Single-vector type I vectors
Inventors
MARTINEZ, Virginia • VAZQUEZ-URIBE, Ruben • TAKOS, Adam • VAN DER HELM, Eric
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Assignees
MemberSNIPR BiomeSNIPR BiomeSNIPR Biome develops precision medicines for infections and diseases linked to antimicrobial resistance and microbiome dysbiosis, using CRISPR-based gene editing to selectively target pathogenic bacteria while sparing the rest of the microbial community. The company applies programmable CRISPR platforms, delivered by engineered phages or probiotics, to address difficult-to-treat infections and modulate the human microbiome. Its programs span discovery through Phase 2 clinical trials, with a significant patent portfolio supporting these innovations.
SNIPR Biome develops precision medicines for infections and diseases linked to antimicrobial resistance and microbiome dysbiosis, using CRISPR-based gene editing to selectively target pathogenic bacteria while sparing the rest of the microbial community. The company applies programmable CRISPR platforms, delivered by engineered phages or probiotics, to address difficult-to-treat infections and modulate the human microbiome. Its programs span discovery through Phase 2 clinical trials, with a significant patent portfolio supporting these innovations.
Publication Number
US-11851663-B2
Publication Date
2023-12-26
Expiration Date
Abstract
The invention relates to the production and use of Cas-encoding sequences and vectors comprising these. Aspects of the invention provide products, vectors, delivery vehicles, uses and methods for producing Cas-encoding sequences in bacterial or archaeal cells.
Core Innovation
A production strain bacterial cell is provided that comprises a nucleic acid vector for introduction into a target bacterial host cell. In the target bacterial host cell, the vector expresses Type I Cas3 and one or more cognate Cascade proteins, and includes a CRISPR array for producing crRNAs or one or more nucleotide sequences encoding one or more guide RNAs, where each spacer sequence is complementary to a target sequence of the target bacterial host cell.
The vector controls expression of Type I Cas3 in the target bacterial host cell with a promoter having a strength weaker than the Anderson Score strength of promoter BBa_J23108. The production strain bacterial cell does not comprise a crRNA or gRNA operable with the Cas3 to target and cut a chromosomal sequence of the production strain cell.
The disclosure also describes delivery of Type I Cas3 and Cascade through delivery vehicles, including phage particles or non-self-replicative transduction particles, and pharmaceutical uses. Additional aspects include promoter attenuation and repression strategies in production strains to manage Cas expression and related effects, including reduced DNA loss and reduced Cas toxicity, mutation, viability loss, and unwanted Cas cutting.
Claims Coverage
1 independent claim is explicitly provided in the partial content. It contains four inventive features that define the production strain bacterial cell, the Type I Cas3 and Cascade expression vector architecture, the promoter strength constraint, and the limitation on operable crRNA/gRNA in the production strain.
Production strain with vector expressing Type I Cas3 and cognate Cascade proteins
A production strain bacterial cell comprising a nucleic acid vector for introduction into a target bacterial host cell for expression of Type I Cas3 and one or more cognate Cascade proteins in the target bacterial host cell, with a first nucleotide sequence encoding a Type I Cas3 and a second nucleotide sequence encoding one or more cognate Cascade proteins.
Promoter strength weaker than Anderson Score BBa_J23108
The first nucleotide sequence is under the control of a promoter for controlling the expression of Type I Cas3 in the target bacterial host cell, wherein the promoter has a strength weaker than the Anderson Score strength of promoter BBa_J23108.
Target host bacterial selection and compatible CRISPR targeting elements
The target bacterial host cell is selected from Fusobacteria, Bacteroides, Staphylococcus, Clostridium, Lactobacillus, Bacillus, Escherichia, Streptococcus, Streptomyces, Pseudomonas, and Klebsiella, and the nucleic acid vector further comprises either a CRISPR array for producing crRNAs in the target bacterial host cell or nucleotide sequences encoding one or more guide RNAs, wherein each spacer sequence is complementary to a target sequence of the target bacterial host cell.
Production strain lacks operable crRNA/gRNA for cutting its own chromosome
The production strain bacterial cell does not comprise a crRNA or gRNA operable with the Cas3 to target and cut a chromosomal sequence of the production strain cell.
The independent claim centers on a production strain carrying a vector that expresses Type I Cas3 and cognate Cascade in a selected target bacterial host, while using a promoter strength constraint weaker than Anderson Score BBa_J23108 and ensuring the production strain itself lacks operable crRNA/gRNA for Cas3 cutting of its chromosome.
Stated Advantages
Documented Applications
No documented applications found
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