# US 10,765,695 B2 · Methods and compositions for selecting siRNA of improved functionality

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- Number: US 10,765,695 B2
- Document ID: US10765695B2
- Status: Granted
- Kind: B2
- Application number: 15/823,415
- Filed: 2017-11-27
- Published: 2018-08-02
- Granted: 2020-09-08
- Earliest claimed: 2002-11-14
- Assignees: Thermo Fisher Scientific Inc.
- Applicants: Thermo Fisher Scientific Inc.
- Inventors: Anastasia Khvorova; Angela Reynolds; Devin Leake; William Marshall; Steven Read; Stephen Scaringe
- Web page: https://patentgrep.com/patent/US10765695B2

## CPC

- A61K 31/713: Double-stranded nucleic acids or oligonucleotides
- C12N 15/1048: {SELEX}
- C12N 15/111: {General methods applicable to biologically active non-coding nucleic acids}
- C12N 15/113: Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; {Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing (when used in plants C12N15/8218)}
- C12N 15/1135: {against oncogenes or tumor suppressor genes}
- C12N 15/1136: {against growth factors, growth regulators, cytokines, lymphokines or hormones}
- C12N 15/1137: {against enzymes (viral enzymes C12N15/1131; receptors C12N15/1138)}
- C12N 15/1138: {against receptors or cell surface proteins}
- C12N 15/8218: {Antisense, co-suppression, viral induced gene silencing [VIGS], post-transcriptional induced gene silencing [PTGS]}
- C12Y 113/12007: Photinus-luciferin 4-monooxygenase (ATP-hydrolysing) (1.13.12.7), i.e. firefly-luciferase
- C12Y 502/01008: Peptidylprolyl isomerase (5.2.1.8), i.e. cyclophilin
- C12N 2310/14: interfering nucleic acids [NA]
- C12N 2320/10: in screening processes
- C12N 2320/11: for the determination of target sites, i.e. of active nucleic acids
- C12N 2320/13: in a process of directed evolution, e.g. SELEX, acquiring a new function
- C12N 2330/31: Libraries, arrays
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C12Y: ENZYMES

## Abstract

Efficient sequence specific gene silencing is possible through the use of siRNA technology. Be selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods compositions, and kits generated through rational design of siRNAs are disclosed, including those directed to the nucleotide sequences for AAT.

## Claims

### Claim 1

An siRNA molecule, wherein said siRNA molecule consists of: (a) a duplex region; and (b) either no overhang regions or at least one overhang region, wherein each overhang region contains six or fewer nucleotides, wherein the duplex region consists of a sense region and an antisense region, wherein said sense region and said antisense region together form said duplex region and said sense region and said antisense region are each 19-30 nucleotides in length and said antisense region comprises a sequence that is the complement of SEQ ID NO: 517.

### Claim 2 (depends on 1)

The siRNA molecule of claim 1, wherein said antisense region and said sense region are each 19-25 nucleotides in length.

### Claim 3 (depends on 2)

The siRNA molecule of claim 2, wherein said antisense region and said sense region are each 19 nucleotides in length.

### Claim 4 (depends on 1)

The siRNA molecule of claim 1, wherein said siRNA molecule has at least one overhang region.

### Claim 5 (depends on 1)

The siRNA molecule of claim 1, wherein said siRNA molecule has no overhang regions.

### Claim 6 (depends on 2)

The siRNA molecule of claim 2, wherein said siRNA molecule has at least one overhang region.

### Claim 7 (depends on 2)

The siRNA molecule of claim 2, wherein said siRNA molecule has no overhang regions.

### Claim 8 (depends on 3)

The siRNA molecule of claim 3, wherein said siRNA molecule has at least one overhang region.

### Claim 9 (depends on 3)

The siRNA molecule of claim 3, wherein said siRNA molecule has no overhang regions.

### Claim 10

A chemically synthesized double stranded siRNA molecule, wherein:
(a) each strand of said double stranded siRNA molecule is 19-30 nucleotides in length; and
(b) one strand of said siRNA molecule comprises a sequence that is the complement of SEQ ID NO: 517.

### Claim 11 (depends on 10)

The chemically synthesized double stranded siRNA molecule of claim 10 wherein each strand of said siRNA molecule is 19-25 nucleotides in length.

### Claim 12 (depends on 11)

The chemically synthesized double stranded siRNA molecule of claim 11, wherein each strand of said siRNA molecule is 19 nucleotides in length.