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  • 51.
    Fondell, Amelie
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Edwards, Katarina
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Physical Chemistry.
    Gustafsson, Erika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Park, John
    Department of Medicine, Division of Haematology-Oncology, Cancer research institute, University of California San Fransisco, USA.
    Öhrngren, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Organic Pharmaceutical Chemistry.
    Strand, Joanna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Unga, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Physical Chemistry.
    Gedda, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Influence of liposome composition on cellular drug delivery and therapeutic effect mediated by Nuclisome-particlesManuscript (preprint) (Other academic)
  • 52.
    Gedda, L
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Bororn neutron capture therapy. Preclinical studies of compounds for tumour targeting.1997Book (Other academic)
  • 53.
    Gedda, L
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Carlsson, J
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Epidermal growth factor-DNA conjugate for two-step targeting: development and introductory in vitro evaluation.1997In: Int J Oncology, Vol. 11, p. 789-Article in journal (Refereed)
  • 54.
    Gedda, L
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Ghaneolhosseini, H
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Nilsson, P
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Nyholm, K
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Pettersson, J
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Sjoberg, S
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Carlsson, J
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    The influence of lipophilicity on binding of boronated DNA-intercalatingcompounds in human glioma spheroids.2000In: Anti-Cancer Drug Des., Vol. 15, p. 277-Article in journal (Refereed)
  • 55. Gedda, L
    et al.
    Ghaneolhosseini, H
    Nilsson, P
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry.
    Nyholm, K
    Pettersson, J
    Sjöberg, S
    Carlsson, J
    The influence of lipophilicity on binding of boronated DNA-intercalating compounds in human glioma speroids2000In: Anti-Cancer Drug Design, no 15, p. 277-286Article in journal (Refereed)
  • 56.
    Gedda, L
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Olsson, P
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Carlsson, J
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Preparation of epidermal growth factor conjugates aimed for boron neutron capture therapy.1996In: Cancer Neutron Capture Therapy for Cancer, Mishima Y, Plenum Press, New York , 1996, Vol. chapter 3, p. 37-Chapter in book (Other academic)
  • 57.
    Gedda, L
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Olsson, P
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Ponten, J
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
    Carlsson, J
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Development and in vitro studies of epidermal growth factor-dextranconjugates for boron neutron capture therapy.1996In: Bioconjug Chem, Vol. 7, p. 584-Article in journal (Refereed)
  • 58.
    Gedda, L
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Silvander, M
    Sjoberg, S
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Tjarks, W
    Carlsson, J
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Cytotoxicity and subcellular localization of boronated phenanthridinium analogues.1998In: Anti-Cancer Drug Des., Vol. 12, p. 671-Article in journal (Refereed)
  • 59.
    Gedda, L
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Silvander, M
    Sjoberg, S
    Tjarks, W
    Carlsson, J
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Two step targeting: EGF-conjugate for delivery of DNA-binding compounds.1997Conference paper (Other academic)
  • 60.
    Gedda, L
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Silvander, M
    Sjoberg, S
    Tjarks, W
    Carlsson, J
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Two-step targeting: EGF-based conjugate for deliverance of nucleus-specific boron compounds.1996In: Seventh International Symposium on Neutron Capture Therapy for Cancer, p. 67-Article, book review (Other academic)
  • 61.
    Gedda, Lars
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Fondell, Amelie
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Lundqvist, Hans
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Park, John
    Department of medicine, Division of Haematology-Oncology, Cancer research institute, University of California San Fransisco, USA.
    Edwards, Katarina
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Experimental radionuclide therapy of HER2-expressing xenografts using two-step targeting Nuclisome-particles2012In: Journal of Nuclear Medicine, ISSN 0161-5505, E-ISSN 1535-5667, Vol. 53, no 3, p. 480-487Article in journal (Refereed)
    Abstract [en]

    The therapeutic potential of Auger-electron emitting radionuclides is strongly dependent on their close vicinity to DNA, since the energy deposition is mainly localized within a few cubic nanometers around the site of decay. Thus, apart from specificity, successful tumor therapy relies on a nuclear delivery strategy. We recently presented a two-step targeting strategy to transport Auger-electron-emitting radionuclides into the cell nucleus by means of nuclide-filled liposomes (Nuclisome particles), that is, polyethylene glycol-stabilized, tumor-cell-targeting liposomes loaded with (125)I-labeled anthracyclines. In the present study, the survival of mice intraperitoneally inoculated with human HER2-expressing SKOV-3 tumor cells and treated with HER2-targeting Nuclisome particles was studied.

    METHODS:

    BALB/c nu/nu mice were inoculated with 10(7) SKOV-3 cells intraperitoneally and thereafter directly injected with Nuclisome particles with increasing specific radioactivity. Groups of 10-12 mice were treated with 0.01 MBq/mouse up to 2 MBq/mouse, and survival was monitored and compared with that in control groups (n = 33). Organs were analyzed for HER2 expression and radiotoxic effects histologically. Absorbed doses were estimated using dose factors from the online Radiation Dose Assessment Resource model.

    RESULTS:

    The results showed a clear correlation between administered radioactive dose and survival. No such dose-dependent survival was observed for mice treated with Nuclisome particles lacking HER2-targeting ability. With HER2-targeting Nuclisome particles, a significant increase in survival, compared with that of untreated control mice, could already be seen at an administered activity of 0.1 MBq/mouse (P = 0.0301). At the highest activity administered, 2 MBq/mouse (P < 0.0001), 70% of the mice survived the study and most were tumor-free. Neither macroscopic nor microscopic radiotoxic side effects were observed. Dosimetric calculations, assuming nonreceptor targeting, revealed that the radioactive doses to normal tissues were low.

    CONCLUSION:

    Taken together the results show that with successful targeting to the tumor-cell nucleus it is possible to obtain a therapeutic effect from Auger-electron-emitting radionuclides administered at radioactive doses low enough to spare normal tissue from radiotoxic side effects.

  • 62. Geitmann, Matthis
    et al.
    Elinder, Malin
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Seeger, Christian
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Brandt, Peter
    de Esch, Iwan J P
    Danielson, U Helena
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Biochemistry.
    Identification of a Novel Scaffold for Allosteric Inhibition of Wild Type and Drug Resistant HIV-1 Reverse Transcriptase by Fragment Library Screening2011In: Journal of Medicinal Chemistry, ISSN 0022-2623, E-ISSN 1520-4804, Vol. 54, no 3, p. 699-708Article in journal (Refereed)
    Abstract [en]

    A novel scaffold inhibiting wild type and drug resistant variants of human immunodeficiency virus type 1 reverse transcriptase (HIV-1RT) has been identified in a library consisting of 1040 fragments. The fragments were significantly different from already known non-nucleoside reverse transcriptase inhibitors (NNRTIs), as indicated by a Tversky similarity analysis. A screening strategy involving SPR biosensor-based interaction analysis and enzyme inhibition was used. Primary biosensor-based screening, using short concentration series, was followed by analysis of nevirapine competition and enzyme inhibition, thus identifying inhibitory fragments binding to the non-nucleoside reverse transcriptase inhibitor (NNRTI) binding site. Ten hits were discovered, and their affinities and resistance profiles were evaluated with wild type and three drug resistant enzyme variants (K103N, Y181C, and L100I). One fragment exhibited submillimolar K(D) and IC(50) values against all four tested enzyme variants. A substructure comparison between the fragment and 826 structurally diverse published NNRTIs confirmed that the scaffold was novel. The fragment is a bromoindanone with a ligand efficiency of 0.42 kcal/mol(-1).

  • 63. Genovese, Giulio
    et al.
    Kaehler, Anna K.
    Handsaker, Robert E.
    Lindberg, Johan
    Rose, Samuel A.
    Bakhoum, Samuel F.
    Chambert, Kimberly
    Mick, Eran
    Neale, Benjamin M.
    Fromer, Menachem
    Purcell, Shaun M.
    Svantesson, Oscar
    Landen, Mikael
    Höglund, Martin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Haematology.
    Lehmann, Sören
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Haematology.
    Gabriel, Stacey B.
    Moran, Jennifer L.
    Lander, Eric S.
    Sullivan, Patrick F.
    Sklar, Pamela
    Groenberg, Henrik
    Hultman, Christina M.
    McCarroll, Steven A.
    Clonal Hematopoiesis and Blood-Cancer Risk Inferred from Blood DNA Sequence2014In: New England Journal of Medicine, ISSN 0028-4793, E-ISSN 1533-4406, Vol. 371, no 26, p. 2477-2487Article in journal (Refereed)
    Abstract [en]

    Background Cancers arise from multiple acquired mutations, which presumably occur over many years. Early stages in cancer development might be present years before cancers become clinically apparent. Methods We analyzed data from whole-exome sequencing of DNA in peripheral-blood cells from 12,380 persons, unselected for cancer or hematologic phenotypes. We identified somatic mutations on the basis of unusual allelic fractions. We used data from Swedish national patient registers to follow health outcomes for 2 to 7 years after DNA sampling. Results Clonal hematopoiesis with somatic mutations was observed in 10% of persons older than 65 years of age but in only 1% of those younger than 50 years of age. Detectable clonal expansions most frequently involved somatic mutations in three genes (DNMT3A, ASXL1, and TET2) that have previously been implicated in hematologic cancers. Clonal hematopoiesis was a strong risk factor for subsequent hematologic cancer (hazard ratio, 12.9; 95% confidence interval, 5.8 to 28.7). Approximately 42% of hematologic cancers in this cohort arose in persons who had clonality at the time of DNA sampling, more than 6 months before a first diagnosis of cancer. Analysis of bone marrow-biopsy specimens obtained from two patients at the time of diagnosis of acute myeloid leukemia revealed that their cancers arose from the earlier clones. Conclusions Clonal hematopoiesis with somatic mutations is readily detected by means of DNA sequencing, is increasingly common as people age, and is associated with increased risks of hematologic cancer and death. A subset of the genes that are mutated in patients with myeloid cancers is frequently mutated in apparently healthy persons; these mutations may represent characteristic early events in the development of hematologic cancers.

  • 64.
    Gustafsson, Ann-Sofie
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medical Radiation Science.
    Radiation response in human cells: DNA damage formation, repair and signaling2015Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Ionizing radiation induces a range of different DNA lesions. In terms of mutation frequency and mammalian cell survival, the most critical of these lesions is the DNA double-strand break (DSB). DSB left unrepaired or mis-repaired may result in chromosomal aberrations that can lead to permanent genetic changes or cell death. The complexity of the DNA damage and the capacity to repair the DSB will determine the fate of the cell. This thesis focuses on the DNA damage formation, repair and signaling after irradiation of human cells.

    Radiation with high linear energy transfer (LET) produces clustered damaged sites in the DNA that are difficult for the cell to repair. Within these clustered sites, non-DSB lesions are formed that can be converted into a DSB and add to the damage complexity and affect DSB repair and the measurement. Heat-labile sites in DNA are converted into DSB at elevated temperatures. We show that heat-released DSB are formed post-irradiation with high-LET ions and increase the initial yield of DSB by 30%-40%, which is similar to yields induced by low-LET radiation.

    DNA-PKcs, a central player in non-homologous end-joining (NHEJ), the major mammalian DSB repair pathway, has been found to be both up- and downregulated in different tumor types. In Paper II we show that low levels of DNA-PKcs lead to extreme radiosensitivity but, surprisingly, had no effect on the DSB repair. However, the fraction of cells in G2/M phase increased two-fold in cells with low levels of DNA-PKcs. The study continued in Paper IV, where cells were synchronized to unmask potential roles of DNA-PKcs in specific cell cycle phases. Irradiation of DNA-PKcs suppressed cells in the G1/S phase caused a delay in cell cycle progression and an increase in accumulation of G2 cells. Further, these cells showed defects in DNA repair, where a significant amount of 53BP1 foci remained after 72 h. This further strengthens the hypothesis that DNA-PKcs has a role in regulation of mitotic progression.

    Several cellular signaling pathways are initiated in response to radiation. One of these downstream signaling proteins is AKT. We identified an interaction between DNA-PKcs and AKT. Knockouts of both AKT1 and AKT2 impaired DSB rejoining after radiation and low levels of DNA-PKcs increased radiosensitivity and decreased DNA repair further.  

    List of papers
    1. Formation and repair of clustered damaged DNA sites in high LET irradiated cells
    Open this publication in new window or tab >>Formation and repair of clustered damaged DNA sites in high LET irradiated cells
    2015 (English)In: International Journal of Radiation Biology, ISSN 0955-3002, E-ISSN 1362-3095, Vol. 91, no 10, p. 820-826Article in journal (Refereed) Published
    Abstract [en]

    PURPOSE: Radiation with high linear energy transfer (LET) produces clustering of DNA double-strand breaks (DSB) as well as non-DSB lesions. Heat-labile sites (HLS) are non-DSB lesions in irradiated cells that may convert into DSB at elevated temperature during preparation of naked DNA for electrophoretic assays and here we studied the initial formation and repair of these clustered damaged sites after irradiation with high LET ions.

