Systems to produce B cells genetically modified to express selected antibodies

Patent metadata

Field	Value
Publication number	US-12589145-B2
Application number	US-202318166151-A
Country	US
Kind code	B2
Filing date	Feb 8, 2023
Priority date	Oct 20, 2017
Publication date	Mar 31, 2026
Grant date	Mar 31, 2026

Abstract

Systems and methods to genetically modify B cells to express selected antibodies are described. The systems and methods can be used to: obviate the need for classical vaccinations; provide protection against infectious agents for which no vaccinations are currently available; provide protection against infectious agents when patients are otherwise immune-suppressed; and/or provide a benefit provided by a therapeutic antibody, such as in the treatment of autoimmune disorders.

First claim

What is claimed is:

1 . A genetic construct comprising or encoding (i) a heavy chain promoter, (ii) a signal peptide, (iii) a full-length light chain of a selected antibody; (iv) a flexible linker or a skipping element; (v) a variable region of a heavy chain of the selected antibody; and (vi) a splice junction comprising the sequence CAGGTAAGT or CAGGTGAGT and 40-80 base pairs of the intron following the last exon of a variable-diversity-joining (VDJ).

2 . The genetic construct of claim 1 , wherein the flexible linker is between the full-length light chain of the selected antibody and the variable region of the heavy chain of the selected antibody.

3 . The genetic construct of claim 1 , wherein the flexible linker is encoded by the nucleotide sequence as set forth in SEQ ID NO: 116; has the amino acid sequence as set forth in one of SEQ ID NOs: 122, 180-184; and/or is a Gly-Ser linker comprising 50-80 amino acids.

4 . The genetic construct of claim 1 , wherein the skipping element is between the full-length light chain of the selected antibody and the variable region of the heavy chain of the selected antibody.

5 . The genetic construct of claim 1 , wherein the skipping element comprises an internal ribosome entry site (IRES) or a self-cleaving peptide having the sequence as set forth in SEQ ID NOs: 176, 177, 178, or 179.

6 . The genetic construct of claim 1 , wherein the heavy chain promoter is IgVH1-69 or J558H10.

7 . The genetic construct of claim 1 , wherein the signal peptide is selected from the sequence as set forth in one of SEQ ID NOs: 118, 134, and 185-194.

8 . The genetic construct of claim 1 , wherein the signal peptide is a signal peptide of a human IgH heavy chain or a human IgL light chain.

9 . The genetic construct of claim 1 , wherein the genetic construct further comprises homology arms.

10 . The genetic construct of claim 9 , wherein the homology arms have the sequence as set forth in SEQ ID NO: 90-101, 110, 125, 127, 140, 142, 143, 153, 170, 171, 173, 174, 278, or 279.

11 . The genetic construct of claim 1 , wherein the genetic construct further encodes a tag having the sequence as set forth in SEQ ID NOs: 122, 195, 196, 197, 198, 199, 200, 201, 202, 203, or 204.

12 . The genetic construct of claim 1 , wherein the selected antibody is
an anti-RSV antibody comprising
(a) a variable heavy chain comprising the sequence as set forth in SEQ ID NO: 138 and a variable light chain comprising the sequence as set forth in SEQ ID NO: 136,
(b) a variable heavy chain comprising the sequence as set forth in SEQ ID NO: 138 and a variable light chain comprising the sequence as set forth in SEQ ID NO: 205,
(c) a variable heavy chain comprising the sequence as set forth in SEQ ID NO: 123 and a light chain comprising the sequence as set forth in SEQ ID NO: 120, or
(d) a variable heavy chain comprising the sequence as set forth in SEQ ID NO: 123 and a variable light chain comprising the sequence as set forth in SEQ ID NO: 206;
an anti-human immunodeficiency virus (HIV) antibody comprising a variable heavy chain comprising the sequence as set forth in SEQ ID NO: 150 and a variable light chain comprising the sequence as set forth in SEQ ID NO: 149; an anti-pertussis antibody comprising a variable heavy chain comprising the sequence as set forth in SEQ ID NO: 235 and a variable light chain comprising the sequence as set forth in SEQ ID NO: 236; an anti-influenza virus antibody comprising a variable heavy chain comprising the sequence as set forth in SEQ ID NO: 159 and a variable light chain comprising the sequence as set forth in SEQ ID NO: 158; anti-Epstein Barr virus (EBV) antibody comprising a variable heavy chain comprising the sequence as set forth in SEQ ID NO: 168 and a variable light chain comprising the sequence as set forth in SEQ ID NO: 166; or an anti-tumor necrosis factor (TNF) antibody comprising a variable heavy chain comprising the sequence as set forth in SEQ ID NO: 254 and a variable light chain comprising the sequence as set forth in SEQ ID NO: 255.

13 . A kit comprising a genetic construct of claim 1 and a gRNA that binds a genomic region comprising SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, or SEQ ID NO: 4.

14 . The kit of claim 13 , wherein the gRNA has the sequence as set forth in
SEQ ID NO: 88, 89, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, or 307 when the genomic region comprises SEQ ID NO: 1; SEQ ID NO: 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, or 327 when the genomic region comprises SEQ ID NO: 2; SEQ ID NO: 87, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, or 346 when the genomic region comprises SEQ ID NO: 3; or SEQ ID NO: 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, or 366 when the genomic region comprises SEQ ID NO: 4.

15 . The kit of claim 13 , further comprising a nuclease.

16 . The kit of claim 15 , wherein the nuclease is Cas9 or Cpf1.

17 . The kit of claim 13 , further comprising a nanoparticle or adeno-associated viral vector.

18 . The kit of claim 15 , wherein the gRNA and nuclease are associated with a nanoparticle and the genetic construct is part of an adeno-associated viral vector.

19 . Am isolated B cell comprising the genetic construct of claim 1 , wherein the genetic construct is inserted into a genomic region having the sequence of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, or SEQ ID NO: 4.

20 . The B cell of claim 19 , wherein the B cell is an antibody-secreting B cell, a memory B cell, a naïve B cell, a B 1 B cell or a marginal zone B cell.

Assignees

• Fred Hutchinson Cancer Center

Inventors

• Taylor Justin J
• Moffett Howell F

Classifications

• C07K16/114

  Lentivirus (G), e.g. human immunodeficiency virus [HIV], feline immunodeficiency virus [FIV] or simian immunodeficiency virus [SIV] · CPC title

• C07K16/108

  Orthomyxoviridae (F), e.g. influenza virus · CPC title

• C07K16/11

  Paramyxoviridae (F); Pneumoviridae (F), e.g. respiratory syncytial virus [RSV] · CPC title

• C12N15/861

  Adenoviral vectors · CPC title

• A61K40/46

  Viral antigens · CPC title