Two patients with history of STEC-HUS, posttransplant recurrence and complement gene mutations

Hemolytic uremic syndrome (HUS) is a disease of microangiopathic hemolytic anemia, thrombocytopenia and acute renal failure. About 90% of cases are secondary to infections by Escherichia coli strains producing Shiga-like toxins (STEC-HUS), while 10% are associated with mutations in genes encoding proteins of complement system (aHUS). We describe two patients with a clinical history of STEC-HUS, who developed end-stage renal disease (ESRD) soon after disease onset. They received a kidney transplant but lost the graft for HUS recurrence, a complication more commonly observed in aHUS. Before planning a second renal transplantation, the two patients underwent genetic screening for aHUS-associated mutations that revealed the presence of a heterozygous CFI mutation in patient #1 and a heterozygous MCP mutation in patient #2, and also in her mother who donated the kidney. This finding argues that the two cases originally diagnosed as STEC-HUS had indeed aHUS triggered by STEC infection on a genetic background of impaired complement regulation. Complement gene sequencing should be performed before kidney transplantation in patients who developed ESRD following STEC-HUS since they may be undiagnosed cases of aHUS, at risk of posttransplant recurrence. Furthermore, genetic analysis of donors is mandatory before living-related transplantation to exclude carriers of HUS-predisposing mutations. Two patients with a clinical history of D+ hemolytic uremic syndrome associated with Shiga-toxin-producing 0157:H7 E. coli and recurrence in the kidney graft carry heterozygous mutations in the genes encoding complement factor I (patient 1) and membrane cofactor protein (patient 2). © Copyright 2013 The American Society of Transplantation and the American Society of Transplant Surgeons

Methylphenidate Analogues as a New Class of Potential Disease-Modifying Agents for Parkinson’s Disease: Evidence from Cell Models and Alpha-Synuclein Transgenic Mice

Parkinson’s disease (PD) is characterized by dopaminergic nigrostriatal neurons degeneration and Lewy body pathology, mainly composed of α-synuclein (αSyn) fibrillary aggregates. We recently described that the neuronal phosphoprotein Synapsin III (Syn III) participates in αSyn pathology in PD brains and is a permissive factor for αSyn aggregation. Moreover, we reported that the gene silencing of Syn III in a human αSyn transgenic (tg) mouse model of PD at a pathological stage, manifesting marked insoluble αSyn deposits and dopaminergic striatal synaptic dysfunction, could reduce αSyn aggregates, restore synaptic functions and motor activities and exert neuroprotective effects. Interestingly, we also described that the monoamine reuptake inhibitor methylphenidate (MPH) can recover the motor activity of human αSyn tg mice through a dopamine (DA) transporter-independent mechanism, which relies on the re-establishment of the functional interaction between Syn III and α-helical αSyn. These findings support that the pathological αSyn/Syn III interaction may constitute a therapeutic target for PD. Here, we studied MPH and some of its analogues as modulators of the pathological αSyn/Syn III interaction. We identified 4-methyl derivative I-threo as a lead candidate modulating αSyn/Syn III interaction and having the ability to reduce αSyn aggregation in vitro and to restore the motility of αSyn tg mice in vivo more efficiently than MPH. Our results support that MPH derivatives may represent a novel class of αSyn clearing agents for PD therapy

Alpha-synuclein/synapsin III pathological interplay boosts the motor response to methylphenidate

