JPWO2019008001A5

JPWO2019008001A5 -

Info

Publication number: JPWO2019008001A5
Application number: JP2019572536A
Authority: JP
Publication date: 2022-05-16

Claims

An artificial nucleic acid molecule comprising at least one coding region, wherein the at least one coding region comprises.
With at least one antigenic peptide or protein,
With at least one T helper epitope,
It encodes with at least one additional amino acid sequence derived from at least one immune response activating signaling protein located in the external plasma membrane.
At least one immune response activation signaling protein located on the external plasma membrane is CTLA4.
An artificial nucleic acid molecule, wherein the at least one additional amino acid sequence comprises or consists of at least one transmembrane domain from CTLA4 .

The artificial nucleic acid molecule of claim 1, wherein the at least one additional amino acid sequence further comprises at least one cytoplasmic domain.

The at least one antigenic peptide or protein is selected or derived from a tumor antigen, virus, bacterium, protozoan, fungus, or allogeneic antigen.
The at least one antigenic peptide or protein optionally comprises or comprises the amino acid sequence corresponding to any of SEQ ID NOs: 3719 to 27945, 76420 to 76439, 76440 to 76474, or fragments, variants or derivatives thereof. , And / or optionally, the nucleic acid sequence according to any of SEQ ID NOs: 27946 to 52172, 76495 to 76514, 52173 to 76399, 76570 to 76589, 76515 to 76549, 76590 to 76624, or a fragment of any of the above sequences. The artificial nucleic acid molecule according to any one of claims 1 and 2, which is encoded by a variant or a derivative.

The tumor antigens are BRAF, PIK3CA, KRAS, IDH1, TP53, NRAS, AKTI, SF3B1, CDKN2A, RPSAP58, EGFR, NY-ESO1, MUC-1, 5T4, Her2, MAGE-A3, LY6K, CEACAM6, CEA, MCA. , KK-LC1, Gastrin, VEGFR2, MMP-7, MPHOSPH1, MAGE-A4, MAGE-A1, MAGE-C1, PRAME, Survival, MAGE-A9, MAGE-C2, FGFR2, WT1, PSA, PSMA, Prostate-specific antigen Precursor, Kitakyushu lung cancer antigen 1, nutrient membrane glycoprotein, cyclin-dependent kinase inhibitor 2A, cyclin-dependent kinase inhibitor 2A, isoform 1/2/3, multiple tumor suppressor 1 / cyclin-dependent kinase 4 inhibitor p16 , GTPase NRas, or the artificial nucleic acid molecule of claim 3, selected from any fragment, variant, or derivative of any of the tumor antigens, or any combination thereof.

The at least one additional amino acid sequence comprises or comprises at least one transmembrane domain and the amino acid sequence corresponding to SEQ ID NO: 76636, or at least one cytoplasmic domain comprising or consisting of fragments, variants, or derivatives thereof. Consisting with, the nucleic acid sequence corresponding to any of SEQ ID NOs: 76659, 76682, 76705, 76728, 76751, 76774, 76797, 76820, 76843, 76866, 76889, 76912, 76935, 76947, 77066, 77004 to 77017, or the above-mentioned sequence. The artificial nucleic acid molecule according to any one of claims 1 to 4, which is encoded by any fragment, variant, or derivative of, preferably SEQ ID NO: 77066.

The transmembrane domain comprises or comprises the amino acid sequence corresponding to SEQ ID NO: 200, or a fragment, variant, or derivative thereof, SEQ ID NO: 408, 616, 824, 1032, 1240, 1448, 1656, 1864, 2072, The artificial nucleic acid molecule according to any one of claims 1 to 5, which is encoded by a nucleic acid sequence corresponding to any of 2280, 2488, 2696, 2904, or a fragment, variant, or derivative of any of the sequences.

The artificial nucleic acid molecule according to any one of claims 1 to 6, further encoding at least one signal peptide and / or at least one linker in the at least one coding region.

The artificial nucleic acid molecule according to any one of claims 1 to 7, which preferably contains at least one coding region of the following formula (I) in the 5'→ 3'direction.

(During the ceremony,
"SIG" encodes a signal peptide,
"L" encodes the linker sequence and
Each "AN" encodes the same or different antigenic peptide or protein, preferably defined in claim 3 or 4.
"IM" encodes a helper epitope,
"TMD / TMCD" preferably encodes an amino acid sequence derived from the transmembrane domain of CTLA4 as defined in claim 6, and optionally, preferably a cytoplasmic domain as defined in claim 5.
m and o are integers independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10, respectively.
b, d, and n are integers independently selected from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10.
c is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10.
e is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10.
a is 1,
p is 1. )

The artificial nucleic acid molecule is RNA,
The artificial nucleic acid molecule according to any one of claims 1 to 8, optionally selected from mRNA, viral RNA, replicon RNA, or circular RNA.

