Publications by authors named "J Christian Lukas"

474 Publications

Homology-directed repair protects the replicating genome from metabolic assaults.

Dev Cell 2021 Feb;56(4):461-477.e7

Protein Signaling Program, Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, Blegdamsvej 3b, DK-2200 Copenhagen, Denmark. Electronic address:

Homology-directed repair (HDR) safeguards DNA integrity under various forms of stress, but how HDR protects replicating genomes under extensive metabolic alterations remains unclear. Here, we report that besides stalling replication forks, inhibition of ribonucleotide reductase (RNR) triggers metabolic imbalance manifested by the accumulation of increased reactive oxygen species (ROS) in cell nuclei. This leads to a redox-sensitive activation of the ATM kinase followed by phosphorylation of the MRE11 nuclease, which in HDR-deficient settings degrades stalled replication forks. Intriguingly, nascent DNA degradation by the ROS-ATM-MRE11 cascade is also triggered by hypoxia, which elevates signaling-competent ROS and attenuates functional HDR without arresting replication forks. Under these conditions, MRE11 degrades daughter-strand DNA gaps, which accumulate behind active replisomes and attract error-prone DNA polymerases to escalate mutation rates. Thus, HDR safeguards replicating genomes against metabolic assaults by restraining mutagenic repair at aberrantly processed nascent DNA. These findings have implications for cancer evolution and tumor therapy.
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http://dx.doi.org/10.1016/j.devcel.2021.01.011DOI Listing
February 2021

Subgroup analysis of nelipepimut-S plus GM-CSF combined with trastuzumab versus trastuzumab alone to prevent recurrences in patients with high-risk, HER2 low-expressing breast cancer.

Clin Immunol 2021 Jan 22;225:108679. Epub 2021 Jan 22.

Cancer Vaccine Development Program, 1422 E. Grayson, 3rd Floor, San Antonio, TX 78208, USA.

HER2-targeted therapy has not benefited patients with low levels of HER2 expression; however, combination therapy may be effective. Primary analysis of a phase IIb trial investigating the HER2-derived vaccine nelipepimut-S (NPS) did not benefit the intention-to-treat population, but subset analysis showed a benefit in triple-negative breast cancer (TNBC) patients. The subset analysis of this multicenter, randomized, single-blind, phase IIb trial identified significant improvement in 36-month disease-free survival (DFS) between NPS (n = 55) and placebo (n = 44) in TNBC (HR 0.25, p = 0.01) and those who express HLA-A24 (HR 0.41, p = 0.05). The TNBC cohort demonstrated improved 36-month DFS in those with HER2 1+ expression (HR 0.17, p = 0.01), HLA-A24 positivity (HR 0.08, p < 0.01), or in those who received neoadjuvant chemotherapy (HR 0.21, p < 0.01). NPS vaccination with trastuzumab was associated with improved 36-month DFS among patients with TNBC. The observed benefit to this high-risk subgroup warrants confirmation in a phase III trial.
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http://dx.doi.org/10.1016/j.clim.2021.108679DOI Listing
January 2021

Challenges of Combining Opioids and P2Y Inhibitors in Acute Coronary Syndrome: Should the Future Be Opioid Free?

Curr Probl Cardiol 2021 Apr 26;46(4):100781. Epub 2020 Dec 26.

Division of Cardiovascular Diseases, Department of Medicine, University of Tennessee Health Sciences Center, Methodist Le Bonheur Healthcare System, Memphis, TN.

Morphine has been long recognized as standard of care in the treatment of acute coronary syndrome (ACS) patients; however, its safety has recently been called into question due to a drug interaction with P2Y inhibitors. Opioids, given in combination with P2Y inhibitors, can reduce antiplatelet effects by slowing gastrointestinal motility and ultimately reducing drug absorption. While there are proposed benefits of opioids in ACS patients, conflicting data regarding clinical outcomes exist. The majority of clinical data slightly favors opioid use in ST-elevation myocardial infarction over non-ST-elevation myocardial infarction, although trends for increased myocardial infarction are present in both settings. Current practice should be aimed at discerning the need for routine opioid use in ACS. Alternative strategies may be needed to overcome these interactions; however, no robust data are currently available to support these treatment options. Future research should be aimed at non-opioid treatment options in ACS, as opioid use remains controversial in this population.
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http://dx.doi.org/10.1016/j.cpcardiol.2020.100781DOI Listing
April 2021

Generation of an iPSC line (AKOSi004-A) from fibroblasts of a female adult NPC1 patient, carrying the compound heterozygous mutation p.Val1023Serfs*15/p.Gly992Arg and of an iPSC line (AKOSi005-A) from a female adult control individual.

Stem Cell Res 2020 Dec 15;50:102127. Epub 2020 Dec 15.

Translational Neurodegeneration Section "Albrecht-Kossel", Department of Neurology, University Medical Center Rostock, 18147 Rostock, Germany; Center for Transdisciplinary Neurosciences Rostock (CTNR), University Medical Center Rostock, 18147 Rostock, Germany. Electronic address:

Niemann-Pick disease Type C (NPC) is a rare progressive neurodegenerative disorder with an incidence of 1:120,000 caused by mutations in the NPC1 or NPC2 gene leading to a massive cholesterol accumulation. Here, we describe the generation of induced pluripotent stem cells (iPSCs) of an affected female adult individual carrying the NPC1 mutation p.Val1023Serfs*15/p.Gly992Arg and an iPSC line from an unrelated healthy female adult control individual. Human iPSCs were derived from fibroblasts using retroviruses carrying the four reprogramming factors OCT4, SOX2, KLF4 and C-MYC. These lines provide a valuable resource for studying the pathophysiology of NPC and for pharmacological intervention.
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http://dx.doi.org/10.1016/j.scr.2020.102127DOI Listing
December 2020

First Line Systemic Treatment for MALT Lymphoma-Do We Still Need Chemotherapy? Real World Data from the Medical University Vienna.

