US 11,352,641 B2 · Circular RNA for translation in eukaryotic cells

Abstract

Methods and constructs for engineering circular RNA are disclosed. In some embodiments, the methods and constructs comprise a vector for making circular RNA, the vector comprising the following elements operably connected to each other and arranged in the following sequence: a.) a 5′ homology arm, b.) a 3′ group I intron fragment containing a 3′ splice site dinucleotide, c.) optionally, a 5′ spacer sequence, d.) a protein coding or noncoding region, e.) optionally, a 3′ spacer sequence, f) a 5′ Group I intron fragment containing a 5′ splice site dinucleotide, and g.) a 3′ homology arm, the vector allowing production of a circular RNA that is translatable or biologically active inside eukaryotic cells. Methods for purifying the circular RNA produced by the vector and the use of nucleoside modifications in circular RNA produced by the vector are also disclosed.

Claims26

Dependent claims

A method for making a cell population comprising circular RNA, said method comprising contacting the cells in the population with a composition comprising:

a) circular RNA produced from a vector comprising the following elements:
i) a 5′ homology arm,
ii) a 3′ Group I self-splicing intron fragment containing a 3′ splice site dinucleotide,
iii) a 5′ spacer sequence,
iv) an internal ribosome entry site (IRES),
v) a protein coding region encoding a chimeric antigen receptor (CAR),
vi) a 3′ spacer sequence,
vii) a 5′ Group I self-splicing intron fragment containing a 5′ splice site dinucleotide, and
viii) a 3′ homology arm,
said vector allowing production of a circular RNA that is translatable or biologically active inside eukaryotic cells, and
b) a delivery vehicle for delivering the circular RNA to the cells, said delivery vehicle comprising a nanocarrier selected from the group consisting of a lipid, a polymer and a lipo-polymeric hybrid,
wherein the circular RNA is delivered into the cells in vitro, trans-arterially, subcutaneously, intratumorally, intramedullary, intranodally, intravenously, intrathecally or intraperitoneally.

9 more elements

A method of making a cell population comprising circular RNA, said method comprising contacting the cells in the population with a composition comprising circular RNA produced from a vector, wherein the circular RNA comprises in the following order, a portion of a 3′ Group I self-splicing intron fragment, an Internal Ribosome Entry Site (IRES), a protein encoding region encoding a chimeric antigen receptor (CAR), and a portion of a 5′ Group I self-splicing intron fragment, wherein said vector allows the production of a circular RNA that is translatable or biologically active inside eukaryotic cells, and a delivery vehicle for delivering the circular RNA to the cells, said delivery vehicle comprising a nanocarrier selected from the group consisting of a lipid, a polymer and a lipo-polymeric hybrid, wherein the circular RNA is delivered into the cells in vitro, trans-arterially, subcutaneously, intratumorally, intramedullary, intranodally, intravenously, intrathecally or intraperitoneally.

Dependencies are inferred from the claim text.

Circular RNA for translation in eukaryotic cells

Life of this patent

Circular RNA for translation in eukaryotic cells

Life of this patent

Abstract

Claims26