| 1. |
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| 2. |
- Judge, Heather M, et al.
(författare)
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Glycoprotein IIb/IIIa and P2Y12 receptor antagonists yield additive inhibition of platelet aggregation, granule secretion, soluble CD40L release and procoagulant responses.
- 2005
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Ingår i: Platelets. - : Informa UK Limited. - 0953-7104 .- 1369-1635. ; 16:7
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Tidskriftsartikel (refereegranskat)abstract
- Glycoprotein IIb/IIIa (GPIIb/IIIa) antagonists, including abciximab and tirofiban, are administered concurrently with clopidogrel, a P2Y12 antagonist, and aspirin in some patients undergoing percutaneous coronary intervention. We studied the effects of, and interactions between, abciximab, tirofiban, aspirin and the P2Y12 antagonist cangrelor on platelet aggregation, alpha and dense granule secretion and procoagulant responses in vitro. Blood was obtained from healthy volunteers. Platelet aggregation, dense granule secretion, alpha granule secretion (PAI-1 and soluble CD40 ligand levels) and procoagulant responses (annexin-V and microparticle formation) were assessed using collagen and thrombin receptor activating peptide (TRAP) as agonists. All the antagonists used singularly inhibited collagen-induced responses. Combinations of abciximab or tirofiban with aspirin and/or cangrelor gave additive inhibition with the greatest effect seen when abciximab or tirofiban was combined with both aspirin and cangrelor. Cangrelor inhibited TRAP-induced responses and, again, there was additive inhibition of these parameters when abciximab or tirofiban were combined with cangrelor. The GPIIb/IIIa receptor plays an important role in amplification of platelet activation such that there are important interactions between GPIIb/IIIa antagonists and inhibitors of both P2Y12 receptor activation and, to a lesser extent, thromboxane A2 generation. These interactions are likely to have important influences on the safety and efficacy of combination anti-platelet therapies.
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| 3. |
- Judge, Heather M, et al.
(författare)
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Relationship between degree of P2Y12 receptor blockade and inhibition of P2Y12-mediated platelet function.
- 2010
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Ingår i: Thrombosis and Haemostasis. - 0340-6245 .- 2567-689X. ; 103:6
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Tidskriftsartikel (refereegranskat)abstract
- The thienopyridine P2Y12 receptor antagonists clopidogrel and prasugrel prevent arterial thrombosis and are routinely used following percutaneous coronary intervention. However, the optimal level of P2Y12 blockade to effectively inhibit platelet function is unknown. These studies utilised the active metabolite of prasugrel (R-138727) to achieve a range of P2Y12 blockade in vitro and assessed several aspects of platelet function. Blood from healthy volunteers was incubated with R-138727 (0-10 microM). P2Y12 receptor number was assessed using a 33P-2MeSADP binding assay. Platelet aggregation (PA) was measured by optical aggregometry with ADP 2-20 microM. VASP phosphorylation, annexin V binding, microparticle formation and P-selectin expression were assessed by flow cytometry. Increasing numbers of unblocked receptors were required for a sustained aggregation response with decreasing concentrations of ADP. A P2Y12 receptor blockade of 60-80% resulted in strong inhibition of final PA response, P-selectin expression, microparticle formation and vasodilator-stimulated phosphoprotein (VASP). PA induced by ADP 2 microM and P-selectin expression were particularly sensitive to low levels of receptor blockade whereas the VASP phosphorylation assay was relatively insensitive, requiring 60% receptor blockade to achieve substantial inhibition. Different assays varied in their ability to discriminate particular ranges of P2Y12 blockade and 80% or greater P2Y12 receptor blockade is required for consistently strong inhibition of several aspects of platelet function. These data guide the interpretation of results from different assays used to monitor the effects of P2Y12 receptor antagonists.
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| 4. |
- Judge, Heather M, et al.
(författare)
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The active metabolite of prasugrel effectively blocks the platelet P2Y12 receptor and inhibits procoagulant and pro-inflammatory platelet responses.
