Targeted and modular exosome loading system
AssigneeNorthwestern University
InventorsJoshua N. Leonard, Michelle E. Hung
Life of this patent
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Abstract
Disclosed are exosomes that include a packaging protein and a cargo RNA in which the packaging protein binds specifically to the cargo RNA. The packaging protein is a fusion protein that includes an RNA-binding domain and an exosome-targeting domain. The cargo RNA includes an RNA-motif that the RNA-binding domain of the fusion protein binds specifically such that the cargo RNA is packaged in the lumen of the exosomes.
Claims11
An exosome comprising a packaging protein and a cargo RNA that binds to the packaging protein, wherein the packaging protein is a fusion protein comprising an RNA-binding domain and an exosome-targeting domain that targets the fusion protein to the membrane of the exosome, wherein the exosome-targeting domain is a domain of a lysosome membrane protein having a luminal N-terminus and a cytoplasmic C-terminus, and wherein the cargo RNA comprises more than one binding loop and the RNA-binding domain of the fusion protein binds specifically to the binding loops of the cargo RNA, and
- further wherein the RNA-binding domain is the RNA-binding domain of MS2 bacteriophage coat protein comprising SEQ ID NO:2 or a variant thereof having at least 80% amino acid sequence identity to SEQ ID NO:2, and wherein the binding loops comprise a sequence and structure selected from the group consisting of:
- where N-N is any two base-paired RNA nucleotides.
A kit comprising:
- (a) a vector for expressing a fusion protein comprising an RNA-binding domain and an exosome-targeting domain that targets the fusion protein to the membranes of exosomes, wherein the exosome-targeting domain is a domain of a lysosome membrane protein having a luminal N-terminus and a cytoplasmic C-terminus, and (b) a vector for expressing a cargo RNA comprising more than one binding loop that bind to the RNA-binding domain of the fusion protein, wherein the RNA-binding domain is the RNA-binding domain of MS2 bacteriophage coat protein comprising SEQ ID NO:2 or a variant thereof having at least 80% amino acid sequence identity to SEQ ID NO:2, and wherein the binding loops comprise a sequence and structure selected from the group consisting of:
- SEQ ID NO:
- 4
- U U
- A A
- N-N
- N-N
- A
- N-N
- N-N
- N-N
- N-N
- SEQ ID NO:
- 5
- C U
- A A
- N-N
- N-N
- A
- N-N
- N-N
- N-N
- N-N,
- and
- SEQ ID NO:
- 6
- U C
- A A
- N-N
- N-N
- A
- N-N
- N-N
- N-N
- N-N,
- where N-N is any two base-paired RNA nucleotides.
Dependencies are inferred from the claim text.