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US 2024/0200229 A1Published application
Compositions and methods for in vivo screening of therapeutics
AssigneeGordian Biotechnology, Inc.
InventorsMartin Borch Jensen, Daniel Fuentes
Life of this patent
Estimating the term…
Abstract
Provided herein are compositions and methods of use thereof for identifying one or more uniquely identifiable therapeutic moieties in vivo by identifying one or more reporters indicative of a cell state. The disclosure provides libraries including one or more expression vectors, each including one or more expression cassette including a PolIII promoter, a molecular therapeutic barcode, and a capture sequence, for example, a P3TM element.
Claims24
A library comprising:
- one or more expression vectors, each comprising an expression cassette comprising:
- i) (a) a polymerase II promoter operably linked to a nucleic acid sequence encoding one or more reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell;
- and (b) a polymerase III promoter operably linked to a nucleic acid sequence encoding an shRNA, a therapeutic moiety barcode, and a capture sequence;
- or
- (ii) (a) a polymerase II promoter operably linked to a nucleic acid sequence encoding one or more reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell;
- and (b) a polymerase III promoter operably linked to a nucleic acid sequence encoding an shRNA, a therapeutic moiety barcode, and a capture sequence, wherein the polymerase III promoter is oriented downstream of the polymerase II promoter on the expression vector.
3 more elements
3-4. (canceled)
A library comprising:
- one or more expression vectors, each comprising:
- (i)(a) a first expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding one or more reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell, and a therapeutic transgene;
- and (b) a second expression cassette comprising a polymerase III promoter operably linked to a therapeutic moiety barcode and a capture sequence;
- (ii) (a) a first expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding one or more reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell, and one or more miRNA;
- and (b) a second expression cassette comprising a polymerase III promoter operably linked to a therapeutic moiety barcode and a capture sequence;
- (iii) (a) a first expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding one or more reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell, and a therapeutic transgene;
- and (b) a second expression cassette comprising a polymerase III promoter operably linked to a therapeutic moiety barcode and a capture sequence, wherein the one or more expression vectors comprise two, three, four, five, or more than five copies of the second expression cassette;
- (iv) (a) a first expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding one or more reporters that collectively are indicative of a likelihood of a cell state of a cell, and a downstream polymerase III promoter operably linked to a nucleic acid sequence encoding an shRNA;
- and (b) a second expression cassette comprising a polymerase III promoter operably linked to a therapeutic moiety barcode and a capture sequence;
- (v) (a) a first expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding one or more reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell, and a downstream polymerase III promoter operably linked to a nucleic acid sequence encoding an shRNA;
- and (b) a second expression cassette comprising a polymerase III promoter operably linked to a therapeutic moiety barcode and a capture sequence, wherein the one or more expression vectors comprise two, three, four, five, or more than five copies of the second expression cassette;
- (vi) (a) a first expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding one or more first reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell, and a downstream polymerase III promoter operably linked to a nucleic acid sequence encoding an sgRNA;
- and (b) a second expression cassette comprising a polymerase III promoter operably linked to a therapeutic moiety barcode and a capture sequence;
- and a second plurality of expression vectors, each comprising:
- (c) a third expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding SaCas9 and a nucleic acid sequence encoding one or more second reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell, wherein the second reporters are different than the first reporters;
- (vii) (a) a first expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding one or more first reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell, and a downstream polymerase III promoter operably linked to a nucleic acid sequence encoding an sgRNA;
- and (b) a second expression cassette comprising a polymerase III promoter operably linked to a therapeutic moiety barcode and a capture sequence;
- and a second plurality of expression vectors, each comprising:
- (c) a third expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding SaCas9 and a nucleic acid sequence encoding one or more second reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell, wherein the second reporters are different than the first reporters, and wherein the one or more first expression vectors comprise two, three, four, five, or more than five copies of the second expression cassette;
- (viii) (a) a first expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding one or more first reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell;
- and (b) a second expression cassette comprising a polymerase III promoter operably linked to a nucleic acid sequence encoding an sgRNA, a therapeutic moiety barcode, and a capture sequence;
- and a second plurality of expression vectors, each comprising:
- (c) a third expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding SaCas9 and a nucleic acid sequence encoding one or more second reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell, wherein the second reporters are different than the first reporters;
- or
- (ix) (a) a first expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding one or more first reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell;
- and (b) a second expression cassette comprising a polymerase III promoter operably linked to a nucleic acid sequence encoding an sgRNA, a therapeutic moiety barcode, and a capture sequence;
- and a second plurality of expression vectors, each comprising:
- (c) a third expression cassette comprising a polymerase II promoter operably linked to a nucleic acid sequence encoding SaCas9 and a nucleic acid sequence encoding one or more second reporters that collectively, when expressed in a cell, are indicative of a likelihood of a cell state of a cell;
- wherein the second reporters are different than the first reporters, and wherein the first plurality of expression vectors comprises two, three, four, five, or more than five copies of the second expression cassette.
26 more elements
(canceled)
10-24. (canceled)
26-28. (canceled)
Dependencies are inferred from the claim text.