Cancers (Basel) 2020 Nov 26;12(12). Epub 2020 Nov 26.

Department of Medicine I, Division of Oncology, Medical University of Vienna, A-1090 Vienna, Austria.

There is no clear therapeutic algorithm for mucosa-associated lymphoid tissue (MALT) lymphoma beyond Helicobacter pylori eradication and while chemotherapy-based regimens are standard for MALT lymphoma patients in need of systemic treatment, it appears of interest to also investigate chemotherapy-free strategies. We have retrospectively assessed MALT lymphoma patients undergoing upfront systemic treatment, classified either as chemotherapy (=classical cytostatic agents +/- rituximab) or immunotherapy (=immunomodulatory agents or single anti-CD20 antibodies) at the Medical University Vienna 1999-2019. The primary endpoint was progression-free survival (PFS). In total, 159 patients were identified with a median follow-up of 67 months. The majority of patients had extragastric disease (80%), but we also identified 32 patients (20%) with Helicobacter pylori negative or disseminated gastric lymphoma. Regarding the type of first line treatment and outcome, 46% (74/159) received a chemotherapy-based regimen and 54% (85/159) immunotherapy including IMiDs lenalidomide/thalidomide (37%), anti-CD20-anitbodies rituximab/ofatumumab (27%), macrolides clarithromycin/azithromycin (27%) and proteasome inhibitor bortezomib (9%). Median PFS was 76 months (95%CI 50-102), and while the overall response (90% vs. 68%, 0.01) and the complete remission rate (75% vs. 43%, 0.01) was significantly higher for chemotherapy, there was no difference in PFS between chemotherapy (median 81 months, 95%CI 47-116) and immunotherapy (76 months, 95%CI 50-103, = 0.57), suggesting comparable long-term outcomes. To conclude, our data show higher response rates with chemo- compared to immunotherapy, but this did not translate into a superior PFS. Given the biological background of MALT lymphoma, and the favorable toxicity profile of novel immunomodulatory treatments, this should be further investigated.
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http://dx.doi.org/10.3390/cancers12123533DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7761357PMC
November 2020

Generation of two induced pluripotent stem cell lines from a female adult homozygous for the Wilson disease associated ATP7B variant p.H1069Q (AKOSi008-A) and a healthy control (AKOSi009-A).

Stem Cell Res 2020 12 5;49:102079. Epub 2020 Nov 5.

Translational Neurodegeneration Section Albrecht-Kossel, Department of Neurology, University Medical Center Rostock, 18147 Rostock, Germany; Center for Transdisciplinary Neurosciences Rostock (CTNR), University Medical Center Rostock, University of Rostock, 18147 Rostock, Germany.

Wilson disease (WD) is a rare, monogenic disorder caused by mutations in the gene ATP7B. A loss of function of the expressed protein leads to excessive hepatic and cerebral copper storage. In this study, we present the generation of two induced pluripotent stem cell (iPSC) lines derived from fibroblasts of a clinically asymptomatic, chelator treated female WD patient carrying the common missense mutation p.H1069Q and an age-matched female healthy control subject. The generated iPSC lines expressed pluripotency markers, showed differentiation potential and retained their parental genotype. Therefore, these cells provide a valuable resource to understand the pathophysiology of WD and can be used as model systems for drug testing.
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http://dx.doi.org/10.1016/j.scr.2020.102079DOI Listing
December 2020

Generation of an iPSC line (AKOSi006-A) from fibroblasts of an NPC1 patient, carrying the homozygous mutation p.I1061T (c.3182 T > C) and a control iPSC line (AKOSi007-A) using a non-integrating Sendai virus system.

Stem Cell Res 2020 12 16;49:102056. Epub 2020 Oct 16.

Translational Neurodegeneration Section, Albrecht-Kossel, Department of Neurology, University Medical Center Rostock, 18147 Rostock, Germany; Center for Transdisciplinary Neurosciences Rostock (CTNR), University Medical Center Rostock, 18147 Rostock, Germany. Electronic address:

Niemann-Pick disease type C1 (NPC1) is a rare inherited lipid storage disorder caused by mutations in the NPC1 gene. Mutations lead to impaired lipid trafficking and subsequently to accumulation of cholesterol and sphingolipids. NPC1-patients present variable multisystemic symptoms, including neurological deficits. Here, we describe the generation of human iPSC lines obtained from fibroblasts of a male individual, carrying the homozygous mutation p.I1061T, and an unrelated and healthy male individual. A non-integrating Sendai virus system, containing KLF4, OCT3/4, SOX2 and C-MYC, was used for reprogramming. These cell lines provide a valuable resource for studying the pathophysiology of multisystemic NPC1-disease.
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http://dx.doi.org/10.1016/j.scr.2020.102056DOI Listing
December 2020

Equilibrium between nascent and parental MCM proteins protects replicating genomes.

Nature 2020 11 21;587(7833):297-302. Epub 2020 Oct 21.

Protein Signaling Program, Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.

Minichromosome maintenance proteins (MCMs) are DNA-dependent ATPases that bind to replication origins and license them to support a single round of DNA replication. A large excess of MCM2-7 assembles on chromatin in G1 phase as pre-replication complexes (pre-RCs), of which only a fraction become the productive CDC45-MCM-GINS (CMG) helicases that are required for genome duplication. It remains unclear why cells generate this surplus of MCMs, how they manage to sustain it across multiple generations, and why even a mild reduction in the MCM pool compromises the integrity of replicating genomes. Here we show that, for daughter cells to sustain error-free DNA replication, their mother cells build up a nuclear pool of MCMs both by recycling chromatin-bound (parental) MCMs and by synthesizing new (nascent) MCMs. Although all MCMs can form pre-RCs, it is the parental pool that is inherently stable and preferentially matures into CMGs. By contrast, nascent MCM3-7 (but not MCM2) undergo rapid proteolysis in the cytoplasm, and their stabilization and nuclear translocation require interaction with minichromosome-maintenance complex-binding protein (MCMBP), a distant MCM paralogue. By chaperoning nascent MCMs, MCMBP safeguards replicating genomes by increasing chromatin coverage with pre-RCs that do not participate on replication origins but adjust the pace of replisome movement to minimize errors during DNA replication. Consequently, although the paucity of pre-RCs in MCMBP-deficient cells does not alter DNA synthesis overall, it increases the speed and asymmetry of individual replisomes, which leads to DNA damage. The surplus of MCMs therefore increases the robustness of genome duplication by restraining the speed at which eukaryotic cells replicate their DNA. Alterations in physiological fork speed might thus explain why even a minor reduction in MCM levels destabilizes the genome and predisposes to increased incidence of tumour formation.
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http://dx.doi.org/10.1038/s41586-020-2842-3DOI Listing
November 2020

