Compositions comprising recombinant probiotic bacteria and methods of use thereof
Inventors
VEDANTAM, Gayatri • VISWANATHAN, Virinchipuram K. • MALLOZZI, Michael
Assignees
US Department of Veterans Affairs • University of Arizona
Publication Number
US-11839649-B2
Publication Date
2023-12-12
Expiration Date
2036-01-22
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Abstract
The invention features probiotic bacteria expressing Clostridium difficile SlpA, or fragment thereof, and its use for the treatment or prevention of Clostridium difficile infection and gut colonization.
Core Innovation
The invention features probiotic bacteria expressing Clostridium difficile SlpA, or a fragment thereof, and uses thereof for the treatment or prevention of Clostridium difficile infection and gut colonization. The probiotic bacteria include Lactococcus lactis and Lactobacillus acidophilus genetically engineered to express a chimeric SlpA polypeptide comprising a bacterial secretion signal, a C. difficile SlpA variable domain, and a Lactobacillus SlpA cell wall binding domain.
The problem being solved is the lack of effective treatments and preventatives for Clostridium difficile infection and colonization, which is a leading cause of antibiotic-associated diarrhea and imposes substantial healthcare costs. Current antibiotics can alter gut microbiome composition and vaccines do not prevent colonization, thereby creating a need for new therapeutic methods.
Claims Coverage
The patent includes two independent claims covering a host cell and a vector, both optimized for expression of a chimeric SlpA polypeptide in probiotic bacteria.
Host cell comprising an optimized nucleic acid sequence encoding a chimeric SlpA polypeptide
A host cell selected from Lactococcus, Lactococcus lactis, Lactobacillus, Lactobacillus acidophilus, or Lactobacillus casei, comprising a nucleic acid sequence optimized for expression (SEQ ID NO:5) encoding a polypeptide containing a bacterial secretion signal, a C. difficile Surface layer protein A (SlpA) variable domain, and a Lactobacillus SlpA cell wall binding domain, wherein the polypeptide optionally has the amino acid sequence SEQ ID NO:4, and the nucleic acid may be integrated into the chromosome.
Vector comprising an optimized nucleic acid sequence encoding a chimeric SlpA polypeptide for probiotic bacteria
A vector comprising a nucleic acid sequence optimized for expression in Lactococcus, Lactococcus lactis, Lactobacillus, Lactobacillus acidophilus, or Lactobacillus casei (SEQ ID NO:5) encoding a polypeptide with a bacterial secretion signal, C. difficile SlpA variable domain, and Lactobacillus SlpA cell wall binding domain, the vector optionally having the amino acid sequence of SEQ ID NO:4, including Lactococcus or Lactobacillus origins of replication, and sequences identical to fragments of the thyA gene region in Lactobacillus for site-specific homologous recombination.
The claims cover probiotic host cells and vectors optimized for expression of a chimeric SlpA polypeptide combining features from C. difficile and Lactobacillus, enabling stable expression and integration into probiotic bacteria for therapeutic use against C. difficile infection and colonization.
Stated Advantages
Recombinant Lactobacillus expressing chimeric SlpA colonizes the gut and protects against virulent Clostridium difficile challenge, extending survival of treated animals.
Stable integration of the chimeric SlpA-encoding nucleic acid into the probiotic genome allows controlled expression and biological containment via auxotrophic survival dependent on thymine supplementation.
The approach offers a novel, non-antibiotic therapeutic strategy for treatment and prevention of C. difficile infection and colonization, potentially reducing antibiotic-associated dysbiosis.
Documented Applications
Treatment or prevention of Clostridium difficile infection and colonization by administering probiotic bacteria expressing chimeric SlpA to the gut of a subject.
Colonization of the gut of a subject with genetically engineered Lactococcus or Lactobacillus strains expressing a chimeric SlpA polypeptide.
Use of recombinant probiotic bacteria for replacement or modulation of a gut microbiome associated with disease.
Administration of probiotic bacteria expressing chimeric SlpA to subjects undergoing antibiotic treatment to prevent C. difficile infection.
Pharmaceutical or therapeutic compositions comprising recombinant probiotic bacteria expressing chimeric SlpA for oral, rectal, or enteral delivery.
Kits containing probiotic bacteria expressing chimeric SlpA and optionally instructions for treating or preventing Clostridium difficile infection or for gut colonization.
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