Process for producing antibody composition by using rna inhibiting the function of alpha1,6-fucosyltransferase

1 . A process for producing an antibody composition using a cell, which comprises using a cell into which a double-stranded RNA comprising an RNA selected from the following (a) or (b) and its complementary RNA is introduced:

(a) an RNA comprising the nucleotide sequence represented by any one of SEQ ID NOs:9 to 30;

(b) an RNA consisting of a nucleotide sequence in which one or several nucleotide(s) is/are deleted, substituted, inserted and/or added in the nucleotide sequence represented by any one of SEQ ID NOs:9 to 30 and having activity of suppressing the function of an enzyme relating to the modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in a complex type N-glycoside-linked sugar chain.

2 . The process according to claim 1 , wherein the enzyme relating to the modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in the complex type N-glycoside-linked sugar chain is α1,6-fucosyltransferase.

3 . The process according to claim 2 , wherein the α1,6-fucosyltransferase is a protein encoded by a DNA selected from the group consisting of the following (a) to (h):

(a) a DNA comprising the nucleotide sequence represented by SEQ ID NO:1;

(b) a DNA comprising the nucleotide sequence represented by SEQ ID NO:2;

(c) a DNA comprising the nucleotide sequence represented by SEQ ID NO:3;

(d) a DNA comprising the nucleotide sequence represented by SEQ ID NO:4;

(e) a DNA which hybridizes with a DNA consisting of the nucleotide sequence represented by SEQ ID NO:1 under stringent conditions and encodes a protein having α1,6-fucosyltransferase activity;

(f) a DNA which hybridizes with a DNA consisting of the nucleotide sequence represented by SEQ ID NO:2 under stringent conditions and encodes a protein having α1,6-fucosyltransferase activity;

(g) a DNA which hybridizes with a DNA consisting of the nucleotide sequence represented by SEQ ID NO:3 under stringent conditions and encodes a protein having α1,6-fucosyltransferase activity;

(h) a DNA which hybridizes with a DNA consisting of the nucleotide sequence represented by SEQ ID NO:4 under stringent conditions and encodes a protein having α1,6-fucosyltransferase activity.

4 . The process according to claim 2 , wherein the α1,6-fucosyltransferase is a protein selected from the group consisting of the following (a) to (l):

(a) a protein comprising the amino acid sequence represented by SEQ ID NO:5;

(b) a protein comprising the amino acid sequence represented by SEQ ID NO:6;

(c) a protein comprising the amino acid sequence represented by SEQ ID NO:7;

(d) a protein comprising the amino acid sequence represented by SEQ ID NO:8;

(e) a protein consisting of an amino acid sequence in which one or more amino acid(s) is/are deleted, substituted, inserted and/or added in the amino acid sequence represented by SEQ ID NO:5 and having α1,6-fucosyltransferase activity;

(f) a protein consisting of an amino acid sequence in which one or more amino acid(s) is/are deleted, substituted, inserted and/or added in the amino acid sequence represented by SEQ ID NO:6 and having α1,6-fucosyltransferase activity;

(g) a protein consisting of an amino acid sequence in which one or more amino acid(s) is/are deleted, substituted, inserted and/or added in the amino acid sequence represented by SEQ ID NO:7 and having α1,6-fucosyltransferase activity;

(h) a protein consisting of an amino acid sequence in which one or more amino acid(s) is/are deleted, substituted, inserted and/or added in the amino acid sequence represented by SEQ ID NO:8 and having α1,6-fucosyltransferase activity;

(i) a protein consisting of an amino acid sequence which has 80% or more homology to the amino acid sequence represented by SEQ ID NO:5 and having α1,6-fucosyltransferase activity;

(j) a protein consisting of an amino acid sequence which has 80% or more homology to the amino acid sequence represented by SEQ ID NO:6 and having α1,6-fucosyltransferase activity;

(k) a protein consisting of an amino acid sequence which has 80% or more homology to the amino acid sequence represented by SEQ ID NO:7 and having α1,6-fucosyltransferase activity;

(l) a protein consisting of an amino acid sequence which has 80% or more homology to the amino acid sequence represented by SEQ ID NO:8 and having α1,6-fucosyltransferase activity.

5 . The process according to claim 1 , wherein the cell into which the RNA having activity of suppressing the function of an enzyme relating to the modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in a complex type N-glycoside-linked sugar chain is introduced is a cell which is resistant to a lectin which recognizes a sugar chain structure in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in an N-glycoside-linked sugar chain.

6 . The process according to claim 5 , wherein the cell is resistant to at least one lectin selected from the group consisting of the following (a) to (d):

(a) a Lens culinaris lectin;

(b) a Pisum sativum lectin;

(c) a Vicia faba lectin;

(d) an Aleuria aurantia lectin.

7 . The process according to claim 1 , wherein the cell is selected from the group consisting of a yeast cell, an animal cell, an insect cell and a plant cell.

8 . The process according to claim 1 , wherein the cell is a cell selected from the group consisting of the following (a) to (i):

(a) a CHO cell derived from Chinese hamster ovary tissue;

(b) a rat myeloma cell line YB2/3HL.P2.G11.16Ag.20 cell;

(c) a mouse myeloma cell line NS0 cell;

(d) a mouse myeloma cell line SP2/0-Ag14 cell;

(e) a BHK cell derived from Syrian hamster kidney tissue;

(f) an antibody-producing hybridoma cell;

(g) a human leukemia cell line Namalwa cell;

(h) an embryonic stem cell;

(i) a fertilized egg cell.