Oxidative Stress and Alterations in the Antioxidative Defense System in Neuronal Cells Derived from NPC1 Patient-Specific Induced Pluripotent Stem Cells.

Int J Mol Sci 2020 Oct 16;21(20). Epub 2020 Oct 16.

Translational Neurodegeneration Section Albrecht Kossel, Department of Neurology, University Medical Center Rostock, University of Rostock, 18147 Rostock, Germany.

Oxidative stress (OS) represents a state of an imbalanced amount of reactive oxygen species (ROS) and/or a hampered efficacy of the antioxidative defense system. Cells of the central nervous system are particularly sensitive to OS, as they have a massive need of oxygen to maintain proper function. Consequently, OS represents a common pathophysiological hallmark of neurodegenerative diseases and is discussed to contribute to the neurodegeneration observed amongst others in Alzheimer's disease and Parkinson's disease. In this context, accumulating evidence suggests that OS is involved in the pathophysiology of Niemann-Pick type C1 disease (NPC1). NPC1, a rare hereditary neurodegenerative disease, belongs to the family of lysosomal storage disorders. A major hallmark of the disease is the accumulation of cholesterol and other glycosphingolipids in lysosomes. Several studies describe OS both in murine in vivo and in vitro NPC1 models. However, studies based on human cells are limited to NPC1 patient-derived fibroblasts. Thus, we analyzed OS in a human neuronal model based on NPC1 patient-specific induced pluripotent stem cells (iPSCs). Higher ROS levels, as determined by DCF (dichlorodihydrofluorescein) fluorescence, indicated oxidative stress in all NPC1-deficient cell lines. This finding was further supported by reduced superoxide dismutase (SOD) activity. The analysis of mRNA and protein levels of SOD1 and SOD2 did not reveal any difference between control cells and NPC1-deficient cells. Interestingly, we observed a striking decrease in catalase mRNA and protein levels in all NPC1-deficient cell lines. As catalase is a key enzyme of the cellular antioxidative defense system, we concluded that the lack of catalase contributes to the elevated ROS levels observed in NPC1-deficient cells. Thus, a restitution of a physiological catalase level may pose an intervention strategy to rescue NPC1-deficient cells from the repercussions of oxidative stress contributing to the neurodegeneration observed in NPC1.
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http://dx.doi.org/10.3390/ijms21207667DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7593914PMC
October 2020

Misfolding of Lysosomal α-Galactosidase a in a Fly Model and Its Alleviation by the Pharmacological Chaperone Migalastat.

Int J Mol Sci 2020 Oct 7;21(19). Epub 2020 Oct 7.

The Shmunis School of Biomedicine and Cancer Research, Life Sciences, Tel Aviv University, Ramat Aviv 69978, Israel.

Fabry disease, an X-linked recessive lysosomal disease, results from mutations in the gene encoding lysosomal α-galactosidase A (α-Gal A). Due to these mutations, there is accumulation of globotriaosylceramide (GL-3) in plasma and in a wide range of cells throughout the body. Like other lysosomal enzymes, α-Gal A is synthesized on endoplasmic reticulum (ER) bound polyribosomes, and upon entry into the ER it undergoes glycosylation and folding. It was previously suggested that α-Gal A variants are recognized as misfolded in the ER and undergo ER-associated degradation (ERAD). In the present study, we used to model misfolding of α-Gal A mutants. We did so by creating transgenic flies expressing mutant α-Gal A variants and assessing development of ER stress, activation of the ER stress response and their relief with a known α-Gal A chaperone, migalastat. Our results showed that the A156V and the A285D α-Gal A mutants underwent ER retention, which led to activation of unfolded protein response (UPR) and ERAD. UPR could be alleviated by migalastat. When expressed in the fly's dopaminergic cells, misfolding of α-Gal A and UPR activation led to death of these cells and to a shorter life span, which could be improved, in a mutation-dependent manner, by migalastat.
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http://dx.doi.org/10.3390/ijms21197397DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7583893PMC
October 2020

Guppies Prefer to Follow Large (Robot) Leaders Irrespective of Own Size.

Front Bioeng Biotechnol 2020 15;8:441. Epub 2020 May 15.

Faculty of Life Sciences, Thaer Institute, Humboldt-Universität zu Berlin, Berlin, Germany.

Body size is often assumed to determine how successfully an individual can lead others with larger individuals being better leaders than smaller ones. But even if larger individuals are more readily followed, body size often correlates with specific behavioral patterns and it is thus unclear whether larger individuals are more often followed than smaller ones because of their size or because they behave in a certain way. To control for behavioral differences among differentially-sized leaders, we used biomimetic robotic fish (Robofish) of different sizes. Live guppies () are known to interact with Robofish in a similar way as with live conspecifics. Consequently, Robofish may serve as a conspecific-like leader that provides standardized behaviors irrespective of its size. We asked whether larger Robofish leaders are preferentially followed and whether the preferences of followers depend on own body size or risk-taking behavior ("boldness"). We found that live female guppies followed larger Robofish leaders in closer proximity than smaller ones and this pattern was independent of the followers' own body size as well as risk-taking behavior. Our study shows a "bigger is better" pattern in leadership that is independent of behavioral differences among differentially-sized leaders, followers' own size and risk-taking behavior.
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http://dx.doi.org/10.3389/fbioe.2020.00441DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7243707PMC
May 2020

An interaction mechanism for the maintenance of fission-fusion dynamics under different individual densities.