- 2008
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Ingår i: Platelets. - : Informa UK Limited. - 0953-7104 .- 1369-1635. ; 19:2
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Tidskriftsartikel (refereegranskat)abstract
- The aim of these studies was to investigate the extent of platelet P2Y(12) receptor inhibition by the thienopyridine active metabolite of prasugrel, R-138727. Blood was taken from healthy volunteers and pre-incubated with R-138727 or cangrelor (AR-C66931MX). Platelet aggregation was assessed in platelet rich plasma (PRP) and whole blood (WB). Vasodilator stimulated phosphoprotein (VASP) phosphorylation, platelet procoagulant activity (annexin V binding and microparticle formation) and calcium mobilisation were measured by flow cytometry. Platelet-leukocyte co-aggregate formation and sCD40L release, both pro-inflammatory responses of platelets, were measured by flow cytometry and ELISA, respectively. P2Y(12) receptor antagonism was determined using a radioligand binding assay ((33)P 2-MeSADP) in resting and stimulated platelets and the effects of clopidogrel administration were also assessed. R-138727 yielded concentration-dependent inhibition of platelet aggregation, VASP phosphorylation inhibition, procoagulant activity and pro-inflammatory responses. In the presence of R-138727 or cangrelor there was increased calcium reuptake following agonist stimulation. R-138727 30 micromol/L and cangrelor 1 micromol/L completely inhibited (33)P 2-MeSADP binding, compared to partial inhibition following clopidogrel administration. Platelet activation and granule secretion did not expose an additional pool of P2Y(12) receptors. Prasugrel's active metabolite effectively blocks the P2Y(12) receptor with the highest concentrations tested yielding complete inhibition of P2Y(12)-mediated amplification of several important platelet responses.
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| 5. |
- Siotia, Anjan, et al.
(författare)
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Utility of a whole blood single platelet counting assay to monitor the effects of tirofiban in patients with acute coronary syndromes scheduled for coronary intervention.
- 2006
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Ingår i: Thrombosis and Haemostasis. - 0340-6245 .- 2567-689X. ; 95:6
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Tidskriftsartikel (refereegranskat)abstract
- This study aimed to establish the utility of a whole-blood single-platelet counting (WBSPC) assay, a measure of microaggregation, in monitoring the effects of tirofiban, comparing this with optical aggregometry (OA) and the Ultegra TRAP cartridge system (UTC), measures of macroaggregation. Fifty-nine patients with acute coronary syndrome scheduled for coronary angiography +/- angioplasty were studied. WBSPC assay (ADP 0.3-100 microM, Sysmex KX21 analyzer), OA (ADP 20 microM) and UTC were performed: before starting tirofiban; 30 min, 4 and 24 h after starting tirofiban; and 1 and 2 h after stopping tirofiban. Thirty minutes after starting tirofiban, there was substantial inhibition of platelet aggregation (40 +/- 30%; WBSPC, 2 minutes after addition of ADP 30 microM) and this remained stable at 4 and 24 h. OA (86 +/- 17%; inhibition of maximal aggregation, ADP 20 microM) and UTC (93 +/- 7%) showed marked inhibition with less inter-individual variation. There was no significant correlation between OA and UTC results (R(2) = 0.006), but fair correlation between OA and WBSPC results (R(2) = 0.37). Greater inhibition of macroaggregation (OA and UTC) was seen compared to microaggregation (WBSPC) such that WBSPC was more discriminating in the therapeutic range when macroaggregation was often completely inhibited. A WBSPC assay of platelet microaggregation shows promise for monitoring GPIIb/IIIa antagonists.
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| 6. |
- Stephens, Lucas, et al.
