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Nucleic acid for treating mite allergy

A nucleic acid and amino acid technology, applied in allergic diseases, applications, antibody medical components, etc., can solve problems such as nucleic acid vaccines that have not yet been reported

Pending Publication Date: 2020-12-18
ASTELLAS PHARMA INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, a nucleic acid vaccine for mite allergy treatment containing multiple mite allergen antigens as well as the organelle stabilization domain and endosomal / lysosomal targeting domain of LAMP-1 has not been reported yet.

Method used

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  • Nucleic acid for treating mite allergy
  • Nucleic acid for treating mite allergy
  • Nucleic acid for treating mite allergy

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0289] [Example 1: Construction of LAMP-Der p 1-Der p 2-Der p 23-Der p 7 plasmid]

[0290] Construction of the LAMP-Der p 1-Der p 2-Der p 23-Der p7 plasmid (an expression vector comprising the following nucleic acid comprising the following Base sequence (that is, base sequence encoding a chimeric protein composed of the amino acid sequence shown in SEQ ID NO: 2): base sequence encoding the signal peptide of LAMP-1 (amino acid sequence 1 to 27 in SEQ ID NO: 2) , the nucleotide sequence encoding the organelle stabilization domain of LAMP-1 (the amino acid sequence of 28 to 380 in SEQ ID NO: 2), the encoding sequences comprising Der p 1, Der p 2, Der p 23 and Der p 7 The base sequence of the original domain (amino acid sequence 383 to 1002 of SEQ ID NO: 2), the base sequence encoding the transmembrane domain of LAMP-1 (amino acid sequence 1006 to 1028 of SEQ ID NO: 2), and the base sequence encoding LAMP-1 The base sequence of the endosome / lysosome targeting domain of 1 (amino ...

Embodiment 2

[0291] [Example 2: Expression of LAMP-Der p 1-Der p 2-Der p 23-Der p 7 chimeric protein]

[0292] Using the 293T cell strain derived from human fetal kidney, the LAMP-Der p 1-Der p 2-Der p 23-Der p7 chimeric protein (the amino acid sequence encoded by the base sequence shown in SEQ ID NO: 1 (i.e. the sequence The in vitro expression of a chimeric protein composed of the amino acid sequence shown in No. 2) was evaluated.

[0293] (1) Cell culture and plasmid introduction

[0294] 293T cells derived from human fetal kidney (Thermo Fisher Scientific Cat. Cat.15070063) in the D-MEM medium (Sigma-Aldrich Company, Cat.D5796) with 3 × 10 5 Cells were seeded into 6-well plates (IWAKI, Cat. 3810-006) in a manner of per well. The inoculated cells were incubated at 37°C, 5% CO 2 After culturing overnight in the presence of LAMP-Der p 1-Der p 2-Der p 23-Der p7 plasmid: Lipofectamine 2000 (Thermo Fisher Scientific Cat. 11668027) = 2.5 (μg): 10 (μL) mixture . Further at 37°C, 5% CO 2...

Embodiment 3

[0304] [Example 3: Induction of IgG2a production by administration of LAMP-Der p 1-Der p 2-Der p 23-Der p 7 plasmid]

[0305]Evaluation of antibody production induction in vivo was performed. For 8 cases in each group, 7-week-old BALB / c female mice (Charles River Japan Co., Ltd., Japan) were intradermally administered with 50 μg of LAMP-Der p 1-Der p 2 three times a week at the start of administration. - 25 μL of PBS solution of Der p 23-Der p 7 plasmid (Day 0, 7 and 14), blood was collected 1 week after the last administration, and a plasma sample was obtained (Day 21). As a comparative control, prepare: LAMP-Der p 23-Der p 7-Der p 2-Der p 1 plasmid (an expression vector comprising the following nucleic acid, which comprises a base sequence that sequentially contains the following base sequence: coding sequence The amino acid sequence of amino acid numbers 1 to 380 of No. 2 (hereinafter referred to as the N-terminal of LAMP-1 in Examples 3 and 4), the base sequence and the c...

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Abstract

The invention is to provide a nucleic acid expected to be useful in treating mite allergy. Provided is a nucleic acid including, in the following order: a base sequence that encodes a signal peptide;a base sequence that encodes an LAMP intra-organelle stabilizing domain; a base sequence that encodes an allergen domain including Derp1, Derp2, Derp23, and Derp7; a base sequence that encodes a transmembrane domain; and a base sequence that encodes an LAMP endosomal / lysosomal targeting domain, wherein the nucleic acid includes a base sequence that encodes a chimeric protein.

Description

technical field [0001] The present invention relates to nucleic acids expected to be useful as active ingredients of pharmaceutical compositions, for example nucleic acids expected to be useful in the treatment of mite allergy. Background technique [0002] Mite allergy is an allergic disease caused by a reaction to an allergen derived from mites. Allergic diseases occur as follows: 1) Allergens taken into the body are phagocytized by antigen-presenting cells and presented to naive T cells by antigens; 2) Naive T cells differentiate into Th2 cells; 3) Th2 cells, etc. Immune cells produce cytokines such as IL-4; 4) B cells produce IgE under the action of IL-4; 5) IgE combined with allergens binds to mast cells. It is known that in patients with allergic diseases, Th2 type immunity is dominant in the counterbalance between Th1 type immunity in which Th1 cells producing IFN-γ and the like participate and Th2 type immunity in which Th2 cells producing IL-4 etc. participate, and...

Claims

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Application Information

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IPC IPC(8): C12N15/62A61K31/7088A61P37/00C12N1/15C12N1/19C12N1/21C12N5/10C12N15/12C12N15/63
CPCA61P37/00A61K31/7088A61K48/005C07K2319/02C07K2319/03C07K2319/06C07K14/43527C07K14/70596A61K38/00C12N15/62A61P37/08A61K39/35A61K2039/5256A61K2039/53A61K2039/57C12N9/50C12Y304/22065C07K19/00C07K14/43531C12N15/63A61K38/177
Inventor 丸井崇则内田征男
Owner ASTELLAS PHARMA INC