intervals and 15-17 days after 2310 rad gamma radiation given at a low dose-rate (70 rad per day). CFUS were still present (although in smaller numbers) up to 210 days after four doses of 375 rad x-rays or 225 rad neutrons. CFUC were also present in liver after four doses of 450 rad x-rays, but their numbers could not be calculated accurately because of the marked inhibitory effect of liver cells on in vitro colony growth. Irradiation with one dose of 450 rad x-rays did not result in the appearance of CFUS in liver, suggesting that hepatic hemopoiesis can be induced by radiation only after repeated or prolonged bone marrow injury.
    • Radiation response and cure rate of human colon adenocarcinoma spheroids of different size: the significance of hypoxia on tumor control modelling.

      Buffa, Francesca M; West, Catharine M L; Byrne, Karen; Moore, James V; Nahum, Alan E; Department of Physics, Institute of Cancer Research and Royal Marsden NHS Trust, London, England, United Kingdom. fbuffa@icr.ac.uk (2001-03-15)
      PURPOSE: To evaluate the adequacy of a Poisson tumor control probability (tcp) model and the impact of hypoxia on tumor cure. METHODS AND MATERIALS: A human colon adenocarcinoma cell line, WiDr, was grown as multicellular spheroids of different diameters. Measurements were made of cell survival and spheroid cure following 300-kV X-ray external beam irradiation in air and nitrogen. Cell survival data were fitted using a two-compartment and an oxygen diffusion model. Spheroid cure data were fitted using the tcp model. RESULTS: Hypoxia was seen only for spheroids greater than 500 microm in diameter. For small spheroids tcp estimates of radiosensitivity and clonogenic number showed excellent agreement with experimentally derived values. For large spheroids, although tcp estimates of radiosensitivity were comparable with measurements, estimates of the clonogenic number were considerably lower than the experimental count. Reoxygenation of large spheroids before irradiation resulted in the tcp estimates of the number of clonogenic cells agreeing with measured values. CONCLUSIONS: When hypoxia was absent, the tcp model accurately predicted cure from measured radiosensitivity and clonogen number. When hypoxia was present, the number of cells capable of regrowth in situ was considerably lower than the number of clonogenic cells that initially survived irradiation. As this counteracted the decreased radiosensitivity, hypoxia was less important for cure than predicted from cell survival assays. This finding suggests that chronic hypoxia may not limit directly the success of radiation therapy.
    • Radiation response of an intraspecific somatic cell hybrid (L5178Y, X-ray resistant line X L5178Y, X-ray sensitive line).

      Dale, B; Paterson Laboratories, Christie Hospital and Holt Radium Institute, Manchester (1979-08)
    • The radiation sensitivity of the haemopoietic microenvironment--effect of dose rate on ectopic ossicle formation.

      Molineux, Graham; Testa, Nydia G; Hendry, Jolyon H; Schofield, Raymond; Department of Experimental Haematology, Paterson Institute for Cancer Research, Christie Hospital and Holt Radium Institute, Withington, Manchester, U.K. (1987-10)
      The haemopoietic microenvironment (HM) consists of a complex mixture of cellular types and extra-cellular matrix. It is essential for prolonged haemopoiesis in both the normal situation and after bone marrow transplantation. The competence of the HM can be assessed by ectopic grafting of femoral marrow. A complete haemopoietic organ develops at the site of implantation. Stem cells (CFU-S) which inhabit the ossicle formed after ectopic implantation can be measured, to assess the function of the engrafted HM to support haemopoiesis. Using this functional endpoint we have examined the radiation sensitivity of the HM at both high and low dose rates, and conclude that high doses of gamma-irradiation delivered at 4 Gy/min or 0.016 Gy/min have widely different effects on the HM, the former proving much more damaging than the latter.
    • Radiation therapy with tositumomab (B1) anti-CD20 monoclonal antibody initiates extracellular signal-regulated kinase/mitogen-activated protein kinase-dependent cell death that overcomes resistance to apoptosis.