16siLoss of dopaminergic nigrostriatal neurons and fibrillary α-synuclein (α-syn) aggregation in Lewy bodies (LB) characterize Parkinson's disease (PD). We recently found that Synapsin III (Syn III), a phosphoprotein regulating dopamine (DA) release with α-syn, is another key component of LB fibrils in the brain of PD patients and acts as a crucial mediator of α-syn aggregation and toxicity. Methylphenidate (MPH), a monoamine reuptake inhibitor (MRI) efficiently counteracting freezing of gait in advanced PD patients, can bind α-syn and controls α-syn-mediated DA overflow and presynaptic compartmentalization. Interestingly, MPH results also efficient for the treatment of attention deficits and hyperactivity disorder (ADHD), a neurodevelopmental psychiatric syndrome associated with Syn III and α-syn polymorphisms and constituting a risk factor for the development of LB disorders. Here, we studied α-syn/Syn III co-deposition and longitudinal changes of α-syn, Syn III and DA transporter (DAT) striatal levels in nigrostriatal neurons of a PD model, the human C-terminally truncated (1−120) α-syn transgenic (SYN120 tg) mouse, in comparison with C57BL/6J wild type (wt) and C57BL/6JOlaHsd α-syn null littermates. Then, we analyzed the locomotor response of these animals to an acute administration of MPH (d-threo) and other MRIs: cocaine, that we previously found to stimulate Syn III-reliant DA release in the absence of α-syn, or the selective DAT blocker GBR-12935, along aging. Finally, we assessed whether these drugs modulate α-syn/Syn III interaction by fluorescence resonance energy transfer (FRET) and performed in silico studies engendering a heuristic model of the α-syn conformations stabilized upon MPH binding. We found that only MPH was able to over-stimulate a Syn III-dependent/DAT-independent locomotor activity in the aged SYN120 tg mice showing α-syn/Syn III co-aggregates. MPH enhanced full length (fl) α-syn/Syn III and even more (1–120) α-syn/Syn III interaction in cells exhibiting α-syn/Syn III inclusions. Moreover, in silico studies confirmed that MPH may reduce α-syn fibrillation by stabilizing a protein conformation with increased lipid binding predisposition. Our observations indicate that the motor-stimulating effect of MPH can be positively fostered in the presence of α-syn/Syn III co-aggregation. This evidence holds significant implications for PD and ADHD therapeutic management.openopenFaustini G.; Longhena F.; Bruno A.; Bono F.; Grigoletto J.; La Via L.; Barbon A.; Casiraghi A.; Straniero V.; Valoti E.; Costantino G.; Benfenati F.; Missale C.; Pizzi M.; Spillantini M.G.; Bellucci A.Faustini, G.; Longhena, F.; Bruno, A.; Bono, F.; Grigoletto, J.; La Via, L.; Barbon, A.; Casiraghi, A.; Straniero, V.; Valoti, E.; Costantino, G.; Benfenati, F.; Missale, C.; Pizzi, M.; Spillantini, M. G.; Bellucci, A

Impact of the COVID-19 outbreak on severe trauma trends and healthcare system reassessment in Lombardia, Italy: an analysis from the regional trauma registry

Backgrounds: The COVID-19 pandemic drastically strained the health systems worldwide, obligating the reassessment of how healthcare is delivered. In Lombardia, Italy, a Regional Emergency Committee (REC) was established and the regional health system reorganized, with only three hospitals designated as hubs for trauma care. The aim of this study was to evaluate the effects of this reorganization of regional care, comparing the distribution of patients before and during the COVID-19 outbreak and to describe changes in the epidemiology of severe trauma among the two periods. Methods: A cohort study was conducted using retrospectively collected data from the Regional Trauma Registry of Lombardia (LTR). We compared the data of trauma patients admitted to three hub hospitals before the COVID-19 outbreak (September 1 to November 19, 2019) with those recorded during the pandemic (February 21 to May 10, 2020) in the same hospitals. Demographic data, level of pre-hospital care (Advanced Life Support-ALS, Basic Life Support-BLS), type of transportation, mechanism of injury (MOI), abbreviated injury score (AIS, 1998 version), injury severity score (ISS), revised trauma score (RTS), and ICU admission and survival outcome of all the patients admitted to the three trauma centers designed as hubs, were reviewed. Screening for COVID-19 was performed with nasopharyngeal swabs, chest ultrasound, and/or computed tomography. Results: During the COVID-19 pandemic, trauma patients admitted to the hubs increased (46.4% vs 28.3%, p < 0.001) with an increase in pre-hospital time (71.8 vs 61.3 min, p < 0.01), while observed in hospital mortality was unaffected. TRISS, ISS, AIS, and ICU admission were similar in both periods. During the COVID-19 outbreak, we observed substantial changes in MOI of severe trauma patients admitted to three hubs, with increases of unintentional (31.9% vs 18.5%, p < 0.05) and intentional falls (8.4% vs 1.2%, p < 0.05), whereas the pandemic restrictions reduced road- related injuries (35.6% vs 60%, p < 0.05). Deaths on scene were significantly increased (17.7% vs 6.8%, p < 0.001). Conclusions: The COVID-19 outbreak affected the epidemiology of severe trauma patients. An increase in trauma patient admissions to a few designated facilities with high level of care obtained satisfactory results, while COVID-19 patients overwhelmed resources of most other hospitals

