Overcoming hypoxia-induced tumor radioresistance in non-small cell lung cancer by targeting DNA-dependent protein kinase in combination with carbon ion irradiation

Background: Hypoxia-induced radioresistance constitutes a major obstacle for a curative treatment of cancer. The aim of this study was to investigate effects of photon and carbon ion irradiation in combination with inhibitors of DNA-Damage Response (DDR) on tumor cell radiosensitivity under hypoxic conditions. Methods: Human non-small cell lung cancer (NSCLC) models, A549 and H1437, were irradiated with dose series of photon and carbon ions under hypoxia (1% O2) vs. normoxic conditions (21% O2). Clonogenic survival was studied after dual combinations of radiotherapy with inhibitors of DNA-dependent Protein Kinase (DNAPKi, M3814) and ATM serine/threonine kinase (ATMi). Results: The OER at 30% survival for photon irradiation of A549 cells was 1.4. The maximal oxygen effect measured as survival ratio was 2.34 at 8 Gy photon irradiation of A549 cells. In contrast, no significant oxygen effect was found after carbon ion irradiation. Accordingly, the relative effect of 6 Gy carbon ions was determined as 3.8 under normoxia and. 4.11 under hypoxia. ATM and DNA-PK inhibitors dose dependently sensitized tumor cells for both radiation qualities. For 100 nM DNAPKi the survival ratio at 4 Gy more than doubled from 1.59 under normoxia to 3.3 under hypoxia revealing a strong radiosensitizing effect under hypoxic conditions. In contrast, this ratio only moderately increased after photon irradiation and ATMi under hypoxia. The most effective treatment was combined carbon ion irradiation and DNA damage repair inhibition. Conclusions: Carbon ions efficiently eradicate hypoxic tumor cells. Both, ATMi and DNAPKi elicit radiosensitizing effects. DNAPKi preferentially sensitizes hypoxic cells to radiotherapy

A solid-phase transfection platform for arrayed CRISPR screens

Arrayed CRISPR‐based screens emerge as a powerful alternative to pooled screens making it possible to investigate a wide range of cellular phenotypes that are typically not amenable to pooled screens. Here, we describe a solid‐phase transfection platform that enables CRISPR‐based genetic screens in arrayed format with flexible readouts. We demonstrate efficient gene knockout upon delivery of guide RNAs and Cas9/guide RNA ribonucleoprotein complexes into untransformed and cancer cell lines. In addition, we provide evidence that our platform can be easily adapted to high‐throughput screens and we use this approach to study oncogene addiction in tumor cells. Finally demonstrating that the human primary cells can also be edited using this method, we pave the way for rapid testing of potential targeted therapies

A solid‐phase transfection platform for arrayed CRISPR screens [Corrigendum]

Since the publication of this study, it has come to our attention that a citation to the study by Bulkescher et al (2017) was omitted from the Introduction. The following sentence should have been included in the introduction: “A previously reported solid‐phase reverse transfection method for proteins (Bulkescher et al , 2017) was used for the delivery of RNPs for three endogenous genes suggesting the potential of solid‐phase reverse transfection for CRISPR/Cas9‐based gene editing, despite its low efficiency”. We apologise for any inconvenience this omission may have caused

Prediction of Means and Variances of Crosses With Genome-Wide Marker Effects in Barley

Background: The expected genetic variance is an important criterion for the selection of crossing partners which will produce superior combinations of genotypes in their progeny. The advent of molecular markers has opened up new vistas for obtaining precise predictors for the genetic variance of a cross, but fast prediction methods that allow plant breeders to select crossing partners based on already available data from their breeding programs without complicated calculations or simulation of breeding populations are still lacking. The main objective of the present study was to demonstrate the practical applicability of an analytical approach for the selection of superior cross combinations with experimental data from a barley breeding program. We used genome-wide marker effects to predict the yield means and genetic variances of 14 DH families resulting from crosses of four donor lines with five registered elite varieties with the genotypic information of the parental lines. For the validation of the predicted parameters, the analytical approach was extended by the masking variance as a major component of phenotypic variance. The predicted parameters were used to fit normal distribution curves of the phenotypic values and to conduct an Anderson-Darling goodness-of-fit test for the observed phenotypic data of the 14 DH families from the field trial.Results: There was no evidence that the observed phenotypic values deviated from the predicted phenotypic normal distributions in 13 out of 14 crosses. The correlations between the observed and the predicted means and the observed and predicted variances were r = 0.95 and r = 0.34, respectively. After removing two crosses with downward outliers in the phenotypic data, the correlation between the observed and predicted variances increased to r = 0.76. A ranking of the 14 crosses based on the sum of predicted mean and genetic variance identified the 50% best crosses from the field trial correctly.Conclusions: We conclude that the prediction accuracy of the presented approach is sufficiently high to identify superior crosses even with limited phenotypic data. We therefore expect that the analytical approach based on genome-wide marker effects is applicable in a wide range of breeding programs