PeerJ 2020 14;8:e8974. Epub 2020 May 14.

Department of Biology and Ecology of Fishes, Leibniz-Institute of Freshwater Ecology and Inland Fisheries, Berlin, Germany.

Animals often show high consistency in their social organisation despite facing changing environmental conditions. Especially in shoaling fish, fission-fusion dynamics that describe for which periods individuals are solitary or social have been found to remain unaltered even when density changed. This compensatory ability is assumed to be an adaptation towards constant predation pressure, but the mechanism through which individuals can actively compensate for density changes is yet unknown. The aim of the current study is to identify behavioural patterns that enable this active compensation. We compared the fission-fusion dynamics of two populations of the live-bearing Atlantic molly () that live in adjacent habitats with very different predator regimes: cave mollies that inhabit a low-predation environment inside a sulfidic cave with a low density of predatory water bugs ( sp.), and mollies that live directly outside the cave (henceforth called "surface" mollies) in a high-predation environment. We analysed their fission-fusion dynamics under two different fish densities of 12 and 6 fish per 0.36 m. As expected, surface mollies spent more time being social than cave mollies, and this difference in social time was a result of surface mollies being less likely to discontinue social contact (once they had a social partner) and being more likely to resume social contact (once alone) than cave mollies. Interestingly, surface mollies were also less likely to switch among social partners than cave mollies. A random walk simulation predicted each population to show reduced social encounters in the low density treatment. While cave mollies largely followed this prediction, surface mollies maintained their interaction probabilities even at low density. Surface mollies achieved this by a reduction in the size of a convex polygon formed by the group as density decreased. This may allow them to largely maintain their fission-fusion dynamics while still being able to visit large parts of the available area as a group. A slight reduction (21%) in the area visited at low densities was also observed but insufficient to explain how the fish maintained their fission-fusion dynamics. Finally, we discuss potential movement rules that could account for the reduction of polygon size and test their performance.
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http://dx.doi.org/10.7717/peerj.8974DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7231501PMC
May 2020

Involving Patient Groups in Drug Research: A Systematic Review of Reasons.

Patient Prefer Adherence 2020 12;14:587-597. Epub 2020 Mar 12.

Institute of Ethics and History of Medicine, University Medicine Greifswald, Greifswald, Germany.

Background: Patients have evolved from mere objects of study to active contributors to drug research in recent decades. Since individual patient's influence to change research processes effectively is limited, patient groups play an important role in the planning and conducting of pharmaceutical studies. Patient group engagement in drug research is usually seen as being beneficial from an ethical viewpoint as well as from the perspective of research practice, while potential disadvantages and risks have been discussed considerably less.

Purpose: A systematic review of reasons was conducted to allow for an overview of the reasons for and against involving patient groups in drug research.

Methods: The literature search was conducted in PubMed and Web of Science. Reasons concerning the influence of patient groups on drug research were extracted and synthesized using qualitative content analysis. The review's main limitation arises from a lack of critical appraisal regarding the quality of the reasons.

Results: A total of 2271 references were retrieved, of which 97 were included in the analysis. Data extraction revealed 91 (73.4%) reasons for and 30 (24.2%) reasons against involving patient organizations in drug research, and 3 (2.4%) ambivalent reasons; amounting to 124 reasons. The main groups of reasons were clustered around the categories: quality of research, acquisition and allocation of resources, and the patient role in research.

Conclusion: This is the first systematic review of reasons concerning the influence of patient groups on drug research. It provides a basis for a continuing debate about the value as well as the limits of involving patient groups. Due to the diversity of research projects there can be no general recommendation for or against patient group involvement. More research is necessary to assess potential advantages and disadvantages of patient groups' influence on other types of research (eg genetics).
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http://dx.doi.org/10.2147/PPA.S232499DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7075437PMC
March 2020

Paraneoplastic Cerebellar Degeneration in Diffuse Large B-cell Lymphoma and Review of Associated Onconeural Antibodies.

Clin Lymphoma Myeloma Leuk 2020 06 19;20(6):e336-e340. Epub 2020 Feb 19.

Department of Hematology and Oncology, Sylvester Comprehensive Cancer Care Center, University of Miami Miller School of Medicine, Miami, FL.

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http://dx.doi.org/10.1016/j.clml.2020.02.010DOI Listing
June 2020

Physiological Tolerance to ssDNA Enables Strand Uncoupling during DNA Replication.

Cell Rep 2020 02;30(7):2416-2429.e7

Center for Chromosome Stability, Institute for Cellular and Molecular Medicine, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen 2200, Denmark. Electronic address:

It has been long assumed that normally leading strand synthesis must proceed coordinated with the lagging strand to prevent strand uncoupling and the pathological accumulation of single-stranded DNA (ssDNA) in the cell, a dogma recently challenged by in vitro studies in prokaryotes. Here, we report that human DNA polymerases can function independently at each strand in vivo and that the resulting strand uncoupling is supported physiologically by a cellular tolerance to ssDNA. Active forks rapidly accumulate ssDNA at the lagging strand when POLA1 is inhibited without triggering a stress response, despite ssDNA formation being considered a hallmark of replication stress. Acute POLA1 inhibition causes a lethal RPA exhaustion, but cells can duplicate their DNA with limited POLA1 activity and exacerbated strand uncoupling as long as RPA molecules suffice to protect the elevated ssDNA. Although robust, this uncoupled mode of DNA replication is also an in-built weakness that can be targeted for cancer treatment.
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http://dx.doi.org/10.1016/j.celrep.2020.01.067DOI Listing
February 2020

Results of a Randomized Phase IIb Trial of Nelipepimut-S + Trastuzumab versus Trastuzumab to Prevent Recurrences in Patients with High-Risk HER2 Low-Expressing Breast Cancer.