    MATERIALS AND METHODS: Induction and repair of DSB were studied in normal human skin fibroblast (GM5758) after irradiation with accelerated carbon and nitrogen ions at an LET of 125 eV/nm. DNA fragmentation was analyzed by pulsed-field gel electrophoresis (PFGE) and by varying the lysis condition we could differentiate between prompt DSB and heat-released DSB.

    RESULTS: Before repair (t = 0 h), the 125 eV/nm ions produced a significant fraction of heat-released DSB, which appeared clustered on DNA fragments with sizes of 1 Mbp or less. These heat-released DSB increased the total number of DSB by 30-40%. This increase is similar to what has been found in low-LET irradiated cells, suggesting that the relative biological effectiveness (RBE) for DSB induction will not be largely affected by the lysis temperature. After 1-2 hours repair, a large fraction of DSB was still unrejoined but there was essentially no heat-released DSB present.

    CONCLUSIONS: These results suggest that high LET radiation, as low LET gamma radiation, induces a significant fraction of heat-labile sites which can be converted into DSB, and these heat-released DSB may affect both induction yields and estimates of repair.

    Keywords
    Clustered damage; DNA damage; DNA DSB repair; DNA repair; double-strand breaks; High LET
    National Category
    Basic Medicine Radiology, Nuclear Medicine and Medical Imaging
    Identifiers
    urn:nbn:se:uu:diva-265135 (URN)10.3109/09553002.2015.1068463 (DOI)000365614800007 ()26136085 (PubMedID)
    Funder
    Swedish Cancer Society
    Available from: 2015-10-22 Created: 2015-10-22 Last updated: 2018-01-11Bibliographically approved
    2. Suppression of DNA-dependent protein kinase sensitize cells to radiation without affecting DSB repair
    Open this publication in new window or tab >>Suppression of DNA-dependent protein kinase sensitize cells to radiation without affecting DSB repair
    2014 (English)In: Mutation research, ISSN 0027-5107, E-ISSN 1873-135X, Vol. 769, p. 1-10Article in journal (Refereed) Published
    Abstract [en]

    Efficient and correct repair of DNA double-strand break (DSB) is critical for cell survival. Defects in the DNA repair may lead to cell death, genomic instability and development of cancer. The catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) is an essential component of the non-homologous end joining (NHEJ) which is the major DSB repair pathway in mammalian cells. In the present study, by using siRNA against DNA-PKcs in four human cell lines, we examined how low levels of DNA-PKcs affected cellular response to ionizing radiation. Decrease of DNA-PKcs levels by 80-95%, induced by siRNA treatment, lead to extreme radiosensitivity, similar to that seen in cells completely lacking DNA-PKcs and low levels of DNA-PKcs promoted cell accumulation in G2/M phase after irradiation and blocked progression of mitosis. Surprisingly, low levels of DNA-PKcs did not affect the repair capacity and the removal of 53BP1 or gamma-H2AX foci and rejoining of DSB appeared normal. This was in strong contrast to cells completely lacking DNA-PKcs and cells treated with the DNA-PKcs inhibitor NU7441, in which DSB repair were severely compromised. This suggests that there are different mechanisms by which loss of DNA-PKcs functions can sensitize cells to ionizing radiation. Further, foci of phosphorylated DNA-PKcs (T2609 and S2056) co-localized with DSB and this was independent of the amount of DNA-PKcs but foci of DNA-PKcs was only seen in siRNA-treated cells. Our study emphasizes on the critical role of DNA-PKcs for maintaining survival after radiation exposure which is uncoupled from its essential function in DSB repair. This could have implications for the development of therapeutic strategies aiming to radiosensitize tumors by affecting the DNA-PKcs function.

    Keywords
    DNA repair, DNA-PKcs, Ionizing radiation, DNA-PK deficiency, NU7441
    National Category
    Medical Genetics
    Identifiers
    urn:nbn:se:uu:diva-237292 (URN)10.1016/j.mrfmmm.2014.06.004 (DOI)000343625700001 ()
    Available from: 2014-12-03 Created: 2014-12-01 Last updated: 2019-03-08Bibliographically approved
    3. The influence of AKT isoforms on radiation sensitivity and DNA repair in colon cancer cell lines
    Open this publication in new window or tab >>The influence of AKT isoforms on radiation sensitivity and DNA repair in colon cancer cell lines
    Show others...
    2014 (English)In: Tumor Biology, ISSN 1010-4283, E-ISSN 1423-0380, Vol. 35, no 4, p. 3525-3534Article in journal (Refereed) Published
    Abstract [en]

    In response to ionizing radiation, several signaling cascades in the cell are activated to repair the DNA breaks, prevent apoptosis, and keep the cells proliferating. AKT is important for survival and proliferation and may also be an activating factor for DNA-PKcs and MRE11, which are essential proteins in the DNA repair process. AKT (PKB) is hyperactivated in several cancers and is associated with resistance to radiotherapy and chemotherapy. There are three AKT isoforms (AKT1, AKT2, and AKT3) with different expression patterns and functions in several cancer tumors. The role of AKT isoforms has been investigated in relation to radiation response and their effects on DNA repair proteins (DNA-PKcs and MRE11) in colon cancer cell lines. The knockout of AKT1 and/or AKT2 affected the radiation sensitivity, and a deficiency of both isoforms impaired the rejoining of radiation-induced DNA double strand breaks. Importantly, the active/phosphorylated forms of AKT and DNA-PKcs associate and exposure to ionizing radiation causes an increase in this interaction. Moreover, an increased expression of both DNA-PKcs and MRE11 was observed when AKT expression was ablated, yet only DNA-PKcs expression influenced AKT phosphorylation. Taken together, these results demonstrate a role for both AKT1 and AKT2 in radiotherapy response in colon cancer cells involving DNA repair capacity through the nonhomologous end joining pathway, thus suggesting that AKT in combination with DNA-PKcs inhibition may be used for radiotherapy sensitizing strategies in colon cancer.

    National Category
    Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
    Identifiers
    urn:nbn:se:uu:diva-221446 (URN)10.1007/s13277-013-1465-9 (DOI)000334495900084 ()
    Available from: 2014-03-31 Created: 2014-03-31 Last updated: 2017-12-05Bibliographically approved
    4. Role of DNA-PKcs in DNA damage response and cell cycle regulation
    Open this publication in new window or tab >>Role of DNA-PKcs in DNA damage response and cell cycle regulation
    (English)Manuscript (preprint) (Other academic)
    National Category
    Cell and Molecular Biology
    Identifiers
    urn:nbn:se:uu:diva-265136 (URN)
    Note

    The catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) is an essential component of non-homologous end joining (NHEJ) which is the major DSB repair pathway in mammalian cells. We have previously reported that suppression of DNA-PKcs sensitize cells to radiation without affecting repair. In the present study we used synchronized cells to unmask potential roles of DNA-PKcs in specific cell-cycle phases. siRNA was used to deplete DNA-PKcs to 10-15 % of normal levels and cell cycle progression and DSB repair was observed in synchronized cells irradiated at different cell cycle phases. Surprisingly, cells irradiated in G2 phase showed similar p-H3 frequency after 72 h as unirradiated control cells, irrespective of their DNA-PKcs status. When cells were irradiated in G1/S phase, a significant increase of mitotic cells in siDNA-PKcs treated cells was seen 72 h later. Further, irradiation in G1/ S phase caused initially (0-12h) the same number of DSB (53BP1 foci), however, over time (>24h) 53BP1 foci remained at relatively high levels in DNA-PKcs depleted cells, indicating presence of unrepaired DSB in the following G1. This suggest that DNA-PKcs has an important regulatory role in G1/S and a key function in mitosis.

    Available from: 2015-10-22 Created: 2015-10-22 Last updated: 2018-01-11
  • 65.
    Halin Lejonklou, Margareta
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Endocrine Tumor Biology.
    Johansson, Térèse
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Endocrine Tumor Biology.
    Ekeblad, Sara
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Endocrine Tumor Biology.
    Skogseid, Britt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Endocrine Tumor Biology.
    Notch signaling factors in the transforming Men1 mouse pancreasManuscript (preprint) (Other academic)
  • 66.
    Hall, Håkan
    et al.
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine.
    Takahashi, Kayo
    Erlandsson, Maria
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Biochemistry and Organic Chemistry.
    Estrada, Sergioa
    Bergström, Elisabeth
    Långström, Bengt
    Pharmacological characterization of 18F-labeled vorozole analogues.Manuscript (Other academic)
    Abstract [en]

    Two 18F-labeled analogues of vorozole ([18F]FVOZ and [18F]FVOO) have been developed as potential tools for the in vivo characterization of aromatase. The purpose of the project was to evaluate the pharmacological properties of these radioligands using a combination of in vitro binding and in vivo distribution studies in the rat and primate. Saturation binding studies with the radioligands in homogenates of rat ovary gave KD and Bmax values of 0.21 ± 0.1 nM and 210 ± 20 fmol/mg, respectively, for [18F]FVOZ, and 7.6 ± 1 nM and 293 ± 12 fmol/mg, respectively, for [18F]FVOO. Organ distribution studies in rats showed the highest accumulation in the adrenal glands, with standardized uptake values (SUVs) of 15 to 20, followed by ovaries and liver with SUVs of approximately 5. The SUVs in the remaining organs were between 0.5 and 1.5. There was probably some defluorination of both radioligands, as the accumulation of radioactivity in bone increased with time. The regional distribution in the brain was studied using ex vivo and in vitro autoradiography. In the brain, specific binding of both [18F]FVOZ and [18F]FVOO were found mainly in the amygdala. PET studies were performed in the Rhesus monkey, and these showed displaceable binding in the amygdala and the preoptic area of the hypothalamus. These studies suggest that [18F]FVOZ might be to be a suitable tracer for the study of aromatase in vitro and in vivo, and could be an alternative to [11C]vorozole in human PET-studies.

  • 67.
    Haylock, Anna-Karin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences.
    Nilvebrant, Johan
    KTH.
    Mortensen, Anja
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Velikyan, Irina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry.
    Nestor, Marika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Falk, Ronny
    KI.
    Generation and evaluation of antibody agents for molecular imaging of CD44v6 expressing cancersManuscript (preprint) (Other academic)
  • 68.
    Heldin, Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Identification and Characterization of Proteins and MicroRNAs that Modulate Receptor Signaling, Vesicular Trafficking and Cell Migration in Vascular Cells2014Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Blood vessels deliver nutrients and oxygen to tissues. Importantly, the functions and growth of blood vessels are commonly altered in disease. The inside of all blood vessels are lined with endothelium, a thin specialized layer of endothelial cells that separate the blood from other tissues. This thesis deals with the identification and functional characterization of proteins and microRNAs that have key roles as modulators of growth factor signaling and directed cell migration of endothelial cells and other vascular cells.

    A previously uncharacterized protein of the exocyst complex, Exocyst complex component 3-like 2 (ExoC3L2) was identified and shown to be highly expressed in endothelial cells of sprouting vessels. Suppression of ExoC3L2 resulted in reduced VEGF-A signaling together with reduced chemotaxis in response to VEGF-A gradients. VEGF-A-signaling via its receptor VEGFR-2 is thus modulated by the exocyst complex and ExoC3L2.

    Expression profiling of highly vascularized tissues were used to identify several microRNAs selectively expressed in blood vessels. miR-145, targeting the transcription factor Fli1, was shown to be expressed in pericytes and mural cells. Elevated levels of miR-145 reduced chemotaxis of both endothelial cells and fibroblasts in response to growth factor gradients. miR-145 depletion in fibroblasts was shown to inhibit chemotaxis in response to PDGF-BB.

    The guanine nucleotide exchange factor FGD5 was shown to be selectively expressed in endothelial cells and to regulate Cdc42 activity. FGD5 was shown to regulate the turnover of activated VEGF-receptors. Suppression of FGD5 impaired endothelial cell chemotaxis, suggesting that FGD5 is required for efficient and sustained VEGF-A signaling.

    Inactivation of RhoD, a regulator of endosomal trafficking, resulted in an increased pool of acetylated and stable microtubules. Knockdown of RhoD in human fibroblasts resulted in a loss of cell polarity. A link between PDGFR-β and RhoD was implicated by the finding that PDGF-BB was shown to trigger formation of GTP-bound RhoD. Chemotaxis towards PDGF-BB was severely inhibited in cells with reduced RhoD expression, suggesting a role for RhoD in chemotaxis via its regulation of microtubule dynamics.