The artificial nucleic acid molecule, preferably RNA, according to any one of claims 1 to 9, wherein the artificial nucleic acid molecule is modified and preferably stabilized.

The G / C content of the at least one coding region is increased compared to the G / C content of the corresponding coding sequence of the corresponding wild-type artificial nucleic acid.
The C content of the at least one coding region is increased compared to the C content of the corresponding coding sequence of the corresponding wild-type artificial nucleic acid, and / or.
The codons in the at least one coding region are adapted for human codon use and the codon goodness-of-fit index (CAI) is preferably increased or maximized in the at least one coding sequence of the artificial nucleic acid. ,
The artificial nucleic acid molecule according to any one of claims 1 to 10, wherein the amino acid sequence encoded by the artificial nucleic acid is preferably unchanged as compared with the amino acid sequence encoded by the corresponding wild-type artificial nucleic acid. RNA.

The at least one coding region includes SEQ ID NOs: 585, 1001, 1417, 1833, 2249, 793, 1209, 1625, 2041, 2457, 2665, 2873, 616, 824, 1032, 1240, 1448, 1656, 1864, 2072, 2280, 2488, 2696, 2904, 76682, 76705, 76728, 76751, 76774, 76797, 76820, 76843, 76866, 76889, 76921, 76935, 76947, 77004 to 77017, 77066, 564, 772, 980, 1188, 1396, The artificial nucleic acid molecule according to claim 11, preferably RNA, comprising or consisting of a nucleic acid sequence corresponding to any of 1604, 1812, 2020, 2228, 2436, 2644, 2852.

The artificial nucleic acid molecule according to any one of claims 1 to 12, preferably RNA, which comprises the following elements in the direction of preferably 5'→ 3'.
a) 5'-cap structure, preferably m7GpppN, ARCA Cap, or Cap1,
b) Optionally, a 5'-UTR element comprising or consisting of, optionally, the nucleic acid sequence corresponding to the nucleic acid sequence according to SEQ ID NO: 3061 or 3063, or a fragment, variant, or corresponding RNA sequence thereof.
c) At least one code sequence as defined in any of claims 1-12,
d) Optionally, preferably comprises or comprises a nucleic acid sequence corresponding to the nucleic acid sequence according to SEQ ID NO: 3065, 3067, 3069, 3071, 3073, 3075, or 3077, or a fragment, variant, or corresponding RNA sequence thereof. 3'-UTR element,
e) Optionally, preferably a poly (A) consisting of 10-1000, 10-500, 10-300, 10-200, 10-100, 40-80, or 50-70 adenosine nucleotides. ) Tail,
f) Optionally, a poly (C) tail consisting of 10-200, 10-100, 20-70, 20-60, or 10-40 cytosine nucleotides, preferably.
g) A histone stem-loop optionally comprising or consisting of the nucleic acid sequence corresponding to SEQ ID NO: 3079 or 3080.

A vaccine comprising at least one artificial nucleic acid molecule according to any one of claims 1 to 13, preferably RNA, and optionally a pharmaceutically acceptable carrier and / or excipient.

It comprises at least two plurality of artificial nucleic acid molecules according to any one of claims 1 to 14, preferably at least two of the plurality of artificial nucleic acid molecules encode different antigenic peptides or proteins, said antigenic peptide. Alternatively, the vaccine according to claim 14, wherein the protein is optionally selected from the antigenic peptide or protein defined in claim 3 or 4, or a fragment, variant, or derivative thereof.

The artificial nucleic acid molecule, preferably RNA,
a) One or more cationic or polycationic compounds, preferably cationic or polycationic polymers, cationic or polycationic peptides or proteins, such as protamine, cationic or polycationic polysaccharides, and / or cations. Complexed with a sex or polycationic lipid, said cationic or polycationic compound is optionally a polymer carrier and / or
b) The vaccine according to any of claims 14 to 15, which is complexed with one or more lipids to form lipid nanoparticles, lipoplexes, and / or preferably liposomes.

The artificial nucleic acid molecule according to any one of claims 1 to 13, preferably RNA, or the vaccine according to any one of claims 14 to 16, optionally a liquid vehicle and / or optionally the artificial nucleic acid molecule or the vaccine. A kit, preferably a kit of parts, comprising technical instructions including information on usage and dosage of the vaccine.

In methods of treating or preventing cancer, optionally for pharmaceutical use; infectious diseases including viral, bacterial, fungal, or protozoal infections; autoimmune diseases; transplant-to-host disease (GvHD); or allergies. The artificial nucleic acid molecule according to any one of claims 1 to 13, preferably RNA, the vaccine according to any one of claims 14 to 16, or the kit according to claim 17.