Clin Cancer Res 2020 06 18;26(11):2515-2523. Epub 2020 Feb 18.

Department of Breast Surgical Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas.

Purpose: Preclinical data provide evidence for synergism between HER2-targeted peptide vaccines and trastuzumab. The efficacy of this combination was evaluated in patients with HER2 low-expressing breast cancer in the adjuvant setting.

Patients And Methods: A phase IIb, multicenter, randomized, single-blinded, controlled trial enrolled disease-free patients after standard therapy completion (NCT01570036). Eligible patients were HLA-A2, A3, A24, and/or A26+, and had HER2 IHC 1+/2+, FISH nonamplified breast cancer, that was node positive and/or hormone receptor-negative [triple-negative breast cancer (TNBC)]. Patients received trastuzumab for 1 year and were randomized to placebo (GM-CSF, control) or nelipepimut-S (NPS) with GM-CSF. Primary outcome was 24-month disease-free survival (DFS). Secondary outcomes were 36-month DFS, safety, and immunologic response.

Results: Overall, 275 patients were randomized; 136 received NPS with GM-CSF, and 139 received placebo with GM-CSF. There were no clinicopathologic differences between groups. Concurrent trastuzumab and NPS with GM-CSF was safe with no additional overall or cardiac toxicity compared with control. At median follow-up of 25.7 (interquartile range, 18.4-32.7) months, estimated DFS did not significantly differ between NPS and control [HR, 0.62; 95% confidence interval (CI), 0.31-1.25; = 0.18]. In a planned exploratory analysis of patients with TNBC, DFS was improved for NPS versus control (HR, 0.26; 95% CI, 0.08-0.81, = 0.01).

Conclusions: The combination of NPS with trastuzumab is safe. In HER2 low-expressing breast cancer, no significant difference in DFS was seen in the intention-to-treat analysis; however, significant clinical benefit was seen in patients with TNBC. These findings warrant further investigation in a phase III randomized trial.
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http://dx.doi.org/10.1158/1078-0432.CCR-19-2741DOI Listing
June 2020

Generation of induced pluripotent stem cell lines AKOSi002-A and AKOSi003-A from symptomatic female adults with Wilson disease.

Stem Cell Res 2020 03 27;43:101708. Epub 2020 Jan 27.

Translational Neurodegeneration Section "Albrecht-Kossel", Department of Neurology, University Medical Center Rostock, 18147 Rostock, Germany; Center for Transdisciplinary Neurosciences Rostock (CTNR), University Medical Center Rostock, University of Rostock, 18147 Rostock, Germany. Electronic address:

Wilson disease (WD) is an inherited, autosomal recessive disorder of copper metabolism caused by mutations in the ATP7B gene. Pathogenic single nucleotide variants (SNVs) lead to functional impairment of the copper transporting ATPase ATP7B, resulting in copper accumulation and toxicity in the liver and brain. We describe the generation of two induced pluripotent stem cell (iPSC) lines derived from fibroblasts of two female WD patients. Patient 1 is compound heterozygous for p.E1064A and p.H1069Q. Patient 2 is homozygous for p.M769V. These iPSCs represent a WD model for pathophysiological studies and pharmacological screening.
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http://dx.doi.org/10.1016/j.scr.2020.101708DOI Listing
March 2020

Assessment of Gene Variant Amenability for Pharmacological Chaperone Therapy with 1-Deoxygalactonojirimycin in Fabry Disease.

Int J Mol Sci 2020 Jan 31;21(3). Epub 2020 Jan 31.

Centogene AG, 18055 Rostock, Germany.

Fabry disease is one of the most common lysosomal storage disorders caused by mutations in the gene encoding lysosomal α-galactosidase A (α-Gal A) and resultant accumulation of glycosphingolipids. The sugar mimetic 1-deoxygalactonojirimycin (DGJ), an orally available pharmacological chaperone, was clinically approved as an alternative to intravenous enzyme replacement therapy. The decision as to whether a patient should be treated with DGJ depends on the genetic variant within the α-galactosidase A encoding gene (). A good laboratory practice (GLP)-validated cell culture-based assay to investigate the biochemical responsiveness of the variants is currently the only source available to obtain pivotal information about susceptibility to treatment. Herein, variants were defined amenable when an absolute increase in enzyme activity of ≥3% of wild type enzyme activity and a relative increase in enzyme activity of ≥1.2-fold was achieved following DGJ treatment. Efficacy testing was carried out for over 1000 identified variants in cell culture. Recent data suggest that about one-third of the variants comply with the amenability criteria. A recent study highlighted the impact of inter-assay variability on DGJ amenability, thereby reducing the power of the assay to predict eligible patients. This prompted us to compare our own α-galactosidase A enzyme activity data in a very similar in-house developed assay with those from the GLP assay. In an essentially retrospective approach, we reviewed 148 gene variants from our former studies for which enzyme data from the GLP study were available and added novel data for 30 variants. We also present data for 18 gene variants for which no data from the GLP assay are currently available. We found that both differences in experimental biochemical data and the criteria for the classification of amenability cause inter-assay discrepancy. We conclude that low baseline activity, borderline biochemical responsiveness, and inter-assay discrepancy are alarm signals for misclassifying a variant that must not be ignored. Furthermore, there is no solid basis for setting a minimum response threshold on which a clinical indication with DGJ can be justified.
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http://dx.doi.org/10.3390/ijms21030956DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7037350PMC
January 2020

Pharmacological Chaperones: A Therapeutic Approach for Diseases Caused by Destabilizing Missense Mutations.

Int J Mol Sci 2020 Jan 13;21(2). Epub 2020 Jan 13.

Istituto di Chimica Biomolecolare-CNR, 80078 Pozzuoli, Italy.

