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The COPE team is very pleased and proud to announce that COPE was chosen as a project success story by the European Commission DG Research & Innovation.
Renal biopsies from donors with acute kidney injury show different molecular patterns according to the post-transplant function.
The utilization of kidneys from donors with acute kidney injury (AKI) is often limited by unpredictable post-transplantation outcomes. The aim of our study was to identify protein mediators implicated in either recovery or failure of these organs. Forty kidney biopsies from donors with (20) and without AKI (20) were selected and then subdivided according to the post-transplant outcome defined as a threshold of 45 ml/min for the eGFR at 1 year from transplantation. Tissue homogenates were analysed by western blot to assess how the levels of 17 pre-selected proteins varied across the four groups. Samples from AKI kidneys with a poor outcome showed a fourfold increase in the levels of PPARg and twofold reduction of STAT1 compared to the other groups (p
Data from VPRBP Functions Downstream of the Androgen Receptor and OGT to Restrict p53 Activation in Prostate Cancer
<div>Abstract<p>Androgen receptor (AR) is a major driver of prostate cancer initiation and progression. O-GlcNAc transferase (OGT), the enzyme that catalyzes the covalent addition of UDP-N-acetylglucosamine (UDP-GlcNAc) to serine and threonine residues of proteins, is often highly expressed in prostate cancer with its expression correlated with high Gleason score. In this study, we have identified an AR and OGT coregulated factor, Vpr (HIV-1) binding protein (VPRBP) also known as DDB1 and CUL4 Associated Factor 1 (DCAF1). We show that VPRBP is regulated by the AR at the transcript level, and stabilized by OGT at the protein level. VPRBP knockdown in prostate cancer cells led to a significant decrease in cell proliferation, p53 stabilization, nucleolar fragmentation, and increased p53 recruitment to the chromatin. In human prostate tumor samples, VPRBP protein overexpression correlated with AR amplification, OGT overexpression, a shorter time to postoperative biochemical progression and poor clinical outcome. In clinical transcriptomic data, VPRBP expression was positively correlated with the AR and also with AR activity gene signatures.</p>Implications:<p>In conclusion, we have shown that VPRBP/DCAF1 promotes prostate cancer cell proliferation by restraining p53 activation under the influence of the AR and OGT.</p></div>
Data from O-GlcNAc Transferase – An Auxiliary Factor or a Full-blown Oncogene?
<div>Abstract<p>The β-linked N-acetyl-d-glucosamine (GlcNAc) is a posttranslational modification of serine and threonine residues catalyzed by the enzyme O-GlcNAc transferase (OGT). Increased OGT expression is a feature of most human cancers and inhibition of OGT decreases cancer cell proliferation. Antiproliferative effects are attributed to posttranslational modifications of known regulators of cancer cell proliferation, such as MYC, FOXM1, and EZH2. In general, OGT amplifies cell-specific phenotype, for example, OGT overexpression enhances reprogramming efficiency of mouse embryonic fibroblasts into stem cells. Genome-wide screens suggest that certain cancers are particularly dependent on OGT, and understanding these addictions is important when considering OGT as a target for cancer therapy. The O-GlcNAc modification is involved in most cellular processes, which raises concerns of on-target undesirable effects of OGT-targeting therapy. Yet, emerging evidence suggest that, much like proteasome inhibitors, specific compounds targeting OGT elicit selective antiproliferative effects in cancer cells, and can prime malignant cells to other treatments. It is, therefore, essential to gain mechanistic insights on substrate specificity for OGT, develop reagents to more specifically enrich for O-GlcNAc–modified proteins, identify O-GlcNAc “readers,” and develop OGT small-molecule inhibitors. Here, we review the relevance of OGT in cancer progression and the potential targeting of this metabolic enzyme as a putative oncogene.</p></div>
Data from Attenuating Adaptive VEGF-A and IL8 Signaling Restores Durable Tumor Control in AR Antagonist–Treated Prostate Cancers
