Recombinant vectors suitable for the treatment of ipex syndrome

Patent metadata

Field	Value
Publication number	US-2022017919-A1
Application number	US-201917295093-A
Country	US
Kind code	A1
Filing date	Nov 19, 2019
Priority date	Nov 20, 2018
Publication date	Jan 20, 2022
Grant date	—

Abstract

IPEX (Immune dysregulation Polyendocrinopathy X linked) syndrome is a primary immunodeficiency caused by mutations in the gene encoding the transcription factor forkhead box P3 (FOXP3), which leads to the loss of function of thymus-derived CD4+CD25+ regulatory T (tTreg) cells. Preclinical and clinical studies suggest that T cell gene therapy approaches designed to selectively restore the repertoire of Treg cells by transfer of wild type FOXP3 gene is a promising potential cure for IPEX. However, there is still a need for a vector that can be used efficiently for the preparation of said Treg cells. The inventors thus compared 6 different lentiviral constructs according to 4 criteria (vector titers, level of transduction of human CD4+ T cells, level of expression of FOXP3 and ΔLNGFR genes, degree of correlation between both expression) and selected one construct comprising a bidirectional PGK-EF1a promoter that showed remarkable efficiency.

First claim

1 . A recombinant nucleic acid molecule comprising a bidirectional PGK-EF1a promoter operably linked to a first transgene in one direction and to a second transgene in the opposite direction, wherein the bidirectional PGK-EF1a promoter comprises a first PGK portion and a second EF1a portion, and wherein
the first transgene is under control of the first PGK portion of the bidirectional PGK-EF1a promoter and encodes a protein that is not constitutively expressed by a T cell, and the second transgene is under control of the second EF1a portion of the bidirectional PGK-EF1a promoter and encodes a transcription factor.

2 . The recombinant nucleic acid molecule of claim 1 wherein the first PGK portion comprises a nucleic sequence having at least 80% of identity with the nucleic acid sequence as set forth in SEQ ID NO:3.

3 . The recombinant nucleic acid molecule of claim 1 wherein the second EF1a portion comprises a nucleic sequence having at least 80% of identity with the nucleic acid sequence as set forth in SEQ ID NO:2.

4 . The recombinant nucleic acid molecule of claim 1 wherein the first PGK portion and the second EF1a portion are separated by a spacer sequence.

5 . The recombinant nucleic acid molecule of claim 4 wherein the spacer sequence comprises a nucleic sequence having at least 80% of identity with the nucleic acid sequence as set forth in SEQ ID NO:4.

6 . The recombinant nucleic acid molecule of claim 1 wherein the bidirectional promoter comprises a nucleic acid sequence having at least 80% of identity with the sequence as set forth in SEQ ID NO:5.

7 . The recombinant nucleic acid molecule of claim 1 wherein the sequences of the first transgene and the second transgene are codon-optimized.

8 . The recombinant nucleic acid molecule of claim 1 wherein the first transgene that is under the control of the first PGK portion of the bidirectional promoter encodes for a low-affinity nerve growth factor receptor (LNGFR).

9 . The recombinant nucleic acid molecule of claim 1 wherein the second transgene that is under the control of the second EF1a portion of the bidirectional promoter encodes for FoxP3.

10 . The recombinant nucleic acid molecule of claim 1 which comprises:
i) a first nucleic acid sequence having at least 80% of identity with the nucleic acid sequence as set forth in SEQ ID: 8,
ii) a second nucleic acid sequence having at least 80% of identity with the nucleic acid sequence acid sequence as set forth in SEQ ID NO:5 and
iii) a third nucleic acid sequence having at least 80% of identity with the nucleic acid sequence as set forth in SEQ ID NO:6.

11 . The recombinant acid molecule of claim 1 which comprises a nucleic acid sequence having at least 80% of identity with the nucleic acid sequence as set forth in SEQ ID NO:11.

12 . A lentiviral vector which comprises the recombinant acid molecule of claim 1 .

13 . (canceled)

14 . A method of producing a population of Treg cells, comprising the step of transfecting or transducing a population of T cells in vitro or ex vivo with the lentiviral vector of claim 12 .

15 . A population of Treg cells obtainable by the method of claim 14 .

16 . A method of treating an autoimmune disease in a subject in need thereof comprising administering to the subject a therapeutically effective amount of the population of Treg cells of claim 15 .

17 . The method of claim 16 wherein the autoimmune disease is IPEX syndrome.

18 . A nucleic acid sequence as set forth in SEQ ID NO:7.

19 . The recombinant nucleic acid molecule of claim 8 wherein an intracytoplasmic part (ΔLNGFR) is truncated from the LNGFR.

Assignees

• Inst Nat Sante Rech Med
• Univ Paris
• Assist Publique Hopitaux Paris Aphp
• Fond Imagine
• Univ Devry Val Dessonne
• Ecole Pratique Des Hautes Etudes
• Medizinische Hochschule Hannover

Inventors

• Andre Isabelle
• Six Emmanuelle
• Bellier Florence
• Delville Marianne
• Cavazzana Marina
• Amendola Mario
• Schambach Axel

Classifications

• C12N15/86 Primary

  Viral vectors · CPC title

• A61K40/416

  Antigens related to auto-immune diseases; Preparations to induce self-tolerance · CPC title

• A61K40/22

  Immunosuppressive or immunotolerising · CPC title

• A61K40/11

  T-cells, e.g. tumour infiltrating lymphocytes [TIL] or regulatory T [Treg] cells; Lymphokine-activated killer [LAK] cells · CPC title

• C12N5/0637

  Immunosuppressive T lymphocytes, e.g. regulatory T cells or Treg · CPC title