Vaccine containing a Shigella bacterium having an enhanced antigenic property

• US 5,858,352 A
• Filed: 05/30/1997
• Issued: 01/12/1999
• Est. Priority Date: 10/05/1994
• Status: Expired due to Fees

• Current Assignee: Bioport Company Limited
• Original Assignee: Antex Biologics Inc.
• Inventors: Pace, John Lee, Frey, Steven Michael, Walker, Richard Ives
• Primary Examiner(s): Naff, David M.
• Assistant Examiner(s): WARE, DEBORAH K
• Application Number:
  US08/866,586
• Time in Patent Office: 592 Days
• Field of Search: 435/252.1, 435/822, 424/93.4, 424/184.1, 424/282.1
• US Class Current: 424/93.4
• CPC Class Codes: A61K 2039/55555 Liposomes; Vesicles, e.g. nanoparticles; Spheres, e.g. nanospheres; Polymers
• CPC Class Codes: A61K 39/105 Delta proteobacteriales, e.g. Lawsonia; Epsilon proteobacteriales, e.g. campylobacter, helicobacter
• CPC Class Codes: A61K 39/107 Vibrio
• CPC Class Codes: A61P 31/04 Antibacterial agents
• CPC Class Codes: C12N 1/20 Bacteria; Culture media therefor
• CPC Class Codes: C12N 1/205 Bacterial isolates
• CPC Class Codes: C12Q 1/045 Culture media therefor
• CPC Class Codes: C12Q 1/18 Testing for antimicrobial activity of a material
• CPC Class Codes: C12R 2001/01 Bacteria or Actinomycetales ; using bacteria or Actinomycetales
• CPC Class Codes: G01N 2333/25 Shigella (G)
• CPC Class Codes: G01N 2469/20 Detection of antibodies in sample from host which are directed against antigens from microorganisms
• CPC Class Codes: G01N 2474/20 Immunohistochemistry assay
• CPC Class Codes: G01N 33/56916 Enterobacteria, e.g. shigella, salmonella, klebsiella, serratia
• CPC Class Codes: G01N 33/56922 Campylobacter
• 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
• CPC Class Codes: Y10S 424/802 Antibody or antigen-binding fragment thereof that binds gram-positive bacteria
• CPC Class Codes: Y10S 435/822 using bacteria or actinomycetales
• CPC Class Codes: Y10S 435/975 Kit

Abstract

Methods using in vitro processes are disclosed for inducing or enhancing expression of enteric bacterial antigens or virulence factors. The methods, therefore, produce antigenically enhanced bacteria for use in vaccines. Also disclosed are methods for using the antigenically enhanced bacteria as well as the vaccines containing the enteric bacteria. Specifically, a whole enteric bacteria or components thereof, are provided by Shigella species. In addition to this microorganism there are other enteric bacteria, such as Campylobacter species and Helicobacter pylori, which are useful for inducing or enhancing expression of enteric bacterial antigens and/or virulence factors.

Claims

1. A vaccine comprising a Shigella bacterium, or an immunogenic fragment or derivative thereof, said Shigella bacterium having an enhanced antigenic property, which is harvested from a culture of a Shigella species grown in vitro in a culture medium with a combination of conditions comprising:

(a) about 0.05% to about 3% bile or about 0.025% to about 0.6% of one or more bile acids or salts thereof;

(b) at a temperature between about 30°

C. and about 42°

C.;

(c) in air or a gas mixture, wherein the gas mixture comprises (i) about 5% to about 20% CO₂ with about 80% to about 95% air;

(ii) about 5% to about 20% CO₂ with about 80% to about 95% N₂ ;

or (iii) about 5% to about 10% O₂ with about 10% to about 20% CO₂ with about 70% to about 85% N₂ ; and

(d) a divalent cation chelator selected from the group consisting of 0 to about 100 μ

M of 1,2-bis(2-aminophenoxy)ethane-N,N,N'"'"',N'"'"'-tetraacetic acid/acetoxymethyl ester, 0 to about 10 mM of ethylene-bis(oxyethylenenitrilo)-tetraacetic acid, and 0 to about 100 μ

M of ethylene-bis (oxyethylenenitrilo)-tetraacetic acid/acetoxymethyl ester,

wherein said Shigella culture is at about early log phase, between early log phase and stationary phase, or at about stationary phase, and the enhanced antigenic property is a higher level of an immunogenic antigen or a new immunogenic antigen when compared to the antigenic property of bacteria from a culture of the Shigella species grown in brain heart infusion broth.

2. The vaccine according to claim 1, wherein the Shigella bacterium is a Shigella species selected from the group consisting of flexneri, sonnei, dysentariae and boydii.

3. The vaccine according to claim 2, wherein the Shigella bacterium is Shigella flexneri strain 2457T and the combination of conditions comprises:

(a) about 0.1% bile salt which is deoxycholate or about 0.8% bile;

(b) the temperature is about 37°

C.;

(c) in air; and

the culture is in early log phase.

4. The vaccine according to claim 1, further comprising a pharmaceutically acceptable carrier or diluent.

5. The vaccine according to claim 1, wherein said Shigella bacterium is inactivated.

6. The vaccine according to claim 5, wherein said Shigella bacterium is inactivated by formalin treatment.

7. The vaccine according to claim 1, wherein said vaccine is suitable for mucosal or parenteral or mucosal and parenteral administration.

8. The vaccine according to claim 1, further comprising an adjuvant.

9. The vaccine according to claim 8, wherein the adjuvant is a nontoxic heat labile toxin E. coli known as mLT.

10. A vaccine comprising a Shigella bacterium, or an immunogenic fragment or derivative thereof, said Shigella bacterium having an enhanced antigenic property, which is harvested from a culture of a Shigella species grown in vitro in a culture medium with a combination of conditions comprising:

(a) a divalent cation chelator selected from the group consisting of about 1.0 to about 25 μ

M of 1,2-bis(2-aminophenoxy)ethane-N,N,N'"'"',N'"'"'-tetraacetic acid/acetoxymethyl ester, about 0.5 to about 10 mM of ethylene-bis(oxyethylenenitrilo)-tetraacetic acid, and about 1 to about 100 μ

M of ethylene-bis(oxyethylenenitrilo)-tetraacetic acidlacetoxymethyl ester;

(b) at a temperature between about 30°

C. and about 42°

C.; and

(c) in air or a gas mixture, wherein the gas mixture comprises (i) about 5% to about 20% CO₂ with about 80% to about 95% air;

(ii) about 5% to about 20% CO₂ with about 80% to about 95% N₂ ;

or (iii) about 5% to about 10% O₂ with about 10% to about 20% CO₂ with about 70% to about 85% N₂,

wherein said Shigella culture is at about early log phase, between early log phase and stationary phase, or at about stationary phase, and the enhanced antigenic property is a higher level of an immunogenic antigen or a new immunogenic antigen when compared to the antigenic property of bacteria from a culture of the Shigella species grown in brain heart infusion broth.

11. The vaccine according to claim 10, further comprising a pharmaceutically acceptable carrier or diluent.

12. The vaccine according to claim 10, wherein said Shigella bacterium is inactivated.

13. The vaccine according to claim 12, wherein said Shigella bacterium is inactivated by formalin treatment.

14. The vaccine according to claim 10, wherein said vaccine is suitable for mucosal or parenteral or mucosal and parenteral administration.

15. The vaccine according to claim 10, further comprising an adjuvant.

16. The vaccine according to claim 15, wherein the adjuvant is a nontoxic heat labile toxin of E. coli known as mLT.