Engineered nucleic acids and methods of use thereof

• US 9,937,233 B2
• Filed: 08/26/2016
• Issued: 04/10/2018
• Est. Priority Date: 08/06/2010
• Status: Active Grant

• Current Assignee: Moderna TX, Inc. (Moderna, Inc.)
• Original Assignee: Moderna TX, Inc. (Moderna, Inc.)
• Inventors: Schrum, Jason P., Bancel, Stephane, Afeyan, Noubar B., Ejebe, Kenechi
• Primary Examiner(s): Nguyen, Quang
• Application Number:
  US15/248,053
• Publication Number:
  US 20170143796A1
• Time in Patent Office: 592 Days
• CPC Class Codes: A61K 2300/00 Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
• CPC Class Codes: A61K 31/395 having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
• CPC Class Codes: A61K 31/495 having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazinesA61K31/48 takes precedence ; with three nitrogen atoms A61K31/53
• 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 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61K 38/13 Cyclosporins
• CPC Class Codes: A61K 38/193 Colony stimulating factors [CSF]
• CPC Class Codes: A61K 38/195 Chemokines, e.g. RANTES
• CPC Class Codes: A61K 38/28 Insulins
• CPC Class Codes: A61K 39/3955 against proteinaceous materials, e.g. enzymes, hormones, lymphokines
• 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: A61K 48/0066 Manipulation of the nucleic acid to modify its expression pattern, e.g. enhance its duration of expression, achieved by the presence of particular introns in the delivered nucleic acid
• CPC Class Codes: A61K 48/0075 characterised by an aspect of the delivery route, e.g. oral, subcutaneous
• CPC Class Codes: A61K 48/0083 characterised by an aspect of the administration regime
• 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 37/00 Drugs for immunological or allergic disorders
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07K 14/62 Insulins
• CPC Class Codes: C07K 14/685 Alpha-melanotropin

Abstract

Provided are compositions and methods for delivering biological moieties such as modified nucleic acids into cells to modulate protein expression. Such compositions and methods include the use of modified messenger RNAs, and are useful to treat or prevent diseases, disorders or conditions, or to improve a subject'"'"'s heath or wellbeing.

Claims

1. A method comprising:

administering to a subject in need thereof a pharmaceutical composition comprising;

i) a synthetic messenger ribonucleic acid (mRNA) encoding a G-CSF polypeptide; and

ii) a pharmaceutically acceptable carrier,

wherein the synthetic mRNA sequence is at least 95% identical to the sequence of SEQ ID NO.;

17.

2. The method of claim 1, wherein the subject in need of G-CSF administration has or is suspected of having a disease, and the synthetic mRNA encoding a G-CSF polypeptide is in an amount sufficient to modulate one or more activities associated with G-CSF to treat the disease.

3. The method of claim 1, wherein the composition further comprises a lipid-based transfection reagent.

4. The method of claim 1, wherein the composition further comprises a cell penetration agent.

5. The method of claim 1, wherein the synthetic mRNA is administered to the subject in a dosage range of about 1.0 μ

g/kg to about 100 mg/kg of body weight.

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

7. The method of claim 1, wherein the administration is repeated two, three, four, five or more than five times.

8. The method of claim 1, wherein the administration occurs once daily.

9. The method of claim 1, wherein the administration occurs once every two days.

10. The method of claim 1, wherein the composition is administered to the subject via an intravenous injection.

11. The method of claim 1, wherein the composition is administered to the subject via a subcutaneous injection.

12. The method of claim 1, further comprising administering an effective amount of AMD3100 (1,1′

-[1,4-phenylene-bis(methylene)]-bis-1,4,8,11-tetraazacyclotetradecane) to the subject after administering the composition.

13. The method of claim 12, wherein the composition is administered at least twice but fewer than ten times prior to administering the AMD3100.

