Organic compounds -- part of the class 532-570 series – Organic compounds – Carbohydrates or derivatives
Reexamination Certificate
1998-10-20
2003-07-15
Priebe, Scott D. (Department: 1632)
Organic compounds -- part of the class 532-570 series
Organic compounds
Carbohydrates or derivatives
C435S069100, C435S069300, C435S320100, C424S093200, C424S185100, C424S268100, C424S272100, C530S300000, C530S350000
Reexamination Certificate
active
06593463
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The invention relates to heterologous gene expression. More particularly, the invention relates to the expression of malaria genes in higher eukaryote cell systems.
2. Summary of the Related Art
Recombinant production of certain heterologous gene products is often difficult in ill vitro cell culture systems or ill vivo recombinant production systems. For example, many researchers have found it difficult to express proteins derived from bacteria, parasites and virus in cell culture systems different from the cell from which the protein was originally derived, and particularly in mammalian cell culture systems. One example of a therapeutically important protein which has been difficult to produce by mammalian cells is the malaria merozoite surface protein (MSP-1).
Malaria is a serious heath problem in tropical countries. Resistance to existing drugs is fast developing and a vaccine is urgently needed. Of the number of antigens that get expressed during the life cycle of
P. falciparum,
MSP-1 is the most extensively studied and promises to be the most successful candidate for vaccination. Individuals exposed to
P. falciparum
develop antibodies against MSP-1, and studies have shown that there is a correlation between a naturally acquired immune response to NISP-1 and reduced malaria morbidity. In a number of studies, immunization with purified native MSP-1 or recombinant fragments of the protein has induced at least partial protection from the parasite (Diggs et al, (1993)
Parasitol Today
9:300-302). Thus MSP-1 is an important target for the development of a vaccine against
P. falciparum.
MSP-1 is a 190-220 kDA glycoprotein. The C-terminal region has been the focus of recombinant production for use as a vaccine. However, a major problem in developing MSP-1 as a vaccine is the difficulty in obtaining recombinant proteins in bacterial or yeast expression systems that are equivalent in immunological potency to the affinity purified native protein (Chang et al., (1992)
J. Immunol.
148:548-555.) and in large enough quantities to make vaccine production feasible.
Improved procedures for enhancing expression of sufficient quantities of MSP-1 would be advantageous.
BRIEF SUMMARY OF THE INVENTION
The present invention provides improved recombinant DNA compositions and procedures for increasing the mRNA levels and protein expression of the malarial surface antigen MSP-1 in cell culture systems, mammalian cell culture systems, or in transgenic mammals. The preferred protein candidate for expression in an expression system in accordance with the invention is a C-terminal derivative of MSP-1 having a DNA coding sequence with reduced AT content, and eliminated mRNA instability motifs and rare codons relative to the recombinant expression systems. Thus, in a first aspect, the invention provides a DNA sequence derived from the sequence shown in SEQ ID NO 2. This derivative sequence is shown in SEQ ID NO 1.
In a second aspect, the invention provides a process for preparing a modified nucleic acid of the invention comprising the steps of lowering the overall AT content of the natural gene encoding MSP-1, eliminating all mRNA instability motifs and replacing all rare codons with a preferred codon of the mammary gland tissue, all by replacing specific codons in the natural gene with codons recognizable to, and preferably preferred by mammary gland tissue and which code for the same amino acids as the replaced codon. This aspect of the invention further includes modified nucleic acids prepared according to the process of the invention.
In a third aspect, the invention also provides vectors comprising modified MSP-1 nucleic acids of the invention and a goat beta casein promoter and signal sequence, and host cells transformed with nucleic acids of the invention.
In a fourth aspect, the invention provides transgenic non-human mammals whose germlines comprise a nucleic acid of the invention.
In a fifth aspect, the invention provides a DNA vaccine comprising a modified MSP-1 gene according to the invention.
REFERENCES:
patent: 5231168 (1993-07-01), Dziegiel et al.
patent: 5736131 (1998-04-01), Bosch et al.
patent: 5795737 (1998-08-01), Seed et al.