<div>Abstract<p>Inhibiting androgen signaling using androgen signaling inhibitors (ASI) remains the primary treatment for castrate-resistant prostate cancer. Acquired resistance to androgen receptor (AR)-targeted therapy represents a major impediment to durable clinical response. Understanding resistance mechanisms, including the role of AR expressed in other cell types within the tumor microenvironment, will extend the clinical benefit of AR-targeted therapy. Here, we show the ASI enzalutamide induces vascular catastrophe and promotes hypoxia and microenvironment adaptation. We characterize treatment-induced hypoxia, and subsequent induction of angiogenesis, as novel mechanisms of relapse to enzalutamide, highlighting the importance of two hypoxia-regulated cytokines in underpinning relapse. We confirmed AR expression in CD34<sup>+</sup> vascular endothelium of biopsy tissue and human vascular endothelial cells (HVEC). Enzalutamide attenuated angiogenic tubule formation and induced cytotoxicity in HVECs <i>in vitro</i>, and rapidly induced sustained hypoxia in LNCaP xenografts. Subsequent reoxygenation, following prolonged enzalutamide treatment, was associated with increased tumor vessel density and accelerated tumor growth. Hypoxia increased AR expression and transcriptional activity in prostate cells <i>in vitro</i>. Coinhibition of IL8 and VEGF-A restored tumor response in the presence of enzalutamide, confirming the functional importance of their elevated expression in enzalutamide-resistant models. Moreover, coinhibition of IL8 and VEGF-A resulted in a durable, effective resolution of enzalutamide-sensitive prostate tumors. We conclude that concurrent inhibition of two hypoxia-induced factors, IL8 and VEGF-A, prolongs tumor sensitivity to enzalutamide in preclinical models and may delay the onset of enzalutamide resistance.</p>Implications:<p>Targeting hypoxia-induced signaling may extend the therapeutic benefit of enzalutamide, providing an improved treatment strategy for patients with resistant disease.</p></div>
New robotic platform for transoral robotic surgery: an IDEAL stage 0 study
ObjectivesThis study aims to assess the feasibility to perform transoral robotic surgery (TORS) with a new robotic platform, the Versius Surgical System (CMR Surgical, UK) in a preclinical cadaveric setting in accordance to stage 0 of the IDEAL-D framework.DesignIDEAL stage 0 preclinical assessment of the Versius Robotic System in TORS in human cadavers.SettingAll procedures were performed in a simulated operating theatre environment at a UK surgical training centre.Participants11 consultant head and neck surgeons from the UK, mainland Europe and the USA took part in TORS procedures on six human cadavers.Interventions3 key index procedures were assessed that represent the core surgical workload of TORS: lateral oropharyngectomy, tongue base resection and partial supraglottic laryngectomy.Main outcome measuresThe primary outcome was the successful completion of each surgical procedure. Secondary outcomes included the optimisation of system setup, instrumentation and surgeon-reported outcomes for feasibility of each component procedural step.Results33 cadaveric procedures were performed and 32 were successfully completed. One supraglottic laryngectomy was not fully completed due to issues dividing the epiglottic cartilage with available instrumentation. Surgeon-reported outcomes met the minimal level of feasibility in all procedures and a consensus that it is feasible to perform TORS with Versius was reached. Available instrumentation was not representative of other robotic platforms used in TORS and further instrument optimisation is recommended before wider dissemination.ConclusionsIt is feasible to perform TORS with the Versius Surgical System (CMR Surgical) within a pre-clinical cadaveric setting. Clinical evaluation is needed and appropriate with the system. Further instrument development and optimisation is desirable.
Should Pulsatile Preservation Be the Gold Standard in Kidney Transplantation?
In recent years, dramatic improvements in kidney transplantation, together with a rising incidence of diseases such as diabetes, have led to an increasing demand for deceased donor kidneys for transplantation. Hence, it has been necessary to expand the kidney donor pool by using organs once considered unsuitable for transplantation. These higher risk kidneys are typically from older donors with additional comorbidities and are more susceptible to injury. Therefore, the transplant community has been focusing efforts in trying to improve the outcomes of these high-risk organs. Preservation by pulsatile machine perfusion has been associated with decreased risk of delayed graft function and renoprotective effects on deceased donor kidneys. The aim of this review is to provide an overview of the principles of this preservation technique and to review the evidence regarding its usage for deceased donor kidneys compared to standard static cold storage.
Development of ex situ normothermic reperfusion as an innovative method to assess pancreases after preservation.