14. The method of claim 1, wherein the composition further comprises a synthetic messenger ribonucleic acid (mRNA) encoding i) a macrophage inflammatory protein (MIP) polypeptide or ii) an antibody that prevents EGF binding to EGFR.

17. The method of claim 1, wherein the mRNA comprises pseudouridine, 5′

-methylcytidine, or a combination thereof.

20. The method of claim 1, wherein 100% of cytidine and uridine nucleosides of the synthetic messenger ribonucleic acid are modified.

23. The method of claim 1, wherein the synthetic messenger ribonucleic acid (mRNA) is identical to the sequence of SEQ ID NO.:

18.

26. The method of claim 1, wherein the synthetic messenger ribonucleic acid (mRNA) is encapsulated in a nanoparticle.

27. The method of claim 1, wherein the synthetic messenger ribonucleic acid (mRNA) is complexed in liposomal form.

28. The method of claim 1, wherein the administration of the synthetic messenger ribonucleic acid (mRNA) encoding a granulocyte colony-stimulating factor (G-CSF) elevates total white blood cell count in the subject.

15. A method of efficient protein production in a cell population, the method comprising contacting a cell population with a modified nucleic acid comprising:

(i) a translatable region encoding a recombinant polypeptide; and

(ii) at least one nucleoside modification, under conditions such that an effective amount of the protein is present in the cell population, wherein the recombinant polypeptide is a granulocyte colony-stimulating factor (G-CSF) polypeptide and wherein the nucleic acid is a synthetic mRNA, and the synthetic mRNA sequence is at least 95% identical to the sequence of SEQ ID NO.;

17.

18. The method of claim 15, wherein the mRNA comprises pseudouridine, 5′

-methylcytidine, or a combination thereof.

21. The method of claim 15, wherein 100% of cytidine and uridine nucleosides of the synthetic messenger ribonucleic acid are modified.

24. The method of claim 15, wherein the synthetic messenger ribonucleic acid (mRNA) is identical to the sequence of SEQ ID NO.:

18.

16. A method for inducing in vivo translation of a recombinant polypeptide in a mammalian subject in need thereof, comprising administering to the subject an effective amount of a composition comprising a nucleic acid comprising:

(i) a translatable region encoding a recombinant polypeptide; and

(ii) at least one nucleoside modification, under conditions such that the nucleic acid is localized into a cell of the subject and the recombinant polypeptide is capable of being translated in the cell from the nucleic acid, wherein the recombinant polypeptide is a granulocyte colony-stimulating factor (G-CSF) polypeptide and wherein the nucleic acid is a synthetic mRNA, and the synthetic mRNA sequence is at least 95% identical to the sequence of SEQ ID NO.;

17.

19. The method of claim 16, wherein the mRNA comprises pseudouridine, 5′

-methylcytidine, or a combination thereof.

22. The method of claim 16, wherein 100% of cytidine and uridine nucleosides of the synthetic messenger ribonucleic acid are modified.

25. The method of claim 16, wherein the synthetic messenger ribonucleic acid (mRNA) is identical to the sequence of SEQ ID NO.:

18.

29. A method of accelerating recovery from neutropenia, the method comprising administering to a subject in need of such recovery a composition comprising a synthetic messenger ribonucleic acid (mRNA) encoding a granulocyte colony-stimulating factor (G-CSF) polypeptide in an amount sufficient to accelerate recovery from neutropenia, wherein the synthetic mRNA sequence is at least 95% identical to the sequence of SEQ ID NO.:

17.

30. The method of claim 29, wherein the subject is human.

31. The method of claim 30, wherein the subject in need of recovery is a cancer patient.

32. The method of claim 31, wherein the cancer patient has undergone chemotherapy or irradiation.

33. The method of claim 32, wherein the cancer patient that has undergone chemotherapy or irradiation is a patient receiving a stem cell transplant.

34. The method of claim 29, wherein the composition further comprises a pharmaceutically acceptable carrier.