scholarly journals Monobac System–A Single Baculovirus for the Production of rAAV

2021 ◽  
Vol 9 (9) ◽  
pp. 1799
Author(s):  
Lionel Galibert ◽  
Aurélien Jacob ◽  
Adrien Savy ◽  
Yohann Dickx ◽  
Delphine Bonnin ◽  
...  
Keyword(s):  
Large Scale ◽  
Viral Vector ◽  
Cell System ◽  
Lower Amount ◽  
Efficient Production ◽  
Baculovirus Genome ◽  
Raav Vector ◽  
System A ◽  
Raav Production

Large-scale manufacturing of rAAV is a bottleneck for the development of genetic disease treatments. The baculovirus/Sf9 cell system underpins the first rAAV treatment approved by EMA and remains one of the most advanced platforms for rAAV manufacturing. Despite early successes, rAAV is still a complex biomaterial to produce. Efficient production of the recombinant viral vector requires that AAV replicase and capsid genes be co-located with the recombinant AAV genome. Here, we present the Monobac system, a singular, modified baculovirus genome that contains all of these functions. To assess the relative yields between the dual baculovirus and Monobac systems, we prepared each system with a transgene encoding γSGC and evaluated vectors’ potency in vivo. Our results show that rAAV production using the Monobac system not only yields higher titers of rAAV vector but also a lower amount of DNA contamination from baculovirus.

scholarly journals Integrase-Defective Lentiviral Vectors for Delivery of Monoclonal Antibodies against Influenza

Viruses ◽  
2020 ◽  
Vol 12 (12) ◽  
pp. 1460
Author(s):  
Zuleika Michelini ◽  
Judith M. Minkoff ◽  
Jianjun Yang ◽  
Donatella Negri ◽  
Andrea Cara ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Influenza A ◽  
Large Scale ◽  
Viral Vector ◽  
Direct Injection ◽  
Lentiviral Vectors ◽  
Attractive Alternative ◽  
Scale Production ◽  
Safety Features

Delivering rapid protection against infectious agents to non-immune populations is a formidable public health challenge. Although passive immunotherapy is a fast and effective method of protection, large-scale production and administration of monoclonal antibodies (mAbs) is expensive and unpractical. Viral vector-mediated delivery of mAbs offers an attractive alternative to their direct injection. Integrase-defective lentiviral vectors (IDLV) are advantageous for this purpose due to the absence of pre-existing anti-vector immunity and the safety features of non-integration and non-replication. We engineered IDLV to produce the humanized mAb VN04-2 (IDLV-VN04-2), which is broadly neutralizing against H5 influenza A virus (IAV), and tested the vectors’ ability to produce antibodies and protect from IAV in vivo. We found that IDLV-transduced cells produced functional VN04-2 mAbs in a time- and dose-dependent fashion. These mAbs specifically bind the hemagglutinin (HA), but not the nucleoprotein (NP) of IAV. VN04-2 mAbs were detected in the serum of mice at different times after intranasal (i.n.) or intramuscular (i.m.) administration of IDLV-VN04-2. Administration of IDLV-VN04-2 by the i.n. route provided rapid protection against lethal IAV challenge, although the protection did not persist at later time points. Our data suggest that administration of mAb-expressing IDLV may represent an effective strategy for rapid protection against infectious diseases.

scholarly journals Plant Milking Technology—An Innovative and Sustainable Process to Produce Highly Active Extracts from Plant Roots

Molecules ◽  
2020 ◽  
Vol 25 (18) ◽  
pp. 4162
Author(s):  
Hanane Chajra ◽  
Aleksander Salwinski ◽  
Agnès Guillaumin ◽  
Benoit Mignard ◽  
Paul Hannewald ◽  
...  
Keyword(s):  
Large Scale ◽  
Morus Alba ◽  
Bioactive Molecules ◽  
Plant Roots ◽  
Efficient Production ◽  
Root Extract ◽  
Nitrogen Deprivation ◽  
Active Ingredients ◽  
Prenylated Flavonoids

