Adhesin-enterotoxin chimera based immunongenic composition against enterotoxigenic Escherichia coli
Inventors
Savarino, Stephen J • Holmes, Randall K • Jobling, Michael G
Assignees
US Department of Navy • University of Colorado Denver
Publication Number
US-9107866-B2
Publication Date
2015-08-18
Expiration Date
2027-01-11
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Abstract
The inventive subject matter relates to an immunogenic composition composed of a chimeric molecule including a conformationally stable adhesin from Escherichia coli fused to a bacterial toxin A subunit, such as cholera toxin A subunit or heat-labile Escherichia coli toxin A subunit. The invention also relates to the adhesin-toxin chimera noncovalently associated with a toxin B subunit of the same or different species as the A subunit. The invention also relates to a method of utilizing an adhesin/toxin fusion composition to elicit an immune response.
Core Innovation
The invention relates to an immunogenic composition composed of a chimeric molecule including a conformationally stable adhesin polypeptide from Escherichia coli fused to a bacterial toxin A subunit, such as cholera toxin A subunit or heat-labile Escherichia coli toxin A subunit. The composition may also include the adhesin-toxin chimera noncovalently associated with a toxin B subunit of the same or different species as the A subunit. The invention further provides a method of utilizing such adhesin/toxin fusion compositions to elicit an immune response against enterotoxigenic Escherichia coli (ETEC).
Enterotoxigenic E. coli (ETEC) causes secretory diarrhea, especially in children and travelers to developing countries, leading to millions of cases and hundreds of thousands of deaths yearly. Existing vaccines have not demonstrated adequate efficacy, partly due to challenges in inducing effective anti-adhesive immunity and achieving proper immunogenicity without toxicity. ETEC adheres to intestinal cells via fimbriae composed of major and minor subunits, with the minor subunits, such as CfaE, identified as key adhesins responsible for binding and thus critical targets for vaccine design.
The invention addresses the problem of instability and poor immunogenicity of adhesin subunits when used alone by conferring conformational stability through fusing the adhesin polypeptide to a bacterial toxin A subunit, optionally associated with a toxin B subunit, thus forming a chimeric immunogen. This chimeric molecule provides protease resistance and maintains conformational integrity, leveraging the adjuvant and immunomodulatory properties of bacterial toxins like cholera toxin or heat-labile E. coli enterotoxin to increase the induction of protective immune responses that inhibit bacterial colonization and neutralize enterotoxin activity.
Claims Coverage
The patent includes multiple inventive features detailed in independent claims that describe immunogenic compositions comprising fusion proteins and methods of inducing immune responses against ETEC.
Fusion protein comprising E. coli fimbrial adhesin operatively connected to toxin A subunit
An immunogenic composition containing a purified fusion protein where an E. coli minor fimbrial adhesin polypeptide or monomer derived from Class 5 or CS3 fimbriae is operatively connected, directly or via a linker and donor strand polypeptide, to a toxin A subunit polypeptide.
Conformational stability imparted by linker and donor strand connection
The adhesin polypeptide is connected at its C-terminus to a linker, which is connected at its C-terminus to a donor strand, and then to the toxin A subunit, conferring conformational stability to the adhesin within the fusion protein.
Fusion protein noncovalently linked to toxin B subunit
The fusion protein may be noncovalently associated with a toxin B subunit polypeptide, forming a chimeric enterotoxin-like molecule where A and B subunits can be from the same or different bacterial species.
Use of bacterial toxin A and B subunits derived from cholera or heat-labile E. coli toxins
The toxin A subunit is selected from group consisting of cholera toxin A, cholera toxin A2, E. coli heat-labile toxin A, or LT A2. The toxin B subunit is derived from cholera toxin B or E. coli heat-labile toxin B.
Inclusion of specific fimbrial adhesins and sequences
The E. coli fimbrial adhesin selected from colonization factor antigen I, CS4, CS14, CS1, PCF071, CS17, CS19, CS2, and CS3, including specific subunits CfaE, CsfD, CsuD, CooD, CosD, CsdD, CsbD, CotD, and CstH, and optionally including histidine tags and signal sequences for expression and purification.
Method of inducing immune response using the fusion protein composition
A method comprising administration of a priming dose and subsequent boosting doses of the immunogenic composition containing the fusion protein, in an amount from 50 μg to 1 mg, by various routes including subcutaneous, transdermal, intramuscular, oral, transcutaneous, or nasal, to elicit an immune response that inhibits ETEC fimbriae adherence and reduces or prevents diarrhea.
Expression of fusion proteins via DNA expression systems in live attenuated bacterial vectors
The fusion proteins can be produced by DNA constructs inserted into live attenuated bacterial vectors such as E. coli, Shigella, Campylobacter, Salmonella, or Vibrio prior to administration as vaccines.
The independent claims collectively cover fusion proteins composed of conformationally stable E. coli adhesins fused to bacterial toxin A subunits, optionally associated with toxin B subunits, and methods for using these chimeras to induce protective immune responses against ETEC in various formulations and administration routes.
Stated Advantages
Conformationally stable adhesin-toxin fusion proteins provide increased protease resistance and immunogenicity of adhesins.
The fusion to bacterial toxin A and B subunits confers immunomodulatory and adjuvant effects, improving vaccine efficacy.
The composition induces functional anti-adhesive antibodies that inhibit ETEC colonization.
The chimera elicits antibody responses that can neutralize heat-labile enterotoxins and reduce diarrheal disease.
Documented Applications
Use of the adhesin-toxin chimeric composition for inducing an immune response against enterotoxigenic Escherichia coli to prevent or reduce diarrheal disease.
Administration as a vaccine either subcutaneously, transdermally, intramuscularly, orally, transcutaneously, or nasally.
Production of the fusion protein via DNA expression in live attenuated bacterial vectors for vaccine delivery.
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