    List of papers
    1. Exocyst Complex Component 3-like 2 (EXOC3L2) Associates with the Exocyst Complex and Mediates Directional Migration of Endothelial Cells
    Open this publication in new window or tab >>Exocyst Complex Component 3-like 2 (EXOC3L2) Associates with the Exocyst Complex and Mediates Directional Migration of Endothelial Cells
    Show others...
    2011 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 286, no 27, p. 24189-24199Article in journal (Refereed) Published
    Abstract [en]

    The exocyst is a protein complex that ensures spatial targeting of exocytotic vesicles to the plasma membrane. We present microarray data obtained from differentiating mouse embryonic stem cell cultures that identify an up-regulation of exocyst complex component 3-like 2 (exoc3l2) mRNA in sprouting blood vessels. Vascular expression of exoc3l2 is confirmed by qPCR analysis of different mouse tissues and immunofluorescence analyses of mouse brain sections. We detect an up-regulation of exoc3l2 mRNA synthesis in primary human endothelial cells in response to VEGFA, and this response is enhanced when the cells are grown on a three-dimensional collagen I matrix. Myc-tagged EXOC3L2 co-precipitates with the exocyst protein EXOC4, and immunofluorescence detection of EXOC3L2 shows partial subcellular colocalization with EXOC4 and EXOC7. Finally, we show that exoc3l2 silencing inhibits VEGF receptor 2 phosphorylation and VEGFA-directed migration of cultured endothelial cells.

    Keywords
    Angiogenes, VEGF, Cell migration, Chemotaxis, Exocyst, Microarray
    National Category
    Biochemistry and Molecular Biology
    Research subject
    Biology with specialization in Molecular Cell Biology
    Identifiers
    urn:nbn:se:uu:diva-145508 (URN)10.1074/jbc.M110.212209 (DOI)000292294900061 ()
    Available from: 2011-02-09 Created: 2011-02-09 Last updated: 2017-12-11Bibliographically approved
    2. Discovery of microvascular miRNAs using public gene expression data: miR-145 is expressed in pericytes and is a regulator of Fli1
    Open this publication in new window or tab >>Discovery of microvascular miRNAs using public gene expression data: miR-145 is expressed in pericytes and is a regulator of Fli1
    Show others...
    2009 (English)In: Genome Medicine, ISSN 1756-994X, E-ISSN 1756-994X, Vol. 1, no 11, p. 108-Article in journal (Refereed) Published
    Abstract [en]

    BACKGROUND

    A function for the microRNA (miRNA) pathway in vascular development and angiogenesis has been firmly established. miRNAs with selective expression in the vasculature are attractive as possible targets in miRNA-based therapies. However, little is known about the expression of miRNAs in microvessels in vivo. Here, we identified candidate microvascular-selective miRNAs by screening public miRNA expression datasets.

    METHODS

    Bioinformatics predictions of microvascular-selective expression were validated with real-time quantitative reverse transcription PCR on purified microvascular fragments from mouse. Pericyte expression was shown with in situ hybridization on tissue sections. Target sites were identified with 3' UTR luciferase assays, and migration was tested in a microfluid chemotaxis chamber.

    RESULTS

    miR-145, miR-126, miR-24, and miR-23a were selectively expressed in microvascular fragments isolated from a range of tissues. In situ hybridization and analysis of Pdgfb retention motif mutant mice demonstrated predominant expression of miR-145 in pericytes. We identified the Ets transcription factor Friend leukemia virus integration 1 (Fli1) as a miR-145 target, and showed that elevated levels of miR-145 reduced migration of microvascular cells in response to growth factor gradients in vitro.

    CONCLUSIONS

    miR-126, miR-24 and miR-23a are selectively expressed in microvascular endothelial cells in vivo, whereas miR-145 is expressed in pericytes. miR-145 targets the hematopoietic transcription factor Fli1 and blocks migration in response to growth factor gradients. Our findings have implications for vascular disease and provide necessary information for future drug design against miRNAs with selective expression in the microvasculature.

    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-119570 (URN)10.1186/gm108 (DOI)19917099 (PubMedID)
    Note

    Article 108

    Available from: 2010-02-26 Created: 2010-02-26 Last updated: 2017-12-12Bibliographically approved
    3. FGD5 promotes VEGF-A signaling via inhibition of VEGF-receptor ubiquitination and degradation
    Open this publication in new window or tab >>FGD5 promotes VEGF-A signaling via inhibition of VEGF-receptor ubiquitination and degradation
    Show others...
    (English)Manuscript (preprint) (Other academic)
    National Category
    Cell and Molecular Biology
    Identifiers
    urn:nbn:se:uu:diva-212946 (URN)
    Available from: 2013-12-16 Created: 2013-12-16 Last updated: 2018-01-11
    4. A RhoD-regulated pathway is needed for PDGF-BB-induced chemotaxis
    Open this publication in new window or tab >>A RhoD-regulated pathway is needed for PDGF-BB-induced chemotaxis
    Show others...
    (English)Manuscript (preprint) (Other academic)
    National Category
    Cell and Molecular Biology
    Identifiers
    urn:nbn:se:uu:diva-212948 (URN)
    Available from: 2013-12-16 Created: 2013-12-16 Last updated: 2018-01-11
  • 69. Hildén, Lars
    et al.
    Väljamäe, Priit
    Johansson, Gunnar
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry.
    Surface character of pulp fibres studied using endoglucanases2005In: Journal of Biotechnology, ISSN 0168-1656, E-ISSN 1873-4863, Vol. 118, no 4, p. 386-397Article in journal (Refereed)
    Abstract [en]

    The endoglucanase Cel5A from Trichoderma reesei and an endoglucanase from Aspergillus sp. (Novozym 476TM from Novozyme A/S) were evaluated as probes for the surface properties of soft- and hardwood chemical pulp fibres. The hydrolysis time curves were in accordance with a two-phase degradation model described by a biexponential function. The kinetic parameters corresponding to the amount of fast and slow degraded parts of the substrate correlated to tensile index, relative bonded area and z-strength of the paper. All paper properties showing a correlation with enzyme kinetic parameters were related to fibre–fibre interactions. Fluorescence labelling of the reducing end groups in pulp fibres followed by enzyme treatment indicated that the fast substrate class corresponds to the population of “loose” cellulose chain ends not tightly associated with the bulk cellulose. The correlation between the parameters of enzyme kinetics and mechanical properties of the paper produced from the corresponding pulp found in this study should allow a rapid evaluation of the raw fibre material used in paper making process.

  • 70.
    Holmfeldt, Linda
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Experimental and Clinical Oncology.
    Mullighan, Charles G
    Generation of human acute lymphoblastic leukemia xenografts for use in oncology drug discovery2015In: Current protocols in pharmacology, ISSN 1934-8290, Vol. 68, p. 14.32.1-14.32.19Article in journal (Refereed)
    Abstract [en]

    The establishment of reproducible mouse models of acute lymphoblastic leukemia (ALL) is necessary to provide in vivo therapeutic test systems that recapitulate human ALL, and for amplification of limited amounts of primary tumor material. A popular assay is the primary xenograft model that utilizes immunocompromised mice. The protocol includes injection of primary patient tumor specimens into mice with subsequent serial passaging of the tumors by retransplants of cells harvested from the mouse bone marrow and spleen. The tumors generated are then used for genomic profiling, ex vivo compound testing, mechanistic studies and retransplantation. Detailed in this unit are procedures for the establishment and maintenance of primary ALL xenograft panels for use in basic research and translational studies.

  • 71. Huber, Stefan M.
    et al.
    Ertem, Mehmed Z.
    Aquilante, Francesco
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Quantum Chemistry.
    Gagliardi, Laura
    Tolman, William B.
    Cramer, Christopher J.
    Generating Cu-II-oxyl/Cu-III-oxo species from Cu-I-alpha-ketocarboxylate complexes and O-2: in silico studies on ligand effects and C-H-activation reactivity.2009In: Chemistry - A European Journal, ISSN 0947-6539, E-ISSN 1521-3765, Vol. 15, no 19, p. 4886-4895Article in journal (Refereed)
    Abstract [en]

    A mechanism for the oxygenation of Cu-I complexes with alpha-keto-carboxylate ligands that is based on a combination of density functional theory and multireference second-order perturbation theory (CASSCF/CASPT2) calculations is elaborated. The reaction proceeds in a manner largely analogous to those of similar Fe-II-alpha-ketocarboxylate systems, that is, by initial attack of a coordinated oxygen molecule on a ketocarboxylate ligand with concomitant decarboxylation. Subsequently, two reactive intermediates may be generated, a Cu-peracid structure and a [CuO](+) species, both of which are capable of oxidizing a phenyl ring component of the supporting ligand. Hydroxylation by the [CuO](+) species is predicted to proceed with a smaller activation free energy. The effects of electronic and steric variations on the oxygenation mechanisms were studied by introducing substituents at several positions of the ligand backbone and by investigating various N-donor ligands. In general, more electron donation by the N-donor ligand leads to increased stabilization of the more Cu-II/Cu-III-like intermediates (oxygen adducts and [CuO](+) species) relative to the more Cu-I-like peracid intermediate. For all ligands investigated the [CuO](+) intermediates are best described as Cu-II-O center dot(-) species with triplet ground states. The reactivity of these compounds in C-H abstraction reactions decreases with more electron-donating N-donor ligands, which also increase the Cu-O bond strength, although the Cu-O bond is generally predicted to be rather weak (with a bond order of about 0.5). A comparison of several methods to obtain singlet energies for the reaction intermediates indicates that multireference second-order perturbation theory is likely more accurate for the initial oxygen adducts, but not necessarily for subsequent reaction intermediates.

  • 72.
    Ihse, Elisabet
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular and Morphological Pathology.
    Two Types of Fibrils in ATTR Amyloidosis: Implications for Clinical Phenotype and Treatment Outcome2011Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Systemic amyloidoses are a group of lethal diseases where proteins aggregate into fibrillar structures, called amyloid fibrils, that deposits throughout the body. Transthyretin (TTR) causes one type of amyloidosis, in which the aggregates mainly infiltrate nervous and cardiac tissue. Almost a hundred different mutations in the TTR gene are known to trigger the disease, but wild-type (wt) TTR is also incorporated into the fibrils, and may alone form amyloid. Patients with the TTRV30M mutation show, for unknown reasons, two clinical phenotypes. Some have an early onset of disease without cardiomyopathy while others have a late onset and cardiomyopathy. It has previously been described that amyloid fibrils formed from TTRV30M can have two different compositions; either with truncated molecules beside full-length TTR (type A) or only-full-length molecules (type B).  In this thesis, the clinical importance of the two types of amyloid fibrils was investigated.

    We found that the fibril composition types are correlated to the two clinical phenotypes seen among TTRV30M patients, with type A fibrils present in late onset patients and type B fibrils in early onset patients.

    The only treatment for hereditary TTR amyloidosis has been liver transplantation, whereby the liver producing the mutant TTR is replaced by an organ only producing wt protein. However, in some patients, cardiac symptoms progress post-transplantationally. We demonstrated that the propensity to incorporate wtTTR differs between fibril types and tissue types in TTRV30M patients, with cardiac amyloid of type A having the highest tendency. This offers an explanation to why particularly cardiac amyloidosis develops after transplantation, and suggests which patients that are at risk for such development.

    By examining patients with other mutations than TTRV30M, we showed that, in contrast to the general belief, a fibril composition with truncated TTR is very common and might even be the general rule. This may explain why TTRV30M patients often have a better outcome after liver transplantation than patients with other mutations.

    In conclusion, this thesis has contributed with one piece to the puzzle of understanding the differences in clinical phenotype and treatment response between TTR amyloidosis patients, by demonstrating corresponding differences at a molecular level.