ZNF93 Increases Resistance to ET-743 (Trabectedin; Yondelis®) and PM00104 (Zalypsis®) in Human Cancer Cell Lines

ET-743 (trabectedin, Yondelis) and PM00104 (Zalypsis) are marine derived compounds that have antitumor activity. ET-743 and PM00104 exposure over sustained periods of treatment will result in the development of drug resistance, but the mechanisms which lead to resistance are not yet understood.Human chondrosarcoma cell lines resistant to ET-743 (CS-1/ER) or PM00104 (CS-1/PR) were established in this study. The CS-1/ER and CS-1/PR exhibited cross resistance to cisplatin and methotrexate but not to doxorubicin. Human Affymetrix Gene Chip arrays were used to examine relative gene expression in these cell lines. We found that a large number of genes have altered expression levels in CS-1/ER and CS-1/PR when compared to the parental cell line. 595 CS-1/ER and 498 CS-1/PR genes were identified as overexpressing; 856 CS-1/ER and 874 CS-1/PR transcripts were identified as underexpressing. Three zinc finger protein (ZNF) genes were on the top 10 overexpressed genes list. These genes have not been previously associated with drug resistance in tumor cells. Differential expressions of ZNF93 and ZNF43 genes were confirmed in both CS-1/ER and CS-1/PR resistant cell lines by real-time RT-PCR. ZNF93 was overexpressed in two ET-743 resistant Ewing sarcoma cell lines as well as in a cisplatin resistant ovarian cancer cell line, but was not overexpressed in paclitaxel resistant cell lines. ZNF93 knockdown by siRNA in CS-1/ER and CS-1/PR caused increased sensitivity for ET-743, PM00104, and cisplatin. Furthermore, ZNF93 transfected CS-1 cells are relatively resistant to ET-743, PM00104 and cisplatin.This study suggests that zinc finger proteins, and ZNF93 in particular, are involved in resistance to ET-743 and PM00104

A New Strategy for Glioblastoma Treatment: In Vitro and In Vivo Preclinical Characterization of Si306, a Pyrazolo[3,4-d]Pyrimidine Dual Src/P-Glycoprotein Inhibitor

20siopenOverexpression of P-glycoprotein (P-gp) and other ATP-binding cassette (ABC) transporters in multidrug resistant (MDR) cancer cells is responsible for the reduction of intracellular drug accumulation, thus decreasing the efficacy of chemotherapeutics. P-gp is also found at endothelial cells' membrane of the blood-brain barrier, where it limits drug delivery to central nervous system (CNS) tumors. We have previously developed a set of pyrazolo[3,4-d]pyrimidines and their prodrugs as novel Src tyrosine kinase inhibitors (TKIs), showing a significant activity against CNS tumors in in vivo. Here we investigated the interaction of the most promising pair of drug/prodrug with P-gp at the cellular level. The tested compounds were found to increase the intracellular accumulation of Rho 123, and to enhance the efficacy of paclitaxel in P-gp overexpressing cells. Encouraging pharmacokinetics properties and tolerability in vivo were also observed. Our findings revealed a novel role of pyrazolo[3,4-d]pyrimidines which may be useful for developing a new effective therapy in MDR cancer treatment, particularly against glioblastoma.openFallacara, Anna Lucia; Zamperini, Claudio; Podolski-Renić, Ana; Dinić, Jelena; Stanković, Tijana; Stepanović, Marija; Mancini, Arianna; Rango, Enrico; Iovenitti, Giulia; Molinari, Alessio; Bugli, Francesca; Sanguinetti, Maurizio; Torelli, Riccardo; Martini, Maurizio; Maccari, Laura; Valoti, Massimo; Dreassi, Elena; Botta, Maurizio; Pešić, Milica; Schenone, SilviaFallacara, Anna Lucia; Zamperini, Claudio; Podolski-Renić, Ana; Dinić, Jelena; Stanković, Tijana; Stepanović, Marija; Mancini, Arianna; Rango, Enrico; Iovenitti, Giulia; Molinari, Alessio; Bugli, Francesca; Sanguinetti, Maurizio; Torelli, Riccardo; Martini, Maurizio; Maccari, Laura; Valoti, Massimo; Dreassi, Elena; Botta, Maurizio; Pešić, Milica; Schenone, Silvi