      Ivanov, Andrei; Krysov, Sergei; Cragg, Mark S; Illidge, Timothy M; School of Cancer and Imaging Sciences, CRUK Paterson Institute for Cancer Research, University of Manchester, United Kingdom. (2008-08-01)
      PURPOSE: The use of targeted radiation therapy (RT) in conjunction with anti-CD20 monoclonal antibodies (mAb) delivers high clinical response rates in B-cell lymphomas as part of radioimmunotherapy. The mechanisms underlying these impressive responses, particularly in patients whose lymphomas have become refractory to chemotherapy, are poorly understood. EXPERIMENTAL DESIGN: In this study, we have investigated the signaling pathways and mode of cell death induced in B-cell lymphoma cells after the combination of RT and either type I (rituximab) or type II (tositumomab/B1) anti-CD20 mAb. RESULTS: Increased tumor cell death was observed when RT was combined with tositumomab, but not rituximab. This additive cell death was found to be mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK)-dependent and could be reversed with mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK) inhibitors, as well as small interfering RNA targeting MEK1/2. Furthermore, we found that this increased death was associated with ERK1/2 nuclear accumulation after tositumomab treatment, which was enhanced in combination with RT. Importantly, although Bcl-2 overexpression resulted in resistance to RT-induced apoptosis, it had no effect on the tumor cell death induced by tositumomab plus RT, indicating a nonapoptotic form of cell death. CONCLUSIONS: These findings indicate that RT and type II anti-CD20 mAb combine to stimulate a prodeath function of the MEK-ERK1/2 pathway, which is able to overcome apoptotic resistance potentially explaining the efficacy of this modality in treating patients with chemoresistant disease.
    • A radiation-controlled molecular switch for use in gene therapy of cancer.

      Scott, S D; Marples, B; Hendry, Jolyon H; Lashford, Linda S; Embleton, M J; Hunter, Robin D; Howell, Anthony; Margison, Geoffrey P; Cancer Research Campaign Section of Genome Damage and Repair, Paterson Institute for Cancer Research, Christie Hospital (NHS) Trust, Manchester, UK. (2000-07)
      Ionising radiation induces the expression of a number of radiation-responsive genes and there is current interest in exploiting this to regulate the expression of exogenous therapeutic genes in gene therapy strategies for cancer. However, the radiation-responsive promoters used in these approaches are often associated with low and transient levels of therapeutic gene expression. We describe here a novel radiation-triggered molecular switching device based on promoter elements from the radiation-responsive Egr-1 gene and the cre-LoxP site-specific recombination system of the P1 bacteriophage. Using this system, a single, minimally toxic dose of radiation induced cre-mediated excision of a lox-P flanked stop cassette in a silenced expression vector and this resulted in amplified levels of CMV-promoter-driven expression of the exogenous tumour-sensitising gene, HSV-tk. This strategy could be used in combination with targeted delivery and tumour-specific promoters to elicit the tumour-targeted and prolonged expression of a variety of tumour-sensitising genes and provide an unprecedented level of control and tumour selectivity.
    • Radiation-hypersensitive cells in small intestinal crypts; their relationships to clonogenic cells.

      Ijiri, K; Potten, Christopher S; Paterson Laboratories, Christie Hospital and Holt Radium Institute, Withington, Manchester, M20 9BX, UK. (1986)
    • Radiation-induced G1 arrest is not defective in fibroblasts from Li-Fraumeni families without TP53 mutations.

      Boyle, John M; Greaves, Martin J; Camplejohn, R S; Birch, Jillian M; Roberts, Stephen A; Varley, Jennifer; CRC Section of Molecular Genetics, Paterson Institute for Cancer Research, Christie CRC Research Centre, Manchester, UK. (1999-04)
      Radiation-induced G1 arrest was studied in four classes of early passage skin fibroblasts comprising 12 normals, 12 heterozygous (mut/wt) TP53 mutation-carriers, two homozygous (mut/-) TP53 mutation-carriers and 16 strains from nine Li-Fraumeni syndrome or Li-Fraumeni-like families in which no TP53 mutation has been found, despite sequencing of all exons, exon-intron boundaries, 3' and 5' untranslated regions and promoter regions. In an assay of p53 allelic expression in yeast, cDNAs from these non-mutation strains behaved as wild-type p53. Using two different assays, we found G1 arrest was reduced in heterozygous strains with mis-sense mutations and one truncation mutation, when compared to the range established for the normal cells. Heterozygous strains with mutations at splice sites behaved like normal cells, whilst homozygous (mut/-) strains showed either extremely reduced, or no, arrest. Strains from all nine non-mutation families gave responses within the normal range. Exceptions to the previously reported inverse correlation between G1 arrest and clonogenic radiation resistance were observed, indicating that these phenotypes are not strictly interdependent.
    • Radiation-induced micronuclei in human fibroblasts in relation to clonogenic radiosensitivity.

