Fargo, ND, United States
Fargo, ND, United States

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Grant
Agency: Cordis | Branch: FP7 | Program: CP-FP | Phase: HEALTH.2011.4.2-2 | Award Amount: 3.93M | Year: 2011

EpoCan aims to develop and implement a comprehensive interdisciplinary strategy to assess the long-term risks of erythropoietin (EPO) and its derivatives (epoetins) on tumour growth progression and thromboembolic events in cancer patients, cardiovascular events, and the development of cancer in chronic kidney disease. Approximately 400,000 patients across Europe receive epoetins treatment each year. Recent meta-analysis data have raised concerns over increased mortality in some patient groups. Hence the urgent need to evaluate the risk-benefit ratio of epoetin treatment and its potential long-term effects. EpoCan brings together a multidisciplinary consortium of 12 world leading academic, industrial and medical partners, with long-standing, complementary expertise in haemostasis, oncology and EPO biology. EpoCan aims to (1) Identify, detect and measure possible long-term hazards of epoetin treatment; (2) Develop novel prognostic tools and new complementary therapeutic reagents: (3) Evaluate the risk-benefit ratio to pave the way for new safety and efficacy criteria. EpoCan will: (a) Utilize a wide array of cellular models to thoroughly analyze EPO/EPO receptor(EPO-R)interaction and signalling, to define the relationship between EPO-R expression in tumour samples and the clinical outcome in cancer patients; (b) Establish and test new, personalized, predictive tools (EPO-R peptide antagonists, novel specific anti-EPO-R monoclonal antibodies, thromboembolic tests); (c) Create new murine models as hosts for tumour implantation subjected to EPO and derivatives established above; (d) Screen and analyze clinical databases; (e) Define models to predict hazardous versus safe/beneficial roles of epoetins in the treatment of cancer and kidney failure associated anaemia. Data obtained will be integrated into coherent models using novel computational algorithms developed for EpoCan. Results are expected to have broad ramifications, with special relevance for clinical oncology.


Zahid M.N.,French Institute of Health and Medical Research | Zahid M.N.,University of Strasbourg | Turek M.,French Institute of Health and Medical Research | Turek M.,University of Strasbourg | And 19 more authors.
Hepatology | Year: 2013

Scavenger receptor class B type I (SR-BI) is a high-density lipoprotein (HDL) receptor highly expressed in the liver and modulating HDL metabolism. Hepatitis C virus (HCV) is able to directly interact with SR-BI and requires this receptor to efficiently enter into hepatocytes to establish productive infection. A complex interplay between lipoproteins, SR-BI and HCV envelope glycoproteins has been reported to take place during this process. SR-BI has been demonstrated to act during binding and postbinding steps of HCV entry. Although the SR-BI determinants involved in HCV binding have been partially characterized, the postbinding function of SR-BI remains largely unknown. To uncover the mechanistic role of SR-BI in viral initiation and dissemination, we generated a novel class of anti-SR-BI monoclonal antibodies that interfere with postbinding steps during the HCV entry process without interfering with HCV particle binding to the target cell surface. Using the novel class of antibodies and cell lines expressing murine and human SR-BI, we demonstrate that the postbinding function of SR-BI is of key impact for both initiation of HCV infection and viral dissemination. Interestingly, this postbinding function of SR-BI appears to be unrelated to HDL interaction but to be directly linked to its lipid transfer function. Conclusion: Taken together, our results uncover a crucial role of the SR-BI postbinding function for initiation and maintenance of viral HCV infection that does not require receptor-E2/HDL interactions. The dissection of the molecular mechanisms of SR-BI-mediated HCV entry opens a novel perspective for the design of entry inhibitors interfering specifically with the proviral function of SR-BI. © 2012 American Association for the Study of Liver Diseases.