A Phase II study of trabectedin single agent in patients with recurrent ovarian cancer previously treated with platinum-based regimens

The objective of this study was to determine the objective response rate in patients with platinum-sensitive and platinum-resistant recurrent ovarian cancer to treatment with trabectedin (Yondelis®) administered as a 3-h infusion weekly for 3 weeks of a 4-week cycle. We carried out a multicentre Phase II trial of trabectedin in patients with advanced recurrent ovarian cancer. Trabectedin (0.58 mg m−2) was administered via a central line, after premedication with dexamethasone, to 147 patients as a 3-h infusion weekly for 3 weeks followed by 1-week rest. Major eligibility criteria included measurable relapsed advanced ovarian cancer and not more than two prior platinum-containing regimens. Patients were stratified according to the treatment-free interval (TFI) between having either platinum-sensitive (⩾6 months TFI) or platinum-resistant disease (<6 months TFI)/platinum-refractory disease (progression during first line therapy). In the platinum-sensitive cohort, 62 evaluable patients with measurable disease had an overall response rate (ORR) of 29.0% (95% CI: 18.2–41.9%) and median progression-free survival (PFS) was 5.1 months (95% CI: 2.8–6.2). Four patients with measurable disease per Response Evaluation Criteria in Solid Tumours (RECIST) criteria had no follow-up scans at the end of treatment. In the platinum-resistant/refractory cohort, 79 patients were evaluable with an ORR of 6.3% (95% CI: 2.1–14.2%). Median PFS was 2.0 months (95% CI: 1.7–3.5 months). Two patients with measurable disease per RECIST criteria had no follow-up scans at the end of treatment. The most frequent (⩾2% of patients) drug-related treatment-emergent grade 3/4 adverse events were reversible liver alanine transferase elevation (10%), neutropaenia (8%), nausea, vomiting, and fatigue (5% each). Trabectedin is an active treatment, with documented responses in patients with platinum sensitive advanced relapsed ovarian cancer, and has a manageable toxicity profile

Merging computational modelling and experimental metabolite identification for toxicity prediction

Xenobiotic metabolism has become of paramount significance in drug research, development, and therapy. Nowadays, medicinal chemists active in the various stages of drug discovery and development are aware that unfavourable metabolic outcomes (e.g., reactive metabolites, enzyme inhibition) may exclude promising candidates from further development due to later toxic effects. Computational evaluation and/or prediction of phase I and II xenobiotics biotransformations may be critical in predicting their pharmacological and toxicological consequences. The knowledge of metabolism pathways is essential in any drug discovery project in order to stabilise the clearance of compounds, and also to decrease or eliminate the formation of potentially toxic intermediates which can induce adverse drug reactions. Although simulation studies of xenobiotic–enzyme interactions can realistically predict phase I biotransformation, phase II reactions are still very difficult to predict due to high number of false positive results. However, when the generation of metabolites in silico is combined with experimental metabolite identification the false positive data can be removed. The paper present an innovative automatic procedure that merge in silico data with experimental data to produce relevant information in order to address ADR problems and poor PK properties