    List of papers
    1. Amyloid fibril composition is related to the phenotype of hereditary transthyretin V30M amyloidosis
    Open this publication in new window or tab >>Amyloid fibril composition is related to the phenotype of hereditary transthyretin V30M amyloidosis
    Show others...
    2008 (English)In: Journal of Pathology, ISSN 0022-3417, E-ISSN 1096-9896, Vol. 216, no 2, p. 253-61Article in journal (Refereed) Published
    Abstract [en]

    Swedish familial systemic amyloidosis with polyneuropathy (FAP) depends on a mutation leading to a methionine-for-valine substitution in transthyretin. The disease appears with different clinical manifestations, including age of onset and involvement of the heart. Liver transplantation is currently the only curative treatment, but progressive cardiomyopathy may occur post-transplant. Two amyloid deposition patterns have previously been described in the heart. In one, the amyloid consists partially of transthyretin fragments and is weakly stainable by Congo red, while in the other, only full-length molecules are found and the fibrils have a strong affinity for Congo red. The present study aimed to see whether these morphological and biochemical variations have clinical implications. Subcutaneous adipose tissue biopsies were taken from 33 patients with Val30Met FAP and examined by microscopy, electrophoresis and western blot. Clinical data included age, sex, duration of disease and echocardiographic determination of the interventricular septum (IVS) thickness. It was found that fibrils composed of only full-length transthyretin were associated with early age of onset (44.8 +/- 12.9 years), no clinical cardiac involvement and a strong affinity for Congo red. In contrast, presence of transthyretin fragments in the amyloid was associated with late age of onset (67.3 +/- 7.0 years), signs of cardiac involvement and weak Congo red staining. For each individual, the same molecular type of amyloid was found in different organs. This is the first report showing that variations in clinical appearance of familial ATTR amyloidosis are associated with specific structural differences in the amyloid fibrils, and therefore may have a molecular cause. The molecular type of amyloid can be determined from a subcutaneous fat tissue biopsy.

    Keywords
    amyloid, transthyretin, fibril, familial, heart, phenotype, age of onset
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-103613 (URN)10.1002/path.2411 (DOI)000259510000015 ()18729067 (PubMedID)
    Available from: 2009-05-20 Created: 2009-05-20 Last updated: 2017-12-13Bibliographically approved
    2. Proportion between wild-type and mutant protein in truncated compared to full-length ATTR: an analysis on transplanted transthyretin T60A amyloidosis patients
    Open this publication in new window or tab >>Proportion between wild-type and mutant protein in truncated compared to full-length ATTR: an analysis on transplanted transthyretin T60A amyloidosis patients
    Show others...
    2009 (English)In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 379, no 4, p. 846-850Article in journal (Refereed) Published
    Abstract [en]

    Familial ATTR amyloidosis is caused by point mutations in the transthyretin gene. The clinical manifestations are highly varied but polyneuropathy and/or cardiomyopathy are generally the main symptoms. The amyloid fibrils can either be composed of only intact ATTR molecules or intact together with fragmented ATTR species. As plasma TTR is almost exclusively synthesized in the liver, liver transplantation is performed in order to eliminate the mutant plasma TTR. The procedure has shown best results among patients with the V30M mutation, while a rapid continued cardiac deposition of wild-type (wt) TTR has been seen for many other mutations. In this paper we investigated the proportion of wtATTR in two TTRT60A patients that underwent liver transplantation; one patient died 3 weeks after surgery, the other patient survived for 12 months. As the role of fragmented TTR species in the pathogenesis is far from understood, we investigated the proportion of wt in these species separately to the full-length molecules, which has not been done before in transplanted patients. The results show a higher proportion of wtTTR in the 12-months-surviving patient than the 3-weeks-surviving patient, but interestingly this difference in wt proportion is mainly seen among the full-length, and not the fragmented, molecules.

    Keywords
    Amyloid, Transthyretin, Familial amyloidotic polyneuropathy, ATTRT60A, Liver transplantation, Fibril, Cardiomyopathy, Wild-type
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-103620 (URN)10.1016/j.bbrc.2008.12.095 (DOI)000263336700009 ()19118530 (PubMedID)
    Available from: 2009-05-20 Created: 2009-05-20 Last updated: 2017-12-13Bibliographically approved
    3. Variation in amount of wild-type transthyretin in different fibril and tissue types in ATTR amyloidosis
    Open this publication in new window or tab >>Variation in amount of wild-type transthyretin in different fibril and tissue types in ATTR amyloidosis
    2011 (English)In: Journal of Molecular Medicine, ISSN 0946-2716, E-ISSN 1432-1440, Vol. 89, no 2, p. 171-180Article in journal (Refereed) Published
    Abstract [en]

    Familial transthyretin (TTR) amyloidosis is caused by a mutation in the TTR gene, although wild-type (wt) TTR is also incorporated into the amyloid fibrils. Liver transplantation (LT) is the prevailing treatment of the disease and is performed in order to eliminate the mutant TTR from plasma. The outcome of the procedure is varied; especially problematic is a progressive cardiomyopathy seen in some patients, presumably caused by continued incorporation of wtTTR. What determines the discrepancy in outcome is not clear. We have previously shown that two structurally distinct amyloid fibrils (with or without fragmented ATTR) are found among ATTRV30M patients. In this study, we investigated the proportion of wtATTR in cardiac and adipose amyloid from patients having either fibril type. It was found that cardiac amyloid more easily incorporates wtTTR than adipose amyloid, offering a potential explanation for the vulnerability of cardiac tissue for continued amyloidosis after LT. In cardiac tissue, fibrils with fragmented ATTR contained a higher wt proportion than fibrils without, suggesting that continued incorporation of wtTTR after LT, perhaps, can take place more easily in these patients. In adipose tissue, a rapid increase in wt proportion after LT indicates that a rather fast turnover of the deposits must occur. A difference in wt proportion between the fibril types was seen post-LT but not pre-LT, possibly caused by differences in turnover rate. Conclusively, this study further establishes the basic dissimilarities between the two fibril types and demonstrates that their role in LT outcome needs to be further investigated.

    Keywords
    Amyloid, Transthyretin, Familial amyloidotic polyneuropathy, Liver transplantation, Wild-type
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-150732 (URN)10.1007/s00109-010-0695-1 (DOI)000288363300008 ()21107516 (PubMedID)
    Available from: 2011-04-05 Created: 2011-04-05 Last updated: 2017-12-11Bibliographically approved
    4. Amyloid fibrils with fragmented ATTR may be the rule in non-Val30Met ATTR amyloidosis
    Open this publication in new window or tab >>Amyloid fibrils with fragmented ATTR may be the rule in non-Val30Met ATTR amyloidosis
    Show others...
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    The clinical phenotype of familial ATTR amyloidosis depends to some extent on the particular mutation, but differences exist also within mutations. We have previously described that two types of amyloid fibril compositions exist among Swedish ATTRV30M amyloidosis patients, one consisting of a mixture of intact and fragmented ATTR (type A) and one composed of only intact ATTR (type B). Patients with type A fibrils have a late age of onset and signs of cardiomyopathy, while patients with type B fibrils have an early onset and much less myocardial involvement.

    The present study aimed to determine if the correlation between fibril type and clinical phenotype is true for familial amyloidosis in general. Cardiac and/or adipose tissue from 48 patients carrying 21 different non-TTRV30M mutations were examined, as well as 7 non-Swedish ATTRV30M patients. Fibril type was determined with western blotting and compared to the patients´ age of onset and degree of cardiomyopathy.

    Non-Swedish V30M patients showed the same correlation as described for Swedish V30M patients, with fibrils of only full-length ATTR (type B) linked to less myocardial involvement. In contrast, all patients with non-V30M mutations had a fibril composition with ATTR fragments (type A). Some of these patients had onset of disease at young age. The vast majority had increased thickness of left cardiac ventricle, but a few individuals had values within normal limits.

    This study shows that a fibril composition with fragmented ATTR is very common in ATTR amyloidosis. It also suggests that fibrils composed of only full-length ATTR is an exception, perhaps only found among young ATTRV30M amyloidosis patients.

    Keywords
    amyloid, transthyretin, familial amyloidotic polyneuropathy, non-TTRV30M, TTRV30M, cardiomyopathy, fibril composition
    National Category
    Cell and Molecular Biology Other Basic Medicine Clinical Laboratory Medicine Biochemistry and Molecular Biology
    Research subject
    Biochemistry; Biology with specialization in Molecular Biology; Biomedical Laboratory Science; Experimental Pathology; Medical Biochemistry; Molecular Biology; Pathology
    Identifiers
    urn:nbn:se:uu:diva-160979 (URN)
    Available from: 2011-11-27 Created: 2011-11-03 Last updated: 2018-01-12
  • 73.
    Ihse, Elisabet
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular and Morphological Pathology.
    Rapezzi, Claudio
    Cardiovascular Dept, University of Bologna, Italy.
    Merlini, Giampaolo
    Amyloid Research and Treatment Center, Scientific Institute Policlinico San Matteo, University of Pavia, Italy.
    Benson, Merrill D
    Dept of Pathology and Laboratory Medicine, Indiana University School of Medicine and Roudebush VA medical Center, Indianapolis, IN, USA.
    Ando, Yukio
    Dept of Diagnostic Medicine, Graduate School of Medical Sciences, Kumamoto University, Japan.
    Suhr, Ole B
    Dept of Public Health and Clinical Medicine, Umeå University.
    Leone, Ornella
    Pathology Department, S.Orsola-Malpighi Hospital, Bologna, Italy.
    Lorenzini, Massimiliano
    Cardiovascular Dept, University of Bologna, Italy.
    Quarta, Candida Cristina
    Cardiovascular Dept, University of Bologna, Italy.
    Obici, Laura
    Amyloid Research and Treatment Center, Scientific Institute Policlinico San Matteo, University of Pavia, Italy.
    Lavatelli, Francesca
    Amyloid Research and Treatment Center, Scientific Institute Policlinico San Matteo, University of Pavia, Italy.
    Liepnieks, Juris
    Dept of Pathology and Laboratory Medicine, Indiana University School of Medicine and Roudebush VA medical Center, Indianapolis, IN, USA.
    Ohshima, Toshinori
    Dept of Neurology, Graduate School of Medical Sciences, Kumamoto University, Japan.
    Jono, Hirofumi
    Dept of Diagnostic Medicine, Graduate School of Medical Sciences, Kumamoto University, Japan.
    Westermark, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular and Morphological Pathology.
    Amyloid fibrils with fragmented ATTR may be the rule in non-Val30Met ATTR amyloidosisManuscript (preprint) (Other academic)
    Abstract [en]

    The clinical phenotype of familial ATTR amyloidosis depends to some extent on the particular mutation, but differences exist also within mutations. We have previously described that two types of amyloid fibril compositions exist among Swedish ATTRV30M amyloidosis patients, one consisting of a mixture of intact and fragmented ATTR (type A) and one composed of only intact ATTR (type B). Patients with type A fibrils have a late age of onset and signs of cardiomyopathy, while patients with type B fibrils have an early onset and much less myocardial involvement.

    The present study aimed to determine if the correlation between fibril type and clinical phenotype is true for familial amyloidosis in general. Cardiac and/or adipose tissue from 48 patients carrying 21 different non-TTRV30M mutations were examined, as well as 7 non-Swedish ATTRV30M patients. Fibril type was determined with western blotting and compared to the patients´ age of onset and degree of cardiomyopathy.

    Non-Swedish V30M patients showed the same correlation as described for Swedish V30M patients, with fibrils of only full-length ATTR (type B) linked to less myocardial involvement. In contrast, all patients with non-V30M mutations had a fibril composition with ATTR fragments (type A). Some of these patients had onset of disease at young age. The vast majority had increased thickness of left cardiac ventricle, but a few individuals had values within normal limits.

    This study shows that a fibril composition with fragmented ATTR is very common in ATTR amyloidosis. It also suggests that fibrils composed of only full-length ATTR is an exception, perhaps only found among young ATTRV30M amyloidosis patients.

  • 74.
    Itsenko, Oleksiy
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Blom, Elisabeth
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Långström, Bengt
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Kihlberg, Tor
    The Use of Lithium Amides in the Palladium-Mediated Synthesis of [Carbonyl-11C]Amides2007In: European Journal of Organic Chemistry, ISSN 1434-193X, E-ISSN 1099-0690, Vol. 2007, no 26, p. 4337-4342Article in journal (Refereed)
    Abstract [en]

    Weakly nucleophilic amines were converted into the corresponding lithium amides and used in either one- or two-pot palladium mediated-reactions with [11C]carbon monoxide and aryl iodides. It was found that palladium acyl complexes may be prepared in a separate step and have sufficient lifetime to be used in a subsequent reaction with a nucleophile. This two-pot procedure was used for the labelling synthesis of eleven amides (nine of which are analogues of WAY-100635, a 5-HT1A radioligand) from weakly nucleophilic amines. The results were compared to a direct one-pot procedure using lithium amides. Both approaches extend the scope of palladium-mediated carbonylation using [11C]carbon monoxide and aryl iodides allowing use of weakly nucleophilic amines.