(författare)
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Archaeological assessment reveals Earth’s early transformation through land use
- 2019
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Ingår i: Science. - : American Association for the Advancement of Science. - 0036-8075 .- 1095-9203. ; 365:6456, s. 897-902
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Tidskriftsartikel (refereegranskat)abstract
- Humans began to leave lasting impacts on Earth’s surface starting 10,000 to 8000 years ago. Through a synthetic collaboration with archaeologists around the globe, Stephens et al. compiled a comprehensive picture of the trajectory of human land use worldwide during the Holocene (see the Perspective by Roberts). Hunter-gatherers, farmers, and pastoralists transformed the face of Earth earlier and to a greater extent than has been widely appreciated, a transformation that was essentially global by 3000 years before the present.Science, this issue p. 897; see also p. 865Environmentally transformative human use of land accelerated with the emergence of agriculture, but the extent, trajectory, and implications of these early changes are not well understood. An empirical global assessment of land use from 10,000 years before the present (yr B.P.) to 1850 CE reveals a planet largely transformed by hunter-gatherers, farmers, and pastoralists by 3000 years ago, considerably earlier than the dates in the land-use reconstructions commonly used by Earth scientists. Synthesis of knowledge contributed by more than 250 archaeologists highlighted gaps in archaeological expertise and data quality, which peaked for 2000 yr B.P. and in traditionally studied and wealthier regions. Archaeological reconstruction of global land-use history illuminates the deep roots of Earth’s transformation and challenges the emerging Anthropocene paradigm that large-scale anthropogenic global environmental change is mostly a recent phenomenon.
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| 7. |
- Williams, John W., et al.
(författare)
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The neotoma paleoecology database, a multiproxy, international, community-curated data resource
- 2018
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Ingår i: Quaternary Research. - : Cambridge University Press. - 0033-5894 .- 1096-0287. ; 89:1, s. 156-177
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Tidskriftsartikel (refereegranskat)abstract
- The Neotoma Paleoecology Database is a community-curated data resource that supports interdisciplinary global change research by enabling broad-scale studies of taxon and community diversity, distributions, and dynamics during the large environmental changes of the past. By consolidating many kinds of data into a common repository, Neotoma lowers costs of paleodata management, makes paleoecological data openly available, and offers a high-quality, curated resource. Neotoma’s distributed scientific governance model is flexible and scalable, with many open pathways for participation by new members, data contributors, stewards, and research communities. The Neotoma data model supports, or can be extended to support, any kind of paleoecological or paleoenvironmental data from sedimentary archives. Data additions to Neotoma are growing and now include >3.8 million observations, >17,000 datasets, and >9200 sites. Dataset types currently include fossil pollen, vertebrates, diatoms, ostracodes, macroinvertebrates, plant macrofossils, insects, testate amoebae, geochronological data, and the recently added organic biomarkers, stable isotopes, and specimen-level data. Multiple avenues exist to obtain Neotoma data, including the Explorer map-based interface, an application programming interface, the neotoma R package, and digital object identifiers. As the volume and variety of scientific data grow, community-curated data resources such as Neotoma have become foundational infrastructure for big data science.
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| 8. |
- Buckland, Robert, 1976-
(författare)
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DNA precursor asymmetries, Mismatch Repair and their effect on mutation specificity
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- In order to build any structure, a good supply of materials, accurate workers and quality control are needed. This is even the case when constructing DNA, the so-called “Code of Life.” For a species to continue to exist, this DNA code must be copied with incredibly high accuracy when each and every cell replicates. In fact, just one mistake in the 12 million bases that comprise the genome of budding yeast, Saccharomyces cerevisiae, can be fatal. DNA is composed of a double strand helix made up of just four different bases repeated millions of times. The building blocks of DNA are the deoxyribonucleotides (dNTPs); dCTP, dTTP, dATP and dGTP. Their production and balance are carefully controlled within each cell, largely by the key enzyme Ribonucleotide Reductase (RNR). Here, we studied how the enzymes that copy DNA, the replicative polymerases α, δ and ε, cope with the effects of an altered dNTP pool balance. An introduced mutation in the allosteric specificity site of RNR in a strain of S. cerevisiae, rnr1-Y285A, leads to elevated dCTP and dTTP levels and has been shown to have a 14-fold increase in mutation rate compared to wild type. To ascertain the full effects of the dNTP pool imbalance upon the replicative polymerases, we disabled one of the major quality control systems in a cell that corrects replication errors, the post-replicative Mismatch Repair system. Using both the CAN1 reporter assay and whole genome sequencing, we found that, despite inherent differences between the polymerases, their replication fidelity was affected very similarly by this dNTP pool imbalance. Hence, the high dCTP and dTTP forced Pol ε and Pol α/δ to make the same mistakes. In addition, the mismatch repair machinery was found to correct replication errors driven by this dNTP pool imbalance with highly variable efficiencies. Another mechanism to protect cells from DNA damage during replication is a checkpoint that can be activated to delay the cell cycle and activate repair mechanisms. In yeast, Mec1 and Rad53 (human ATR and Chk1/Chk2) are two key S-phase checkpoint proteins. They are essential as they are also required for normal DNA replication and dNTP pool regulation. However the reason why they are essential is not well understood. We investigated this by mutating RAD53 and analyzing dNTP pools and gene interactions. We show that Rad53 is essential in S-phase due to its role in regulating basal dNTP levels by action in the Dun1 pathway that regulates RNR and Rad53’s compensatory kinase function if dNTP levels are perturbed.In conclusion we present further evidence of the importance of dNTP pools in the maintenance of genome integrity and shed more light on the complex regulation of dNTP levels.