Concurrence of form and function in developing networks and its role in synaptic pruning

A fundamental question in neuroscience is how structure and function of neural systems are related. We study this interplay by combining a familiar auto-associative neural network with an evolving mechanism for the birth and death of synapses. A feedback loop then arises leading to two qualitatively different types of behaviour. In one, the network structure becomes heterogeneous and dissasortative, and the system displays good memory performance; furthermore, the structure is optimised for the particular memory patterns stored during the process. In the other, the structure remains homogeneous and incapable of pattern retrieval. These findings provide an inspiring picture of brain structure and dynamics that is compatible with experimental results on early brain development, and may help to explain synaptic pruning. Other evolving networks—such as those of protein interactions—might share the basic ingredients for this feedback loop and other questions, and indeed many of their structural features are as predicted by our model.We are grateful for financial support from the Spanish MINECO (project of Excellence: FIS2017-84256-P) and from “Obra Social La Caixa”

"Im Alltag der Reform": Eine Oral-History-Studie zu den Gründungs- und Anfangsjahren der Laborschule Bielefeld

Devantié R, Lücker F, Makowski T, Zenke CT. "Im Alltag der Reform": Eine Oral-History-Studie zu den Gründungs- und Anfangsjahren der Laborschule Bielefeld. In: Freke N, Kullmann H, Lücker F, Lütje-Klose B, Schwab S, Zenke CT, eds. Laborschulforschung 2016. Bielefeld; 2016

Die Laborschule als inklusiver Raum

Zenke CT, Dorniak M, Walter J. Die Laborschule als inklusiver Raum. In: Freke N, Kullmann H, Lücker F, Textor A, Zenke CT, eds. Laborschulforschung 2015. Anträge und Berichte zum Forschungs- und Entwicklungsplan. Werkstattheft. Vol 50. Bielefeld: Wissenschaftliche Einrichtung Laborschule; 2015: 125-132

The Role of ATR Inhibitors in Ovarian Cancer: Investigating Predictive Biomarkers of Response

Ataxia telangiectasia and Rad-3 related kinase (ATR) signals DNA lesions and replication stress (RS) to the S and G2/M checkpoints and DNA repair pathways making it a promising target to exploit the dysregulated DNA damage response in cancer. ATR inhibitors (ATRi) are under clinical investigation as monotherapy and in combination with other anticancer agents. Molecular determinants of sensitivity to ATRi are common in ovarian cancer, suggesting the therapeutic potential of ATRi. We investigated the cytotoxicity of the ATRi, VE-821, in a panel of human ovarian cancer cell lines. High grade serous (HGS) cell lines were significantly more sensitive to VE-821 than non-HGS (p ≤ 0.0001) but previously identified determinants of sensitivity (TP53, ATM and BRCA1) were not predictive. Only low RAD51 (p = 0.041), TopBP1 (p = 0.026) and APOBEC3B (p = 0.015) protein expression were associated with increased VE-821 sensitivity. HGS cells had increased levels of RS (pRPASer4/8 and γH2AX nuclear immunofluorescence), and elevated RS predicted sensitivity to VE-821 independently of the cell line subtype. These data suggest that functional assessment of RS biomarkers may be a better predictive biomarker of ATRi response than any single aberrant gene in ovarian cancer and potentially other cancers.Medicine, Faculty ofNon UBCMedicine, Department ofObstetrics and Gynaecology, Department ofReviewedFacultyResearche

„Im Alltag der Reform“. Eine Interview-Studie zu den Gründungs- und Anfangsjahren der Laborschule Bielefeld. Fortsetzungsantrag

Zenke CT, Devantié R, Freke N, Lücker F, Stieber G. „Im Alltag der Reform“. Eine Interview-Studie zu den Gründungs- und Anfangsjahren der Laborschule Bielefeld. Fortsetzungsantrag. In: Dorniak M, Gold J, Textor A, Zenke CT, Zentarra D, eds. Laborschulforschung 2019. Anträge und Berichte zum Forschungs- und Entwicklungsplan (Werkstatthefte, 54). Bielefeld; 2019: 81-86