Antibacterial composition effective in treating gram negative bacterial infections and method for preparing the same
Inventors
Yoon, Seong Jun • JUN, Soo Youn • PARK, Jeong Won • Kim, Ji Hyun • KIM, Saet Byeol • Kang, Sang Hyeon
Assignees
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Publication Number
US-11548917-B1
Publication Date
2023-01-10
Expiration Date
Abstract
A pharmaceutical composition for treating Gram negative bacteria-associated infections includes an antibacterial protein that includes at least one selected from the group consisting of a protein having the amino acid sequence as set forth in SEQ ID NO: 1, a protein having the amino acid sequence as set forth in SEQ ID NO: 2, a protein having the amino acid sequence as set forth in SEQ ID NO: 3, a protein having the amino acid sequence as set forth in SEQ ID NO: 4, a protein having the amino acid sequence as set forth in SEQ ID NO: 5, a protein having the amino acid sequence as set forth in SEQ ID NO: 6, a protein having the amino acid sequence as set forth in SEQ ID NO: 7, and a protein having the amino acid sequence as set forth in SEQ ID NO: 8. A method of preparing the antibacterial protein is also disclosed.
Core Innovation
The disclosure describes an antibacterial protein set comprising antibacterial proteins AP-P-1 through AP-P-8, each defined by an amino-acid sequence as set forth in SEQ ID NO: 1 through SEQ ID NO: 8. The antibacterial proteins are engineered for enhanced Gram-negative outer-membrane penetration, and the document further describes a corresponding pharmaceutical composition that includes at least one of the antibacterial proteins for treating Gram-negative bacterial infections.
The problem addressed is Gram negative bacteria-associated infections, including pneumonia, peritonitis, urinary tract infections, bloodstream infections, wound or surgical site infections, and meningitis. The described approach is positioned for effective activity against Gram-negative bacteria, including antibiotic-resistant Gram-negative bacteria.
The document also describes a method of preparing the antibacterial proteins by culturing Escherichia coli cells including plasmids that comprise sequences as set forth in SEQ ID NO: 17 through SEQ ID NO: 24. The method includes inducing the expression of the antibacterial protein, recovering an inclusion body, solubilizing the inclusion body, refolding the antibacterial protein, and purifying the antibacterial protein.
In supported embodiments, the antibacterial proteins show Gram-negative-specific activity, including bacteriolysis and activity against Pseudomonas aeruginosa, Acinetobacter baumannii, and Klebsiella pneumoniae, while not showing activity against other tested bacteria. Experimental support described includes purification of the proteins and in vivo therapeutic efficacy in a mouse infection model with improved survival and reduced mortality.
Claims Coverage
The partial content includes two independent claims: clm-00001 and clm-00012. Both independent claims are grounded in sequence-defined antibacterial proteins (SEQ ID NO: 1-8) and, for the method claim, specific plasmid sequences (SEQ ID NO: 17-24), with the composition claim additionally covering Gram-negative infection indications and formulation constraints.
Gram-negative antibacterial protein pharmaceutical composition
A pharmaceutical composition for treating Gram negative bacteria-associated infections, comprising an antibacterial protein that includes at least one selected from the group consisting of a protein having the amino acid sequence as set forth in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8.
E. coli inclusion body processing method for sequence-defined antibacterial proteins
A method of preparing an antibacterial protein that includes at least one selected from the group consisting of a protein having the amino acid sequence as set forth in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8, comprising culturing Escherichia coli cells including a plasmid that comprises a sequence as set forth in SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, or SEQ ID NO: 24; inducing the expression of the antibacterial protein; recovering an inclusion body; solubilizing the inclusion body; refolding the antibacterial protein; and purifying the antibacterial protein.
Across the independent claims, the core claimed subject matter is a pharmaceutical composition containing at least one sequence-defined antibacterial protein (SEQ ID NO: 1-8) for Gram negative bacteria-associated infections, and a corresponding E. coli-based preparation method that uses plasmids containing sequence sets (SEQ ID NO: 17-24) followed by inclusion body recovery, solubilizing, refolding, and purification.
Stated Advantages
Effective against antibiotic-resistant Gram-negative bacteria.
Gram-negative-specific lysis, with no activity against other tested bacteria.
Improved survival and reduced mortality in a mouse infection model.
Documented Applications
Treating Gram negative bacteria-associated infections including pneumonia, peritonitis, urinary tract infections, bloodstream infections, wound or surgical site infections, and meningitis.
Antibacterial activity against antibiotic-resistant Gram-negative bacteria, including Pseudomonas aeruginosa, Acinetobacter baumannii, and Klebsiella pneumoniae.
Preparing antibacterial proteins by culturing Escherichia coli cells and processing inclusion bodies to obtain purified antibacterial protein.
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