      O'Driscoll, M C; Scott, David; Orton, C J; Kiltie, Anne E; Davidson, Susan E; Hunter, Robin D; West, Catharine M L; Cancer Research Campaign Section of Genome Damage and Repair, Paterson Institute for Cancer Research, Christie Hospital (NHS) Trust, Manchester, UK. (1998-12)
      As part of our programme for developing predictive tests for normal tissue response to radiotherapy, we have investigated the efficacy of the cytokinesis-block micronucleus (MN) assay as a means of detecting interindividual differences in cellular radiosensitivity. A study was made of nine fibroblast strains established from vaginal biopsies of pretreatment cervical cancer patients and an ataxia telangiectasia (A-T) cell strain. Cells were irradiated in plateau phase, replated and treated with cytochalasin B 24 h later. MN formation was examined 72 h after irradiation as the number of MN in 100 binucleate cells. The method yielded low spontaneous MN yields (<7 per 100 cells), and mean induced MN frequencies after 3.5 Gy varied between cell strains from 18 to 144 per 100 cells. However, in repeat experiments, considerable intrastrain variability was observed (CV = 32%), with up to twofold differences in MN yields, although this was less than interstrain variability (CV = 62%). An analysis was made of the relationship between MN results and previously obtained clonogenic survival data. There was a significant correlation between MN yields and clonogenic survival. However, when the A-T strain was excluded from the analysis, the correlation lost significance, mainly because of one slow-growing strain which was the most sensitive to cell killing but had almost the lowest MN frequency. With current methodology, the MN assay on human fibroblasts does not appear to have a role in predictive testing of normal tissue radiosensitivity.
    • Radiation-induced micronucleus induction in lymphocytes identifies a high frequency of radiosensitive cases among breast cancer patients: a test for predisposition?

      Scott, David; Barber, J B; Levine, Edward; Burrill, W; Roberts, Stephen A; Paterson Institute for Cancer Research, Christie CRC Research Centre, Manchester, UK. (1998-02)
      Enhanced sensitivity to the chromosome-damaging effects of ionizing radiation is a feature of many cancer-predisposing conditions. We previously showed that 42% of an unselected series of breast cancer patients and 9% of healthy control subjects showed elevated chromosomal radiosensitivity of lymphocytes irradiated in the G2 phase of the cell cycle. We suggested that, in addition to the highly penetrant genes BRCA1 and BRCA2, which confer a very high risk of breast cancer and are carried by about 5% of all breast cancer patients, there are also low-penetrance predisposing genes carried by a much higher proportion of breast cancer patients, a view supported by recent epidemiological studies. Ideally, testing for the presence of these putative genes should involve the use of simpler methods than the G2 assay, which requires metaphase analysis of chromosome damage. Here we report on the use of a simple, rapid micronucleus assay in G0 lymphocytes exposed to high dose rate (HDR) or low dose rate gamma-irradiation, with delayed mitogenic stimulation. Good assay reproducibility was obtained, particularly with the HDR protocol, which identified 31% (12 out of 39) of breast cancer patients compared with 5% (2 out of 42) of healthy controls as having elevated radiation sensitivity. In the long term, such cytogenetic assays may have the potential for selecting women for intensive screening for breast cancer.
    • Radiation-induced mitotic delay: duration, dose and cell position dependence in the crypts of the small intestine in the mouse.

      Chwalinski, S; Potten, Christopher S; Paterson Laboratories, Christie Hospital & Holt Radium Institute, Manchester, M20 9BX, U.K. (1986-05)
      The cells of the proliferative compartment in the crypt of the small intestine undergo a step by step differentiation and/or maturation from stem cells to the functional cells on the villi. The consequent hierarchical organization of the proliferative cell population can be related to the actual position of cells within the crypt. The stem cells are found near the bottom of the crypt with the more mature cells occurring at increasingly higher positions. The sensitivity of proliferative cells in the crypt of small intestine to radiation-induced mitotic delay was investigated at each position within the crypt. Using the stathmokinetic method (vincristine accumulation), the following were noted. The yield of mitotic figures 3 h immediately after irradiation showed a strong cell position dependence with the cells at the base of the crypt being most inhibited and those at the top of the proliferative compartment least affected. The mitotic yields were largely unaffected for the first 15 min suggesting that there is a transition point (Tp) for radiosensitivity which is located about 15 min before metaphase for all crypt cells. Cells located less than 15 min from metaphase are unaffected while those more than 15 min from metaphase are inhibited from further cell cycle progression. After this initial delay all proliferative cells were inhibited in their progression through G2 but some recovered more quickly than others. The ratio of the time of division delay (Td) in stem cells to that in cells at the top of the proliferative compartment was about 3:1. In absolute values Td after 1.0 Gy was about 1 h and 2.8 h, for cells at the top of the crypt and at the base, respectively. After 2.5 Gy the corresponding values were less than 3 h and between 5 and 6 h for the mid-crypt and crypt base respectively. There is thus a dependence on dose for the duration of the mitotic inhibition which for the cells at the top of the crypt is similar to the widely quoted average value 1 h per Gy, but the duration depends strongly on cell position. Thus not all proliferative cells respond in the same way. The duration is shorter the closer the proliferative cells are to their last cell division in the proliferative hierarchy in the crypt and longest for cells situated where the stem cells are to be expected.
    • Radiation-induced p53 and p21WAF-1/CIP1 expression in the murine intestinal epithelium: apoptosis and cell cycle arrest.

