Die Bedeutung von Interferon alpha und Interferon gamma auf den Verlauf der Marekschen Krankheit beim Haushuhn

Marek´s disease virus (MDV) is a herpesvirus causing a lymphoproliferative disorder of domestic chickens worldwide. The early signs of disease like apathy, anorexy and the transient paralysis as well as the symptoms at the later stages (Neoplasia and death) causes large economical problems in the chicken industry. With the development of an effetive Vaccine, the first practical vaccine against an oncogenic disease of any type in any spieces, most of the serious problems were solved. However recent reports describing an increased virulence resulting in vaccine breaks, indicates a demand for better vaccines. To be able to enhance the prophylaxis against Marek´s disease virus, the understanding of the interaction between host and virus needs to be expanded. Interferons are cytokines with antiviral activity. On the basis of their structural and functional features they can be divided into type I (IFN-alpha and IFN-beta) and type II (IFN-gamma) interferons. Type I plays an important role during the early phase of viral infections, while IFN-gamma has a prominent role later during the infection through its immunmodulatory functions. In this study it was shown that chicken embryocellcultures responds to the infection with either an apathogenic strain of MDV (CVI 988) or a highly pathogenic strain (RB1B), with the secretion of an antiviral factor. Neutralisation with cytokine specific antibodies revealed that this activity could only be attributed to IFN-alpha but not IFN-beta. In addition, synthesis of IFN-gamma was not observed in any of the experiments. Treatment of MDV infected embryocellcultures with either recombinant chicken IFN-alpha (rChIFN-alpha) or recombinant chicken IFN-gamma (rChIFN-gamma) led to a significant reduction in plaque numbers. This antiviral effect was still observed when cultures were treated 48 hours after infection. In order to understand the functional role of the IFNs in vivo, leghorn birds were repeatedly treated with rChIFN-alpha or rChIFN-gamma for ten weeks starting on the day of hatch. The birds were then challanged on the second day of life with a highly virulent MDV-strain (vv+, EU-1). Treatment with IFN-gamma delayed the mortality to a very small degree. In contrast, IFN-alpha treatment delayed the mortality for 3 weeks. However, prevention of the disease was not achieved regardless of the used IFN-alpha dose. The functional relevance of IFN-alpha was further underscored by experiments where birds were treated with a neutralizing monoclonal antibody to ChIFN-alpha and thereafter infected with the EU-1 strain at the second day of life. Birds in this group died 3 weeks earlier than those in the untreated control group. These data shows that Interferons play a role in the progression of Mareks disease, however, regardless the IFN-dose applicated, the disease could not be prevented by the cytokines on their own.Die durch des Marek Disease Virus (MDV) ausgelöste Mareksche Krankheit ist eine der bedeutesten Krankheiten der Geflügelwirtschaft. Zu den frühen Krankheitserscheinungen gehören Apathie, Anorexie, und eine transiente Paralyse, im weiteren Verlauf der Erkrankung treten Tumoren auf. Die Krankheit ist durch eine hohe Mortalität gekennzeichnet. Die Impfprophylaxe stellt die wichtigste Bekämpfungsmaßnahme dar. Da zunehmend pathogenere Feldstämme auftreten kommt es aber immer wieder zu Impfdurchbrüchen. Um bessere Bekämpfungsmaßnahmen entwickeln zu können, sind detaillierte Kenntnisse der Wirt-Virus-Interaktion notwendig. Interferone (IFN) sind antiviral wirksame Botenstoffe des Immunsystems. Sie werden aufgrund ihrer Eigenschaften in Typ I (IFN-alpha und IFN-beta) und Typ II (IFN-gamma) Interferone unterteilt. Typ I IFNs spielen insbesondere in der frühen Phase einer Virusinfektion eine wichtige Rolle, während IFN-gamma als potenter Immunmodulator auch in späteren Phasen der Infektion von Bedeutung ist. In dieser Arbeit konnte in in vitro Versuchen gezeigt werden, dass Embryofibroblastenkulturen auf Infektionen mit dem Impfvirus CVI 988 oder dem hochpathogenen Virusisolat RB1B mit der Synthese eines antiviral wirksamen Faktors reagieren. Neutralisationsexperimente mit spezifischen Antikörpern zeigten, dass es sich dabei um IFN-alpha nicht aber IFN-beta handelte. Ebenso wenig konnte eine Synthese von IFN-gamma in diesen Kulturen nachgewiesen werden. Die Behandlung von infizierten Fibroblastenkulturen mit rekombinantem Hühner IFN- alpha (rChIFN-alpha) oder rekombinanten Hühner IFN-gamma (rChIFN-gamma) führte zu einer hochgradigen Reduktion der Plaquezahlen. Diese konnte auch dann beobachtet werden, wenn die Interferone erst 48 Stunden nach der Infektion der Kulturen zugesetzt wurden.Diese in vitro Studien wurden durch in vivo Versuche ergänzt, bei denen leghorn Hennen über einen Zeitraum von 10 Wochen wiederholt entweder mit rChIFN-alpha oder rChIFN-gamma behandelt und mit einem hochpathogenen MDV-1 Isolat (EU-1) infiziert wurden. Während die Behandlung mir rChIFN-gamma nur zu einer geringgradigen Verzögerung der Mortalität führte, konnte mit rChIFN-alpha eine deutliche Verzögerung erzielt werden. Einen Prävention der MD wurde aber in keinem Fall erzielt. Die funktionelle Bedeutun

