Anellovirus compositions and methods of use
Inventors
Delagrave, Simon • Diaz, Fernando Martin • Kahvejian, Avak • Lebo, Kevin James • Nawandar, Dhananjay Maniklal • Pitts, Jared David • Tedstone, Ryan D. • Weinstein, Erica Gabrielle • Yozwiak, Nathan Lawrence
Assignees
Ring Therapeutics Inc • Flagship Pioneering Innovations V Inc • Flagship Pioneering Inc
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Publication Number
US-11446344-B1
Publication Date
2022-09-20
Expiration Date
Abstract
This invention relates generally to viral vectors and viral particles based on Anelloviruses, which can be used to deliver an agent (e.g., an exogenous effector or an endogenous effector, e.g., a therapeutic effector) to a cell (e.g., a cell in a subject to be treated therapeutically). Described herein are anellosomes, anellovectors, and compositions and uses thereof.
Core Innovation
The invention relates to anellovirus-based delivery systems in the form of synthetic anellosomes or anellovectors that comprise a proteinaceous exterior comprising an Anellovirus ORF1 polypeptide. The proteinaceous exterior encapsulates a genetic element that includes a promoter operably linked to a heterologous sequence encoding a therapeutic polypeptide or therapeutic nucleic acid, and the genetic element comprises an Anellovirus 5′ UTR.
The genetic element does not comprise any sequence encoding Anellovirus ORF1, ORF2, or ORF3, or allows nonfunctional forms in certain variants. In related embodiments, the genetic element includes nucleic acid sequences having sequence-identity constraints to portions of an Anellovirus 5′ UTR while still excluding Anellovirus ORF1, ORF2, and ORF3.
The disclosure further describes sequence-identity criteria for Anellovirus ORF1 subsequences by domain, including an Arg-rich region, a jelly-roll domain, a hypervariable region, an N22 domain, and a C-terminal domain, with reference to tabled sequence information. The document also describes engineered anellovirus genomes and characterized anellosomes, including deletion mapping, conserved anellovirus regions and clade categories, variable-length transgene payloads, and payload placement in NCR, miRNA, UTR, and GC-rich regions.
Claims Coverage
The independent claims covered across the input items center on a particle having an Anellovirus ORF1 proteinaceous exterior that encapsulates a promoter-linked genetic element containing an Anellovirus 5′ UTR while excluding, or allowing only nonfunctional versions of, Anellovirus ORF1, ORF2, and ORF3 sequences. Additional claim features include sequence-identity constraints, ORF1 source grouping, and payload-type limitations; in total, the combined claim sets describe multiple overlapping inventive features.
Anellovirus ORF1 proteinaceous exterior encapsulating a promoter-driven therapeutic genetic element with Anellovirus 5′ UTR
A particle comprising a proteinaceous exterior comprising an Anellovirus ORF1 polypeptide and a genetic element encapsulated by the proteinaceous exterior, wherein the genetic element comprises a promoter operably linked to a heterologous sequence encoding a therapeutic polypeptide or nucleic acid, comprises an Anellovirus 5′ UTR, and does not comprise any sequence encoding Anellovirus ORF1, ORF2, or ORF3.
Anellovirus 5′ UTR sequence-identity constraint
The genetic element comprises a nucleic acid sequence comprising at least 90% sequence identity to about 60 consecutive nucleotides of an Anellovirus 5′ UTR.
Nonfunctional Anellovirus ORF1, ORF2, and ORF3 allowance
The genetic element does not comprise any sequence encoding Anellovirus ORF1, ORF2, or ORF3, or comprises nucleic acid sequences encoding nonfunctional Anellovirus ORF1, ORF2, and ORF3.
Promoter-driven therapeutic peptide
The genetic element comprises a promoter operably linked to a heterologous sequence encoding a therapeutic peptide, and the therapeutic peptide comprises a peptide hormone or a peptide ligand.
Therapeutic polypeptide class selection
The therapeutic polypeptide comprises a hormone, a cytokine, an enzyme, an antibody molecule, a transcription factor, a receptor, a ligand, a membrane transporter, a secreted protein, or a nuclease.
Anellovirus ORF1 polypeptide corresponding to Alphatorquevirus, Betatorquevirus, or Gammatorquevirus ORF1
The proteinaceous exterior comprises an Anellovirus ORF1 polypeptide whose ORF1 corresponds to the ORF1 of an Alphatorquevirus, Betatorquevirus, or Gammatorquevirus.
Anellovirus 5′ UTR identity constraint of at least 95% to specified nucleotide ranges
The genetic element comprises a nucleic acid sequence that is at least 95% identical to one of several specified nucleotide ranges from particular SEQ ID NOs.
Across the independent claims, the inventive particle structure centers on an Anellovirus ORF1 proteinaceous exterior encapsulating a genetic element with a promoter linked to a heterologous therapeutic sequence, an included Anellovirus 5′ UTR, and exclusion of Anellovirus ORF1, ORF2, and ORF3 encoding sequences or allowance of nonfunctional ORF-encoding sequences. Additional narrowing includes sequence-identity requirements for the 5′ UTR, ORF1 source grouping, and therapeutic payload type constraints.
Stated Advantages
Substantially non-immunogenic behavior.
Substantially non-pathogenic behavior.
Limited or non-detectable bacterial infection.
Low integration frequency described as non-integrating.
Infect capability for mammalian cells, including in vitro.
Replication potential characterized and quantified, contrasted with replication-deficient constructs.
Includes described immune-response/efficacy comparisons using antibody prevalence relative to AAV.
Substantially non-immunogenic viral delivery.
Non-pathogenic viral delivery.
Documented Applications
Therapeutic use to modulate biological function via delivery of a promoter-driven heterologous therapeutic payload using anellosome or synthetic anellosome particles.
Delivery to specific cell, tissue, and organ targets including bone marrow, blood, heart, GI, skin, liver cells, epithelial cells, and retina photoreceptors.
Delivery to immune cell categories described as immune effector cells including T cell, NK cell, and macrophage engulfment context.
Production and characterization of engineered anellosomes.
Quantification using qPCR for genome equivalents.
In vivo expression in mice, including luciferase expression.
Evaluation of host genome integration and functional miRNA cargo.
Downstream characterization and comparisons including whole-genome sequencing and AAV comparison.
Therapeutic payload delivery using anellovirus-based particles to support payload expression and functional effects in cellular contexts.
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