The term "pharmacological chaperone" was introduced 20 years ago. Since then the approach with this type of drug has been proposed for several diseases, lysosomal storage disorders representing the most popular targets. The hallmark of a pharmacological chaperone is its ability to bind a protein specifically and stabilize it. This property can be beneficial for curing diseases that are associated with protein mutants that are intrinsically active but unstable. The total activity of the affected proteins in the cell is lower than normal because they are cleared by the quality control system. Although most pharmacological chaperones are reversible competitive inhibitors or antagonists of their target proteins, the inhibitory activity is neither required nor desirable. This issue is well documented by specific examples among which those concerning Fabry disease. Direct specific binding is not the only mechanism by which small molecules can rescue mutant proteins in the cell. These drugs and the properly defined pharmacological chaperones can work together with different and possibly synergistic modes of action to revert a disease phenotype caused by an unstable protein.
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http://dx.doi.org/10.3390/ijms21020489DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7014102PMC
January 2020

Proteostasis regulators modulate proteasomal activity and gene expression to attenuate multiple phenotypes in Fabry disease.

Biochem J 2020 01;477(2):359-380

Translational Neurodegeneration Section "Albrecht-Kossel", Department of Neurology, University Medical Center Rostock, University of Rostock, 18147 Rostock, Germany.

The lysosomal storage disorder Fabry disease is characterized by a deficiency of the lysosomal enzyme α-Galactosidase A. The observation that missense variants in the encoding GLA gene often lead to structural destabilization, endoplasmic reticulum retention and proteasomal degradation of the misfolded, but otherwise catalytically functional enzyme has resulted in the exploration of alternative therapeutic approaches. In this context, we have investigated proteostasis regulators (PRs) for their potential to increase cellular enzyme activity, and to reduce the disease-specific accumulation of the biomarker globotriaosylsphingosine in patient-derived cell culture. The PRs also acted synergistically with the clinically approved 1-deoxygalactonojirimycine, demonstrating the potential of combination treatment in a therapeutic application. Extensive characterization of the effective PRs revealed inhibition of the proteasome and elevation of GLA gene expression as paramount effects. Further analysis of transcriptional patterns of the PRs exposed a variety of genes involved in proteostasis as potential modulators. We propose that addressing proteostasis is an effective approach to discover new therapeutic targets for diseases involving folding and trafficking-deficient protein mutants.
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http://dx.doi.org/10.1042/BCJ20190513DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6993862PMC
January 2020

Ectopic thyroid with benign and malignant findings: A case series.

Int J Surg Case Rep 2020 18;66:33-38. Epub 2019 Nov 18.

Department of Radiology, Na Homolce Hospital, Prague, Czech Republic.

Introduction: Ectopia is the most common sporadically occurring thyroid heterotopy. We present three cases of ectopic thyroid tissue with compression of the upper aerodigestive tract. The first case involved ectopic thyroid tissue in the lingual area of a 60-year-old male with dysphagia, swelling at the base of the tongue, and stomatolalia. The second case was a 66-year-old female with papillary thyroid carcinoma (PTC) in a thyroglossal duct cyst. The third patient was a 50-year-old female with aberrant thyroid tissue in the right submandibular region, with a cribriform-morular variant of PTC (CMV-PTC).

Methods: After resecting the heterotopic tissue and verifying the presence of PTC, the second and third cases underwent total thyroidectomy, and the third patient also underwent radioactive iodine ablation (RAI). Postoperative athyreosis was compensated by permanent levothyroxine substitution.

Results: The diagnosis of ectopic thyroid tissue is challenging. Clinical examination together with imaging methods play a key role, especially postoperative histological examination along with scintigraphy and single photon emission computed tomography (SPECT). Ultrasonography should be used to exclude normally localized thyroid tissue and to distinguish other tumorous diseases. In the pre-operative examination, ultrasound-guided fine-needle aspiration biopsy (US-FNAB) often results in technically-difficult sampling and non-diagnostic cytology.

Conclusion: Resection is the most suitable therapy for clinical symptoms of a foreign body in the upper aerodigestive tract and inflammatory complications; total thyroidectomy follows in case of malignant transformation. Thyroid heterotopy is a rare pathological condition, yet it should be taken into consideration during differential diagnosis of tumorous oropharyngeal and neck lesions.
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http://dx.doi.org/10.1016/j.ijscr.2019.11.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6909043PMC
November 2019

Generation of the Niemann-Pick type C2 patient-derived iPSC line AKOSi001-A.

Stem Cell Res 2019 12 15;41:101606. Epub 2019 Oct 15.

Translational Neurodegeneration Section "Albrecht-Kossel", Department of Neurology, University Medical Center Rostock, Gehlsheimer Straße 20, Rostock 18147, Germany; Center for Transdisciplinary Neurosciences Rostock (CTNR), University Medical Center Rostock, University of Rostock, Rostock 18147, Germany. Electronic address:

Niemann-Pick disease Type C (NPC) is a rare progressive neurodegenerative disorder with an incidence of 1:120,000 caused by mutations in the NPC1 or NPC2 gene. Only 5% of NPC patients suffer from mutations of the NPC2 gene. Here we demonstrate the generation of a Niemann-Pick disease Type C2 (NPC2) patient-derived induced pluripotent stem cell line. This cell line is capable to differentiate into derivatives of the neuronal lineage, providing a valuable tool to study pathogenic mechanisms of NPC2.
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http://dx.doi.org/10.1016/j.scr.2019.101606DOI Listing
December 2019

Stabilization of chromatin topology safeguards genome integrity.

Nature 2019 10 23;574(7779):571-574. Epub 2019 Oct 23.

Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.

