Namalwa cells

In addition to recombinant biopharmaceuticals, animal cell culture is used to produce various other biologically based pharmaceuticals. Chief amongst these are a variety of vaccines and hy-bridoma cell-produced monoclonal antibodies (Chapter 13). Earlier interferon preparations were also produced in culture by a particular lymphoblastoid cell line (the Namalwa cell line), which was found to synthesize high levels of several IFN-a s naturally (Chapter 8). 

After this period, there was an accelerated use of animal cells. Namalwa cells (Finter et al., 1991) were used to produce alpha-interferon by Wellcome in 1986. Vero cells (a cell line derived from monkey) were used for anti-rabies vaccine. Hybridomas were considered acceptable for mAb production and the use of genetically modified CHO (Chinese hamster ovary) cells was authorized for the production of tissue plasminogen activator (tPA) by Genentech in 1986. 

Human B-lymphoblastoid cell line Namalwa is derived from a Burkitt s lymphoma tumour biopsy and can be grown for an indefinitely long period of time (Klein et al., 1979 Biliau et ah, 1973). Although they contain Epstein-Barr (EB) nuclear antigen, Namalwa cells do not release EB virus. The cell line is used for the production of IFN-a. Namalwa cells can be induced to produce IFN by adding Sendai virus to cells pretreated with 2 mM sodium butyrate (Baker et al, 1980 Johnston, 1980) and can be primed with Sendai virus in order to increase the levels of IFN-a produced. 

The Namalwa cell line is cited in a US and/or other patents and must not be used to infringe patent claims. 

Johnston MD (1980) Enhanced production of interferon from human lymphoblastoid (Namalwa) cells pre-treated with sodium butyrate. Journal of General Virology 50 191-194. 

Musgrave SC, Douglas Y, Layton G, Merrett J, Scott MF Caulcott CA (1993) Optimisation of alpha-interferon expression in Namalwa cells. In Kaminogawa S, Ametani A Hachimura S (eds) Animal Cell Technology Basic and Applied Aspects, Kluwer Academic, Dordrecht, The Netherlands, pp. 223-230. 

The way to create a serum-free culture is to adapt the cells to the serum-free medium. In our laboratory, we tried to adapt a human lymphoblastoid cell line, Namalwa, from a medium containing 10% serum to serum-free. We were able to adapt Namalwa cell to a ITPSG serum-free medium which contained insulin, transferrin, sodium pyruvate, selenious acid and galactose in RPMI-1640. In the case of cell adaptation for production of autocrine growth factor, we were able to grow the cell line in serum- and protein-free media as well as in K5 62-K1 (T1) which produces an autocrine growth factor, LGF-1 (leukemia derived growth factor-1). 

The specific standard methods of a new perfusion culture will now be described for growth and maintenance of mammalian cells in suspension cultures at high density. The biofermenter was used for high density culture of Namalwa cells with serum-free medium as the model. In 1980, the parent Namalwa cells were obtained fi-om Mr. F. Klein of Frederick Cancer Research Center, Frederick, Maryland, U. S. A. In our laboratories, we were able to adapt the cells to a serum and albumin-fi ee medium and named the cells KJM-1. ITPSG and ITPSG+F68 used a serum-free medium containing insulin, 3 g/ml Transferrin, 5 g/ml sodium pyruvate, 5 mM seienious acid,... 

The biofermenter BF-F500 system consisted of a 1.5 1 culture vessel, 2 1 medium reservoir and effluent bottle (2 1 glass vessels) for fresh and expended media which were connected to the perfusion (culture) vessel by a peristaltic pump. As shown in Fig. 14, the fermenter systems have a conical shape sedimentation column in the center of the fermenter, and an impeller on the bottom of the sedimentation column. The Namalwa cells, KJM-1, were cultivated by continuous cultivation in the biofermenter. In Fig. 15, the culture has been inoculated at 1 to 2 x 10 cells/ml with an initial flow rate of approximately 10 ml/h, sufficient to support the population growth. At densities of 7 x 10 -1.5 x 10" cells/ml, we have used a nutrient flow rate of 1340 ml/h using ITPSG and ITPSG-F68 serum-free media. The flow rate of fresh media was increased step-wise from 240 to 960 ml/d in proportion to the increase in cell density. This resulted in an increase of 4 to 10 fold in cell density compared to the conventional batch culture systems. This system was then scaled up to a 45 1 SUS316L unit mounted on an auto-sterilization sequence system with a medium reservoir and an effluent vessel of 901 each. 

Figure 15. High density Namalwa cell culture in serum-free medium.

Some cell lines, such as Namalwa, can grow satisfactorily in medium in which the only protein is albumin. Other cell lines show distinct protein requirements, such as albumin, transferrin, and insulin, or the addition of polypeptide growth factors, isolated from non-serum sources that have shown stimulation of many cell types in culture. Some cells have very fastidious growth requirements and their stability and productivity may be reduced significantly in serum-free media. 

Namalwa Human tumour cell line Interferon-a production ... 

Baker PN, Morser J Burke DC (1980) Effects of sodium butyrate on a human lymphoblastoid cell line (Namalwa) and its interferon production. Journal of Interferon Research 1 (1) 71-77. 

Miyaji H, Harada N, Mizukami T, Sato S, Fujiyoshi N Itoh S (1990) Expression of human lymphotoxin in namalwa KJM-1 cells adapted to serum-free medium. Cytotechnology 4 39-43. 

Cellular Origin. IFNa is produced by monocytes, macrophages, lymphocytes, and cellular lines such as Namalwa and KGl (Figure 22-27). IFNp is produced by fibroblasts and epithelial cells. However, many other cells can also produce these cytokines, including hematopoietic cells... 

Recently, a cDNA encoding a GM3-specific a2-8 SAT (SAT-2 or GD3 synthase) has been isolated from an expression cDNA library of human melanoma cell line WM266-4 by enrichment of Namalwa KJM-1 cells highly expressing GD3 (Sasaki et al., 1994). Anti-GD3 antibody is used for fluorescence-activated... 

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