Delivery and formulation of engineered nucleic acids

• US 9,533,047 B2
• Filed: 07/22/2014
• Issued: 01/03/2017
• Est. Priority Date: 03/31/2011
• Status: Active Grant

• Current Assignee: Moderna TX, Inc. (Moderna, Inc.)
• Original Assignee: Moderna TX, Inc. (Moderna, Inc.)
• Inventors: de Fougerolles, Antonin, Elbashir, Sayda M.
• Primary Examiner(s): Kelly, Robert M
• Application Number:
  US14/337,513
• Publication Number:
  US 20150017211A1
• Time in Patent Office: 896 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: A61K 31/7088 Compounds having three or more nucleosides or nucleotides
• CPC Class Codes: A61K 31/7115 Nucleic acids or oligonucleotides having modified bases, i.e. other than adenine, guanine, cytosine, uracil or thymine
• CPC Class Codes: A61K 38/193 Colony stimulating factors [CSF]
• CPC Class Codes: A61K 38/4846 Factor VII (3.4.21.21); Factor IX (3.4.21.22); Factor Xa (3.4.21.6); Factor XI (3.4.21.27); Factor XII (3.4.21.38)
• CPC Class Codes: A61K 47/18 Amines; Amides; Ureas; Quaternary ammonium compounds; Amino acids; Oligopeptides having up to five amino acids
• CPC Class Codes: A61K 47/22 Heterocyclic compounds, e.g. ascorbic acid, tocopherol or pyrrolidones
• CPC Class Codes: A61K 47/28 Steroids, e.g. cholesterol, bile acids or glycyrrhetinic acid
• CPC Class Codes: A61K 48/00 Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
• CPC Class Codes: A61P 1/00 Drugs for disorders of the alimentary tract or the digestive system
• CPC Class Codes: A61P 11/00 Drugs for disorders of the respiratory system
• CPC Class Codes: A61P 13/00 Drugs for disorders of the urinary system
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 19/00 Drugs for skeletal disorders
• CPC Class Codes: A61P 21/00 Drugs for disorders of the muscular or neuromuscular system
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 3/00 Drugs for disorders of the metabolism of the blood or the extracellular fluid A61P7/00
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/02 : Immunomodulators
• CPC Class Codes: A61P 43/00 : Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 5/00 : Drugs for disorders of the endocrine system
• CPC Class Codes: A61P 7/00 : Drugs for disorders of the blood or the extracellular fluid
• CPC Class Codes: A61P 9/00 : Drugs for disorders of the cardiovascular system
• CPC Class Codes: C07K 14/535 : Granulocyte CSF; Granulocyte-macrophage CSF
• CPC Class Codes: C12N 15/67 : General methods for enhancing the expression
• CPC Class Codes: C12N 15/87 : Introduction of foreign genetic material using processes not otherwise provided for, e.g. co-transformation
• CPC Class Codes: C12N 2310/335 : Modified T or U
• CPC Class Codes: C12N 9/644 : Coagulation factor IXa (3.4.21.22)

Abstract

Provided are formulations, compositions and methods for delivering biological moieties such as modified nucleic acids into cells to modulate protein expression. Such compositions and methods include the delivery of biological moieties, and are useful for production of proteins.

Claims

1. A method of producing a polypeptide of interest in a cell in a subject in need thereof, comprising administering to the subject a pharmaceutical composition comprising a modified messenger RNA (mmRNA), in a lipid nanoparticle (LNP) formulation, said lipid nanoparticle formulation comprising a lipid selected from the group consisting of DLin-DMA, DLin-K-DMA, DLin-KC2-DMA, DLin-MC3-DMA, 98N12-5, and C12-200;

a cholesterol; and

a PEG-lipid;

such that the mmRNA is introduced into the cell, wherein the mmRNA comprises a translatable region encoding the polypeptide of interest and comprises the modified nucleoside 1-methyl-pseudouridine, and wherein the pharmaceutical composition comprises an effective amount of the mmRNA providing for increased polypeptide production and substantially reduced innate immune response in the cell, as compared to a composition comprising a corresponding unmodified mRNA.

2. The method of claim 1, wherein the mmRNA comprises the modified nucleoside 1-methyl-pseudouridine in combination with 5-methyl-cytidine.

3. The method of claim 2, wherein the mmRNA comprises at least about 95% 1-methyl-pseudouridine in place of uracils and at least about 95% 5-methyl-cytidine in place of cytosines.

4. The method of claim 2, wherein the mmRNA comprises about 100% 1-methyl-pseudouridine in place of uracils and about 100% 5-methyl-cytidine in place of cytosines.

5. The method of claim 1, wherein the innate immune response is reduced from 80% to 99.9% as compared to the immune response induced by a corresponding unmodified mRNA.

6. The method of claim 1, wherein the mmRNA reduces the expression or activity level of a Type 1 interferon compared to the corresponding unmodified mRNA.

7. The method of claim 1, wherein the mmRNA and LNP are formulated at a total lipid to mmRNA weight ratio of between 10:

1 and 30;

1.

8. The method of claim 1, wherein the pharmaceutical composition is administered by an intravenous route.

10. The method of claim 1, wherein the pharmaceutical composition is administered by a subcutaneous route.

11. The method of claim 1, wherein the subject is a human.

12. The method of claim 1, where the pharmaceutical composition is administered at a total dose of about 0.1 mg/kg to about 40 mg/kg.

13. The method of claim 1, wherein administration occurs on a schedule selected from the group consisting of three times a day, twice a day, once a day, every other day, every third day, weekly, biweekly, every three weeks, every four weeks, and monthly.

14. The method of claim 12, wherein the total dose is administered by multiple administrations.

15. The method of claim 14, wherein the multiple administrations occur on a schedule selected from the group consisting of three times a day, twice a day, once a day, every other day, every third day, weekly, biweekly, every three weeks, every four weeks, and monthly.

16. The method of claim 1, wherein the increased polypeptide production is observed in a tissue or bodily fluid within 8 hours.

17. The method of claim 1, wherein the increased polypeptide production is observed in a tissue or bodily fluid within 2 hours.

9. The method of claim P, wherein the pharmaceutical composition is administered by an intramuscular route.