Mechanisms of growth inhibition of primary prostate epithelial cells following gamma irradiation or photodynamic therapy including senscence, necrosis, and autophagy, but not apoptosis

In comparison to more differentiated cells, prostate cancer stem-like cells are radioresistant, which could explain radio-recurrent prostate cancer. Improvement of radiotherapeutic efficacy may therefore require combination therapy. We have investigated the consequences of treating primary prostate epithelial cells with gamma irradiation and photodynamic therapy (PDT), both of which act through production of reactive oxygen species (ROS). Primary prostate epithelial cells were cultured from patient samples of benign prostatic hyperplasia and prostate cancer prior to treatment with PDT or gamma irradiation. Cell viability was measured using MTT and alamar blue assay, and cell recovery by colony-forming assays. Immunofluorescence of gamma-H2AX foci was used to quantify DNA damage, and autophagy and apoptosis were assessed using Western blots. Necrosis and senescence were measured by propidium iodide staining and beta-galactosidase staining, respectively. Both PDT and gamma irradiation reduced the colony-forming ability of primary prostate epithelial cells. PDT reduced the viability of all types of cells in the cultures, including stem-like cells and more differentiated cells. PDT induced necrosis and autophagy, whereas gamma irradiation induced senescence, but neither treatment induced apoptosis. PDT and gamma irradiation therefore inhibit cell growth by different mechanisms. We suggest these treatments would be suitable for use in combination as sequential treatments against prostate cancer

"Petit spot" rejuvenated volcanism superimposed on plume-derived Samoan shield volcanoes: Evidence from a 645-m drill core from Tutuila Island, American Samoa

Author Posting. © American Geophysical Union, 2019. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Geochemistry Geophysics Geosystems 20(3), (2019): 1485-1507, doi:10.1029/2018GC007985.In 2015 a geothermal exploration well was drilled on the island of Tutuila, American Samoa. The sample suite from the drill core provides 645 m of volcanic stratigraphy from a Samoan volcano, spanning 1.45 million years of volcanic history. In the Tutuila drill core, shield lavas with an EM2 (enriched mantle 2) signature are observed at depth, spanning 1.46 to 1.44 Ma. These are overlain by younger (1.35 to 1.17 Ma) shield lavas with a primordial “common” (focus zone) component interlayered with lavas that sample a depleted mantle component. Following ~1.15 Myr of volcanic quiescence, rejuvenated volcanism initiated at 24.3 ka and samples an EM1 (enriched mantle 1) component. The timing of the initiation of rejuvenated volcanism on Tutuila suggests that rejuvenated volcanism may be tectonically driven, as Samoan hotspot volcanoes approach the northern terminus of the Tonga Trench. This is consistent with a model where the timing of rejuvenated volcanism at Tutuila and at other Samoan volcanoes relates to their distance from the Tonga Trench. Notably, the Samoan rejuvenated lavas have EM1 isotopic compositions distinct from shield lavas that are geochemically similar to “petit spot” lavas erupted outboard of the Japan Trench and late stage lavas erupted at Christmas Island located outboard of the Sunda Trench. Therefore, like the Samoan rejuvenated lavas, petit spot volcanism in general appears to be related to tectonic uplift outboard of subduction zones, and existing geochemical data suggest that petit spots share similar EM1 isotopic signatures.Reviews from Kaj Hoernle and three anonymous reviewers are gratefully acknowledged. M. G. J. acknowledges support from the American Samoa Power Authority and National Science Foundation grants OCE‐1736984 and EAR‐1624840. The Tutuila drill core was the brainchild of Tim Bodell, without whom we would still have no stratigraphic record of Tutuila volcanism. The support of Utu Abe Malae and Matamua Katrina Mariner was instrumental to the project's success. We dedicate this paper to the memory of Abe Malae and his efforts to support science and education in American Samoa. Images of the entire drill core are available online (escholarship.org/uc/item/6gg6p61w). All data presented are either part of this study or previously published and are referenced in text.2019-08-1

Smoking cessation counselling training in the pre-clerkship curriculum of Canadian medical schools: A national survey