To safeguard genome integrity in response to DNA double-strand breaks (DSBs), mammalian cells mobilize the neighbouring chromatin to shield DNA ends against excessive resection that could undermine repair fidelity and cause damage to healthy chromosomes. This form of genome surveillance is orchestrated by 53BP1, whose accumulation at DSBs triggers sequential recruitment of RIF1 and the shieldin-CST-POLα complex. How this pathway reflects and influences the three-dimensional nuclear architecture is not known. Here we use super-resolution microscopy to show that 53BP1 and RIF1 form an autonomous functional module that stabilizes three-dimensional chromatin topology at sites of DNA breakage. This process is initiated by accumulation of 53BP1 at regions of compact chromatin that colocalize with topologically associating domain (TAD) sequences, followed by recruitment of RIF1 to the boundaries between such domains. The alternating distribution of 53BP1 and RIF1 stabilizes several neighbouring TAD-sized structures at a single DBS site into an ordered, circular arrangement. Depletion of 53BP1 or RIF1 (but not shieldin) disrupts this arrangement and leads to decompaction of DSB-flanking chromatin, reduction in interchromatin space, aberrant spreading of DNA repair proteins, and hyper-resection of DNA ends. Similar topological distortions are triggered by depletion of cohesin, which suggests that the maintenance of chromatin structure after DNA breakage involves basic mechanisms that shape three-dimensional nuclear organization. As topological stabilization of DSB-flanking chromatin is independent of DNA repair, we propose that, besides providing a structural scaffold to protect DNA ends against aberrant processing, 53BP1 and RIF1 safeguard epigenetic integrity at loci that are disrupted by DNA breakage.
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http://dx.doi.org/10.1038/s41586-019-1659-4DOI Listing
October 2019

Determination of the Pathological Features of NPC1 Variants in a Cellular Complementation Test.

Int J Mol Sci 2019 Oct 19;20(20). Epub 2019 Oct 19.

Translational Neurodegeneration Section "Albrecht-Kossel", Department of Neurology, University Medical Center Rostock, University of Rostock, 18147 Rostock, Germany.

Niemann-Pick Type C (NP-C) is a rare disorder of lipid metabolism caused by mutations within the and genes. NP-C is a neurovisceral disease leading to a heterogeneous, multisystemic spectrum of symptoms in those affected. Until now, there is no investigative tool to demonstrate the significance of single variants within the genes. Hence, the aim of the study was to establish a test that allows for an objective assessment of the pathological potential of gene variants. Chinese hamster ovary cells defective in the gene accumulate cholesterol in lysosomal storage organelles. The cells were transfected with NPC1-GFP plasmid vectors carrying distinct sequence variants. Filipin staining was used to test for complementation of the phenotype. The known variant p.Ile1061Thr showed a significantly impaired cholesterol clearance after 12 and 24 h compared to the wild type. Among the investigated variants, p.Ser954Leu and p.Glu1273Lys showed decelerated cholesterol clearance as well. The remaining variants p.Gln60His, p.Val494Met, and p.Ile787Val showed a cholesterol clearance indistinguishable from wild type. Further, p.Ile1061Thr acquired an enhanced clearance ability upon 25-hydroxycholesterol treatment. We conclude that the variants that caused an abnormal clearance phenotype are highly likely to be of clinical relevance. Moreover, we present a system that can be utilized to screen for new drugs.
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http://dx.doi.org/10.3390/ijms20205185DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6834306PMC
October 2019

Transformed mucosa-associated lymphoid tissue lymphomas: A single institution retrospective study including polymerase chain reaction-based clonality analysis.

Br J Haematol 2019 08 24;186(3):448-459. Epub 2019 May 24.

Department of Medicine I, Clinical Division of Oncology, Medical University of Vienna, Vienna, Austria.

Given the lack of consistent data regarding the clinico-pathological features and clonal lymphomagenesis of patients with mucosa-associated lymphoid tissue (MALT) lymphoma and histological transformation (HT), we have systematically analysed 379 patients (32% gastric, 68% extra-gastric; median follow-up 52 months) diagnosed with HT at the Medical University Vienna 1999-2017, and reassessed tissues of identified patients by polymerase chain reaction (PCR)-based clonality analysis. HT was documented in 12/379 patients (3·2%) and occurred at a median time of 22 months (range; 6-202 months) after diagnosis of MALT lymphoma. By PCR-based clonality analysis, we detected a clear-cut clonal relationship of MALT lymphoma and diffuse large B-cell lymphoma (DLBCL) in 8 of 11 analysed cases proving that the large majority of DLBCL following MALT lymphoma are clonally-related and constitute a real transformation. Interestingly, HT occurred within the first 2·5 years after diagnosis in patients with clonal relationship, whereas time to aggressive lymphoma was longer in patients identified as clonally-unrelated (most likely secondary) lymphoma (82-202 months), suggesting that HT is an early event in this disease. Survival of patients with HT was poor with 6/12 dying at 1·5-33 months after HT, however, patients with localized gastric transformation had a superior outcome with only 1/6 dying due to progression of lymphoma.
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http://dx.doi.org/10.1111/bjh.15953DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6771836PMC
August 2019

Silent but significant - A synonymous SNV alters prognosis in Pompe disease.

EBioMedicine 2019 May 12;43:20-21. Epub 2019 Apr 12.

Department of Neurology, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA; Program in Medical and Population Genetics, Broad Institute of MIT and Harvard, Cambridge, MA, USA.

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http://dx.doi.org/10.1016/j.ebiom.2019.04.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6562060PMC
May 2019

Role of endoplasmic reticulum stress and protein misfolding in disorders of the liver and pancreas.

Adv Med Sci 2019 Sep 9;64(2):315-323. Epub 2019 Apr 9.

