# US 11,439,692 B2 · Method of treating diseases associated with MYD88 pathways using CRISPR-GNDM system

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- Number: US 11,439,692 B2
- Document ID: US11439692B2
- Status: Granted
- Kind: B2
- Application number: 16/614,145
- Filed: 2018-05-17
- Published: 2021-08-26
- Granted: 2022-09-13
- Earliest claimed: 2017-05-17
- Assignees: Modalis Therapeutics Corporation
- Applicants: Modalis Therapeutics Corporation
- Inventors: Tetsuya Yamagata; Yuanbo Qin; Iain Robert Thompson; Haruhiko Morita
- Web page: https://patentgrep.com/patent/US11439692B2

## CPC

- A61K 38/465: {acting on ester bonds (3.1), e.g. lipases, ribonucleases}
- A61P 1/16: for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
- A61K 31/7105: Natural ribonucleic acids, i.e. containing only riboses attached to adenine, guanine, cytosine or uracil and having 3'-5' phosphodiester links
- A61P 29/00: Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
- A61P 31/00: Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P 35/00: Antineoplastic agents
- A61P 37/06: Immunosuppressants, e.g. drugs for graft rejection
- C07K 14/47: from mammals
- C07K 14/705: Receptors; Cell surface antigens; Cell surface determinants {(tumour specific antigens C07K14/4748)}
- C12N 9/226: {Class 2 CAS enzyme complex, e.g. single CAS protein}
- C12N 15/11: DNA or RNA fragments; Modified forms thereof (DNA or RNA not used in recombinant technology, C07H21/00); {Non-coding nucleic acids having a biological activity}
- C12N 15/90: Stable introduction of foreign DNA into chromosome
- A61K 48/00: Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
- C07K 2319/09: containing a nuclear localisation signal
- C07K 2319/42: containing a HA(hemagglutinin)-tag
- C12N 2740/16043: viral genome or elements thereof as genetic vector
- 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)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

The present invention provides a method of treating a disease associated with activated MYD88 signaling in a subject, including suppressing MYD88 expression in a subject by targeting an expression regulatory region of MYD88 gene by using a CRISPR-Guide Nucleotide Directed Modulation (GNDM), without affecting the expression of an adjacent ACAA1 gene. Also, provided is a CRISPR-GNDM system for suppressing MYD88, including expression comprising (a) a protein selected from the group consisting of dCas9 or dCpf1, a fusion protein of dCas9 or dCpf1 and Kruppel associated box (KRAB) and (b) a guide RNA (gRNA) targeting an expression regulatory region of MYD88 gene.

## Claims

### Claim 1

A CRISPR-GNDM system for suppressing MYD88 expression comprising (a) a protein selected from the group consisting of dCas9 or dCpf1, and a fusion protein of dCas9 or dCpf1 and Kruppel associated box (KRAB) or other transcription repressor, and (b) a guide nucleotide (gN) targeting an expression regulatory region of MYD88 gene.

### Claim 2 (depends on 1)

The CRISPR-GNDM system according to claim 1, wherein the protein of (a) is selected from the group consisting of dCas9 or dCpf1, and a fusion protein of dCas9 or dCpf1 and Kruppel associated box (KRAB).

### Claim 3 (depends on 1)

The CRISPR-GNDM system according to claim 1, wherein the expression regulatory region is a region having the nucleotide sequence set forth in SEQ ID NO: 65.

### Claim 4 (depends on 1)

The CRISPR-GNDM system according to claim 1, wherein the expression regulatory region is a region having the nucleotide sequence set forth in positions 1,405-7,352 of SEQ ID NO: 65.

### Claim 5 (depends on 1)

The CRISPR-GNDM system according to claim 1, wherein the gN comprises a nucleotide sequence set forth in any one of SEQ ID NOs:5 to 35.

### Claim 6 (depends on 1)

The CRISPR-GNDM system according to claim 1, wherein the gN comprises a nucleotide sequence set forth in any one of SEQ ID NOs:9 to 35.

### Claim 7

A method of treating a disease associated with MYD88 activation in a subject, comprising suppressing MYD88 expression in the subject by targeting an expression regulatory region of MYD88 gene using a CRISPR-GNDM system.

### Claim 8 (depends on 7)

The method according to claim 7, wherein the MYD88 expression is suppressed without affecting the expression of an adjacent ACAA1 gene.

### Claim 9 (depends on 7)

The method according to claim 7, wherein the disease is selected from the group consisting of an inflammatory disease, an autoimmune disease, an infectious disease and a cancer.

### Claim 10 (depends on 7)

The method according to claim 7, wherein the disease is NASH.

### Claim 11 (depends on 7)

The method according to claim 7, wherein the disease is a hematological malignancy.

### Claim 12 (depends on 7)

The method according to claim 7, wherein the CRISPR-GNDM system comprises (a) a protein selected from the group consisting of dCas9 or dCpf1, and a fusion protein of dCas9 or dCpf1 and Kruppel associated box (KRAB) or other transcription repressor, and (b) a guide nucleotide (gN) targeting an expression regulatory region of MYD88 gene.

### Claim 13 (depends on 12)

The method according to claim 12, wherein the protein of (a) is selected from the group consisting of dCas9 or dCpf1, and a fusion protein of dCas9 or dCpf1 and Kruppel associated box (KRAB).

### Claim 14 (depends on 12)

The method according to claim 12, wherein the expression regulatory region is a region having the nucleotide sequence set forth in SEQ ID NO: 65.

### Claim 15 (depends on 12)

The method according to claim 12, wherein the expression regulatory region is a region having the nucleotide sequence set forth in positions 1,405-7,352 of SEQ ID NO: 65.

### Claim 16 (depends on 12)

The method according to claim 12, wherein the gN comprises a nucleotide sequence set forth in any one of SEQ ID NOs:5 to 35.

### Claim 17 (depends on 12)

The method according to claim 12, wherein the gN comprises a nucleotide sequence set forth in any one of SEQ ID NOs:9 to 35.