  • 75.
    Jansson-Löfmark, Rasmus
    et al.
    Sahlgrenska Academy at University of Gothenburg, Unit for Pharmacokinetics and drug metabolism, Göteborg, Sweden.
    Römsing, Susanne
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Center for Clinical Research Dalarna.
    Albers, E.
    Sahlgrenska Academy at University of Gothenburg, Unit for Pharmacokinetics and drug metabolism, Göteborg, Sweden.
    Ashton, M.
    Sahlgrenska Academy at University of Gothenburg, Unit for Pharmacokinetics and drug metabolism, Göteborg, Sweden .
    Determination of eflornithine enantiomers in plasma by precolumn derivatization with o-phthalaldehyde-N-acetyl-l-cysteine and liquid chromatography with UV detection2010In: BMC Biomedical chromotography, ISSN 0269-3879, E-ISSN 1099-0801, Vol. 24, no 7, p. 768-773Article in journal (Refereed)
    Abstract [en]

    A bioanalytical method for indirect determination of eflornithine enantiomers in 75 mu L human plasma has been developed and validated. L- and D-eflornithine were derivatized with o-phthalaldehyde and N-acetyl-L-cysteine to generate diastereomers which were separated on two serially connected Chromolith Performance columns (RP-18e 100 x 4.6 mm i.d.) by a isocratic flow followed by a gradient flow for elution of endogenous compounds. The diastereomers were detected with UV (340 nm). The between-day precisions for L- and D-eflornithine in plasma were 8.4 and 2.3% at 3 mu m, 4.0 and 5.1% at 400 mu m, and 2.0 and 3.7% at 1000 mu m. The lower limit of quantification was determined to be 1.5 mu m, at which precision was 14.9 and 9.9% for 1- and D-eflornithine, respectively.

  • 76. Johansson, E. M. J.
    et al.
    Odelius, M.
    Gorgoi, M.
    Karis, Olof
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Materials Science, Surface and Interface Science.
    Ovsyannikov, R.
    Schafers, S.
    Svensson, Svante
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Materials Science, Surface and Interface Science.
    Siegbahn, Hans
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Materials Science, Surface and Interface Science.
    Rensmo, Håkan
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Materials Science, Surface and Interface Science.
    Valence Electronic Structure Of Ruthenium Based Complexes Probed By Photoelectron Spectroscopy At High Kinetic Energy (Hike) And Modeled By Dft Calculations2008In: Chemical Physics Letters, ISSN 0009-2614, E-ISSN 1873-4448, Vol. 464, no 4-6, p. 192-197Article in journal (Refereed)
    Abstract [en]

    The valence electronic structure of a series of molecular films containing ruthenium polypyridine complexes has been investigated by photoelectron spectroscopy (PES) at high kinetic energy (HIKE) using hard X-ray. The experiment shows the possibility to experimentally probe the metal contribution to the valence spectra in a bulk sensitive mode. Specifically to directly follow the Ru 4d contribution to the highest occupied molecular orbitals of such complexes. The experimental spectra are accurately modeled by DFT calculations only if a crystal structure environment is taken into account showing the importance of intermolecular interaction for modeling the electronic structure of such complexes.

  • 77. Kakehashi, Rie
    et al.
    Karlsson, Göran
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Physical Chemistry.
    Almgren, Mats
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Physical Chemistry.
    Stomatosomes, blastula vesicles and bilayer disks: morphological richness of structures formed in dilute aqueous mixtures of a cationic and an anionic surfactant2009In: Journal of Colloid and Interface Science, ISSN 0021-9797, E-ISSN 1095-7103, Vol. 331, no 2, p. 484-493Article in journal (Refereed)
    Abstract [en]

    Cryogenic transmission electron microscopy (cryoTEM) was used to study the structures formed in mixts. of sodium dodecylsulfate (SDS) and dodecyltrimethylammonium bromide (DTAB) in dil. aq. solns. with 0-300 mM NaBr. The DTAB mole fraction, X, was in the range 0.2-0.4, limited at 25 DegC by pptn. of solid DTA-DS at X = 0.38 without salt to X = 0.25 at 300 mM NaBr. At a total surfactant concn. of 100 mM the samples sepd. into two liq. phases (the bottom phase birefringent) within a narrow (+-0.01 mol fraction units) compn. range. At the mid-point X varied from 0.32 without salt to 0.22 at 300 mM NaBr. Elemental anal. of C, S, O, and N in the sepd. phases of a sample with 100 mM NaBr and X = 0.26 showed the top phase to contain almost only SDS at a low concn., 14 mM, and the bottom phase 175 mM total surfactant, with X = 0.27 . Elemental anal. on samples without added salt gave erratic results, indicating problems in the phys. sepn. of the phases. The cryoTEM survey of the sepd. phases revealed similar problems. Without salt both phases showed similar structures, whereas the top phase in the sample with added salt was void of structures larger than small micelles. The cryoTEM survey revealed a variety of structures being simultaneously present in most samples. A general trend with increasing X was an evolution from globular micelles, over disks, bands, branched bands transforming into sparse webs, perforated bilayer structures, and finally smooth bilayers. Increasing salt and total surfactant concns. resulted in the emergence of structures with smaller mean curvature at lower X. Perforated bilayers were found in samples with 100 mM or less of added salt, and usually persisted to DTAB contents where ppts. appeared. The porous bilayers seemed to derive from sparse webs of band-like structures, and the hole size decreased with increasing X and salt concn. Two types of recurrent structures were noticed: blastula aggregates, seemingly an intermediate structure transforming crumpled bilayers into vesicles of similar size (diam. 400-500 ANG.), obsd. over a broad range of conditions, and at 100 mM total surfactant concn. and 50 mM added salt or more a type of regular disks with a diam. of 180 +- 30 ANG.

  • 78.
    Karlsson, Oskar
    et al.
    Karolinska Inst, Dept Clin Neurosci, Ctr Mol Med, Stockholm, Sweden; Harvard TH Chan Sch Publ Hlth, Dept Environm Hlth, Boston, MA USA.
    Rodosthenous, Rodosthenis S
    Harvard TH Chan Sch Publ Hlth, Dept Environm Hlth, Boston, MA USA.
    Jara, Calvin
    Icahn Sch Med Mt Sinai, Dept Prevent Med, New York, NY 10029 USA.
    Brennan, Kasey J
    Harvard TH Chan Sch Publ Hlth, Dept Environm Hlth, Boston, MA USA.
    Wright, Robert O
    Icahn Sch Med Mt Sinai, Dept Prevent Med, New York, NY 10029 USA; Icahn Sch Med Mt Sinai, Dept Pediat, Kravis Childrens Hosp, New York, NY 10029 USA.
    Baccarelli, Andrea A
    Harvard TH Chan Sch Publ Hlth, Dept Environm Hlth, Boston, MA USA.
    Wright, Rosalind J
    Icahn Sch Med Mt Sinai, Dept Prevent Med, New York, NY 10029 USA; Icahn Sch Med Mt Sinai, Dept Pediat, Kravis Childrens Hosp, New York, NY 10029 USA.
    Detection of long non-coding RNAs in human breastmilk extracellular vesicles: Implications for early child development.2016In: Epigenetics, ISSN 1559-2294, E-ISSN 1559-2308, Vol. 11, no 10, p. 721-729Article in journal (Refereed)
    Abstract [en]

    Breastmilk has many documented beneficial effects on the developing human infant, but the components of breastmilk that influence these developmental pathways have not been fully elucidated. Increasing evidence suggests that non-coding RNAs encapsulated in extracellular vesicles (EVs) represent an important mechanism of communication between the mother and child. Long non-coding RNAs (lncRNAs) are of particular interest given their key role in gene expression and development. However, it is not known whether breastmilk EVs contain lncRNAs. We used qRT-PCR to determine whether EVs isolated from human breastmilk contain lncRNAs previously reported to be important for developmental processes. We detected 55 of the 87 screened lncRNAs in EVs from the 30 analyzed breastmilk samples, and CRNDE, DANCR GAS5, SRA1 and ZFAS1 were detected in >90% of the samples. GAS5, SNHG8 and ZFAS1 levels were highly correlated (Spearman's rho>0.9; P<0.0001), which may indicate that the loading of these lncRNAs into breastmilk EVs is regulated by the same pathways. The detected lncRNAs are important epigenetic regulators involved in processes such as immune cell regulation and metabolism. They may target a repertoire of recipient cells in offspring and could be essential for child development and health. Further experimental and epidemiological studies are warranted to determine the impact of breastmilk EV-encapsulated lnRNAs in mother to child signaling.

  • 79.
    Karlsson, Susanne
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science.
    Modin, Judit
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Becker, Hans-Christian
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science.
    Hammarström, Leif
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science.
    Grennberg, Helena
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    How Close Can You Get?: Studies of Ultrafast Light-Induced Processes in Ruthenium-[60] Fullerene Dyads with Short Pyrazolino and Pyrrolidino Links2008In: Inorganic Chemistry, ISSN 0020-1669, E-ISSN 1520-510X, Vol. 47, no 16, p. 7286-7294Article in journal (Refereed)
    Abstract [en]

    Two pyrazoline- and one pyrrolidine-bridged Ru(II)bipyridine-[60]fullerene dyads have been prepared and studied by ultrafast time-resolved spectroscopy. A silver-assisted synthesis route, in which Ag(I) removes the chlorides from the precursor complex Ru(bpy)(2)Cl-2 facilitates successful coordination of the [60]fullerene-substituted third ligand. Upon light excitation of the ruthenium moiety, the emission was strongly quenched by the fullerene. The main quenching mechanism is an exceptionally fast direct energy transfer (k(obs) > , 1 x 10(12) s(-1) in the pyrazoline-bridged dyads), resulting in population of the lowest excited triplet state of fullerene. No evidence for electron transfer was found, despite the extraordinarily short donor-acceptor distance that could kinetically favor that process. The observations have implications on the ongoing development of devices built from Ru-polypyridyl complexes and nanostructured carbon, such as C-60 or nanotubes.

  • 80.
    Karlsson, Susanne
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science, Chemical Physics.
    Streich, Daniel
    Johansson, Olof
    Anderlund, Magnus
    Becker, Hans-Christian
    Hammarström, Leif
    Double-pulse Excitation of a Mn2-Ru(II)-Naphthalenediimide Triad: Challenges for Accumulative Electron TransferManuscript (preprint) (Other academic)
  • 81. Kempf, Stefan J.
    et al.
    Buratovic, Sonja
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Organismal Biology, Environmental toxicology.
    von Toerne, Christine
    Moertl, Simone
    Stenerlöw, Bo
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Hauck, Stefanie M.
    Atkinson, Michael J.
    Eriksson, Per
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Organismal Biology, Environmental toxicology.
    Tapio, Soile
    Ionising Radiation Immediately Impairs Synaptic Plasticity-Associated Cytoskeletal Signalling Pathways in HT22 Cells and in Mouse Brain: An In Vitro/In Vivo Comparison Study2014In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 9, no 10, p. e110464-Article in journal (Refereed)
    Abstract [en]

    Patients suffering from brain malignancies are treated with high-dose ionising radiation. However, this may lead to severe learning and memory impairment. Preventive treatments to minimise these side effects have not been possible due to the lack of knowledge of the involved signalling pathways and molecular targets. Mouse hippocampal neuronal HT22 cells were irradiated with acute gamma doses of 0.5 Gy, 1.0 Gy and 4.0 Gy. Changes in the cellular proteome were investigated by isotope-coded protein label technology and tandem mass spectrometry after 4 and 24 hours. To compare the findings with the in vivo response, male NMRI mice were irradiated on postnatal day 10 with a gamma dose of 1.0 Gy, followed by evaluation of the cellular proteome of hippocampus and cortex 24 hours post-irradiation. Analysis of the in vitro proteome showed that signalling pathways related to synaptic actin-remodelling were significantly affected at 1.0 Gy and 4.0 Gy but not at 0.5 Gy after 4 and 24 hours. We observed radiation-induced reduction of the miR-132 and Rac1 levels; miR-132 is known to regulate Rac1 activity by blocking the GTPase-activating protein p250GAP. In the irradiated hippocampus and cortex we observed alterations in the signalling pathways similar to those in vitro. The decreased expression of miR-132 and Rac1 was associated with an increase in hippocampal cofilin and phospho-cofilin. The Rac1-Cofilin pathway is involved in the modulation of synaptic actin filament formation that is necessary for correct spine and synapse morphology to enable processes of learning and memory. We suggest that acute radiation exposure leads to rapid dendritic spine and synapse morphology alterations via aberrant cytoskeletal signalling and processing and that this is associated with the immediate neurocognitive side effects observed in patients treated with ionising radiation.