Kwilas S.,U.S. Army | Kishimori J.M.,U.S. Army | Josleyn M.,U.S. Army | Jerke K.,U.S. Army | And 3 more authors.
Current Gene Therapy | Year: 2014

Sin Nombre virus (SNV) and Andes virus (ANDV) cause most of the hantavirus pulmonary syndrome (HPS) cases in North and South America, respectively. The chances of a patient surviving HPS are only two in three. Previously, we demonstrated that SNV and ANDV DNA vaccines encoding the virus envelope glycoproteins elicit high-titer neutralizing antibodies in laboratory animals, and (for ANDV) in nonhuman primates (NHPs). In those studies, the vaccines were delivered by gene gun or muscle electroporation. Here, we tested whether a combined SNV/ANDV DNA vaccine (HPS DNA vaccine) could be delivered effectively using a disposable syringe jet injection (DSJI) system (PharmaJet, Inc). PharmaJet intramuscular (IM) and intradermal (ID) needle-free devices are FDA 510(k)-cleared, simple to use, and do not require electricity or pressurized gas. First, we tested the SNV DNA vaccine delivered by PharmaJet IM or ID devices in rabbits and NHPs. Both IM and ID devices produced high-titer anti-SNV neutralizing antibody responses in rabbits and NHPs. However, the ID device required at least two vaccinations in NHP to detect neutralizing antibodies in most animals, whereas all animals vaccinated once with the IM device seroconverted. Because the IM device was more effective in NHP, the Stratis® (PharmaJet IM device) was selected for follow-up studies. We evaluated the HPS DNA vaccine delivered using Stratis® and found that it produced high-titer anti-SNV and anti-ANDV neutralizing antibodies in rabbits (n=8/group) as measured by a classic plaque reduction neutralization test and a new pseudovirion neutralization assay. We were interested in determining if the differences between DSJI delivery (e.g., high-velocity liquid penetration through tissue) and other methods of vaccine injection, such as needle/syringe, might result in a more immunogenic DNA vaccine. To accomplish this, we compared the HPS DNA vaccine delivered by DSJI versus needle/syringe in NHPs (n=8/group). We found that both the anti-SNV and anti-ANDV neutralizing antibody titers were significantly higher (p-value 0.0115) in the DSJI-vaccinated groups than the needle/syringe group. For example, the anti-SNV and anti-ANDV PRNT50 geometric mean titers (GMTs) were 1,974 and 349 in the DSJI-vaccinated group versus 87 and 42 in the needle/syringe group. These data demonstrate, for the first time, that a spring-powered DSJI device is capable of effectively delivering a DNA vaccine to NHPs. Whether this HPS DNA vaccine, or any DNA vaccine, delivered by spring-powered DSJI will elicit a strong immune response in humans, requires clinical trials. © 2014 Bentham Science Publishers.


Hooper J.W.,U.S. Army | Brocato R.L.,U.S. Army | Kwilas S.A.,U.S. Army | Hammerbeck C.D.,U.S. Army | And 7 more authors.
Science Translational Medicine | Year: 2014

Polyclonal immunoglobulin-based medical products have been used successfully to treat diseases caused by viruses for more than a century. We demonstrate the use of DNA vaccine technology and transchromosomal bovines (TcBs) to produce fully human polyclonal immunoglobulins (IgG) with potent antiviral neutralizing activity. Specifically, two hantavirus DNA vaccines [Andes virus (ANDV) DNA vaccine and Sin Nombre virus (SNV) DNA vaccine] were used to produce a candidate immunoglobulin product for the prevention and treatment of hantavirus pulmonary syndrome (HPS). A needle-free jet injection device was used to vaccinate TcB, and hightiter neutralizing antibodies (titers >1000) against both viruses were produced within 1 month. Plasma collected at day 10 after the fourth vaccination was used to produce purified a α-HPS TcB human IgG. Treatment with 20,000 neutralizing antibody units (NAU)/kg starting 5 days after challenge with ANDV protected seven of eight animals, whereas zero of eight animals treated with the same dose of normal TcB human IgG survived. Likewise, treatment with 20,000 NAU/kg starting 5 days after challenge with SNV protected immunocompromised hamsters from lethal HPS, protecting five of eight animals. Our findings that the a-HPS TcB human IgG is capable of protecting in animal models of lethal HPS when administered after exposure provides proof of concept that this approach can be used to develop candidate next-generation polyclonal immunoglobulinbased medical products without the need for human donors, despeciation protocols, or inactivated/attenuated vaccine antigen. Copyright 2014 by the American Association for the Advancement of Science.