We have used an original technology (Plant Milking Technology) based on aeroponic cultivation of plants associated with the gentle recovery of active ingredients from roots. Extraction of bioactive molecules was achieved by soaking the roots, still attached to the living plants, into a nontoxic solvent for a 2 h period. This nondestructive recovery process allows using the same root biomass for successive harvesting dates, in a recyclable way. We have applied this technology to Morus alba L. (mulberry tree), an emblematic tree of the Traditional Chinese Medicine (TCM). Trees were aeroponically grown in large-scale devices (100 m2) and were submitted to nitrogen deprivation to increase the content in active molecules (prenylated flavonoids). The Plant Milking technology applied to Morus alba L. allowed to produce an extract enriched in prenylated compounds (18-fold increase when compared to commercial root extract). Prenylated flavonoids (moracenin A and B, kuwanon C, wittiorumin F, morusin) presented a high affinity for the aged-associated collagenase enzyme, which was confirmed by activity inhibition. In accordance, M. alba extract presents efficient properties to regulate the skin matrisome, which is critical during skin aging. The benefits have been especially confirmed in vivo on wrinkle reduction, in a clinical study that involved aged women. Plant Milking technology is an optimal solution to produce active ingredients from plant roots, including trees, that meet both customer expectations around sustainability, as well as the need for an efficient production system for biotechnologists.

Efficient large-scale production and concentration of HIV-1-based lentiviral vectors for use in vivo

2003 ◽  
Vol 12 (3) ◽  
pp. 221-228 ◽  
Author(s):  
Jason E. Coleman ◽  
Matthew J. Huentelman ◽  
Sergey Kasparov ◽  
Beverly L. Metcalfe ◽  
Julian F. R. Paton ◽  
...  
Keyword(s):  
Animal Studies ◽  
Large Scale ◽  
Lentiviral Vector ◽  
High Titer ◽  
Lentiviral Vectors ◽  
Efficient Production ◽  
Scale Production ◽  
Peripheral Tissues ◽  
Large Scale Production

The aim of this study was to develop an efficient method for packaging and concentrating lentiviral vectors that consistently yields high-titer virus on a scale suitable for in vivo applications. Transient cotransfection of 293T packaging cells with DNA plasmids encoding lentiviral vector components was optimized using SuperFect, an activated dendrimer-based transfection reagent. The use of SuperFect allowed reproducible and efficient production of high-titer lentiviral vector at concentrations greater than 1 × 107transducing units per ml (TU/ml) and required less than one-third of the total amount of DNA used in traditional calcium phosphate transfection methods. Viral titers were further increased using a novel concentration protocol that yielded an average final titer of 1.4 × 1010TU/ml. Lentiviruses produced using these methods exhibited efficient transduction of central nervous system and peripheral tissues in vivo. The method is reproducible and can be scaled up to facilitate the use of these vectors in animal studies.

1004: In-Vivo Transfection of the Rat Corpus Cavernosum Using a Non-Viral, Linear Polyethylenimine Gene Delivery System: A Novel Application

2006 ◽  
Vol 175 (4S) ◽  
pp. 323-324 ◽  
Author(s):  
Joseph Dall'era ◽  
Sweaty Koul ◽  
Jesse Mills ◽  
Jeremy Myers ◽  
Randall B. Meacham ◽  
...  
Keyword(s):  
Gene Delivery ◽  
Delivery System ◽  
Corpus Cavernosum ◽  
Gene Delivery System ◽  
System A ◽  
In Vivo Transfection ◽  
Linear Polyethylenimine

Purification of the Antihemophilic Factor by Gel Filtration on Agarose

1969 ◽  
Vol 22 (03) ◽  
pp. 577-583 ◽  
Author(s):  
M.M.P Paulssen ◽  
A.C.M.G.B Wouterlood ◽  
H.L.M.A Scheffers
Keyword(s):  
Factor Viii ◽  
Plasma Proteins ◽  
Large Scale ◽  
Gel Filtration ◽  
Large Scale Preparation ◽  
Scale Preparation ◽  
Antihemophilic Factor