Synthesis of IFN-β by Virus-Infected Chicken Embryo Cells Demonstrated with Specific Antisera and a New Bioassay

Transcripts of interferon-α(IFN-α) and IFN-β genes are present in virus-infected chicken cells, but because of a lack of appropriate assays and reagents, it was unclear if biologically active IFN-β is secreted. We have established a nonviral bioassay for the sensitive detection of chicken IFN (ChIFN). This assay is based on a quail cell line that carries a luciferase gene that is controlled by the IFN-responsive chicken Mx promoter. Luciferase activity was strongly stimulated when the indicator cells were incubated with ChIFN-α, ChIFN-β, or ChIFN-γ but not with chicken interleukin-1β (ChIL-1β). Unlike the classic antiviral assay that preferentially detects ChIFN-α, the Mx-luciferase assay detected ChIFN-α and ChIFN-β with similar sensitivity. With the help of this novel assay and with rabbit antisera specific for either IFN-α or IFN-β, we analyzed the composition of IFN in supernatants of virus-infected chicken embryo cells. Virtually all IFN produced in response to Newcastle disease virus (NDV) was IFN-α. However, IFN produced in response to influenza A or vaccinia virus (VV) was a mixture of usually more than 80% IFN-α and up to 20% IFN-β. Thus, IFN-α and IFN-β both contribute to the cytokine activity in supernatants of virus-infected chicken cells. Furthermore, the infecting virus appears to determine the IFN subtype composition

IFNα and IFNγ Impede Marek’s Disease Progression

Marek’s disease virus (MDV) is an alphaherpesvirus that causes Marek’s disease, a malignant lymphoproliferative disease of domestic chickens. While MDV vaccines protect animals from clinical disease, they do not provide sterilizing immunity and allow field strains to circulate and evolve in vaccinated flocks. Therefore, there is a need for improved vaccines and for a better understanding of innate and adaptive immune responses against MDV infections. Interferons (IFNs) play important roles in the innate immune defenses against viruses and induce upregulation of a cellular antiviral state. In this report, we quantified the potent antiviral effect of IFNα and IFNγ against MDV infections in vitro. Moreover, we demonstrate that both cytokines can delay Marek’s disease onset and progression in vivo. Additionally, blocking of endogenous IFNα using a specific monoclonal antibody, in turn, accelerated disease. In summary, our data reveal the effects of IFNα and IFNγ on MDV infection and improve our understanding of innate immune responses against this oncogenic virus

Design of TRUST, a non-interventional, multicenter, 3-year prospective study investigating an integrated patient management approach in patients with relapsing-remitting multiple sclerosis treated with natalizumab