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| 9. |
- Buckland, Robert J, et al.
(författare)
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Increased and Imbalanced dNTP Pools Symmetrically Promote Both Leading and Lagging Strand Replication Infidelity
- 2014
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Ingår i: PLOS Genetics. - : Public Library of Science (PLoS). - 1553-7390 .- 1553-7404. ; 10:12
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Tidskriftsartikel (refereegranskat)abstract
- The fidelity of DNA replication requires an appropriate balance of dNTPs, yet the nascent leading and lagging strands of the nuclear genome are primarily synthesized by replicases that differ in subunit composition, protein partnerships and biochemical properties, including fidelity. These facts pose the question of whether imbalanced dNTP pools differentially influence leading and lagging strand replication fidelity. Here we test this possibility by examining strand-specific replication infidelity driven by a mutation in yeast ribonucleotide reductase, rnr1-Y285A, that leads to elevated dTTP and dCTP concentrations. The results for the CAN1 mutational reporter gene present in opposite orientations in the genome reveal that the rates, and surprisingly even the sequence contexts, of replication errors are remarkably similar for leading and lagging strand synthesis. Moreover, while many mismatches driven by the dNTP pool imbalance are efficiently corrected by mismatch repair, others are repaired less efficiently, especially those in sequence contexts suggesting reduced proofreading due to increased mismatch extension driven by the high dTTP and dCTP concentrations. Thus the two DNA strands of the nuclear genome are at similar risk of mutations resulting from this dNTP pool imbalance, and this risk is not completely suppressed even when both major replication error correction mechanisms are genetically intact.
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| 10. |
- Geoghegan, Fintan, et al.
(författare)
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Bioenergetics of acquired cisplatin resistant H1299 non-small cell lung cancer and P31 mesothelioma cells
- 2017
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Ingår i: Oncotarget. - : IMPACT JOURNALS LLC. - 1949-2553. ; 8:55, s. 94711-94725
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Tidskriftsartikel (refereegranskat)abstract
- Acquired cisplatin resistance is a common feature of tumours following cancer treatment with cisplatin and also of non-small cell lung cancer (H1299) and mesothelioma (P31) cell lines exposed to cisplatin. To elucidate the cellular basis of acquired cisplatin resistance, a comprehensive bioenergetic analysis was undertaken. We demonstrate that cellular oxygen consumption was significantly decreased in cisplatin resistant cells and that the reduction was primarily due to reduced mitochondrial activity as a result of reduced mitochondrial abundance. The differential mitochondrial abundance was supported by data showing reduced sirtuin 1 (SIRT1), peroxisome-proliferator activator receptor-gamma co-activator 1-alpha (PGC1 alpha), sirtuin 3 (SIRT3) and mitochondrial transcription factor A (TFAM) protein expression in resistant cells. Consistent with these data we observed increased reactive oxygen species (ROS) production and increased hypoxia inducible factor 1-alpha (HIF1 alpha) stabilization in cisplatin resistant cells when compared to cisplatin sensitive controls. We also observed an increase in AMP kinase subunit alpha 2 (AMPK alpha 2) transcripts and protein expression in resistant H1299 cells. mRNA expression was also reduced for cisplatin resistant H1299 cells in these genes, however the pattern was not consistent in resistant P31 cells. There was very little change in DNA methylation of these genes, suggesting that the cells are not stably reprogrammed epigenetically. Taken together, our data demonstrate reduced oxidative metabolism, reduced mitochondrial abundance, potential for increased glycolytic flux and increased ROS production in acquired cisplatin resistant cells. This suggests that the metabolic changes are a result of reduced SIRT3 expression and increased HIF-1 alpha stabilization.
