Paramyxovirus-derived RNP
OwnerKaio Kitazato, Tsugumine Shu +8inventors
InventorsKaio Kitazato, Tsugumine Shu, Hidekazu Kuma, Yasuji Ueda, Makoto Asakawa, Mamoru Hasegawa, Akihiro Iida, Takahiro Hirata, Makoto Inoue, Yumiko Tokusumi
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Abstract
A functional RNP containing negative-strand single-stranded RNA derived from Sendai virus, which has been modified so as not to express at least one envelope protein, has been successfully prepared. An RNP comprising a foreign gene is prepared and inserted into a cell with the use of a cationic liposome, thereby successfully expressing the foreign gene.
Claims42
A method of producing a complex, wherein said method comprising the steps of:
- (a) forming a complex comprising (i) a negative-strand single-stranded RNA derived from a paramyxovirus, or the complementary strand thereof, and (ii) proteins encoded by and binding to said negative-strand single-stranded RNA, or its complement;
- (b) amplifying the complex of step (a) in cells expressing an envelope protein;
- and
- (c) recovering the complex of step (b);
- wherein the negative-strand single-stranded RNA, or its complement, is modified so as to express NP, P, and L proteins, but not at least one protein selected from the group consisting of (i) F protein, (ii) HN or H protein, and (iii) M protein, wherein the cells, expressing the envelope protein, include a gene encoding the envelope protein incorporated into their chromosomes, and wherein expression of the envelope protein is induced by a DNA recombinase.
A method of producing a complex, said method comprising the steps of:
- (a) transcribing a DNA encoding a negative-strand single-stranded RNA, or the complementary strand thereof, in a first cell expressing at least NP protein, P protein, L protein, and an envelope protein, wherein said negative-strand single-stranded RNA, or its complement, is modified to not express at least one protein selected from the group consisting of (i) F protein, (ii) HN or H protein, and (iii) M protein to form a complex comprising said negative-strand single-stranded RNA, or its complement, and proteins encoded by said negative-strand single-stranded RNA, or its complement;
- (b) amplifying said complex by co-culturing said first cell with a second cell that comprises a gene encoding an envelope protein in its chromosome and expresses said envelope protein, wherein the expression of said envelope protein is induced by a DNA recombinase;
- and
- (c) recovering the complex of step (b).
A helper cell comprising at least a gene selected from the group consisting of (i) F gene, (ii) HN or H gene, and (iii) M gene of a paramyxovirus in its chromosomes, wherein expression of the gene is induced by a DNA recombinase.
A method of attenuating cytotoxicity caused by gene transfer with a paramyxovirus vector, wherein the method is characterized by using a paramyxovirus vector deficient in at least one gene selected from the group consisting of (i) F gene, (ii) HN or H gene, and (iii) M gene.
A paramyxovirus vector comprising G protein of vesicular stomatitis virus, wherein said paramyxovirus vector does not comprise a paramyxovirus gene encoding a G protein.
A method of producing bone marrow cells expressing a foreign gene, said method comprising the step of contacting a paramyxovirus vector carrying the foreign gene with a bone marrow cell.
Dependencies are inferred from the claim text.