Background: Natalizumab provides rapid and high-efficacy control of multiple sclerosis disease activity with long-term stabilization. However, the benefits of the drug are countered by a risk of developing progressive multifocal leukoencephalopathy in patients infected with the John Cunningham Virus. Close monitoring is required in patients with increased progressive multifocal leukoencephalopathy risk receiving natalizumab in the long-term for an optimal benefit-risk evaluation. Standardized high-quality monitoring procedures may provide a superior basis for individual benefit and risk evaluation and thus improve treatment decisions. The non-interventional study TRUST was designed to capture natalizumab effectiveness under real-life conditions and to examine alternate approaches for clinical assessments, magnetic resonance imaging monitoring and use of biomarkers for progressive multifocal leukoencephalopathy risk stratification. Methods/Design: TRUST is a non-interventional, multicenter, prospective cohort study conducted at approximately 200 German neurological centers. The study is intended to enroll 1260 relapsing-remitting multiple sclerosis patients with ongoing natalizumab therapy for at least 12 months. Patients will be followed for a period of 3 years, irrespective of treatment changes after study start. Data on clinical, subclinical and patient-centric outcomes will be documented in order to compare the effectiveness of continuous versus discontinued natalizumab treatment. Furthermore, the type and frequency of clinical, magnetic resonance imaging and biomarker assessments, reasons for continuation or discontinuation of therapy and the safety profile of natalizumab will be collected to explore the impact of a systematic patient management approach and its potential impact on patient outcome. Specifically, the role of biomarkers, the use of expert opinions, the impact of high-frequency magnetic resonance imaging assessment for early progressive multifocal leukoencephalopathy detection and the role of additional radiological and clinical expert advice will be explored. Discussion: TRUST was initiated in spring 2014 and enrollment is anticipated to be completed by mid 2016. Annual interim analyses will deliver continuous information and transparency with regard to the patient cohorts and the completeness and quality of data as well as closely monitor any safety signals in the natalizumab-treated cohort. The study’s results may provide insights into opportunities to improve the benefit-risk assessment in clinical practice and support treatment decisions

Känslighetsanalys av Flödeskommitténs riktlinjer för dimensionering av hel älv

En känslighetsanalys av Flödeskommittens riktlinjer för beräkning av dimensionerande flöden tillämpade på flermagasinssystemet i Ljusnan redovisas. Simuleringarna har gjorts med hjälp av ett modell.system baserat på HBV-modellen, som kompletterats med regleringsstrategier. Det 14 560 km 2 stora avrinningsområdet delades in i 16 delområden. Vid tre kraftverksdammar studerades känsligheten hos de dimensionerande vattennivåerna med hänsyn tagen till ändringar i de föreskrivna riktlinjerna, regleringsstrategier och modellparametrar. Förändringar i den dimensionerande nederbörden, snömagasinet eller utskovskapaciteten hade betydande inverkan på högsta vattenståndet i alla tre magasinen. Ändringar i regleringsstrategierna hade mindre effekt. Den högsta recessionsparametern i HBV-modellen, Ko, hade stor inverkan på de dimensionerande vattennivåerna. Efter det att magasinet fyllts, syntes ett klart samband mellan högsta tillrinning och maximal vattennivå. Studien visar, att det är svårt att förutbestämma den integrerade effekten av extrem nederbörd, snösmältning, markfuktighetstillstånd och reglering i ett system

Replication of Modified Vaccinia Virus Ankara in Primary Chicken Embryo Fibroblasts Requires Expression of the Interferon Resistance Gene E3L