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| 11. |
- Hoch, Nicolas C, et al.
(författare)
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Molecular basis of the essential s phase function of the rad53 checkpoint kinase
- 2013
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Ingår i: Molecular and Cellular Biology. - 0270-7306 .- 1098-5549. ; 33:16, s. 3202-3213
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Tidskriftsartikel (refereegranskat)abstract
- The essential yeast kinases Mec1 and Rad53, or human ATR and Chk1, are crucial for checkpoint responses to exogenous genotoxic agents, but why they are also required for DNA replication in unperturbed cells remains poorly understood. Here we report that even in the absence of DNA-damaging agents, the rad53-4AQ mutant, lacking the N-terminal Mec1 phosphorylation site cluster, is synthetic lethal with a deletion of the RAD9 DNA damage checkpoint adaptor. This phenotype is caused by an inability of rad53-4AQ to activate the downstream kinase Dun1, which then leads to reduced basal deoxynucleoside triphosphate (dNTP) levels, spontaneous replication fork stalling, and constitutive activation of and dependence on S phase DNA damage checkpoints. Surprisingly, the kinase-deficient rad53-K227A mutant does not share these phenotypes but is rendered inviable by additional phosphosite mutations that prevent its binding to Dun1. The results demonstrate that ultralow Rad53 catalytic activity is sufficient for normal replication of undamaged chromosomes as long as it is targeted toward activation of the effector kinase Dun1. Our findings indicate that the essential S phase function of Rad53 is comprised by the combination of its role in regulating basal dNTP levels and its compensatory kinase function if dNTP levels are perturbed.
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| 12. |
- Huang, Junchi, et al.
(författare)
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Osteoclasts directly influence castration-resistant prostate cancer cells
- 2022
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Ingår i: Clinical and Experimental Metastasis. - : Springer Nature. - 0262-0898 .- 1573-7276. ; 39:5, s. 801-814
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Tidskriftsartikel (refereegranskat)abstract
- Metastasis to bone is the leading cause of death from prostate cancer. Interaction between tumor cells and bone cells can promote progression and influence tumor phenotype. It is known that prostate cancer cells support osteoclast differentiation, and degradation of bone matrix by osteoclasts releases growth factors stimulating tumor cell proliferation and invasion. In the present study osteolytic (PC-3) and osteoblastic (LNCaP-19) castration-resistant prostate cancer (CRPC) cells were co-cultured with mature osteoclasts or their precursor cells (RAW 264.7) to characterize direct effects of mature osteoclasts on CRPC cells. Osteoclasts increased proliferation and decrease apoptosis of CRPC cells as assessed with flow cytometry. RNA sequencing revealed that osteolytic CRPC cells were more responsive to osteoclast stimulation regarding gene expression, but the overall induced expression patterns were similar between the prostate cancer cell lines. Genes related to DNA repair were upregulated by osteoclasts, while genes related to endoplasmic reticulum stress-induced apoptosis and cholesterol synthesis were downregulated. The results of this study shows that osteoclasts directly influence CRPC cells, increasing proliferation, decreasing apoptosis, and affecting gene expression pathways that can affect sensitivity to DNA damage and endoplasmic reticulum function. This suggests targeting of osteoclasts to be a possible way to affect efficacy of other drugs by combination regimens in treating prostate cancer metastases.
