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Una oligomers having reduced off-target effects in gene silencing

a technology of una oligomers and gene silencing, applied in the field of gene silencing techniques, can solve the problems of difficult to carry out, rna interference is the occurrence of off-target effects, and the silencing technique is difficult to achieve, so as to reduce ttr and off-target effects

Inactive Publication Date: 2015-10-29
ARCTURUS THERAPEUTICS
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

The patent describes a new type of active agent called UNA oligomers that can be used for gene silencing with reduced off-target effects. These oligomers can be particularly useful for treating transthyretin-related amyloidosis, a disease caused by the buildup of a certain protein. The UNA oligomers have a more specific structure and can inhibit the expression of the target gene with at least 10% less off-target effects than traditional methods like siRNA. This new technology has the potential to improve the effectiveness and safety of gene silencing therapies.

Problems solved by technology

A major drawback of gene silencing techniques such as RNA interference is the occurrence of off-target effects.
Off-target effects occur when a gene-silencing agent has an effect on a gene product to which it is not targeted, and is therefore unwanted.
Off-target effects loom as one of the main hurdles for developing gene silencing therapeutics.
These conventional methods have been able to reduce off-target effects, however, they can be difficult to carry out.

Method used

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  • Una oligomers having reduced off-target effects in gene silencing
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  • Una oligomers having reduced off-target effects in gene silencing

Examples

Experimental program
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Effect test

example 1

[0169]This example shows that UNA oligomers dramatically reduce off target activity of the passenger strand in gene silencing by RNA interference. The reduction in passenger strand off target activity can depend on the positioning of various UNA monomers in the oligomer. In this example, it is shown that the presence of a combination of UNA monomers in three positions in a UNA oligomer, more specifically, in the passenger strand at the 5′ end and at the 3′ end, as well as in the guide strand at the 3′ end, greatly reduced off target knockdown activity by the passenger strand.

[0170]UNA oligomers targeted to ApoCIII having reduced off-target effects are shown in Table 3. As used herein, a duplex oligomer is represented with the passenger strand above, and the guide strand below. The end group numbering will depend on the identity of the terminal monomer, as described above.

TABLE 3UNA oligomers ATX98 and ATX100SEQ IDNO:OLIGOMER33ATX1-ÃAAAGGGACAGUAUUCUCAÛmU-3′34983′-mUÛUUUUCCCUGUCAUAAGA...

example 2

[0175]FIG. 3 shows that certain UNA oligomers had at least comparable knockdown levels of activity to conventional siRNAs for TTR mRNA expression. ATX13, ATX14, ATX15, ATX16, ATX17, ATX21 and ATX25 were targeted to the 3′-UTR of human TTR, and therefore were targeted to both wild-type V30V and V30M mutant TTR.

[0176]In particular, UNA oligomer ATX13 having a UNA monomer in the first strand located at the 1 (5′) end, and UNA oligomer ATX15 having a UNA monomer in the first strand located at the 1 (5′) end and two UNA monomers in the second strand located at the 3 (3′) end in the 20th and 21st positions counting from the 5′ end, had at least comparable knockdown levels of activity as compared to conventional siRNA ATS-91.

[0177]Further, UNA oligomer ATX21 having a UNA monomer in the first strand located at the 1 (5′) end, one UNA monomer in the first strand located at the 3 (3′) end in the 20th position counting from the 5′ end, and one UNA monomer in the second strand located at the 3 ...

example 3

[0180]FIG. 5 shows the protocol for measuring off-target (OT) effects. A Luciferase reporter assay using PSICHECK vector was established. For each measurement, 4 plasmids were constructed: guide strand GSCM and guide strand GSSM for second strand or antisense knockdown, and passenger strand PSCM and passenger strand PSSM for first strand or sense strand knockdown. The reporter plasmid was co-transfected with UNA oligomer into HeLa cells. In this system, if the UNA oligomer binds to the target sequence inserted in 3-UTR of luciferase, then the chemiluminescent signal is reduced or disappeared.

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Abstract

This invention provides UNA oligomers for gene silencing with reduced off-target effects. The UNA oligomers can have a first strand and a second strand, each of the strands being 19-29 monomers in length, the monomers being UNA monomers and various nucleic acid monomers. Embodiments include pharmaceutical compositions and methods for treating or preventing TTR-related amyloidosis with reduced off-target effects by administering a UNA oligomer to a subject.

Description

SEQUENCE LISTING[0001]This application includes a Sequence Listing submitted electronically herewith as an ASCII file created on Mar. 24, 2015, named ARC1246WO_SL.txt, which is 30,972 bytes in size, and is hereby incorporated by reference in its entirety.BACKGROUND OF THE INVENTION[0002]A major drawback of gene silencing techniques such as RNA interference is the occurrence of off-target effects. Off-target effects occur when a gene-silencing agent has an effect on a gene product to which it is not targeted, and is therefore unwanted. Off-target effects loom as one of the main hurdles for developing gene silencing therapeutics.[0003]One method to reduce off-target effects is to intelligently design the structure of the gene silencing agent to avoid effects on genes other than the desired target. In some cases, the gene silencing agents could be pooled so that the concentration of agents having a particular off-target effect would be reduced. In other cases, the gene silencing agent ...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): C12N15/113A61K47/14A61K31/713
CPCC12N15/113A61K31/713C12N2310/32C12N2310/14A61K47/14C12N2310/323A61P25/00A61P25/02A61P25/28A61P43/00A61P5/14A61P9/00
Inventor TACHIKAWA, KIYOSHIPAYNE, JOSEPH E.CHIVUKULA, PADMANABH
Owner ARCTURUS THERAPEUTICS
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