Highly attenuated modified vaccinia virus Ankara (MVA) serves as a candidate vaccine to immunize against infectious diseases and cancer. MVA was randomly obtained by serial growth in cultures of chicken embryo fibroblasts (CEF), resulting in the loss of substantial genomic information including many genes regulating virus-host interactions. The vaccinia virus interferon (IFN) resistance gene E3L is among the few conserved open reading frames encoding viral immune defense proteins. To investigate the relevance of E3L in the MVA life cycle, we generated the deletion mutant MVA-ΔE3L. Surprisingly, we found that MVA-ΔE3L had lost the ability to grow in CEF, which is the first finding of a vaccinia virus host range phenotype in this otherwise highly permissive cell culture. Reinsertion of E3L led to the generation of revertant virus MVA-E3rev and rescued productive replication in CEF. Nonproductive infection of CEF with MVA-ΔE3L allowed viral DNA replication to occur but resulted in an abrupt inhibition of viral protein synthesis at late times. Under these nonpermissive conditions, CEF underwent apoptosis starting as early as 6 h after infection, as shown by DNA fragmentation, Hoechst staining, and caspase activation. Moreover, we detected high levels of active chicken alpha/beta IFN (IFN-α/β) in supernatants of MVA-ΔE3L-infected CEF, while moderate IFN quantities were found after MVA or MVA-E3rev infection and no IFN activity was present upon infection with wild-type vaccinia viruses. Interestingly, pretreatment of CEF with similar amounts of recombinant chicken IFN-α inhibited growth of vaccinia viruses, including MVA. We conclude that efficient propagation of MVA in CEF, the tissue culture system used for production of MVA-based vaccines, essentially requires conserved E3L gene function as an inhibitor of apoptosis and/or IFN induction

Melflufen plus dexamethasone (dex) in patients (pts) with relapsed/refractory multiple myeloma (RRMM) exposed/refractory to prior alkylators: A pooled analysis of the O-12-M1 and HORIZON studies.

International audience8048 Background: Melphalan flufenamide (melflufen) is a first-in-class peptide-drug conjugate (PDC) that leverages aminopeptidases and rapidly releases alkylating agents inside tumor cells. Melflufen has a mechanism of action distinct from other alkylating agents (Slipicevic et al. AACR 2020. Abs. 1843). In the O-12-M1 (NCT01897714) and HORIZON (OP-106; NCT02963493) studies, melflufen plus dex showed meaningful efficacy and a clinically manageable safety profile in pts with RRMM (Richardson et al. Lancet Haematol. 2020;7:5; Richardson et al. J Clin Oncol. 2020;Dec 9 [Epub]). This pooled analysis examines pts from these studies exposed to prior alkylators. Methods: Both the O-12-M1 and HORIZON studies included pts with RRMM who received ≥ 2 prior lines of therapy (LoTs) and had a primary endpoint of overall response rate (ORR). Secondary endpoints included progression-free survival (PFS) and safety. Data from the 2 studies were pooled and analyzed according to previous exposure and refractoriness to alkylators before study entry. Refractoriness to prior alkylator therapy was defined as disease that failed to achieve a minimal response or progressed while on therapy, or within 60 d of last therapy. Results: Of 202 pts (HORIZON: n = 157, cutoff January 14, 2020; O-12-M1: n = 45, cutoff October 29, 2019), 178 (88%) had been exposed to alkylators in ≥ 1 prior LoT (see Table for subgroups). Pts exposed and refractory to alkylators in ≥ 2 LoTs had the highest number of pts refractory to an alkylator in the last LoT (61%), and 82% were refractory to an alkylator within 12 mo of study entry. Meaningful response rates were seen in all subgroups, except for pts who were exposed and refractory to alkylators in ≥ 2 prior LoTs (see Table). PFS trended toward being shorter with higher exposure and refractoriness to prior alkylators. Results should be interpreted with caution due to limited pt numbers. Grade 3/4 adverse events (AEs) were similar between pts exposed to prior alkylators (O-12-M1: 85%; HORIZON: 89%) and the overall population (O-12-M1: 84%; HORIZON: 89%). The most common AEs were hematologic, but were mostly reversible and clinically manageable. Nonhematologic AEs were infrequent and primarily grade 1/2. Conclusions: Melflufen in combination with dex showed meaningful efficacy and a clinically manageable safety profile in pts with RRMM exposed/refractory to prior alkylators. Clinical trial information: NCT02963493 and NCT01897714. [Table: see text