SummaryFactor VIII can be isolated from plasma proteins, including fibrinogen by chromatography on agarose. The best results were obtained with Sepharose 6B. Large scale preparation is also possible when cryoprecipitate is separated by chromatography. In most fractions containing factor VIII a turbidity is observed which may be due to the presence of chylomicrons.The purified factor VIII was active in vivo as well as in vitro.

The Effect of Active Site-inhibited Factor VIIa on Tissue Factor-initiated Coagulation Using Platelets before and after Aspirin Administration

1997 ◽  
Vol 78 (04) ◽  
pp. 1202-1208 ◽  
Author(s):  
Marianne Kjalke ◽  
Julie A Oliver ◽  
Dougald M Monroe ◽  
Maureane Hoffman ◽  
Mirella Ezban ◽  
...  
Keyword(s):  
Tissue Factor ◽  
Active Site ◽  
Large Scale ◽  
Thrombin Generation ◽  
Factor Viia ◽  
Dependent Manner ◽  
Antithrombotic Agent ◽  
Before And After ◽  
Ic50 Values

SummaryActive site-inactivated factor VIIa has potential as an antithrombotic agent. The effects of D-Phe-L-Phe-L-Arg-chloromethyl ketone-treated factor VIla (FFR-FVIIa) were evaluated in a cell-based system mimicking in vivo initiation of coagulation. FFR-FVIIa inhibited platelet activation (as measured by expression of P-selectin) and subsequent large-scale thrombin generation in a dose-dependent manner with IC50 values of 1.4 ± 0.8 nM (n = 8) and 0.9 ± 0.7 nM (n = 7), respectively. Kd for factor VIIa binding to monocytes ki for FFR-FVIIa competing with factor VIIa were similar (11.4 ± 0.8 pM and 10.6 ± 1.1 pM, respectively), showing that FFR-FVIIa binds to tissue factor in the tenase complex with the same affinity as factor VIIa. Using platelets from volunteers before and after ingestion of aspirin (1.3 g), there were no significant differences in the IC50 values of FFR-FVIIa [after aspirin ingestion, the IC50 values were 1.7 ± 0.9 nM (n = 8) for P-selectin expression, p = 0.37, and 1.4 ± 1.3 nM (n = 7) for thrombin generation, p = 0.38]. This shows that aspirin treatment of platelets does not influence the inhibition of tissue factor-initiated coagulation by FFR-FVIIa, probably because thrombin activation of platelets is not entirely dependent upon expression of thromboxane A2.

Enhanced Stability and Controlled Delivery of MOF Encapsulated Vaccines and Their Immunogenic Response In Vivo

2018 ◽  
Author(s):  
Michael Luzuriaga ◽  
Raymond P. Welch ◽  
Madushani Dharmawardana ◽  
Candace Benjamin ◽  
Shaobo Li ◽  
...  
Keyword(s):  
Viral Vector ◽  
Controlled Delivery ◽  
Conformational Structure ◽  
Spectroscopy Analysis ◽  
Zeolitic Imidazolate Framework ◽  
Structural Conformation ◽  
Depth Study ◽  
Viral Nanoparticle

<div><div><div><p>Vaccines have an innate tendency to lose their structural conformation upon environmental and chemical stressors. A loss in conformation reduces the therapeutic ability to prevent the spread of a pathogen. Herein, we report an in-depth study of zeolitic imidazolate framework-8 (ZIF-8) and its ability to provide protection for a model viral vector against dena- turing conditions. The immunoassay and spectroscopy analysis together demonstrate enhanced thermal and chemical stability to the conformational structure of the encapsulated viral nanoparticle. The long-term biological activity of this virus-ZIF composite was investigated in animal models to further elucidate the integrity of the encapsulated virus, the bio-safety, and immunogenicity of the overall composite. Additionally, histological analysis found no observable tissue damage in the skin or vital organs in mice, following multiple subcutaneous administrations. This study shows that ZIF-based protein composites are strong candidates for improved preservation of proteinaceous drugs, are biocompatible, and capable of controlling the release and adsorption of drugs in vivo.</p></div></div></div>