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| 13. |
- Jia, Shaodong, et al.
(författare)
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Determination of deoxyribonucleoside triphosphate concentrations in yeast cells by strong anion-exchange high-performance liquid chromatography coupled with ultraviolet detection
- 2015
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Ingår i: DNA Replikation. - New York : Springer-Verlag New York. - 9781493925957 - 9781493925964 ; , s. 113-121
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Bokkapitel (refereegranskat)abstract
- DNA polymerase assays are commonly used for the detection of deoxyribonucleoside triphosphates (dNTPs) in biological samples. For better specificity and accuracy, high-performance liquid chromatography (HPLC) methods have been developed for the analysis of the four dNTPs in complex samples. Here we describe a simple method using isocratic strong anion-exchange (SAX) chromatographic separation coupled with ultraviolet detection (UV) for the analysis of the four dNTPs in budding yeast Saccharomyces cerevisiae. This method can be applied to other species of yeast or bacteria.
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| 14. |
- Johnson, Paul C D, et al.
(författare)
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Abundant variation in microsatellites of the parasitic nematode Trichostrongylus tenuis and linkage to a tandem repeat.
- 2006
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 148:2
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Tidskriftsartikel (refereegranskat)abstract
- An understanding of how genes move between and within populations of parasitic nematodes is important in combating the evolution and spread of anthelmintic resistance. Much has been learned by studying mitochondrial DNA markers, but autosomal markers such as microsatellites have been applied to only a few nematode species, despite their many advantages for studying gene flow in eukaryotes. Here, we describe the isolation of 307 microsatellites from Trichostrongylus tenuis, an intestinal nematode of red grouse. High levels of variation were revealed at sixteen microsatellite loci (including three sex-lined loci) in 111 male T. tenuis nematodes collected from four hosts at a single grouse estate in Scotland (average He = 0.708; mean number of alleles = 12.2). A population genetic analysis detected no deviation from panmixia either between (F(ST) = 0.00) or within hosts (F(IS) = 0.015). We discuss the feasibility of developing microsatellites in parasitic nematodes and the problem of null alleles. We also describe a novel 146-bp repeat element, TteREP1, which is linked to two-thirds of the microsatellites sequenced and is associated with marker development failure. The sequence of TteREP1 is related to the TcREP-class of repeats found in several other trichostrongyloid species including Trichostrongylus colubriformis and Haemonchus contortus.
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| 15. |
- Rentoft, Matilda, et al.
(författare)
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Heterozygous colon cancer-associated mutations of SAMHD1 have functional significance
- 2016
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 113:17, s. 4723-4728
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Tidskriftsartikel (refereegranskat)abstract
- Even small variations in dNTP concentrations decrease DNA replication fidelity, and this observation prompted us to analyze genomic cancer data for mutations in enzymes involved in dNTP metabolism. We found that sterile alpha motif and histidine-aspartate domain-containing protein 1 (SAMHD1), a deoxyribonucleoside triphosphate triphosphohydrolase that decreases dNTP pools, is frequently mutated in colon cancers, that these mutations negatively affect SAMHD1 activity, and that severalSAMHD1mutations are found in tumors with defective mismatch repair. We show that minor changes in dNTP pools in combination with inactivated mismatch repair dramatically increase mutation rates. Determination of dNTP pools in mouse embryos revealed that inactivation of oneSAMHD1allele is sufficient to elevate dNTP pools. These observations suggest that heterozygous cancer-associatedSAMHD1mutations increase mutation rates in cancer cells.
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| 16. |
- Valencia, Liliana, et al.
