MUTATED E. COLI HEAT-LABILE ENTEROTOXIN

• US 20080102078A1
• Filed: 07/18/2007
• Published: 05/01/2008
• Est. Priority Date: 10/27/2006
• Status: Active Grant

• Current Assignee: Development Center for Biotechnology
• Original Assignee: Development Center for Biotechnology
• Inventors: Yuan, Ta-Tung, Hsu, Yu-Shen, Lin, Young-Sun
• Granted Patent: US 8,110,197 B2
• Time in Patent Office: Days
• US Class Current: 424/189.1
• CPC Class Codes: A61K 2039/5252 inactivated (killed)
• CPC Class Codes: A61K 2039/543 intranasal
• CPC Class Codes: A61K 2039/55544 Bacterial toxins
• CPC Class Codes: A61K 39/12 Viral antigens
• CPC Class Codes: A61K 39/145 Orthomyxoviridae, e.g. influenza virus
• CPC Class Codes: A61K 39/39 characterised by the immunostimulating additives, e.g. chemical adjuvants
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07K 14/245 Escherichia (G)
• CPC Class Codes: C12N 2760/16134 Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
• CPC Class Codes: Y02A 50/30 Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

This invention relates to a mutant E. coli heat-labile enterotoxin (LT) subunit A that can be used as an adjuvant. This subunit A mutant contains an amino acid substitution at a position corresponding to position 61 of a wild-type LT. An LT containing this mutated subunit A exhibits reduced toxicity compared to its wild type counterpart.

Claims

1. An isolated polypeptide comprising a mutated E. coli heat-labile enterotoxin subunit A, a position of which corresponds to position 61 of SEQ ID NO:

5 and is an amino acid residue other than S, T, and F, wherein an E. coli heat-labile enterotoxin containing the mutated subunit A has a reduced toxicity compared to a wild-type E. coli heat-labile enterotoxin containing SEQ ID NO;

5.

2. The isolated polypeptide of claim 1, wherein the E. coli heat-labile enterotoxin containing the mutated subunit A has a toxicity less than 10⁻

6-fold that of the wild-type E. coli heat-labile enterotoxin.

3. The isolated polypeptide of claim 1, wherein the amino acid residue is selected from a group consisting of K, R, H, and Y.

4. The isolated polypeptide of claim 1, wherein the mutated subunit A has an amino acid sequence selected from a group consisting of SEQ ID NOs:

2, 4, 8, and 10.

5. A vaccine comprising an antigen and an adjuvant, the adjuvant being a mutated E. coli heat-labile enterotoxin subunit A, a position of which corresponds to position 61 of SEQ ID NO:

5 and is an amino acid residue other than S, T, or F, wherein an E. coli heat-labile enterotoxin containing the mutated subunit A has a reduced toxicity compared to a wild-type E. coli heat-labile enterotoxin containing SEQ ID NO;

5.

6. The vaccine of claim 5, wherein the antigen is from a bacterium or a virus.

7. The vaccine of claim 6, wherein the bacterium is selected from a group consisting of Streptococcus pneumoniae, Escherichia coli, Helicobacter pylori, Neisseria meningitidis, and Haemophilus influenza b.

8. The vaccine of claim 6, wherein the virus is selected from a group consisting of Influenza virus, human immunodeficiency virus, hepatitis A virus, hepatitis B virus, hepatitis C virus, human papillomavirus, enterovirus, and rotavirus.

9. The vaccine of claim 5, wherein the E. coli heat-labile enterotoxin containing the mutated subunit A has a toxicity less than 1⁻

6-fold that of the wild-type E. coli heat-labile enterotoxin.

10. The vaccine of claim 5, wherein the subunit A mutant contains K, R, Y, or H at the position corresponding to position 61 of SEQ ID NO:

5.

11. The vaccine of claim 5, wherein the subunit A mutant contains K at the position corresponding to position 61 of SEQ ID NO:

5.

12. The vaccine of claim 5, wherein the subunit A mutant contains R at the position corresponding to position 61 of SEQ ID NO:

5.

13. The vaccine of claim 5, wherein the subunit A mutant contains an amino acid sequence selected from a ground consisting of SEQ ID NOs:

2, 4, 8, and 10.

14. The vaccine of claim 5, further comprising LT subunit B, wherein the subunit A mutant and the subunit B form whole E. coli heat-labile enterotoxin.

15. An isolated nucleic acid comprising a nucleotide sequence encoding a mutated E. coli heat-labile enterotoxin subunit A, its position corresponding to position 61 of SEQ ID NO:

5 being an amino acid residue other than S, T, or F, wherein an E. coli heat-labile enterotoxin containing the mutated subunit A has reduced toxicity compared to a wild-type E. coli heat-labile enterotoxin containing SEQ ID NO;

5.

16. The nucleic acid of claim 15, wherein the E. coli heat-labile enterotoxin containing the mutated subunit A has a toxicity less than 10⁻

6-fold that of the wild-type E. coli heat-labile enterotoxin.

17. The nucleic acid of claim 15, wherein the amino acid residue is selected from a group consisting of K, R, Y, and H.

18. The nucleic acid of claim 15, wherein the nucleotide sequence is selected from a group consisting of SEQ ID NOs:

1, 3, 7, and 9.

19. A vector comprising the nucleic acid of claim 15.

20. A transformed cell comprising the vector of claim 19.