9 . The process according to claim 1 , wherein the cell is a transformant into which a gene encoding an antibody molecule is introduced.

10 . The process according to claim 9 , wherein the antibody molecule is selected from the group consisting of the following (a) to (d):

(a) a human antibody;

(b) a humanized antibody;

(c) an antibody fragment comprising the Fc region of (a) or (b);

(d) a fusion protein comprising the Fc region of (a) or (b).

11 . The process according to claim 9 , wherein the antibody molecule belongs to an IgG class.

12 . The process according to claim 1 , wherein the antibody composition is an antibody composition having higher antibody-dependent cell-mediated cytotoxic activity than an antibody composition produced by a parent cell into which a double-stranded RNA comprising an RNA selected from the following (a) or (b) and its complementary RNA is not introduced:

(a) an RNA comprising the nucleotide sequence represented by any one of SEQ ID NOs:9 to 30;

(b) an RNA consisting of a nucleotide sequence in which one or several nucleotide(s) is/are deleted, substituted, inserted and/or added in the nucleotide sequence represented by any one of SEQ ID NOs:9 to 30 and having activity of suppressing the function of an enzyme relating to the modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetyiglucosamine in the reducing end through α-bond in a complex type N-glycoside-linked sugar chain.

13 . The process according to claim 12 , wherein the antibody composition having higher antibody-dependent cell-mediated cytotoxic activity is an antibody composition which comprises antibody molecules having complex type N-glycoside-linked sugar chains in the Fc region, and in which a ratio of sugar chains in which fucose is not bound to N-acetylglucosamine in the reducing end in the sugar chains among the complex type N-glycoside-linked sugar chains is higher than that of an antibody composition produced by the parent cell.

14 . The process according to claim 13 , wherein the complex type N-glycoside-linked sugar chains are sugar chains in which 1-position of fucose is not bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in the sugar chains.

15 . The process according to claim 12 , wherein the antibody composition having higher antibody-dependent cell-mediated cytotoxic activity is an antibody composition which comprises antibody molecules having complex type N-glycoside-linked sugar chains in the Fc region, and in which the ratio of sugar chains in which fucose is not bound to N-acetylglucosamine in the reducing end in the sugar chains among the complex type N-glycoside-linked sugar chains is 20% or more.

16 . The process according to claim 12 , wherein the antibody composition having higher antibody-dependent cell-mediated cytotoxic activity is an antibody composition which comprises antibody molecules having complex type N-glycoside-linked sugar chains in the Fc region, and in which the complex type N-glycoside-linked sugar chains are sugar chains in which fucose is not bound to N-acetylglucosamine in the reducing end.

17 . A cell into which an RNA capable of suppressing the function of an enzyme relating to the modification of a sugar chain in which 1position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in a complex type N-glycoside-linked sugar chain is introduced, and which is used in the process according to claim 1 .

18 . The cell according to claim 17 , wherein the enzyme relating to the modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in a complex type N-glycoside-linked sugar chain is α1,6-fucosyltransferase.

19 . A cell in which an RNA selected from RNAs of the group consisting of the nucleotide sequences represented by any one of SEQ ID NOs:9 to 30 is introduced or expressed.

20 . A double-stranded RNA consisting of an RNA selected from the following (a) or (b) and its complementary RNA:

(a) an RNA comprising the nucleotide sequence represented by any one of SEQ ID NOs:9 to 30;

(b) an RNA consisting of a nucleotide sequence in which one or several nucleotide(s) is/are deleted, substituted, inserted and/or added in the nucleotide sequence represented by any one of SEQ ID NOs:9 to 30 and having activity of suppressing the function of an enzyme relating to the modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in a complex type N-glycoside-linked sugar chain.

21 . A DNA corresponding to the RNA described in claim 20 and a complementary DNA to the DNA.

22 . A recombinant DNA which is obtainable by introducing a DNA corresponding to the RNA described in claim 20 and a complementary DNA to the DNA into a vector.

23 . The recombinant DNA according to claim 22 , which expresses the double-stranded RNA consisting of an RNA selected from the following (a) or (b) and its complementary RNA:

(a) an RNA comprising the nucleotide sequence represented by any one of SEQ ID NOs:9 to 30;

(b) an RNA consisting of a nucleotide sequence in which one or several nucleotide(s) is/are deleted, substituted, inserted and/or added in the nucleotide sequence represented by any one of SEQ ID NOs:9 to 30 and having activity of suppressing the function of an enzyme relating to the modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in a complex type N-glycoside-linked sugar chain.

24 . A transformant which is obtainable by introducing the recombinant DNA according to claim 22 into a cell.

25 . A method for constructing a cell which is resistant to a lectin which recognizes a sugar chain structure in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in a complex type N-glycoside-linked sugar chain, which comprises introducing or expressing the double-stranded RNA described in claim 20 in a cell.

26 . The method according to claim 25 , wherein the cell which is resistant to a lectin which recognizes a sugar chain structure in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in a complex type N-glycoside-linked sugar chain is resistant to at least one lectin selected from the group consisting of the following (a) to (d):

(a) a Lens culinaris lectin;

(b) a Pisum sativum lectin;

(c) a Vicia faba lectin;

(d) an Aleuria aurantia lectin.

27 . A method for suppressing the function of an enzyme relating to the modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in a complex type N-glycoside-linked sugar chain, which comprises using an RNA selected from RNAs of the group consisting of the nucleotide sequences of any one of SEQ ID NOs:9 to 30.

28 . The method according to claim 27 , wherein the enzyme relating to the modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in a complex type N-glycoside-linked sugar chain is α1,6-fucosyltransferase.