Background: Cigarette use is Canada’s leading cause of preventable disease, disability, and death. The Medical Council of Canada requires that physicians be able to address tobacco-use, however smoking cessation counselling (SCC) training remains largely neglected in the pre-clerkship curricula of many Canadian medical schools.Methods: Between July and October of 2015, Canada’s 17 medical schools were invited to participate in an administrative survey: The Canadian Medical School Assessment of Smoking Cessation Counselling in the Pre-Clerkship Curriculum. Each was asked to comment on the presence of 28 tobacco-related topics in their curricula, including: time devoted to source material; year(s) of training during which medical students were exposed to related content; methods of teaching and examination; and, the attitudes, policies, and barriers relevant to the presence of smoking cessation counselling (SCC) training in the curriculum.A second short survey: Assessing Medical Students Attitudes toward Smoking Cessation Education was distributed to 100 University of Ottawa medical students to assess comfort level and perceived confidence toward addressing smoking cessation with patients.Results: Eleven of 17 medical schools completed the administrative survey. The results demonstrated substantial deficits and inconsistencies in the delivery of SCC training in the pre-clerkship curricula of Canada’s medical schools.  The short survey revealed perceived discomfort regarding smoking cessation discussion, consistent with the potential curriculum deficits suggested in the larger national survey.Conclusion: The results of both surveys suggest an unfortunate oversight given the devastating impact of tobacco-related diseases. Institutional commitment and enhanced inter-university collaboration could facilitate the development of a national undergraduate medical education program to enhance the delivery of SCC training within the pre-clerkship curricula of Canadian medical schools

Inhibition of the glucocorticoid receptor results in an enhanced miR-99a/100-mediated radiation response in stem-like cells from human prostate cancers

Radiation therapy is a major primary treatment option for both localized early stage prostate cancer, and for advanced, regionally un-resectable, cancer. However, around 30% of patients still experience biochemical recurrence after radiation therapy within 10 years. Thus, identification of better biomarkers and new targets are urgently required to improve current therapeutic strategies. The miR-99 family has been shown to play an important role in the regulation of the DNA damage response, via targeting of the SWI/SNF chromatin remodeling factors, SMARCA5 and SMARCD1 in cell line models. In the present study, we have demonstrated that low expression of miR-99a and miR-100 is present in cell populations which are relatively radiation insensitive, for example in prostate cancer stem cells and in castration-resistant prostate cancer. Additionally, treatment of cells with the synthetic glucocorticoid, Dexamethasone resulted in decreased miR-99a and 100 expression, suggesting a new mechanism of miR-99a and 100 regulation in androgen-independent prostate cells. Strikingly, treatment of prostate cells with the glucocorticoid receptor inhibitor, Mifepristone was found to sensitize prostate cells to radiation by increasing the levels of miR-99a and miR-100. These results qualify the miR99 family as markers of radiation sensitivity and as potential therapeutic targets to improve efficiency of radiotherapy

Inhibition of the PI3K/AKT/mTOR pathway activates autophagy and compensatory Ras/Raf/MEK/ERK signalling in prostate cancer

The PI3K/AKT/mTOR pathway is frequently activated in advanced prostate cancer, due to loss of the tumour suppressor PTEN, and is an important axis for drug development. We have assessed the molecular and functional consequences of pathway blockade by inhibiting AKT and mTOR kinases either in combination or as individual drug treatments. In established prostate cancer cell lines, a decrease in cell viability and in phospho-biomarker expression was observed. Although apoptosis was not induced, a G1 growth arrest was observed in PTEN null LNCaP cells, but not in BPH1 or PC3 cells. In contrast, when the AKT inhibitor AZD7328 was applied to patient-derived prostate cultures that retained expression of PTEN, activation of a compensatory Ras/MEK/ERK pathway was observed. Moreover, whilst autophagy was induced following treatment with AZD7328, cell viability was less affected in the patient-derived cultures than in cell lines. Surprisingly, treatment with a combination of both AZD7328 and two separate MEK1/2 inhibitors further enhanced phosphorylation of ERK1/2 in primary prostate cultures. However, it also induced irreversible growth arrest and senescence.Ex vivo treatment of a patient-derived xenograft (PDX) of prostate cancer with a combination of AZD7328 and the mTOR inhibitor KU-0063794, significantly reduced tumour frequency upon re-engraftment of tumour cells.The results demonstrate that single agent targeting of the PI3K/AKT/mTOR pathway triggers activation of the Ras/MEK/ERK compensatory pathway in near-patient samples. Therefore, blockade of one pathway is insufficient to treat prostate cancer in man

Modeling of Biochemical States of DNA Replication Using Hidden Markov Models

In nanopore experiments, DNA replication facilitated by $\phi 29$ DNA polymerase (DNAP) can be observed at the single molecule level. The biochemical state of the DNA-DNAP complex was studied by setting the complex atop a $\alpha$-hemolysin nanopore and applying an electric voltage. The movement of the DNA strand relative to the nanopore was observed on a single base pair level by the ionic current blockade. The time trace of the recorded ionic current amplitude from these experiments was used to study the biochemical states. Given that the recorded amplitude of the ionic current was an indirect measurement of the true amplitude level, which in turn was an indirect measurement of the true biochemical state, the experiments were modeled as a Hidden Markov Chain (HMC). When the DNA position of two biochemical states relative to the nanopore is the same, the states yield the same current amplitude level. To extract the dynamic transition rates between biochemical states that are not distinguishable in amplitude level, two methodologies were applied to study the HMC. The first was a fully Bayesian model, for which Markov chain Monte-Carlo (MCMC) simulations were used to infer the reaction rates in a system of three biochemical states with two observed amplitude levels. The second model adopted concepts of Viterbi training or the segmental k-means algorithm to find point estimates of the transition rates. Given the low transition probabilities, the properties of the second model led to a substantial bias in inference. The bias was addressed by first using a meta-model to describe the relation between the generating transition rates and the biased inference. Then the inverse problem of the meta-model was solved to reduce the bias in the inference. The meta-model was a fully Bayesian Gaussian process model, built by creating a series of computer-generated datasets around the given dataset. Improved inference was obtained by drawing posterior samples from the inverse of the meta-model. Since both the MCMC simulations and bias reduction techniques resulted in simulated posteriors, the two methods were used to confirm each other. In the comparison of these two methods, the approach of the second model plus bias reduction has the advantage of achieving similar inference accuracy at a much lower computational cost

