Cell vector system

Data on the consistency of fermentation conditions and culture growth, and on the maintenance of product yield should be presented. Criteria for the rejection of culture lots should be established. The maximum number of cell doublings or passage levels to be permitted during production should be specified, based on information on the stability of the host-cell/vector system on serial subculture up to and beyond the level used in production. 

To circumvent this problem, vectors that are based on lentiviruses have been developed. In contrast to prototypic retroviruses, lentiviruses do not require cell division for integration. Gene-therapy vectors have been developed from a broad spectrum of lentiviruses including human immunodeficiency vims (HIV), simian and feline immunodeficiency vims as well as visna/maedi vims. The most widely used lentiviral vector system is based on HIV-1. These vectors can efficiently transduce a broad spectrum of dividing and nondividing cells including neurons, hepatocytes, muscle cells, and hematopoietic stem cells [1,2]. 

HIV phenotype A type of resistance testing for human immunodeficiency virus (HIV) in which a patient s blood sample is obtained, and the patient s HIV genes that encode for reverse transcriptase and protease are removed and placed in an HIV viral vector. This viral vector is replicated in a cell culture system with varying concentrations of antiretrovirals. A drug concentration-viral inhibition curve is developed and the concentration needed to inhibit 50% of the patient s virus is reported. This is used to predict resistance versus susceptibility. 

Philipps, B., Forstner, M. and Mayr, L.M. (2005) A baculovirus expression vector system for simultaneous protein expression in insect and mammalian cells. Biotechnology Progress, 21 (3), 708-711. 

A list of the various vectors capable of introducing genes into recipient cells has been provided in Table 14.2. These vectors are conveniently categorized as being viral-based or non-viral-based systems. The main vector systems developed, thus far, are discussed in somewhat more detail below. 

The second approach used in first-principles tribological simulations focuses on the behavior of the sheared fluid. That is, the walls are not considered and the system is treated as bulk fluid, as discussed. These simulations are typically performed using ab initio molecular dynamics (AIMD) with DFT and plane-wave basis sets. A general tribological AIMD simulation would be run as follows. A system representing the fluid would be placed in a simulation cell repeated periodically in all three directions. Shear or load is applied to the system using schemes such as that of Parrinello and Rahman, which was discussed above. In this approach, one defines a (potentially time-dependent) reference stress tensor aref and alters the nuclear and cell dynamics, such that the internal stress tensor crsys is equal to aref. When crsys = aref, the internal and external forces on the cell vectors balance, and the system is subject to the desired shear or load. 

The first two antibodies our group tried to express in this vector system within mammalian cells had mistakes in the genetic material that coded for variable regions and, consequently, did not express any antibody at all. The first antibody that was produced by recombinant technology at IDEC using this vector system was in early 1991. It was supposed to be a chimeric anti-CD4 antibody, but the antibody secreted by the cells did not bind to CD4. CD4 is a surface molecule on the leukocytes known as T cells. A primatized antibody (a chimeric antibody where the variable domains are isolated from a cynomol-gus monkey see Figure 32.2) to CD4 was produced later that year. That antibody ended up in clinical trials in humans at the same time as Rituxan. 

Therefore, evidence exists to support numerous possible mechanisms for the uptake promoted by TAT peptides. An alternative explanation is that TAT is a sticky opportunistic peptide that has the ability to bind to the cell surface and exploit multiple mechanisms in order to enter the cytoplasm (92). Clearly, the mechanism of internalization requires further study. Nevertheless, it has been observed that transfection with CPPs requires less lipid and therefore proves to be less cytotoxic to cells in vitro and in vivo, making it a promising vector system for future gene therapy (95,104). 

Parkes R, Meng QH, Siapati KE, et al. High efficiency transfection of porcine vascular cells in vitro with a synthetic vector system. J Gene Med 2002 4(3) 292-299. 

Uduehi A, Mailhos C, Truman H, et al. Enhancement of integrin-mediated transfection of haematopoietic cells with a synthetic vector system. Biotechnol Appl Biochem 2003 38(Pt 3) 201-209. 

Sorge, ]., Wright, D., Erdman, V. and Cutting, A. (1984) Amphotropic retrovirus vector system for human cell gene transfer. Mol. Cell. Biol. 4, 1730-1737. 

Insect cells in culture are also hosts for recombinant protein production. Production of recombinant proteins in the baculovirus expression vector system is the most common system. Titers of recombinant protein as high as 11 g/L have been obtained. 

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