(författare)
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Identification of novel genetic variants in the mutational hotspot region 14kb upstream of the LCT gene in a Mexican population
- 2017
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Ingår i: Scandinavian Journal of Clinical and Laboratory Investigation. - : Taylor & Francis. - 0036-5513 .- 1502-7686. ; 77:5, s. 311-314
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Tidskriftsartikel (refereegranskat)abstract
- Several polymorphic loci linked to lactase persistence (LP) have been described, all located in a small mutational hotspot region far upstream (approximate to 14kb) of the lactase (LCT) gene. One is typically found in Europeans, LCT -13910C>T, several others are found in East Africans and Arabs, e.g. LCT -13907C>G and LCT -13915T>G. The possibility of similar loci, specific to populations in South and Central America, has not received much attention so far. To identify possible novel polymorphisms in the mutational hotspot region, we sampled 158 subjects from a rural area in South-Central Mexico. DNA was isolated from serum, and Sanger sequencing of a 501bp region spanning the LCT -13910C>T hotspot was successfully performed in 150 samples. The frequency of the European-type LCT -13910T-allele was q=0.202, and 35% of the population was thus lactase-persistent (CT or TT). Sixteen novel genetic variants were found amongst 11 of the subjects, all were heterozygotes: seven of the subjects were also carriers of at least one LCT -13910T-allele. Thus, the mutational hotspot region is also a hotspot in the rural Mexican population: 11/150 subjects carried a total of 16 previously unknown private mutations but no novel polymorphism was found. The relationship between such novel genetic variants in Mexicans and lactase persistence is worthy of more investigation.
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| 17. |
- Watt, Danielle L., et al.
(författare)
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Genome-wide analysis of the specificity and mechanisms of replication infidelity driven by imbalanced dNTP pools
- 2016
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Ingår i: Nucleic Acids Research. - : Oxford University Press (OUP). - 0305-1048 .- 1362-4962. ; 44:4, s. 1669-1680
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- The absolute and relative concentrations of the four dNTPs are key determinants of DNA replication fidelity, yet the consequences of altered dNTP pools on replication fidelity have not previously been investigated on a genome-wide scale. Here, we use deep sequencing to determine the types, rates and locations of uncorrected replication errors that accumulate in the nuclear genome of a mismatch repair-deficient diploid yeast strain with elevated dCTP and dTTP concentrations. These imbalanced dNTP pools promote replication errors in specific DNA sequence motifs suggesting increased misinsertion and increased mismatch extension at the expense of proofreading. Interestingly, substitution rates are similar for leading and lagging strand replication, but are higher in regions replicated late in S phase. Remarkably, the rate of single base deletions is preferentially increased in coding sequences and in short rather than long mononucleotides runs. Based on DNA sequence motifs, we propose two distinct mechanisms for generating single base deletions in vivo. Collectively, the results indicate that elevated dCTP and dTTP pools increase mismatch formation and decrease error correction across the nuclear genome, and most strongly increases mutation rates in coding and late replicating sequences.
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| 18. |
- Welén, Karin, 1970, et al.
(författare)
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A Phase 2 Trial of the Effect of Antiandrogen Therapy on COVID-19 Outcome : No Evidence of Benefit, Supported by Epidemiology and In Vitro Data
- 2022
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Ingår i: European Urology. - : Elsevier. - 0302-2838 .- 1873-7560. ; 81:3, s. 285-293
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Tidskriftsartikel (refereegranskat)abstract
- Background: Men are more severely affected by COVID-19. Testosterone may influence SARS-CoV-2 infection and the immune response.Objective: To clinically, epidemiologically, and experimentally evaluate the effect of antiandrogens on SARS-CoV-2 infection.Designs, settings, and participants: A randomized phase 2 clinical trial (COVIDENZA) enrolled 42 hospitalized COVID-19 patients before safety evaluation. We also conducted a population-based retrospective study of 7894 SARS-CoV-2–positive prostate cancer patients and an experimental study using an air-liquid interface three-dimensional culture model of primary lung cells.Intervention: In COVIDENZA, patients were randomized 2:1 to 5 d of enzalutamide or standard of care.Outcome measurements: The primary outcomes in COVIDENZA were the time to mechanical ventilation or discharge from hospital. The population-based study investigated risk of hospitalization, intensive care, and death from COVID-19 after androgen inhibition.Results and limitations: Enzalutamide-treated patients required longer hospitalization (hazard ratio [HR] for discharge from hospital 0.43, 95% confidence interval [CI] 0.20–0.93) and the trial was terminated early. In the epidemiological study, no preventive effects were observed. The frail population of patients treated with androgen deprivation therapy (ADT) in combination with abiraterone acetate or enzalutamide had a higher risk of dying from COVID-19 (HR 2.51, 95% CI 1.52–4.16). In vitro data showed no effect of enzalutamide on virus replication. The epidemiological study has limitations that include residual confounders.Conclusions: The results do not support a therapeutic effect of enzalutamide or preventive effects of bicalutamide or ADT in COVID-19. Thus, these antiandrogens should not be used for hospitalized COVID-19 patients or as prevention for COVID-19. Further research on these therapeutics in this setting are not warranted.Patient summary: We studied whether inhibition of testosterone could diminish COVID-19 symptoms. We found no evidence of an effect in a clinical study or in epidemiological or experimental investigations. We conclude that androgen inhibition should not be used for prevention or treatment of COVID-19.
