Abstract
We have used transgenic mouse technology to establish immortalized hepatoma cell lines stably secreting heterologous proteins, such as human α1-antitrypsin and human factor IX. Hepatocyte-specific regulatory DNA sequences were used to target both the expression of anonc gene and the gene coding for the human protein to the liver of transgenic mice which eventually developed hepatocellular carcinomas. Tumour cells were subsequently established as permanent cell lines, which maintained a differentiated phenotype under specific culture conditions, being capable of producing biologically active and correctly processed human α1-antitrypsin and factor IX. Moreover, a preliminary analysis has shown that certain cell lines express elevated total cytochrome P450 activity. These cells could therefore represent a useful alternative to the use of animals or primary cultures in drug safety testing.
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References
Anson DS, Austen DEG and Brownlee GG (1985) Expression of human clotting factor IX from recombinant DNA clones in mammalian cells. Nature 315: 683–685.
Austen DEG and Rhymes IL (1975) A Laboratory Manual of Blood Coagulation (Blackwell Scientific).
Bernard O, Cory S, Gerondakis S, Webb E and Adams JM (1983) Sequence of the murine and human cellularmyc oncogenes and two modes ofmyc transcription resulting from chromosome translocation in B lymphoid tumours. EMBO J 2: 2375–2383.
Costanzi C and Gillespie D (1987) Fast blots: immobilization of DNA and RNA from cells. In: Berger SL, Kimmel AR (eds) Guide to Molecular Cloning Techniques, Vol 152. Orlando: Academic Press, pp. 582–587.
Crystal RG (1989) The α1-antitrypsin gene and its deficiency states. Trends Genet 5: 411–417.
Dalemans W, Perraud F, Le Meur M, Gerlinger P, Courtney M and Pavirani A (1990) Heterologous protein expression by trans-immortalised differentiated liver cell lines derived from transgenic mice. Biologicals 18: 191–198.
Deschatrette J and Weiss MC (1974) Characterization of differentiated and dedifferentiated clones of a rat hepatoma. Biochimie 56: 1603–1611.
Furie B and Furie BC (1988) The molecular basis of blood coagulation. Cell 53: 505–518.
Hogan B, Costantini F and Lacy E (1986) Manipulating the mouse embryo, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, pp. 332.
Jallat S, Perraud F, Dalemans W, Balland A, Dieterle A, Faure T, Meulien P and Pavirani A (1990a) Characterization of recombinant human Factor IX expressed in transgenic mice and in derived trans-immortalized hepatic cell lines. EMBO J 9: 3295–3301.
Jallat S, Meulien P, Pavirani A and Perraud F (1990b) Patent Cooperation Treaty (PCT)/FR90/00606.
Kaufman RJ, Wasley LC, Furie BC, Furie B and Shoemaker CB (1986) Expression, purification and characterization of recombinant γ-carboxylated Factor IX synthesized in chinese hamster ovary cells. J Biol Chem 261: 9622–9628.
Long GL, Chandra T, Woo SLC, Davie EW and Kurachi K (1984) Complete sequence of the cDNA for human α1-antitrypsin and the gene for the S variant. Biochemistry 23: 4828–4837.
Morel F, Beaune PH, Ratanasavanh D, Flinois JP, Yang CS, Guengerich FP and Guillouzo A (1990) Expression of cytochrome P450 enzymes in cultured human hepatocytes. Eur J Biochem 191: 437–444.
Omura T and Sato R (1964) The carbon monoxide-building pigment of liver microsomes. II Solubilization, purification and properties. J Biol Chem 239: 2379–2385.
Palmiter RD, Chen HY and Brinster RL (1982) Differential regulation of metallothionein-thymidine kinase fusion genes in transgenic mice and their offspring. Cell 29: 701–710.
Pavirani A, Meulien P, Harrer H, Schamber F, Dott K, Villeval D, Cordier Y, Wiesel ML, Mazurier C, Van de Pol H, Piquet Y, Cazenave JP and Lecocq JP (1987) Choosing a host cell for active recombinant factor VIII production using vaccinia virus. Bio/Technology 5: 389–392.
Pavirani A, Skern T, Le Meur M, Lutz Y, Lathe R, Crystal RG, Fuchs JP, Gerlinger P and Courtney M (1989) Recombinant proteins of therapeutic interest expressed by lymphoid cell lines derived from transgenic mice. Bio/Technology 7: 1049–1054.
Perraud F, Dalemans W, Gendrault JL, Dreyer D, Ali-Hadji D, Faure T and Pavirani A (1991) Characterization of transimmortalised hepatic cell lines established from transgenic mice. Exp Cell Res 195: 59–65.
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Perraud, F., Dalemans, W., Ali-Hadji, D. et al. Novel cell lines derived from transgenic mice expressing recombinant human proteins. Cytotechnology 9, 69–75 (1992). https://doi.org/10.1007/BF02521733
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DOI: https://doi.org/10.1007/BF02521733