Expression of functional recombinant human factor IX in milk of mice

Lisauskas, S.; Cunha, Nicolau Brito da; Vianna, Giovanni R.; Mendes, Érica Araújo; Ramos, Gustavo L.; Maranhão, Andréa Queiroz; Brígido, Marcelo M.; Almeida, Jussara O. S. C.; Baptista, Heloisa Allegro; Motta, Fabiana Louise; Pesquero, João Bosco; Aragão, F. J. L.; Rech, E. L.

doi:10.1007/s10529-008-9818-y

Cited by 10 publications

(5 citation statements)

References 21 publications

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“…Clinical use demonstrated the safety and haemostatic effect of rh‐FIX [35,36]. Recently, the FIX gene could be expressed in the mammary glands of transgenic mice [37]. Secretion of coagulant active FIX at 3% of the total soluble proteins has been achieved, opening up the prospect of commercial production in transgenic animals.…”

Section: Coagulation Factorsmentioning

confidence: 99%

Recombinant plasma proteins

Burnouf

2010

Vox Sanguinis

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For almost 50 years, the fractionation of human plasma has been the sole possible source of a wide range of therapeutic proteins--such as coagulation factors, anticoagulants, immunoglobulins, and albumin--essential to the treatment of serious congenital or acquired bleeding or immunological diseases. In the last 20 years, the application of recombinant technologies to mammalian cell cultures has made possible--although with some limitations in productivity, costs, and immunogenic risks--to produce and commercialize complex plasma glycoproteins for human therapeutic applications and has opened the way to the development of new molecular entities. Today, the advanced exploration of alternative cell lines and enhanced cell culture systems, as well as the development of alternative expression technologies, such as transgenic animals, is opening a new era in the production of the full range of recombinant plasma protein therapeutics. In this review, we examine the achievements and ongoing challenges of the recombinant DNA technology as a platform for the production of plasma proteins and the advantages and limitations of such products compared to fractionated plasma proteins.

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Section: Coagulation Factorsmentioning

confidence: 99%

Recombinant plasma proteins

Burnouf

2010

Vox Sanguinis

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“…Despite the efficacy of this approach, coagulation therapy based on purified hFIX from pooled plasma is closely associated with high costs and prion and virus contamination risks (Lisauskas et al 2008). In contrast, the adjusted continuous infusion of licensed recombinant hFIX (BeneFix) has proved to be inherently free of human blood-borne pathogens and efficacious in the treatment of Type B Christmas disease over the last 10 years (Lambert et al 2007).…”

Section: Introductionmentioning

confidence: 99%

“…We recently demonstrated the functional secretion of hFIX in the milk of transgenic mice, reaching levels of up to 3% of the total soluble protein (TSP; Lisauskas et al 2008). Despite the correct post-translational processing of the recombinant hFIX synthesised in bacteria and animal systems, the production of biopharmaceuticals in these platforms may incur significantly high overall costs that are due-at least partially-to a long production timescale, low scale-up capacity and expensive storage conditions (Kusnadi et al 1997;Ma et al 2003).…”

Section: Introductionmentioning

confidence: 99%

Accumulation of functional recombinant human coagulation factor IX in transgenic soybean seeds

et al. 2010

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The seed-based production of recombinant proteins is an efficient strategy to achieve the accumulation, correct folding, and increased stability of these recombinant proteins. Among potential plant molecular farming systems, soybean [Glycine max (L.) Merrill] is a viable option for the production of recombinant proteins due to its high protein content, known regulatory sequences, efficient gene transfer protocols, and a scalable production system under greenhouse conditions. We report here the expression and stable accumulation of human coagulation factor IX (hFIX) in transgenic soybean seeds. A biolistic process was utilised to co-introduce a plasmid carrying the hFIX gene under the transcriptional control of the α' subunit of a β-conglycinin seed-specific promoter and an α-Coixin signal peptide in soybean embryonic axes from mature seeds. The 56-kDa hFIX protein was expressed in the transgenic seeds at levels of up to 0.23% (0.8 g kg(-1) seed) of the total soluble seed protein as determined by an enzyme-linked immunosorbent assay (ELISA) and western blot. Ultrastructural immunocytochemistry assays indicated that the recombinant hFIX in seed cotyledonary cells was efficiently directed to protein storage vacuoles. Mass spectrometry characterisation confirmed the presence of the hFIX recombinant protein sequence. Protein extracts from transgenic seeds showed a blood-clotting activity of up to 1.4% of normal plasma. Our results demonstrate the correct processing and stable accumulation of functional hFIX in soybean seeds stored for 6 years under room temperature conditions (22 ± 2°C).

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“…Общепринятым вектором для создания животных-продуцентов в молоке рекомбинантного белка является pBC1 Milk Expression Vector Kit ("Invitrogen", США) на основе криптической плазмиды Bacillus subtilis [2]. С его помощью созданы конструкции для экспрессии рекомбинантных белков в молоке разных видов животных, как крупных сельскохозяйственных (козы [3,4], коровы [5] и свиньи [6]), так и лабораторных (мыши [7][8][9][10]).…”

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Multiplex PCR test-system development for the genotyping of transgenic mice with pBC1 MILK expression vector-based transgene

Kalmykov

Kusov

Deykin

et al. 2019

Доклады Академии наук

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Developed multiplex PCR test-system for identification of transgenic cis-regulatory elements (promoter, insulator) from genetic constructions based on pBC1 Milk expression vector incorporated into the Mus musculus genome. Test-system was validated by genotyping producent strains generated by linearized vector DNA microinjection into fertilized zygote with occasional transgene genome integration, expressing the following transgenes in mammary gland, in milk: Human lactoferrin protein in different length, heat-shock protein HSP-70, Firefly luciferase protein, lysozyme protein. Reaction conditions selection described in details for better reproducibility.

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Expression of functional recombinant human factor IX in milk of mice

Cited by 10 publications

References 21 publications

Recombinant plasma proteins

Recombinant plasma proteins

Accumulation of functional recombinant human coagulation factor IX in transgenic soybean seeds

Multiplex PCR test-system development for the genotyping of transgenic mice with pBC1 MILK expression vector-based transgene

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