GluN2A NMDA Receptor Enhancement Improves Brain Oscillations, Synchrony, and Cognitive Functions in Dravet Syndrome and Alzheimer's Disease Models.

NMDA receptors (NMDARs) play subunit-specific roles in synaptic function and are implicated in neuropsychiatric and neurodegenerative disorders. However, the in vivo consequences and therapeutic potential of pharmacologically enhancing NMDAR function via allosteric modulation are largely unknown. We examine the in vivo effects of GNE-0723, a positive allosteric modulator of GluN2A-subunit-containing NMDARs, on brain network and cognitive functions in mouse models of Dravet syndrome (DS) and Alzheimer's disease (AD). GNE-0723 use dependently potentiates synaptic NMDA receptor currents and reduces brain oscillation power with a predominant effect on low-frequency (12-20 Hz) oscillations. Interestingly, DS and AD mouse models display aberrant low-frequency oscillatory power that is tightly correlated with network hypersynchrony. GNE-0723 treatment reduces aberrant low-frequency oscillations and epileptiform discharges and improves cognitive functions in DS and AD mouse models. GluN2A-subunit-containing NMDAR enhancers may have therapeutic benefits in brain disorders with network hypersynchrony and cognitive impairments

Immunochip analysis identifies multiple susceptibility loci for systemic sclerosis

In this study, 1,833 systemic sclerosis (SSc) cases and 3,466 controls were genotyped with the Immunochip array. Classical alleles, amino acid residues, and SNPs across the human leukocyte antigen (HLA) region were imputed and tested. These analyses resulted in a model composed of six polymorphic amino acid positions and seven SNPs that explained the observed significant associations in the region. In addition, a replication step comprising 4,017 SSc cases and 5,935 controls was carried out for several selected non-HLA variants, reaching a total of 5,850 cases and 9,401 controls of European ancestry. Following this strategy, we identified and validated three SSc risk loci, including DNASE1L3 at 3p14, the SCHIP1-IL12A locus at 3q25, and ATG5 at 6q21, as well as a suggested association of the TREH-DDX6 locus at 11q23. The associations of several previously reported SSc risk loci were validated and further refined, and the observed peak of association in PXK was related to DNASE1L3. Our study has increased the number of known genetic associations with SSc, provided further insight into the pleiotropic effects of shared autoimmune risk factors, and highlighted the power of dense mapping for detecting previously overlooked susceptibility loci

Evaluating the Role of Ultrasound in Prostate cancer (ERUP) trial – Phase 1 early experience of micro-ultrasound in the UK

Purpose: To evaluate if the use of micro-ultrasound (microUS) can detect significant prostate (csPCa) pathology when compared to histology obtained during a transperineal prostate biopsy.Methods: Patients suspected of having prostate cancer, who had a prebiopsy MRI and could tolerate a transrectal examination were prospectively recruited. All patients had a microUS scan prior to their biopsy. The findings of MRI, microUS, and histology were risk stratified in accordance with local pathways. Comparison of assigned risk scores were made using histology as the reference standard.Results: Data from 101 patients were evaluated. Histology showed that csPCa was detected in 48.5% (n = 49/101) of patients. Moderate inter-rater agreement was found in both MRI and MicroUS with К of 0.31 in both modalities. High risk findings were identified in 81% (n = 82/101) patients at MRI and in 66% (n = 67/101) patients at microUS. Sensitivity and specificity of MRI was found to be 87% and 34.6% and for microUS 73.3% and 53.8% respectively.Conclusion: A limitation of this study was that the biopsy was not performed with microUS which may have resulted in unidentified cancers and lowered the apparent accuracy of the technique. However, we conclude that whilst microUS was diagnostic, MRI demonstrated higher sensitivity in our local population and remains the pre-biopsy imaging modality of choice. However, the higher specificity of microUS identified does indicate that it may be of value when MRI is contraindicated. The role of microUS, within an active surveillance pathway for prostate cancer, warrants further investigation