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| 19. |
- Welen, Karin, et al.
(författare)
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COVIDENZA - A prospective, multicenter, randomized PHASE II clinical trial of enzalutamide treatment to decrease the morbidity in patients with Corona virus disease 2019 (COVID-19): a structured summary of a study protocol for a randomised controlled trial.
- 2021
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Ingår i: Trials. - : Springer Science and Business Media LLC. - 1745-6215. ; 22:1
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Tidskriftsartikel (refereegranskat)abstract
- The main goal of the COVIDENZA trial is to evaluate if inhibition of testosterone signalling by enzalutamide can improve the outcome of patients hospitalised for COVID-19. The hypothesis is based on the observation that the majority of patients in need of intensive care are male, and the connection between androgen receptor signalling and expression of TMPRSS2, an enzyme important for SARS-CoV-2 host cell internalization.Hospitalised COVID-19 patients will be randomised (2:1) to enzalutamide plus standard of care vs. standard of care designed to identify superiority.Included participants, men or women above 50 years of age, must be hospitalised for PCR confirmed COVID-19 symptoms and not in need of immediate mechanical ventilation. Major exclusion criteria are breast-feeding or pregnant women, hormonal treatment for prostate or breast cancer, treatment with immunosuppressive drugs, current symptomatic unstable cardiovascular disease (see Additional file 1 for further details). The trial is registered at Umeå University Hospital, Region Västerbotten, Sweden and 8 hospitals are approved for inclusion in Sweden.Patients randomised to the treatment arm will be treated orally with 160 mg (4x40 mg) enzalutamide (Xtandi®) daily, for five consecutive days. The study is not placebo controlled. The comparator is standard of care treatment for patients hospitalised with COVID-19.The primary endpoints of the study are (time to) need of mechanical ventilation or discharge from hospital as assessed by a clinical 7-point ordinal scale (up to 30 days after inclusion).Randomisation was stratified by center and sex. Each strata was randomized separately with block size six with a 2:1 allocation ratio (enzalutamide + "standard of care": "standard of care"). The randomisation list, with consecutive subject numbers, was generated by an independent statistician using the PROC PLAN procedure of SAS version 9.4 software (SAS Institute, Inc, Cary, North Carolina) BLINDING (MASKING): This is an open-label trial.The trial is designed to have three phases. The first, an exploration phase of 45 participants (30 treatment and 15 control) will focus on safety and includes a more extensive laboratory assessment as well as more frequent safety evaluation. The second prolongation phase, includes the first 100 participants followed by an interim analysis to define the power of the study. The third phase is the continuation of the study up to maximum 600 participants included in total.The current protocol version is COVIDENZA v2.0 as of September 10, 2020. Recruitment started July 29, 2020 and is presently in safety pause after the first exploration phase. Recruitment is anticipated to be complete by 31 December 2021.Eudract number 2020-002027-10 ClinicalTrials.gov Identifier: NCT04475601 , registered June 8, 2020 FULL PROTOCOL: The full protocol is attached as an additional file, accessible from the Trials website (Additional file 1). In the interest in expediting dissemination of this material, the familiar formatting has been eliminated; this Letter serves as a summary of the key elements of the full protocol.
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