Static cold storage (SCS) is the standard method for pancreas preservation prior to transplantation; however, it does not permit organ assessment. Normothermic reperfusion (NR) is utilized clinically for other organs to assess viability. Our aim was to develop NR using normothermic machine perfusion technique to simulate reperfusion at the time of transplantation, enabling evaluation of oxygenated hypothermic machine perfusion (HMPO2) as a newer strategy to optimize pancreas preservation. 13 porcine pancreases procured after circulatory death were divided into 3 groups: 4 pancreases preserved using SCS, and 2 groups preserved by HMPO2 (n = 4 and n = 5, differing by type of preservation solution). Duration of perfusion or cold storage was 6 hours before the 1-hour assessment using NR. Outcome measures were perfusion characteristics, biochemistry and change in tissue water mass as oedema assessment. During NR, the HMPO2 groups demonstrated better perfusion characteristics, normal macroscopic appearances, decreased water mass and one HMPO2 group demonstrated a response to glucose stimulation. Conversely, the SCS group showed an increased water mass and developed early macroscopic appearances of oedema, interstitial haemorrhage and minimal portal outflow. This study suggests that ex situ assessment of pancreases by NR is promising, and that HMPO2 may be better than SCS.
Encyclopedia of Inflammatory Diseases
Reactive oxygen species (ROS) is a collective term given to a group of oxygen-containing intermediates, many of which react with biomolecules such as DNA, lipids, or proteins. ROS include (but are not limited to) hydrogen peroxide (H2O2), the superoxide radical anion (O2˙−), the hydroxyl radical (˙OH), and singlet oxygen (1O2). ROS sometimes have one or more unpaired electrons – as denoted by a superscript dot “˙”.
Comparison of in-gel and in-solution proteolysis in the proteome profiling of organ perfusion solutions.
PURPOSE: The organ perfusion solution (perfusate), collected at clinically and temporally significant stages of the organ preservation and transplantation process, provides a valuable insight into the biological status of an organ over time and prior to reperfusion (transplantation) in the recipient. The objective of this study was to assess two bottom-up proteomics workflows for the extraction of tryptic peptides from the perfusate. EXPERIMENTAL DESIGN: Two different kinds of perfusate samples from kidney and liver trials were profiled using liquid chromatography-mass spectrometry (LC-MS/MS). The preparation of clean peptide mixtures for downstream analysis was performed considering different aspects of sample preparation; protein estimation, enrichment, in-gel and urea-based in-solution digestion. RESULTS: In-solution digestion of perfusate allowed identification of the highest number of peptides and proteins with greater sequence coverage and higher confidence data in kidney and liver perfusate. Key pathways identified by gene ontology analysis included complement, coagulation and antioxidant pathways, and a number of biomarkers previously linked to ischemia-reperfusion injury were also observed in perfusate. CONCLUSIONS: This study showed that in-solution digestion is a more efficient method for LC-MS/MS analysis of kidney and liver organ perfusion solutions. This method is also quicker and easier than in-gel digestion, allowing for greater sample throughput, with fewer opportunities for experimental error or peptide loss.
The Effect of Continuous Liver Normothermic Machine Perfusion on the Severity of Histological Bile Duct Injury.
Static Cold Storage (SCS) injures the bile duct, while the effect of Normothermic Machine Perfusion (NMP) is unknown. In a sub-study of the COPE trial on liver NMP, we investigated the impact of preservation type on histological bile duct injury score (BDIS). Transplants with at least one bile duct biopsy, either at end of preservation or 1 h post-reperfusion, were considered. BDIS was determined by assessing peribiliary glands injury, stromal and mural loss, haemorrhage, and thrombosis. A bivariate linear model compared BDIS (estimate, CI) between groups. Sixty-five transplants and 85 biopsies were analysed. Twenty-three grafts were preserved with SCS and 42 with NMP, with comparable baseline characteristics except for a shorter cold ischemic time in NMP. The BDIS increased over time regardless of preservation type (p = 0.04). The BDIS estimate was higher in NMP [8.02 (7.40-8.65)] than in SCS [5.39 (4.52-6.26), p < 0.0001] regardless of time. One patient in each group developed ischemic cholangiopathy, with a BDIS of 6 for the NMP-preserved liver. In six other NMP grafts, BDIS ranged 7-12 without development of ischemic cholangiopathy. In conclusion, BDIS increases over time, and the higher BDIS in NMP did not increase ischemic cholangiopathy. Thus, BDIS may overestimate this risk after liver NMP.