Sin Nombre virus (SNV; family Bunyaviridae, genus Hantavirus) causes a hemorrhagic fever known as hantavirus pulmonary syndrome (HPS) in North America. There have been approximately 200 fatal cases of HPS in the United States since 1993, predominantly in healthy working-age males (case fatality rate 35%). There are no FDA-approved vaccines or drugs to prevent or treat HPS. Previously, we reported that hantavirus vaccines based on the full-length M gene segment of Andes virus (ANDV) for HPS in South America, and Hantaan virus (HTNV) and Puumala virus (PUUV) for hemorrhagic fever with renal syndrome (HFRS) in Eurasia, all elicited high-titer neutralizing antibodies in animal models. HFRS is more prevalent than HPS (>20,000 cases per year) but less pathogenic (case fatality rate 1-15%). Here, we report the construction and testing of a SNV full-length M gene-based DNA vaccine to prevent HPS. Rabbits vaccinated with the SNV DNA vaccine by muscle electroporation (mEP) developed high titers of neutralizing antibodies. Furthermore, hamsters vaccinated three times with the SNV DNA vaccine using a gene gun were completely protected against SNV infection. This is the first vaccine of any kind that specifically elicits high-titer neutralizing antibodies against SNV. To test the possibility of producing a pan-hantavirus vaccine, rabbits were vaccinated by mEP with an HPS mix (ANDV and SNV plasmids), or HFRS mix (HTNV and PUUV plasmids), or HPS/HFRS mix (all four plasmids). The HPS mix and HFRS mix elicited neutralizing antibodies predominantly against ANDV/SNV and HTNV/PUUV, respectively. Furthermore, the HPS/HFRS mix elicited neutralizing antibodies against all four viruses. These findings demonstrate a pan-hantavirus vaccine using a mixed-plasmid DNA vaccine approach is feasible and warrants further development. © 2013 The Authors.


Brocato R.,U.S. Army | Josleyn M.,U.S. Army | Ballantyne J.,Aldevron | Vial P.,University for Development | Hooper J.W.,U.S. Army
PLoS ONE | Year: 2012

Andes virus (ANDV) is the predominant cause of hantavirus pulmonary syndrome (HPS) in South America and the only hantavirus known to be transmitted person-to-person. There are no vaccines, prophylactics, or therapeutics to prevent or treat this highly pathogenic disease (case-fatality 35-40%). Infection of Syrian hamsters with ANDV results in a disease that closely mimics human HPS in incubation time, symptoms of respiratory distress, and disease pathology. Here, we evaluated the feasibility of two postexposure prophylaxis strategies in the ANDV/hamster lethal disease model. First, we evaluated a natural product, human polyclonal antibody, obtained as fresh frozen plasma (FFP) from a HPS survivor. Second, we used DNA vaccine technology to manufacture a polyclonal immunoglobulin-based product that could be purified from the eggs of vaccinated ducks (Anas platyrhynchos). The natural "despeciation" of the duck IgY (i.e., Fc removed) results in an immunoglobulin predicted to be minimally reactogenic in humans. Administration of ≥5,000 neutralizing antibody units (NAU)/kg of FFP-protected hamsters from lethal disease when given up to 8 days after intranasal ANDV challenge. IgY/IgYΔFc antibodies purified from the eggs of DNA-vaccinated ducks effectively neutralized ANDV in vitro as measured by plaque reduction neutralization tests (PRNT). Administration of 12,000 NAU/kg of duck egg-derived IgY/IgYΔFc protected hamsters when administered up to 8 days after intranasal challenge and 5 days after intramuscular challenge. These experiments demonstrate that convalescent FFP shows promise as a postexposure HPS prophylactic. Moreover, these data demonstrate the feasibility of using DNA vaccine technology coupled with the duck/egg system to manufacture a product that could supplement or replace FFP. The DNA vaccine-duck/egg system can be scaled as needed and obviates the necessity of using limited blood products obtained from a small number of HPS survivors. This is the first report demonstrating the in vivo efficacy of any antiviral product produced using DNA vaccine-duck/egg system.


Grant
Agency: Department of Defense | Branch: Army | Program: STTR | Phase: Phase I | Award Amount: 100.00K | Year: 2013

In this Phase I STTR proposal we will determine the ruggedness of the genetic immunization technique in the production of duck egg derived immunoglobulin; specifically the natural F(ab")2 analog, IgY^Fc. We have already demonstrated that potently neutralizing duck egg antibodies generated with an early candidate Andes virus DNA vaccine, delivered via intramuscular electroporation, can protect after lethal challenge. We now seek to optimize the elements necessary to deploy a commercially viable platform system for the rapid production of passive immunity products as countermeasures to emerging viral threats. Mammalian or avian (Mallard duck) codon optimized Andes virus DNA vaccine variants of the original will be administered to ducks utilizing electroporation or a needle-free device. An increased potency and/or response frequency along with a decreased response time would represent a significant and enabling progression in the field. The work will be coupled with a small parallel study in sheep with Andes and Junin virus DNA vaccine candidates. Endpoints as measures of success will be determined by known correlates of protection using plaque reduction neutralizing tests and pseudoviral assays. Simple reactogenicity studies will also be performed with all full-length and despeciated immunoglobulin"s and the results compared to licensed polyclonal and monoclonal products.