Naphthoquinones and derivatives for chemotherapy: perspectives and limitations of their anti-trypanosomatids activities

2020 ◽  
Vol 26 ◽  
Author(s):  
Luíza Dantas-Pereira ◽  
Edézio F. Cunha-Junior ◽  
Valter V. Andrade-Neto ◽  
John F. Bower ◽  
Guilherme A. M. Jardim ◽  
...  
Keyword(s):  
Large Scale ◽  
Neglected Tropical Diseases ◽  
Folk Medicine ◽  
Leishmania Species ◽  
Parasitic Infections ◽  
Mechanistic Analysis ◽  
Leishmania Spp ◽  
Nanomolar Range

: Chagas disease, Sleeping sickness and Leishmaniasis, caused by trypanosomatids Trypanosoma cruzi, Trypanosoma brucei and Leishmania spp., respectively, are considered neglected tropical diseases, and they especially affect impoverished populations in the developing world. The available chemotherapies are very limited and a search for alternatives is still necessary. In folk medicine, natural naphthoquinones have been employed for the treatment of a great variety of illnesses, including parasitic infections. This review is focused on the anti-trypanosomatid activity and mechanistic analysis of naphthoquinones and derivatives. Among all the series of derivatives tested in vitro, naphthoquinone-derived 1,2,3-triazoles were very active on T. cruzi infective forms in blood bank conditions, as well as in amastigotes of Leishmania spp. naphthoquinones containing a CF3 on a phenyl amine ring inhibited T. brucei proliferation in the nanomolar range, and naphthopterocarpanquinones stood out for their activity on a range of Leishmania species. Some of these compounds showed a promising selectivity index (SI) (30 to 1900), supporting further analysis in animal models. Indeed, high toxicity to the host and inactivation by blood components are crucial obstacles to be overcome to use naphthoquinones and/or their derivatives for chemotherapy. Multidisciplinary initiatives embracing medicinal chemistry, bioinformatics, biochemistry, and molecular and cellular biology need to be encouraged to allow the optimization of these compounds. Large scale automated tests are pivotal for the efficiency of the screening step, and subsequent evaluation of both the mechanism of action in vitro and pharmacokinetics in vivo are essential for the development of a novel, specific and safe derivative, minimizing adverse effects.

Implantable neural interfaces

2018 ◽  
Author(s):  
Stefano Vassanelli
Keyword(s):  
Brain Function ◽  
Large Scale ◽  
Ionic Channels ◽  
Patch Clamp Technique ◽  
Advantages And Disadvantages ◽  
Optogenetic Stimulation ◽  
Physical Interfaces ◽  
The Brain

Establishing direct communication with the brain through physical interfaces is a fundamental strategy to investigate brain function. Starting with the patch-clamp technique in the seventies, neuroscience has moved from detailed characterization of ionic channels to the analysis of single neurons and, more recently, microcircuits in brain neuronal networks. Development of new biohybrid probes with electrodes for recording and stimulating neurons in the living animal is a natural consequence of this trend. The recent introduction of optogenetic stimulation and advanced high-resolution large-scale electrical recording approaches demonstrates this need. Brain implants for real-time neurophysiology are also opening new avenues for neuroprosthetics to restore brain function after injury or in neurological disorders. This chapter provides an overview on existing and emergent neurophysiology technologies with particular focus on those intended to interface neuronal microcircuits in vivo. Chemical, electrical, and optogenetic-based interfaces are presented, with an analysis of advantages and disadvantages of the different technical approaches.

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