patent: 1997 48649 (1998-04-01), None
patent: 0 682 115 (1995-11-01), None
patent: WO 91 08216 (1991-06-01), None
patent: WO 94 28930 (1994-12-01), None
patent: WO 97/30159 (1997-08-01), None
patent: WO 97 31115 (1997-08-01), None
patent: WO 98 14583 (1998-04-01), None
Chattergoon et al, 1997, FASEB J., 11: 753-763.*
Ebert et al, 1991, Biotechnology, 9: 935-838.*
Holder et al, 1985, Nature, 317: 270-273.*
Jongwutiwes et al, 1993, Mol. Biochem. Parasitol., 59: 95-100.*
Ledley et al, 1991, Hum. Gene Ther., 2: 77-83.*
McDonnell et al, 1996, The New England J. Med., 334(1): 42-45.*
Orkin et al, 1995, Report and Recommendations of Panel to Assess NIH Investment in Gene Therapy Res.*
Pharmacia Catalogue, 1995, p. 277.*
Wang et al, 1989, J. Biol. Chem., 264(35): 21116-21121.*
Zinkernagel et al, 1993, Fundamental Immunology, 3rd edition, Raven Press.*
Akashi et al, 1994, Blood, 83(11): 3182-3187.*
Gordan et al.; Genetic Transformation Mouse Enbryos by Microinjection of purified DNA; (1980);Proc. Natl. Acad. Sci.; 77: pp. 7380-7384.
Gorden et al.; Integration and Stable Germ Line Transmission of Genes Injected into Mouse Pronuclei; (1981);Science; 214: pp. 1244-1246.
Brinster et al.; Factors Affecting the Efficiency of Intoducing Foreing DNA into Mice by Microinjecting Eggs; (1985);Proc. Natl. Acad. Sci.; 82: pp. 4438-4442.
Palmiter et al.; Transgenic Mice; (1985);Cell; 41: pp. 343-345.
Wall et al.; Development of Porcine Ova That Were Centrifuged to permit Visualizaton of Pronuclei and Nuclei1; (1985);Biol. Reprod.; 32: pp. 645-651.
Shaw et al.; A Conserved AU Sequence from the 3′Untranslated Region of GM-CSF mRNA Mediates Selective mRNA Degradation; (1986);Cell; 46: pp. 659-667.
Simons et al.; Gene Transfer into Sheep; (1988); Bio/Technology; 6: pp. 179-183.
Chang et al.; Generalized Immunological Recognition of the Major Merozoite Surface Antigen (gp195) ofPlasmodium falciparum; (1989);Proc. Natl. Acad. Sci.; 86: pp. 6343-6347.
Vilotte et al.; Efficient Tissue-Specific Expression of Bovine &agr;-lactalbumin in Transgenic Mice; (1989);Eur. J. Biochem.; 186: pp. 43-48.
Buhler et al.; Rabbit &bgr;-Casein Promotor Directs Secretion of Human Interleukin-2 into the Milk of Transgenic Rabbits; (1990);Bio/Technology; 8: pp. 140-143.
Ebert et al.; Transgenic Production of A Variant of Human Tissue-Type Plasminogen Activator in Goat Milk Generation of Transgenic Goats . . . ; (1991);Bio/Technology; 9: pp. 835-838.
Krimenfort et al.; Generation of Transgenic Dairy Cattle Using ‘In Vitro’Embryo Production; (1991)Bio/Technology; 9: pp. 844-847.
Wright et al.; High Level Expression of Active Human Alpha-1-Antitrypsin in the Milk of Transgenic Sheep; (1991);Bio/Technology; 9: 830-834.
Chang et al.; A Carboxyl-Terminal Fragment ofPlasmodium falciparumgp195 Expressed by a Recombinant Baculovirus Induces Antibodies that . . . ; (1992);J. Immunol; 149: pp. 548-555.
Soulier et al.; Expression Analysis of Ruminant &agr;-Lactalbumin in Transgenic Mice: Developmental Regulation and General Location of Important Cis-Regulatory . . . ; (1992);FEBS Letters: 297(1-2): pp. 13-18.
Wall et al.; Making Transgenic Livestock: Genetic Engineering on a Large Scale: (1992);J. Cell. Biochem.; 49: pp. 113-120.
Diggs et al.; The Major Merozoite Surface Protein as a Malaria Vaccine Target; (1993); Parasitology Today; 9(8): pp. 300-302.
Campbell et al.; Sheep Coloned by Nuclear Transfer from a Cultured Cell Line; (1996);Nature; 380: pp. 64-66.
Chang et al.; A Recombinant Baculovirus 42-Kilodalton C-Terminal Fragment ofPlasmodium falciparumMerozoite Surface Protein 1 Protects Aotus Monkeys . . . ; (1996);Infecton&Immunity; 64(1): pp. 253-262.
Hochi et al.; Secretion of Bovine &agr;-Lactalbumin Into the Milk of Transgenic Rats, (1992)Molecular Reproduction and Development33: 160-164.
Chen Li How
Meade Harry
Fish & Richardson P.C.
Genzyme Transgenics Corporation
Paras, Jr. Peter
Priebe Scott D.
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