PubMed | U.S. Army, Aldevron and Rega Institute for Medical Research
Type: Journal Article | Journal: Journal of virology | Year: 2016

Several members of the Arenaviridae can cause acute febrile diseases in humans, often resulting in lethality. The use of convalescent-phase human plasma is an effective treatment in humans infected with arenaviruses, particularly species found in South America. Despite this, little work has focused on developing potent and defined immunotherapeutics against arenaviruses. In the present study, we produced arenavirus neutralizing antibodies by DNA vaccination of rabbits with plasmids encoding the full-length glycoprotein precursors of Junn virus (JUNV), Machupo virus (MACV), and Guanarito virus (GTOV). Geometric mean neutralizing antibody titers, as measured by the 50% plaque reduction neutralization test (PRNT(50)), exceeded 5,000 against homologous viruses. Antisera against each targeted virus exhibited limited cross-species binding and, to a lesser extent, cross-neutralization. Anti-JUNV glycoprotein rabbit antiserum protected Hartley guinea pigs from lethal intraperitoneal infection with JUNV strain Romero when the antiserum was administered 2 days after challenge and provided some protection (30%) when administered 4 days after challenge. Treatment starting on day 6 did not protect animals. We further formulated an IgG antibody cocktail by combining anti-JUNV, -MACV, and -GTOV antibodies produced in DNA-vaccinated rabbits. This cocktail protected 100% of guinea pigs against JUNV and GTOV lethal disease. We then expanded on this cocktail approach by simultaneously vaccinating rabbits with a combination of plasmids encoding glycoproteins from JUNV, MACV, GTOV, and Sabia virus (SABV). Sera collected from rabbits vaccinated with the combination vaccine neutralized all four targets. These findings support the concept of using a DNA vaccine approach to generate a potent pan-arenavirus immunotherapeutic.Arenaviruses are an important family of emerging viruses. In infected humans, convalescent-phase plasma containing neutralizing antibodies can mitigate the severity of disease caused by arenaviruses, particularly species found in South America. Because of variations in potency of the human-derived product, limited availability, and safety concerns, this treatment option has essentially been abandoned. Accordingly, despite this approach being an effective postinfection treatment option, research on novel approaches to produce potent polyclonal antibody-based therapies have been deficient. Here we show that DNA-based vaccine technology can be used to make potently neutralizing antibodies in rabbits that exclusively target the glycoproteins of several human-pathogenic arenaviruses found in South America, including JUNV, MACV, GTOV, and SABV. These antibodies protected guinea pigs from lethal disease when given post-virus challenge. We also generated a purified antibody cocktail with antibodies targeting three arenaviruses and demonstrated protective efficacy against all three targets. Our findings demonstrate that use of the DNA vaccine technology could be used to produce candidate antiarenavirus neutralizing antibody-based products.


Grant
Agency: Department of Defense | Branch: Army | Program: STTR | Phase: Phase II | Award Amount: 725.55K | Year: 2013

In this Phase II STTR proposal we will demonstrate the scalability and ruggedness of the production system necessary for the commercialization of the goose egg-derived immunoglobulin product that is a countermeasure to the lethal (40 %) Hantavirus Pulmonary Syndrome caused by Andes Virus (ANDV). A major objective will be creating a Quality System (QS) around the global process that aligns it with standards commensurate with FDA guidance. Related objectives will be the development of critical assays and the harmonization of the related Standard Operating Procedures into the QS. Pharmacokinetic and repeat dose studies will be performed that will allow us to better evaluate the quantities to be used and routes of delivery for the anti-ANDV product in the challenge study. Endpoints as measures of success would be the reproducible production of 5 g lots of low impurity anti-ANDV and the use of such lots affording clinically significant protection up to five days after challenge. These results would indicate the commercial viability of a platform that is based on the use of genetic immunization in geese as a means of creating therapies for Category-A pathogens for which no countermeasures or vaccines exist.

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