Department of Medicine A, University Medicine Greifswald, Greifswald, Germany. Electronic address:

The endoplasmic reticulum (ER) is the site of synthesis and folding of membrane and secretory proteins. The fraction of protein passing through the ER represents a large proportion of the total protein in the cell. Protein folding, glycosylation, sorting and transport are essential tasks of the ER and a compromised ER folding network has been recognized to be a key component in the disease pathogenicity of common neurodegenerative, metabolic and malignant diseases. On the other hand, the ER protein folding machinery also holds significant potential for therapeutic interventions. Many causes can lead to ER stress. A disturbed calcium homeostasis, the generation of reactive oxygen species (ROS) and a persistent overload of misfolded proteins within the ER can drive the course of adisease. In this review the role of ER-stress in diseases of the liver and pancreas will be examined using pancreatitis and Wilson´s disease as examples. Potential therapeutic targets in ER-stress pathways will also be discussed.
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http://dx.doi.org/10.1016/j.advms.2019.03.004DOI Listing
September 2019

Pre-Therapeutic Total Lesion Glycolysis on [F]FDG-PET Enables Prognostication of 2-Year Progression-Free Survival in MALT Lymphoma Patients Treated with CD20-Antibody-Based Immunotherapy.

Mol Imaging Biol 2019 12;21(6):1192-1199

Department of Medicine I, Medical University of Vienna, Vienna, Austria.

Purpose: Standardized uptake values (SUV), total metabolic tumor volumes (TMTV), and total lesion glycolysis (TLG) based on positron emission tomography with 2-deoxy-2-[F]fluoro-D-glucose ([F]FDG/positron emission tomography (PET) are established outcome predictors in FDG-avid lymphomas. We therefore investigated whether these biomarkers also have prognostic value in extranodal marginal zone B cell lymphoma of the mucosa-associated lymphoid tissue (MALT lymphoma), with a focus on patients treated with anti-CD20 antibody-based immunotherapy.

Procedures: Pre-therapeutic [F]FDG/PET scans of 61 treatment-naïve MALT lymphoma patients, including 35 scheduled for anti-CD20 antibody-based immunotherapy, were included in this retrospective study. SUVmean, SUVmax, TMTV, and TLG were measured and tested for 2-year progression-free survival (PFS) prognostication, using Cox regression analyses. Receiver operating characteristic curves were used to determine optimal cutoffs for prognostic [F]FDG/PET parameters, and Kaplan-Meier estimates with log rank tests were performed.

Results: After 2 years, progression had occurred in 12/61 patients (CD20-anitbody group 6/35). TLG emerged as the only significant prognostic factor for 2-year PFS in the multivariate analyses with forward selection, both in entire cohort (hazard ratio HR, 1.001; 95 % CI, 1.001-1.002; P < 0.0001) and in the CD20-antibody group (HR, 1.001; 95 % CI, 1.001-1.002; P = 0.001). However, in the entire population, where 8/26 patients with a TLG > 90 (30.8 %) vs. 4/35 patients with a TLG ≤ 90 (11.4 %) showed progression within the 2-year observation period, TLG-based separation of risk groups failed (HR, 0.35; 95 % CI, 0.10-1.15; P = 0.069); whereas in the CD20-antibody group, where 6/16 patients with a TLG > 90 (37.5 %) vs. 0/19 patients with a TLG ≤ 90 (0.0 %) showed progression, risk group separation was successful (HR, 0.010; 95 % CI, 0.0001-8.068; P = 0.003).

Conclusions: TLG may improve early risk stratification of MALT lymphoma patients treated with CD20-antibody-based immunotherapy.
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http://dx.doi.org/10.1007/s11307-019-01329-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7604829PMC
December 2019

Initial safety analysis of a randomized phase II trial of nelipepimut-S + GM-CSF and trastuzumab compared to trastuzumab alone to prevent recurrence in breast cancer patients with HER2 low-expressing tumors.

Clin Immunol 2019 04 25;201:48-54. Epub 2019 Feb 25.

Department of Breast Surgical Oncology, The University of Texas MD Anderson Cancer Center, 1400 Pressler Street, Houston, TX 77030, USA. Electronic address:

The development of HER2-targeted therapy has decreased recurrence rates and improved survival, transforming the natural history of HER2-positive breast cancer. However only a minority of breast cancer patients benefit as these agents are not used in patients with tumors expressing low levels of HER2. Preclinical data suggests a synergistic action of HER2-targeted vaccination with trastuzumab. We report the initial safety interim analysis of a phase II trial that enrolled patients with HER2 low-expressing (IHC 1+/2+) breast cancer who were clinically disease-free after standard therapy. Patients were randomized to receive the HER2-peptide vaccine nelipepimut-S + GM-CSF with trastuzumab (vaccine arm) or trastuzumab + GM-CSF (control arm) and were followed for recurrence. A planned analysis that occurred after enrollment of 150 patients showed no significant differences in toxicity between the two arms, including cardiac toxicity. The clinical efficacy of this combination will be reported 6 months after the final patient was enrolled.
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http://dx.doi.org/10.1016/j.clim.2019.02.011DOI Listing
April 2019

53BP1 nuclear bodies enforce replication timing at under-replicated DNA to limit heritable DNA damage.

Nat Cell Biol 2019 04 25;21(4):487-497. Epub 2019 Feb 25.

Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.

Failure to complete DNA replication is a stochastic by-product of genome doubling in almost every cell cycle. During mitosis, under-replicated DNA (UR-DNA) is converted into DNA lesions, which are inherited by daughter cells and sequestered in 53BP1 nuclear bodies (53BP1-NBs). The fate of such cells remains unknown. Here, we show that the formation of 53BP1-NBs interrupts the chain of iterative damage intrinsically embedded in UR-DNA. Unlike clastogen-induced 53BP1 foci that are repaired throughout interphase, 53BP1-NBs restrain replication of the embedded genomic loci until late S phase, thus enabling the dedicated RAD52-mediated repair of UR-DNA lesions. The absence or malfunction of 53BP1-NBs causes premature replication of the affected loci, accompanied by genotoxic RAD51-mediated recombination. Thus, through adjusting replication timing and repair pathway choice at under-replicated loci, 53BP1-NBs enable the completion of genome duplication of inherited UR-DNA and prevent the conversion of stochastic under-replications into genome instability.
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http://dx.doi.org/10.1038/s41556-019-0293-6DOI Listing
April 2019