  • 82.
    Larsson, Dhana E
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Analyses of Dose-Response and Mechanistic Action of Different Anti-Cancer Drugs for Neuroendocrine Tumor Cell Lines2011Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Cancer is a disease with poor response rates on available treatments. Problems with resistance and intolerance against cancer drugs are major reasons for failure of the drugs. The need to discover new cancer drugs is important. In this thesis screening of new cancer drugs and evaluation of their mechanism of action are discussed. The aim of the thesis was to find new compounds active against neuroendocrine tumors (NETs).

    In paper I, we screened 1280 substances on two bronchial carcinoid cell lines and one pancreatic carcinoid cell line. Eleven of these compounds were found to have antitumor activity at low concentrations. The most active agents were brefeldin A, emetine, bortezomib and idarubicin, having IC50 values (the concentration of the drug where > 50% of the cells die) < 1μM. In addition, sanguinarine, Bay11-7085, mitoxantrone, doxorubicin, β-lapachone, NSC 95397 and CGP- 74514A were active with IC50 values < 10 μM.

     In paper II, additional studies have been undertaken to investigate the combination effect of the most active drugs with conventional cytotoxic drugs used in clinical practice. If synergistic or additive effects are found, drugs with different mechanism of action and toxicity profiles may be combined, making it possible to reduce the toxic effects yet maintaining the antitumor activity.

    In paper III, studies were undertaken to find the mechanistic action, apoptosis or necrosis, of the drugs NSC 95397, brefeldin A, bortezomib and sanguinarine in NETs. All four drugs were shown to induce caspase-3 activity and nuclear fragmentation/condensation in the neuroendocrine tumor cell lines, indicating that their antitumor activity was induction of apoptosis.

    In paper IV, the mechanism of action was studied for CGP-74514A and emetine. Both drugs worked by induction of apoptosis. In addition, their cytotoxic activity was studied in a three-dimensional model, the in vitro hollow fiber model. The Hollow Fiber model permits more realistic simulation of in vivo drug effects in a controlled system providing data that more accurately reflects biological responses. Our results showed that the hollow fiber model may be suitable for studies of new drugs in the neuroendocrine tumor cell lines.

    List of papers
    1. Identification and evaluation of potential anti-cancer drugs on human neuroendocrine tumor cell lines
    Open this publication in new window or tab >>Identification and evaluation of potential anti-cancer drugs on human neuroendocrine tumor cell lines
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    2006 (English)In: Anticancer Research, ISSN 0250-7005, E-ISSN 1791-7530, Vol. 26, no 6B, p. 4125-4129Article in journal (Refereed) Published
    Abstract [en]

    The aim of this study was to investigate drug sensitivity in neuroendocrine tumor cell lines. Materials and Methods: In vitro drug sensitivity screening was performed using the fluorometric microculture cytotoxicity assay in one human pancreatic carcinoid and two human bronchial carcinoid cell lines. In addition, a normal human retinal pigment epithelial cell line was used for comparison. A total of 18 drugs with different mechanisms of action were tested. Results: The most active agents were brefeldin A, emetine, bortezomib and idarubicin, having IC50 values < 1 μM in all four cell lines. In addition, the three tumor cell lines showed sensitivity for sanguinarine, Bay11-7085, mitoxantrone, doxorubicin, β-lapachone, NSC 95397 and CGP-74514A. Conclusion: The cell lines were sensitive for several drugs acting in different ways, covering a broad spectrum of mechanisms of action. Some of these compounds may possibly be used in clinical trials and show therapeutic effect in patients with neuroendocrine tumors.

    Keywords
    Drug sensitivity, FMCA, Neuroendocrine tumors
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-24875 (URN)000243040300005 ()17201123 (PubMedID)
    Available from: 2007-02-08 Created: 2007-02-08 Last updated: 2017-12-07Bibliographically approved
    2. Combination analyses of anti-cancer drugs on human neuroendocrine tumor cell lines
    Open this publication in new window or tab >>Combination analyses of anti-cancer drugs on human neuroendocrine tumor cell lines
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    2009 (English)In: Cancer Chemotherapy and Pharmacology, ISSN 0344-5704, E-ISSN 1432-0843, Vol. 65, no 1, p. 5-12Article in journal (Refereed) Published
    Abstract [en]

    PURPOSE:

    There is a large need for better pharmacological treatment of neuroendocrine tumors. The aim of this study was to investigate and quantify the cytotoxic potentiating effects resulting from a combination of five substances, NSC 95397, emetine, CGP-74514A hydrochloride, Brefeldin A and sanguinarine chloride, chosen from a previous screening of 1,280 pharmacologically active agents on neuroendocrine tumor cells, with standard cytotoxic agents currently used in the treatment of neuroendocrine tumors.

    METHOD:

    The human pancreatic carcinoid cell line BON-1, human typical bronchial carcinoid cell line NCI-H727 and the human atypical bronchial carcinoid cell line NCI-H720 were used. Combinations between doxorubicin, etoposide, oxaliplatin, docetaxel, and each one of the five agents were studied and simultaneous exposures were explored using the median-effect method.

    RESULTS:

    Most of the combinations of NSC-95397 and emetine with doxorubicin, etoposide, docetaxel, and oxaliplatin showed synergism, and their remaining combinations were additive. Almost all of the CGP-74514A hydrochloride interactions were additive, while brefeldin A and sanguinarine displayed less synergy but more additive and antagonistic interactions in combination with the standard drugs.

    CONCLUSION:

    The synergistic and additive interactions make NSC-95397, emetine, and CGP-74514A hydrochloride potential candidates for incorporation into combination chemotherapy regimens and these drugs might be the suitable candidates for further clinical studies in patients with bronchial carcinoids and pancreatic endocrine tumors.

    Keywords
    Neuroendocrine tumors, Combination, Chemotherapy, Drug-sensitivity
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-102564 (URN)10.1007/s00280-009-0997-6 (DOI)000269916800002 ()19381631 (PubMedID)
    Available from: 2009-05-08 Created: 2009-05-08 Last updated: 2017-12-13Bibliographically approved
    3. The Cytotoxic Agents NSC-95397, Brefeldin A, Bortezomib and Sanguinarine Induce Apoptosis in Neuroendocrine Tumors In Vitro
    Open this publication in new window or tab >>The Cytotoxic Agents NSC-95397, Brefeldin A, Bortezomib and Sanguinarine Induce Apoptosis in Neuroendocrine Tumors In Vitro
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    2010 (English)In: Anticancer Research, ISSN 0250-7005, E-ISSN 1791-7530, Vol. 30, no 1, p. 149-156Article in journal (Refereed) Published
    Abstract [en]

    The aim of this study was to investigate the apoptosis resulting from NSC 95397, brefeldin A, bortezomib and sanguinarine in neuroendocrine tumor cell lines. Materials and Methods: A multiparametric high-content screening assay for measurement of apoptosis was used. The human pancreatic carcinoid cell line, BON-1, human typical bronchial carcinoid cell line NCI-H727 and the human atypical bronchial carcinoid cell line NCI-H720 were tested. After incubation with cytotoxic drugs, the DNA-binding dye Hoechst 33342, fluorescein-tagged probes that covalently bind active caspase-3 and chloromethyl-X-rosamine to detect mitochondrial membrane potential were added. Image acquisition and quantitative measurement of fluorescence was performed using automated image capture and analysis instrument ArrayScan. In addition, nuclear morphology was examined on microscopic slides stained with May-Grunewald-Giemsa. Results: A time- and dose-dependent activation of caspase-3 and increase in nuclear fragmentation and condensation were observed for the drugs using a multiparametric apoptosis assay. These results were confirmed with nuclear morphological examination on microscopic slides. Conclusion: NSC 95397, brefeldin A, bortezomib and sanguinarine induced caspase-3 activation with modest changes in nuclear morphology.

    Keywords
    Apoptosis, caspase activity, NSC 95397, brefeldin A
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-137871 (URN)000274537400020 ()20150630 (PubMedID)
    Available from: 2010-12-16 Created: 2010-12-16 Last updated: 2017-12-11Bibliographically approved
    4.
    The record could not be found. The reason may be that the record is no longer available or you may have typed in a wrong id in the address field.
  • 83.
    Li, Su-Chen
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Endocrine oncology.
    Khan, Mohid
    Caplin, Martyn
    Meyer, Tim
    Öberg, Kjell
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Endocrine oncology.
    Giandomenico, Valeria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Endocrine oncology.
    Circulating microRNA detection in small intestinal neuroendocrine tumor patients treated with somatostatin analogs2014Article in journal (Refereed)
  • 84.
    Lignell, Martin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science, Chemical Physics.
    Tegler, Lotta
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Becker, Hans-Christian
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science, Chemical Physics.
    Hydrated and Dehydrated Tertiary Interactions - Opening and Closing - of a Four-helix Bundle Peptide2009In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 97, no 2, p. 572-580Article in journal (Refereed)
    Abstract [en]

    The structural heterogeneity and thermal denaturation of a dansyl-labeled four-helix bundle homodimeric peptide has been studied with steady state and time-resolved fluorescence spectroscopy and with circular dichroism. At room temperature the fluorescence decay of the polarity-sensitive dansyl, located in the hydrophobic core region, can be described by a broad distribution of fluorescence lifetimes, reflecting the heterogeneous microenvironment. However, the lifetime distribution is nearly bimodal, which we ascribe to the presence of two major conformational subgroups. Since the fluorescence lifetime reflects the water content of the four-helix bundle conformations we can use the lifetime analysis to monitor the change of hydration state of the hydrophobic core of the four-helix bundle. Increasing the temperature from 9 °C to 23 °C leads to an increased population of molten-globule-like conformations with a less ordered helical backbone structure. The fluorescence emission maximum remains constant in this temperature interval, and the hydrophobic core is not strongly affected. Above 30 °C the structural dynamics involve transient openings of the four-helix bundle structure as evidenced by the emergence of a water-quenched component and less negative CD. Above 60 °C the homodimer starts to dissociate, as shown by the increasing loss of CD and narrow, short-lived fluorescence lifetime distributions.

  • 85.
    Lindahl, Sofia
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Ekman, Anna
    Khan, Samiullah
    Wennerberg, Christina
    Börjesson, Pål
    Sjöberg, Per J.R.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Nordberg Karlsson, Eva
    Turner, Charlotta
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Exploring the possibility of using a thermostable mutant of β-glucosidase for rapid hydrolysis of quercetin glucosides in hot water2010In: Green Chemistry, ISSN 1463-9262, E-ISSN 1463-9270, Vol. 12, no 1, p. 159-168Article in journal (Refereed)
    Abstract [en]

    The antioxidant quercetin was extracted from yellow onion waste and converted to its aglycone form by a combination of subcritical water extraction and enzymatic hydrolysis. The hydrolytic step was catalysed by a double residue (N221S, P342L) mutant of the thermostable beta-glucosidase (TnBgl1A), isolated from the thermophile Thermotoga neapolitana and cloned and produced in E. coli. The activity of wt TnBgl1A was shown to be dependent on the position of the glucosylation on the quercetin backbone, favouring hydrolysis of quercetin-4'-glucoside over quercetin-3-glucoside. The mutated variant of the enzyme harboured a mutation in the +2 sub-site (N221S) and showed increased catalytic efficiency in quercetin-3-glucoside hydrolysis and also to a certain extent hydrolysis of quercetin-4'-glucoside. The mutated enzyme was used directly in yellow onion extracts, prepared by subcritical water extraction, resulting in complete hydrolysis of the glucosylated flavonoids quercetin-3,4'-diglucoside, quercetin-4'-glucoside, quercetin-3-glucoside, isorhamnetin-4'-glucoside and isorhamnetin-3,4'-diglucoside. To complete hydrolysis within five minutes, 3 mg of TnBgl1A_N221S was used per gramme of onion (dry weight). A life cycle assessment was done to compare the environmental impact of the new method with a conventional solid-liquid extraction-and-hydrolysis method utilising aqueous methanol and hydrochloric acid. Comparison of the methods showed that the new method is preferable regarding primary energy consumption and global warming potential. Another advantage of this method is that handling of toxic chemicals (methanol and HCl) is avoided. This shows that combined subcritical water extraction/enzyme hydrolysis is both a fast and sustainable method to obtain quercetin from onion waste.

  • 86.
    Lindgren, N. Johan V.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Expanding the Amino Acid Alphabet by Design: Enhanced and Controlled Catalytic Activity in Folded Polypeptide Catalysts2009Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    This thesis addresses structure and reactivity of polypeptide catalysts in reactions that mimic the hydrolysis of RNA and DNA. A designed helix-loop-helix motif was used as a scaffold where the amino acid residues were systematically varied. The reactivity of a previously reported catalyst, HNI, was evaluated and the catalytic residues of the active site, Arg and His, were replaced in a stepwise manner by the artificial amino acid Gcp. Gcp has a guanidinocarbonyl pyrrole side chain, i.e. a side chain that mimics that of Arg but with a lower pKa. Gcp was used to replace both histidine and arginine in the polypeptide catalysts and was able to bind phosphate as well as carry out general base catalysis.

