Multiple host range bacteriophage with hybrid tail fibres
Inventors
Fairhead, Heather • Wilkinson, Adam • Pitts, Katy • BARNARD, Anne • SEVERI, Emmanuele • Anderson, Neil
Assignees
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Publication Number
US-11492601-B2
Publication Date
2022-11-08
Expiration Date
Abstract
Modified bacteriophage, uses thereof, and compositions containing the modified bacteriophage are described. The compositions are useful for human treatment and may treat various conditions, including bacterial infections.
Core Innovation
The invention relates to a modified bacteriophage that is capable of infecting a plurality of different target bacteria and is non-lytic. The modified bacteriophage includes an α/β small acid-soluble spore protein (SASP) gene encoding a SASP that is toxic to the target bacteria, and includes an inactivated lysis gene. The bacteriophage expresses a hybrid host range determinant protein configured to broaden infectivity across multiple target bacteria.
The hybrid host range determinant protein comprises a tail fibre protein having a receptor binding region for binding to the target bacteria and a region linking the receptor binding region to the body of the bacteriophage. The receptor binding region is a C-terminal receptor binding region, and the region linking the C-terminal receptor binding region to the body is an N-terminal region. Each of the C-terminal and N-terminal regions is from a different bacteriophage than the other.
In particular embodiments, the N-terminal region comprises amino acids 1 to 628 of the tail fibre protein and the C-terminal region comprises amino acids 629 to 964 of the tail fibre protein based on the amino acid sequence of bacteriophage Phi33. The C-terminal region has no more than 96% amino acid sequence identity with the C-terminal region of bacteriophage Phi33, and the N-terminal region has at least 95% amino acid sequence identity with the N-terminal region of bacteriophage Phi33. The disclosed framework further enables hybrid tail fibre transplantation between Phi33 and different PB1-like phages to produce hybrid tail fibre proteins.
The invention also targets Pseudomonas, including a modified bacteriophage capable of infecting a plurality of different Pseudomonas target bacteria, while maintaining the non-lytic character and the toxic α/β SASP gene. The described platform is implemented with a hybrid host range determinant protein derived from different bacteriophages, and sequence identity constraints are used to define allowable hybrid tail fibre designs. The partial content further indicates engineered SASPject derivatives/mixes and uses for medicaments and decontamination.
Claims Coverage
The independent claims in the provided set are clm-00001, clm-00004, and clm-00005. Across these independent claims, three main inventive features are repeated: a non-lytic modified bacteriophage with an inactivated lysis gene carrying a toxic α/β SASP gene, and a hybrid host range determinant tail fibre protein with N-terminal (linking) and C-terminal (receptor binding) regions from different bacteriophages, including Phi33-based segment boundaries and sequence-identity constraints.
Non-lytic modified bacteriophage with inactivated lysis and toxic α/β SASP gene
A modified bacteriophage capable of infecting a plurality of different target bacteria, wherein the bacteriophage includes an α/β small acid-soluble spore protein (SASP) gene encoding a SASP which is toxic to the target bacteria; wherein the bacteriophage is non-lytic; and wherein the bacteriophage comprises an inactivated lysis gene.
Hybrid host range determinant tail fibre with donor-defined N- and C-terminal regions
The bacteriophage expresses a hybrid host range determinant protein which comprises an amino acid sequence from a plurality of different bacteriophages; the hybrid host range determinant protein comprises a tail fibre protein comprising a receptor binding region for binding to the target bacteria and a region linking the receptor binding region to the body of the bacteriophage; wherein the receptor binding region is a C-terminal receptor binding region and the region linking the C-terminal receptor binding region to the body of the bacteriophage is an N-terminal region; and wherein each of the C-terminal and N-terminal regions are from a different bacteriophage than each other.
Phi33-based tail fibre segment boundaries and identity constraints for hybridization
The N-terminal region comprises amino acids 1 to 628 of the tail fibre protein and the C-terminal region comprises amino acids 629 to 964 of the tail fibre protein, based on the amino acid sequence of bacteriophage Phi33; and/or the C-terminal region has no more than 96% amino acid sequence identity with the C-terminal region of bacteriophage Phi33; and wherein the N-terminal region has at least 95% amino acid sequence identity with the N-terminal region of bacteriophage Phi33.
Pseudomonas-targeting scope for non-lytic SASP-expressing hybrid HRD phage
A modified bacteriophage capable of infecting a plurality of different Pseudomonas target bacteria, which bacteriophage includes an α/β small acid-soluble spore protein (SASP) gene encoding a SASP which is toxic to the target bacteria; wherein the bacteriophage is non-lytic; and wherein the bacteriophage expresses a hybrid host range determinant protein implemented as a hybrid tail fibre protein having a C-terminal receptor binding region and an N-terminal region from different bacteriophages with Phi33-based segment boundaries and sequence identity constraints.
Hybrid tail fibre determinant with N- and C-terminal regions from different bacteriophages under Phi33 identity thresholds
A modified bacteriophage capable of infecting a plurality of different target bacteria, wherein the bacteriophage expresses a hybrid host range determinant protein with an N-terminal region and a C-terminal region from different bacteriophages; wherein the N-terminal region comprises amino acids with at least 95% identity to amino acids 1 to 628 of the tail fibre protein of bacteriophage Phi33 and the C-terminal region comprises amino acids corresponding to amino acids 629 to 964 of the tail fibre protein of bacteriophage Phi33; and wherein the C-terminal region has no more than 96% amino acid sequence identity with the C-terminal region of amino acids 629 to 964 of bacteriophage Phi33.
The independent claims define a non-lytic modified bacteriophage that carries an inactivated lysis gene and a toxic α/β SASP gene, and that expresses a hybrid host range determinant tail fibre protein. The tail fibre protein is constructed from donor bacteriophages by assigning an N-terminal linking region and a C-terminal receptor binding region from different bacteriophages. The claims further restrict the hybrid designs using Phi33-based segment boundaries and specified amino acid sequence identity thresholds, with one independent claim explicitly limiting the target bacteria to Pseudomonas.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Not explicitly described in patent.
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