      Wilson, James W; Pritchard, D Mark; Hickman, John A; Potten, Christopher S; CRC Epithelial Biology Laboratory, Paterson Institute for Cancer Research, Manchester, United Kingdom. jwilson@picr.man.ac.uk (1998-09)
      p53-dependent expression of p21WAF-1/CIP1 has been studied in murine intestinal epithelium after exposure to ionizing radiation. In un-irradiated small intestine, neither p53 nor p21WAF-1/CIP1 could be detected by immunohistochemistry. After irradiation (8 Gy), there was a time- and dose-dependent increase in the expression of both proteins. In the small bowel, the positional expression of p53 and p21WAF-1/CIP1 was similar but not coincident. Both proteins could be observed throughout the crypts with greatest frequency of expression over the first 15 cell positions, which includes the stem cell population (approximately positions 3 to 5) and the proliferating, transit cell population (approximately positions 5 to 15). p53-positive cells were primarily distributed toward the base of the crypt relative to p21WAF-1/CIP1. Subdivision of the p53-positive cell population revealed that the cells with strongest p53 immunoreactivity were positioned farther toward the base of the crypt, and their distribution was approximately coincident with the frequency distribution of apoptotic cells. Cells that were either weakly or moderately immunoreactive for p53 were located toward the middle of the crypt and were approximately coincident with the distribution of p21WAF-1/CIP1. The numbers of both p53- and p21WAF-1/CIP1-positive cells declined steadily with time, and by 6 days after irradiation there were very few immunoreactive cells to observe. Radiation-induced increase in p53 and p21WAF-1/CIP1 expression was not detected in mice homozygously null for p53. Expression of p21WAF-1/CIP1 and incorporation of tritiated thymidine were found to be mutually exclusive. In the large bowel, p21WAF-1/CIP1 and p53 expression were observed along the entire length of the colonic crypts after irradiation (8 Gy), and, unlike in the small intestine, this expression was not only maintained but increased over 72 hours. p21WAF-1/CIP1 immunoreactivity was detected in large intestine epithelium up to 6 days after irradiation. The differential expression of p21WAF-1/CIP1, observed between the large and small bowel and within the small intestinal crypts, is discussed.
    • Radiation-induced reduction reactions with bovine serum albumin.

      Schuessler, H; Davies, J V; Institut fur Radiologie der Universitat Erlangen-Nurnberg, D-8520 Erlangen, FR Germany (1983-03)
      The reduction of bovine serum albumin (BSA) by hydrated electrons, formate and ethanol radicals leads to the formation of S-S-radicals. The reaction of the hydrated electrons becomes faster, if BSA was preirradiated in the presence of formate or ethanol, but the rate does not change if radiolysis is done in the presence of t-butanol or without scavenger. The reduction of BSA by formate radicals is 10 times slower but gives a higher yield of S-S-radicals than by hydrated electrons. The rate and yield of the reduction by the formate radicals are increased by preirradiation of BSA. The reduction by ethanol radicals is five times slower than by formate radicals and gives lower yields of S-S-radicals which are increased by preirradiation. All rates and yields of the reduction reactions are decreased by changing the pH from 6.3 to 7.3.
    • Radiation-induced tumorous outgrowths in young gametophytes of osmunda regalis.

      Haigh, M V; Howard, Alma; Paterson Laboratories, Christie Hospital and Holt Radium Institute, Manchester (1973)
    • The radical ions and photoionization of bile pigments.

      Land, Edward J; Sloper, R W; Truscott, T G; Paterson Laboratories, Christie Hospital and Holt Radium Institute, Manchester, M20 9BX (1983-12)
      The semireduced, semioxidized, and OH(.)-adduct radicals of bilirubin (BR) and biliverdin (BV) have been characterized using pulse radiolysis techniques. Laser flash photolysis (265-nm) of these pigments led to monophotonic photoionization with quantum yields of 0.08 for BR and 0.03 for BV. No evidence for triplet formation or for photoisomerization was found after 265-nm laser excitation. However, 347-nm excitation of BR in chloroform led to simultaneous photoisomerization and radical formation, but the radicals are thought to have originated from a pathway other than photoionization. The relevance of these observations to BR photoreactivity is discussed. BR radical ions in alkaline solution did not react with tryptophan (TrpH), but the semioxidized TrpH radical oxidized BR with k = 4.3 X 10(8) dm3 mole-1 sec-1. When human serum albumin (HSA) was oxidized using radiolytically generated azide radicals, a radical transformation involving TrpH and TyrOH residues occurred with k = 3.8 X 10(3) sec-1. When BR was complexed with the protein the transformation rate was reduced to 1.6 X 10(3) sec-1. This was interpreted in terms of a conformational change in the protein. Identification of the probable residues involved provided information about the primary BR binding site which was consistent with an earlier report.