    The parent polypeptide HNI was shown to catalyse phosphoryl transfer reactions of phosphodiesters in an active site with two His and two Arg residues. The performance of the active site was improved by the introduction of two Tyr residues to form the catalyst HJ1 designed to provide nucleophilic catalysis in the hydrolysis of DNA model substrates.

    To improve the catalytic activity beyond that of HJI, Gcp was introduced to replace Arg and His residues in the HN1 scaffold. The designed catalyst JL3 was capable of a 150-fold rate enhancement compared to HNI in the reaction of the substrate HPNP, representing the first step in RNA hydrolysis. Mechanistic studies of JL3 catalysis suggested that the pKa value of the Gcp residue in the folded polypeptides was around 5. In combination with the observation of a solvent kinetic isotope effect of 1.7 the Gcp residue was proposed to provide general base catalysis and transition state stabilisation in the reaction of uridine 3′-2,2,2-trichloroethylphosphate, a realistic RNA model with a leaving group pKa of 12.5. The JL3 polypeptide catalyst followed saturation kinetics with a kcat/KM of 1.08 x 10-3 M-1s-1.

    The introduction of a designed photoswitchable amino acid in a catalytic polypeptide allowed the activity of the polypeptide in the reaction with an activated ester to be under photochemical control. Photoisomerization of this switch altered the structure of the polypeptide and affected the catalytic activity of the polypeptide catalyst.

    The chemical synthesis of designed molecules expands the amino acid alphabet and makes it possible to downsize enzymatic functions. It opens up possibilities for the production of novel biocatalysts that can catalyse natural as well as non-natural reactions.

    List of papers
    1.
    The record could not be found. The reason may be that the record is no longer available or you may have typed in a wrong id in the address field.
    2. Downsizing of Enzymes by Chemical Methods: Arginine Mimics with Low pK(a) Values Increase the Rates of Hydrolysis of RNA Model Compounds
    Open this publication in new window or tab >>Downsizing of Enzymes by Chemical Methods: Arginine Mimics with Low pK(a) Values Increase the Rates of Hydrolysis of RNA Model Compounds
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    2009 (English)In: Angewandte Chemie International Edition, ISSN 1433-7851, E-ISSN 1521-3773, Vol. 48, no 36, p. 6722-6725Article in journal (Refereed) Published
    Keywords
    de novo design, homogeneous catalysis, hydrolysis, peptides, phosphodiesters
    National Category
    Chemical Sciences
    Identifiers
    urn:nbn:se:uu:diva-100486 (URN)10.1002/anie.200900595 (DOI)000269361300034 ()19655357 (PubMedID)
    Available from: 2009-04-01 Created: 2009-04-01 Last updated: 2017-12-13Bibliographically approved
    3. Design and Synthesis of Folded Polypeptides that Carry the Guandiniumcarbonyl Pyrrole Residue – a Versatile Amino Acid with High Helix Propensity
    Open this publication in new window or tab >>Design and Synthesis of Folded Polypeptides that Carry the Guandiniumcarbonyl Pyrrole Residue – a Versatile Amino Acid with High Helix Propensity
    (English)Manuscript (Other (popular science, discussion, etc.))
    Identifiers
    urn:nbn:se:uu:diva-100488 (URN)
    Available from: 2009-04-01 Created: 2009-04-01 Last updated: 2010-01-14
    4. Photochemical Regulation of an Artificial Hydrolase by a Backbone Incorporated Tertiary Structure Switch
    Open this publication in new window or tab >>Photochemical Regulation of an Artificial Hydrolase by a Backbone Incorporated Tertiary Structure Switch
    2009 (English)In: Chemistry - A European Journal, ISSN 0947-6539, E-ISSN 1521-3765, Vol. 15, no 2, p. 501-505Article in journal (Refereed) Published
    Abstract [en]

    A stilbene chromophore has been incorporated into the turn region of a 42 amino acid peptide, linking two helical peptide sections. Spatial proximity between these sections, as well as aggregation into dimers, is required to facilitate the catalytic function of this artificial hydrolase. Photomodulation of the hydrolase activity results in an increase of the activity of 42 % upon switching from the trans to the cis isomer of the chromophore. This is rationalized by a change in the aggregation state of the peptidomimetic, which is supported by diffusion coefficients obtained from PFG-NMR experiments. The results show that incorporation of a small, relatively flexible chromophore into a large peptide is capable of inducing a considerable change in tertiary structure and thus, functionality.

    Keywords
    aggregation, chromophores, enzyme catalysis, peptidomimetics
    National Category
    Organic Chemistry
    Research subject
    Organic Chemistry
    Identifiers
    urn:nbn:se:uu:diva-99821 (URN)10.1002/chem.200801808 (DOI)000262491500025 ()
    Available from: 2009-03-31 Created: 2009-03-20 Last updated: 2017-12-13Bibliographically approved
  • 87.
    Lindh, Jonas
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Organic Pharmaceutical Chemistry. ORGFARM.
    Sjöberg, Per J. R.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Larhed, Mats
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Organic Pharmaceutical Chemistry.
    Synthesis of aryl ketones by palladium(II)-catalyzed decarboxylative addition of benzoic acids to nitriles2010In: Angewandte Chemie International Edition, ISSN 1433-7851, E-ISSN 1521-3773, Vol. 49, no 42, p. 7733-7737Article in journal (Refereed)
  • 88.
    Lindskog, Cecilia
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    The potential clinical impact of the tissue-based map of the human proteome2015In: Expert Review of Proteomics, ISSN 1478-9450, E-ISSN 1744-8387, Vol. 12, no 3, p. 213-215Article in journal (Other academic)
    Abstract [en]

    Since the first draft of the human genome sequence was published, several attempts have been made to map the human proteome, the functional representation of the genome. One such initiative is the Human Protein Atlas project, which recently released a tissue-based map of the human proteome. The Human Protein Atlas is based on the combination of transcriptomics and antibody-based proteomics for mapping the human proteome down to the single cell level. The comprehensive publicly available database contains more than 13 million unique immunohistochemistry images and provides an excellent resource for exploration and investigation of future drug targets and disease biomarkers.

  • 89. Liu, C
    et al.
    Marioni, R E
    Hedman, Åsa K
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Molecular epidemiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Pfeiffer, L
    Tsai, P-C
    Reynolds, L M
    Just, A C
    Duan, Q
    Boer, C G
    Tanaka, T
    Elks, C E
    Aslibekyan, S
    Brody, J A
    Kühnel, B
    Herder, C
    Almli, L M
    Zhi, D
    Wang, Y
    Huan, T
    Yao, C
    Mendelson, M M
    Joehanes, R
    Liang, L
    Love, S-A
    Guan, W
    Shah, S
    McRae, A F
    Kretschmer, A
    Prokisch, H
    Strauch, K
    Peters, A
    Visscher, P M
    Wray, N R
    Guo, X
    Wiggins, K L
    Smith, A K
    Binder, E B
    Ressler, K J
    Irvin, M R
    Absher, D M
    Hernandez, D
    Ferrucci, L
    Bandinelli, S
    Lohman, K
    Ding, J
    Trevisi, L
    Gustafsson, Stefan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Molecular epidemiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Sandling, Johanna K.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Molecular Medicine. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Stolk, L
    Uitterlinden, A G
    Yet, I
    Castillo-Fernandez, J E
    Spector, T D
    Schwartz, J D
    Vokonas, P
    Lind, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Cardiovascular epidemiology.
    Li, Y
    Fornage, M
    Arnett, D K
    Wareham, N J
    Sotoodehnia, N
    Ong, K K
    van Meurs, J B J
    Conneely, K N
    Baccarelli, A A
    Deary, I J
    Bell, J T
    North, K E
    Liu, Y
    Waldenberger, M
    London, S J
    Ingelsson, Erik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Molecular epidemiology. Uppsala University, Science for Life Laboratory, SciLifeLab. Department of Medicine, Division of Cardiovascular Medicine, Stanford University School of Medicine, Stanford, CA, USA.
    Levy, D
    A DNA methylation biomarker of alcohol consumption.2018In: Molecular Psychiatry, ISSN 1359-4184, E-ISSN 1476-5578, Vol. 23, p. 422-433Article in journal (Refereed)
    Abstract [en]

    The lack of reliable measures of alcohol intake is a major obstacle to the diagnosis and treatment of alcohol-related diseases. Epigenetic modifications such as DNA methylation may provide novel biomarkers of alcohol use. To examine this possibility, we performed an epigenome-wide association study of methylation of cytosine-phosphate-guanine dinucleotide (CpG) sites in relation to alcohol intake in 13 population-based cohorts (ntotal=13 317; 54% women; mean age across cohorts 42-76 years) using whole blood (9643 European and 2423 African ancestries) or monocyte-derived DNA (588 European, 263 African and 400 Hispanic ancestry) samples. We performed meta-analysis and variable selection in whole-blood samples of people of European ancestry (n=6926) and identified 144 CpGs that provided substantial discrimination (area under the curve=0.90-0.99) for current heavy alcohol intake (⩾42 g per day in men and ⩾28 g per day in women) in four replication cohorts. The ancestry-stratified meta-analysis in whole blood identified 328 (9643 European ancestry samples) and 165 (2423 African ancestry samples) alcohol-related CpGs at Bonferroni-adjusted P<1 × 10(-7). Analysis of the monocyte-derived DNA (n=1251) identified 62 alcohol-related CpGs at P<1 × 10(-7). In whole-blood samples of people of European ancestry, we detected differential methylation in two neurotransmitter receptor genes, the γ-Aminobutyric acid-A receptor delta and γ-aminobutyric acid B receptor subunit 1; their differential methylation was associated with expression levels of a number of genes involved in immune function. In conclusion, we have identified a robust alcohol-related DNA methylation signature and shown the potential utility of DNA methylation as a clinically useful diagnostic test to detect current heavy alcohol consumption.

  • 90.
    Ljungström, Olle
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Ekman, Pia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Glucagon-induced phosphorylation of pyruvate kinase (type L) in rat liver slices1977In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 78, no 4, p. 1147-1155Article in journal (Refereed)
    Abstract [en]

    The effect of glucagon on the phosphorylation of pyruvate kinase in 32P-labelled slices from rat liver was investigated. Pyruvate kinase was isolated by immunoadsorbent chromatography. The enzyme was partially phosphorylated in the absence of added hormone (0.2 mol of phosphate/mol of enzyme subunit). Upon incubation with 10−7 M glucagon, the incorporation of [32P]phosphate was 0.6–0.7 mol/mol of enzyme subunit. Concomitantly, the concentration of intracellular cyclic 3′,5′-AMP increased from 0.3 to 3.2 μM. The phosphorylation inhibited the enzyme activity at low concentrations of phosphoenolpyruvate (60% at 0.5 mM). Almost maximal phosphorylation of the enzyme was reached within 2 min after the addition of glucagon. The concentration of hormone giving half maximal effect on the pyruvate kinase phosphorylation was about 7×10−9M. The inactivation of the enzyme paralleled the increase in phosphorylation. It is concluded that pyruvate kinase is phosphorylated in the intact liver cell.

  • 91.
    Loog, Mart
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Järv, Jaak
    Tartu University Estland.
    Engström, Lorentz
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Influence of enzyme inactivation on the kinetics of peptide phosphorylation by protein kinase C1997In: Proceedings of the Estonian Academy of Sciences, ISSN 1736-6046, E-ISSN 1736-7530, Vol. 46, no 3, p. 102-111Article in journal (Refereed)
  • 92.
    Lundström, Yasmin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical and experimental pathology.
    Lundström, Patrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical and experimental pathology.
    Popova, Svetlana
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical and experimental pathology.
    Lindblom, Rickard P.F.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Thoracic Surgery. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care.
    Alafuzoff, Irina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical and experimental pathology.
    Detection of Changes in Immunohistochemical Stains Caused by Postmortem Delay and Fixation Time2019In: Applied immunohistochemistry & molecular morphology (Print), ISSN 1541-2016, E-ISSN 1533-4058, Vol. 27, no 3, p. 238-245Article in journal (Refereed)
    Abstract [en]

    In this study, we have systematically assessed the influence of postmortem delay (PMD) and fixation time (FT) on the immunohistochemical (IHC) staining outcome. The IHC method is frequently applied on surgical and postmortem samples in diagnostics and research. To replicate the routine situation, brain tissues from pigs were exposed to either storage in a refrigerator (+8°C), that is, PMD (1 to 168 h), or fixed in 10% buffered formalin, that is, FT (18 to 94 d). Subsequently, the tissue was routinely processed into paraffin blocks to enable construction of tissue microarrays (TMA). Sections cut from the TMA blocks were stained applying 13 different antibodies directed against neuronal and glial antigens. Immunoreactivity applying 5 antibodies was influenced by prolonged PMD and applying 2 antibodies by prolonged FT. None of the staining outcomes related to the PMD or FT were predictable. Loss of TMA cores during processing was primarily influenced by pretreatment and by tissue characteristics (gray/white matter). The test model described here confirmed that these 2 variables, PMD and FT, indeed influence the IHC outcome. The PMD and FT are particularly of importance while assessing tissue samples obtained at autopsy. The result above is also of importance while comparing the IHC outcomes seen in the postmortem setting (various PMD/FT) with surgical samples or with IHC outcome seen in experimental animal setting (controlled PMD/FT). Thus, we suggest that the test model described here is considered when assessing the reliability of the IHC outcome when analyzing tissues with various characteristics.

  • 93.
    Lännerström, Linda
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Family Medicine and Preventive Medicine. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Centrum för klinisk forskning i Sörmland (CKFD).
    Wallman, Thorne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Family Medicine and Preventive Medicine. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Centrum för klinisk forskning i Sörmland (CKFD).
    Holmström, Inger K
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Health Services Research.
    Losing independence: the lived experience of being long-term sick-listed2013In: BMC Public Health, ISSN 1471-2458, E-ISSN 1471-2458, Vol. 13, p. 745-Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Sickness absence is a multifaceted problem. Much is known about risk factors for being long-term sick-listed, but there is still little known about the various aftermaths and experiences of it. The aim of this qualitative study was to describe, analyze and understand long-term sickness-absent people's experiences of being sick-listed.

    METHODS: The design was descriptive and had a phenomenological approach. Sixteen long-term sickness-absent individuals were purposively sampled from three municipalities in Sweden in 2011, and data were collected through semi-structured, individual interviews. The interview questions addressed how the participants experienced being sick-listed and how the sick-listing affected their lives. Transcribed interviews were analysed using Giorgi's phenomenological method.

    RESULTS: The interviews revealed that the participants' experiences of being sick-listed was that they lost their independence in the process of stepping out of working society, attending the mandatory steps in the rehabilitation chain and having numerous encounters with professionals. The participants described that their life-worlds were radically changed when they became sick-listed. Their experiences of their changing life-worlds were mostly highly negative, but there were also a few positive experiences. The most conspicuous findings were the fact that stopping working brought with it so many changes, the participants' feelings of powerlessness in the process, and their experiences of offensive treatment by and/or encounters with professionals.

    CONCLUSIONS: Sick-listed persons experienced the process of being on long-term sickness absent as very negative. The negative experiences are linked to consequences of stopping to work, consequences of social insurance rules and to negative encounters with professionals handling the sickness absence. The positive experiences of being sick-listed were few in the present study. There is a need to further examine the extent of these negative experiences are and how they affect sick-listed people's recovery and return to work. Long-term sickness absence; sick leave; experiences; interviews; phenomenology; Sweden.

  • 94.
    Malmström, David
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Pelzing, Matthias
    Bruker Bioscience Melbourne, Australia.
    Capillary Electrophoresis-Mass Spectrometry (CZE-MS) Analysis of Intact Monoclonal Antibodies using bare fused silicaManuscript (preprint) (Other academic)
  • 95.
    Mardoukhi, Sahar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Kartläggning av inkomna konsumentfrågor om läkemedel och biverkningar till Läkemedelsupplysningen på Läkemedelsverket2013Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
    Abstract [sv]

    Kartläggning av inkomna konsumentfrågor om läkemedel och biverkningar till Läkemedelsupplysningen på Läkemedelsverket

    Sahar Mardoukhi

    Handledare: Elin Kimland, Med Dr, Leg ssk, farmaceut. Examinator: Professor Margareta Hammarlund-Udenaes

    Institutionen för farmaceutisk biovetenskap. Avdelningen för farmakokinetik och läkemedelsterapi. 30 hp

     

    Introduktion: Läkemedel är ett viktigt verktyg för att behandla patienter men kan ibland ge upphov till läkemedelsbiverkningar. Under de senaste decennierna har myndigheter ansträngt sig för att kunna bidra till en säkrare läkemedelsanvändning för konsumenter genom att bl. a. etablera möjlighet till  rapportering av biverkningar för konsumenter. Ett annat initiativ är att det finns en Läkemedelsupplysningen (LMU) inom Sverige som besvarar frågor om läkemedel och läkemedelsnära produkter som riktar sig till allmänheten.

    Syfte: Denna studie syftar till att öka kunskapen om konsumenternas frågor om läkemedelsbiverkningar genom att kartlägga och analysera inkomna frågor till LMU.

    Material och metoder: Retrospektiv epidemiologisk kartläggning av inkomna frågor om läkemedelsbiverkningar till LMU baserad på mätning under en vecka per kvartal januari till juni 2013. En statistisk analys med χ2-test (P-värde <0,05) utfördes för att studera eventuella skillnader mellan de två utvalda kvartalsperioder.

    Resultat: Totalt dokumenterades 968 frågor i undersökningen varav 513 frågor från första kvartalsperioden och 455 frågor från andra kvartalsperioden. Drygt en tredje del (38%) av frågorna berörde ATC-kod N (läkemedel för behandling av sjukdomar, symtom från nervsystemet). Omkring 5 % av frågor gällande biverkningar som inte var tidigare dokumenterade i produktinformationen. Ingen signifikant skillnad observerades för de olika kategorier mellan de två kvartalsperioderna dock observerades en signifikant skillnad för antalet biverkningsfrågor mellan båda kvartalsperioderna.

    Konklusion: Kartläggning av inkomna konsumentfrågor om läkemedelsbiverkningar kan utgöra en källa för ökad kunskap utifrån ett konsumentperspektiv. Majoriteten av frågorna handlade om läkemedlen som ofta förskrivs och är väldokumenterade. Andelen frågor om ej tidigare listade biverkningar var låg.

  • 96.
    Mortensen, Anja C
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Spiegelberg, Diana
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences.
    Haylock, Anna-Karin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences.
    Lundqvist, Hans
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Nestor, Marika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Preclinical evaluation of a novel engineered recombinant humananti-CD44v6 antibody for potential use in radio-immunotherapy2018In: International Journal of Oncology, ISSN 1019-6439Article in journal (Refereed)
    Abstract [en]

    CD44v6 is overexpressed in a variety of cancers,rendering it a promising target for radio-immunotherapy (RIT).In this study, we have characterized a novel engineered recombinantmonoclonal anti-CD44v6 antibody, AbN44v6, andassessed its potential for use in RIT using either 177Lu or 131Ias therapeutic radionuclides. In vitro affinity and specificityassays characterized the binding of the antibody labeled with177Lu, 125I or 131I. The therapeutic effects of 177Lu-AbN44v6 and131I-AbN44v6 were investigated using two in vitro 3D tumormodels with different CD44v6 expression. Finally, the normaltissue biodistribution and dosimetry for 177Lu-AbN44v6 and125I-AbN44v6/131I-AbN44v6 were assessed in vivo using amouse model. All AbN44v6 radioconjugates demonstratedCD44v6-specific binding in vitro. In the in vitro 3D tumormodels, dose-dependent therapeutic effects were observedwith both 177Lu-AbN44v6 and 131I-AbN44v6, with a greatersignificant therapeutic effect observed on the cells with a higherCD44v6 expression. Biodistribution experiments demonstrateda greater uptake of 177Lu-AbN44v6 in the liver, spleen and bone,compared to 125I-AbN44v6, whereas 125I-AbN44v6 demonstrateda longer circulation time. In dosimetric calculations, thecritical organs for 177Lu-AbN44v6 were the liver and spleen,whereas the kidneys and red marrow were considered the criticalorgans for 131I-AbN44v6. The effective dose was in the order of0.1 mSv/MBq for both labels. In conclusion, AbN44v6 boundspecifically and with high affinity to CD44v6. Furthermore,in vitro RIT demonstrated growth inhibition in a CD44v6-specific activity-dependent manner for both radioconjugates,demonstrating that both 177Lu-AbN44v6 and 131I-AbN44v6 maybe promising RIT candidates. Furthermore, biodistributionand dosimetric analysis supported the applicability of bothconjugates for RIT. The CD44v6-specific therapeutic effectsobserved with radiolabeled AbN44v6 in the 3D tumor modelsin vitro, combined with the beneficial dosimetry

  • 97.
    Naessén, Tord
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Women's and Children's Health.
    Kushnir, Mark M.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Chaika, Andrey
    Donetsk State Medical University, Ukraine.
    Nosenko, Jelena
    Donetsk State Medical University, Ukraine.
    Mogilevkina, Iryna
    Donetsk State Medical University, Ukraine.
    Rockwood, Alan L.
    ARUP Institute for Clinical and Experimental Pathology, Utah.
    Carlström, Kjell
    Karolinska Institute.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Kirilovas, Dmitrijus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Women's and Children's Health.
    Steroid profiles in ovarian follicular fluid in women with and without polycystic ovary syndrome, analyzed by liquid chromatography-tandem mass spectrometry.2010In: Fertility and Sterility, ISSN 0015-0282, E-ISSN 1556-5653, Vol. 94, no 6, p. 2228-2233Article in journal (Refereed)
    Abstract [en]

    OBJECTIVE

    To compare steroid concentrations and steroid product-to-precursor ratios in ovarian follicular fluid (FF) from women with polycystic ovary syndrome (PCOS) and from regularly menstruating women in their early follicular phase, using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Polycystic ovary syndrome involves abnormal regulation of the steroidogenic enzymes, leading to arrest of follicle development.

    DESIGN

    Case-control study.

    SETTING

    University hospital clinic.

    PATIENT(S)

    Follicular fluid from size-matched ovarian follicles (5-8 mm) in 27 nonstimulated women with PCOS and in 21 women without PCOS was sampled. Thirteen steroids were quantitated from 40 muL of FF, using LC-MS/MS.

    INTERVENTION(S)

    None.

    MAIN OUTCOME MEASURE(S)

    Concentrations of steroids in the FF and product-to-precursor ratios (enzyme activity) were compared between the groups.

    RESULT(S)

    In women with PCOS, ovarian FF contained higher concentrations of individual and total androgens, lower individual and total estrogens (E), and a lower total E-to-androgen ratio, compared with regularly menstruating women. The product-to-precursor concentration ratios indicated higher CYP17-linked and lower CYP19-linked (aromatase) enzyme activity. Receiver operating characteristic plots indicated the early CYP17 step (17-OH5P/5P) being highly important for the prevalence of PCOS (c = 0.95).

    CONCLUSION(S)

    The women with PCOS had higher ovarian CYP17-linked and lower CYP19-linked (aromatase) enzyme activity, confirming previous data. Multiple steroid assessments from minute volumes including FF from nonstimulated ovaries, using LC-MS/MS, might be useful in research, clinical endocrinology, and in IVF.

  • 98.
    Nilsson, P
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Gedda, L
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Sjostrom, A
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Carlsson, J
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Effects of dextranation on the uptake of peptides in micrometastases:studies on binding of EGF in tumor spheroids.2001In: Tumour Biol, Vol. 22, p. 229-Article in journal (Refereed)
  • 99.
    Norberg, Thomas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Organic Chemistry II.
    Synthesis of phosphodiester-linked microbial polysaccharide structures2009In: Progress in the synthesis of complex carbohydrate chains of plant and microbial polysaccharides / [ed] N. E. Nifantiev, Trivandrum: Transworld Research Network , 2009, p. 309-325Chapter in book (Other academic)
    Abstract [en]

    This chapter reviews work published until mid-2008 on the synthesis of phosphodiester-linked microbial polysaccharide structures, with emphasis on the development during the last two decades. Different techniques and chemistries that have been used to create the phosphodiester linkages are discussed, and syntheses of oligomeric (2-10 repeating units) phosphodiester-linked fragments of natural polysaccharides from different sources are presented.

  • 100. Nordberg, Agneta
    et al.
    Hartvig, Per
    Lundkvist, Hans
    Antoni, Gunnar
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry.
    Ulin, Johan
    Långström, Bengt
    Uptake and distribution of (+)-R and (-)-S N-[methyl-11C]nicotine in brains of rhesus    monkey- an attempt to study nicotine receptors in vivo.1989In: J neurol trans 1, p. 195-205Article in journal (Refereed)
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